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The CLEAPSS

Recipe Book

Introduction to this Edition

Recipe Cards were first produced in 1991 but very quickly demand led to an increase in the scope and

range of the information included. Over the ensuing 20 years we have added more and more

information whilst retaining the card format which users found useful.

This 2011 edition sees a change from cards to A4 sheets. The A5 card has become too small to contain

all the information required for some topics, and we believe that there is no further scope for reducing

the size of the print.

We have extended the number of separate entries from 74 to 106 to allow for more information on

some topics and to make other information easier to locate. We have added colour and some

photographs, where these are useful. The loose leaf A4 format also allows plenty of room for you to

insert recipes and instructions of your own.

Risk Assessment

Where a risk assessment includes control measures these have been incorporated into the instructions,

as describe in our Guidance Leaflet G90, Making and Recording Risk Assessments in School Science.

Each recipe therefore includes model risk assessments but does not include factors that you have to

thinks about, such as technician and teacher experience, and prep room conditions.

As in the previous edition, there is no mention of general “bench solutions”. In the past such solutions

were often much more concentrated than required and posed unnecessary hazards and risks. The

principle we follow (as in COSHH) is that the concentration of any reagent should be the lowest at

which the procedure works satisfactorily to give the intended result. Sometimes, therefore, we suggest

solutions which may appear to be an odd concentration but this nevertheless is the most suitable. For

example, 0.4 M sodium hydroxide solution is IRRITANT, whereas 0.5 M is CORROSIVE.

Making solutions

The recipes also make use of the laboratory jug as a measuring tool. Although apparently not

particularly accurate, we routinely achieve concentrations of between 1.95 and 2.05 M when using a jug

to prepare 2M sulfuric(VI) acid from concentrated sulfuric(VI) acid during the Practical Techniques in

Chemistry course. Clearly, using a jug can produce solutions with concentrations that are sufficiently

accurate for many laboratory purposes.

You will find more detail about making solutions in section 7.6 of the Handbook. If you run into

difficulties not covered in either the Recipe Book or the Handbook, phone CLEAPSS on 01895 251496

but do try the index first.

Contents

Recipe sheet Number

Agar 1

Alcohol/water and

propanone/water solutions

2

Alginate beads 3

Aluminium solutions 4

Ammonia gas 5

Ammonia solution (ammonium

hydroxide)

6

Ammonium chloride 7

Ammonium vanadate(V) solution 8

Azo dyes 9

Barium solutions 10

Benedict’s qualitative reagent 11

Benedict’s quantitative reagent 12

Biochemical indicators and tests 13

Bismuth nitrate(V) solution 14

Biuret reagent 15

Brodie’s fluid 16

Bromine water 17

Buffer solutions 18

Calcium chloride and nitrate(V)

solutions

19

Calcium hydroxide solution 20

Carbon dioxide 21

Cerium(IV) solutions 22

Chemiluminesence reactions 23

Chlorine gas 24

Chlorine water 25

Chromatography solvents and

locating agents

26

Chromium(III) chloride and

chrome alum solutions

27

Citric acid 28

Clock reactions

29

Recipe sheet Number

Cobalt(II) chloride solution and

thermochromic liquid

30

Copper(II) solutions 31

Crude oil alternative 32

2,4-Dinitrophenylhydrazine

solution

33

3,5-Dinitrosalicylic acid 34

Drosophila food base 35

Electroplating solutions 36

Enzymes 37

Etching solutions 38

Ethanoic acid 39

Fehling’s solutions 40

Fixatives used before preserving

biological specimens

41

Gases less commonly used in

schools 42

Hydrochloric acid 43

Hydrogen gas 44

Hydrogen peroxide 45

Indicators (acid-base) 46

Indicator (universal) 47

Indicators (carbon dioxide) 48

Indicators for redox, precipitation

and complexometric titrations 49

Iodine solution 50

Iron(II) solutions 51

Iron(III) solutions 52

Lead(II) nitrate(V) 53

Lithium chloride 54

Magnesium sulfate(VI) 55

Manganese(II) sulfate(VI) 56

Mercury solutions 57

Methanal solution 58

Methanoic acid 59

Recipe sheet Number

Nickel sulfate(VI) 60

Nitric(V) acid 61

Nylon rope experiment 62

Oscillating reactions 63

Oxygen gas 64

Phosphoric(V) acid 65

Plant mineral requirement

solutions

66

Potassium and sodium

phosphates

67

Potassium chloride 68

Potassium chromate(VI) 69

Potassium dichromate(VI) 70

Potassium hydroxide 71

Potassium iodide 72

Potassium manganate(VII) 73

Preservatives used after fixing

biological specimens

74

Ringer’s and other saline

solutions for physiological use

75

Sandell’s solution 76

Silver nitrate(V) 77

Slime 78

Soap and bubble solutions 79

Sodium carbonate 80

Sodium chlorate(I) solution 81

Sodium chloride 82

Sodium ethanoate 83

Recipe sheet Number

Sodium hydrogencarbonate 84

Sodium hydroxide 85

Sodium silicate, the crystal

(chemical) garden and silicate

gels

86

Sodium thiosulfate 87

Stains for bacterial activity 88

Stains for cell contents 89

Stains for electrophoresis 90

Stains for fungal material 91

Stains for metabolic activity 92

Stains for plant material 93

Standard solutions for titration 94

Strontium chloride 95

Sulfur dioxide 96

Sulfur dioxide solution 97

Sulfuric(VI) acid 98

Testing for gases 99

Testing for negative ions 100

Testing for positive ions 101

Testing for organic functional

groups

102

Tin(II) chloride 103

Water (sea and hard) 104

Winkler’s method for dissolved

oxygen

105

Zinc sulfate(VI) 106

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1 Agar

For microbiological activities using purchased media, follow the instructions on the bottle. The recipes are grouped into agars for microbiology and agars for other activities. All agars for microbiological work need to be sterilised before and after use. General Hazards Agar inhaled as a fine powder may cause an allergic reaction or other respiratory

problems. The use of agar that could isolate human pathogens (eg, blood agar) should be avoided.

Control measures

Use a balance in a non-working fume cupboard, ie, not switched on, with the sash down, to weigh out agar. Use heatproof gloves to protect from scalding when handling freshly-sterilised molten agar.

Procedure for preparing technical agar (also called agar-agar)

Mix 1.5 g of agar with 10 ml of water into a paste. Slowly add more water with stirring until the volume is 100 ml. Heat the mixture with stirring on a boiling water bath to 95 °C in the required container. This preliminary heating can be omitted if the agar is going to be sterilised immediately, unless it is necessary to decant the agar into smaller containers. In acid media, the amount of agar should be increased from 1.5 to 2 g. If the solidified agar in any recipe is too sloppy or too firm, repeat the procedure using slightly more or less agar. The agar gel is not stable in strongly alkaline solutions.

Usual sterilising conditions

(If required) Autoclave the container(s) with the made-up suspension(s) for 15 minutes at 15 psi (121 °C).

Agars for microbiology

China blue lactose agar

Use 3.6 g of China blue lactose agar powder in 100 ml of distilled water. If this mix proves too thin for ‘rough’ handling by students, then thicken by adding 0.5 g of agar-agar (just thickener, no nutrients).

Crystal violet agar. A selective (against Gram positive) medium for soil bacteria

In a fume cupboard which is not switched on, add 0.005 g of crystal violet (HARMFUL, DANGEROUS FOR THE ENVIRONMENT) to 1 litre of liquid nutrient agar solution. The resultant solution is low hazard.

Glucose nutrient agar

Add 0.5% w/v of glucose to molten nutrient agar.

Malt agar for fungi Mix 2 g of malt extract with 2 g of agar with 10 ml of water into a paste. Slowly add more water with stirring until the volume is 100 ml. Autoclave the suspension at 10 psi (115 °C) for 10 minutes.

Mannitol yeast extract agar for growing root nodule bacteria

Mix 10 g agar in 1 litre of water, and dissolve in a boiling water bath. Add 0.5 g K2HPO4, 0.2 g MgSO4.7H2O, 0.2 g NaCl, 0.2 g CaCl2.6H2O,10 g mannitol, and 0.4 g yeast extract. Dispense as required and sterilise by autoclaving before use.

Nitrogen-free mineral salts agar for growing nitrogen-fixing bacteria

Dissolve 0.05 g FeCl3.6H2O in 500 ml distilled water. Add 2 g K2HPO4, 0.25 g MgSO4.7H2O and 10 g glucose. Check the pH and adjust, if necessary, to 8.3 using 0.1 M NaOH. Pour into a bottle containing 1 g CaCO3 and 7.5 g agar powder. Mix and autoclave at 121 °C for 20 minutes. Before pouring plates, swirl to thoroughly mix the CaCO3 and agar.

Nutrient agar for bacteria

Mix 2 g of ‘Bovril’, 0.5 g of sodium chloride and 1.5 g of agar with 10 ml of water into a paste. Slowly add more water with stirring until the volume is 100 ml. Heat/sterilise the suspension as in ‘Usual sterilising conditions’ above.

Starch malt agar for growth of fungi and digestion of starch

Mix 3 g of light malt powder (from home-brewing shops), 0.5 g of peptone (to promote growth) in 20 ml of water. Also make a paste containing 1 g of soluble starch in 10 ml of hot water. Add these two solutions to 1.5 g of agar with stirring and slowly add more water with stirring until the volume is 100 ml. Stir before decanting into smaller containers (if required) and sterilising. Autoclave the suspension at 10 psi (115 °C) for 10 minutes.

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Agars for other activities These should be made up as required, and not stored for long periods to avoid any unwanted microbial activity. Dispose of as soon as possible after the activity. Sterilise in an autoclave any which are suspected of microbial contamination.

Agar for starch synthesis

Mix 0.5 g of glucose-1-phosphate and 1.5 g of agar with 10 ml of water. Slowly add more water with stirring until the volume is 100 ml. Boiling, not sterilising, should be sufficient.

Electrolytic agar Add one gram of sodium sulfate(VI) or other electrolyte to the hot agar solution before pouring.

Ferroxyl agar gel for rusting experiments

Add 1.4 g of agar, 2 g of sodium chloride, 0.1 g of potassium hexacyanoferrate(III) and 1 ml of phenolphthalein solution to the 100 ml of water and warm, with stirring, to 95 °C. Pour the solution into Petri dishes.

Indicator agar Add 1 ml of the chosen indicator solution to the agar solution before pouring.

Mayonnaise agar for lipase activity (1)

Dilute 4 g salad cream or mayonnaise with 5 ml water and add 1 ml 0.1 M sodium hydroxide solution (IRRITANT). Add about 1 ml of this alkaline mixture to a solution of bromocresol green dye (about 0.003 g in 100 ml water) until the mixture just turns blue-green and stir to ensure even distribution. Boil the resultant mixture with 2 g agar, cool to 50-60 °C then pour thin layers in Petri dishes. The plates will need to be incubated at 30 °C for 24 hours before being examined for orange-yellow areas produced by lipases.

Mayonnaise agar for lipase activity (2)

Alternatively, mayonnaise agar can be made up without the dye and, after incubation, the plates can be flooded with 0.4 M copper(II) sulfate solution and left for 30 minutes before being examined. Clear areas in the blue-green matrix indicate where lipases have broken down the fatty acids in the mayonnaise.

Milk agar for protease activity

Stir together 2 g low-fat milk powder (Marvel is recommended as it contains very little fat), 1 g agar and 100 ml water. Heat as for technical agar and pour into Petri dishes in very thin layers. Proteases should produce clear patches by breaking down proteins in the milk within 30 minutes or so.

Phenolphthalein indicator agar

Use 2 g agar in 100 ml boiling distilled (or deionised) water in a beaker. Add 10 ml of 0.2 M sodium carbonate solution and 5 ml of phenolphthalein into the beaker and stir well. Carbon dioxide in the atmosphere causes the colour to fade on storage, so the agar is better prepared shortly before it is required.

Starch agar for amylase activity

Mix a paste containing 1 g of soluble starch in 10 ml of cold water. Add 1.5 g of agar, stir well and slowly add more water with stirring until the volume is 100 ml. Heat as for technical agar above.

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2 Alcohol/water and propanone/water solutions

Quoted flash points appear to vary slightly between sources. To prepare molar solutions, liquids can be weighed in tared containers. 2 M propanone solution is used in a rate of reaction experiment called the iodination of propanone.

To prepare 100 ml of an x% (v/v) solution of ethanol in water

• Add x ml of ethanol to a 100 ml measuring cylinder. • Add water up to the 100 ml mark. • Label the solution. If it is highly flammable, then it needs the hazard warning label but, if flammable, a

label is not needed. However, the hazard classification needs to be written on any risk assessment.

% ethanol 0 10 20 30 40 50 60 70 80 90 100 Flash point (°C) - 49 36 29 26 24 22 21 20 17 13 Density (g cm-3) at 20 °C 1.00 0.98 0.97 0.96 0.95 0.93 0.91 0.89 0.86 0.83 0.78 Hazard - FLAMMABLE HIGHLY FLAMMABLE

To prepare 1 litre of 1 M methanol solution

• Weigh out 32 g of methanol in a tared container or measure out 41 ml of methanol in a measuring cylinder.

• Add this to a 1000 ml measuring cylinder or 1 litre measuring jug. • Add water up to the 1 litre mark. • Label the solution harmful.

To prepare 1 litre of 1 M ethanol solution

• Weigh out 46 g of ethanol in a tared container or measure out 58 ml of ethanol in a measuring cylinder.

• Add this to a 1000 ml measuring cylinder or 1 litre measuring jug. • Add water up to the 1 litre mark. • The solution is low hazard.

To prepare 1 litre of 2 M propanone solution

• Weigh out 116 g of propanone in a tared container or measure out 147 ml of propanone in a measuring cylinder.

• Add this to a 1000 ml measuring cylinder or 1 litre measuring jug. • Add water up to the 1 litre mark. • The solution is low hazard.

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3 Alginate beads

In studies of enzymes or the physiology of yeast cells, a valuable technique is to immobilise the enzyme or cells inside beads of sodium alginate. The beads containing the enzyme/cells can then be used as usual or packed into a column (eg, a syringe barrel) and a suitable substrate passed over them. The products are collected at the bottom of the column and the immobilised enzymes or cells can be used again. When making up the alginate and enzyme solutions it is essential to use purified water; otherwise calcium ions in the water will cause the alginate to ‘set’ prematurely. Alginate beads can usually be stored overnight, covered and refrigerated but are unlikely to keep longer than their non-immobilised components. Always trial practicals to confirm activity of organisms or enzymes.

Preparing immobilised enzymes/cells in alginate beads

• Make up a solution of the enzyme to be studied (see Recipe sheet 37 for enzymes), or a suspension of yeast cells using purified water.

• Sprinkle 2 g of sodium alginate in 100 ml of warm, purified water and mix using a mechanical stirrer. Allow the solution to cool. Initially the mixture will form glutinous lumps, but it becomes smooth over time.

• Dissolve 3 g of calcium chloride-6-water in 200 ml of purified water in a 250 ml beaker. • Mix 2 ml of the enzyme/suspension with 8 ml of the 2% sodium alginate solution. Variations on these

proportions may be used. • Draw this up into a 10 ml syringe. • Add the sodium alginate/enzyme (or cell) mixture one drop at a time to the calcium chloride solution

making sure the tip of the syringe is held above the solution in the beaker. • Allow the beads to harden for a few minutes before straining them out of the beaker.

Hardened alginate beads in calcium chloride solution

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4 Aluminium solutions

Hydrated aluminium salts such as the chloride (AlCl6.6H2O, M = 241.5 g mol-1), sulfate(VI) (Al2(SO4)3.16H2O, M = 630 g mol-1) and nitrate(V) (Al(NO3)3.9H2O, M = 375 g mol-1) absorb water (ie, they are hygroscopic) and become damp on storage. Do check these chemicals before use. Aluminium potassium sulfate(VI), also known as potassium aluminium sulfate, alum and potash alum is easily stored and suitable for all activities where aluminium ions are required for testing. However, it is not all that soluble in water, although it does make large octagonal crystals. Aluminium solutions are acidic.

Aluminium potassium sulfate(VI)

Formula: AlK(SO4)2.12H2O Molar mass: 474.39 g mol-1 Solubility: 11 g per 100 ml

General Hazards See Hazcard 2B. Aluminium salts in water are acidic. Never use anhydrous aluminium chloride to make solutions. It reacts violently with water producing toxic fumes of hydrogen chloride.

Mass (g) of solid to be used

Concentration required

Volume (ml) of solution required Hazard warning label 100 250 1000

0.01 M Ten-fold dilution of the 0.1 M solution -

0.1 M 4.74 11.86 47.44 -

0.2 M 9.49 23.72 94.88 -

Saturated (20 °C) 12 29 114 -

Aluminium chloride (hydrated)

Formula: AlCl3.6H2O Molar mass: 241.43 g mol-1 Solubility: 83 g per 100 ml

General Hazards See Hazcard 2A. Aluminium salts in water are acidic.





0.1 M 2.41 6.03 24.14 -

0.5 M 12.07 30.18 120.72 -

Saturated (20 °C) 90 225 900 IRRITANT

Preparing solutions of aluminium salts

• Wear eye protection. • Measure out the indicated quantity of the aluminium salt. • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

water to the required level. • Pour into a labelled bottle.

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5 Ammonia gas

Ammonia is less dense than air and very soluble in water so it has to be collected by the downward displacement of air (upward delivery). Theoretically, 2.1 ml of fresh concentrated (‘880’) ammonia solution produces 1 litre of gas, although this is never achieved in practice, so always use an excess. Older stocks of concentrated ammonia will be less concentrated. If the fountain experiment is to be carried out, a round bottom flask is substituted for the gas jar in the diagram below. Many standard text books use a calcium oxide drying tube. There is no real need for this. General Hazards See Hazcards 5 & 6. Ammonia begins to bubble off from ‘880’ ammonia at about

55 °C so heat gently and use anti-bumping granules to allow the ammonia to boil off gradually.

Preparing ammonia gas

• Use a fume cupboard. Wear goggles. • Add 10 ml of concentrated ammonia to the boiling tube

(CORROSIVE). • Set up the equipment as shown on the right. • Warm the boiling tube gently for about 5 minutes. • Knowing when the gas jar is full is not easy. Placing

moist red litmus at the neck of the inverted flask is not a good indicator that the flask is full of gas. It is a matter of judgement and experience. It might be better to use a fresh boiling tube of concentrated ammonia for each gas jar or flask required.

• When the collection is finished, place a cover slip or, better still, a large bung into the opening of the gas jar. Store the gas jar upside down because ammonia is lighter than air.

• The residual ammonia solution can be poured down the sink in a fume cupboard with plenty of water.

Heatgently

10 ml of concentratedammonia solution

Inverted gas jarheld by a clamp

Anti-bumping granules

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6 Ammonia solution (ammonium hydroxide)

The concentration of 35% (w/v) ammonia solution, (also known as ‘880’ ammonia), is 18.1 mol dm-3. Educational suppliers commonly supply this concentration although other concentrated solutions, eg, 25% (w/v) are available. If kept for long periods, the concentration of ammonia solutions decreases because of leakage of gas from the container. If the concentrated ammonia is several years old it would be wise to test the concentration of the solution before diluting further. It is sensible not to store diluted ammonia solutions for long periods. It is better to prepare diluted solutions only when required. Solutions less than 1 M should be made by further dilution of 1 M ammonia solution and are best made fresh before use. If you have other concentrations of concentrated ammonia, then contact CLEAPSS for more advice. Some of the values below are different from previous Recipe Cards as more precise information is now available. Formula: NH3 Molar mass: 17.03 g mol-1 Solubility: infinite General Hazards Concentrated ammonia; see Hazcards 5 & 6.

Take great care when opening bottles of concentrated ammonia on hot days.

Volume of ‘880’ ammonia to be used



0.1 M Ten-fold dilution of the 1 M solution -

0.5 M Two-fold dilution of 1 M solution -

1 M 28 55 138 -

2 M 55 111 276 -

5 M 138 276 691 IRRITANT

Preparing ammonia solutions

• Use a fume cupboard if concentrated ammonia or solutions more concentrated than 5 M ammonia are used. Wear goggles (a face shield is preferable when handling large volumes) and chemical resistant gloves.

• Measure the indicated quantity of ammonia solution in an appropriate measuring cylinder. • Add the liquid to about two thirds of the final volume of water in an appropriate beaker or laboratory

jug. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

water to the required level. • Pour into a labelled bottle and mix well. Add a hazard warning if appropriate.

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7 Ammonium chloride

Formula: NH4Cl Molar mass: 53.5 g mol-1 Solubility: 36 g per 100 ml General Hazards See Hazcard 9A.





0.5 M 2.68 6.69 26.75 -

1.0 M 5.34 13.38 53.50 -

Saturated (20 °C) 40 100 400 -

Preparing ammonium chloride solution

• Measure out the indicated quantity of ammonium chloride. • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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8 Ammonium vanadate(V) solution

Also known as ammonium polytrioxovanadate(V) and ammonium metavanadate. Ammonium vanadate solution is used to illustrate the various oxidising states exhibited by vanadium. Ammonium vanadate is not very soluble in pure water and has to be dissolved in alkali first, before making acidic for use. Formula: NH4VO3 Molar mass: 117 g mol-1 Solubility: 0.52 g per 100 ml General Hazards Ammonium vanadate(V) is TOXIC. See Hazcards 9B, 91 and 98A.

Preparing 100 ml of 0.1 M ammonium vanadate solution

• Wear goggles. • Dissolve 1.17 g of ammonium vanadate(V) in 20 ml of 2 M sodium hydroxide in a beaker. (The odour

of ammonia may be detected but it will cause no harm.) • Transfer the solution to a 100 ml measuring cylinder. • Add 1 M sulfuric(VI) acid to bring the total volume to 100 ml. • The yellow solution is irritant because of the presence of sulfuric(VI) acid.

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9 Azo dyes

It is a myth that all azo dyes are carcinogens. The dyes can be prepared in schools as long as sensible laboratory procedures are observed. On the whole, the more soluble a dye and the presence of sulfonic acid groups on the benzene ring, then the safer the dye (eg, Orange II and methyl orange). See Guide G195 for preparation of azo dyes from ethyl 4-aminobenzoate. The recipes below can be used safely in schools. General Hazards Sulfanilic acid (4-aminobenzenesulfonic acid), see Hazcard 4B. Sodium nitrate(III)

(sodium nitrite); see Hazcard 93. Naphthalen-2-ol; see Hazcard 70. N,N-dimethylphenylamine; see Hazcard 4B. Sodium hydroxide; see Hazcard 91. Ethanoic acid; see Hazcard 38A. Sodium carbonate; see Hazcard 95A. Methyl orange; see Hazcard 32.

Preparing azo dyes as described is a two-stage process.

Stage 1: Preparing the diazonium salt of sulfanilic acid

Solution A: • Add to a boiling tube, 0.6 g of sulfanilic acid,

0.2 g of anhydrous sodium carbonate and 10 ml of purified water.

• Warm the mixture to boiling and then cool under the tap. Add 0.3 g of sodium nitrate(III) and agitate the test tube until the salt dissolves.

Solution B: • In another boiling tube, cool down 8 ml of

1 M hydrochloric acid using an ice-water bath.

• Add solution A to solution B. The diazonium salt settles out. It is more stable then many others and will keep for some hours.

• Test for the presence of free nitrous acid with starch-iodide paper. If the paper does not turn blue, add a little more sodium nitrate(III). Swirl the contents of the boiling tube and divide the contents into two test tubes.

Stage 2 (a): Preparing Orange II (IRRITANT) and dying cotton with it

• Wear goggles. Wear disposable nitrile gloves. • In a test tube, dissolve 0.25 g of naphthalen-2-ol* in 4 ml of 1 M sodium hydroxide solution by

warming. • Cool the solution in an ice-water bath and pour it into a Petri dish. • Using forceps, add white cotton wool or cotton cloth (eg, bandage material) to the Petri dish to soak up

the solution. • Add the contents of the test tube containing the diazonium salt onto the cotton. The dye will appear. • Remove the cotton with forceps from the Petri dish and rinse it under cold water to remove any solid

dye. This is a soluble dye so do not do this for too long. Allow the cotton to dry. * Other phenols, benzene diols, cresols and naphthols can be used with the diazonium salt of sulfanilic acid to produce other dyes.

Stage 2 (b): Preparing methyl orange (TOXIC)

• Wear goggles. Wear disposable nitrile gloves. • In a test tube, add 0.2 ml of N,N-dimethylphenylamine to 0.2 ml of glacial ethanoic acid and agitate the

mixture well. • Add the contents of the test tube containing the diazonium salt to the test tube containing

N,N-dimethylphenylamine. Leave for 5 minutes for the red dye to form. • Add 2 ml of 2 M sodium hydroxide solution, heat to boiling and allow the solution to cool. • The solid orange sodium salt of methyl orange should form.

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10 Barium solutions

Purified water should be used to avoid cloudiness caused by the precipitation of barium sulfate(VI). Barium chloride is classified as TOXIC if swallowed and barium nitrate(V) as HARMFUL if swallowed. All solutions of barium nitrate(V) are LOW HAZARD. Barium chloride is more soluble in water than barium nitrate(V). General Hazards Barium chloride; see Hazcard 10A. Barium nitrate(V); see Hazcard 11.

Barium chloride

Formula: BaCl2.2H2O Molar mass: 244.26 g mol-1 Solubility: 26 g per 100 ml Mass (g) of solid to be used



0.01 M Ten-fold dilution of 0.1 M solution -

0.1 M 2.44 6.11 24.43 HARMFUL

0.5 M 12.21 30.53 122.13 HARMFUL

Saturated (20 °C) 36 90 260 TOXIC

Barium nitrate

Formula: Ba(NO3)2 Molar mass: 261.37 g mol-1 Solubility: 9 g per 100 ml Mass (g) of solid to be used




0.1 M 2.61 6.53 26.14 -

Saturated (20 °C) 9 45 90 -

Preparing barium salts solutions

• Wear eye protection. Wear disposable nitrile gloves when weighing and preparing the solution. • Measure out the indicated quantity of the solid barium salt. • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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11 Benedict’s qualitative reagent

Benedict’s solution or DNSA (see Recipe sheet 34) should be used in place of Fehling’s solution to test for reducing sugars because it is less hazardous. Glucose, lactose and maltose are reducing sugars and give a positive test. Sucrose is a non-reducing sugar and does not give a positive result. Benedict’s solution is less satisfactory in testing for non-reducing sugars and aldehydes in organic chemistry in which case Fehling’s or Sandell’s solutions will be needed. No hazard warning symbol is required on the bottle as the concentrations of each of the constituents are low. This solution is not suitable for colorimetric work. See Recipe sheet 12 for Benedict’s quantitative reagent or Recipe sheet 34 for DNSA. To differentiate between reducing sugars, enzyme tests are required. General Hazards Sodium carbonate; see Hazcard 95A. Copper sulfate(VI); see Hazcard 27C.

Procedure to produce 1 litre of solution

• Wear eye protection. • Measure out 170 g of trisodium citrate-2-water and 100 g of anhydrous sodium carbonate (or 256 g of

hydrated sodium carbonate). • Add the solids to about 850 ml of purified water in a 1 litre beaker. • Stir to dissolve, warming if necessary. • Add 17.4 g of copper(II) sulfate(VI)-5-water and stir to dissolve. • Pour the solution into an appropriate measuring cylinder and dilute to the final volume. • Filter if necessary. • Pour into a labelled bottle and mix well.

Procedure for carrying out the test

• The material under test is mixed with about 1 ml of water in a test tube or vial, and about 3 ml of Benedict's reagent is added.

• Place the test tube in a boiling water bath for about 5 minutes. • The colour should progress from blue (with no glucose present) to green, yellow, orange, red, and

then brick red or brown as glucose concentration increases.

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12 Benedict’s quantitative reagent

The procedure detects the loss of blue colour as the sugar reduces the copper(II) ions to copper(I). It can be used either in volumetric or colorimetric methods. The addition of thiocyanate forms a complex and prevents the precipitation of copper(I) oxide. See also Recipe sheet 34 for DNSA which is an alternative test for reducing sugars. General Hazards Sodium carbonate; see Hazcard 95A. Copper sulfate(VI); see Hazcard 27C.

Potassium thiocyanate; Hazcard 95C. Potassium hexacyanoferrate(II) see Hazcard 79. The volumetric method uses hot liquids.

Preparing 1 litre of Benedict’s quantitative solution (BQS)

• Wear eye protection. • Measure out 75 g of anhydrous (or 160 g of hydrated) sodium carbonate, 200 g of trisodium citrate-2-

water and 125 g of potassium thiocyanate. Add these solids to 700 ml of boiled distilled water in a 1 litre beaker. Stir to dissolve, reheating if necessary.

• Measure out 18 g of copper(II) sulfate(VI), dissolve in 100 ml of purified water and add this, with constant stirring, to the solution made in step 1.

• Add 0.25 g of potassium hexacyanoferrate(II) and pour the solution into a suitable volumetric flask. • Dilute to the final volume with purified water and mix well. For the colorimetric procedure below, dilute

35 ml of this solution to 100 ml with water. (No ‘hazard warning’ label is required at these concentrations.)

Using the solution: volumetric procedure

• Place a 0.3% solution of glucose in a burette. • Place 10 ml of BQS into a 100 ml conical flask, heat to boiling and add about 2 g of anhydrous sodium

carbonate. • Add the glucose solution in 0.5 ml amounts, boiling each time until the blue or green colour just

disappears. Record the total volume of glucose added. When nearing the endpoint it is advisable to boil for 30 seconds to avoid overshooting.

• Repeat the procedure to confirm your result. • To measure the concentration of glucose in an unknown solution, repeat the procedure with glucose

solutions of unknown concentration. • By comparing the volume obtained on titration of the known and unknown samples, the %

concentration of the unknown can be calculated. % 0.3 x

A = titration volume for known glucose solution B = titration volume for unknown glucose solution (Some methods suggest adding a few drops of 1% aqueous methylene blue solution to act as an indicator. It should be added when enough glucose solution has been added to turn the original solution a very pale blue colour.)

Using the solution: colorimetric procedure

• Place a 1% solution of glucose in a burette. • Place water in another burette and prepare a series of glucose solutions of varying concentration from

0 to 1%. • In a series of labelled fresh test tubes, add 1 ml of each solution prepared in step 2, to 10 ml of BQS.

Place these in a boiling water bath for a few minutes. • Allow the solutions to cool and any precipitates to settle. • Use a red or yellow filter in the colorimeter. Obtain the absorbance of light through each solution. • Use these readings to construct a glucose concentration calibration curve. • Use the calibration curve to identify unknown concentrations of glucose solution.

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13 Biochemical indicators and tests

Amino acid, polypeptide and protein tests

See also Recipe Sheet 15 for the Biuret test.

Cole’s modification of Millon’s reagent

Test for soluble proteins

Wear goggles and gloves. Dissolve 10 g of mercury(II) sulfate(VI) in 100 ml of 2 M sulfuric(VI) acid (solution A). Separately, dissolve 1 g of sodium nitrate(III) (nitrite) in 100 ml of water (solution B). Before use, mix two volumes of solution A with one of solution B. A reddish-brown colouration or precipitate indicates the presence of soluble proteins.

See Hazcards 62, 93, 98A. Solution A and the combined solution are TOXIC.

Marquis’s reagent

Test for alkaloids

Wear goggles and gloves. Use a fume cupboard. Add 2 drops of 40% methanal solution to 3 ml of concentrated sulfuric(VI) acid just before use. This is a spot test and various colours appear. See http://en.wikipedia.org/wiki/Marquis_reagent.

See Hazcards 63 and 98. Label the solution CORROSIVE and TOXIC.

Ninhydrin For amine groups

Wear eye protection. Dissolve 0.1 g of ninhydrin in 100 ml of water. A purple colour, known as Ruhemann's purple, is produced when ninhydrin reacts with primary and secondary amines, indicating the presence of amino acids. Can be used as a spray or a dip, but needs to be heated, in an oven at 110 °C or with a hairdryer, for the colours to appear.

See Hazcard 66. No hazard warning is required on the solution.

Sakaguchi test

Test for proteins containing arginine

Wear goggles. Dissolve 5 g of sodium hydroxide in 100 ml of water (solution A). Dissolve 1 g of napthalen-1-ol in 100 ml of water (solution B). For the test, one drop of sodium chlorate(I) solution (10-14% available chlorine, see Recipe sheet 81) is required as well. Proteins containing arginine appear an intense red colour.

See Hazcards 70, 89 & 91.

Carbohydrate tests

See also 3,5-dinitrosalacylic acid (DNSA), Benedict’s solution, Sandell’s solution and Fehling’s solution.

Molisch’s solution

For all carbo-hydrates

Wear goggles. Dissolve 5 g of napthalen-1-ol in 100 ml of ethanol. The solution containing a possible carbohydrate is combined with a small amount of Molisch's reagent in a test tube. After mixing, a small amount of concentrated sulfuric(VI) acid is slowly added down the sides of the sloping test tube, without mixing, to form a lower layer. Look for a purple ring at the interface of the two layers.

See Hazcards 40A, 70 & 98. Label the solution HIGHLY

FLAMMABLE & HARMFUL.

Periodic acid Schiff (PAS) reaction

For poly-saccharides

Wear goggles and chemical-resistant gloves. Dissolve 1 g of iodic(VII) acid (periodic acid) in 100 ml of water. Used in conjunction with Schiff’s reagent. Changes from colourless to purple.

Iodic(VII) acid is CORROSIVE & OXIDISING. No hazard warning is required on the solution.

http://en.wikipedia.org/wiki/Marquis_reagent�

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Cellular respiration

Janus green B

Wear eye protection and disposable nitrile gloves when making up the solution. Dissolve 0.3 g of the dye in 100 ml of purified water. Dilute this solution ten times with water before use. Colour changes are from blue to salmon pink. See Guidance leaflet PS88 for more details including a practical activity.

No hazard label is required on the solution.

Methylene blue

Wear eye protection, and gloves to avoid staining the skin. Dissolve 1 g solid in 100 ml water and add 0.6 g sodium chloride. The blue indicator turns colourless as the dye is reduced.

See Hazcards 32 and 40. No hazard label is required on the solution.

TTC Dissolve 1 g of 2,3,5-triphenyl tetrazolium chloride (TTC) in 100 ml of water. (A 0.5% solution is less expensive and gives just as good results but takes longer. It works well with maize seedlings.) Turns red as the dye is reduced.

Low hazard.

Tasting and genetics studies Phenylthio-carbamide

This is also called PTC, phenylthiourea or PTU. Dissolve 0.05 g in 100 ml of water. Filter paper is soaked in the solution and then hung up to dry before cutting into strips. See Handbook 15.13 for details on tasting investigations.

The solid is VERY TOXIC; see Hazcard 35. The solution is low hazard.

Vitamin C

DCPIP solution

Also called 2,6-dichlorophenol indophenol, and phenol-indodichlorophenol. Dissolve 0.1 g of dye in 100 ml of water. The standard vitamin C solution should also be 0.1% (w/v). Add the blue dye until the colour does not disappear.

Both are low hazard chemicals.

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14 Bismuth nitrate(V) solution

The solutions need to be kept acidic to avoid the formation of insoluble basic salts. Formula: Bi(NO3)3.5H2O Molar mass: 395 g mol-1

General Hazards See Hazcards 67 & 73B.

Preparing 100 ml of 0.1 M of bismuth nitrate(V) solution

• Wear goggles. • Dissolve 5.34 g of bismuth(III) nitrate(V)–5-water in 70 ml of 1 M nitric(V) acid. • Transfer the solution to a 100 ml measuring cylinder. • Make up to 100 ml with 1 M nitric(V) acid. • Label the solution CORROSIVE.

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15 Biuret reagent

Biuret reagent tests for proteins. Previous methods and recipes for this solution have used more concentrated solutions than that described below. This meant that Y7 and 8 pupils were precluded from doing this experiment according to the advice on Hazcard 91 (sodium hydroxide). However, the procedure below uses 0.1 M sodium hydroxide and 0.01 M copper(II) sulfate(VI) solutions and teachers might find that further dilution is possible. It may be necessary to make acidic samples alkaline before the test for proteins is carried out. Coomassie blue can also be used to test for proteins. It is available in a test kit with instructions for use. It’s advantages are that it does not require heating and is very sensitive.

Qualitative Biuret Reagent This does not keep so only prepare what is required. General Hazards Sodium hydroxide (solid) and 2 M solution. See Hazcards 91. Copper sulphate, see

Hazcard 27C.

Preparing 1 litre of Qualitative Biuret reagent

• Wear goggles. • Weigh out 0.75 g of copper(II) sulfate(VI)-5 -water. • Prepare 1 litre of 2 M potassium or sodium hydroxide solution. • Dissolve the copper(II) sulfate(VI) in the alkali and label the solution CORROSIVE.

Procedure and use of Biuret solution suitable for Y7 and 8

• Wear suitable eye protection. • Prepare 0.01 M copper(II) sulfate(VI) solution (Recipe Sheet 31). • Prepare 0.1 M sodium hydroxide solution (Recipe Sheet 85). • Place the sample of a liquid to be tested in a test tube to a depth of 10 mm. • Add the same volume of 0.1 M sodium hydroxide solution and agitate the test tube. • Add a few drops of the 0.01 M copper(II) sulfate(VI) solution and agitate the test tube. • A purple or pink colouration indicates the presence of protein.

Quantitative Biuret Reagent This does keep. General Hazards Sodium hydroxide (solid) and a 2 M solution see Hazcard 91. Copper(II) sulfate(VI)

see Hazcard 27C.

• Wear goggles. • Dissolve 1.5 g of copper(II) sulfate(VI)-5-water crystals and 6 g of potassium sodium tartrate-4-water in

500 g of purified water. • Add 375 ml of 2 M sodium hydroxide with stirring. • If a precipitate occurs, add 1 g of potassium iodide. • Pour this mixture into a 1000 ml volumetric flask and dilute to 1 litre. Mix well. Label this solution

CORROSIVE as it is a 0.75 M sodium hydroxide solution.

For quantitative analysis, a series of standards can be produced with solutions of varying % dilutions of a protein such as albumen, using a colorimeter with a 540 nm (green) filter. Unknown proteins solutions can then be compared against these standards.

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16 Brodie’s fluid

Also known as manometer or manometric fluid. Water strongly adheres to glass so requires an emulsifier to avoid water droplets ‘sticking’ to the walls of the manometer and affecting the readings, and to reduce capillary action in small bore manometers. General Hazards Sodium azide is VERY TOXIC and contact with acids liberates a very toxic gas. (See

Hazcard 95B). For eosin, see Hazcard 32. Once in solution, the solution is LOW

HAZARD. Sodium azide may be omitted but the fluid will not keep as well and there will be mould growth.

A. Preparing 1 litre of Brodie’s fluid

• Wear eye protection. • Weigh out 46 g of sodium chloride, 10 g of sodium tauroglycocholate (bile salts), 0.2 g sodium azide

(optional) and 0.5 g of eosin. • Make up to 1 litre with distilled or deionised water.

B. Brodie’s fluid substitute for short term use

• 44 g of sodium bromide. • 1 g of liquid detergent (eg washing-up liquid). • 0.3 g of Evans blue.

C. Simple version for short-term use

• Use water with a food dye such as cochineal and a few drops of detergent.

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17 Bromine water

The solubility of bromine in water is 4 g, (ie, 1.25 cm3 ) in 100 g of water at room temperature. This would be a 4% (w/v) solution. And its concentration would be 0.25 M. Solutions equal to or greater than 0.06 mol dm-3 (ie, a 0.3% v/v solution) are TOXIC. Solutions equal to or more concentrated than 0.006 mol dm-3 (0.1% w/v or 0.03% v/v) but more dilute than 0.06 mol dm-3 should be labelled HARMFUL. Aqueous solutions of bromine should be prepared just before use. If stored for long periods, especially though the summer, the solution becomes paler as bromine vapour is lost. Do not make this solution for the first time without seeking practical advice from a more-experienced colleague. General Hazards Bromine is VERY TOXIC and CORROSIVE (see Hazcard 15A). Hazcard 15B deals with

bromine water. 0.025 M bromine water has a considerable vapour of bromine gas above it. It should be dispensed from a fume cupboard. More-dilute solutions can be used in a well-ventilated room but staff should discourage any direct inhalation of the vapour. Sodium chlorate(I) is CORROSIVE (see Hazcard 89), 2 M hydrochloric acid; see Hazcard 47A, Potassium bromate(V) is TOXIC, (see Hazcard 80).

Preparing 1 litre of 0.02 M solution (HARMFUL) of bromine water

Method 1 • Use a fume cupboard. Wear goggles or a face shield and chemical-resistant gloves. • Using a disposable plastic pipette, add 1 ml of bromine to 500 ml of water in a 1 litre beaker. • Add a stirrer bar and place on a magnetic stirrer to dissolve the bromine. This can take over

20 minutes. • Dilute to 1 litre with water. • Alternatively, an ampoule containing 1 ml of bromine can be crushed under 500 ml of water, stirred

using a magnetic stirrer until it dissolves and made up to 1 litre with water. The solution must be decanted so that no broken glass is present.

Method 2

• Use a fume cupboard. Wear goggles or a face shield. Consider wearing gloves. • Dissolve 4.76 g of potassium bromide in 76 ml of water, add 14 ml of 10% (available chlorine) sodium

chlorate(I) solution (CORROSIVE) and 10 ml of 2 M hydrochloric acid (IRRITANT). Dilute to 1 litre with water.

Method 3 This reaction is slow and it is better to leave the solution 24 hours before it is used.

• Use a fume cupboard. Wear eye protection. • Add 1.12 g of potassium bromate(V) (TOXIC), 12 g of potassium bromide and 14 ml of 2 mol dm-3

hydrochloric acid (IRRITANT) into a 1 litre jug or measuring cylinder. • Add water to 1 litre.

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18 Buffer solutions

Buffer solutions retain their pH on addition of small amounts of acid, alkali or on dilution. Unless stated otherwise, buffers are low hazard but eye protection should be worn during preparation. The pH value of a buffer is slightly altered by temperature but this is not usually significant. The values used in the Recipe Sheet are for 20 °C. A definition for pH suitable for schools is minus the logarithm (to the base 10, ie log10) of the hydrogen ion concentration in an aqueous solution’. For a more advanced explanation, consult http://en.wikipedia.org/wiki/PH. There is also a pOH scale and pH + pOH =14. A pH meter should be calibrated with standard buffers before it is used. These may be prepared from tablets or commercial solutions. Use distilled or deionised water to make up buffers. Single-component buffers are very quick to make up but the values may not always be convenient. The majority of buffers involve two components mixed in certain proportions. For accurate work, it is wise to check their pH with a calibrated pH meter. A universal buffer mixture can be used to obtain an array of buffer solutions from pH 2 to 11. The stock solution can, usefully, be stored for several months. Buffers (especially those between pH 4 and pH 7) do not keep well. Moulds develop which affect pH readings and block the junctions on pH probes. Commercial buffer solutions contain a mould inhibitor which allows longer storage. To save time buffers can be stored as pre-weighed dry components or as frozen solutions. Before use, make sure frozen solutions are returned to room temperature, that all components are fully dissolved, and check the pH. Special biological buffers (eg, TRIS) are available that do not use phosphate(V) or ethanoate ions which might interfere with some biochemical processes. They are named Good buffers after their developer, Norman Good. Different enzymes are inhibited by different reagents, so check the protocol and choose the correct buffer system.

Single component buffers

pH 1.7 buffer

Dissolve 1.27 g of potassium hydrogen-ethanoate-1-ethanedioic acid-2-water (potassium tetroxalate) in 10 ml of hot water and make up to 100 ml with cold water.

Low hazard See Hazcard 36A.

pH 4 buffer Dissolve 1.01 g of potassium hydrogenphthalate in 10 ml of hot water and make up to 100 ml with cold water. The solution does not keep well because of mould growth.

Low hazard See Hazcard 13B.

pH 7 buffer Dissolve 0.77 g of ammonium ethanoate in 100 ml of cold water. The solution does not keep well because of mould growth.

Low hazard See Hazcard 9B.

pH 9.2 buffer

Dissolve 0.38 g of sodium tetraborate-10-water in 100 ml of water. Low hazard See Hazcard 14.

pH 12.6 buffer

Use saturated limewater solution. Low hazard but wear eye protection. See Hazcard 18.

Temperature effects on single component buffers

Temperature / °C 5 10 15 20 25 30 40 50 60 70 80 90

pH 4 buffer 4.01 4 4 4 4.01 4.01 4.03 4.06 4.09 4.12 4.16 4.2

pH 9.2 buffer 9.39 9.33 9.27 9.23 9.18 9.14 9.07 9.02 8.97 8.93 8.89 8.85

http://en.wikipedia.org/wiki/PH�

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Buffers for pH probe calibration

pH 4 buffer Dissolve 1.012 g of potassium hydrogenphthalate in 10 ml of hot water and make up to 100 ml with cold water. The solution does not keep well because of mould growth.

See Hazcard 13B

pH7 buffer 0.339 g of citric acid and 4.306 disodium hydrogenphosphate(V)-12-water (or 1.707 g of anhydrous salt) dissolved in water and made up to 100 ml with water.

See Hazcard 9B.

pH10 buffer 0.477 g sodium tetraborate-10-water and 18.3 ml of 0.1 M sodium hydroxide (from a burette) is made up to 100 ml with water.

See Hazcards 14 and 91.

NB Although masses are given to 3 decimal places, it would be acceptable to work to 2 decimal places.

Double component buffers

To prepare these buffers, make up the individual components, then mix the quantities given for the required pH. For accurate work, confirm the values with a calibrated pH meter and probe.

Using a solid and a solution.

General Hazards As 0.1 M sodium hydroxide solution is IRRITANT, wear eye protection when preparing the buffers. All the resulting buffers are low hazard. See Hazcards 13B, 36C and 91.

3 10.21 g potassium hydrogen phthalate and 223 ml of 0.1 M hydrochloric acid

Dilute each mixture to 1

litre solution with distilled or

deionised water.

4 10.21 g potassium hydrogen phthalate

5 10.21 g potassium hydrogen phthalate and 226 ml of freshly-made 0.1 M sodium hydroxide solution

6 6.81 g potassium dihydrogen phosphate and 56 ml of freshly-made 0.1 M sodium hydroxide solution

7 6.81 g potassium dihydrogen phosphate and 291 ml of 0.1 M sodium hydroxide solution

8 6.81 g potassium dihydrogen phosphate and 467 ml of freshly-made 0.1 M sodium hydroxide solution

9 4.77 g sodium tetraborate-10-water and 46 ml of 0.1 M hydrochloric acid

10 4.77 g sodium tetraborate-10-water and 183 ml of freshly-made 0.1 M sodium hydroxide solution

11 2.1 g sodium bicarbonate and 227 ml of freshly-made 0.1 M sodium hydroxide solution

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Mixing two prepared solutions Consult the relevant Recipe Sheets for the preparation of the solutions.

General Hazards

All these buffers are low hazard. See relevant Hazcards for more information.

The citric acid/disodium hydrogenphosphate(V) buffer system

A is 0.2 M disodium hydrogenphosphate(V) and B is 0.1 M citric acid.

pH A (ml) B (ml) pH A (ml) B (ml) pH A (ml) B (ml)

2.2 2.00 98.00 4.2 41.40 58.60 6.2 66.10 33.90

2.4 6.20 93.80 4.4 44.10 55.90 6.4 69.25 30.75

2.6 10.90 89.10 4.6 46.75 53.25 6.6 72.75 27.25

2.8 15.85 84.15 4.8 49.30 50.70 6.8 77.25 22.75

3.0 20.55 79.45 5.0 51.50 48.50 7.0 82.35 17.65

3.2 24.70 75.30 5.2 53.60 46.40 7.2 86.95 13.05

3.4 28.50 71.50 5.4 55.75 44.25 7.4 90.85 9.15

3.6 32.20 67.80 5.6 58.00 42.00 7.6 93.65 6.35

3.8 35.50 64.50 5.8 60.45 39.55 7.8 95.75 4.25

4.0 38.55 61.45 6.0 63.15 36.85 8.0 97.25 2.75

The ethanoic acid/sodium ethanoate buffer system

The ammonia/ammonium chloride buffer system

Add the stated volume of A (0.2 M sodium ethanoate solution) to the stated volume B

(0.2 M ethanoic acid)

Add the stated volume of A (0.2 M ammonia) to the stated volume of B (0.2 M ammonium

chloride solution)

pH A (ml) B (ml) pH A (ml) B (ml)

3.8 12.0 88.0 8.4 12.5 87.5

4.0 18.0 82.0 8.6 18.5 81.5

4.2 26.5 73.5 8.8 26.0 74.0

4.4 37.0 63.0 9.0 36.0 64.0

4.6 49.0 51.0 9.25 50.0 50.0

4.8 60.0 40.0 9.4 58.5 41.5

5.0 70.5 29.5 9.6 69.0 31.0

5.2 79.0 21.0 9.8 78.0 22.0

5.4 85.5 14.05 10.0 85.0 15.0

The sodium hydrogencarbonate/sodium hydroxide) system buffer system

The phosphate(V) buffer system. (potassium salts may also be used)

Add the stated volume of A (0.1 M sodium hydroxide) to 50 ml of 0.05 M sodium

hydrogencarbonate and dilute to 100 ml (Wear eye protection.)

Add the stated volume of A [0.2 M sodium dihydrogenphosphate(V)] to the stated

volume of B [0.2 M disodium hydrogenphosphate(V)]

pH A (ml) pH A (ml) B (ml) 9.6 5.00 6.0 87.7 12.3 9.8 6.20 6.5 68.5 31.5 10.0 10.70 7.0 39.0 61.0 10.2 13.80 7.5 16.0 84.0 10.4 16.50 8.0 5.3 94.7 10.6 19.10 10.8 21.20 11.0 22.70

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Universal Buffer solutions

The Universal Buffer: Recipe 1

• Wear eye protection. • To prepare the stock solution, add 9.9 g of disodium hydrogenphosphate-12-water, (or 4.6 g disodium

hydrogenphosphate-2-water) (see Hazcard 72), 7.0 g of citric acid (see Hazcard 9) and 3.5 g of boric acid (see Hazcard 14) to 250 ml of 1 M sodium hydroxide solution (see Hazcard 91) and dilute to 1000 ml with distilled/deionised water. This solution keeps well. Label the solution IRRITANT.

To make any buffer between 3.5 and 10

• Wear eye protection. • Place 0.1 M hydrochloric acid in a burette. • Place 20 ml of the stock solution in a 250 ml beaker on a magnetic stirrer. Clamp a pH probe into the

solution. • Add the acid to the solution in the beaker with constant stirring until the required pH is obtained. If too

much acid is added, use a little more stock solution to increase the pH value. • Add distilled/deionised water to make the final solution 100 ml.

The Universal Buffer: Recipe 2

• Wear eye protection. • To 500 ml of water, add 2 ml of concentrated ethanoic acid (see Hazcard 38A), 3 ml of 85%

phosphoric(V) acid (see Hazcard 72), and 2.4 g of boric acid (see Hazcard 14), and make the solution up to one litre with distilled/deionised water.

• Prepare 0.2 M sodium hydroxide solution (see Hazcard 91), and label the solution IRRITANT.

To make any buffer between 2 and 11

• Wear eye protection. • Put 0.2 M sodium hydroxide solution in a burette. • Place 100 ml of the acid solution in a 400 ml beaker on a magnetic stirrer. Clamp a calibrated pH

probe into the solution. • Add 0.2 M sodium hydroxide to the acid solution until the required pH is obtained.

Buffer solutions: Biological Two component buffers are often used for biological systems. However, if an enzyme is affected by one, or more, of the component ions, eg, phosphate(V), ethanoate or ammonium ions, specialist buffers are required. The cheapest specialist biological buffer is tris(hydroxymethylamino)methane (IRRITANT) known as TRIS. The solutions made up from the data below are low hazard.

TRIS buffers

pH Volume of 0.1 M TRIS solution

Volume of 0.1 M hydrochloric acid

7.0 100 93.2 7.5 100 80.6 8.0 100 58.4 8.5 100 29.4 9.0 100 11.4

A full range of biological buffers can be found at www.sigmaaldrich.com/life-science/core-bioreagebnts/bioogical-buffers.html. For products and applications such as extraction of materials or electrophoresis, see protocols at www.ncbe.reading.ac.uk.

http://www.sigmaaldrich.com/life-science/core-bioreagebnts/bioogical-buffers.html�

http://www.ncbe.reading.ac.uk/�

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19 Calcium chloride and nitrate(V) solutions

Hydrated calcium chloride and nitrate(V) absorb water from the atmosphere. On occasions, the solid completely dissolves to leave a clear solution. Distilled or deionised water should be used to make solutions. In hard water areas, solutions may be cloudy if tap water is used. Do not use anhydrous calcium chloride to make solutions. General Hazards See Hazcard 19A

Formula: CaCl2.6H2O Molar mass: 219.08 g mol-1 Solubility: 74 g per 100 ml

Preparing 100 ml of 0.1 M of calcium chloride solution

• Wear eye protection. • Dissolve 2.19 g of calcium chloride-6-water in 70 ml of water. • Transfer the solution to a 100 ml measuring cylinder. • Make up to 100 ml with water. • The solution is low hazard.

Formula: Ca(NO3)2.4H2O Molar mass: 236.15 g mol-1 Solubility: 121 g per 100 ml

Preparing 100 ml of 0.1 M of calcium nitrate(V) solution

• Wear eye protection. • Dissolve 2.36 g of calcium nitrate-4-water in 70 ml of water. • Transfer the solution to a 100 ml measuring cylinder. • Make up to 100 ml with water. • The solution is low hazard.

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20 Calcium hydroxide solution (Limewater)

Saturated calcium hydroxide solution, commonly called lime water, is a 0.02 M aqueous solution of calcium hydroxide with a pH of 12.4. It does not keep for long periods in large bottles because it reacts with carbon dioxide from the atmosphere. It would be wise to start afresh each year. Lime water, in small bottles designed for class use, will quickly cease to work. Class sets need to be tested before handing out to the class. 1 M hydrochloric acid can be used to clean bottles that previously contained limewater.

Formula: Ca(OH)2 Molar mass: 74.09 g mol-1 Solubility: 0.15 g per 100 ml General Hazards Calcium hydroxide solid; see Hazcard 18. Splashed droplets of limewater in the eye

have caused quite severe irritation, so the solution should also be labelled and treated as an IRRITANT even though strictly its dilution is such that it is not formally classed as hazardous.

Preparing make 2.5 L of lime water in a bottle

• Wear eye protection. • Place 5 g of calcium hydroxide in a 600 ml beaker and half-fill it with water. • Stir the suspension and pour it via a funnel into a 2.5 litre bottle. • Fill the bottle with water, stopper and shake it. • Leave the bottle overnight for the suspension to settle. • When required, decant the limewater solution, slowly without agitating the sediment, into smaller

bottles for use in lessons. • Add more water and/or calcium hydroxide suspension when the level becomes low.

Preparing a large continual supply

• If a large stock is required, keep an excess of calcium hydroxide in an aspirator protected by a soda-lime tube, as shown below, and top up with distilled water as necessary.

• Use about 100 g calcium hydroxide for a 10 litre aspirator. It might take a week to fully settle.

Absorption tubefilled with soda lime

Calcium hydroxide

Limewater

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21 Carbon dioxide

Carbon dioxide is prepared by the action of dilute hydrochloric or nitric acid on calcium carbonate, usually as marble chips. If powdered calcium carbonate is used, the rate of gas production may be too rapid to be easily controlled. The gas is collected over water or by downward delivery. (Downward delivery uses a tube into the bottom of an upright gas jar. It is also known as upward displacement of air and relies on the fact that carbon dioxide is more dense than air.) Although carbon dioxide is slightly soluble in water (at room temperature, the solubility of carbon dioxide is about 6.4 cm3 of carbon dioxide per 100 ml of water), the rate at which it dissolves is slow. However, if necessary, the gas can be collected over warm water in which it is less soluble. Collection by downward delivery is quicker but it is difficult to ascertain exactly when the gas jar is full. However, downward delivery is necessary for burning magnesium in carbon dioxide, in order to avoid water interfering with the reaction. Soda water is a saturated solution of carbon dioxide in water (carbonic acid). General Hazards See Hazcards 20 & 47A.

• Wear eye protection. • In the 250 ml Büchner flask, add several lumps of marble chips. • Add enough water to immerse the base of the thistle funnel tube. • Set up the apparatus as shown in the diagram. • The inverted measuring cylinder should be full with water. • Now add 5 ml of 5 M hydrochloric acid (IRRITANT) down the thistle funnel. • Once the measuring cylinder is full of gas, it indicates the apparatus has been completely purged.

Remove it and replace with an inverted gas jar full of warm water. • Keep collecting gas jars of gas. When full of gas, either slide a gas jar cover in place or put a large

bung into the gas jar. • If the rate of gas production slows down but more gas is needed, add further 5 ml portions of 5 M

hydrochloric acid.

Add 5 mol dm hydrochloric acid-3

250 ml measuring cylinder

Warm water

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22 Cerium(IV) solutions

Cerium(IV) compounds have a variable water content, which makes preparing accurate concentration solutions very difficult. This is why the mass given in the recipe is approximate.

Cerium(IV) solutions, which have an intense yellow colour, are used in redox titrations.

Cerium(IV) solutions are more stable in solution than potassium manganate(VII) solutions.

Cerium(IV) solutions need to be prepared in dilute sulfuric(VI) acid. [Hydrochloric acid must be avoided as cerium(IV) ions oxidise the chloride ions slowly to chlorine.]

Cerium(IV) solutions should be standardised against sodium ethanedioate solution before use.

General Hazards The solids are irritating to the eyes, respiratory system and skin.

Preparing 1 litre of 0.1 M cerium(IV) ions

• Wear eye protection. • Measure out about 64 g of ammonium cerium(IV) sulfate(VI)-2-water or 40 g of cerium(IV) sulfate(VI)-

4-water. • Add the solid to about 500 ml of 1 M sulfuric(VI) acid in a beaker. • Stir to dissolve, warming if necessary. • Pour the solution into a 1-litre volumetric flask and make it up to the mark with water. • Pour into a labelled bottle. • The solution is low hazard.

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23 Chemiluminesence reactions

Luminol (3-aminobenzene-1,2-dicarboxylic hydrazide or 3-aminophthalhydrazide) in alkaline solution is oxidised to the 4-aminobenzene-1,2-dicarboxylate ion with the evolution of oxygen. Goggles should be worn when preparing the solutions if using solid sodium hydroxide but spectacles are suitable for the demonstrations using the solutions. Recipe D is ideal for a chemiluminescent fountain demonstration using ammonia (see picture below).

Four recipes illustrating chemiluminesence reactions

A • Solution A: Dissolve 0.4 g of luminol and 4 g of potassium hydroxide or sodium hydroxide in 1000 ml of water. Label the container IRRITANT.

• Solution B: Dilute 50 ml of fresh (10-14% available chlorine) sodium chlorate(I) solution to 1000 ml. Label the container IRRITANT.

• In a darkened room, mix equal volumes of solutions A & B together. The addition of a pellet of potassium hydroxide or sodium hydroxide may produce more light.

See Hazcards 4B, 89, 91. Wear goggles to prepare the solution.

B • Dissolve 0.2 g of luminol and 1 g of potassium hydroxide or sodium hydroxide in 1000 ml of water. To a known volume of this solution add an equal volume of 20 vol hydrogen peroxide solution.

• In a darkened room, sprinkle a few crystals of potassium hexacyanoferrate(III) onto the surface of the solution and observe the trails of light.

• The addition of a pellet of potassium hydroxide or sodium hydroxide may produce more light.

See Hazcards 4B, 50, 79, 91. Wear goggles to prepare the solution.

C • Solution A: Dissolve 0.2 g luminol, 4 g of anhydrous sodium carbonate, 24 g of sodium hydrogencarbonate, 0.5 g of ammonium carbonate and 0.4 g of hydrated copper(II) sulfate(VI) in 1000 ml water.

• Solution B: Prepare 1000 ml of 5 vol hydrogen peroxide solution. Mix equal volumes of A and B in a dark room or in a box with spy holes.

See Hazcards 4B, 27C, 50, 95A.

D • Solution A: Mix together 0.2 g luminol, 11 g of anhydrous sodium carbonate, 8 g of sodium hydrogencarbonate, 0.5 g of ammonium carbonate and 0.4 g of copper(II) sulfate(VI) in 1000 ml water. Add 25 ml of 10 vol hydrogen peroxide.

• Solution B: Dissolve 0.1 g of cobalt(II) chloride and 0.1 g of sodium nitrate(III) (ie, sodium nitrite) in 1000 ml of water.

• Mix equal volumes of A and B in a dark room or in a box with spy holes.

See Hazcards 4B, 9A, 25, 27C, 50, 89, 93, 95A.

The chemiluminescent fountain demonstration

The flask is originally filled with ammonia. Two containers labelled A and B which contain 1 litre each of solution A & B in Recipe D are used to supply the solution in the fountain. The solutions are connected via a T-piece into a single tube which is inserted into the flask.

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24 Chlorine gas

Version 1: Concentrated hydrochloric acid on potassium manganate(VII) The reaction of concentrated hydrochloric acid on potassium manganate(VII) is the more common and reliable of the two methods. However it uses more-hazardous starting chemicals. It produces chlorine quickly. The glassware is difficult to clean at the end and more-concentrated hydrochloric acid is required to remove manganese(IV) oxide stains. Do not make this solution for the first time without seeking practical advice from a more-experienced colleague. General Hazards See Hazcards 22A, 20 & 81. Double check that you are using concentrated

hydrochloric acid and not concentrated sulfuric(VI) acid. Explosions have resulted from using the wrong acid.

• Use a fume cupboard. Make sure it is working. Wear goggles. • Set up the equipment as in the illustration below. Make sure that the thistle funnel tube nearly reaches

the base of the flask. • Place at least 5 g of potassium manganate(VII) (OXIDISING & HARMFUL) in the Büchner flask. • Pour the concentrated hydrochloric acid (CORROSIVE) down the thistle funnel. • Collect the gas in a gas jar. When the contents of the gas jar are clearly green, assume the gas jar is

full. Or (optional), plug the gas jar with a large wad of mineral wool, or cotton wool, and put some damp blue litmus paper on top. It takes time for the chlorine to diffuse through the wool and bleach the litmus. When the bleaching is complete the jar will be full. Remove it and replace the mineral wool with a gas jar cover or a large bung.

• More gas jars of gas can be collected. • To dispose of the reaction mixture, pour it down the fume cupboard sink with lots of water. The stained

glass may need treating with a little concentrated hydrochloric acid before washing further.

Mineral wool

Moist blue litmus

At least 5 g of Potassium Manganate(VII)

Wooden block

Gas jar

Delivery tube as fardown as possible

In a fume cupboard, pour concentrated hydrochloric acid down the thistle funnel

Büchner flask

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Version 2: 5 M hydrochloric acid added to sodium chlorate(I) solution This reaction is slower than first version and is easier to control. The glassware is easy to clean at the end of the procedure. Sodium chlorate(I) solution is difficult to store and must be less than 1 year old to obtain a good supply of chlorine. Domestic bleach is not suitable. As the reaction subsides, either the acid or the sodium chlorate(I) solution can be alternatively added to the reaction vessel to supply more gas. The gas preparation arrangement (shown below) is known as the Andrews method. It is an alternative arrangement to using a thistle funnel. The arrangement in version 1 can also be used. Do not make this solution for the first time without seeking practical advice from a more-experienced colleague. General Hazards See Hazcards 22A, 20 & 89. Sodium chlorate(I) is a solution provided by the

supplier often as sodium hypochlorite. Sodium chlorate(V) is a solid. Do not get them mixed up.

• Use a fume cupboard. Make sure it is working. Wear goggles. • Place at least 50 ml sodium chlorate(I) solution (CORROSIVE) in the Büchner flask. • Set up the equipment as in the illustration below. The magnetic stirrer is optional but it does appear to

make the procedure more efficient. • Pour the 5 M hydrochloric acid (IRRITANT) into the separating funnel. • Turn the stirrer on and drip the acid into the flask. The solution will now bubble as the chlorine is

produced. • Collect the gas in a gas jar. When the contents of the gas jar are clearly green, assume the gas jar is

full. Or (optional), plug the gas jar with a large wad of cotton or mineral wool and put some damp blue litmus paper on top. It takes time for the chlorine to diffuse through the cotton wool and bleach the litmus. When the bleaching is complete the jar will be full. Remove it and replace the mineral wool with a gas jar cover or a large bung.

• To dispose of the reaction mixture, pour it down the fume cupboard sink with lots of water.

Separating funnel

5 M hydrochloric acid

Magnetic stirrer

Sodium chlorate(i)solution

Mineral wool

Moist blue litmus

Gas jar

Delivery tube as fardown as possible

Rubber tubing

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25 Chlorine water

The solubility of chlorine in water is about 0.6 g in 100 ml of water at room temperature. This would be a 0.6% (w/v) solution. Its concentration would be 0.085 M. An aqueous solution of chlorine should be prepared just before use. It does not keep for more than just a few days and should not be stored. It is very difficult to make up solutions of known concentration. All that is required for displacement reactions is that the solution works. It should, therefore, be trialled before use. Do not make this solution for the first time without seeking practical advice from a more experienced colleague. Three methods are described below. General Hazards Chlorine gas is TOXIC and CORROSIVE (see Hazcard 22A). Hazcard 22B deals with

chlorine water. Freshly-made chlorine water is formally LOW HAZARD but it has a considerable vapour of chlorine gas above it. It is better dispensed from a fume cupboard. More dilute solutions can be used in a well-ventilated room but staff should discourage direct inhalation of the vapour.

Method 1 • Use a fume cupboard. Wear goggles or a face shield and chemical-resistant gloves. • Bubble chlorine gas into 250 ml water in a gas jar until the solution goes light green. Use the

equipment for preparing chlorine gas but fill the gas jar half-full with water.

Method 2 Sodium chlorate(I) solution is an aqueous solution. Do not become confused with sodium chlorate(V), which is a solid.

10% w/v available chlorine sodium chlorate(I) solution does not store well. Over two years it may become completely useless.

• Use a fume cupboard. Wear goggles or a face shield and chemical-resistant gloves. • Place 10 ml of 10% w/v available chlorine sodium chlorate(I) solution (CORROSIVE) in 1 litre beaker. • Add about 80 ml of water and 10 ml of 2 M hydrochloric acid. Stir well. • Dilute to a suitable volume with more water.

Method 3 Sodium dichloroisocyanurate is used for purifying water.

• Use a fume cupboard. Wear eye protection. • Add 3 g of sodium dichloroisocyanurate (OXIDISING; HARMFUL), to 100 cm3 of water. When the solution

is clear, add 100 cm3 of 1 mol dm-3 hydrochloric acid.

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26 Chromatography solvents and locating agents

A mixture of compounds is placed on a stationary phase such as paper, or silica on thin layer plates. Chromatography involves passing a solvent through the stationary phase which separates the components of the mixture due to subtle differences in a compound's partition coefficient between the mobile solvent and the stationary phase. A locating agent is used to emphasise where the components of the mixture end up after the chromatography has finished. This is particularly important for colourless components of mixtures. Control measures for all of the following solutions

Solutions are better made up in a fume cupboard. Wear eye protection. There must be no sources of ignition in the vicinity.

Substances separated

Amino acids Solvent: Butan-1-ol, glacial ethanoic acid, water; 6:1:2 by volume.

Locator: Apply ninhydrin and heat in an oven at 110 °C or with a hairdryer.

See Hazcards 38A, 66, 84B. Label solvent HARMFUL and CORROSIVE.

Analgesics, eg, aspirin, paracetamol

Solvent: Ethyl ethanoate, hexane, ethanoic acid; 10:9:1 by volume.

Locator: UV light or iodine vapour. Do not look directly at UV radiation sources.

Label the solvent HIGHLY FLAMMABLE and CORROSIVE. See Hazcards 43A, 54A.

Anthocyanins (plant pigments)

Solvent: 100 ml 50% aqueous methanol solution with 1 ml of ethanoic acid or 1 ml concentrated hydrochloric acid in 100 ml ethanol.

Locator: Natural colour. (The components are light sensitive so run the chromatograms in the dark if possible and quickly note or photograph the results.)

Label the solvent HIGHLY FLAMMABLE

and TOXIC. See Hazcards 38A, 40B.

Inks from Biro pens

Solvent: Butan-1-ol, ethanol, water; 3:1:1 by volume. The addition of a few drops of 880 ammonia is said to produce a better chromatogram.

Locator: Natural colour.

Label the solvent HIGHLY FLAMMABLE and HARMFUL. See Hazcards 6, 40A, 84B.

Chlorophyll Solvent: Propanone, petroleum spirit (100-120 °C); 1 :9 by volume or cyclohexane, propanone, ethoxyethane; 2:1:1 by volume.

Locator: Natural colour of dyes and UV light. Do not look directly at UV light. (The components are light sensitive so the results should be noted or photographed quickly.)

Label the solvents HIGHLY FLAMMABLE. See Hazcards 45A, 85 or 42, 45B, 85.

Lipstick Solvent: 3-Methylbutan-1-ol,propanone and water; 2:1:9 by volume.

Locator: Natural colour.

The solvent is LOW

HAZARD See Hazcards 84C, 85.

Metal ions Solvent: Propanone, hydrochloric acid (conc), Distilled water; 17:2:1 by volume.

Locator: Use conc ammonia solution followed by 0.1% dithio-oxamide (rubeanic acid).

Label solvent HIGHLY

FLAMMABLE and

IRRITANT Label the locator CORROSIVE. See Hazcards 35, 47A, 85.

Nitration of methyl benzoate

For separating the crude products obtained from the nitration of methyl benzoate. Solvent: Ethoxyethane, pet ether (80/100 °C); 1:9 by volume. Locator: UV light. Do not look directly at UV radiation sources.

Label the solvent HIGHLY FLAMMABLE and HARMFUL. See Hazcards 42, 45A.

Sugars Solvent: Ethyl ethanoate, pyridine, water, 8:2:1 by volume. Locator: Dab Benedict’s solution on the chromatogram and dry in

an oven at 110 °C.

Label the solvent HIGHLY FLAMMABLE. See Hazcards 4C, 8, 43A.

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27 Chromium(III) chloride and chrome alum solutions

The colour of chromium(III) solutions varies with temperature and storage. Use cold water to dissolve the salt and do not heat the solutions. Prepare just before use.

General Hazards Chromium (III) salts are IRRITANT. See Hazcard 24.

Formula: CrCl3.6H2O Molar mass: 266.5 g mol-1 Solubility: 58 g per 100 ml

Preparing 100 ml of 0.1 M chromium(III) chloride solution

• Wear eye protection. • Dissolve 2.67 g of chromium(III) chloride-6-water in 70 ml of cold pure water. • Make up to 100 ml with pure water. • The solution is low hazard.

Formula: CrK(SO4)2.12H2O Molar mass: 499.4 g mol-1 Solubility: 22 g per 100 ml

Preparing 100 ml of 0.1 M chromium(III) potassium sulfate(VI) solution

• Wear eye protection. • Dissolve 5.0 g of chromium(III) potassium sulfate(VI)-12-water in 70 ml of cold pure water. • Make up to 100 ml with pure water. • The solution is low hazard.

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28 Citric acid

Also known as 2-hydroxypropane-1,2,3-tricarboxylic acid. It can purchased in either an anhydrous or monohydrate form. Values in italics below are for the monohydrate form. General Hazards See Hazcard 36C.

Formula: C6H8O7 Molar mass: 192.12 g mol-1 Solubility: 133 g per 100 ml Formula: C6H8O7.H2O Molar mass: 210.14 g mol-1 Solubility: 133 g per 100 ml





0.1 M 1.92 (2.10) 4.80 (5.35) 19.21 (21.01) -

0.5 M 9.60 (10.51) 24.02 (26.27) 96.06 (105.07) -

1.0 M 19.21 (21.01) 48.03 (53.54) 192.12 (210.14) -


Procedure

• Wear eye protection. • Measure out the indicated quantity of the solid citric acid. • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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29 Clock reactions

These are used for rate of reaction investigations. Solutions should be made a day before use and stored in the place where they will be used to attain room temperature. Clock reactions need to be tried out beforehand so that concentrations can be ‘tweaked’ to give reasonable times. Summer and winter temperature variations can also alter reaction times. The suggested amounts are for student activities. These reactions, however, make excellent demonstrations and the volumes can be significantly increased. The asterisk by the name of a solution indicates that this reagent is involved in the main reaction and its concentration can be varied. However, the amount of each solution must remain as specified. Varying reaction temperature will alter the time lapse for the clock. The addition of transition metal ions, eg, copper(II) ions, as a catalyst can also be investigated. However, you need to trial the experiment as the sequence that components are added is very important.

The hydrogen peroxide/potassium iodide reaction (The Harcourt-Essen reaction)

Solution A 1% starch solution (indicator) Solution B* 4 ‘vol’ hydrogen peroxide Solution C* 0.1 M hydrochloric acid Solution D 0.01 M sodium thiosulfate Solution E* 0.05 M potassium iodide Add 5 drops of solution A to a suitable beaker plus 10 ml each of solutions B to E in that order.

See Hazcards 47A & B, 67, 95C. Wear eye protection.

The thiosulfate/acid reaction (disappearing cross)

Solution A 0.1 M sodium thiosulfate Solution B 1 M hydrochloric or 0.05 M sulfuric(VI) acid Add 1 ml of the acid for every 10 ml of sodium thiosulfate. Once the run is complete, dispose of the contents in 0.5 M sodium carbonate solution to stop the reaction and neutralise sulfur dioxide.

See Hazcards 47A, 95C & 98. Wear eye protection. Do not use temperatures above 50 °C.

The potassium iodate/sodium metabisulfite reaction (The Landolt iodine clock)

Solution A 1% starch solution (indicator) Solution B* 0.025 M potassium iodate(V) solution (5.35 g per 1000 ml of solution) Solution C* 0.025 M sodium metabisulfite solution (4.75 g per 1000 ml of solution. Prepare in a fume cupboard.) Add 5 drops of solution A to a suitable beaker plus 10 ml each of solutions B and C.

See Hazcards 80, 92. Wear eye protection.

The reaction between iron(III) ions and iodide ions

Solution A 1% starch solution (indicator) Solution B 0.01 M sodium thiosulfate Solution C* 0.025 M iron(III) chloride using 0.1 M hydrochloric acid to dilute a more concentrated iron(III) solution Solution D* 0.025 M potassium iodide Mix 5 drops of solution A followed by 10 ml each of solutions B, C and D in that order.

See Hazcards 41, 47B, 55, 67, 95C. Wear eye protection.

The potassium iodide/potassium persulfate reaction

Solution A Dissolve 0.25 g of sodium thiosulfate in 100 ml of 1% starch solution Solution B* Dissolve 13.50 g of potassium persulfate in 1000 ml of solution Solution C* 0.1 M potassium iodide Mix 5 ml of solution A with 50 ml each of solutions B and C in that order.

See Hazcards 47B, 95B, 95C. Wear eye protection.

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30 Cobalt(II) chloride solution and thermochromic liquid

General Hazards Cobalt salts are TOXIC (See Hazcard 25). Reported carcinogenic and sensitisation warnings about cobalt compounds have become more severe in recent years. CLEAPSS has not received any reports from educational establishments of cobalt salts causing harm. The concerns for health are in mining and metallurgical applications used on a daily basis and in large doses.

Formula: CoCl2.6H2O Molar mass: 237.9 g mol-1 Solubility: 97 g per 100 ml

Preparing 100 ml of 0.1 M cobalt(II) chloride solution

• Wear eye protection. Wear disposable gloves and consider weighing the solid in non-working fume cupboard with the sash window low enough to stop any particles being inhaled.

• Dissolve 2.38 g of cobalt(II) chloride-6-water in 70 ml of water. • Make up to 100 ml with pure water. • The solution should be labelled TOXIC.

Formula: Co(NO3)2.6H2O Molar mass: 291 g mol-1 Solubility: 155 g per 100 ml

Preparing 100 ml of 0.1 M cobalt(II) nitrate solution

• Use the method above but dissolve 2.91 g of cobalt(II) nitrate-6-water in 70 ml of water. Formula: CoSO4.7H2O Molar mass: 281.1 g mol-1 Solubility: 97 g per 100 ml

Preparing 100 ml of 0.1 M cobalt(II) sulfate solution

• Use the method above but dissolve 2.81 g of cobalt(II) sulfate(VI)-7-water in 70 ml of water.

Thermochromic liquid Used in models of hot-water systems.

• Dissolve 40 g of hydrated cobalt(II) chloride in 1 litre of ethanol without heating. • Add drops of water and stir until pink. A drop or two of ammonia solution will precipitate the hydroxide

which shows the flow in the model. • Label the solution TOXIC & HIGHLY FLAMMABLE.

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31 Copper(II) solutions

It is advisable to use distilled or deionised water to make these solutions. Solutions of copper(II) salts are sometimes cloudy. If this is the case, add 1 ml of 1 M sulfuric acid and stir. Continue this procedure until the solution is clear. The Health & Safety Executive provide hazard classifications for solutions by % (w/v). We generally measure concentration in mols per dm-3 rather than % (w/v), which leads to inconsistencies in hazard classification between different copper salts with the same concentration. In the tables below the hazard classifications in brackets are consistent with that for copper sulfate(VI), the most commonly used copper(II) salt, but are not strictly correct. General Hazards Copper(II) salts are HARMFUL if swallowed. See Hazcard 27A, B & C.

Formula: CuSO4.5H2O Molar mass: 249.68 g mol-1 Solubility: 32 g per 100 ml Mass (g) of solid to be used




0.1 M 2.50 6.24 24.97 -

0.5 M 12.49 31.21 124.84 -

1.0 M 24.97 62.42 249.68 HARMFUL

Saturated (20 °C) 40 100 400 HARMFUL

Formula: CuCl2.2H2O Molar mass: 170.48 g mol-1 Solubility: 76 g per 100 ml Mass (g) of solid to be used




0.1 M 1.71 4.26 17.05 -

0.5 M 8.52 21.31 85.24 -

1.0 M 17.05 42.62 170.48 (HARMFUL)


Formula: Cu(NO3)2.3H2O Molar mass: 241.6 g mol-1 Solubility: 138 g per 100 ml Mass (g) of solid to be used




0.1 M 2.42 6.03 24.15 -

0.5 M 12.08 30.19 120.75 -

1.0 M 24.15 60.34 241.50 (HARMFUL)


Procedure

• Wear eye protection. • Measure out the indicated quantity of copper(II) salt. • Add the solid to about two thirds of the final volume of water in a beaker or jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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32 Crude oil alternative

Real crude oil (and petrol) contain benzene in concentrations greater than 0.1% and must not be used in school science (COSHH Regulations). A synthetic mixture can be prepared using mainly aliphatic hydrocarbons to illustrate the principle of fractional distillation of crude oil in industry. An alternative name for petroleum spirit is petroleum ether. On storing for several months, the lighter fractions evaporate away so the procedure should be tested to see that enough low boiling point fractions can be distilled off. Further details on the procedure can be found in L195, Safer Chemicals, Safer Reactions and section 13 of the Handbook. Standard fractional distillation equipment using a fractionating column is unsuitable as very high temperatures are required. Use apparatus similar to the diagram below.

General Hazards Petroleum spirit is HIGHLY FLAMMABLE and HARMFUL (see Hazcard 45A).

Preparing 100 ml of synthetic crude oil

• Wear eye protection. Do not prepare the mixture near sources of ignition. • Mix together 55 ml of liquid paraffin (medicinal), 20 ml of paraffin oil (kerosene), 11 ml of white spirit,

4 ml of petroleum ether (100-120 °C), 4 ml of petroleum ether (80-100 °C) and 6 ml of petroleum ether (60-80 °C).

• Add a squeeze of black oil paint (eg, Winsor and Newton’s Ivory Black) from a tube and stir well. • After adding to a labelled bottle, shake the mixture well. Label the container HIGHLY FLAMMABLE &

HARMFUL. • Always shake the mixture well before use. The paint will separate out if the bottle is left still for some

time. Note The recipe is not written in stone. Another possibility, if you have an even lower fraction of petroleum spirit, is:

Mix together 50 ml of liquid paraffin (medicinal), 20 ml of paraffin oil (kerosene), 10 ml of white spirit, 5 ml of petroleum ether (100-120 °C), 5 ml of petroleum ether (80-100 °C), 5 ml of petroleum spirit (60-80 °C) and 5 ml of petroleum spirit (40-60 °C).

75 x 10 mm test tubesto collect the fractions

1 ml of water in a test tube to compare volumes(optional)

Side-arm boiling tube

Mineral fibre with 6 ml of ‘crude oil’

Heat Block of wood to hold the test tubes

0 – 360 °C thermometer

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33 2,4-Dinitrophenylhydrazine solution

The solid is also known as 2,4-DNP and the solution is called Brady’s reagent. It is used to identify organic compounds with carbonyl groups by producing orange or yellow insoluble derivatives which, if purified by recrystallisation, give sharp melting points. Solid 2,4-DNP is supplied moist as there may be an explosion risk if very dry solid is handled. Unopened bottles of 2,4-DNP already contain 20% to 33% water. Two recipes for preparing Brady’s reagent are given below. The new recipe using phosphoric acid has been shown to work and is safer. General Hazards 2,4-DNP is EXPLOSIVE and TOXIC by inhalation, skin contact and if swallowed

(see Hazcard 30). For information about the alcohols see Hazcard 40A. Both concentrated sulfuric(VI) acid and phosphoric(V) acid are CORROSIVE (see Hazcards 98A and 72 respectively).

Formula: (NO2)2C6H3NHNH2 Molar mass: 198.14 g mol-1

Procedure for making about 100 ml of Brady’s reagent

Traditional method New method

• Wear eye protection, gloves and work in a fume cupboard.

• Stir 2.7 g of 2,4-DNP with 96 ml of methanol. Cautiously and slowly add 4 ml of concentrated sulfuric(VI) acid.

• Usually the solid dissolves to form a clear solution with a little stirring. If cloudy, filter.

• Wear eye protection and gloves. • Stir 2 g of 2,4-DNP with 50 ml of 85%

phosphoric(V) acid. • Add 50 ml of ethanol.

Label the solution TOXIC and HIGHLY FLAMMABLE.

Procedure for using Brady’s reagent

• Wear eye protection, and gloves. • Place 0.5 ml (if liquid) or 0.5 g (if solid) of the carbonyl compound to be identified in a test tube. • Add 5 ml of Brady’s reagent. [If crystals do not appear then add a little 2 M sulfuric(VI) acid.] • Cool the solution in an ice bath for 15 to 30 minutes. • Filter off the crystals (vacuum filtration is quicker). • Wash the crystals with water and recrystallise them before obtaining a melting point.

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34 3,5-Dinitrosalicylic acid

Also called DNSA or DNS. 3,5-Dinitrosalicylic acid is an alternative to Benedict’s solution for testing glucose and other reducing sugars. It can be used either qualitatively (yellow to red) or quantitatively using colorimetry with a green filter absorbing at 565 nm. It is more sensitive to glucose than Benedict’s solution. General Hazards 3,5-Dinitrosalicylic acid is HARMFUL by ingestion and irritating to the eyes and skin.

Preparing DNSA reagent

• Wear goggles. • Add 1.0 g of 3,5-dinitrosalicylic acid to 50 ml of water and warm gently to dissolve. • Slowly add 30 g sodium potassium tartrate tetrahydrate, (KNaC4H4O6·4H2O). The mixture thickens

(like custard). • Add 20 ml of 2 M sodium hydroxide solution and dilute the mixture with water to a final volume of

100 ml. • Label the solution IRRITANT.

Experimental procedure

• Wear eye protection. • Mix 0.3 ml of the sample with 3 ml of DNSA reagent. Glass vials are ideal for this. • Heat in hot water (from a kettle) for 5 minutes till a yellow colour develops. The samples do not need

to be boiled. • Remove the vial with tongs and allow to cool in a cold water bath. • The colorimeter will need to be set up with the green filter (about 565 nm). • Use reagent with water as a blank. • There are no further colour changes beyond a glucose concentration of 0.04 M, ie 0.72% (w/v).

Therefore, prepare a glucose standard curve with this concentration as the maximum. • Samples which are believed to contain higher amounts of glucose will need to be diluted before

testing.

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35 Drosophila food base

The food-base should pour easily when hot but set firmly when cold. It should also remain firm when the adult flies emerge.

Preparing the food base

• Soak 72 g of oatmeal in 120 ml of pure water. • Dissolve 35 g of treacle (black molasses) in 40 ml of pure water. • Boil 6 g of agar in 400 ml of pure water. • The mould inhibitor is made by dissolving 0.1 g of methyl 4-hydroxybenzoate (Nipagin; see

Hazcard 52) in a small volume of 95% (v/v) industrial denatured alcohol. • Mix all the ingredients and heat in a boiling-water bath, with constant stirring, for at least 15 minutes.

Add more water if the mixture does not pour easily. • Using a funnel, pour the mixture into either milk bottles or specimen tubes, making sure that the

medium does not drip down the sides of the vessels. • Autoclave at 15 psi (121 °C) for 15 minutes. • Allow the autoclave to cool naturally after sterilisation is complete; otherwise the medium will boil up

and contaminate the walls of the vessels.

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36 Electroplating solutions

There are many formulations for plating solutions. Some are hazardous and where they are not covered by a model (general) risk assessment, either here or elsewhere, a special risk assessment will be needed before use by teachers / technicians or students. Help can be obtained from CLEAPSS.

Use Recipe for 100 ml of solution Notes

Copper electroplating

Use 1 M copper(II) sulfate(VI) solution See Hazcard 27C. Label the solution HARMFUL.

Nickel electroplating

Dissolve 5 g of ammonium nickel(II) sulfate(VI) in 80 ml of water and dilute to 100 ml.

See Hazcard 65B. Label the solution HARMFUL.

Silver electroplating

Dissolve 1.6 g of silver nitrate(V) and 32 g of potassium iodide in 100 ml of water. Add 3 drops of concentrated sulfuric(VI) acid. Use a carbon rod as the anode.

See Hazcards 87 and 98A. Label the solution IRRITANT.

Zinc electroplating Dissolve 33 g of zinc sulfate(VI)-7-water in 50 ml of water and dilute to 100 ml. Add five drops of 2 M sulfuric(VI) acid and 2 “spatulas” full of boric acid. Boric acid crystals dissolve more easily than the powder. This is an old recipe and the word “spatula” is used which is not helpful. The solubility of boric acid is 6 g per 100 ml of water. 6 g is probably a sensible amount to add.

See Hazcards 14, 98A and 108B. It would be advisable to label the solution HARMFUL.

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37 Enzymes

This sheet is applicable to amylase, lipase, trypsin, pepsin etc. Enzyme preparations are available in the form of powders, capsules, tablets or concentrated solutions. Stock enzymes should be stored in the refrigerator and discarded when out of date or no longer active. Enzyme activity varies considerably with age, source and storage conditions. It is therefore essential to perform preliminary trials to check whether the enzyme is working and whether the endpoint is reached in an appropriate time. Allow time to order fresh stock in case the sample does not yield satisfactory results. If the concentration required is not specified on the protocol for the activity or suggested by the supplier, begin with 1% (w/v), test, and adjust accordingly. It is often preferable to obtain enzymes from a natural source. Purchased enzymes may have a microbial origin and are sometimes very heat stable or have unusual pH profiles so that investigations of the influence of temperature or pH on enzyme activity yield unexpected results. For example, saliva is often more reliable as a source of amylase than commercial products, which may also be contaminated with sugar. Health risks are minimal if each pupil uses his/her own saliva and rinses out contaminated glassware afterwards, prior to soaking in sodium chlorate(I) solution before normal washing up. Follow procedures in the Handbook. Note that for many enzymes, the pH will need to be adjusted to the optimum; check the details provided or the protocol. Enzyme powders may, in extreme cases, cause sensitisation. It is best to make up solutions in a fume cupboard which is not switched on. This helps to protect the operator from breathing in the powder in case it disperses. An operating fume cupboard may cause the powder to blow around. In case of a spill, the fume cupboard should be switched on to vent the powder. General Hazards See Hazcard 33. All enzyme powders are HARMFUL and may cause sensitisation.

Any solutions greater than 1% should be considered as IRRITANT.

Preparing 100 ml solution of 1% enzyme solution

• Wear eye protection and disposable nitrile gloves when making up solutions from the powders. • Avoid inhaling the powder. It may be wise to carry out the weighing and preparation in a fume

cupboard (with the fan switched off to avoid the draught affecting the balance). • Measure out 1 g of the enzyme. • Add the solid to about 70 ml of water or buffer (if needed for the activity) in a beaker. • Stir to dissolve (do not warm the solution). • Pour the solution into an appropriate measuring cylinder/volumetric flask. • Dilute to the final volume with pure water. • Pour into a labelled bottle and mix well. Store in the refrigerator or on ice during use. • Carry out the procedure that the students will undertake and consult with the teacher to confirm that

the results are satisfactory. It may be necessary either to dilute the solution further with more water or add more of the enzyme.

Sodium tauroglycocholate (Bile salts)

General Hazards See Hazcard 95C. Although low hazard, bile salts have an unpleasant odour and may be better prepared in the fume cupboard.

Preparing 100 ml of 3% bile salt solution

• Weigh out 3 g of bile salts. • Dissolve the salt in 70 ml of water. • Make up to 100 ml.

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38 Etching solutions

There are many formulations for etchants. Some are hazardous and where they are not covered by a model (general) risk assessment, here or elsewhere, a special risk assessment will be needed before use by teachers / technicians or students. Help can be obtained from CLEAPSS.

Use Recipe for 100 ml of solution Notes

Aluminium etching

Use 2 M sodium hydroxide solution at 60-70 °C. See Hazcard 91, Wear goggles.

Copper etching Dissolve 20 g of iron(III) chloride-6-water in 80 ml of water and 20 ml of concentrated hydrochloric acid (36% w/v).

See Hazcards 47A and 55C. Label the solution IRRITANT.

Iron etching 1 (Fry’s reagent)

Dissolve 59 g of copper(II) choride-2-water in 40 ml of water and add 60 ml of concentrated hydrochloric acid (36% w/v).

See Hazcards 27A and 47A. Label the solution CORROSIVE.

Iron etching 2 (Nital solution)

Add 2 ml of concentrated nitric(V) acid to 98 ml of ethanol. Under no circumstances use more than 2 ml of nitric(V) acid in making up 100 ml of solution.

See Hazcards 40A and 67. Label the solution HIGHLY FLAMMABLE & HARMFUL. Do not store.

Lead etching Wear goggles and chemical resistant gloves. Before use, mix together 50 ml of 5 M nitric(V) acid and 50 ml of 15% (w/v) ammonium molybdate(VI) solution. Apply with a swab for 30 s before rinsing.

See Hazcards 9A and 67. Label the solution CORROSIVE.

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39 Ethanoic acid

Also known as glacial acetic acid. The concentration of this solution is 17.4 mol dm-3. On cold days, the acid freezes to form a ‘glacial’ solid. In this case, place the bottle in warm water for several hours before making the diluted sample, but make sure the label does not come off. Formula: CH3COOH Molar mass: 60.05 g mol-1

General Hazards See Hazcard 38A. Ethanoic acid is CORROSIVE. It has a very sharp odour.

Volume (ml) of concentrated acid to be used




0.1 M Ten-fold dilution of 1 M solution -

0.5 M 14 29 71 -

1 M 29 57 143 -

2 M 57 114 286 IRRITANT

Procedure

• Wear goggles (a face shield is preferable for large volumes) and chemical resistant gloves. • Use the fume cupboard. • Measure out the indicated volume of ethanoic acid in a measuring cylinder. • Add the liquid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir the mixture well. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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40 Fehling’s solutions

Benedict’s solution or DNSA are safer alternatives when testing for sugars. Fehling's solution can only be used to test for aliphatic aldehydes, whereas Tollen's reagent (see Recipe Sheet 102) can be used to test for both aliphatic and aromatic aldehydes. Fehling's solutions A & B need to be made and stored separately. They are mixed just prior to use. Sandell’s solution (which is little known and not cited in exam school texts; see Recipe Sheet 76) can be made and stored for long periods. The concentration of sodium hydroxide is only 0.4 M in the solution (Fehling’s solution B is close to 4M).

General Hazards Copper(II) sulfate(VI) crystals are HARMFUL if swallowed (see Hazcard 27C). 2M sulfuric(VI) acid, solid sodium hydroxide and 0.5 M sodium hydroxide solution are CORROSIVE (see Hazcards 91 and 98A).

Preparing 100 ml of Fehling’s solution

Fehling’s solution A

• Wear eye protection. • Measure out 7 g of copper(II) sulfate(VI)-5-water and dissolve this in 60 ml of water. • If the solution appears cloudy add 10 ml of 2 M sulphuric acid. • Make up to 100 ml with water. Label the bottle.

Fehling’s solution B

• Wear goggles. • Measure out 15.4 g of sodium hydroxide and 35 g of potassium sodium tartrate. • Add the solids to 60 ml of water on a magnetic stirrer. Make up to 100 ml with water. Label the bottle

and add a CORROSIVE warning. Solutions A and B should be mixed in equal volumes, just prior to use. During the test, do not heat directly over a Bunsen burner: use a boiling water bath.

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41 Fixatives used before preserving biological specimens

Fixatives kill cells whilst retaining their structure. Although safer preservatives are available as alternatives to methanal, it is necessary to use methanal in many fixatives. (See Recipe Sheet 74 for ‘preservatives’.) More information about the procedures for fixing and preserving specimens can be found in the Laboratory Handbook, section 15. Methanal solution for fixing animal tissue

Dilute 10 ml of 40% methanal solution to 100 ml. Leave specimen in this solution for 16-48 hours before preserving.

For methanal, see Hazcard 63. Label the final solution HARMFUL. Use a fume cupboard, wear eye protection and disposable nitrile gloves.

Formalin-alcohol fixative for plant tissue

Mix together 90 ml of ethanol with 10 ml of 40% methanal solution. Leave specimen in this solution for 12 hours before preserving.

For ethanol; see Hazcard 40A. For methanal, see Hazcard 63. Label the solution HIGHLY FLAMMABLE and HARMFUL. Use a fume cupboard, wear eye protection and disposable nitrile gloves.

Formalin-aceto-alcohol (FAA) fixative for plant tissue

Mix together 5 ml of 40% methanal and 5 ml of glacial ethanoic acid in 90 ml of 70% ethanol. Leave specimen in this solution for 12 hours before preserving.

For ethanol; see Hazcard 40A. For methanal, see Hazcard 63. For ethanoic acid, see Hazcard 38. Label the solution HIGHLY FLAMMABLE & HARMFUL. Use a fume cupboard, wear eye protection and disposable nitrile gloves.

Acetic alcohol (Clarke’s fluid) (a cytological fixative especially for chromosomes)

Mix 25 ml of glacial ethanoic acid with 75 ml of absolute alcohol (Industrial denatured alcohol is normally adequate).

For ethanol; see Hazcard 40A. For ethanoic acid, see Hazcard 38. Label the solution HIGHLY FLAMMABLE

and CORROSIVE. Use a fume cupboard, wear eye protection and disposable nitrile gloves.

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42 Gases less commonly used in schools

Gas Hazard information

Minimum amount of reagents to prepare 1000 ml of gas

Carbon monoxide

See Hazcard 21 Concentrated sulfuric(VI) acid is slowly added to 2.9 g of solid sodium methanoate. Collect over water.

Dinitrogen monoxide

See Hazcard 68 5.8 g of hydroxyammonium chloride in 30 ml of water is added to a warm solution containing 81 g of ammonium iron(III) sulfate(VI) in 160 ml of water. Collect over warm water.

Hydrogen chloride

See Hazcard 49 2.1 ml of concentrated sulfuric(VI) acid is slowly added to excess solid sodium chloride. Collect by downward delivery (a tube into the bottom of an upright gas jar).

Hydrogen sulfide

See Hazcard 51 42 ml of 2 M hydrochloric acid is added to excess iron(II) sulfide. Collect over hot water.

Nitrogen See Hazcard 93 2.9 g of sodium nitrate(III) (nitrite) dissolved in a little water is warmed gently with excess ammonium chloride. Pass the gas through water to remove any oxides of nitrogen formed as a side reaction, and then collect.

Nitrogen dioxide

See Hazcard 68 8 ml of concentrated nitric(V) acid (70%) is added to an excess of copper turnings. Collect by downward delivery (a tube into the bottom of an upright gas jar).

Nitrogen monoxide

See Hazcard 68 30 ml of half-concentrated nitric(V) acid (35%) is added to an excess of copper turnings. Collect over water. OR 2.9 g of sodium nitrate(III) (nitrite) dissolved in 6 ml of water is added to 12 g of hydrated iron(II) sulfate(VI) and enough 5 M hydrochloric acid is added to cover the crystals. Collect over water.

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43 Hydrochloric acid

Hydrochloric acid is available from all major educational suppliers as 35 - 38% (w/w) solution with a specific gravity of 1.18. If you wish to know the dilution factors for other commercial concentrations (eg, 32%), then contact CLEAPSS. For accurate work, the concentration should always be found by titrating against a standard solution of sodium carbonate. The concentration of concentrated hydrochloric aid is about 11.7 mol dm-3. If kept for long periods, the concentration of 35 - 38% (w/w) hydrochloric acid decreases as the gas diffuses into the atmosphere. Do not make dilute solutions for the first time without seeking practical advice from a more experienced colleague. Formula: HCl Density: 1.18 g cm-3 Molar mass: 36.46 g mol-1 General Hazards See Hazcard 47A. Hydrochloric acid is CORROSIVE. It has a very sharp odour.




0.1 M Ten-fold dilution of 1 M solution -

0.4 M 17 34 84 -

0.5 M 21 42 105 -

1 M 42 84 209 -

2 M 84 167 417 IRRITANT Procedure

• Wear goggles (a face shield is preferable when handling large volumes) and chemical resistant gloves.

• Use a fume cupboard. Take care opening a bottle on a hot day. • Measure out the indicated quantity of concentrated hydrochloric acid in a measuring cylinder. • Add the liquid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir the mixture well. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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44 Hydrogen gas General Hazards See Hazcards 27C, 48 & 98. Hydrogen is extremely flammable. There must be no

Bunsen burner being used within 1 m of the apparatus. Note that the first gas collected consists of air and hydrogen. This is an explosive mixture and should be discarded.

• Wear goggles. • Place several granules of zinc in the

generator. • The reaction needs to be speeded up at the

start, to flush out air quickly. • Add about 5 ml of 1 M copper sulfate(VI)

(HARMFUL) down the thistle funnel. This reacts with zinc, coating it with copper which acts as a catalyst.

• Add 2 M sulfuric(VI) acid (CORROSIVE) down the thistle funnel.

• Collect the gas in the 250 ml measuring cylinder. Once the measuring cylinder is full, its contents (largely air) must be discarded.

• Now gas jars, boiling tubes, syringes and soap bubbles of pure hydrogen can be prepared.

Dry hydrogen [for burning or reducing copper(II) oxide]

• Use a U-tube filled loosely with calcium chloride granules (not powder) and collect the first gases in measuring cylinder one size larger than the hydrogen generator.

• Once the measuring cylinder is full, disconnect at A, and attach other equipment but before using the hydrogen, leave time to flush the air out of this newly-attached equipment. The gas collected in the measuring cylinder can be discarded.

250 ml measuring cylinder

Zinc

Hydrogengenerator

250 mlplastic bottle

1 to 5 ml of 1Mcopper(II) sulfate(VI) plus2M sulfuric acid

A 500ml measuring cylinder; to ensure all air is flushed through

U-tube

Calcium chloride

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45 Hydrogen peroxide

In the UK, the concentration of hydrogen peroxide is given in terms of the volume of oxygen it can produce; thus 1 ml of ‘10 vol’ hydrogen peroxide is capable of producing 10 ml of oxygen gas at 0 °C and 1atm. The maximum concentration of purchased hydrogen peroxide is ‘100 vol’ and even this solution will become more dilute with time. The concentration of 100 ‘vol’ solution is 8.3 mol dm-3. Commercial hydrogen peroxide solutions contain an inhibitor to slow down decomposition. During the preparation of diluted solutions, the inhibitor is diluted so solutions will deteriorate quite quickly. They should be prepared when required and not stored for long periods. For accurate work you will need to standardise the solution against a known solution of potassium manganate(VII). For activities with catalase, start with a 1 M solution and adjust subsequent concentrations depending on observed results. Formula: H2O2 Molar mass: 34.02 g mol-1

General Hazards See Hazcard 50. ‘100 vol’ hydrogen peroxide is HARMFUL.


Volume (ml) of solution required Hazard warning

label 250 1000 2500

0.08 M 1 vol 0.29% Twenty-fold dilution of the 20 ‘vol’ solution -

0.1 M 1.2 vol 0.34% Ten-fold dilution of the 1 M solution -

0.34 M 4 vol 1.14% Five-fold dilution of the 20 ‘vol’ solution -

1 M 12 vol 3.3% 29 115 288 -

1.7 M 20 vol 5.7% 50 200 500 IRRITANT

3.4 M 40 vol 11.4% 100 400 1000 IRRITANT

Procedure

• Wear eye protection and gloves. • Measure out the indicated quantity of 100 vol hydrogen peroxide in a measuring cylinder. • Add the liquid to about two thirds of the final volume of water in a beaker or laboratory jug and stir. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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46 Indicators (acid-base)

General Hazards See Hazcard 32. The hazards of many dyes and indicators are not well known. Many are made up in ethanol which is HIGHLY FLAMMABLE & HARMFUL (see Hazcard 40A).

Indicator Quantity

(g)

Volume of

IDA

Acid Alkali pH range

Bromophenol blue 0.4 200 Yellow Blue 3.0 - 4.5

Methyl orange 0.4 200 Red Yellow 3.0 - 4.5

Bromocresol green 0.4 200 Yellow Blue 4.0 - 5.4

Methyl red 0.2 300 Red Yellow 4.0 - 6.0

Bromothymol blue 0.4 200 Yellow Blue 6.0 - 7.5

Phenol red 0.4 200 Yellow Red 6.6 - 8.0

Neutral Red 0.4 200 Red Yellow 6.8 - 8.0

Cresol red 0.2 200 Yellow Purple 7.2 - 8.8

Thymol blue 0.4 200 Yellow Violet 8.0 - 9.6

Phenolphthalein 1.0 600 Colourless Mauve 8.0 - 10.0

Thymolphthalein 2.0 1000 Colourless Blue 9.3 - 10.5

Procedure to prepare 1000 ml of indicator solution

• Wear eye protection and use a fume cupboard when dispensing from bottles of solid indicators to avoid raising dust. Use gloves to avoid staining the skin.

• Dissolve the quantity of indicator in the volume of industrial denatured alcohol (IDA) given in the table in a suitable beaker.

• Transfer solution to a 1000 ml measuring cylinder and make up to 1000 ml with water.

Litmus solution • Dissolve 1 g of litmus in 1000 ml of water. Other recipes suggest up to 10 g of litmus but this may

depend upon the age of the sample. Filter if necessary.

Home-made indicators from fruits and vegetables Coloured aqueous solutions can be made from red cabbage, flower petals, blackberries, beetroot, etc, frozen in ice-cube trays and stored for months.

Screened methyl orange The normal colour change of some indicators is not always clear and may be altered by ‘screening’. The most common screened indicator used in schools is screened methyl orange. Dissolve 1 g of methyl orange and 2.6 g of xylene cyanol FF in 1000 ml of water. The end point is a grey colour.

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47 Indicator (universal)

General Hazards See Hazcard 32. The hazards of many dyes and indicators are not well known. Ethanol is HIGHLY FLAMMABLE & HARMFUL (see Hazcard 40A).

Procedure to prepare 1000 ml of Yamada Universal indicator solution

• Wear eye protection and use a fume cupboard (not switched on) to avoid raising dust when dispensing from bottles of solid indicators. Use gloves to avoid staining the skin.

• Place 0.25 g of bromothymol blue, 0.025 g of thymol blue, 0.063 g of methyl red and 0.5 g of phenolphthalein in an appropriate beaker.

• Add 500 ml of ethanol and stir to dissolve the dyes. • Add 500 ml of pure water. • Pour into a labelled 1000 ml bottle and mix well. • Label this solution (which is red) FLAMMABLE. • The colours of this solution when added to buffers are shown below.

3 3.5 4 4.5 5 5.5 6 6.5 7 7.5 8 8.5 9 9.5 10 10.5 11

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48 Indicators (carbon dioxide)

General Hazards See Hazcard 32. The hazards of many dyes and indicators are not well known. Many are made up in ethanol which is HIGHLY FLAMMABLE & HARMFUL (see Hazcard 40A).

Hydrogencarbonate (bicarbonate) indicator: Preparation of the stock solution: • Wear eye protection. • Use a fume cupboard (not switched on) to avoid raising dust when dispensing from bottles of solid

indicators. Use gloves to avoid staining the skin. • Dissolve 0.20 g of thymol blue and 0.1 g of cresol red in 20 ml of ethanol. • Dissolve 0.85 g of sodium hydrogencarbonate in about 200 ml of freshly-boiled distilled water. • Add the ethanolic dye solution and dilute to 1000 ml with water.

For use:

• Dilute the stock solution ten times with freshly-boiled distilled water. • Bubble air through the diluted solution to equilibrate it with atmospheric carbon dioxide. • When ready for use, the solution should be a deep cherry red colour in a bottle (in a test tube the

colour of the solution is less intense). If difficulties are experienced (eg, if distilled water is too acidic), try adding a pinch more sodium hydrogencarbonate. Avoid breathing over open vessels of the diluted indicator; the carbon dioxide exhaled may alter its pH.

Bromothymol blue indicator This can be used as an alternative to the above indicator in carbon dioxide uptake/release studies in biology. Some authorities report fewer difficulties in obtaining good results. The colour change is to yellow (more acid) and blue (more alkaline).

• Wear eye protection. • Use a fume cupboard (not switched on) to avoid raising dust when dispensing from bottles of solid

indicators. Use gloves to avoid staining the skin. • Dissolve 0.02 g of bromothymol blue in 20 ml ethanol before diluting to 1 litre with water.

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49 Indicators for redox, precipitation and complexometric titrations

Cerium(IV) titrations

Dissolve 1.49 g of 1:10-phenanthroline in 100 ml of water and add 0.70 g of iron(II) sulfate(VI). (Ferroin indicator).

Wear eye protection. 1:10-phenanthroline is TOXIC. The solution is low hazard.

Dissolve 0.1 g of N-phenylanthranilic acid in 5 ml of 0.1 M sodium hydroxide. Dilute to 100 ml with water.

See Hazcard 78.

Dichromate(VI) titrations

Dissolve 0.1 g of N-phenylanthranilic acid in 5 ml of 0.1 M sodium hydroxide. Dilute to 100 ml with water.

See Hazcard 78.

Dissolve 0.2 g of sodium or barium diphenylamine-4-sulfonate [diphenylamine-4-sulfonic acid (sodium salt)] in 100 ml of water.

See Hazcard 10B. Sodium diphenylamine-4-sulfonate is an IRRITANT. Wear eye protection.

Starch indicator for thiosulfate/iodine titrations (and detecting iodine)

Mix 1 g of soluble starch with a small amount of pure water and stir it vigorously to form a paste. Boil 100 ml of pure water and add it to the paste with stirring and allow to cool.

This ‘solution’ (it is really a suspension) does not keep. Dispose of it if it starts to go cloudier or ‘lumpy’.

Silver nitrate(V) titrations

Dissolve 5 g of potassium chromate(VI) in 100 ml of pure water. See Hazcard 78. Wear goggles. Label this solution TOXIC.

Dissolve 0.1 g of eosin in a mixture of 70 ml of ethanol and 30 ml of water.

See Hazcard 32 & 40A. Label this solution HIGHLY FLAMMABLE & HARMFUL. Precipitate becomes pink.

Dissolve 0.1 g of fluorescein (sodium salt) in 100 ml of water. See Hazcard 32 & 40A. Label this solution HARMFUL. Precipitate becomes pink.

(For Volhard’s method.) Dissolve 10 g of ammonium iron(III) sulfate(VI).12-water in 90 ml of water and add enough 2 M nitric acid to clear the solution and prevent hydrolysis. Make up to 100 ml. Use 1 ml per titration.

See Hazcard 55C. Colour change: white ppt to orange red colouration.

EDTA titrations

Dissolve 1 g of the Eriochrome black T (Solochrome black) in 100 ml of ethanol or grind 1 g of the solid dye with 100 g of sodium chloride. Use about 0.2 g of this solid mixture for each titration.

See Hazcard 32 & 40A. Label this solution HARMFUL. (Colour change is red to blue.)

Grind 1 g of murexide with 100 g of sodium chloride. Use 0.2 g for each titration.

See Hazcard 32 & 40A. Label this solution HARMFUL. (Colour change is red to blue.)

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50 Iodine solution

Iodine is only sparingly soluble in water (0.3 g/L). It is usual to dissolve it in aqueous potassium iodide solution (KI) or organic solvents such as ethanol. The procedure will take time even with stirring. A 0.01 M solution is suitable as a test reagent for starch. The concentration of solutions decreases with storage. Check that the solutions work before use in the laboratory. Formula: I2 Molar mass: 253.80 g mol-1

General Hazards See Hazcard 54. The organic solvents used are also hazardous and the relevant Hazcard should be consulted.

Mass (g) of solid to be used Concentration

required Volume (ml) of solution required Hazard

warning label

100 250 1000


0.1 M 8 g of KI + 2.54 g of I2 24 g of KI + 6.35 g of I2 80 g of KI + 25.38 g of I2 -

Procedure

• Wear eye protection and work in a well-ventilated room. Wear disposable nitrile gloves. • Measure out the indicated quantity of potassium iodide (KI) into an appropriate beaker. • Moisten the potassium iodide with a few drops of water. • Measure out the indicated quantity of iodine and add it to the moistened potassium iodide. • Add a small volume of water and stir. When no more iodine appears to dissolve, add some more water

and stir. Keep repeating this procedure until all the iodine has dissolved. • Pour the solution into a measuring cylinder and dilute to the final volume. Make sure there are no bits

of iodine remaining. If there are, return the solution to the beaker and leave it on a magnetic stirrer for several minutes.

• Add the solution to a labelled bottle and mix well.

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51 Iron(II) solutions

Comments about iron(II) salts Iron(II) sulfate(VI)-7-water crystals are green but they sometimes turn white through loss of water of crystallisation (efflorescence) or brown through oxidation during storage. Iron(II) chloride-4-water should also be green but it is even more susceptible to oxidation than iron sulfate(VI), changing to a brown solid. It is often quoted in experimental worksheets for displacement reactions but iron(II) sulfate(VI) or diammonium iron(II) sulfate(VI) work just as well. Neutral solutions of iron(II) salts turn brown very quickly on standing because of oxidation by air. Iron(II) salts are best prepared in an acidic solution rather than water. It is safer to use this procedure than to start adding concentrated sulfuric(VI) acid at the end, as is often stated in older recipe books. Diammonium iron(II) sulfate(VI) is more stable to air during storage. It is the preferred salt for titration experiments (see Recipe sheet 52). Do not to store these acidic iron(II) solutions for long periods. A clear green colour gradually turns to dirty green because of air oxidation. General Hazards See Hazcards 55B and 98A. Label the final solution IRRITANT because of the acid

present.

Iron(II) sulfate(VI)-7-water Also known as ferrous sulfate heptahydrate. Formula: FeSO4.7H2O Molar mass: 278.01 g mol-1 Solubility: 48 g per 100 ml



0.01 M Ten-fold dilution of a 0.1 M solution with 0.1 M sulfuric(VI) acid -

0.1 M 2.78 6.95 27.80 IRRITANT

1 M 27.80 69.50 278.01 IRRITANT


Iron(II) chloride-4-water Do not use the anhydrous salt. Formula: FeCl2.4H2O Molar mass: 198.8 g mol-1 Solubility: 68 g per 100 ml



0.01 M Ten-fold dilution of a 0.1 M solution with 0.1 M hydrochloric acid -

0.1 M 1.99 4.97 19.88 IRRITANT

1 M 19.88 49.7 198.80 IRRITANT


Procedure

• Wear eye protection. • Measure out the indicated quantity of iron(II) salt. • Add the solid to about two thirds of the final volume of 1 M sulfuric(VI) acid for the sulfate(VI) or 1 M

hydrochloric acid for the chloride in a beaker or laboratory jug. • Stir to dissolve (do not warm the solution). • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

either 1 M sulfuric(VI) acid for the sulfate(VI) or 1 M hydrochloric acid for the chloride to the required level.

• Pour into a labelled bottle and mix well. Add a hazard warning if appropriate.

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Diammonium iron(II) sulfate(VI) Also known as ammonium ferrous sulfate(VI) or Mohr’s salt.

Formula: (NH4)2SO4.FeSO4.6H2O Molar mass: 392.14 g mol-1 Solubility: 36 g per 100 ml

General Hazards 1 M sulfuric(VI) acid; Hazcard 98A. Iron(II) salts; Hazcard 55B.



0.01 M Ten-fold dilution of a 0.1 M solution with 0.1 M sulfuric(VI) acid IRRITANT

0.1 M 3.92 9.80 39.21 IRRITANT

0.5 M 19.61 49.02 196.07 IRRITANT


Procedure

• Wear eye protection. • Measure out the indicated quantity of diammonium iron(II) sulfate(VI). • Add the solid to about two thirds of the final volume of 1 M sulfuric(VI) acid in a beaker or laboratory

jug. • Stir to dissolve (do not warm the solution). • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

1 M sulfuric(VI) acid to the required level. • Pour into a labelled bottle and mix well. Add hazard warning.

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52 Iron (III) solutions

Aqueous solutions of iron(III) salts do not keep if made up solely in water. If required for a longer period, an acid should be added. Even in acid solution, there is gradual deterioration and a solid often separates out on storage. Iron(III) chloride (also known as ferric chloride) is available in two forms, hydrated iron(III) chloride and anhydrous iron(III) chloride. The anhydrous solid should not be used to make iron(III) solutions. It does not dissolve but reacts exothermically with water, to form iron(III) hydroxide and hydrogen chloride gas. Iron(III) chloride-6-water does not store well as it absorbs water (hygroscopic). Iron(III) sulfate(VI) has a variable amount of water of crystallisation (9-water is just one of the varieties on sale). It does not dissolve completely and the solution will need to be filtered (or the solid allowed to settle) before use. Ammonium iron(III) sulfate(VI) is much easier to use and is suitable for most applications. The lilac salt stores quite well but the surface sometimes becomes brown/yellow if the solid has become damp. The brown/yellow solid should be removed before weighing. The solid also deteriorates if exposed to heat and light. The solution is brown, Solutions should not be heated to accelerate dissolving as colloidal solutions can form. See the etching solution on Recipe sheet 38 if the iron(III) chloride is to be used for this purpose.

Iron (III) chloride Also known as ferric chloride. Formula: FeCl3.6H2O Molar mass: 270.30 g mol-1 Solubility: 92 g per 100 ml

General Hazards See Hazcard 55C. Do make sure you are using then hydrated salt and not the anhydrous salt.



0.01 M Ten-fold dilution of a 0.1 M solution with 0.2 M hydrochloric acid -

0.1 M 2.70 6.76 27.03 IRRITANT

1 M 27.03 67.58 270.30 IRRITANT


Procedure

• Wear eye protection. • Measure out the indicated quantity of iron(III) chloride-6-water. • Add the solid to about two thirds of the final volume of 1 M hydrochloric acid in a beaker or laboratory

jug. • Stir to dissolve (do not warm the solution). • Pour the solution from the beaker into an appropriate measuring cylinder or a laboratory jug and add

1 M hydrochloric acid to the required level. • Pour into a labelled bottle and mix well. Add a hazard warning if appropriate.

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Iron(III) sulfate(VI) Formula: Fe2(SO4)2.9.H2O Molar mass: 562 g mol-1

General Hazards See Hazcards 55C and 98. 1 M sulfuric(VI) acid. Label the final solution IRRITANT

because of the acid present.




0.1 M 5.6 14.1 56.2 IRRITANT

0.5 M 28.1 70.3 281 IRRITANT

1 M 56.2 140.5 562 IRRITANT

Ammonium iron(III) sulfate(VI) Formula: NH4Fe(SO4)2.12.H2O Molar mass: 482.19 g mol-1 Solubility: 124 g per 100 ml General Hazards See Hazcards 55C and 98. 1 M sulfuric(VI) acid. Label the final solution IRRITANT

because of the acid present.




0.1 M 4.82 12.05 48.22 IRRITANT

0.5 M 24.11 96.4 241.10 IRRITANT


Procedure

• Wear eye protection. • Measure out the indicated quantity of the iron(III) salt. • Add the solid to about two thirds of the final volume of 1 M sulfuric(VI) acid in a beaker or laboratory

jug. • Stir to dissolve (do not warm the solution). • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

1 M sulfuric(VI) acid to the required level. • Pour into a labelled bottle and mix well. Add a hazard warning if appropriate.

Ammonium iron(III) citrate and blueprints Wear eye protection. Dissolve 3 g of iron(III) chloride-6-water and 3 g of citric acid in 50 ml of water in a 250 ml plastic beaker. Add about 4.5 g of ammonium carbonate in small amounts, stirring each time. There is quite a lot of frothing and the solution turns green. This solution will keep. Blue print solution: Add 5 g of potassium hexacyanoferrate(III) and stir. This addition has to be made just before it is required.

Procedure

The solution is painted onto paper in dull indoor light (preferably in a cupboard) and hung to dry over paper towels or blotting paper. An opaque object (eg, scissors) is placed on the dry paper and left in the light for a day or two although, on a sunny day, 20 minutes may well be enough for light to convert soluble iron(III) salts to insoluble iron(II) salts. Wearing gloves, rinse the paper in a bowl of water to remove the solution leaving a colourless image on a Prussian Blue background. This can be messy and may stain clothing and unprotected hands. The paper is again hung to dry over absorbent paper.

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53 Lead(II) nitrate(V)

Tap water contains sulfate and carbonate ions which will precipitate out as insoluble lead compounds if lead ions are added. Lead salts should, thereore, be prepared in distilled or deionised water. Lead(II) nitrate(V) is used in preference to lead(II) ethanoate (acetate) as it appears to be slightly less hazardous. Disposal issues: see section 7.5 of the Handbook. Formula: Pb(NO3)2 Molar mass: 331.21 g mol-1 Solubility: 56 g per 100 ml

General Hazards See Hazcards 57A & B.



0.005 M Twenty-fold dilution of a 0.1 M solution with water or ten-fold dilution of 0.05 M solution

-

0.01 M Ten-fold dilution of a 0.1 M solution with water TOXIC

0.05 M 1.66 16.55 TOXIC

0.1 M 3.31 8.28 33.12 TOXIC

1 M 33.12 82.80 331.21 TOXIC

Saturated (20 °C) 56 140 560 TOXIC

Procedure

• Wear goggles and disposable nitrile gloves. If necessary, weigh materials on a balance placed in a fume cupboard which is not switched on and the sash pulled down.

• Measure out the indicated quantity of lead(II) nitrate(V). • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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54 Lithium chloride

Formula: LiCl Molar mass: 42.39 g mol-1 Solubility: 84 g per 100 ml

General Hazards Lithium salts are HARMFUL if swallowed and irritants. See Hazcards 47A and 58.





0.5 M 2.12 5.30 21.20 -

1.0 M 4.24 10.60 42.39 -


Procedure

• Measure out the indicated quantity of lithium chloride. • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

water to the required level. • Pour into a labelled bottle. Add a hazard warning if appropriate.

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55 Magnesium sulfate(VI)

Magnesium sulfate(VI)-7-water loses water of crystallisation slowly on standing to the air and becomes powdery (efflorescent). However, it is more convenient to make solutions from the hydrated salt than the anhydrous salt which is also available from suppliers. The anhydrous salt readily absorbs water from the atmosphere during storage. Formula: MgSO4.7H2O Molar mass: 246.47 g mol-1 Solubility: 71 g per 100 ml

General Hazards See Hazcard 59B.




0.1 M 2.47 6.16 24.65 -

0.5 M 12.32 30.81 123.24 -

1.0 M 24.65 61.62 246.47 -

Saturated (20 °C) 75 190 750 -

Procedure

• Measure out the indicated quantity of hydrated magnesium sulfate(VI). • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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56 Manganese(II) sulfate(VI)

Manganese(II) sulfate(VI) forms many hydrates. The 4 water and 1 water varieties appear to be the most readily available. The values in brackets in the table below are for the monohydrate. Formula: MnSO4.4H2O Molar mass: 223.07 g mol-1 Solubility: 95 g per 100 ml Formula: MnSO4.H2O Molar mass: 169.02 g mol-1 Solubility: 40 g per 100 ml

General Hazards Manganese(II) sulfate(VI) is HARMFUL. See Hazcard 60.




0.1 M 2.23 (1.69) 5.58 (4.23) 22.31 (16.90) -

0.5 M 11.15 (8.45) 27.89 (21.13) 111.54 (84.51) -

1.0 M 22.31 (16.90) 55.77 (42.26) 223.07 (169.02) HARMFUL

Saturated (20 °C) 100 (45) 250 (115) 1000 (450) HARMFUL

Procedure

• Measure out the indicated quantity of hydrated manganese(II) sulfate(VI). • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

water to the required level. • Pour into a labelled bottle. Add a hazard label if appropriate.

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57 Mercury solutions

Only make what is needed. Diluting to 0.04 M reduces the level of hazard to TOXIC. The main issue with mercury salts is that they should not be put down the drain. The waste should be collected, stored as toxic waste and removed to sealed landfill by a registered hazardous waste collector. Although labelled toxic and having a notorious history, using an inorganic salt will not kill you. However, the risk assessment for the activity must establish that the use of the mercury compound is really necessary and that no alternative is possible.

General Hazards Mercury salts are VERY TOXIC. See Hazcards 62 & 67.

Mercury(I) nitrate [also known as dimercury(I) nitrate(V)]

Make up in dilute nitric(V) acid to produce a clear solution. Formula: Hg2(NO3)2.2H2O Molar mass: 561.22 g mol-1 Solubility: 2 g per 100 ml Procedure for preparing 100 ml of 0.02 M of dimercury(I) nitrate(V)-2-water solution [which is 0.04 M with respect to the mercury(I) ion]

• Wear goggles. • Dissolve 1.12 g of the solid in 70 ml of 1 M nitric(V) acid. • Transfer the solution to a 100 ml measuring cylinder. • Make up to 100 ml with 1 M nitric(V) acid. • Label the solution CORROSIVE and TOXIC.

Mercury(II) chloride

Formula: HgCl2 Molar mass: 271.54 g mol-1 Solubility: 7.4 g per 100 ml Procedure for preparing 100 ml of 0.04 M of mercury(II) chloride solution

• Wear goggles. • Dissolve 1.09 g of mercury(II) chloride-2-water in 70 ml of water. • Transfer the solution to a 100 ml measuring cylinder. • Make up to 100 ml with water. • Label the solution TOXIC.

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58 Methanal solution

Also known as formalin or formaldehyde solution. The commercial solution is a 40% (w/v) solution of methanal in water with stabiliser added. Methanal is a gas at room temperature but it dissolves in water. If the bottle top is not secure then it will gradually lose gas and its concentration will fall. The concentration of the 40% solution is 11.3 mol dm-3. Formula: HCHO Molar mass: 30.03 g mol-1

General Hazards See Hazcard 63. It can cause burns, conjunctivitis or sensitisation by skin contact.

Volume of liquid required



0.3% 0.1 M Ten-fold dilution of a 0.1 M solution with water IRRITANT

1% 0.3 M 13 25 63 HARMFUL

3% 1 M 38 75 190 HARMFUL

4% 1.3 M 50 100 250 HARMFUL

Procedure

• Use a fume cupboard to handle the concentrated solution; wear goggles (a face shield is preferable when handling large volumes) and wear disposable nitrile gloves.

• Measure out the indicated volume of 40% methanal solution in a measuring cylinder. • Add the liquid to about two thirds of the final volume of water in a suitable beaker or laboratory jug and

stir. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

water to the required level. • Pour into a labelled bottle and mix well. Add the appropriate hazard warning.

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59 Methanoic acid

Also known as formic acid. It is usually supplied as a 90% (w/w) solution. The concentration of this solution is 23.9 mol dm-3. Formula: HCOOH Density: 1.22 g cm-3 Molar mass: 46.02 g mol-1

General Hazards See Hazcard 38A. Methanoic acid is CORROSIVE. It has a sharp odour.


Volume (ml) of solution required Hazard warning label 250 500 1000 2500

0.01 M Ten-fold dilution of the 0.1 M solution -0.1 M Ten-fold dilution of 1 M solution -1 M 10.5 21 42 105 -2 M 21 42 84 210 IRRITANT

Procedure

• Wear goggles (a face shield is preferable for large volumes) and chemical-resistant gloves. • Work in a fume cupboard. • Measure out the indicated quantity of methanoic acid in a measuring cylinder. • Add the liquid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir the mixture well. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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60 Nickel sulfate(VI)

Nickel sulfate(VI)-6-water is the most common nickel sulphate salt sold now, but do check the label on the bottle because the 7-water hydrate is also available. Formula: NiSO4.6H2O Molar mass: 262.86 g mol-1 Solubility: 65 g per 100 ml

General Hazards The hazards of nickel salts are being constantly reviewed. Sensitisation may be a problem for some people. See Hazcard 65B.

Procedure for preparing 100 ml of 0.1 M of nickel(II) sulfate(VI) solution

• Wear eye protection. • Dissolve 2.63 g of nickel(II) sulfate(VI)-6-water in 70 ml of water. • Make up to 100 ml with pure water. • The solution should be labelled TOXIC.

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61 Nitric(V) acid

The commercial solution is about 70% (w/v) and has a specific gravity of 1.42. Its concentration is 15.8 mol dm-3. Concentrated nitric(V) acid should be a colourless solution and stored in dark bottles. When exposed to light, the solution goes brown and TOXIC nitrogen dioxide gas is produced (see Hazcard 68). Nitrogen dioxide will leak from bottles and the solution will become less concentrated. Also available from some suppliers is 95-100% (fuming) nitric(V) acid, which is even more hazardous as it is unstable. It is unsuitable for schools to store this for long periods. Contact CLEAPSS if you find some. Do not make dilute solutions for the first time without seeking practical advice from a more-experienced colleague. Formula: HNO3 Molar mass: 63.01 g mol-1

General Hazards See Hazcard 67. If in contact with the skin, this becomes yellow and layers of skin peel off.



0.01 M Ten-fold dilution of a 0.1 M solution with water -

0.1 M Ten-fold dilution of a 1 M solution with water IRRITANT

0.4 M 13 25 63 IRRITANT

1 M 32 63 158 CORROSIVE

5M 158 317 792 CORROSIVE

Procedure

• Wear goggles (a face shield is preferable when handling large volumes) and chemical-resistant gloves.

• Work in a fume cupboard. • Measure out the indicated volume of concentrated nitric(V) acid in a measuring cylinder. • Add the liquid to about two thirds of the final volume of water in suitable beaker or laboratory jug. • Stir the solution (which will become warm). • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

water to the required level. • Pour the solution into a labelled bottle and mix well. Add a hazard warning if appropriate.

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62 Nylon rope experiment

Although used in the past, tetrachloromethane (carbon tetrachloride) (see Hazcard 100) and 1,1,1-trichloroethane (see Hazcard 103), should no longer be chosen as the solvent. An alternative is cyclohexane. Hexanedioyl chloride (adipoyl chloride) may be used but is more unstable in storage, so it is normally better to buy decanedioyl (sebacoyl) chloride. The acid chlorides are liquids at room temperature. They can be weighed out in a suitable container. Suppliers also provide the prepared solutions. These are used directly. Do check the labels.

General Hazards The acid chlorides and hexane-1,6-diamine are CORROSIVE (see Hazcards 3B & 41) and cyclohexane is HIGHLY FLAMMABLE & HARMFUL (see Hazcard 45B).

Procedure using cyclohexane as the solvent

• Wear eye protection. Wear disposable nitrile gloves when pulling out the nylon thread. The room should be well ventilated and there must be no sources of ignition.

• Dissolve 2.2 g of hexane-1,6-diamine in 50 ml of distilled water in a beaker (label it ‘A’). • Dissolve 1.5 g of decanedioyl chloride or hexanedioyl chloride in 50 ml of cyclohexane in another

beaker (label it ‘B’) and add HIGHLY FLAMMABLE & HARMFUL warning signs.

Large-scale version • Carefully pour the solution from beaker B onto solution A down a glass rod. The two liquid phases do

not mix; resist any attempt to stir the mixture at all. • Use forceps to pull out the nylon formed at the interface of the two solutions. • Disposal: see Handbook section 7.5.

Small-scale version

• Place sufficient solution A to cover the base in a Petri dish. • Carefully pour solution B onto solution A in a Petri dish down a glass rod. The two liquid phases do not

mix; resist any attempt to stir the mixture at all. • Use forceps to pull out the nylon formed at the interface of the two solutions. • The thread can pulled over glass rods or pulleys smeared with a lubricant such as WD40 (see pictures

below.) • Disposal: see Handbook section 7.5.

It is also possible to reduce the scale further and use 5 ml beakers. This is particularly suitable for use by students.

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63 Oscillating reactions

More information about these reactions can be found in Chemical Demonstrations (Vol 2), Shakashiri, 1985, University of Wisconsin Press, ISBN 0299101304. The entire four volume series is a good source of information.

The ‘blue-bottle’ reaction

• Wear goggles to prepare the sodium hydroxide solution. Wear eye protection for the experiment.

• In a 500 ml flask, dissolve 10 g of glucose in 300 ml of 0.4 M sodium hydroxide solution. Add 6 drops of a 0.2% aqueous methylene blue solution. Fit a rubber bung.

• When the solution becomes colourless, it can be shaken to turn it blue again after which it will again turn colourless.

Other dyes and mixtures of dyes that can be used are: • 1% resazurin solution (pale blue to purple pink). • 1% indigo carmine (yellow to red brown and to green with vigorous shaking). • 0.2% phenosafranine (pink to colourless). • Phenosafranine and methylene blue (pink to purple and to blue with vigorous

shaking) - amounts need to be found by trial and error. • Methyl red and methylene blue (yellow to green) – amounts need to be found by

trial and error.

See Hazcards 32 & 91. The solution should be labelled IRRITANT.

The Belousov-Zhabotinski (B-Z) reaction

All reagents in the Belousov-Zhabotinski (B-Z) reactions must be free of chloride ions, which inhibit the reaction. • Wear eye protection. • Place 100 ml of 1 M sulfuric(VI) acid in a beaker on a magnetic stirrer. • Add 2.86 g of malonic acid. Let it dissolve. • Add 1.04 g of potassium bromate(V). Let it dissolve. • Add 0.11 g of cerium(IV) ammonium nitrate(V) (IRRITANT). Let it dissolve. • Add about 0.5 ml of ferroin indicator (see Recipe Sheet 49). If the red/green

oscillations slow down, more potassium bromate(V) can be added.

See Hazcards 6B,

80 & 98A.

The Briggs-Rausher (B-R) reaction

All reagents in the Briggs-Rausher (B-R) reaction must be free of chloride ions, which inhibit the reaction. • Solution A: 10 vol hydrogen peroxide solution. • Solution B: 6% (w/v) potassium iodate(V) solution in 0.1 M sulfuric(VI) acid. • Solution C: 1% starch solution. • Solution D: a solution containing 2% (w/v) malonic acid and 0.45% (w/v)

manganese sulfate(VI). • Add the solutions to a beaker on an magnetic stirrer in the following order and

quantities: 20 ml of solution A, 20 ml of solution B, 1 ml of solution C and 20 ml of solution D.

See Hazcards 6B, 50 & 80. The diluted solutions do not require a hazard warning.

D. The nitrogen gas oscillating reaction Use a fume cupboard (nitrogen dioxide, VERY TOXIC, is produced). Wear eye protection. • Solution A: Dissolve 2.6 g of ammonium sulfate(VI) in 10 ml of 0.2 M sulfuric(VI)

acid. • Solution B: Dissolve 2.8 g of sodium nitrate(III) (nitrite) in 10 ml of water. • Mix the two solutions in a 100 ml beaker and stir quickly. Transfer to a 50 ml

measuring cylinder and after a minute the oscillations become quite distinct.

See Hazcards 68 & 93.

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64 Oxygen gas

Oxygen is prepared by adding hydrogen peroxide solution onto a catalyst, manganese(IV) oxide. The reaction is very vigorous and exothermic and chemicals have been known to shoot up the thistle funnel or the bung has been forced out of the flask. The rate of the reaction depends on the surface area of the catalyst. It is better to use granules rather than a fine powder. If only powder is available, then only a small amount is required. Potassium iodide (1 g) produces oxygen at a steadier rate. The solution first saturates itself with oxygen and then the gas is released. Hydrogen peroxide decomposes slowly during storage. Suppliers add an inhibitor to slow the process, but once this is used up, decomposition accelerates. If more hydrogen peroxide than expected is required, then this is a sure sign that the hydrogen peroxide is ‘old’ and new stock is required. 5 ml of 100 vol solution of hydrogen peroxide produces 500 ml of oxygen, ie, 5 x 100 ml. General Hazards See Hazcards 50, 60 & 69.

• Wear eye protection. • In the 250 ml Büchner flask, add 0.1 g of manganese(IV) oxide (HARMFUL) powder or 1 g of granules. • Fill the measuring cylinder with water ready to collect the first 250 ml of gas produced. The measuring

cylinder allows you to be confident that apparatus is well flushed out before oxygen is collected. • Pour at least 25 ml of cold water, enough to cover the bottom, into the thistle funnel. • Now add 5 ml of 100 vol hydrogen peroxide (HARMFUL). • Once the measuring cylinder is full of gas, remove it and replace with a gas jar full of water. • Keep collecting gas jars of gas. When full of gas, place a glass cover on the gas jar. • If the rate of gas being produced slows down, add further 5 ml portions of 100 vol hydrogen peroxide.

Add 5 ml of 100 volhydrogen peroxide

250 ml measuringcylinder

250-ml Büchner Flask

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65 Phosphoric(V) acid

Also known as orthophosphoric acid. The usual concentrated commercial solution is about 85% (w/v). The concentration of this solution is 14.7 mol dm-3. Formula: H3PO4 Density: 1.7 g cm-3 Molar mass: 98 g mol-1

General Hazards See Hazcard 72.


Volume (ml) of solution required Hazard warning

label 500 1000 2500


0.1 M 3.5 7.0 17.0 -

0.5 M 17 34 85 -

1 M 34 68 170 IRRITANT

Procedure

• Wear goggles and chemical-resistant gloves. • Measure out the indicated volume of concentrated phosphoric(V) acid in a measuring cylinder. • Add the liquid to about two thirds of the final volume of water in a suitable beaker or laboratory jug. • Stir the solution. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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66 Plant mineral requirement solutions

A Sach’s culture solution (complete recipe)

Dissolve the following salts in 1000 ml of pure water:

• 0.25 g of calcium sulfate(VI)-2-water • 0.25 g of calcium phosphate(V)-2-water [CaH4(PO4)2.2H2O] • 0.25 g of magnesium sulfate(VI)-7-water • 0.08 g of sodium chloride • 0.70 g of potassium nitrate(V) • 0.005 g of iron(III) chloride-6-water

B Sach’s culture solution with mineral deficiencies

Deficient in calcium: 0.2 g of potassium sulfate(VI) replaces calcium sulfate(VI)-2-water and 0.71 g of sodium dihydrogenphosphate(V)-2-water replaces calcium phosphate(V) in recipe A.

Deficient in iron: Omit iron(III) chloride-6-water in recipe A.

Deficient in magnesium: 0.17 g of potassium sulfate(VI) replaces magnesium sulfate(VI) in recipe A.

Deficient in nitrogen: 0.52 g of potassium chloride replaces potassium nitrate(V) in recipe A.

Deficient in phosphorus: 0.16 g of calcium nitrate(V)-4-water replaces calcium phosphate(V) in recipe A.

Deficient in potassium: 0.59 g of sodium nitrate(V) replaces potassium nitrate(V) in recipe A.

Deficient in sulfur: 0.16 g of calcium chloride replaces calcium sulfate(VI) and 0.21 g of magnesium chloride-6-water replaces magnesium sulfate(VI) in recipe A.

C Knop’s culture solution Solution 1 Dissolve the following salts in 1000 ml of pure water:

• 1 g of magnesium sulfate(VI) • 1 g of potassium nitrate(V) • 1 g of dipotassium hydrogenphosphate(V) [K2HPO4]

Solution 2

• Dissolve 4 g of calcium nitrate(V) in 1000 ml of pure water. Add solution 1 to solution 2 before use. This recipe is suitable for algae. For flowering plants, further dilution by 2, 3 or 4 times may produce better results. This should be determined by experiment.

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67 Potassium and sodium phosphates

These solutions are used to make buffer solutions (see Recipe Sheet 18 for Buffers). Masses are given to 3 decimal places but for many purposes, such accuracy is not required. Adjust to 2 decimal places. Make sure you are using the correct salt. It is best to check with the formula (see below). Sodium and potassium dihydrogen phosphate(V) solutions are best prepared just before use. During storage there may be mould growth. Potassium dihydrogen phosphate(V)

KH2PO4 0.2 M Dissolve 27.218 g of the solid in 500 ml of water and add water to 1000 ml.

Sodium dihydrogen phosphate(V)

NaH2PO4 0.2 M Dissolve 23.996 g of the solid in 500 ml of water and add water to 1000 ml.

Sodium dihydrogen phosphate(V)-1-water

NaH2PO4.H2O 0.2 M Dissolve 27.598 g of the solid in 500 ml of water and add water to 1000 ml.

Dipotassium hydrogen phosphate(V)

K2HPO4 0.2 M Dissolve 34.836 g of the solid in 500 ml of water and add water to 1000 ml.

Dipotassium hydrogen phosphate(V)-3-water

K2HPO4.3H2O 0.2 M Dissolve 45.644 g of the solid in 500 ml of water and add water to 1000 ml.

Disodium hydrogen phosphate(V)

Na2HPO4 0.2 M Dissolve 28.392 g of the solid in 500 ml of water and add water to 1000 ml.

Disodium hydrogen phosphate(V)-7-water

Na2HPO4.7H20 0.2 M Dissolve 53.614 g of the solid in 500 ml of water and add water to 1000 ml.

Potassium phosphate(V)

K3PO4 0.2 M Dissolve 42.454 g of the solid in 500 ml of water and add water to 1000 ml.

Sodium phosphate(V) Na3PO4 0.2 M Dissolve 32.788 g of the solid in 500 ml of water and add water to 1000 ml.

Sodium phosphate(V)-12-water

Na3PO4.12H2O 0.2 M Dissolve 76.024 g of the solid in 500 ml of water and add water to 1000 ml.

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68 Potassium chloride

Formula: KCl Molar mass: 74.55 g mol-1 Solubility: 11 g per 100 ml Mass (g) of solid to be used




1.0 M 7.46 18.64 74.55 -

3.0 M (used for filing pH probes)

22.34 55.91 223.65 -

Saturated (20 °C) 40 100 400 -

Procedure

• Measure out the indicated quantity of potassium chloride. • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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69 Potassium chromate(VI)

Formula: K2CrO4 Molar mass: 194.19 g mol-1 Solubility: 64 g per 100 ml General Hazards See Hazcard 78.




0.01 M Ten-fold dilution of the 0.1 M solution TOXIC

0.1 M 1.94 4.86 19.42 TOXIC

1.0 M 19.42 48.58 194.19 VERY TOXIC

Saturated (20 °C) 68 158 630 VERY TOXIC

Procedure

• Wear chemical-resistant gloves and goggles. Use a fume cupboard if the solid is a fine powder. • Measure out the indicated quantity of potassium chromate(VI). • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

water to the required level. • Pour into a labelled bottle. Add the hazard warning.

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70 Potassium dichromate(VI)

Potassium dichromate solutions are best made up in 0.1 M sulfuric(VI) acid rather than in pure water. This can be used as acidified potassium dichromate(VI) solution. The solution is used to test for the presence of alcohols. See Recipe Sheet 102 for ‘Test reagents for organic functional groups’. Formula: K2Cr2O7 Molar mass: 294.18 g mol-1 Solubility: 12 g per 100 ml General Hazards See Hazcard 78.




0.01 M Ten-fold dilution of the 0.1 M solution with 0.1 M sulfuric(VI) acid TOXIC

0.1 M 2.94 7.36 29.42 TOXIC

Saturated (20 °C) 12 30 120 VERY TOXIC

Procedure

• Wear chemical-resistant gloves and goggles. Use a fume cupboard if the solid is a fine powder. • Measure out the indicated quantity of potassium dichromate(VI). • Add the solid to about two thirds of the final volume of 0.1 M sulfuric(VI) acid in a beaker or laboratory

jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

0.1 M sulfuric(VI) acid to the required level. • Pour into a labelled bottle. Add the hazard warning.

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71 Potassium hydroxide

Solutions absorb carbon dioxide on standing. Solutions lower than 0.1 M are most significantly affected and should, therefore, be freshly made. Potassium hydroxide solution is best stored in plastic screw-cap bottles. It is known to etch glass when stored for long periods and polystop caps can allow carbon dioxide to enter. Equipment with glass joints, eg, burettes and bottles, can seize up over time. Formula: KOH Molar mass: 56.11 g mol-1 Solubility: 112 g per 100 ml General Hazards See Hazcard 91. When added to water so much heat is evolved that boiling could

occur. Do not make this solution for the first time without seeking practical advice from a more experienced colleague. A choking mist is often formed as the solid dissolves in water. While this is not a serious safety risk, it is unpleasant and it is wise to use a fume cupboard where possible. If saturated potassium hydroxide has to be prepared after carrying out an exhaustive risk assessment, then start from 5 M potassium hydroxide solution and add pellets a little at a time. Do not store this solution.





0.1 M Ten-fold dilution of the 1 M solution IRRITANT

0.4 M 2.24 g 5.61 g 22.44 g IRRITANT

1.0 M 5.61 g 14.03 g 56.11 g CORROSIVE

5.0 M 28.06 g 70.14 g 280.55 g CORROSIVE

Procedure

• Wear chemical-resistant gloves and goggles. Use a fume cupboard if the solid is a fine powder. • Measure out the indicated quantity of potassium hydroxide. • Add the solid in stages to about two thirds of the final volume of water in a beaker or laboratory jug. If

concentrated solutions are being made, ice should be used in place of water. • Stir carefully to dissolve before adding the next group of pellets. It may be necessary to cool the

solution between additions. (Ice could be added.) • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

water to the required level. • Pour into a labelled plastic bottle. Add a hazard warning if appropriate.

A 10% (w/v) alcoholic solution of potassium hydroxide Solutions absorb carbon dioxide on standing. Solutions lower than 0.1 M are most significantly affected and should, therefore, be freshly made.

• Wear chemical-resistant gloves and goggles. Use a fume cupboard if the solid is a fine powder. • Measure out 10 g of potassium hydroxide. • Place 70 ml of the alcohol (ethanol or methanol) in a glass 250 ml beaker. • Insert a magnetic stirrer bar and place the beaker on a magnetic stirrer. • Add the potassium hydroxide solution in 3 parts, allowing the solid to dissolving before adding the next

amount. The process is exothermic but not as extreme as that with water. • Place the solution in a 100 ml measuring cylinder and make up to the 100 ml mark with the alcohol. • Pour into a labelled plastic bottle. Label the solution CORROSIVE and HIGHLY FLAMMABLE.

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72 Potassium iodide

Solutions of potassium iodide often go yellow on keeping. If this occurs, add 0.1 M sodium thiosulfate solution dropwise, or a crystal of the solid with stirring, until the solution becomes colourless. Formula: KI Molar mass: 166.00 g mol-1 Solubility: 144 g per 100 ml Mass (g) of solid to be used




0.1 M 1.66 4.15 16.60 -

1.0 M 16.60 41.50 166.00 -

Saturated (20 °C) 150 375 1500 -

Procedure

• Wear eye protection. • Measure out the indicated quantity of potassium iodide. • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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73 Potassium manganate(VII) Also known as potassium permanganate. Solutions must be acidic. Solutions are safer made up in dilute sulfuric(VI) acid rather than adding concentrated sulfuric(VI) acid at a later stage. Solutions do not keep well unless the container is scrupulously clean. They slowly reacts with water, forming manganese(IV) oxide which badly stains glass and plastic equipment. Solutions are best kept in dark bottles, shielded from light. Light increases the rate of decomposition and staining of equipment. Equipment can be cleaned by filling the bottle either with 1 M sulfuric(VI) acid with a small amount of hydrogen peroxide solution (20 or 100 vol) or, if this does not work, add 2 M hydrochloric acid and leave it for several hours (or days) in a fume cupboard. Use a 0.002 M solution for carrying out tests for unsaturation in alkenes. It is not possible to make a 1 M solution. Formula: KmnO4 Molar mass: 158.03 g mol-1 Solubility: 6 g per 100 ml General Hazards See Hazcard 81. Mass (g) of solid to be used Concentration

required Conc of acid for dilution


0.002 M 0.1 M Ten-fold dilution of the 0.02 M solution with 0.1 M sulphuric(VI) acid

-

0.02 M 0.1 M 1.58 3.16 7.90 - 0.1 M 1 M 7.90 15.80 39.51 IRRITANT

Saturated 1 M 32 64 160 IRRITANT Procedure

• Wear eye protection. • It would be advisable to wear gloves as the chemical stains the skin brown. • Measure out the indicated quantity of potassium manganate(VII). • Add the solid to about two thirds of the final volume of the suggested dilute sulfuric(VI) acid in a

beaker or laboratory jug. • Stir to dissolve. It is better not to heat but dissolving can take some time, and it is difficult to see when

the solid has all dissolved. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

dilute sulfuric(VI) acid to the required level. • Pour into a labelled bottle.

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74 Preservatives used after fixing biological specimens

Preservatives prevent chemical decomposition of the fixed material (see Recipe Sheet 41). More information about the procedures for fixing and preserving specimens can be found in the CLEAPSS Handbook, section 15.8. Preserving animal tissue (Recipe 1)

Add 10 ml of odourless preservative (eg, Phenoxetol or Opresol) to 50 ml of propane-1,2-diol. Dilute this mixture to 100 ml with purified water. (Another version of this recipe includes and additional 1 g of sodium chloride.)

Wear eye protection. See Hazcard 37. Animal tissue should not be preserved for more than 1 month in these solutions unless it is fixed first. Label the solution IRRITANT.

Preserving animal tissue (Recipe 2)

Mix together 10 g of 4-methoxybenz-aldehyde (p-anisaldehyde), 50 ml of Teepol and 50 ml of propane-1,2-diol and make up this mixture to 1000 ml with purified water.

Wear eye protection. See Hazcard 37. Animal tissue should not be preserved for more than 1 month in these solutions unless it is fixed first. Label the solution IRRITANT. 4-Methoxybenzaldehyde is HARMFUL if swallowed. Label the solution HARMFUL.

Preserving plant tissue Use the formalin-aceto-alcohol (FAA) recipe on the fixative recipe sheet 41. To preserve the red or green colour add copper(II) sulfate(VI)-5-water crystals; the amount is not too critical.

Use a fume cupboard. Wear eye protection. For ethanol; see Hazcard 40A. For methanal, see Hazcard 63. For ethanoic acid, see Hazcard 38. For copper(II) sulfate(VI), see Hazcard 27C. Label the solution HIGHLY FLAMMABLE, IRRITANT and HARMFUL.

Kaiserling’s preservative

Add 36 g of sodium ethanoate, 5 ml of a 5% camphor solution in ethanol and 72 ml of propane-1,2,3-triol (glycerol). Add this mixture to 120 ml of purified water.

Wear eye protection. Camphor is HARMFUL. For propane-1,2,3-triol, see Hazcard 37. For sodium ethanoate see Hazcard 38A. for ethanol, see Hazcard 40A. Label the solution HARMFUL and HIGHLY FLAMMABLE.

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75 Ringer’s and other saline solutions for physiological use

Saline solutions are suitable for temporary microscopial preparations and short experiments. For more prolonged investigations, a suitable Ringer’s solution should be used. Ringer’s solutions For frogs Dissolve 6.50 g of sodium chloride, 0.12 g of calcium chloride-6-water (IRRITANT), 0.14 g of

potassium chloride and 0.20 g of sodium hydrogencarbonate in 1000 ml of water. For mammals Dissolve 8 g of sodium chloride, 0.2 g of calcium chloride-6-water (IRRITANT), 0.2 g of

potassium chloride, 0.19 g of magnesium chloride-6-water, 0.05 g of sodium dihydrogenphosphate and 1 g of sodium hydrogencarbonate in 1000 ml of water.

For locusts and other insects

Dissolve 7.60 g of sodium chloride, 0.22 g of calcium chloride-6-water, 0.75 g of potassium chloride, 0.19 g of magnesium chloride-6-water, 0.48 g of sodium dihydrogen phosphate, and 0.37 g of sodium hydrogencarbonate in 1000 ml of water.

For earthworms

Dissolve 6 g of sodium chloride, 0.2 g of calcium chloride-6-water, 0.12 g of potassium chloride and 0.1 g of sodium hydrogencarbonate in 1000 ml of water.

For marine crustaceans

Dissolve 31 g of sodium chloride, 1.37 g of calcium chloride-6-water (IRRITANT), 0.99 g of potassium chloride, 2.35 g of magnesium chloride-6-water, 0.48 g of sodium dihydrogenphosphate, and 0.22 g of sodium hydrogencarbonate in 1000 ml of water.

Saline solutions For invertebrate tissues Dissolve 7.5 g of sodium chloride in 1000 ml of water.

For amphibian tissues Dissolve 6.4 g of sodium chloride in 1000 ml of water.

For mammalian tissues Dissolve 9 g of sodium chloride in 1000 ml of water.

For mammalian blood Dissolve 6 g of sodium chloride in 1000 ml of water.

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76 Sandell’s solution

Sandell’s solution is a safer alternative than Fehling’s testing for sugars. Sandell’s solution (which is little known) can be stored for a long time. The advantage in its use is that the concentration of sodium hydroxide is 0.4 M in the solution, much lower than in Fehling’s solution B, which is close to 4M. The method, by A. Sandell, was published in the Journal of Chemical Education, April 1994, p346. SSERC has shown that this solution is stable enough to store and works with both aldehydes and monosaccharides. It is used in the same way as Benedict’s solution. General Hazards Copper(II) sulfate(VI) crystals are HARMFUL if swallowed (see Hazcard 27C).

2M sulfuric(VI) acid, solid sodium hydroxide and 0.5 M sodium hydroxide solution are CORROSIVE (see Hazcards 91 and 98A). EDTA (disodium salt) is low hazard (see Hazcard 3B).

Procedure to make 100 ml of Sandell’s solution

• Wear goggles when preparing the solution. • Measure out 0.80 g of copper(II) sulfate(VI)-5-water and 1.80 g of EDTA (disodium salt), and dissolve

in 80 ml of water. • Add 20 ml of 2 M sodium hydroxide (CORROSIVE). • Make up to 100 ml with water. Label the bottle IRRITANT.

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77 Silver nitrate(V)

This is probably the most expensive chemical used in schools; use it with care, retrieving unused stock. Pure water (ie, distilled or deionised) should be used to avoid cloudiness of solutions. Tollen’s reagent [ammoniacal silver nitrate(V) solution], (see Recipe sheet 102) should not be stored. It should be made and disposed of in situ by the students. Formula: AgNO3 Molar mass: 169.87 g mol-1 Solubility: 216 g per 100 ml General Hazards See Hazcard 87. The solid and more-concentrated solutions will stain skin and other

organic material black. This stain is difficult to remove. Work surfaces also stain. Mass (g) of solid to be used



0.01 M Ten-fold dilution of the 0.1 M with water -

0.1 1.70 4.25 16.99 -

0.2 3.40 8.50 33.98 IRRITANT

0.5 8.49 21.23 84.94 CORROSIVE

Saturated (20 °C) 220 550 2200 CORROSIVE

Procedure

• Wear eye protection. • Measure out the indicated quantity of silver nitrate(V). • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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78 Slime

Different sources of polyvinyl alcohol vary in their average molar mass; some are about 115000 g mol-1, while others are much less, at around 17000 g mol-1 (and less expensive). The larger the average molar mass, the better the slime. PVA wood glue is cheaper still but contains polyvinyl acetate and other chemicals such as binders. This will affect the nature of the slime and it may become more solid. Custard powder can also be used with PVA glue to form bouncing custard balls. Slime is a gel. It is thought that borate ions react with hydroxyl groups in the polymer of vinyl alcohol to form cross links, with the elimination of water. These cross links probably involve hydrogen bonds which continually form and break under flow. The ‘sliminess’ of the gel can be adjusted by altering the amount of sodium borate used. Viscosity changes can be detected by timing the passage of a ball bearing through the ‘slime’ in a measuring cylinder. If acid is added to the ‘slime’, the gel collapses to give a free-flowing solution. Although the slime can be stored in a labelled sealed bag and placed in a refrigerator, it can develop mould if kept for long periods. Disposal: Add enough 1 M sulfuric(VI) acid to just allow the solution to become free-flowing, dilute and pour down a foul-water drain. These chemicals, and the made-up slime, should not be taken home. General Hazards See Hazcard 14. However, since publication of Hazcards, the toxicity of boron

compounds has been the subject of a considerable review and debate in the European Union. Some suppliers will now label the bottle TOXIC with R63 (Possible risk of harm to the unborn child) warning. However only solutions above 8% (w/v) will be labelled TOXIC. Sodium tetraborate should be weighed in a fume cupboard to avoid raising dust.

Procedure using polyvinyl alcohol (PVA) PVA with a molar mass above 85,000 g mol-1 PVA with a molar mass below 85,000 g mol-1

Wear eye protection. Hot water from a kettle may be useful to start with. Those with sensitive skin may be affected by the weak alkalinity of the solutions. In this case, disposable gloves should be worn.

• Pour about 100 ml of water (no hotter than 90 °C) into a 400 ml beaker, add 4 g of high-mass polyvinyl alcohol, stirring rapidly. Add food colouring and/or fluorescent dye, eg, fluorescein, for added (disgusting) effect.

• Heat to 90 °C (but do not boil) and keep stirring as required until the polymer dissolves. Allow the solution to cool. All of this may take some time (Solution A.).

• Dissolve 0.80 g of sodium tetraborate-10-water (0.42 g of anhydrous sodium tetraborate) in 20 ml of water. (Solution B.)

• Pour about 100 ml of water (no hotter than 90 °C) into a 400 ml beaker, add 8 g of low-mass polyvinyl alcohol, stirring rapidly. Add food colouring and/or fluorescent dye, eg, fluorescein, for added (disgusting) effect.

• Heat to 90 °C (but do not boil) and keep stirring as required until the polymer dissolves. Allow the solution to cool. All of this may take some time (Solution A.).

• Dissolve 1.6 g of sodium tetraborate-10-water (0.84 g of anhydrous sodium tetraborate) in 20 ml of water. (Solution B.)

• Add solution B to A with vigorous stirring. Let the gel form before removing it and washing with water.

Using PVA glue: Dilute it to about 25%; there is no need to heat it but mix it well with water. Add sodium tetraborate as prepared above for use with low-mass PVA. Change the concentration of PVA glue to alter the ‘slime’ quality.

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79 Soap and bubble solutions

Alcoholic soap solution can be titrated against tap water in a stoppered bottle until a froth, stable for 15 seconds, is obtained. The concentration of soap solution and/or volume of water involved in the titration will depend on the hardness of the local tap water. Solutions should be tested before being presented to a class so that minor adjustments to the formulation can be made. The alcoholic soap solution is called either Clarke’s or Wanklyn’s soap solution. General Hazards See Hazcards 37 & 40A. Do not heat soap solutions made up in ethanol with a

naked flame. Use a hot plate or surround the beaker with hot water.

Procedure to make a soap solution suitable for titrating

• Dissolve 5 g of soap flakes (eg, Lux) in 500 ml of ethanol by stirring the suspension on a hot plate. This may take some time.

• After cooling, pour the solution into a 1 litre volumetric flask and dilute to 1000 ml with pure water. Mix the solution well.

• The titration with tap water to obtain a permanent froth should be rehearsed so that adjustments to the volume of tap water or concentration of the soap solution can be made.

Bubble mixture Recipes may differ with different makes of liquid detergent. The recipe below works best using Fairy Liquid. (other detergents will also work but you may need to experiment with quantities. Procedure to make a bubble solution

• Mix together by volume, 10 parts liquid detergent, 85 parts pure water and 5 parts propane-1,2,3-triol (glycerol).

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80 Sodium carbonate

Sodium carbonate is sold both as anhydrous sodium carbonate or sodium carbonate-10-hydrate. Anhydrous sodium carbonate absorbs water from the atmosphere (deliquescence). Hydrated sodium carbonate loses water to the atmosphere (efflorescence); look for a white powder as opposed to a ‘glassy’ solid. Do not use technical grade sodium carbonate to make up solutions. There is a lot of sediment. Solutions may be cloudy if hard tap water is used. Formula: Na2CO3 Molar mass: 105.99 g mol-1 Solubility: 22 g per 100 ml Formula: Na2CO3.10H2O Molar mass: 286.14 g mol-1 Solubility: 50 g per 100 ml

General Hazards See Hazcard 95A.

Anhydrous sodium carbonate Mass (g) of solid to be used



0.05 M Two-fold dilution of 0.1 M solution -

0.1 M 2.65 5.30 10.60 -

0.5 M 13.25 26.50 53.00 -

1 M 26.50 53.00 105.99 -

Saturated 55 110 220 IRRITANT

Hydrated sodium carbonate Mass (g) of solid to be used



0.05 M Two-fold dilution of 0.1 M solution -

0.1 M 7.15 14.31 28.61 -

0.5 M 35.77 71.54 143.07 -

1 M 71.54 143.07 286.14 -

Saturated 125 250 500 IRRITANT

Procedure

• Wear eye protection. • Measure out the indicated quantity of anhydrous or hydrated sodium carbonate. • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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81 Sodium chlorate(I) solution

Also known as sodium hypochlorite solution, sodium chlorate(I) or bleach. Do not get confused with the solid sodium chlorate(V). The solution sold by laboratory suppliers and described as ‘10-14% available chlorine’ contains 100 000 to 140 000 ppm of chlorine and has a molarity of 1.5 M. It has a limited shelf-life and may not be as active after a year in storage. Some technicians use domestic products such as bleach but please read the containers carefully as some bleaches use hydrogen peroxide and not sodium chlorate(I). Domestic bleach is usually 0.5 M or less and represents poorer value per mole but may be suitable if only needed infrequently. However, there are also thickeners and detergents in domestic bleach that may cause problems. See also Handbook section 15.12. Solutions used for disinfecting should always be freshly prepared. If using this as a disinfectant, purchase or prepare fresh stocks after 3 months or so. Formula: NaClO Molar mass: 74.45 g mol-1 General Hazards See Hazcard 89.

Concentration required Preparation Use 1% 10000 ppm 0.15 M See below for procedure.

No need for further dilution. Disinfectant for blood spills or dirty conditions.

0.25% 2500 ppm 0.038 M Mix, by volume, 1 part of the 1% solution with 3 parts of water.

Disinfectant for micro-biological discard pots.

0.1% 1000 ppm 0.015 M Mix by volume, 1 part of the 1% solution with 9 parts of water.

Disinfectant for general use, eg, sterilising solution.

0.01% 100 ppm 0.0015 M Dilute the 0.1% solution further by mixing by volume, 1 part of the 0.1% solution with 9 parts of water.

Disinfectant for sterilising mouthpieces, swabbing skin.

Procedure to make 1000 ml of 1% available chlorine solution

• Wear gloves and goggles or a face shield. • Measure out 100 ml of 10-14% sodium chlorate(I) solution into a 1000 ml measuring cylinder. • Add water to the 1000 ml level. • Pour the solution into an appropriate labelled bottle and mix well. The solution is low hazard. • Depending on use, dilute, if necessary, from a 1% solution according to the table above.

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82 Sodium chloride

Formula: NaCl Molar mass: 58.44 g mol-1 Solubility: 36 g per 100 ml Mass (g) of solid to be used




0.5 M 2.92 7.31 29.22 -

1.0 M 5.84 14.61 58.44 -

Saturated (20 °C) 37 93 370 -

Procedure

• Measure out the indicated quantity of sodium chloride. • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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83 Sodium ethanoate

Sodium ethanoate is sold as the anhydrous salt or as the trihydrate.

Anhydrous sodium ethanoate Formula: NaCH3COO Molar mass: 82.03 g mol-1 Solubility: 46 g per 100 ml Mass (g) of solid to be used



0.2 M 5-fold dilution of the 1 M solution -

1.0 M 8.20 20.51 82.03 -

Saturated (20 °C) 50 125 500 -

Hydrated sodium ethanoate Formula: NaCH3COO.3H2O Molar mass: 136.08 g mol-1 Solubility: 76 g per 100 ml Mass (g) of solid to be used



0.2 M 5-fold dilution of the 1 M solution -

1.0 M 13.61 34.02 136.08 -

Saturated (20 °C) 80 200 800 -

Procedure

• Measure out the indicated quantity of sodium ethanoate. • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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84 Sodium hydrogencarbonate

Formula: NaHCO3 Molar mass: 84.01 g mol-1 Solubility: 10 g per 100 ml Mass (g) of solid to be used




0.5 4.20 11.50 42.00 -

1.0 M 8.40 21.00 84.01 -

Saturated (20 °C) 12 30 120 -

Procedure

• Measure out the indicated quantity of sodium hydrogencarbonate. • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve. Do not warm as decomposition may take place. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


Sodium hydrogencarbonate - alkaline pyrogallol solution for absorbing or removing oxygen gas Although safer than using sodium hydroxide with pyrogallol (benzene-1,2,3-triol), the absorption of oxygen is slower and may be too slow for some procedures In such circumstances, there is no alternative to the use of sodium hydroxide. The procedure described below limits the absorption of oxygen from the air. General Hazards See Hazcard 12.

Procedure

• Wear eye protection when preparing the solution. • Prepare a saturated solution of sodium hydrogencarbonate using freshly-boiled pure water. • Place a wide-bore glass tube into the water and then pour liquid paraffin onto the water’s surface. • Add a few crystals of benzene-1,2,3-triol (pyrogallol) down the glass tube and then slowly withdraw it.

(1 g of benzene-1,2,3-triol is capable of absorbing 190 cm3 of oxygen.)

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85 Sodium hydroxide

Solutions absorb carbon dioxide on standing. Solutions lower than 0.1 M are most significantly affected and should, therefore, be freshly made. Sodium hydroxide solution is better stored in plastic screw-cap bottles. It is know to etch glass when stored for long periods and polystop caps can allow carbon dioxide to enter. Equipment with glass joints, eg, burettes and bottles, can seize up over time. Formula: NaOH Molar mass: 40.00 g mol-1 Solubility: 108 g per 100 ml General Hazards See Hazcard 91. When added to water, heat is evolved such that boiling could

occur. Do not make this solution for the first time without seeking practical advice from a more-experienced colleague. A choking mist is often formed as the solid dissolves in water. While this is not a serious safety risk, it is unpleasant and it is wise to use a fume cupboard where possible. If after carrying out an exhaustive risk assessment, there is no alternative to preparing saturated sodium hydroxide, then start from 5 M sodium hydroxide solution and add pellets a little at a time. Do not store this solution.

Mass (g) of solid to be used Concentration

required Volume (ml) of solution required Hazard warning

label 250 1000 2500 0.01 M Ten-fold dilution of the 0.1 M solution -

0.1 M Ten-fold dilution of the 1 M solution IRRITANT

0.4 M 4.00 16.00 40.00 IRRITANT

1.0 M 10.00 40.00 100.00 CORROSIVE

5.0 M 50.00 200.00 500.00 CORROSIVE

Procedure

• Wear chemical-resistant gloves and goggles. Use a fume cupboard if the solid is a fine powder. • Measure out the indicated quantity of sodium hydroxide. • Add the solid in stages to about two thirds of the final volume of water in a beaker or laboratory jug. If

concentrated solutions are being made, ice should be used in place of water. • Stir carefully to dissolve before adding the next group of pellets. It may be necessary to cool the

solution between additions. (Ice could be added.) • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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86 Sodium silicate, the crystal (chemical) garden and silicate gels

The solution is also known as water glass. It is more convenient to buy a solution which has a specific gravity of about 1.5, rather than the solid, and dilute it down. It is, however, possible to make a solution from the solid. Expensive glassware can be very difficult to clean after using this reagent. In this case, it is better to use old jam or coffee jars and dispose of them after use. General Hazards See Hazcard 95B. The solution is strongly alkaline. Consult the relevant Hazcards

when using metal salts to add to the chemical garden. Procedure to make 250 ml of sodium silicate solution for a chemical garden from the commercially-available solution

• Wear eye protection. • Add 200 ml of commercial sodium silicate to 600 ml of pure water in a 1-litre beaker. • Pour into a labelled bottle. Although the solution is deemed low hazard, it might be prudent to label the

bottle IRRITANT as the solution is strongly alkaline. Procedure to make 250 ml of 8% sodium silicate solution from the solid

• Wear eye protection. • Add 20 g of sodium silicate to a 600 ml beaker and add 250 ml of pure water. • Add a magnetic stirrer bar and place the mixture on a heater/stirrer. • Maintain a temperature of about 45 °C until the solid appears to dissolve. • Filter through Whatman No.1 filter paper. • Pour into a labelled bottle. Although the solution is deemed low hazard, it might be prudent to label the

bottle IRRITANT as the solution is strongly alkaline. The crystal (chemical) garden

• Wear eye protection and consider wearing disposable nitrile gloves. • Pour the prepared sodium silicate into a 600 ml beaker (a labelled coffee jar is also suitable), cover it

with a clock glass or sheet of glass (check edges are not sharp) and allow the liquid to settle. (After adding crystals, it is better not to move the container so any moving should be done at this stage.)

• Now add small (eg, rice-grain size) crystals of manganese(II) sulfate(VI), copper(II) sulfate(VI), chromium(III) chloride, iron(II) sulfate(VI), iron(III) chloride, cobalt(II) chloride, tin(II) chloride, aluminium sulfate(VI), nickel(II) sulfate(VI) or any other soluble salt you wish to try.

• Leave the container undisturbed for several days. Label the container IRRITANT. Growing lead(II) iodide crystals in gels

• Dilute 16 ml of commercial sodium silicate solution to 100 ml with pure water (solution A). • Place 15 ml of 1 M ethanoic acid (see Recipe Sheet 39) in a beaker and add 0.67 g of potassium

iodide (solution B). • Add 15 ml of solution A to solution B, stirring vigorously. Pour the mixture into a boiling tube and allow

it to set (5 to 60 minutes). • Dissolve 0.66 g of lead(II) nitrate(V) in 2 ml of water and pour this onto the top of the gel. Label the

container IRRITANT. • Observe the spread of crystals of lead(II) iodide over the next few weeks.

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87 Sodium thiosulfate

Also known as ‘hypo’ and it is usually supplied as hydrated crystals. Distilled water is naturally acidic (dissolved carbon dioxide) and this may result in the slow formation of sulfur over several hours. This is avoided by adding ‘a pinch’ of sodium hydrogencarbonate or sodium sulfate(IV) to the solution to neutralise the acidity. A 25% solution is suitable to keep with bromine for use in case of accidents. (See Hazcard 95C.) Formula: Na2S2O3.5H20 Molar mass: 248.17 g mol-1 Solubility: 70 g per 100 ml Mass (g) of solid to be used




0.1 M 2.48 6.20 24.82 -

0.5 M 12.41 31.02 124.08 -

1.0 M 24.82 62.04 248.17 -

Saturated (20 °C) 75 188 750 -

Procedure

• Measure out the indicated quantity of sodium thiosulfate-5-water. • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve. Do not warm the solution. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add

water to the required level. • Add about 1 g of sodium hydrogencarbonate or sodium sulfate(IV) (sulfite) (HARMFUL) for every

1000 ml of solution, to help to preserve it. • Pour the solution into a labelled bottle and mix well.

A saturated solution The saturated solution requires little water. Weigh out the sodium thiosulfate as indicated and add it to hot (almost boiling) water with a volume in millilitres equal to about half the weight of solid in grams. Stir to dissolve and leave to cool to room temperature. If there is no solid present in the cooled solution, add one crystal of solid to it. If this causes further solid to form, then the solution is saturated and may be left in contact with the solid. If not, add further solid.

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88 Stains for bacterial activity

Gram Stain For details on using this stain see Guidance Leaflet GL 95

To prepare the crystal violet solution, dissolve 2 g of the dye in 100 ml of industrial denatured alcohol (Solution A) and 1 g of ammonium ethanedioate in 100 ml of distilled/deionised water (Solution B). Add 25 ml of solution A to 100 ml of solution B.

To prepare the Safranin counterstain, dissolve 0.5 g of safranin in 100 ml of distilled/deionised water.

To prepare Gram’s (Lugol’s) iodine, dissolve 1 g of iodine and 2 g of potassium iodide in 300 ml of distilled or deionised water.

See Hazcards 32, 36A, 40A and 85. IDA is HIGHLY FLAMMABLE. There should be no sources of ignition in the vicinity Wear eye protection during the preparation. Ensure the room is well ventilated. Wear chemical-resistant gloves when preparing the crystal violet stain.

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89 Stains for cells

Aceto carmine Add 0.5 g of carmine to 55 ml of purified water in a

conical flask. Boil and add 45 ml of concentrated ethanoic acid. Plug the flask with cotton wool, boil again, cool and filter.

For ethanoic acid, see Hazcard 38. Label the stain CORROSIVE. Wear goggles. Use a fume cupboard.

Ehrlich’s haematoxylin for animal histology

Dissolve 0.64 g of haematoxylin in 32 ml of ethanol heated on a water bath, and filter. Separately, dissolve 0.32 g of aluminium potassium sulfate(VI) in 32 ml of purified water. Mix this solution with the haematoxylin solution and add 32 ml of propane-1,2,3-triol (glycerol) and 4 ml of glacial ethanoic acid. Leave exposed to daylight for 6 weeks to ‘ripen’ or add 1 ml of 0.2 M potassium manganate(VII) solution.

For aluminium potassium sulfate(VI), see Hazcard 3B. For ethanol; see Hazcard 40A. For ethanoic acid, see Hazcard 38. For propane-1,2,3-triol, see Hazcard 37. For ethanoic acid, see Hazcard 38. For potassium manganate(VII), see Hazcard 81. Wear eye protection and disposable nitrile gloves when making up the solution. Label the stain HIGHLY

FLAMMABLE.

Eosin for cytoplasmic staining

Dissolve 1 g eosin in 100 ml tap water. Used as a counterstain to Ehrlich’s haematoxylin. For alcoholic solution, substitute 75% ethanol for water.

Wear eye protection. Low hazard unless ethanol is used as the solvent.

Feulgen's stain for DNA

Dissolve 0.45 g of basic fuchsin in 87 ml of boiling water. Cool to 50 °C and add 30 ml of 1 M hydrochloric acid and 3 g of potassium metabisulfite. Leave in the dark for 24 hours to bleach. Add 0.5 g of decolourising charcoal, shake and filter into a stoppered bottle. Store in a cool, dark place.

For potassium metabisulfite, see Hazcard 97. For hydrochloric acid , see Hazcard 47A. Wear eye protection. Use a fume cupboard. No hazard label is required.

Leishman's stain for blood cells

Dissolve 0.15 g of the solid stain in 100 ml of methanol.

For methanol, see Hazcard 40. Prepare the solution in a fume cupboard wearing eye protection and disposable nitrile gloves. Label the solution TOXIC and

HIGHLY FLAMMABLE.

Methyl green pyronin for DNA and RNA

1 g of methyl green pyronin is dissolved in 100 ml of distilled/deionised water. DNA is stained green and RNA is stained red.

Wear eye protection and disposable nitrile gloves.

Methylene blue for cheek cells

Wear eye protection, and gloves to avoid staining the skin. Dissolve 0.1 to 1 g in 100 ml of water.

For methylene blue, see Hazcard 32. The solution is low hazard.

Orcein acetic (Acetic orcein) for chromosomes

The stock solution contains 2.2 g of orcein dissolved in 100 ml of glacial ethanoic acid. Dilute 10 ml of this solution with 12 ml of water before use. This diluted solution does not keep.

For ethanoic acid, see Hazcard 38. Wear goggles and chemical-resistant gloves. Use a fume cupboard. Label the concentrated solution CORROSIVE.

Sudan III or IV solutions for lipids

Dissolve 0.5 g of dye in 70 ml of ethanol and 30 ml of water, using a warm water bath, and filter.

Ethanol is HIGHLY FLAMMABLE (see Hazcards 32 and 40). Label the solution HIGHLY

FLAMMABLE. Wear eye protection.

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90 Stains for electrophoresis

Methylene blue is less sensitive than Azure A and not so convenient to use in electrophoresis but it is cheaper and adequate for some activities. Azure A for DNA Dissolve 0.04 g in 100 ml of 20% ethanol (ie, 0.04%

w/v) to apply after running the gel. Destaining is not needed.

Low hazard.

Colloidal Coomassie Blue for proteins

See www.ncbe.reading.ac.uk/NCBE/SCIENCEYEAR/recipes.html for a recipe, or purchase from this organisation. Note that this stain needs to be made up at least 24 hours before use, and requires the addition of sodium chloride solution to stabilise it.

Low hazard.

Methylene blue for DNA

Dissolve 0.05 g in 200 ml water (ie, make up to 0.025% w/v) to apply after running the gel. Destaining is needed.

See Hazcard 32, The solution is low hazard.

http://www.ncbe.reading.ac.uk/NCBE/SCIENCEYEAR/recipes.html�

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91 Stains for fungal material

Lactophenol Dissolve 10 g of phenol in 10 ml of water (do not heat). Add 10 ml of propan-1,2,3-triol (glycerol) and 10 ml lactic acid.

For propan-1,2,3-triol, see Hazcard 38C. For phenol, see Hazcard 70.Wear goggles. Wear disposable nitrile gloves. Use a fume cupboard. Label the solution TOXIC and CORROSIVE.

Cotton blue (aniline blue) for fungi

Dilute 8 ml of 0.1% cotton blue solution in lactophenol and 4 ml of 0.1% basic fuchsin solution in lactophenol with 88 ml of lactophenol.

See Hazcard 38C. Fuchsin is HARMFUL. Wear suitable eye protection and gloves. Label the solution TOXIC.

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92 Stains for metabolic activity

Janus green B (Diazine green) for mitochondria

Dissolve 0.3 g of the dye in 100 ml of purified water. Dilute this solution ten times with water before use. The colour change is blue (with oxygen) to pink (anaerobic). See Guidance Leaflet PS 89 for more information.

Wear eye protection and disposable nitrile gloves when making up the solution. No hazard label is required.

Methylene blue for cell contents

For living organisms: dissolve 1 g solid in 100 ml water and add 0.6 g sodium chloride. For dead tissue: dissolve 0.23 g solid in 23 ml of ethanol and dilute to 100 ml with water.

For ethanol, see Hazcard 40. For methyl blue see Hazcard 32. Wear eye protection and disposable nitrile gloves. No hazard label is required.

Neutral red Dissolve 0.1 g of neutral red in 100 ml of the appropriate isotonic saline solution (see Recipe Sheet 75). Dilute the solution ten times further with the saline solution.

Wear eye protection and disposable nitrile gloves.

Resazurin solution to test for milk freshness

Dissolve one tablet or 0.005 g of resazurin powder in 50 ml of water. (1 ml of this is added to 10 ml of a sample of milk. A colour change from blue to pink to white indicates how many bacteria are present.)

Resazurin is IRRITANT.

TTC, to show respiratory activity

Dissolve 1 g of 2,3,5-triphenyl tetrazolium chloride (TTC) in 100 ml of water. (A 0.5% solution is less expensive and gives just as good results but takes longer. It works well with maize seedlings.) Produces a red colouration.

Low hazard.

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93 Stains for plant material

Aniline (phenyl-ammonium) sulfate stain for lignin

Mix 89 ml of ethanol, 10 ml of 0.05 M sulfuric acid and 1 g of phenylammonium sulfate [aniline sulfate(VI)].

For phenylammonium sulfate(VI): see Hazcard 4. Wear eye protection and disposable nitrile gloves when making up the solution. Label the stain HIGHLY

FLAMMABLE.

FABIL for plant tissues

Prepare 3 solutions: 0.5 % solution of aniline blue in lactophenol, 0.5% solution of basic fuchsin in lactophenol and a solution containing 0.3 g of iodine and 0.6 g of potassium iodide in 100 ml of lactophenol. When required, mix in the proportions of 4:1:5 and allow to stand overnight. Filter before use. (Cell contents stain blue, cellulose walls stain light blue and lignin stains yellow.)

For lactophenol, see Hazcard 38C; for iodine, see Hazcard 54. Wear goggles and chemical-resistant gloves. Label the stain TOXIC.

Iodine stain Use 0.01 M iodine solution. See Recipe Sheet 50.

Phloroglucinol for pentoses and lignin

Dissolve 5 g of phloroglucinol (benzene-1,3,5-triol) in 75 ml of ethanol and 25 ml of water. Ligneous tissue should be well-flooded and staining continued for about 4 minutes after which 1 drop of concentrated hydrochloric acid should be added.

Phloroglucinol is an IRRITANT. Ethanol is HIGHLY FLAMMABLE (see Hazcards 12, 40 and 47). Label solution HIGHLY

FLAMMABLE. Wear eye protection.

Schulze’s solution for cellulose

Dissolve by warming 20 g of anhydrous zinc chloride in 8.5 ml of water and allow the mixture to cool. In a separate container, dissolve 1 g of potassium iodide and 0.5 g of iodine in 20 ml of water. Add this solution dropwise to the zinc chloride solution until iodine precipitate persists on agitation.

Zinc chloride is CORROSIVE and iodine is HARMFUL (see Hazcards 54 and 108). Wear eye protection, and chemical-resistant gloves, and carry out the procedure in a fume cupboard. Label the solution CORROSIVE.

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94 Standard solutions for titration

A primary standard solution is one with a concentration that is accurately known. (They do not have to be 0.1 M) They are used to determine the concentration of other reagents undergoing titration. Solids used should be of high purity (ie, more than 99.5% pure). Hydrated salts should not be used. The solid should not alter composition during weighing (which is why sodium hydroxide is not used; it absorbs water and carbon dioxide). Distilled or deionised water must be used. If possible, use a balance reading to 3 decimal places.

0.1 M sodium carbonate solution for standardising strong acid solutions

Procedure to prepare 1000 ml of solution

• Heat the anhydrous sodium carbonate in an evaporating basin with a gentle flame for 30 minutes and allow it to cool in a desiccator.

• Measure 10.599 g of solid into a clean 250 ml beaker and add 150 ml of boiled pure water. • Stir the solution until the solid dissolves and pour it via a funnel into a 1000 ml volumetric flask. • Rinse the stirring rod and the beaker with water, pouring the washings into the volumetric flask.

Repeat this twice more. • Add water via the funnel into the flask so it is just up to the required mark. Add the stopper and mix the

solution well. • Titrate against a strong acid using methyl red as the indicator. The colour of the endpoint is orange.

0.1 M potassium hydrogenphthalate solution for standardising strong alkali solutions


• Measure 20.433 g of the solid into a clean 250 ml beaker and add 150 ml of boiled pure water. • Stir the solution until the solid dissolves and pour it via a funnel into a 1000 ml volumetric flask. • Rinse the stirring rod and the beaker with water, pouring the washings into the volumetric flask.


solution well. • Titrate against a strong alkali using phenolphthalein as the indicator. The colour of the endpoint is pale

mauve which should last for at least 30 s.

0.1 M sodium ethanedioate solution for standardising potassium manganate(VII) solution


• Measure 13.40 g of solid into a clean 250 ml beaker. Add 150 ml of cold pure water. Stir the solution until the solid dissolves and pour it via a funnel into a 1000 ml volumetric flask.

• Rinse the stirring rod and the beaker with water, pouring the washings into the volumetric flask. Repeat this twice more.

• Add water via the funnel into the flask so it is just up to the required mark. Add the stopper and mix the solution well.

• Place the potassium manganate(VII) in the burette. Titrate against hot standard sodium ethanedioate solution until there is a permanent mauve colour.

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0.0167 M potassium iodate(V) solution for standardising sodium thiosulfate solution


• Dry the potassium iodate(V) at 180 °C for 30 minutes before cooling and using. • Measure 3.574 g of solid into a clean 250 ml beaker and add 150 ml of cold pure water. • Stir the solution until the solid dissolves and pour it via a funnel into a 1000 ml volumetric flask. • Rinse the stirring rod and the beaker with water, pouring the washings into the volumetric flask.


solution well. • To use, begin with a measured volume (10, 20 or 25 ml) of potassium iodate(V) solution. Add 10 ml of

1 M potassium iodide solution and 10 ml of 1 M sulfuric(VI) acid to produce iodine against which sodium thiosulfate is titrated. Use a 1% starch solution as the indicator when the solution is pale yellow. And from the results it is possible to calculate the concentration of the thiosulfate solution.

0.1 M calcium carbonate solution for standardising EDTA solution


• Dry the precipitated calcium carbonate at 150 °C for 1 hour. • Measure 10.00 g of cooled solid into a clean 250 ml conical flask and add enough 2 M hydrochloric

acid to dissolve it, placing a filter funnel over the flask to stop spray. • Make up this solution in a 1-litre volumetric flask.

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95 Strontium chloride

Strontium chloride hexahydrate will absorb water from the atmosphere. Formula: SrCl2.6H2O Molar mass: 266.6 g mol-1 Solubility: 88 g per 100 ml General Hazards See Hazcard 19A.

Procedure for preparing 100 ml of 0.1 M strontium chloride solution

• Wear eye protection. • Dissolve 2.67 g of strontium chloride-6-water in 70 ml of water. • Make up to 100 ml with pure water. • The solution is low hazard.

NB: If a school has only strontium carbonate, the solution can be made by adding 1.48 g of the carbonate to 100 ml of 0.1 M hydrochloric acid.

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96 Sulfur dioxide

For acid rain see the Recipe sheet 97. This is a particularly nasty gas to prepare and use. Do make sure that the procedure is trialled with help from a more-experienced colleague and that the fume cupboard is working satisfactorily.

General Hazards See Hazcards 47A, 78, 92 & 97. Sulfur dioxide is TOXIC by inhalation. There is no safe Workplace Exposure Limit for sulfur dioxide. As a guide, the HSE has advised that levels, must be less than 2.7 mg m-3 (ie, 1 ppm). Any preparation of sulfur dioxide should be carried out in a fume cupboard.

• Use a fume cupboard. Make sure it is working. Wear goggles. • Place between 10 and 20 g of sodium metabisulfite (HARMFUL) in a 250 cm3 conical flask. • Set up the equipment as shown below. • Pour enough 2 M hydrochloric acid (IRRITANT) to just cover the bottom of the thistle funnel tube. Place

some moist acidified potassium dichromate(VI) paper (TOXIC) above the gas jar. • Heat the conical flask gently and collect the gas by downward delivery. When the acidified potassium

dichromate(VI) paper turns green, the gas jar is full. • Add 5 M hydrochloric acid to the metabisulfite if more sulfur dioxide is required. • The leftover reagents can be poured down the fume cupboard sink with plenty of water. • It is possible to carry out the fountain experiment. A round-bottom flask is substituted for the gas jar.

Heat

Add at least 50 cm3 of2 M hydrochloric acid

Loose mineralwool plug

Acidified potassium dichromate(VI) paper

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97 Sulfur dioxide solution

Sulfur dioxide is a particularly nasty gas to prepare and use. Do make sure that the procedure is trialled with help from a more-experienced colleague and that the fume cupboard is working satisfactorily.

General Hazards See Hazcards 47A, 78, 92 & 97. Sulfur dioxide is TOXIC by inhalation. There is no safe Workplace Exposure Limit for sulfur dioxide. As a guide, the HSE has advised that levels must be less than 2.7 mg m-3 (ie, 1 ppm). Any preparation of sulfur dioxide should be carried out in the fume cupboard.

Method 1 • Use a fume cupboard. Make sure it is working Wear goggles. • Place between 10 and 20 g of sodium metabisulfite (HARMFUL) into a 250 cm3 conical flask. • Set up the equipment as shown below. The inverted filter funnel should be held just below the surface

of the water in the beaker. • Pour enough 2 M hydrochloric acid (IRRITANT) to just cover the bottom of the thistle funnel tube in the

flask. • Heat the conical flask gently to produce the gas, which will dissolve as it meets the water in the

beaker. The inverted funnel prevents suck back. • Add 5 M hydrochloric acid if more sulfur dioxide is required. • The leftover reagents can be poured down the fume cupboard sink with plenty of water.

Method 2 In a fume cupboard, dissolve 9.5 g of sodium metabisulfite (HARMFUL) in 100 ml of water. Add 100 ml of 0.5 M sulfuric(VI) acid and make up to 250 ml. Acid Rain Pupils should open containers in a fume cupboard and particular attention should be paid to known asthmatics. For acid rain, use 0.0001 M sulfuric(VI) acid rather than a sulphur dioxide solution. For an acidic atmosphere, use a 0.1 M solution of sodium metabisulfite in water. This will generate enough SO2 gas to provide an acid atmosphere.

Heat

Add at least 50 cm3 of2 M hydrochloric acid

Inverted funnel just below watersurface

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98 Sulfuric(VI) acid

The procedure below uses ice made from distilled or deionised water. Make sure you have a supply of this in the freezer. Do not make dilute solutions for the first time without seeking practical advice from a more-experienced colleague. Formula: H2SO4 Molar mass: 98.07 g mol-1 General Hazards See Hazcard 98A. Addition of acid to water produces a lot of heat. NEVER ADD

WATER TO THE ACID; serious accidents have occurred when this has been done.



0.01 M Ten-fold dilution of a 0.1 M solution with water -

0.1 M Ten-fold dilution of a 1 M solution with water -

0.4 M 11 22 54 -

1 M 27 54 136 IRRITANT

Battery acid 115 230 575 CORROSIVE

5M 135 270 680 CORROSIVE

Procedure

• Wear goggles (a face shield is preferable when handling large volumes) and chemical-resistant gloves.

• Measure out the indicated volume of concentrated sulfuric(VI) acid in a measuring cylinder. • Fill the beaker or laboratory jug half to two thirds full with ice, add 200 ml of water and a stirrer bar. • Set the stirrer running on a magnetic stirrer and add the concentrated sulfuric(VI) acid slowly onto the

ice. • Keep stirring the solution until the ice melts. • Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add


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99 Testing for gases

Ammonia Use damp red litmus paper

Damp red litmus goes blue. Universal indicator can be used as well.

Wear eye protection.

Place concentrated hydrochloric acid close by

White smoke of ammonium chloride (IRRITANT) forms. Wear eye protection.

Carbon dioxide Use lime water See Recipe Sheet 20 for calcium hydroxide. A white

precipitate of calcium carbonate forms which, in time, goes colourless as calcium hydrogencarbonate forms.

Wear eye protection if blowing into limewater through a straw.

Carbon monoxide Light it in a fume cupboard

The gas catches fire with a blue flame forming carbon dioxide.


Chlorine Use damp blue litmus paper

Damp blue litmus goes red on formation of chloric(I) acid. This then bleaches the paper.

Wear eye protection. Use a fume cupboard.

Dinitrogen monoxide Add a lighted or glowing splint

The gas supports combustion and can relight a glowing splint, although it is not as active as oxygen.


Hydrogen Add a lighted splint If the hydrogen is mixed with air there is a ‘pop’ but if

the gas is pure it catches fire and there is no ‘pop’. Wear eye protection.

Hydrogen chloride Use damp blue litmus paper

Damp blue litmus goes red. Universal indicator can be used as well.


Place concentrated ammonia solution close by

White smoke of ammonium chloride forms. Wear eye protection.

Hydrogen sulfide Moisten filter paper with 0.05 M lead(II) nitrate(V) solution

The paper immediately turns black with the formation of lead sulfide.

Wear eye protection. Use tongs or forceps to hold the paper.

Nitrogen There is no positive test for nitrogen.

Nitrogen dioxide Observe the colour The only brown gas there is. Wear eye protection.

Nitrogen monoxide Add the gas to air Brown nitrogen dioxide is formed. Wear eye protection.

Oxygen Add a glowing splint This relights a glowing splint, sometimes with a ‘pop’. Wear eye protection.

Sulfur dioxide Moisten filter paper with 0.05 M acidified potassium dichromate(VI) solution

The paper goes green with the formation of chromium(III) ions.

Wear eye protection. Use a fume cupboard.

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100 Testing for negative ions

Bromide ions 0.05 M silver nitrate(V) solution

Dissolve 0.85 g of silver nitrate(V) in 100 ml of distilled/deionised water. A cream precipitate forms which is soluble after the addition of concentrated ammonia.

See Hazcard 6 & 87. Wear eye protection.

Carbonate ions 2 M hydrochloric acid Adding the acid produces a lot of fizzing. The carbon

dioxide should be identifiable with lime water. See Hazcard 47A. Wear eye protection.

Chloride ions 0.05 M silver nitrate(V) solution

Dissolve 0.85 g of silver nitrate(V) in 100 ml of water, which must be pure. A white precipitate forms which is soluble after the addition of 2 M ammonia solution.


Ethanoate and methanoate ions 0.1 M iron(III) chloride solution

Add 2 M ammonia solution to 0.1 M iron(III) chloride solution until a precipitate just appears. Now add more of the original iron(III) chloride in drops with stirring until the precipitate just dissolves. Addition of this neutral iron(III) chloride to both ethanoates and methanoates gives a red solution.

See Hazcard 6 & 55C Wear eye protection.

Iodide ions 0.05 M silver nitrate(V) solution

Dissolve 0.85 g of silver nitrate(V) in 100 ml of distilled/deionised water. A yellow precipitate forms which is not soluble after the addition 2 M ammonia solution.


Nitrate(III) (nitrite) ions 2 M hydrochloric acid Adding the acid produces a pale blue liquid and a brown

gas (nitrogen dioxide, TOXIC). Immediately, add water to quench the reaction.

See Hazcard 47A, 68 and 93. Wear eye protection.

Nitrate(V) ions Brown ring test Dissolve the nitrate(V) in about 1 ml of water and add

0.5 g of iron(II) sulfate(VI)-7-water. Hold the test tube at an angle and carefully pour about 1 ml of concentrated sulfuric(VI) acid down the side of the test tube so it forms a layer at the bottom. A brown ring indicates nitrate(V) ions present.

Wear goggles. See Hazcards 55B & 98.

Devada’s alloy Add 2 ml of suspected nitrate(V) to 5 ml of 1 M sodium hydroxide solution. Add a little Devarda’s alloy (contains aluminium, copper and zinc) and warm. Test for ammonia gas.

Wear goggles. See Hazcard 1 & 91.

Phosphate ions 0.5 M ammonium molybdate(VI) solution (HARMFUL)

Dissolve 4 g of ammonium molybdate(VI) in 4 ml of concentrated ammonia and 6 ml of water. Add 12 g of ammonium nitrate(V) and dilute to 100 ml. When testing a sample, acidify the sample first with 0.5 ml of 1 M nitric(V) acid and then add 2 ml of the ammonium molybdate(VI) solution.

See Hazcards 6, 8, 9A & 67.

Sulfate(VI) ions 0.1 M barium chloride solution (HARMFUL)

Dissolve 2.44 g in 100 ml of distilled/deionised water. A white precipitate is formed. Sulfate(IV) ions do not give a precipitate but it can form in time due to oxidation by dissolved oxygen.

See Hazcard 10A.

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101 Testing for positive ions

Metal ions in general

Reagent Recipe and observations Hazards and notes 1 Flame tests Barium ions Greenish flame

Calcium ions Brick red flame Copper(II) ions Bluish flame Lead(II) ions White flame Lithium Crimson red flame Potassium ions Lilac flame (best seen thro’ cobalt glass) Rubidium ions Blue flame Strontium ions Crimson flame Sodium ions Persistent yellow flame.

See Hazcards 10A, 19A & 47B. The Hazcards also have the procedure for the tests. There is also the borax bead test on Hazcard 14.

2 Add 0.4 M sodium hydroxide to a small amount of the solution under test

Dissolve 1.6 g of sodium hydroxide in 100 ml of water. Precipitates form, some coloured and others colourless, some of which dissolve in excess alkali solution. Aluminium ions White precipitate which dissolves in excess alkali Ammonium ions No precipitate but odour of ammonia Barium ions White precipitate Calcium ions White precipitate Cobalt(II) ions Blue precipitate Copper(II) ions Blue precipitate Iron(II) ions Green precipitate Iron(III) ions Brown precipitate Lead(II) ions White precipitate which dissolves in excess alkali Magnesium ions White precipitate Nickel(II) ions Green precipitate Silver ions Brown precipitate Zinc ions White precipitate which dissolves in excess alkali Potassium ions No precipitate Sodium ions No precipitate

See Hazcard 91. The concentration of the metal solutions needs to be 0.1 M.

3 0.1 M sodium, potassium or ammonium sulphide solution

Dissolve 2.4 g of sodium sulfide-9-water or 1.0 g of potassium sulfide in 100 ml of water. 4 ml of ammonium sulfide solution can be diluted to 100 ml with water. Some metal ions form precipitates with the solutions. Refer to an (old!) A-level text for details.

See Hazcard 51. The test solutions are IRRITANT. Use a fume cupboard as the odour is repulsive.

Aluminium ions Alizarin solution Dissolve 0.1 g of alizarin in 100 ml of water. Addition of 2 M

ammonia solution to an aluminium salt in the presence of alizarin gives a pink precipitate or ‘lake’.

See Hazcard 32.

Ammonium ions 0.1 M sodium hexanitrocobalt-ate(III) solution (HARMFUL)

Dissolve 4.04 g in 100 ml of water. A yellow precipitate forms with ammonium ions.

See Hazcard 95A. It is an unstable solution with a maximum shelf life of 3 weeks.

0.4 M sodium hydroxide solution (IRRITANT)

Warm with the test solution. Ammonia gas will be given off.


Calcium ions 0.25 M ammonium ethanedioate

Dissolve 3.5 g of ammonium ethanedioate in 100 ml of water. Ammonium ethanedioate gives a white precipitate of calcium ethanedioate. The precipitate can be filtered off and dried. It decomposes under heat to give calcium oxide. The oxide can be weighed to estimate quantitatively the amount of calcium present in a sample.

See Hazcard 36A.

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Cobalt(II) ions Thiooxamide (rubeanic acid) (HIGHLY FLAMMABLE, HARMFUL)

Dissolve 0.5 g in 100 ml of ethanol. A brown precipitate is produced with cobalt ions.

See Hazcards 35 and also 40A.

0.1 M ammonium or potassium thiocyanate

Dissolve 0.76 g of ammonium thiocyanate or 0.97 g of potassium thiocyanate in 50 ml of water and make up to 100 ml. A blue solution forms with cobalt(II) ions.

See Hazcard 9B or 95C.

Copper(II) ions 2M ammonia solution Dilute 11 ml of 880 ammonia to 100 ml with water. A blue

precipitate is formed which dissolves in excess ammonia solution to form a deep blue solution.

See Hazcards 6. Prepare in a fume cupboard.

Thiooxamide (rubeanic acid) (HIGHLY FLAMMABLE, HARMFUL)

Dissolve 0.5 g in 100 ml of ethanol. A black precipitate forms with copper(II) ions.



Dissolve 0.76 g of ammonium thiocyanate or 0.97 g of potassium thiocyanate in water 50 ml of water and make up to 100 ml. A black precipitate forms.

See Hazcard 9B or 95C. The solution is LOW HAZARD.

Iron(II) ions 0.1 M potassium hexacyanoferrate(III)

Dissolve 3.29 g in 100 ml of water. A light blue precipitate forms.


1,10 phenanthroline solution

Dissolve 1.49 g of 1,10 phenanthroline in 100 ml of water. The reagent gives a red colour when mixed with iron (III) ions.

See Hazcard 55B. Solid Phenanthroline is TOXIC if swallowed. No hazard warning is required on the solution.

Iron(III) ions 0.1 M potassium hexacyanoferrate(III)

Dissolve 3.29 g in 100 ml of water. A dark blue precipitate forms.


Ferroxyl indicator (HIGHLY FLAMMABLE)

Dissolve 2 g of sodium chloride, 0.1 g potassium hexacyanoferrate(III) and 1.0 g of phenolphthalein in 50 ml of ethanol, and mix with 100 ml of water. Very useful for detecting the rusting process.

See Hazcards 32, 40A and 79.


Dissolve 0.76 g of ammonium thiocyanate or 0.97 g of potassium thiocyanate in 50 ml of water and make up to 100 ml. A dark red solution forms. [NB there is no colouration with iron(II) ions]

See Hazcard 9B or 95C. The solution is LOW HAZARD.

Lead(II) ions Dithizone Dissolve 0.005 g in 100 ml of dichloromethane or

methylbenzene. The reagent gives a red colouration with lead(II) ions.

See Hazcards 28, 32 and 46. Label the solution according to which solvent is used.

Potassium iodide Prepare a 0.1 M solution. A bright yellow precipitate forms.

See Hazcard 47B.

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Nickel(II) ions Thiooxamide (rubeanic acid) (HIGHLY FLAMMABLE, HARMFUL)

Dissolve 0.5 g in 100 ml of ethanol. A violet precipitate forms.


Butanedione dioxime (dimethylglyoxime) (HIGHLY FLAMMABLE & HARMFUL)

Dissolve 1 g in 100 ml of ethanol. A red precipitate forms. See Hazcard 53.

Potassium ions 0.1 M Sodium hexanitrocobalt-ate(III) (HARMFUL)

Dissolve 4.04 g in 100 ml of water. A yellow precipitate forms.

See Hazcard 95A. It is an unstable solution with a maximum shelf life of 3 weeks.

Zinc(II) ions 2M ammonia solution Dilute 11 ml of 880 ammonia to 100 ml with water. A white

precipitate forms, which dissolves in excess ammonia solution to form a colourless solution.

See Hazcards 6. Prepare in a fume cupboard. The solution is LOW HAZARD.

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102 Testing for organic functional groups

Alcohols - primary, secondary & tertiary

Lucas reagent (CORROSIVE)

Dissolve 68 g of anhydrous zinc chloride in 43 g of concentrated (36%) hydrochloric acid. The time taken for turbidity to appear is used to differentiate between the three classes of alcohols: • no visible reaction: primary alcohol, • solution turns cloudy in 3 - 5 minutes: secondary alcohol, • solution turns cloudy immediately, and/or phases separate:

tertiary or benzyl alcohol.

See Hazcards 47A & 108A.

Potassium dichromate(VI) test

Identifies primary and secondary alcohols. To five drops of the alcohol, add 10 drops of 1 M sulfuric(VI) acid and 4 drops of 0.1 M potassium dichromate(VI) solution. Place the solution in boiling hot water from a kettle. The colour change is orange to green.

See Hazcard 78 & 98A.

Potassium manganate(VII) test

Identifies primary and secondary alcohols. To five drops of the alcohol, add 10 drops of 1 M sulfuric(VI) acid and 4 drops of 0.002 M potassium manganate(VII) solution. Place the solution in boiling hot water from a kettle. The colour change is purple to colourless.

See Hazcard 81 & 98A.

Aldehydes and ketones

Hydrogen sulfite test

Dissolve 20 g of sodium metabisulfite in 100 ml of hot water. White precipitate appears with the reagent is added to a carbonyl compound.

See Hazcard 92.

Tollen’s reagent It does not work with ketones. This reagent is ammoniacal silver nitrate(V), it must be made immediately before use and not stored. The breakdown products are EXPLOSIVE. Add one drop of 0.4 M sodium hydroxide solution to 1 ml of 0.1 M silver nitrate(V). Add 1 M ammonia solution until the precipitate just redissolves. On warming with an aldehyde, a silver mirror forms.

See Hazcard 87. Immediately dispose of liquid after use down the foul-water drain.

Schiff’s reagent It does not work quickly with ketones. Dissolve 0.1 g of fuchsin in 100 ml of water and add 0.9 g of sodium metabisulfite followed by 2 ml of 2 M nitric(V) acid, so it is colourless. Addition of an aldehyde restores the colour. Be aware that sulfur dioxide, a TOXIC gas, may be evolved.

See Hazcards 32, 67 & 92. Wear eye protection.

The iodoform test Works with methyl ketones (CH3CO-), methyl secondary alcohols, ethanol and ethanal. To fives drops of the liquid under test, add 5 drops of 0.01 M iodine, followed by enough 1 M sodium hydroxide solution until the brown colour is removed. Warm in boiling water bath and look for a yellow precipitate of iodoform.

See Hazcards 54A & 91.

Carboxylic acids

‘Neutral’ iron(III) chloride

Add 2 M ammonia solution to 0.1 M iron(III) chloride solution until a precipitate just appears. Now add more of the original iron(III) chloride in drops with stirring until the precipitate just dissolves. Add this to the liquid under test, and look for a red solution. [Methanoic acid reacts with acidified potassium manganate(VII) and potassium dichromate(VI) in a similar manner to alcohols.]

See Hazcards 6 & 55C.

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114 © CLEAPSS 2011

Double bonds or unsaturation Bromine water Add 0.002 M bromine water which will go clear. See Hazcard 15B.

Potassium manganate(VII) (acidified)

Add an acidified 0.002 M solution of potassium manganate(VII) which will go clear. If it goes brown, it is not acidified enough.


Potassium manganate(VII) (alkaline)

Alkaline potassium manganate(VII) solution is not stable so it must be made in situ. Add enough solid anhydrous sodium carbonate to make the acidified solution alkaline. Add alkaline 0.002 M solution of potassium manganate(VII) which should go green.


OH groups in alcohols and acids Phosphorus pentachloride

To about 2 ml of the liquid or solid under test, add phosphorus pentachloride with a small spatula. There is a vigorous reaction and hydrogen chloride is produced.

See Hazcard 74.

Phenol ‘Neutral’ iron(III) chloride

Add 2 M ammonia solution to 0.1 M iron(III) chloride solution until a precipitate just appears. Now add more of the original iron(III) chloride in drops with stirring until the precipitate just dissolves. Add this to the material under test, and look for a purple colouration.

See Hazcards 6 & 55C.

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103 Tin(II) chloride

Also known as stannous chloride. Solutions stronger than 0.2 M will be difficult to prepare. This reagent has to be dissolved in concentrated hydrochloric acid before diluting further with water. The final solution will be 1 M with respect to hydrochloric acid. The solution may be cloudy, so either leave it for some hours before decanting off the clear solution or filter through fine filter paper. Formula: SnCl2.2H20 Molar mass: 225.63 g mol-1 Solubility: 84 g per 100 ml General Hazards See Hazcards 102 & 47A.





0.1 M 2.26 5.64 22.56 -

Procedure

• Wear goggles and chemical-resistant gloves. Use a fume cupboard. • Measure out the indicated quantity of tin(II) chloride-2-water. • Add the solid to about 10% of the final volume of concentrated hydrochloric acid in a beaker. • Stir to dissolve. Do not warm the solution. • Pour the solution into a measuring cylinder or laboratory jug and dilute to the final volume with

distilled/deionised water. • Leave to clear. • Pour the solution into a labelled bottle and mix well.

CLEAPSS Recipe Book

116 © CLEAPSS 2011

104 Water (sea and hard)

It is possible to obtain sea-water salt from several suppliers. For acid rain, see Recipe Sheet 97.

A. Sea water

Dissolve the following salts in 250 ml of pure water and dilute to 1000 ml.

• 23.99 g of sodium chloride • 0.74 g of potassium chloride • 2.24 g of calcium chloride-6-water • 10.89 g of magnesium chloride-6-water • 4.01 g of anhydrous sodium sulfate(VI) [OR 9.10 g of sodium sulfate(VI)-10-water] • 0.20 g of sodium hydrogencarbonate • 0.09 g of sodium bromide • 0.03 g of boric acid (if required) • 0.01 g of strontium chloride (if required)

B. Permanent hard water • Measure 0.45 g of calcium sulfate(VI)-2-water and add it to 1000 ml of water in a bottle and leave it

overnight to dissolve.

C. Temporary hard water • Filter 130 ml of freshly-made calcium hydroxide solution (see Recipe Sheet 20). • Pass carbon dioxide though the solution so that calcium carbonate first precipitates and then, with

further carbon dioxide, re-dissolves to form calcium hydrogencarbonate. This may take some time. • Dilute this clear solution to 1000 ml.

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© CLEAPSS 2011 117

105 Winkler’s method for dissolved oxygen

This technique is better than other methods (eg, using phenosafranine) because the oxygen in the water is ‘fixed’ once the manganese(II) hydroxide is formed. However, the solutions are hazardous to prepare and use, and should be added to the water sample as soon as possible. General Hazards See Hazcards 60, 91 & 98. Great care must be taken in making the 8 M sodium

hydroxide solution. Gloves and eye protection (a face shield is preferable) should be used. The preparation is very slow and must not be rushed.

Preparation of the solutions used in Winkler’s method

100 ml of 8 M

sodium hydroxide

solution

• Wear goggles and chemical-resistant gloves. Consider using a fume cupboard. • Add 8 g of sodium hydroxide pellets to 70 ml of water in a 250 ml beaker. • Once it has dissolved, cool the solution in an ice bath. • When the temperature of the solution has dropped to below room temperature

add another 8 g. Stir until it has dissolved and cool again. • Repeat this procedure until a total of 32 g of sodium hydroxide has been added. • Add the solution to a 100 ml measuring cylinder and add water to the 100 ml

mark. • Stir carefully into this solution 14 g of potassium iodide. • Pour the solution into a labelled bottle and, when cool, mix well. Attach a

CORROSIVE hazard warning.

The manganese(II)

solution

• Dissolve 50 g of manganese(II) sulfate(VI)-4-water or 45 g of manganese(II) chloride-4-water in 80 ml of pure water and dilute to 100 ml.

• Pour it into a labelled bottle, mixing well. Add a HARMFUL hazard warning.

For the titration • Prepare 0.01 M sodium thiosulfate solution (see Recipe Sheet 87) which can be standardised against potassium iodate solution (see Recipe Sheet 94) for accurate work.

The indicator • Prepare 1% starch solution. See Recipe Sheet 49.

Winkler’s method for dissolved oxygen Procedure for using these solutions

• Wear goggles. • Collect 250 ml of water in a 250 ml stoppered bottle, ensuring there is no air trapped inside. • Use a 1 ml pipette (or marked teat pipette) to transfer 1 ml of the manganese(II) solution to the water

sample, injecting the liquid below the surface of the water, and taking care not to blow any more air into the water.

• Use a similar technique to transfer 1 ml of the alkaline potassium iodide solution to the water sample. • Replace the stopper on the bottle without trapping any air and agitate the bottle to mix the reagents. • Allow the contents to stand for 10 minutes. • Now the brown manganese(III) hydroxide has precipitated, the contents of the flask can be transferred

to a larger flask or bottle. • Add 1.5 ml of concentrated sulfuric(VI) acid which will dissolve the precipitate and liberate iodine. • Titrate this solution with the standardised sodium thiosulfate solution, adding 2 ml of starch solution

near the end-point. (1 ‘mol’ of oxygen is equivalent to 4 ‘mol’ of thiosulfate solution.)

CLEAPSS Recipe Book

118 © CLEAPSS 2011

106 Zinc sulfate(VI)

On standing, zinc sulfate(VI)-7-water loses water of crystallisation slowly to the air and becomes powdery. However, it is more convenient to make solutions from the hydrated salt than the anhydrous salt which is also available from suppliers. The anhydrous salt readily absorbs water from the atmosphere during storage. Formula: ZnSO4.7H2O Molar mass: 287.54 g mol-1 Solubility: 96 g per 100 ml General Hazards Zinc sulfate(VI) is HARMFUL. See Hazcard 108B.




0.1 M 2.88 7.19 28.75 -

1.0 M 28.75 71.88 287.54 HARMFUL


Procedure

• Measure out the indicated quantity of hydrated zinc sulfate(VI). • Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug. • Stir to dissolve, warming if necessary. • Either pour the solution from the beaker into an appropriate measuring cylinder and add water to the

required level or add water to the required level in the laboratory jug. • Pour into a labelled bottle.

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© CLEAPSS 2011 119

Index Name Recipe

Sheet

Acetic acid 39

Acetic alcohol 41

Acetic orcein 89

Aceto carmine 89

Acid atmosphere 97

Acid base indicators 46

Acid rain 97

Agar 1

Alcohol tests 102

Alcohol/water solutions 2

Alcoholic potassium hydroxide

71

Alcoholic solutions 2

Aldehyde tests 102

Alginate beads 3

Alizarin solution 101

Alkaline pyrogallol 84

Alum 4

Aluminium chloride 4

Aluminium etching 38

Aluminium potassium sulfate(VI)

4

Aluminium solutions 4

Amino acid chromatography

26

Amino acid tests 13

Ammonia gas 5

Ammonia solution 6

Ammonia test 99

Ammonium cerium(IV) sulfate(VI)

22

Ammonium chloride 7

Ammonium ethanedioate

101

Ammonium hydroxide 6

Ammonium iron(III) citrate

52

Ammonium iron(III) sulfate(VI)

52

Ammonium sulfide 101

Ammonium thiocyanate 101

Ammonium vanadate 8

Amylase 37

Analgesics chromatography

26

Andrews arrangement for gas preparation

24

Name Recipe Sheet

Aniline blue 91

Aniline sulfate(VI) 93

Animal tissue preservative

74

Anion tests 100

Aspirator 20

Azo dyes 9

Azure A 90

Barium chloride 10

Barium diphenylamine-4-sulfonate indicator

49

Barium nitrate(V) 10

Barium solutions 10

Basic agar 1

Belousov-Zhabotinski reaction

63

Benedict’s qualitative reagent

11

Benedict’s quantitative reagent

12

Bile salts 37

Biological buffers 18

Biro ink chromatography

26

Bismuth nitrate(V) 14

Biuret reagent 15

Bleach 81

Blue-bottle reaction 63

Blueprints 52

B-R reaction 63

Brady’s reagent 33

Briggs-Rausher reaction 63

Brodie’s fluid 16

Bromide ion test 100

Bromine water 17

Bromocresol green 46

Bromophenol blue 46

Bubble mixture 79

Buffers 18

Butanedione dioxime 101

B-Z reaction 63

Calcium carbonate (standard solution)

94

Calcium chloride 19

Calcium hydroxide 20

Calcium nitrate(V) 19

CLEAPSS Recipe Book

120 © CLEAPSS 2011

Name Recipe Sheet

Carbohydrate tests 13

Carbon dioxide 99

Carbon dioxide gas 21

Carbon dioxide indicator 48

Carbon monoxide preparation

42

Carbon monoxide test 99

Carbonate ion test 100

Carbonyl group test 33

Carboxylic acid test 102

Cation tests 101

Cellular respiration 13

Cerium(IV) solutions 22

Cerium(IV) sulfate(VI) 22

Cheek cells 89

Chemical garden 86

Chemiluminesence 23

China blue agar 1

Chloride ion test 100

Chlorine gas preparation

24

Chlorine test 99

Chlorine water 25

Chlorophyll chromatography

26

Chromatography solvents

26

Chrome alum 27

Chromium(III) chloride 27

Citric acid 28

Clarke’s fluid 41

Clarke’s soap solution 79

Clock reactions 29

Cobalt(II) chloride 30

Cobalt(II) nitrate(V) 30

Cobalt(II) thiocyanate 30

Cole’s reagent 13

Colloidal Coomassie blue

90

Complexometric indicators

49

Coomassie blue 90

Copper electroplating 36

Copper etching 38

Copper(II) chloride 31

Copper(II) nitrate(V) 31

Copper(II) solutions 31

Copper(II) sulfate(VI) 31

Name Recipe Sheet

Cotton blue 91

Cotton dying 9

Cresol red 46

Crude oil alternative 32

Cytological fixative 41

DCPIP 13

Diammonium iron(II) sulfate(VI)

51

Diazine green 92

Diazonium salts 9

2,6- dichlorophenol indo-phenol

13

Dichromate(VI) titration indicators

49

Dimethylglyoxime 101

Dinitrogen monoxide preparation

42

Dinitrogen monoxide test

99

2,3,5- Dinitrophenylhydrazine 33

3,5- Dinitrosalicylic acid 34

Dipotassium hydrogen phosphate(V)

67

Dipotassium hydrogen phosphate(V) -3- water

67

Disappearing cross reaction

29

Disinfectants 81

Disodium hydrogen phosphate(V)

67

Disodium hydrogen phosphate(V) -7- water

67

Dissolved oxygen 105

Dithizone 101

2,4- DNP 33

DNS 34

DNSA 34

Double bond test 102

Drosophila food base 35

Dyes 9

Dying cotton 9

EDTA titration indicators 49

Ehrlich’s haematoxylin 89

Eiosin indicator 49

Electrolytic agar 1

Enzymes 37

Enzymes immobilised 3

Eosin 89

Eriochrome black 49

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Name Recipe Sheet

Etching solutions 38

Ethanoate ion test 100

Ethanoic acid 39

Ethanol/water mixtures 2

Ethanolic potassium hydroxide

71

FAA 41, 74

FABIL 93

Fehling's solution 40

Ferroin indicator 49

Ferroxyl agar 1

Ferroxyl indicator 101

Feulgen's stain 89

Fixatives in biology 41

Flame tests 101

Fluorescein indicator 49

Formaldehyde 58

Formalin alcohol fixative 41

Formalin-aceto fixative 41

Formalin-aceto-alcohol 74

Formic acid 59

Glucose nutrient agar 1

Glucose quantitative test

12

Glucose test 11, 34

Gram Stain 88

Harcourt-Essen reaction 29

Hard water 104

Home-made indicators 46

Hydrochloric acid 43

Hydrogen 99

Hydrogen carbonate indicator

48

Hydrogen chloride preparation

42

Hydrogen chloride test 99

Hydrogen gas 44

Hydrogen peroxide 45

Hydrogen peroxide/ potassium iodide reaction

29

Hydrogen sulfide preparation

42

Hydrogen sulfide test 99

Hydrogen-sulfite test 102

2- hydroxy-1,2,3-tricarboxylic acid

27

Immobilised enzymes 3

Name Recipe Sheet

Indicator agar 1

Iodide ion test 100

Iodine solution 50

Iodine stain 93

Iodoform test 102

Iron etching 38

Iron(II) chloride 51

Iron(II) solutions 51

Iron(II) sulfate(VI) 51

Iron(III) chloride 52

Iron(III) ions and iodide ion clock reaction

29

Iron(III) solutions 52

Iron(III) sulfate(VI) 52

Janus green B 13, 92

Kaiserling’s preservative

74

Ketone tests 102

Knop’s culture solution 66

Lactophenol 91

Landolt iodine reaction 29

Lead etching 38

Lead nitrate(V) 53

Leishman's stain 89

Lime water 20

Lipase 37

Lipstick chromatography

26

Lithium chloride 54

Litmus 46

Locating agents 26

Lucas reagent 102

Lugol’s iodine 88

Luminol 23

Magnesium sulfate(VI) 55

Malt agar 1

Manganese(II) sulfate(VI)

56

Manometric fluid 16

Mayonnaise agar 1

Mercury(I) nitrate(V) 57

Mercury(II) chloride 57

Metal ion chromatography

26

Methanal solution 58

Methanoate ion test 100

Methanoic acid 59

Methanol solution 2

CLEAPSS Recipe Book

122 © CLEAPSS 2011

Name Recipe Sheet

Methanolic potassium hydroxide

71

Methyl benzoate nitration

26

Methyl green pyronin 89

Methyl orange 46

Methyl orange preparation

9

Methyl red 46

Methylene blue 13, 89, 90, 92

Milk agar 1

Millon’s reagent 13

Mohr’s salt 51

Molisch’s solution 13

Murexide 49

Neutral Red 46, 92

Neutral iron(III) chloride 102

Nickel electroplating 36

Nickel(II) sulfate(VI) 60

Ninhydrin 13

Nitrate(III) ion test 100

Nitrate(V) test 100

Nitration of Methyl benzoate

26

Nitric(V) acid 61

Nitrogen preparation 42

Nitrogen test 99

Nitrogen dioxide preparation

42

Nitrogen dioxide test 99

nitrogen gas oscillating reaction

63

Nitrogen monoxide preparation

42

Nitrogen monoxide test 99

N-phenylanthranilic acid indicator

49

Nutrient agar for bacteria

1

Nylon rope experiment 62

OH group test 102

Orange II preparation 9

Orcein acetic 90

Organic functional group tests

102

Orthophosphoric acid 65

Oscillating reactions 63

Oxygen preparation 64

Oxygen test 99

Name Recipe Sheet

Oxygen levels in water 105

PAS reaction 13

Pepsin 37

Periodic acid Schiff reaction

13

Permanently hard water 104

pH buffers 18

pH probe calibration buffers

18

Phenanthroline solution 101

Phenol red 46

phenol-indodichlorophenol

13

Phenolphthalein 46

Phenolphthalein indicator agar

1

Phenylammonium sulfate(VI)

93

Phenylthiocarbamide 13

Phenylthiourea 13

Phloroglucinol 93

Phosphate(V) ion test 100

Phosphoric(V) acid 65

Phosphorus pentachloride

102

Physiological saline solutions

75

Plant mineral requirement solutions

66

Plant tissue preservative

74

Polypeptide tests 13

Potassium chloride 68

Potassium chromate 69

Potassium dichromate(VI)

70

Potassium dihydrogen phosphate(V)

67

Potassium hexacyanoferrate(III)

101

Potassium hydrogen phthalate (standard solution)

94

Potassium hydroxide 71

Potassium iodate (standard solution)

94

Potassium iodate/sodium metabisulfite reaction

29

Potassium iodide 72, 101

Potassium iodide/potassium persulfate clock reaction

29

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Name Recipe Sheet

Potassium manganate(VII)

73

Potassium phosphate(V)

67

Potassium sulfide 101

Potassium thiocyanate 101

Precipitation indicators 49

Preservatives for animal tissues

74

Propanone solution 2

Protein tests 13

PTC 13

PTU 13

PVA 78

Pyrogallol 84

Red cabbage indicator 46

Redox indicators 49

Reducing sugar test 11, 12

Resazurin solution 93

Ringer’s solution 75

Rubeanic acid 101

Sach’s culture solutions 66

Safranin counterstain 88

Sakaguchi test 13

Saliva 37

Sandell’s solution 76

Schiff’s reagent 102

Schulze’s solution 93

Screened methyl orange

46

Sea water 104

Silicate gels 86

Silver electroplating 36

Silver nitrate(V) 77

Silver nitrate(V) titration indicators

49

Slime 78

Soap 79

Sodium carbonate 80

Sodium carbonate (standard solution)

94

Sodium chlorate(I) 81

Sodium chloride 82

Sodium dihydrogen phosphate(V)

67

Sodium dihydrogen phosphate(V) -1- water

67

Sodium diphenylamine-4-sulfonate indicator

49

Name Recipe Sheet

Sodium ethandioate (standard solution)

94

Sodium ethanoate 83

Sodium hexanitrocobaltate(III)

101

Sodium hydrogencarbonate

84

Sodium hydroxide 85

Sodium hydroxide test 101

Sodium hypochlorate 81

Sodium phosphate(V) 67

Sodium phosphate(V) -12- water

67

Sodium silicate 86

Sodium sulfide 101

Sodium tauroglycocholate

37

Sodium thiosulfate 87

Solochrome black 49

Stains for cells 89

Stains for electrophoresis

90

Stains for fungal material

91

Stains for metabolic activity

92

Stains for plant material 93

Standard solutions 94

Starch agar 1

Starch indicator 49

Starch malt agar 1

Sterilising conditions 1

Strontium choride 95

Sudan III or IV solutions 89

Sugar chromatography 26

Sulfate(IV) ion test 100

Sulfate(VI) ion test 100

Sulfur dioxide preparation

96

Sulfur dioxide test 99

Sulfur dioxide solution 97

Sulfuric(VI) acid 98

Technical agar 1

Temporary hard water 104

Tests for gases 99

Tests for negative ions 100

Tests for positive ions 101

Thermochromic liquid 30

Thiooxamide 101

CLEAPSS Recipe Book

124 © CLEAPSS 2011

Name Recipe Sheet

Thiosulfate/acid reaction

29

Thiosulfate/iodine titration indicator

49

Thymol blue 46

Thymolphthalein 46

Tin(II) chloride 103 Tollen’s reagent 102

2,3,5- Triphenyl tetrazolium chloride

13

Trypsin 37

TTC 13

Universal buffer 18

Universal indicator 47

Name Recipe Sheet

Unsaturation in organic chemistry

102

Vitamin C test 13

Wanklyn’s soap solution 79

Washing soda 80

Water 104

Water solutions for plants

66

Winkler’s method 105

Yamada indicator 47

Zinc electroplating 36

Zinc sulphate 106

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