ENVIRONMENTAL ENGINEERING LABORATORY Spring Semester … · ENVIRONMENTAL ENGINEERING LABORATORY...

ENVIRONMENTAL ENGINEERING LABORATORY

Spring Semester 2006

Instructors

Dr. Brian Wrenn Assistant Professor

Department of Civil Engineering [email protected]

935-8144

Dr. Lars Angenent Assistant Professor

Department of Chemical Engineering [email protected]

935-5663

January 2006

(Corrected) Schedule*

Date G1 G2 Reports Due 01-18-06

01-25-06 Course overview

02-01-06 Review of Statistics (Wrenn)

02-08-06 A-1 D

02-15-06 A-2 E

02-22-06 Experimental Design (Angenent) D (G2)

03-01-06 B-1 C-1 A (G1), E (G2)

03-08-06 B-2 C-2

03-15-06 Spring Break

03-22-06 C-1 B-1

03-29-06 C-2 B-2 B (G1), C (G2)

04-05-06 F A-1

04-12-06 D A-2 C (G1), B (G2)

04-19-06 E F F (G1)

04-26-06 Grad Projects D (G1), A (G2)

05-03-06 E (G1), F (G2)

05-10-06 Oral Presentation of Final Projects (Grad, B, F) Grad Projects

*Experiments designated in bold are supervised by Prof. Angenent; experiments designated in italics are supervised by Prof. Wrenn. Please turn in prelaboratory assignments and reports to the correct professor.

Lab Groups

Name Group Carolyn Maus Jessica Mohatt

G1

Adam Hargrove Abhas Singh

Kelsey Suddard G2

ENVIRONMENTAL ENGINEERING LABORATORY CHE/CE/ENVE 408/508

PREFACE

Welcome to the Environmental Engineering Laboratory. The objective of this course is to provide you with an introduction to several important tools that are commonly used in the analysis and characterization of environmental systems. These tools include analytical instruments, such as gas chromatographs and atomic absorption spectrophotometers, that are commonly used to collect data from natural and engineered systems. They also include simpler—but no less important—analytical methods that are based wet chemical analysis (e.g., titrations) or simple instruments (e.g., colorimeters), some of which are used in the field. Rigorous training in the use of these analytical instruments or methods is not our goal; instead, this course will emphasize a more general principle that is central to the quantitative chemical analysis of environmental systems: data interpretation. In particular, we hope to demonstrate how the methods used for data collection constrain the interpretation of results and to provide you with experience in the application of some statistical tools that can be used for data analysis. The experiments that you will conduct were not necessarily chosen due to their importance in environmental science and engineering, although they all play important roles in some aspect of these fields. Rather, they were chosen because they build upon the knowledge and experience that you should have gained in other courses (esp., ChE/EnvE 443 – Environmental Chemistry). We hope that these experiments provide a foundation for independent learning regarding both instrumentation and environmental analysis.

This course is continually evolving. It was originally designed by Prof. Jay Turner, and many of these experiments are derived from his original designs. Although we wish to acknowledge and are grateful for his contribution, he is not responsible for any deficiencies in the current course. The current suite of experiments attempts to use classical applications of analytical chemistry in environmental engineering (e.g., measurement of water/soil quality parameters) to illustrate fundamental principles involved in the collection and analysis of data for the characterization of environmental systems. We recognize that the course can and should be improved, and we would greatly appreciate your constructive criticism.

This manual is divided into three sections. Section I describes course logistics and basic laboratory safety practices. Section II summarizes the operating procedures for major instrumentation used in one-or-more experiments. Finally, Section III contains detailed descriptions of each experiment.

Lars Angenent and Brian A. Wrenn Spring Semester 2006 i


TABLE OF CONTENTS

PREFACE..................................................................................................................i

TABLE OF CONTENTS...........................................................................................ii

SECTION I: LOGISTICS AND LABORATORY SAFETY Course Logistics ............................................................................................1

Experiments .............................................................................................1 Recommended Reading ...........................................................................1 Attendance ...............................................................................................1 Prelaboratory Assignments ......................................................................1 Final Reports............................................................................................2 Grades ......................................................................................................3 Cooperative Learning ..............................................................................3 Late Policy ...............................................................................................3 Graduate Student Projects........................................................................4 Final Oral Presentations...........................................................................4 Lab Notebooks .........................................................................................4

Laboratory Protocols......................................................................................6 Student Liability for Breakage.......................................................................7 Laboratory Safety ..........................................................................................7

The Basics................................................................................................8 Safety Equipment in the Jens Laboratory ................................................8 Chemical Storage and Disposal ...............................................................8 Hazards Specific to this Laboratory.........................................................9

SECTION II: INSTRUMENTATION Operation of the pH Meter.............................................................................10 UV/VIS Spectrometer Operation...................................................................9

SECTION III: JENS LABORATORY LAYOUT AND EMERGENCY EXIT ROUTES Layout ............................................................................................................17 Emergency Exit Routes..................................................................................18

SECTION IV: LABORATORY EXPERIMENTS A. Atomic Absorption and Metals Precipitation ..........................................A-1 B. Adsorption................................................................................................B-1 C. Biochemical Oxygen Demand/Chemical Oxygen Demand.....................C-1 D. Henry's Law .............................................................................................D-1 E. Mass Transfer and Kinetics ......................................................................E-1 F. Total Petroleum Hydrocarbons (TPH) .....................................................F-1

Lars Angenent and Brian A. Wrenn Spring Semester 2006 ii


LOGISTICS AND LABORATORY SAFETY CLASS WEB SITE URL: classes.cec.wustl.edu/~ce408 PASSWORD FOR PROTECTED ACROBAT FILES: ClearSkies

Course Logistics Experiments:

This course consists of six experiments, A through F, which will be conducted over the semester. Two class periods will be devoted to a review of important statistical principles and methods that are important tools for analysis of experimental or observational data. Three experiments are two weeks in duration and three are designed to be completed in one week. The experiments follow a round-robin format. Lab groups consist of two to three students. Laboratory partners and your position in the round robin will be assigned by the instructor. The group assignments and schedule of experiments is given in the first page of this manual. Some rescheduling of experiments may be necessary due to the instructors’ travel.

Required Textbook: • this manual

Recommended Reading (on Reserve in Cupples II, Rm. 208): • Enke, C.G. 2001. The Art and Science of Chemical Analysis, John Wiley & Sons, New

York, NY. • Miller, J.N. and J.C. Miller. 2000. Statistics and Chemometrics for Analytical Chemistry,

Fourth Edition. Pearson Education Limited, Harlow, England. • Holman, J.P. 2001. Experimental Methods for Engineers, Seventh Edition, McGraw-Hill,

New York, NY.

Attendance: Attendance is mandatory in this course. Because of the round-robin nature of this course, it

is difficult to make up a missed laboratory session, but we will attempt to be flexible when an unavoidable conflict arises. These experiments are designed to be conducted by a group of two people. So, you have a responsibility to your lab partner to come prepared to every lab. If you absolutely can’t make a class you must notify the instructor and your partner(s) in advance and arrange to reschedule. Unexcused absences will result in a grade of zero for your final report.

Prelaboratory Assignments: A prelaboratory assignment (prelab) is due each week by 5:00 p.m. on the day before your

scheduled laboratory period (Tuesday). Prelab assignments are worth 15 points per lab session. The assignment questions are in the lab manual with each experiment. They should be printed neatly like any homework assignment. Prelabs are to be done individually.

In addition to the prelabs, your lab notebook must demonstrate that you are ready to conduct the experiment before you begin. This means that you should have recorded the Purpose and Procedure for the lab and have the Results section prepared to record data. Each group member must keep his/her own lab notebook, and they will be inspected before you are allowed to begin the lab. Although one group member may record data associated with specific activities, all group members should copy the data to their own notebooks before leaving lab.

Lars Angenent and Brian A. Wrenn Spring Semester 2006 1


Final Reports: A final report is required for each of the six experiments. Final reports are due two weeks

after the experiment is completed and are to be written and submitted as a group (i.e., one report per group). These reports are formal and should be TYPED. (Note: Where equations are required, use an appropriate equation editor or neatly hand write the equations in your report. Points will be deducted for inadequate presentation of equations.) Final reports are worth 50 points for one-week labs and 100 points for two-week labs. They must include the following (point totals given in parentheses; double for two-week experiments):

I. Title Page – Include the course number, the experiment letter and name, the date submitted, the names of all group members, and the instructor to whom you are submitting the report.

II. Executive Summary (5 points) – This should be 1/2 to 1 page long (maximum). An Executive Summary is used in all reports submitted to the U.S. Environmental Protection Agency. It is not an introduction; it is a concise overview of the experiment, including significant results and conclusions. Avoid excessive verbosity in this section.

III. Deviations From the Method - Reference your lab manual and describe any procedural modifications that you made during the experiment. If there were no deviations, reference the lab manual. (Note: No specific point allocation is assigned to this section, but failure to provide this information will result in a reduction of your total grade that is proportional to the severity of the omission.)

IV. Results (20 pts) - Include a brief introduction that describes the types of measurements that were made and how they contributed to achieving the objectives of the experiment. Use tables and graphs wherever possible. All data that is presented in this section should be referenced in the accompanying text. At a minimum, describe what each figure and graph shows. Include any statistical calculations, standard curves, linear regressions, etc. where appropriate. You will be graded on the quality of your results and data analysis (i.e., correct use of appropriate methods) and the clarity of your presentation.

V. Discussion (15 points) - Briefly reiterate the context for this experiment (i.e., how it is important in environmental engineering or science) and answer any questions that are posed in the course manual. This section should allow you to demonstrate your understanding of the purpose, results, and applications of the experiment. Place your results in context with the collective understanding of the field. An excellent final report should include a thorough demonstration of what you learned during the laboratory and make connections to principles that you learned in other courses. Average reports meet the minimum requirements with good quality data and appropriate analysis. In general, you should: • Discuss potential sources of error in your results or important interferences in the

method(s) that you used. • Demonstrate your understanding of the purpose and experimental methods you used. For

example, describe the function of Quality Assurance procedures such as method blanks, standards, and surrogate standards (e.g., recovery of standard spikes).

• Discuss the engineering applications of your results or observations. • Use references when needed (especially other textbooks). Do not reference the lab

manual.



VI. Appendix (10 pts) - Show all of your calculations in a clear and concise manner. You may handwrite this section, and raw data may be included by photocopying from your lab notebook, but be sure that your handwriting is legible. The keys to this section are clarity, completeness, and organization. If you used a computer spreadsheet, be sure to show one complete example calculation for each formula.

Grades: Your course grade will be based on your laboratory final reports (60%), the prelab

assignments (15%), the final oral presentation (10%), and your lab notebook (15%), which must be turned in during the final oral presentations (May 12, 2004). Graduate student projects (see below) will be treated as an additional experiment and graded as follows: draft project proposal (10 pts), experimental design (20 pts), report (50 pts).

Cooperative learning: The laboratory final report will be prepared as a group. We will use cooperative learning to

structure group functioning to prevent an unequal work load by group members. Research has shown that learning benefits the most from cooperative learning and that a gain in knowledge retention is anticipated. Conceptually, cooperative learning is ideal for engineering education, because team effort and group-based problem solving are required in the practicing world. One of the main instruments of cooperative learning is positive interdependence, which ensures that group members are dependent on each other for the successful completion of the project (in this case the lab). In addition, self assessment of the group function is performed by filling out forms to improve the interaction over the period of the course. Students can “fire and hire” after several attempts to motivate freeloading group members have failed and if such freeloading behavior had been documented in the self assessment forms. Finally, each of the lab reports that are handed in as a group, will be graded by the instructors and each of the group members grades themselves and the other group member(s). The latter will result in each group member obtaining a different grade if an unequal work load between group members is found. Note: If a group member has not contributed to the lab report, her/his name should not appear on the lab report. For this lab, 60% of the grade is based on your group work, and you and your team members are grading yourselves as part of this grade. A handout containing the assessment forms will be given to you during the first class and a pdf file will be available on the web-site. Every student should fill out an individual effort rating form, and each group should jointly fill out a self-assessment form. The self-assessment form must be included with the lab report and the individual effort rating should be given to the instructors directly.

Late Policy: With prelabs due every week and final reports due every two weeks it is very important to

turn assignments in on time so that you do not fall behind in this class. Note that few lab reports are submitted during the first half of the course, but reports are due almost every week in the second half. So, appropriate time management is critical to success in this course. Any assignment turned in late will receive a grade reduction as follows:

1 day late -- deduct 10% 2 days late -- deduct 50% 3 or more days -- deduct 100%



Graduate-Student Projects: Students enrolled in ChE/EnvE 508 must do one extra project to receive graduate credit for

this class. The grade, which will be based on a written report and an oral presentation, will be averaged in as if you did one extra laboratory experiment. The instructors are available meet with all graduate students to discuss potential projects. Projects will be initiated by the students, but the topics must be approved by the instructors. A draft proposal for graduate projects is due on March 1, 2006 and will be returned during the first lab session after spring break. This draft proposal should include a brief description of the objectives of your proposed project, the methods that you plan to use, and an estimate of the number of experimental units that expect to use. A detailed experimental plan must be submitted by April 5, 2006. If special equipment or reagents are required, you should discuss these issues with the instructors to make sure everything you need is available. Graduate students and undergraduates taking the course for graduate credit can make arrangements to use the Jens Lab for their special projects at any time after their experimental plan is approved, but the instructors will be available to provide advice from 4-8 p.m. on April 26, 2006.

Final Oral Presentation: There is no final exam in this course, but all students will give a final presentation on one

experiment during the regularly scheduled exam period (4-7 p.m. on May 10, 2006). Undergraduate students will make presentations based on their last experiment, and graduate students will present the results of their special projects. Undergraduate lab partners will give a single group presentation, but graduate presentations will be made by each individual. Presentations should be about 15 minutes with a 5-10 minute discussion period to follow.

All presentations should include a brief introduction describing the importance of the experiment to the field of environmental engineering and a clear statement of its objectives. (You may want to speak to environmental engineering professionals to provide context or practical applications for the techniques you used or the process that you investigated.) Undergraduates can concentrate on presentation and interpretation of results because all students will have conducted the experiments that you will be describing. Graduate students must briefly describe the methods that they used. Results and conclusions should be presented clearly and in a logical manner. Presentation of results in a chronological order (i.e., first I did A, then I measured B, etc.) is seldom the best way to organize a talk or report. As part of your conclusions, you might suggest ways to improve the experiment in the future (e.g., better analytical methods may be available, the reaction conditions or time may be suboptimal). All students are encouraged to contribute to the discussion following each presentation. This may be based on your experiences with the experiment or similar processes, or you might be able to provide some insight from other courses or experiences that you have had.

The presentations will be given in the Environmental Engineering Program conference room (Cupples II, Room 212). A computer projector and a white board are available in the room. Please use visual aids. (PowerPoint presentations are encouraged.)

Lab Notebooks: Record keeping in the laboratory is extremely important. You must keep a bound lab

notebook for recording results, observations, and deviations from the method. Your notebooks will be collected during the final presentations and graded (15% of your total grade).



It is of utmost importance to write down your objectives, activities, observations, data, and results in a lab notebook. Details are very helpful when you need to perform a similar experiment years from now, if somebody else in the laboratory has to make similar measurements for another purpose, or if you wish to reinterpret your results at some later date. Do not worry that you are recording too much information; you cannot rely on your memory. You should also record any mistakes you made and problems you encountered. When your advisor or a colleague explains and shows you a new procedure, write every step down carefully. Make sure the person that explains things remain at a slow speed of showing you, so that you can write everything down. You will have to be instructed again if you forget to do this. Somebody else needs to be able to read your notebook and should be able to perform the same test or protocol you just described, while also having access to a standard operating procedure (SOP). The SOP is given to you as part of the lab manual. You should not copy the SOP in your lab notebook.

Some standard procedures that you will have to follow: 1. The notebook must be permanently bound and the pages numbered (a lab notebook will be

given to you, but you will have to number the pages yourself). 2. Leave the first 2-4 pages blank initially, and use them for a Table of Contents that you fill as

you go along. Tip: It is OK to have white spaces in the lab notebook, you simply draw a line to connect the parts of the lab session.

3. Date every page and sign after you are done for the day (end of lab session). 4. Start each new lab session on a new page. 5. Handwriting must be legible. All the writing should be done in ink and are part of the

permanent record: neatly cross out mistakes with a single line, so that it remains readable, don’t tear out pages.

6. You may glue in drawings, graphs, Excel data sheets, handouts, etc. Sign or leave circles around the corners of the paper that you glue in.

7. The lab notebook should be current; so, write while performing work, or finish the notebook entries immediately after finishing your work. We will check your lab notebook for accuracy and to make sure it is current.

8. It is reasonable to make corrections. Delete enries by drawing a single straight line through text. The deleted text should remain readable.

9. Everything should be recorded including: a) Title of experiment and date b) Note the members in the lab who assisted you. c) Purpose: your objectives in one sentence. d) Introduction: what you are trying to do and why you are trying to do it. e) Calculations: some are needed before starting the lab session. f) Procedures: this should not be an exact copy of the SOP, but reworked so it is easy for

you to follow and full of tips on how to do it. It also should include where materials are located and which equipment was used (and its location).

g) Information: clearly identify the chemicals you used, the calculations and procedures for mixing the chemicals if this was performed (e.g., recipes, pH adjustments, tips on how to mix the chemicals [i.e., mixing, heating, what water to use, etc]). Write down the actual



time when you start and end an experiment: for example, use “3:15 pm to 5:15 pm” rather than “2 hours”.

h) Equipment manufacturer, model, specific details (e.g., reference number, if available) of all the equipment that you used.

i) Any deviations from the SOP and why such deviations were made. j) Observations: everything that happens during the experiment that may be important for

the results or the interpretation of the results. k) Results: raw data and calculated values, sample calculations are needed because you may

forget how they were done (for example a conversion of units). l) Discussion and conclusions: interpret your results and draw conclusions on your

experiment. 10. If you need to add information later, please date, sign, and mark what was added. If it the

new information is recorded in a different part of the notebook, indicate the pages on which the related information can be found (add a similar page reference to the new information on an appropriate page in the section containing the original data).

11. You may refer to a page number when you do a test over again in a similar manner. 12. Pictures from gels must be taped in the notebook as well, but make sure you label them with

meaningful names so that you unambiguously identify it later. 13. Never record data on loose pieces of paper. 14. Date all the information taped in your notebook. Don’t leave data just stored in the

computer because this is not a permanent record!!

Lab notebooks are works in progress. So, sometimes they are not particularly tidy, but they needs to be readable by others. Be guided by the principle that you will forget everything that you do; do you remember what you did exactly 2 years ago?

Lab notebooks will be graded at the end of the semester, but the instructors will monitor them throughout the semester to determine whether you came prepared for each lab session. Before every lab session, the Purpose, Introduction, some necessary calculations, some of the Procedures, and some Tables (for recording data) must be prepared in the lab notebook. The degree to which your notebooks are properly prepared for each lab session will influence the grade you receive. During the first lecture, examples of notebooks will be shown. These examples will be shown on the course webpage as pdf files.

Laboratory Protocols Good laboratory technique must be used to obtain reproducible, accurate data. Prior to a

given analysis, you should: • determine the type of information that is desired (i.e., qualitative vs. quantitative); • if quantitative analysis, estimate the required analytical precision; • obtain an estimate (e.g., an educated guess) of the concentrations of species of interest in the

sample • determine which instruments are suitable for the desired analysis • identify potential interferences due to the properties of the sample to be analyzed • identify positive and negative controls if required

Different protocols for routine aspects such as glassware cleaning and water purification are justified for different types of analyses. For example, you do not want to spend excessive time



(and money) to achieve extremely high reagent purity when performing an analysis that does not warrant such resolution. In contrast, analysis of species at the ppm (parts per million) or ppb (parts per billion) level often requires very meticulous attention to every detail of the experimental protocol to avoid sample contamination. A crucial step of any analysis is the careful planning and analysis of sample blanks and matrix spikes to ensure that the integrity of the analysis is not sacrificed by inadequate sample preparation and handling, and matrix effects due to sample composition.

Label, label, label! Make sure you label all solutions and samples. Labeling tape and a pen (e.g., "Sharpie") is recommended. Print your name and lab session on anything that is to be saved for the next lab session. Any improperly labeled or unlabeled reagents or samples will be discarded. All labels must include the contents, concentration, date, and your name. When labeling contents, avoid use of abbreviations and chemical symbols. If necessary, you may write the full contents of complex solutions on a separate sheet of paper, but this must remain with the samples at all times! This is an EPA requirement, and we can be fined for improperly labeled containers.

Student Liability for Breakage Students will be held responsible for damage or breakage resulting from negligence or

careless handling. Failure to observe specific precautions noted in the text, posted near the equipment, or explained by the instructor will constitute negligence. Normal wear and tear is, of course, excluded. The following procedure will be observed: • Before beginning an experiment, you should check the equipment for defects. Report any

obvious damage to one of the instructors before you start work. • The instructors will inspect the equipment before and after class and will note any new

damage. If new damage is noticed, the instructors will assume that the most recent users are responsible.

This procedure is intended to provide a record of defects to facilitate prompt repair and improve equipment reliability. It will also give each student an incentive to exercise reasonable care in the laboratory.

Laboratory Safety The safety and protocol procedures described in this section are specific for this course as

offered in the Jens Laboratory and should not be considered as a comprehensive laboratory safety manual. Always observe appropriate safety measures when working in or visiting any laboratory. A more thorough discussion of laboratory safety may be found in Standard Methods For The Examination of Water and Wastewater, 20th Edition. Wash U’s lab safety training presentation is accessible through the EH&S website at: http://www.ehs.wustl.edu/training/trainingpreshill.htm

Safety is a priority in this course. The need for rigid safety protocols may not be obvious during every activity in this laboratory, but safe work habits are essential to preventing accidents. All students and instructors will be required to take the appropriate precautions at all times. Never assume that bench-scale apparatus, or small amounts of chemicals, do not represent a hazard. Even a few drops of acid can cause serious injury.



In case of fire, first dial the emergency number (5-5555) before trying to fight the fire. Second, evacuate the area. Only then should you try to fight the fire, and then only if you can do so safely. Remember, even the most disastrous fire starts small. Do not hesitate to call the fire department. Appendix A contains a map showing the escape routes from the Jens Laboratory.

If you must evacuate the lab, proceed out either exit to the northwest corner of the floor and up the steps to the building exit (see map on p. 18).

•

If someone is hurt in the laboratory, tell one of the instructors immediately. Call the emergency number (5-5555) if assistance is required. Report even minor incidences to the instructors so that corrective actions may be taken to improve safety in the future.

Safety Basics: Dress appropriately for this class. Do not wear your best or expensive clothes. Many of the

chemicals used here can stain or eat holes in clothing. Don’t wear extremely loose-fitting clothing that can fall into things or knock over containers. Always wear closed-toed shoes (no sandals of any type!), long pants, a lab coat, and safety glasses or goggles.

Anyone caught eating, drinking, or smoking in the laboratory will be summarily executed (i.e., you will recieve a grade of zero for that experiment). Always wash your hands well before and especially after you have worked in the laboratory. Never pipet by mouth! Pay close attention to safety procedures outlined in each specific experiment.

Emergency telephone numbers are posted by the phone in the middle of the lab. The phone is for emergency use only. Personal phone calls (in-coming or out-going) are not allowed.

Safety Equipment in the Jens Laboratory: Safety goggles and glasses, disposable nitrile gloves, and lab coats are available in the Jens

Lab and must be used. Of course, you may bring your own glasses/goggles/coat if you prefer.

Type ABC fire extinguishers are located at both exits of the laboratory (see map p. 17). A fire blanket is attached to the coat cabinet. An eyewash and a safety shower are located in the center of the laboratory (near the atomic absorption spectrophotometer; p. 17). The hoses attached to the faucets also make excellent eye washes. A chemical spill kit is available. Notify an instructor immediately of any spill in the laboratory.

The fume hoods are to remain in operation constantly. Do not turn them off. Certain procedures must be performed in the fume hoods; be sure to do so. Be courteous with the limited space in the hood and always clean up after yourself as you go. (Remember: you know the identity of the white powder that you left on the balance, but the next user does not. Potassium cyanide and glucose are both white powders.)

Chemical Storage and Disposal: Reagents needed for each experiment will be stored near the area in which the experiment is

to be performed. Your instructor will inform you of any exceptions. Always return the chemicals to their proper storage place. If you are in doubt, ask! Storing incompatible chemicals together is dangerous.



Chemical disposal procedures will be outlined in each experiment, where necessary. Many of the wastes will be collected in containers. All will be well labeled and convenient. Please follow the disposal procedures carefully.

Hazards Specific to this Laboratory: Many of the procedures in this laboratory require care and thought, but a few specific

hazards need to be mentioned here. 1) Use extreme caution with the dichromate and acid solution when performing the Chemical

Oxygen Demand test. All acids require careful use, but this is a strong oxidizer and contains toxic compounds, Cr(VI) and Hg(II), and it will be heated to a very high temperature (150oC). Use of the closed reflux method minimizes much of the danger associated with splattering, but poses the additional hazard of generating high pressures in sealed vessels.

2) The gas cylinders in the laboratory contain high pressure and some contain explosive gases (esp, H2). They are the responsibility of the instructors. Do not attempt to adjust them in any way.

3) Some of the electronic equipment is high voltage. Do not attempt any procedure beyond that which is called for in the method. Do not place anything on top of the equipment. Bring only your notebooks and prepared samples to the equipment benches.



INSTRUMENTATION

Operation of the pH Meter The measurement system for pH consists of the meter and two probes - a pH electrode and

an automatic temperature corrector (ATC) probe. (Note: As of this writing, the ATC probe does not work, but it must be connected anyway.) The pH electrode bulb senses the concentration of hydrogen ions (H+, protons) in solution by differential adsorption to the two sides of the glass membrane, thus establishing an electrical potential difference (i.e., a voltage) across the membrane. This glass bulb is very fragile. Although the probe is surrounded by a plastic guard, it does not provide complete protection. Therefore, handle the pH electrode carefully. Never touch the glass bulb. This can impart a static electrical charge to the outer surface of the glass that will disrupt its ability to measure the adsorption of protons. Remove excess water by gently shaking the probe or inserting a Kimwipe between the bulb and plastic guard to wick it away.

Dual-point Calibration: 1. Plug in the meter. If it is already plugged in, and one of the error codes is displayed, press

the "Reset" key. 2. pH buffers are stored in plastic bottles near each meter. Select the pH 7 (yellow) buffer and

either the pH 4 (red) or pH 10 (blue) buffer depending on the pH range you expect to measure. Fill a small 20-mL screw-cap vial about half full with an appropriate buffer. If you are not the first user, vials containing buffer may already be present. In general, the buffer can be reused, but make certain that it is not too old. Ask your lab mates or the instructor if you do not know its history. (Note: pH 10 buffer is particularly susceptible to degradation due to its tendency to absorb CO2 from air. This buffer should be replaced if it is more than several hours old.)

3. If all solutions are equilibrated to room temperature, you can leave the ATC probe in the storage solution. If the temperature of the solution whose pH you wish to measure is significantly different from room temperature, you must use the ATC probe to obtain a correct measurement. (As noted above, the ATC probe is currently nonfunctional.)

4. Rinse the pH electrode and ATC probe using distilled water. Always rinse into a waste beaker; do not rinse into the buffer solutions! Gently shake the probes to remove residual water drops. Gently wick away excess water using a Kimwipe if necessary, but do not touch the glass bulb.

5. Immerse the pH electrode into the pH 7 buffer. If you are using the ATC probe, the meter will display the solution temperature. Stir the solution with a magnetic stirring bar; gently tap the probe body to dislodge any air bubbles clinging to the probe tip. Press the "pH 7" key a few times until a stable reading is obtained (it may take a few minutes for the probes to reach equilibrium with the surrounding solution). If the meter displays "E1--", ensure you are using the pH 7 buffer and press the "Reset" key to start again.

6. Remove the pH electrode and ATC probe from the buffer and rinse with distilled water. Gently shake the probes to remove residual water drops.

7. Immerse both probes in the second buffer (pH 4 or pH 10, depending on the pH of your sample). Stir the solution with a magnetic stirring bar; gently tap the probe body to dislodge



any air bubbles clinging to the probe tip. Wait for the pH to stabilize then press the appropriate "pH 4" or "pH 10" key; the meter now displays the pH for the buffer.

8) Press the "pH" key; the meter calibrates the response and is ready for pH measurement of other solutions. If the meter displays "E2--", ensure the proper buffer is being used and the proper pH key is being depressed. In the event of an error message, repeat the calibration starting from step #3.

9) Measure the pH of the pH 7 standard to check the calibration. If the pH is not 7.00 + 0.02, repeat the calibration procedure (be certain that the probe reading has stabilized before you recalibrate and during each step of the recalibration procedure).

pH Measurement: 1) Remove both probes from the previous solution and rinse with distilled water. Gently shake

the probe to remove residual water drops or wick away excess water with a Kimwipe. 2) Insert both probes into the solution of interest; the meter displays the pH if no keys were

depressed between the calibration and current measurement. If other keys were depressed (e.g., to measure temperature), return to the pH display by pressing the "pH" key. Stir the solution with a magnetic stirring bar, and gently tap the probe body to dislodge any air bubbles clinging to the probe tip. The reading may take a few moments to stabilize; record the pH when the reading is relatively stable.

3) Repeat this procedure for additional measurements. 4) Between measurements of different samples, keep the probe tips immersed in KCl storage

solution. To avoid cross-contamination of your samples, always rinse the probe tips and probe side walls.

NOTE: If the "Reset" key is pressed at any time after the meter calibration, it will be necessary to repeat the calibration!

Storage: 1) Rinse the probes with distilled water and immerse the tips in a beaker containing KCl

solution. Leave immersed in the KCl solution until the meter is used again. 2) Discard the used buffer solutions.

Specifications: Equipment: pH meter, Model 6209, Jenco Electronics, Ltd., Taipei, Taiwan. pH electrode, #C2506A-12A-A023BC, Broadley-James Corp., Santa Ana, CA. ATC probe, #600A, A. Daigger & Co., Inc., Wheeling, IL.

Variable Range Resolution Accuracy pH 0 to 14 0.01 ± 0.1% temperature -10 to +250 _C 0.1_C ± 0.5 _C

For more details, refer to: Operation Manual for Models 6201, 6209 pH Meters, Jenco Electronics, Ltd. •

• Electrode Instructions, Broadley-James Corporation.



UV/VIS Spectrometer Operation The Perkin-Elmer Lambda 2 UV/VIS spectrometer is a double beam unit which can be

operated in several modes. Conventional fixed-wavelength analysis can be performed, and the instrument can be programmed to scan portions of the UV/visible wavelength range. We also have PC-based software (PECSS) which can drive the unit, create data files that can be printed or opened in other applications (e.g., MS Excel), automatically perform calibration calculations, etc. You will have an opportunity to use several of these functions in this course.

Recall that while the absorbance measurements are absolute, they are related to concentration solely by comparison to data collected for known standards (i.e., the extinction coefficient of the species of interest under the conditions of your measurement must be known). Therefore, standards must be carefully prepared, and they must bracket the expected absorbances.

•

• All measurements must be made within the linear range and within an appropriate range of the instrument. For this spectrophotometer, the sensitivity of the photomultiplier restricts measurements to absorbances < 2. All samples and standards should be prepared at the lab bench. Bring only cuvettes or test tubes containing samples (in a rack) and your notebook to the analytical bench. Please do not place any objects on top of the UV/VIS unit.

•

Manual Operation at Fixed Wavelength: 1) Turn on the main power using the green switch on top of the instrument (upper-right-hand

corner). The unit responds with LAMBDA 2 VERS. 5.4K BUSY

Wait until the response changes to xxx.x nm x.xxx ABS INPUT: > <

where the x's are numbers. 2) Press GOTO. Use the numeric pad to enter the desired wavelength and press ENTER. Wait

until the response returns to yyy.y nm x.xxx ABS INPUT: > <

where yyy.y is the wavelength you selected. If you enter an incorrect wavelength, start again by pressing GOTO.

3) Open the hinged door. Place an analytical blank cuvette in the reference cell holder (rear position) and an analytical blank cuvette in the sample cell holder (front position). The blanks should contain the same solvent as your samples. For disposable cuvettes, always insert the cuvettes with the arrow in the direction of the beam path (that is, the arrow should face to the left or to the right, not towards you or away from you). The same requirement applies for quartz cuvettes except that a blue mark replaces the arrow. Also, make sure the sides through which the light will pass are clean and free of any smudges or water droplets that will scatter light. Close the hinged door. Press BACK CORR to zero the instrument response. The display should change to

yyy.y nm 0.000 ABS INPUT: > <



4) Replace the cuvette in the sample cell holder with a cuvette containing a sample. Close the door and allow the absorbance response to stabilize. Record the absorbance and repeat for additional samples. Note that you do not need to press any keys to obtain readings; the response is continuous.

5) Should you inadvertently press the START key or another key, press STOP twice to return to the desired screen display.

6) Turn off the main power at the end of the evening.

Manual Operation at Multiple Wavelengths (WAVPR/MAN): 1) Absorbance measurements can be made at up to 20 different wavelengths on each sample

using the manual wavelength program, WAVPR/MAN. The instrument automatically cycles through all designated wavelengths beginning at the longest wavelength and progressing toward the shortest. The instrument can store a separate background correction factor at each wavelength.

2) Program the wavelengths that will be measured as follows: • Press the METHOD key. The display will read “Select Method > <” Key in “3” to

select the WAVPR/MAN method and press the ENTER key. • The display will read “<-->/PARAM/START”. Press the PARAMETER key. • The display will read “Modify Method” and “PARAM/->. Press the PARAMETER key. • You will then cycle through a variety of parameters, which you can change either using

the numeric keypad (for numeric information) or the cursor keys (to scroll through a list of options). When you have entered the value you want for a given parameter, move to the next parameter by pressing the PARAMETER key. When you are finished changing parameters, press the STOP key. The parameters that you can set and the values that you should select are given in Table 1.

Table 1: Parameters for WAVEPR/MAN Parameter Value Action Key

ORDINATE MODE ABS (absorbance) right cursor # WAVELENGTHS # (must be <20) ENTER

WAVEL. 1 λ1 ENTER, right cursor WAVEL. 2* λ2 ENTER, right cursor WAVEL. n* λn ENTER FACTOR 1 ENTER

RESPONSE (seconds) 2 ENTER LAMP UV, Vis, or UV+Vis right cursor

BACK CORR. YES right cursor SAMPLES/BATCH # ENTER FIRST SAMPLE # 1 ENTER

CYCLES 1 ENTER *λ1 > λ2 > λn

3) At the display “METH 3 WAVEPR/MAN”, press the START key to begin the analysis. The instrument will then display “BACK CORR.” and “PRESS START”. Insert a cuvette



containing the solvent in which your samples are dissolved (usually DI water) in both positions, close the sample compartment cover, then press the START key. The grating will turn to the first wavelength, perform a brief background correction, then move to the second wavelength and repeat the process. You will hear the grating motor whenever the grating is turning. After measuring for “RESPONSE” seconds (see Table 1) at each new wavelength, the instrument will display the wavelength and the background corrected absorbance of the solution in the cuvette in the sample position. At this point, the absorbance should read 0.000 + 0.001 at all wavelengths. Note: Use quartz cuvettes if any measurements will be made at wavelengths <400 nm.

4) The display will read “BATCH NUMBER 1” and “PRESS START”.

5) After the background correction is complete, rinse the cuvette with your first sample or standard, fill the cuvette with the same solution, and place it in the sample position. Close the sample compartment cover, and press the START key. The display will read “SAMPLE 1” and “PRESS START”. Press start to read the absorbance of the first standard at all selected wavelengths. Record the absorbance when they are displayed.

6) The display will read “SAMPLE 2” and “PRESS START”. Repeat step 5 until all samples and standards have been measured.

7) Turn off the main power when you are finished using the instrument for the evening.

Operation with PECSS Software Startup:

Start the program by typing “pecss” at the DOS prompt from the root directory. This will initialize the software and display a menu of options. Note that the bottom of the screen displays “Ready for next command”.

Acquiring UV-Vis Absorbance Spectra (SCAN Mode): The SCAN mode is used to obtain an absorbance spectrum. From the “Ready for next

command” statement, type “scan”.

1. A screen will appear which prompts you to set several parameters. Enter the values provided in Table 2, and then hit the “enter (return)” key to advance to the screen for the scan summary. • Be sure to provide a unique file name (8 characters maximum) for all spectra. The

default is User001, which will automatically increment with each new spectrum that is acquired, but you should provide a file name that is meaningful for later reference. You will have to copy your files to a floppy disk and import them into Excel or some other program to plot the spectra.

2. After entering these parameters, you will be prompted at the bottom of the screen with the following options: Y (scan), N(do not scan), A (autozero / background correction), and C (change parameters). After each scan, this list of options will reappear at the bottom of the screen. • Be sure to autozero/background correct before performing any scans or if the instrument

has been used in another mode since performing the last background correction in scan mode.



Table 2: SCAN program parameters for obtaining absorbance spectra

Parameter Value Comments

Scan Options: reg X (X, Y, Z) register for storing A vs. λ data start λmax maximum wavelength (nm) end λmin minimum wavelength (nm) int 0.5 wavelength interval for readings (∆λ)

ncycl 1 # cycles per spectrum

Instrument Options: ord A ordinate variable (e.g., absorbance)

speed 480 scanning speed (nm/min) smooth 2 bandwidth (nm) lamps 3 UV and visible lamps both “on”

rec 0 instrument recorder “off”

Output Options:

sample ID filidxxx 8-character filename; defined by user y(min) 0 lower bound on ordinate y(max) upper bound on ordinate (e.g., 1.0, 2.0) asave Y(es)/N(o) autosave the spectrum (N) aprint Y(es)/N(o) autoprint the spectrum (N)

3. When finished with the SCAN mode, type “N” at the “Ready for next command” prompt. Press “Esc” key to return to main menu.

4. Use the “VIEW” command to display the spectrum. The syntax is as follows:

VIEW reg (X, Y, Z) min (Amin) max (Amax) start (λmax) end (λmin)

You can overlay two or more spectra by invoking “VIEW” more than once, specifying each register (X, Y, or Z) separately. Be sure the values for Amin, Amax, λmax, and λmin are the same in all command lines.

5. Make ASCII files containing your spectral data using the “JCAMP” command. The command syntax is:

JCAMP reg (X, Y, Z) format (S, U) filid (8-character file name)

Select “S” (scaled) as the file format. This will produce a space-delimited ASCII file, filid.dx, which will be stored in the directory \PECSS\DATA on the hard drive. You can copy these files to disk and open them in Excel to plot the spectra. • Type “dos” in the command line to work from the DOS prompt. Copy your file to a

floppy disk by typing: “copy \data\*.dx a:\*.dx” where * is your 8-character (max) file name.

• When you are finished copying the files containing your spectra to your floppy disk, type “exit” to return to the main menu.



Measurements at Multiple Wavelengths (WAVPRG Mode): The WAVPRG mode is used to record and display the absorbance value at specified

wavelength(s). From the “Ready for next command” prompt, type “WAVPRG”.

1. A screen will appear which prompts you to set several parameters. Enter the values provided in Table 3, and then hit the “enter (return)” key to advance to the screen for the wave program summary.

2. After entering these parameters, you will be prompted at the bottom of the screen with the following options: Y (read absorbance), N (do not read absorbance), A (autozero / background correction), and C (change parameters). After each reading, this list of options will appear at the bottom of the screen.

3. When finished with the WAVPRG mode, type “N” at the “Ready for next command” prompt.

Table 3: WAVPROG parameters for measuring absorbance at two (or more) wavelengths

Parameter Value Comments

Scan Options:

method leave blank ncycl 1 # cycles per sample nwave 2 # of wavelengths monitored

Wavelength Options: absc λ1, λ2 wavelengths to be monitored

Instrument Options:

ord A ordinate variable (absorbance) resp 2 response time (s)

lamps 3 UV and visible lamps both “on”

Output Options:

sample ID defined by operator asave Y autosave the data aprint N autoprint the data

ShutDown: To exit the PECSS program and return to DOS type “stop” at the “Ready for next command”

prompt. Turn off main power when finished using the instrument. JENS LABORATORY LAYOUT AND EMERGENCY EXIT ROUTES

Laboratory Layout



north hallway

acid desk sink service elevator

hoodfire extinguisher

hood

hood

AAS

GC-FIDGC-PID

FTIR

desk sink

sink

sink

eye wash safety shower

lab coats

gas cylinder

trench drain UV-Vis shaker desk

desk

Aquatic Chemistry Laboratory

east hallway

(Giammar) BOD incubator

refrigerator

fire extinguisher

stairs

south hallway




Emergency Exit Routes

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