Laboratory Manual: CHEM 311 Environmental Chemical Analysis
Laboratory Manual: CHEM 311 Environmental Chemical Analysis
Laboratory Manual: CHEM 311 Environmental Chemical Analysis
ENVIRONMENTAL CHEMICAL
ANALYSIS
LABORATORY MANUAL
Sept. 7th Introduction to a Chemical Analysis Laboratory: Good Laboratory Practices, Data
Analysis, Technical Reports and Full Lab Reports.
Sept. 14th Introduction to Metering Devices (pH, Turbidity, Conductivity and DO)
Calibration, Precision and Data Reporting
Data Tables Due: Sept. 20th
Full Lab and Technical Reports are due 11 calendar days following the completion of the lab. A late
penalty of 10% per week applies for reports up to two weeks, after which they will NOT be accepted.
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TABLE OF CONTENTS
INTRODUCTION Page
List of Experiments iv
Lab Reports v
Typical Marking Scheme vi
Cheating and Plagiarism vii
General Laboratory Procedures viii
How to Read an Analytical Method ix
Laboratory Safety xi
General Laboratory References xii
Introduction to Good Laboratory Practices xiii
LAB 0 – Good Laboratory Practices Exercise xxvii
EXPERIMENTS
APPENDICES
Glossary of Terms 65
iii
LIST OF EXPERIMENTS
9. Metals in Sediment
(Technical Report/20)
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FULL LAB REPORTS
CHEM 311 Lab Reports are submitted as stand-alone formal reports (unless otherwise noted) that are to
be written in an impersonal voice in typed format. Your lab report should outline the principles of the
chemistry and/or instrumentation employed, calibration techniques, data handling, an estimate of
experimental uncertainty and a general awareness of the context and significance of the results.
TITLE PAGE AND IDENTIFICATION: Course number. Name of student. Name of partner. Date.
Unknown #. The title should provide the reader with both the analyte and the matrix studied and give
some indication of the technique employed. E.g., The Analysis of Fluoride Ion in Toothpaste Using an
Ion Selective Electrode.
PRINCIPLE OF METHOD: Describe the principles involved in relating the measured quantity (e.g.,
volume of titrant, absorbance, potential etc.) to the analyte concentration. For wet chemical techniques,
include the stoichiometry of chemical reactions that will be used in the calculation of results. For
instrumental techniques, describe the principle of operation of the instrument itself. Schematic diagrams
may be useful for instrumental methods. Do not describe details of the procedure here.
DATA: Tabulate data with descriptive headings and footnotes providing details. Data tables should be
able to stand alone providing enough information that the reader could carry out necessary calculations
without having to go hunting for additional information. For example, in a data table summarizing
titration volumes, be sure to include the titrant concentration and the sample volume.
DISCUSSION: State your result/s and give some context for the magnitude (high, medium or low). For
example, report the levels of Fluoride ion in commercial toothpaste, other foodstuffs or drinking water.
Be sure to convert to common concentration units, if necessary. Comment on the precision (RSD) and/or
accuracy (% bias) of the method using your data and the reported values given in Standard Methods.
Discuss possible interferents and other sources of error. Conclusion paragraph should clearly report final
results for all samples with 95% CL and n (# of replicates).
REFERENCES: All references cited in the report should be listed as numbered endnotes in the style
adopted by Analytical Chemistry.
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TYPICAL MARKING SCHEME (Full Report)
The following represents a typical marking scheme. Actual marking schemes for a particular lab
may vary.
Mark Max.
Mark
Technique/Preparation – preparedness, timeliness and ability to work
1
carefully in a clean an organized manner.
Principle of Method – explain the type of analysis, include relevant
chemical equations and/or theory of instrumental operation, including
calibration technique. Addresses the theory that relates the measured 3
signal to a meaningful quantitative result. Does not include procedural
details.
Data – complete, clearly presented tables including all pertinent
3
information and uncertainty in measurements.
Calculations – correct, organized, clearly presented including error
analysis to give uncertainty in the final result. Include calibration 3
curves, if any.
Results – level of agreement between your result/s and the known or
3
true value for an unknown or environmental sample.
Discussion – clearly state your result/s and give some context for the
magnitude (high, medium or low). Comment on the precision (RSD)
and/or accuracy (% bias) of the method using your data and the
reported values given in Standard Methods. Discuss possible 4
interferents and other sources of error. Conclusion paragraph should
clearly report final results for all samples with 95% CL and n (# of
replicates).
Literature Comparison – include brief overview of essential aspects
of an alternate method for the same analyte or alternate analyte using 2
the same method. Use Standard Methods, text or library references.
Layout/Organization – includes pertinent information on title page,
proper section headings, labelled figures and/or graphs, all sources of 1
information (references) properly cited as end-notes.
TOTAL 20
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GENERAL LABORATORY PROCEDURES
General Procedures
1. Labs are conducted in pairs. You will need to be organized and divide tasks to complete
the labs in the allocated time.
2. Glassware will be provided on an as needed basis during the lab period. Students should
come to the lab prepared with a list of required glassware and an organized work plan.
4. At the end of the laboratory period: All glassware should be thoroughly washed
(including a final rinse with deionized water) and left on the return cart. All electrical
apparatus should be switched off and unplugged. All taps should be turned fully off and
all waste should be placed in the appropriate waste container.
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HOW TO READ AN ANALYTICAL METHOD
Preparing Standard Solutions
The procedure for a particular experiment states “make up a series of standards from your
stock solution, from X to Y concentration.” How do you proceed?
There are several important pieces of information hidden in the above instructions.
1. “make up … from your stock solution”: all of your standards will originate from
this solution. In some cases, where a wide range of concentrations is required, you
may need to prepare a ‘sub-stock’ solution (diluted stock), which you will then use
to make your most dilute standards.
2. “a series of standards”: in some cases you will be told to make a certain number of
standards; in others it is left to your discretion. Generally, four or five standards
are used to prepare a calibration curve. You must prepare at least three standards.
3. “standards … from X to Y”: this is the concentration range that your standards
will cover. Your most dilute standard will have a concentration of X. Your most
concentrated standard will have a concentration of Y. Units will depend on the
experiment.
4. the dilutions you use to make your standards must be calculated to ‘fit’ the
glassware available. You will have access to 1, 2, 5, 10, 20, 25 and 50 mL
volumetric pipettes. You also have 25, 50, 100, 200, 250, 500 and 1000 mL
volumetric flasks. Calculate your dilutions so they ‘fit’ this equipment. (e.g., 1.5
mL into a 150 mL flask is an impossible dilution with your equipment. How else
could you get the same dilution factor?)
5. All dilutions used to make standards are done using volumetric glassware.
Analytical Shorthand
Rather than spell out exactly how quantities should be measured every time, analytical
chemists use a shorthand based on the precise use of language and significant figures.
Read through the following examples and ‘translate’. If you can’t see the difference
between the instructions, ask your lab instructor!
Your translation should include: the type of equipment used, the technique used, and the
amount of reagent used.
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Units
A word on units: you will spend a lot of your time as an analytical chemist converting
between units. If you have worked in an analytical lab, you already know about it. Set
up a list of conversions for yourself or create an Excel spreadsheet to do this for you. It
will save you a lot of time and needless errors later.
mg/L
g/mL
moles/L
mg/kg
mg/g
wt/wt %
mg/L
ng/mL
The Plan
You will need to have an experimental plan organized prior to arriving in the lab.
1. The information you have in the lab manual must be reprocessed to create an
“analytical method”, i.e., a plan. Some of that reprocessing is described above, for
example preparing calibration standards.
2. Create your plan using numbered steps or a flow chart, so you can track where you
are. Set it up, if you like, so you can check off each step you complete.
3. Number the steps so you can make the most efficient use of your time in the lab.
4. Be aware of time requirements. For example, if your standard will be made from a
chemical in the drying oven, it will take some time for it to cool. This might be your
first step.
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LABORATORY SAFETY
A chemical laboratory is a potentially dangerous environment; the hazards of fire, cuts, burns
and poisoning being most prevalent. It is a safe practice to assume all chemical reagents are
potentially hazardous. While the use of particularly toxic or carcinogenic reagents is generally
avoided, some of the reagents in this lab are dangerous. Check the MSDS and consult your
instructor for more information. The first line of defence for skin contact is to flush with plenty
of water. Two eyewash stations are provided for the immediate flushing of eye splashes. In the
event of an accident, contact your instructor immediately. Safety rules will work only if you
obey them and encourage others to obey them. Please familiarise yourself with the following
regulations.
Personal Safety
There must be no smoking or eating in the laboratory.
Students must wear safety glasses at all times. Safety glasses are available. Contact lenses
should be removed prior to entering the laboratory. Prescription glasses may be worn, but
should be covered with safety glasses.
Students are recommended to wear laboratory coats in the laboratory.
Many of the chemicals in the laboratory are poisonous whether taken orally or absorbed
through the skin. If any chemical is swallowed the supervisor should be summoned
immediately. If any chemical comes into contact with the skin it should be washed off
immediately with plenty of water.
While heating a substance in a test tube, care should be taken to ensure that the mouth of the
test tube is not pointing at anyone. A student should never look down into a test tube that is
being heated.
Concentrated acids and bases; strong oxidising and reducing agents; flammable solvents and
toxic chemicals should be treated with respect.
Always wash your hands prior to existing the lab and before eating.
Fire
In the event of fire, the flames should be extinguished with one of the extinguishers in the
laboratory and the supervisor notified immediately.
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Material Safety Data Sheets (MSDS)
Material Safety Data Sheets summarize physical and chemical properties of all chemical
reagents used in this laboratory. In addition, the MSDS sheets contain information on the
hazards and toxicity effects. MSDS can be found in the prep room and should be consulted if
there is any question regarding the safety of materials encountered.
1. Standard Methods for the Examination of Water and Wastewater (21st Ed.), APHA, 2005.
2. Water Analysis Handbook, (2nd Ed.), Hach Co., Loveland, 1992.
3. Environmental Sampling and Analysis for Technicians, M. Csuros, Lewis Publishers, Boca
Raton, 1994.
4. Drinking Water Chemistry: A Laboratory Manual, B.A. Hauser, Lewis Publishers, Boca
Raton, 2001.
5. Laboratory Manual for the Examination of Water, Wastewater and Soil (3rd Ed.), H.H. Rump,
Wiley-VCH, New York, 1999.
6. Water Quality and Pond Soil Analysis for Aquaculture, C.E. Boyd, C.S. Tucker, Auburn
University, 1992.
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INTRODUCTION TO GOOD LABORATORY PRACTICES (GLPs)
The following notes on proper experimental technique and use of equipment are collectively
known as GLPs. It is assumed that the student is familiar with and always practices the following
procedures outlined in this section. Your laboratory instructor will be evaluating your experimental
technique. Failure to practice the following will lead to poor results and poor technique. Both of
these are graded. Exceptions to standard GLPs must be noted in your lab report.
Care must be taken to ensure that glassware is thoroughly clean before use. However, recognize
that soap can be a serious chemical contaminant. Do not clean your glassware with soap unless
specifically instructed. In general, glassware will be supplied clean, acid washed and thoroughly
rinsed with deionized water. Properly cleaned glassware is indicated by the presence of an
unbroken film of water on the surface. It is seldom necessary to dry glassware before use; in fact
this practice should be discouraged because it wastes time, can be a cause of contamination and
result in changes in volumetric glassware.
Always pre-rinse burets and volumetric pipettes with the titrant or solution to be transferred prior
to use. Note that rinsing is most effectively accomplished with a greater number of small
portions, rather than a smaller number of large portions.
Rinse all glassware with tap and distilled water after use. Never let reagents dry in volumetric
glassware.
2. Housekeeping
Good housekeeping is important for the safety and convenience of everyone, including the
cleaning staff, who are not chemists. The analytical lab is a shared space. It is a busy place, and
any mess you don't clean up will inconvenience many people. A mess will not impress your lab
instructor and you certainly won't impress any future employers with poor, unsafe work habits.
Don't leave equipment and chemicals scattered over the benchtop; return them when you are
finished using them. This allows others to use the same equipment, and prevents any accidents
involving or resulting from your mess. Fumehood space is often at a premium, so clear out as
soon as you no longer need to work there. When you are finished working, in the fumehood
especially, clean the entire area with a damp paper towel. Clean up all chemical spills
immediately, especially when the balances or other instruments are involved. Wash the outside
of reagent bottles when you are finished using them. Drips and small spills may go unnoticed
until they have had some time to react and cause a burn, at which point it is too late.
Never leave experiments unattended. If you must, especially in the case of reactions and
digestions or anything that involves the use of hotplates and stirrers, inform your lab instructor
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and have them or another student keep an eye on your experiment. Even if you have to leave the
lab for only a minute to go to the bathroom, inform your lab instructor.
When using chemicals and supplies, use all of one container before opening another one, unless
specifically directed to do otherwise by your lab instructor. Inform your lab instructor of any
chemicals or supplies that are running low, including cylinders of compressed gases and printer
supplies, so that there will be enough for the next student to complete their experiment. Also
inform your lab instructor as soon as possible if anything is missing or if anything is broken, so
that replacements can be obtained as quickly as possible.
Dispose of all refuse in the appropriate waste container as soon as possible; if in doubt ask your
lab instructor. Questions are not ‘stupid’; unlike endangering the lives and health of others. At
the end of the lab, return all chemicals and equipment where they belong and clean up your work
area. Double-check everywhere you worked, make sure that all equipment and supplies are
returned, and all waste appropriately disposed (including any bench and fumehood space, the
balances, instruments and sinks where you worked). As far as possible, shut down and turn off
all equipment that you have worked with as soon as you have finished.
There are three grades of water available for use in the lab:
· tap
· distilled/ reverse osmosis
· deionized
The three grades of water are progressively more expensive to produce. In general, use only the
minimum purity of water necessary and do not waste water, particularly the more expensive grades.
Distilled water will suffice for most uses.
Successful analytical work depends on the purity and quality of the available reagents. A freshly
opened reagent container can be used with confidence; whether or not the stated assay values for
purity and impurities remain valid depends entirely on how the container has been handled since
being opened. The purity of the chemical reagents available (and the quality of your results and,
therefore, your marks) depends on strict adherence to the following rules.
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Use small pre-cleaned beakers (in 10 and 20 mL sizes) for pouring out reagents. Any solid or
liquid not used should be disposed of appropriately.
1 The possibility of contamination can be minimized by choosing the smallest bottle that
will supply the required quantity of reagent. Ensure that the purity of the reagent is
sufficient to prevent contamination and interferences and to reduce the blank to a
minimum. In general, try to use only reagents whose purity you can be certain of (check
assay values on the label).
2 Replace the top of the reagent container immediately after removal of the reagent.
3 Hold stoppers between the fingers; stoppers should never be set down on the bench or
anywhere else except in the neck of the appropriate flask or reagent bottle.
4 Unless specifically directed to the contrary, never return any excess reagent or solution to
a reagent bottle. Contamination of the entire bottle by returning excess reagent is a false
economy - considerable time can be spent determining the source of any contamination
(and consequent poor results), and the entire bottle then has to be disposed of and a new
bottle obtained.
5 Before taking a bottle of chemical to the weighing room, first clean your spatula with
deionized water. Scrub it dry with a Kimwipe. Now take your solid chemical to the
weighing room and weigh it.
6 Keep reagent storage areas and the balances clean. Clean up any spilled chemicals
immediately.
7 Do not use a new or unopened bottle of reagent without first obtaining the permission of
the lab instructor.
5. Handling Solids
5.1 Balances
Both analytical and top-loading balances are available. Use the top-loading balances
wherever possible, i.e., whenever the weight does not have to be exact or where an
uncertainty of 0.01 g will suffice. For all work where the weight has to be known as
exactly as possible, use the analytical balances.
Electronic Balances:
Most of you will have used the self-taring electronic balances in other courses. In
general:
• Keep the balance pan and surrounding areas clean.
• Weigh solids into a container that is as light as possible.
• An initial tare weight is usually unneeded, since you can tare the container to zero.
• CLEAN UP when you are done
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Detail: weighing solids
Ensure that the solid is of fairly uniform texture and will readily pour. If necessary,
shake the capped reagent bottle and/or tap it on a wooden surface to break up any large
lumps to allow the reagent to pour freely. Remove the cap and pour a slight excess into a
clean, dry beaker or weighing boat. Immediately replace the cap on the reagent bottle
and tighten it. Weigh reagent from the beaker or weighing boat into the desired
container(s). If necessary, obtain additional solid from the reagent bottle as described
above. When finished, return the sealed reagent bottle and properly dispose of the excess
reagent.
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likely to occur, the interior of the desiccator now being under a partial vacuum. Both of
these conditions can cause the contents of the desiccator to be physically lost or
contaminated. Although it defeats the purpose somewhat, it may be best to allow some
cooling to occur before finally sealing the lid. It also helps to break the seal several times
during cooling to relieve any vacuum that may be forming.
6. Measurement of Liquids
The reliable measurement of volume is usually performed with the pipet, the buret and the
volumetric flask (weighing is accurate but time-consuming). Pipets and burets are ordinarily
designed and calibrated to deliver specified volumes, whereas volumetric flasks are calibrated on
a "to contain" basis. Volumetric equipment is marked by the manufacturer to indicate not only
the manner of calibration (usually with a TD for "to deliver" or a TC for "to contain") but also
the temperature for which the calibration strictly refers. The volume occupied by a given mass
of liquid varies with temperature, as does the volume of the container holding the liquid. As a
general rule, volumetric glassware should not be heated because the calibration can be
permanently altered.
6.2 Pipets
There are different types of pipets. Most of the work in this course involves the use of
volumetric pipets which deliver a fixed volume of liquid and graduated pipets. Carefully
inspect graduated pipets to see if the graduations extend to the tip of the pipet - if they do
not then the volume of the tip is not calibrated and the pipet should be drained only as far
as the lowest calibration line. Liquids are drawn into pipets through the application of a
slight vacuum. Never pipet anything by mouth, use a pipet bulb.
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Carefully inspect the pipet for damage, especially the tip. Consult your lab instructor if in
doubt about damaged pipets.
In general, when planning dilutions, attempt to use the largest volumetric pipet that is
practical. For example, making a 1/10 dilution with a 5 mL pipet and 50 mL flask will be
less accurate than making the same dilution using a 10 mL pipet and a 100 mL flask. For
even better accuracy, you could use a 25 mL pipet and a 250 mL flask! These guidelines
apply primarily to your standard solutions, and particularly to any that will be used for
subsequent dilutions. If in doubt about your ‘dilution plan’, speak to your instructor
before the lab.
Note that only the upper part of the pipet should be handled. Do not touch or hold the
bottom of the pipet to avoid contaminating the liquid being pipetted (and yourself!).
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Table 1: Tolerances of Class A transfer (volumetric) pipets
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6.3 Micropipettors
Micropipettors are commercially available in various sizes; some deliver a fixed volume
while others are adjustable within a given range. Common volumes for micropipettors in
an analytical lab are 5 uL to 2000 uL. Micropipettors use disposable plastic tips. To
prevent contamination of these tips, insert the clean pipettor (not your hand) into bag
containing the tips and using your hand on the outside of the bag, slide a tip into place.
Once the disposable tip is firmly in place, the micropipettor is ready for use. Depress the
button at the top of the micropipettor to the first stop position and place the tip into the
liquid to be transferred. Release the button to pull up the solution and then remove the
micropipettor. Place the tip into the receiving vessel and then depress the button at the top
of the micropipettor to the second stop position. Depressing to the first stop ejects most of
the solution drawn up, while the second stop ‘blows out’ the remaining solution. Some
micropipettors have a third stop position which ejects the tip.
Fixed volume micropipettors are more accurate than the adjustable micropipettors.
Typical values for accuracy and precision are given for Eppendorf™ brand
micropipetteor.
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Table 2: Fixed volume micropipettor specifications
Pipette volume (L) Accuracy (%) Precision (%)
5 ± 1.5 < 0.8
10 ± 1.0 < 0.5
50 ± 0.7 < 0.3
> 100 ± 0.6 < 0.2
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6.5 Volumetric Flasks
Fill the flask until the bulb of the flask is almost full. Stop and swirl the solution to
achieve adequate mixing. Bring the liquid level almost to the mark and allow time for
solutions to drain from the neck of the flask (and for thermal expansion to room
temperature, if necessary). Use a pasteur pipet to carefully dilute to the mark. Firmly
stopper the flask, and invert repeatedly (do not shake) to assure uniform mixing. For
storage beyond one day, transfer the contents to a clean, dry storage bottle or one that has
been thoroughly rinsed with several portions of the solution from the flask. If you are
using aqueous solvents, it is unnecessary to dry glassware; rinse glassware with deionized
water. In general, volumetric glassware (pipets and volumetric flasks) should not be
placed in a hot oven to dry as the thermal expansion may distort accuracy.
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Burets are designed to deliver liquids primarily for the purpose of volumetric titrations.
They are still widely used in an analytical laboratory and considered a ‘reference’ method
for many analytes. A standard 50 mL buret is divided into 1 mL graduations with 0.1 mL
sub-divisions. Leveling the eye with the bottom of the meniscus, an analyst should be
able to interpolate volume readings to within 0.02 mL with a high degree of
reproducibility.
Filling burets
• Load 50 mL burets using a funnel.
• Rinse inside of buret including the valve and tip, with three small portions of titrant
to be used.
• Fill the buret and run some titrant into a waste beaker checking for air bubbles in the
tip.
• Make sure all air bubbles have cleared.
• Let the solution level stabilize and record the initial volume to nearest ±0.02 mL.
Titrating
• Control the stopcock valve with your non-dominant hand. This allows for finer
control of the valve and swirling of the receiving flask with your dominant hand.
• If you know (by calculation or experience) the approximate volume to be delivered,
you can add ~80% quickly and then slow down as you approach the endpoint.
• If the endpoint is to be determined by a colour change, you will observe a temporary
change which disappears with swirling as you get close to the endpoint.
• To add less than a drop of titrant as the endpoint approaches, carefully adjust the
stopcock valve until titrant just begins to flow. Close the valve and wash the hanging
drop into the receiving flask with a wash bottle.
• Repeat the above step until the endpoint colour change is just barely visible and
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permanent for more than 30 sec.
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Follow the instructions below for the use of digital titrators:
1. Choose the appropriate titrant Reagent Cartridge for the analyte and method chosen.
Be sure to check both the titrant identity and concentration. Record the sample
volume and the digit multiplier to be used.
2. With the titrator plunger fully retracted, slide the Reagent Cartridge into place.
3. Remove the Reagent Cartridge tip cap and replace with a Delivery Tube.
4. Depress the button on the plunger slider and slide the plunger until it meets resistance.
5. With the titrator in the vertical position (tip up) continue to slide the plunger by hand
or by cranking the Delivery Knob to remove all air bubbles from the Reagent
Cartridge and the Delivery tube. This will require wasting some of the titrant.
6. Wipe the outside of the Delivery Tube with a Kimwipe™ to remove excess titrant.
7. Re-set the Digital Counter to zero.
8. Titrate the sample to the specified endpoint.
9. Record the number on the Digit Counter and convert this to concentration of analyte
using the multiplier appropriate for the particular method.
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8. Sample Sequencing
Analytical laboratories have strict guidelines for sample analysis, which are required to meet
various quality control standards. For CHEM 311, you will use the following sequence in all
experiments, unless otherwise noted. In some cases, you may not have separate calibration and
method blanks. Try to run at least one sample in triplicate; this will allow you to calculate precision
estimates.
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LAB 0: GOOD LABORATORY PRACTICES EXERCISE
OBJECTIVE
EXPERIMENTAL PROCEDURE
[HSO3NH2]Stock =
Next, prepare a ~0.1M standard solution of sulfamic acid by diluting the appropriate volume of
stock solution with deionized water in another 100 mL volumetric flask. Note: the final
concentration should be known as precisely as possible (it is a standard solution), but need not be
exactly 0.1000M (i.e., 0.09981 M or 0.1022 M are completely acceptable values).
[HSO3NH2]Standard =
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Finally, standardize the unknown sodium hydroxide solution provided by titrating a 25 mL
aliquot of standard sulfamic acid with the NaOH solution. Use 3 drops of indicator solution
(phenolphthalein) to visualize the endpoint of the titration by the appearance of a persistent light
pink color. The stoichiometry of the reaction is:
[NaOH] =
* from the titration of 25.00 mL of ~0.1 M sulfamic acid standard solutions using a
phenolphthalein endpoint
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