Nutrient Analysis Using Colorimetry
Torrey LindboSWRP, PSU-ESR
Safety Equipment
GlovesGogglesMaterial Safety Data Sheets
Other Considerations: Alcohol Hand Wash Table or Tarp (spill prevention)
Colorimetry
Colorimetry: method for quantifying the concentration of a chemical constituent by analyzing change in color Based on color change for standard
solutions of known concentration Light usually filtered to specific
wavelength
Visible Light Spectrum = ROYGBIV Red ~ 700 nm Violet ~ 400 nm
Colorimeter
Color change measured using colorimeterColorimeter determines amount of transmitted light absorbed by sample
Transmitted Light Detector
Light sourceTungsten Lamp
Lens Sample VialFilter
Light Paths
Refracted
Absorbed
Transmitted
Reflected
Absorbance (A) is logarithmically related to transmittance (T)A = - log T
Beer’s Law
A = εLcWhere,A = absorbanceε = absorptivity; experimentally
derived constant for each substrateL = path length of light through samplec = concentration of substance in
solution
Beer’s Law
Concentration (c)
A = εLc
Absorbance (A)
Slope = εL
Hach DR/850 Colorimeter
Has pre-programmed filters and calibration curvesCap doubles as light coverSample color change compared to “blank” – what ZERO looks likeBlank may be stream sample with no reagent added, or it may be deionized water with reagent
7 9
4 5
1
I/0
2
8
6
3
0 READ
DR/850 Colorimeter
TIMER Enter
Hach DR/700 Colorimeter
Each test requires a “module” that needs to be turned to the correct program number. Example: Nitrate module = 50.01 Low range procedure = program 50.07.1 High range procedure = program 50.05.1
Need to perform Lamp Intensity AdjustmentUse film canister as light cover
“Zeroing” the Colorimeter
Concentration (c)
Beer’s LawA = εLc
c = εL / A
A
Slope = εL(constant)
SampleAbsorbance
BlankAbsorbance
Other DR/850 Features
Timer Beeps when reaction periods for tests are
over Limits colorimeter use; no other tests can
be run while timer is running
Store Can store up to 99 readings
Download Hach makes infrared computer interface
Hach Glassware
Cleaning is crucial first stepAcid Washing Soak in ~10% acid bath Choose acid depending upon
tests (i.e., don’t use nitric acid if you’re testing for nitrate)
Rinse thoroughly with DI water Etching on Hach glassware
may erode after long soaks10 mL
20 mL
25 mL
Colorimetric Analyses
Soluble Reactive Phosphorus (SRP)Total PhosphorusAmmonia NitrogenNitrate NitrogenChlorine
Phosphorus
Clean glassware crucialSRP test is quick and easy (2 minute reaction time) – good for flexibility with colorimeter sharingTotal Phosphorus requires digestion Hot plate Strong Acid and Base Requires fume hood This test should be run in the lab
Phosphorus
Equipment Needed 3-5 vials for samples + 1 for blank PhosVer3 reagent powder pillows Non-toxic waste container DR/850 Colorimeter
Procedure Overview 10 mL sample + reagent in each sample 10 mL sample and no reagent for blank Shake and wait 2 minutes
The Meniscus
Meniscus: the curved, lens-like surface of water in glassGlass has a negative charge that attracts the positive H from waterRead volumes from bottom of meniscus
10 mL
Nitrate Nitrogen
Vigorous Shaking2 Procedures: Low (< 0.5 mg/L) and High (0.6 – 20 mg/L) RangeToxic waste – Cadmium (heavy metal) used to reduce nitrateTime requirement (i.e., Low Range takes 20+ mins.) Shake 3 min, wait 2, add reagent 2,
shake, wait 15 min.
Low Range Nitrate Procedure
Equipment Needed: 6-10 vials for samples + 1 for blank NitraVer6 and NitriVer3 reagent
powder pillows
Low Range Procedure: 30 mL sample + NitraVer6 in each
sample (shake 3 mins) 10 mL treated sample + NitriVer3 10 mL sample and no reagent for
blank
High Range Nitrate Procedure
Equipment Needed: 3-5 vials for samples + 1 for blank NitraVer5 reagent powder pillows Toxic waste container
High Range Procedure: 10 mL sample + NitraVer5 in each
sample 10 mL sample and no reagent for blank
Ammonia Nitrogen
Timing – measure each sample immediately after reaction time Time table; stagger reagent addition
to each vial by 1 minute
Blank is deionized water + reagents (only need 1 blank) DI water source must be PURE
Ammonia Nitrogen
Equipment Needed: 3-5 vials for samples + 1 for blank Ammonia Salicylate and Ammonia
Cyanurate reagent powder pillows Toxic waste container
Procedure Overview: 10 mL sample + reagents in each
sample 10 mL DI water + reagents for blank
Chlorine
Sample needs to be collected in glassBest measured immediately in field
Chlorine
Equipment Needed 3-5 vials for samples + 1 for blank DPD chlorine reagent powder pillows Non-toxic waste container DR/850 Colorimeter
Procedure Overview 10 mL sample + reagent in each sample 10 mL sample and no reagent for blank Shake and wait 2 minutes
Waste Management
Appendix A, pages 1-3Collection Plastic Bottles work fine Clearly Labeled Non-Toxic, Ammonia and Nitrate all
should be stored separately
Organization “Callister Carrier” = cardboard 6-pack
container
Non Toxic Waste
DO, pH, Alkalinity, SRP, ChlorineDisposal Neutralize pH (baking soda) Pour down drain with excess water
Ammonia Waste
Several constituents of concern Maximum concentrations in manual
No way to safely reduce volumeMust be disposed of at a hazardous waste facility METRO or PSU Waste Consortium Talk to your local waste handler,
administrator or local college for other disposal sites
Nitrate Waste
Cadmium is only constituent of concernManual has instructions for precipitating heavy metalsMost teachers bring entire waste container to hazardous waste facility
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