CHM 2354

LABORATORY OF ANALYTICAL CHEMISTRY LABORATORY MANUAL

Winter 2014

Department of Chemistry University of Ottawa

IMPORTANT NOTICE TO ALL STUDENTS. Safety glasses/goggles are mandatory in all Chemistry laboratories. Students can purchase this safety item from the storeroom - MRN 308. Students will NOT be allowed to work in the laboratory without their safety glasses. Lab coats are mandatory. No contact lenses, sandals, or open shoes may be worn in the laboratories.
AVIS IMPORTANT A TOUS LES ETUDIANTS. Les lunettes protectrices sent obligatoires dans tous les laboratoires de chimie. Les etudiants peuvent acheter cet article securitaire dans notre magazin MRN 308. Aucun etudiant ne sera admis pour travailler dans le laboratoire sans ses lunettes.

For all the laboratory sessions you will require:

Safety glasses and Lab coat Lab manual 2 Good quality bound notebooks (Hard cover preferred, but spiral bound accepted) Calculator Graph Paper (if your notebook has none) Permanent marker Ruler A University computer account YOU WILL START THE LABORATORY WITH EXP. 1 ON THE VERY FIRST SESSION. BE PREPARED.

Contents
General Information Laboratory Schedule Laboratory Notebooks Calculations and Statistical Analysis in Analytical Chemistry Experiment #1. Calibration of Apparatus Experiment #2. Analysis of Aspirin Requirements for Formal Report for Experiment 2 Experiment #3. The Determination of Sodium Carbonate in Soda Ash Experiment #4. The Determination of Chloride by the Volhard Method Experiment #5. An investigation of interferences in the determination of Ca2+ in water by Flame Atomic Absorption Spectrophotometry Experiment #6. Determination of iron in multi-vitamin tablets by colorimetry and verification of their consistency between two production batches. 3 4 5-8 10-15 16-24 25-30 31-34 35-38 39-42 43-49 50-54

General Information
Time and Location: Lab (1) Monday 8:30 am 12:30 pm, MRN 3rd floor Lab (2) Monday 2:30 pm - 6:30 pm, MRN 3rd floor Absenteeism: Attendance is compulsory at all laboratory sessions. A student is required to produce a medical certificate from their doctor or campus health center if absent from any session. No other excuses will be accepted. Students who do not have a valid reason for missing a lab session will not be allowed to make up the time. Students who miss more than two lab sessions will not receive a grade for the course. Grading: 75% Accuracy (60%) and Precision (15%) of Results The accuracy mark will be based on the closeness of your reported value to the actual value The precision mark is based on the standard deviation in your unknown value If you miscalculate your final result, you will be allowed to recalculate it and hand it in for a re-grade (once!) with a 10% penalty to your mark. Resubmissions will only be accepted up to 2 weeks after handing in your result. No resubmissions will be accepted after that point. Laboratory Notebook Long Report for Experiment 2 TA evaluation

15% 5% 5%

There is no final exam for this course. Required Text: Laboratory Manual

Other suggested textbooks: Exploring Chemical Analysis, 5th Ed. Daniel C. Harris Quantitative Chemical Analysis 3rd Ed. or later, D.C. Harris.

Ensure that you read all relevant material in this manual and prepared any work required before attending a laboratory session.

Lab Schedule You will be performing experiments throughout the semester by following the schedule provided below. Take note of what bay your lab bench is located in on the first day, and cycle through the experiments in this manual according to the schedule. It is possible that you will be performing experiments based on theory and material that has not yet been covered in class. All topics and techniques are nonetheless covered in your required textbook, and outlined in the Required Reading for every prelab. Week Bay A Bay B Bay C Bay D Bay E Bay F Jan 6 No lab session Jan 13 Expt 1 and general orientation 20 5 2 2 2 2 5 27 5 2 2 2 2 5 Feb 3 3 5 3 3 3 3 10 3 5 3 3 3 3 17 Study Break 24 4 4 5 4 4 4 Mar 3 4 4 5 4 4 4 10 2 3 4 5 6 2 17 2 3 4 5 6 2 24 6 6 6 6 5 6 31 6 6 6 6 5 6
*

Please write your final result for each experiment on the result sheets provided by your TA before leaving lab. Sign your name next to your result to testify to the following: The result you are submitting is the same as that written in your notebook You have not in any way falsified data or fudged calculations You have double-checked your calculations for accuracy

You must record the day and time of your submission on the result sheet. Late submission or changes/corrections to submitted results will result in a 10% decrease in your result mark. Such changes must be recorded on a new result sheet and signed by you. They may not been sent to your TA by email.

Format of Course: This course is designed to teach you skills in handling substances and apparatus in a quantitative manner. It may be necessary for two students to share the equipment contained in an assigned locker but each student performs the experiments independently unless otherwise stated. There are six experiments. The first is devoted mainly to calibration of glassware. The object of the remaining five experiments is to analyze "unknown" samples for various components. Each student will be given a coded sample for each analysis. Do not interchange samples or code numbers with the person who shares your locker. A failure for that analysis is likely to result if you are not careful about this. If you repeat an analysis, you must obtain a fresh sample and sample code. Also, if you spoil an analysis through spilling, etc., you should abandon it and start over with a fresh sample. Laboratory Notebooks: You are required to keep 2 bound, hardcover laboratory notebooks. (You will use alternate notebooks for alternate experiments). You are required to write all instructions into this book in a way that you will be able to follow them in the lab. Do not copy the instructions verbatim, or use a photocopy. You will not be allowed to use the laboratory manual in the lab. Notebooks should be kept according to the standards of a research laboratory notebook, given below. Adherer to them strictly or you will be harshly penalized! An example of a good notebook entry can be found on page 8. DO the following: Use the first 2 pages of the notebook as a table of contents. Each time you record a new experiment, write the experiment title, date, and page number in the table of contents. Number the pages of the notebook Recorded all data, notes, and observations directly into your notebook in ink. If you make an error in a notebook entry, cross through the error with a single line so that the information underneath the line is still legible. If you skip a page in the notebook, draw a single diagonal line through that page. Start each new experiment on a new page. All work in your notebook should be legible and set out in a way that someone who has not performed the lab will be able to understand your results. Provide references for any published procedures that you have followed or other sources of information DO NOT do the following: Write data or calculations on loose-leaf sheets of paper then copy them into your notebook later. Record data in pencil Scribble through errors, erase errors, or use whiteout in your notebook. (In a research lab, such practices could result in the loss of a patent!)

The following information should be recorded in your notebook for each experiment: 1) Title and date of experiment 2) Before coming to lab, the following information should be written into your notebook: Answers to prelab questions found at the beginning of each experiment A flow chart and instructions for the laboratory experiment (you will not be allowed to use your laboratory manual during the lab period). See page 7 for sample flow chart. Sample calculations (e.g. rough titres) Prepare any data tables that you will need to use during the lab. 3) Experimental measurements, data, and observations: Sample number (for unknowns) All measurements (with units indicated) Data should be written clearly and legibly in tables (see sample notebook page) Any changes to the experimental procedure Errors made while performing the experiment or problems encountered 4) Calculations and results: A sample of each type of calculation, including units and uncertainty Calculations of standard deviation Overall result and error with standard deviation (uncertainty) should be reported with appropriate number of significant figures. 5) A brief discussion of your results and their significance 6) Any rough notes or calculations should also be recorded in the notebook (not on scrap paper). This is the diary of your work and your thought processes. Your notebook must be handed in at the end of the lab period on the last day of each experiment. Late submissions will be penalized: 10% off for the 1st day late, 20% off if more than 1 day late to 1 week late, 40% off if more than 1 week late.

Sample Flow Chart:

Dry sample 1hr ! 110oC

Put glass filter in gooch crucible

Make sure holes in crucible covered Weigh 3x 0.025g sample (accurately) Place in 400 mL Beaker (unscratched)

Crucibles in beaker with watch glass lid [Heat for 45 min at 150oC. Cool 20 mins and weigh (accurately)] Repeat

Add 20 mL 1:1 HCl (10 mL HCl added to 10 mL water)

No

Successive weighings < 0.0003 g difference?

Etc.

Yes Done

Sample Notebook Entry:

Laboratory Etiquette and Safety: Tidiness is essential for good work. Spilled chemicals must be cleaned up immediately. Balances must be kept especially clean. They are very expensive. Most chemicals spilled on the balances are corrosive. Corrosion of the weigh pan will result in errors in masses weighed - this will affect your results. All masses must be taken on a watch glasses, in glass vessels, or in we weighing boats. Do not use paper for weighing. Acid-containing solutions must never be heated on an open bench. Use a fume hood, with the sash lowered, and keep your head away from the opening. Out of consideration to your locker partner, as well as in your own interest, keep locker and equipment clean. Mistakes are frequently made when incorrect chemicals are used. Make sure that you know which chemical you require. The labels on the bottles are not always the same as in the laboratory manual. Be careful - similar names are not always the same compounds.

IF AT ANY TIME YOU ARE UNSURE OF WHAT TO DO, ASK AN INSTRUCTOR!

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CALCULATIONS AND STATISTICAL ANALYSIS IN ANALYTICAL CHEMISTRY

Suggested Reading: Exploring Chemical Analysis, 5th Ed. Daniel C. Harris, Chapters 3 and 4.

I. TYPES OF ERROR Two types of errors normally affect chemical analyses. 1) Random error Random error causes data to be scattered more or less symmetrically around a mean value. This type of error is caused by the many incontrollable variables that are an inevitable part of every physical or chemical measurement. It estimates the precision of a determination and is calculated as standard deviation. The uncertainty of a measurement is directly related to this type of error. 2) Systematic error Systematic error a set of data to differ from the expected true value. There are three types of systematic errors: instrumental, method or personal error. The magnitude of this error estimates the accuracy of the result and can be calculated as absolute or relative error: absolute error = |accepted value found value| relative error = (abs. error)/ (accepted value) x 100 The magnitude of random error can be decreased by increasing the number of replicates while that of the systematic error, by checking the instruments (calibration), the method (interferences, trying other methods), analyzing reference materials, etc. II. REVIEW OF SIGNIFICANT FIGURES The significant figures in a digit include all of the certain digits and the first uncertain digit. e.g. For a burette reading where the liquid is approximately half way between 30.2 mL and 30.3 mL, the volume can be reported as 30.25 mL. The last digit (5) is the only uncertain digit. The other digits are certain. In reporting standard deviations, usually only 1 significant digit is needed. e.g. The above burette reading should be reported as 30.25 0.02 mL Significant Figures in Addition and Subtraction 24.3 + 1.34 + 1.6 = 27.24 = 27.2 24.3 and 1.6 have only 1 significant digit after the decimal. Therefore, the answer can only have 1 significant digit after the decimal.

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Significant Figures in Multiplication and Division The final answer should be rounded so that it contains the same number of significant digits as the original number with the smallest number of significant digits. e.g. (24 x 4.02)/100.0 = 0.965 = 0.96 Rounding If the number after the last significant digit is a 5, the round to the nearest even number: e.g. If we can only have 2 significant digits: 3.45 is rounded to 3.4 3.55 is rounded to 3.6 Since burettes can be read to 2 decimal places, and the electronic balance and volumetric glassware is accurate to 4 significant figures, it is reasonable to quote 4 significant figures in calculations. To avoid rounding errors, you should keep more figures on your calculator, but do not write these down in the report. Always round your answer at the end of the calculation to no more than 4 significant figures. The number of significant figures in your final answer will depend on your calculated standard deviation for the replicate analyses. For example, 38.72% 0.1% should be written as 38.7 0.1% since the error is in the third figure, so only 3 figures are significant. Standard deviations should be quoted to 1 significant figure only. III. STATISTICAL ANALYSIS For every experiment, you will be expected to calculate the mean result and the standard deviation of the result (by both methods: standard deviation of replicate results and cumulative standard deviation). If a result appears to be an outlier, you can conduct a Q-test to determine whether the result can be discarded. In your experiment 2 report, you will also be required to conduct t-tests and create a Shewhart Chart. A. Mean:
N

x=

! xi
i =1

x is the measured value N is the number of replicate measurements

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B. Standard Deviation for Replicate Results If several measurements have been taken, the standard deviation in the result is determined using this formula:

s=

" ( xi ! x ) 2
i =1

N !1

This equation can be alternatively (and equivalently) expressed as:

'N x $ %( i " N & i =1 # 2 ( xi ! N s = i =1 N !1

C. Cumulative Standard Deviation for an Individual Result For a single result, the standard deviation (or error) in the result is determined as the accumulation of errors (or standard deviations) in the individual measurements that contributed to the final result. The method for calculating the error in a single result depends upon the mathematical operations performed in calculating the result. In the following examples, y is the result, and sy is the error in the result. i) Addition and subtraction: y = a + b c (where y is the result)
2 2 s y = sa + s2 b + sc

ii) Multiplication and division:

y = (a x b)/c

sy
y = ax

&s # &s # = $ a ! + $ b ! + ... y %a" %b"

sy y sa a

iii) Exponential:

=x

iv) Logarithm:

y = log10a

s y = 0.434

sa a

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D. Relative Standard Deviation This is an alternative way of expressing standard deviation

RSD =

s x

It can also be expressed as a percent (often called coefficient of variation)

s % RSD = ! 100% x
If the %RSD for the replicate results is less than the cumulative %RSD for each reading, then it is meaningless, since the %RSD for replicate results cannot be more precise than the cumulative %RSD, which indicates the level of error from the equipment used. In every report you should calculate and discuss these two measures. E. G-test If any of your results appear to be outliers (i.e. a single value that is considerably larger or smaller than the rest of the results), determine whether to reject this result using a G test. The appropriate formula is:

G=

questionable value - x s

where x is mean of all values s is standard deviation of all values Determine Gt from the table of G values (Harris, pg 90) for the appropriate number of replicate results (You will want to use the 95% confidence level). If Gcalculated is greater than Gtable then the questionable value may be rejected. You should have a minimum of 4 values to perform a G test. Once you have rejected a value you cannot use the G test to reject a second value.

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F. t-Test for comparing 2 experimental means

t=
where

x1 ! x 2 s

N1 N 2 N1 + N 2

(1)

N1 is the number of replicate measurements in data set 1 N2 is the number of replicate measurements in data set 2

s is the pooled standard deviation, described by formula (2) below

x 1 is the mean value from data set 1 x 2 is the mean value from data set 2

s=

set1

" ( xi ! x1 ) 2 + " ( x j ! x 2 ) 2
set 2

N1 + N 2 ! 2

(2)

The value of s is a pooled standard deviation making use of both sets of data. The value of t from equation (1) is to be compared to the value of t in Table 1 below for N1 + N2 2 degrees of freedom. If the calculated t is greater than the tabulated t, the two results are significantly different at the confidence level in question. Table 1: Values of t for Various Levels of Probability (Also see Harris et al. p. 84)

G. t-test for comparing an experimentally-determined mean to a true value

t=
where

x ! xt s/ N

(3)

is the experimental mean

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xt is the true value s is the standard deviation in the experimental data N is the number of replicate measurements The value of t from equation (3) is to be compared to the value of t in Table 1 above for N-1 degrees of freedom. If the calculated t is greater than the tabulated t, the two results are significantly different at the confidence level in question. H. Shewhart Charts: Population Statistics The Shewhart Chart or Control Chart (Harris, pg. 107) is used to indicate whether multiple measurements taken are subject to random errors only, and that no systematic errors or loss of accuracy have occurred during the process. An example of this type of chart is shown below. The data plotted is for a thiosulfate standard solution, the concentration of which was determined by taking several measurements over a period of time. The solid line is the target concentration of the thiosulfate, - this is the accepted concentration of the standard. The dotted lines are confidence limits for the target value. The upper and lower warning lines are at 2s (95.5% confidence limit) and the upper and lower action lines are at 3s (97% confidence limit). If the process is under statistical control, then all measurements will be within the upper and lower action lines. Any point between the warning line and action line is subject to a large random error, whereas a systematic error is indicated by a point outside of the action lines, or two or more points between the warning and action lines on the same side of the mean. 10 successive points on the same side of the mean are also indicative of a systematic error. Whenever a systematic error is suspected, the process is halted and corrective measures are taken. Control Chart for Thiosulfate Standard
Thiosulfate Standard
1.05

Concentration / M

1.03 1.01 0.99 0.97 0.95 0 1 2 3 4 5 6 7 8 9 10 11 Measurem ent Num ber Target Line Action Line Warning Line

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EXPERIMENT 1 CALIBRATION OF APPARATUS

Suggested Reading: Exploring Chemical Analysis, 5th Ed. Daniel C. Harris, Chapter 2. PRELAB QUESTIONS 1. When and why would an analytical balance, that reads to 0.1 mg, not be used for weighing substances used in an analysis? 2. Define: (a) weighing by difference, and (b) absolute weight. What is the difference between these? Which is commonly used in analysis and why? 3. On scientific grounds, why is it preferable to use only one particular balance throughout the whole course? 4. Volumetric glassware that is used for accurate work should never be heated in an oven in order to dry it because glass exhibits volume hysteresis upon changing its temperature. Explain this. How can an analyst make use of wet burettes, pipettes, etc. 5. After having dried a solid sample in a weighing bottle, it was placed on the pan of an analytical balance while still warm (~ 40oC). The operator could not obtain a steady balance reading (i.e. the weight appeared to drift). Why? In which direction was the drift? 6. What are the three corrections that are required in order to determine the volume of a vessel by weighing its water content? 7. The figure below shows the mean errors in the burette volume at different apparent volumes. Error = apparent volume - true volume Apparent volumes are the volumes read directly from the burette. True volume is the actual volume that is delivered from the burette. You have just finished three different titrations and your volume readings are: I II III V1 = 12.43 cm3 V2 = 20.06 cm3 V3 = 34.51 cm3

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Using the plot below determine the true volumes that were delivered from the burette: V1corr, V2corr, and V3corr.

8. You are calibrating a burette. Your readings are: Volume Interval /mL 0 - 10.02 0 - 19.97 0 - 30.04 0 - 40.03 0 - 49.91 Corrected mass of water at 20oC /g 9.980 19.897 30.012 40.001 49.650

A. From the corrected mass of water in column 2, calculate the true volume of water delivered in each measurement. (Use table 3.7 page 50 for conversion of mass to volume) B. From the true volume obtained in step A and the corresponding apparent volume found in column 1 of the table above, calculate the error associated with each measurement. Error = apparent volume - true volume C. Plot your calculated errors as a function of the apparent volume (as shown in the graph in problem #7). 9. A typical burette is shown below. Read the burette scale and calculate the volume of liquid added from the burette, assuming the initial reading of the burette was 0.24 mL. Make sure you read the scale to 2 decimal places.

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10. Calculate the standard deviation for the following data: 12.3423 mL, 12.3411 mL, 12.3454 mL, 12.3425 mL

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EXPERIMENT 1 CALIBRATION OF APPARATUS

Objective: To learn techniques for calibrating high precision equipment, and to develop good technique for measuring masses and volumes using this equipment. Some points of technique Various regulatory bodies as well as major industries recognize and define Standard Procedures. These analytical methods have been proven in research programs to give reliable results under normal circumstances and in capable hands. A precision of 0.1 - 0.2% and an accuracy of ~ 0.3% can be expected from the better volumetric and gravimetric procedures. If these goals are to be met, certain points of technique must be adhered to. The most serious common flaw is found in dirty volumetric glassware. If droplets of liquid remain adhering to the walls of a burette or a pipette, the accuracy cited above is impossible. The glassware should be washed with detergent, rinsed with tap water and finally rinsed with a little distilled water. It is not necessary to rinse with copious amounts of distilled water. Do not dry the glassware with paper towels. Small pieces of lint may contaminate your experiment. Wet glassware can be rinsed with the solution you are using to prevent any changes in the solution concentration. When a high degree of precision and accuracy is required, the following high-precision equipment should be used: analytical balance, burettes, pipettes, volumetric flasks. Use of other equipment (such as top-loading balance, graduated cylinders, beakers) will result in lower precision and accuracy. I. The Analytical Balance Electronic analytical balances are simple to use, and are capable of weighing accurately to 0.1mg (0.0001 g). A description of a balance is given on pp. 37 in Harris. A description of general weighing technique is as follows: 1. Switch on the electronic balance (if not already on) and allow the balance to warm up. 2. Calibrate the balance. Consult your TA for directions. 3. Always label your weighing bottles, flasks etc. before you use them. Use a glass marker or a permanent marker to do this. 4. Never touch the vessels being weighed with your hands. This will transfer grease and moisture to the vessel or result in the solution being warmed, and result in inaccuracy. 5. Never try to weigh an object that is heavier than the weight limit of the balance (~ 200g for the analytical balances)

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A. Weighing by difference: 1. Place the substance to be weighed in a dry weighing bottle, and weigh the bottle, lid and contents. 2. Hold the bottle above a beaker, remove stopper and transfer the sample with the aid of a spatula to a beaker. If some powder adheres to the spatula, tap it against the rim of the beaker and, if necessary, rinse adhering crystals into the receiver by a stream from a wash bottle. Replace stopper and re - weigh the bottle and contents. Repeat for subsequent samples. Number the beakers. Weights should be recorded in a your laboratory notebook. B. Using a weighing boat: 1. Plastic weighing boats may also be used to weigh samples. A clean dry weighing boat is placed on the pan and the scale is tared. 2. The appropriate amount of sample is transferred to the weighing boat, and the accurate reading taken. 3. Transfer the sample directly to the beaker, and wash out the weighing boat with a little water. Care must be taken to insure that all solid and washing solution is transferred directly to the beaker. Accurate weights cannot be made on paper or other porous material. Check the zero setting of a balance before and after a weighing. If there is a difference, check for a fault or average the empty pan readings and subtract this from the weight. II. Volumetric Apparatus (Harris, pp. 43 - 46) For accurate analytical work it is necessary to standardize volumetric glassware such as pipettes, burettes and volumetric flasks. Glassware is manufactured in two grades of accuracy - Classes A and B, which are marked on the glass. Class A is sufficiently accurate for most work. The tolerances permitted for Class A equipment is given in Harris, pp. 43. Tolerance is usually defined as twice the standard deviation for the replicate measurements. With large vessels, errors of the order of tenths of a mL are but a small percentage of the total volume and are of little concern. Thus the 250 and 500 mL volumetric flasks are accurate enough for most purposes. When using volumetric flasks to make up solutions, always make sure any solid is dissolved in about half the volume of liquid before dilution to the mark. Always add the last little bit of liquid with a pasteur pipette, not from a wash bottle or you will add too much liquid. Mix well by inverting the flask several times. The technique of pipetting should be taught by an instructor. Several points are as follows (see Fig. 2-6, pg. 41 in Harris reproduced below): 1. Keep the tip well below the surface of the liquid while withdrawing an aliquot. Draw the liquid up to slightly above the calibration line. Always use a rubber bulb to do this - never mouth pipette. 2. Lift the pipette from the solution and wipe the end with tissue. Do not allow the tissue to touch the hole, or you will draw out some of the liquid by capillary action. 3. Adjust the liquid to the calibration line with your index finger, never your thumb. Your finger should be dry while doing this. Avoid the error of parallax by making sure your eye and the

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calibration line is at the same level. Touch the inside of the vessel with the tip to remove any adhering drop of liquid. 4. Transfer the pipette to the receiving vessel. Retard the outflow with your finger so that the delivery time is 10-15 seconds. Keep the tip clean as you lay the pipette down. Do not heat pipettes in the drying oven. If you use a rubber bulb for filling, do not insert the pipette stem into the bulb. Merely hold the stem against the bulb at its opening. Ensure that no liquid enters the rubber bulb. This will happen if you suck up some air into the pipette. 5. Do not blow out a pipette; it should retain some liquid as in Fig. 2.22d. When delivering liquid, finally touch the tip to the wall of the receiving vessel.

Figure 2.22 The technique of using a burette involves attention to the following points:

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1. Do not fill exactly to 0.00 mL as this is a waste of time. It is far more accurate to read the volume in the burette than to try to adjust the level. 2. Take readings at the bottom of the meniscus with the aid of a half-blackened card or one with black lines upon it. Make sure you are at eye level. (Ask for a demonstration) 3. Wipe the outside of the burette with tissue after filling and always remove the funnel from the top before you take any measurements. 4. Watch for slow leaks from the stopcock and check for any bubbles around the stopcock. These can be removed by rapidly turning the stopcock 180o until no more bubbles are observed. 5. Watch for air entering the stopcock. 6. Use your left hand to operate the stopcock while the handle faces your right. You will find this awkward at first but excellent later as you can then swirl the conical flask with your right hand while titrating. This technique is shown in the diagram below.

Experimental Procedure In this exercise, the burette will be calibrated at 10 mL intervals and the 25 mL pipette will be calibrated as well. Read the pertinent sections of the textbook before proceeding (p. 48-49 Harris). You will see that the calibration procedure, pp. 49, entails weighing the quantity of distilled water that is contained or delivered by the vessel. Glassware is calibrated by the manufacturer at a standard temperature of 20oC. The user upon recalibrating more precisely, must note the temperature of the water being used and carry out calculations making corrections for buoyancy. In using a volumetric vessel, the temperature of the liquid should be noted. Table 28.3 f (given below) shows the corrections that the analyst should make when using a vessel at a temperature other than the temperature of calibration. It will be seen that at times other than mid-summer, the variation of laboratory temperature is unlikely to cause a variation greater than 0.1%. For this experiment, glass beakers can be dried in the oven, but you must ensure that they are at room temperature when you come to use them. Do not dry them with paper towels.

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A. Calibrating the Pipette 1) Fill a 250 mL beaker with water from the sink (use both hot and cold water to adjust the water temperature as close as possible to room temperature). Allow the water to reach thermal equilibrium with its surroundings (i.e. room temperature). This will take around 20 minutes. Check the temperature every 5 minutes until it remains constant. Make a note of each temperature reading in your lab notebook. 2) Weigh a clean 50 mL erlenmeyer flask (it does not have to be dry). 3) Using correct pipetting techniques, transfer 25 mL of your temperature-equilibrated water to the erlenmeyer flask. 4) Reweigh the erlenmeyer flask with the water in it. Calculate the mass of the water delivered (the difference between the masses of the empty and filled flask). 5) Record the water temperature at the time the calibration is done. 6) For each mass of water delivered, determine the true volume of water corrected to 20oC. The true volume can be determined using Table 28-3 (above) as follows: Find the temperature of your water in column 1 of the table. In column 3, find the volume (corrected to 20oC) that corresponds to the temperature in column 1. 7) Empty the contents of the erlenmeyer back into your 250 mL beaker to be reused in later trials. 8) Repeat steps 2 through 7 two more times. It is not necessary to dry the erlenmeyer flask between trials, but it should be reweighed after its contents are emptied and before the next sample of water is added. From your three calibrations, calculate the mean true volume delivered by the pipette and the standard deviation from the mean. Compare your results with the manufacturers tolerance. (Note:

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the manufacturers tolerance is two times the standard deviation.) A table of the relevant values is given below. Manufacturers tolerance of Class A volumetric glassware
Volumetric glassware 5 mL pipette 25 mL pipette 50 mL burette 100 mL volumetric flask 500 mL volumetric flask Manufacturers tolerance, mL 0.01 0.03 0.05 0.08 0.20

B. Calibration of the Burette 1) Fill the burette with temperature-equilibrated water, and make sure that no air bubbles are trapped in the tip of the burette. Drain water from the burette as needed to remove air bubbles from the tip. 2) Fill the burette to near the 0 mL mark. Do NOT adjust the volume to exactly 0.00 mL. Attempting to do so will introduce bias into the measurement and will waste time. Read and record the volume accurately to 2 decimal places. 3) In order to check for leaks, wait 5 minutes then record the volume again. If there is no leakage, the 2 readings should be identical. 4) Weigh a clean 50 mL erlenmeyer flask (it does not have to be dry). 5) Take an initial reading of the volume of water in your burette (measured to 2 places past the decimal), then let 10 mL of water run into the erlenmeyer flask. Take the final reading from the burette (measured to 2 places past the decimal). To double-check your reading technique, have your demonstrator also determine the final reading. Compare your reading to the demonstrators reading: they should agree within 0.01 mL. Determine the total volume of water added to the erlenmeyer flask (this is the difference between the initial reading and the final reading). 6) Weigh the erlenmeyer flask to determine the mass of the water delivered. 7) Empty the contents of the erlenmeyer flask back into your 250 mL beaker to be reused. 8) Refill the burette to near the 0 mL mark. 9) Repeat steps 4 through 8 two more times, each time draining 10 mL of water into the erlenmeyer. 10) Carry out steps 4 through 8 three times for each of the following volumes: 20 mL, 30 mL, 40 mL, and 50 mL. 11) For each mass of water delivered, determine the true volume of water corrected to 20oC using Table 28-3. Make a plot such as the one shown in problem 7 in the prelab questions. The y-axis should be the error and x-axis should be the apparent volume. The graph should be smooth, and your errors should not be significantly larger than about 0.10 mL.

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EXPERIMENT 2 ANALYSIS OF ASPIRIN

Suggested Reading: Exploring Chemical Analysis, 5th Ed. Daniel C. Harris, Chapter 10. Prelab Questions 1. Find the MSDS sheet for aspirin as described below. 2. Define primary and secondary standards. What are used in this experiment as the primary and secondary standards. 3. Why is it necessary to use carbonate-free NaOH. 4. Calculate the approximate volume of HCl you will use in the back titration step if the mass of ASA is 325 mg per tablet. (The procedure is as described in the experiment below using 0.10 M HCl and 0.10 M NaOH) 5. Find the appropriate equations that you will need for the statistical analysis section. Include descriptions of all variables, and under which circumstances you will use each equation. 6. Illustrate the titration of Potassium hydrogen phthalate with NaOH using an appropriate titration curve and indicate on this curve the correct choice of the indicator. 7. With reference to the acid strength and using titration curves, discuss why it is possible to use phenolphthalein as indicator during the back titration with HCl when acetate is also present. On a titration curve show the equivalence point of the OH- and the acetate titration and show where the phenolphthalein indicator will change colour. Materials Safety Data Sheets (MSDS). When using unfamiliar chemicals in the lab, it is important to know about the potential hazards of the chemicals and what precautions to take to minimize exposure to these chemicals. The MSDS are a valuable tool in finding out information about a particular chemical. MSDS for most chemicals are available on the internet. The quickest way to find a particular chemical is to find the Chemical Abstracts number (CAS #) before looking for the MSDS. This is because a MSDS may be filed under numerous chemical names, however each chemical has a single CAS number. A valuable web site to use to find a CAS number, (and many other useful sources of information on the chemical) is ChemFinder. At the website, http://chemfinder.camsoft.com/ you can search for compounds in a variety of ways. The search machine will return to you information about the chemical of interest, as well as several other links to sites with chemical information, some of which may contain MSDS. Once you have the CAS number, you can also use the Sigma-Aldrich Web

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server (http://www.sigma-aldrich.com/SAWS.nsf/msdshelp?OpenForm) to search for MSDS. The disadvantage of this site is that it requires the user to register first, although registration is free. Many other websites are available - just search for MSDS using a suitable search engine. Material safety data is also available in the reference section of the library. Before coming to class, find the MSDS sheet for aspirin.

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EXPERIMENT 2 ANALYSIS OF ASPIRIN

Objective
During the course of this experiment, you will determine the quantity of acetyl salicylic acid (ASA) in an aspirin headache tablet. To achieve this, you will dissolve a headache tablet with the aid of a NaOH solution, then analyze the resulting solution by titrating the excess NaOH with hydrochloric acid. Pharmaceutical manufacturers stipulate that the tablets will contain a certain mass of ASA per tablet. You will determine this, and calculate the concentration of ASA as a percentage weight (%w/w).

Theory
Aspirin tablets contain ASA (MW 180.15 g.mol-1) and fillers, used to bind the tablet and to dilute the drug when small quantities of drug are required. In this case, the fillers are cornstarch and SiO2. NaOH is standardized using a weak organic acid salt, potassium hydrogen phthalate: KH(C8H4O4) + NaOH " NaK(C8H4O4) + H2O The molecular weight of KH(C8H4O4) is 204.23 g/mol. This acid reacts with NaOH as shown above. Phenolphthalein indicator may be used for this titration since NaOH is a strong base, and at the end point the pH of the solution will be in the region of 8 - 9, the pH range where phenolphthalein indicator changes colour. This standardized solution of NaOH is then used to standardize the hydrochloric acid solution used. The reaction of NaOH with acetyl salicylic acid is as follows:

O O OH O CH3 OH

2 OHO

CH3CO2-

H2O

The acetyl salicylic acid is dissolved in an excess of NaOH, then the excess of NaOH is back titrated with a standardized solution of hydrochloric acid. Since the titration is one of a strong acid with a strong base, phenolphthalein may be used as the indicator. Only the excess NaOH is titrated by the HCl, as the acetate formed by the hydrolysis reaction is not titrated since phenolphthalein is used as the indicator and the end point of the titration of acetate with HCl occurs at a lower pH than the end point of the phenolphthalein.

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Sampling When samples are chosen for analysis, the actual result obtained depends on how similar the samples are to each other. Good samples should be as similar as possible. These samples are called representative samples the samples are representative of the material being analyzed (i.e. the concentration of the analyte in the sample accurately reflects the concentration of the analyte in the bulk). When the material is homogeneous and finely dived into a powder, it is fairly easy to take representative samples, but most materials are heterogeneous and the composed of pieces of different sizes. In this experiment you will be using two different sampling methods. One is to randomly select pieces of the material and analyze them, the second is to combine many pieces of the material, grind them down to a homogeneous powder and to analyze a portion of that powder.

Experimental The analysis of aspirin by back titration

Preparation of Carbonate Free NaOH Prepare a solution of sodium hydroxide as follows: 1. Boil 1.5 L of distilled water and allow to cool. 2. Roughly weigh 5 g of NaOH pellets (MW 39.997 g/mol) on a watch glass. Sodium Hydroxide is very caustic. Ensure that you clean up spills immediately. 3. Rinse off surface Na2CO3 with a small quantity of distilled water from a wash bottle. Discard the rinsings. Do not allow the pellets to sit in the liquid or you will dissolve too much of the NaOH and your solution will be too weak. 4. Dissolve the pellets by adding them to 5 mL boiled distilled water in a small beaker. This will give a 50% solution of NaOH in which Na2CO3, is practically insoluble. If some undissolved Na2CO3 remains, allow this to settle and carefully decant 4 to 5 mL of the liquid or remove it with a pipette into a little distilled water that has been freshly boiled and cooled. Avoid transferring any of the carbonate in this step. 5. Transfer the solution to a 1 litre plastic storage bottle, and add approximately 995 mL of the boiled water (enough to almost fill the bottle), stopper the bottle well and mix thoroughly by inversion. The solution will now be about 0.1M. 6. Always store you solution in a plastic bottle. Before storage, squeeze the bottle slightly as you put on the cap to expel as much air as possible. This will minimize the amount of CO2 that is absorbed into the solution. Standardization of NaOH using Potassium Hydrogen Phthalate 1. Rinse your burette with about 5 mL of the carbonate-free 0.1 M NaOH solution prepared above. Ensure that the burette tip is rinsed as well. Discard the solution. 2. Fill the burette with NaOH solution and remove any air bubbles from the tip or sides of the burette. Adjust the solution level to approximately 0 mL. 3. Remove the drop at the end of the burette by touching it to the side of a waste beaker. 4. Read the starting volume accurately. The burette should be read to a precision of 1/5 of the smallest division i.e. to a precision of 0.02 mL. 5. Set up a table in your lab notebook.
Mass KHphthalate /g Moles KHphthalate /mol Vol. NaOH /mL Conc. NaOH /M

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6. Using a weighing boat or weighing paper, weigh with precision a quantity of 0.7 to 0.9 g of predried potassium hydrogen phthalate (stored in the desiccator) to the nearest 0.1 mg and rinse into a erlenmeyer flask. 7. Calculate your rough titre (volume of NaOH that must be added to reach the equivalence point) based on the mass of the standard. 8. Add to each sample 50 to 75 mL of boiled distilled water. Stopper the flasks and swirl to dissolve the solid. 9. Add 3 drops of phenolphthalein 10. Add 75% of the calculated volume of the NaOH (from step 7) from the burette into the conical flask with rapid agitation. Rinse the sides of the flask with distilled water. 11. Add the NaOH dropwise, mixing well between each drop until a faint pink colour is seen which persists for 30 seconds. (This colour may fade after a few minutes owing to the uptake of carbon dioxide. If it doesnt fade within a few minutes, then you have over titrated your solution). If you have some difficulty seeing the pale pink colour of the indicator, a doubled quantity (6 drops) may be employed without serious error being introduced. Ensure that any drops on the end of the burette tip are added to the solution, and accurately read the final volume on the burette. 12. Repeat this standardization 2 more times using new samples of potassium hydrogen phthalate for each trial. 13. A blank titration must be carried out and any blank titre should be subtracted from all titrations. This is done by titrating 5 mL of 0.5 M sodium chloride with the NaOH solution. Treat the solution as above. (It will only take a drop or two of the NaOH solution if any at all.) 14. Calculate the molarity of the NaOH solution by taking the mean of your three trials. The deviation of these values from the average should not exceed 0.5% relative error if you are to proceed to the next part of the experiment. Preparation of 0.1 M Hydrochloric Acid 1. Add 8.6 mL of concentrated hydrochloric acid (11.6 M) to 500mL (use a measuring cylinder) of tap water in a glass storage bottle and make up to 1 litre by adding approximately 490mL water. Concentrated acid should always be added to water, never the other way around. 2. Mix well by inverting the sealed bottle several times. Standardization of 0.1 M Hydrochloric Acid 1. Rinse the 25 mL pipette 3 times with small portions of HCl. 2. Pipette 25 mL of the hydrochloric acid into an erlenmeyer flask. 3. Add 2 -3 drops of phenolphthalein to the solution in the erlenmeyer flask and titrate with standardized 0.1 M NaOH as described above. 4. Repeat the titration until 3 titrations differ by no more than 0.5%. 5. Calculate the concentration of the HCl. Preparation of Acetyl Salicylate Solution 1. Weigh with precision 3 aspirin tablets and place each tablet in a separate 100 mL volumetric flask. (The flasks must be clean and dry. Remember not to dry the flasks in an oven.)

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2. Label each flask clearly. 3. Add approximately 75 mL of 0.1 M NaOH to each flask and shake well until the tablet is dissolved. A small quantity of white residue may not dissolve. 4. Fill the flask to the 100 mL calibration mark with 0.1 M NaOH. 5. Shake well to mix contents. Back titration of the excess NaOH 1. 2. 3. 4. 5. 6. 7. 8. Empty the burette and wash well with tap water, then rinse with distilled water. Rinse the burette with HCl before filling. Use the same procedure as previously used with the NaOH. Pipette 25 mL of the acetyl salicylate solution into an erlenmeyer flask. If there is any residue on the bottom of the flask, be careful not to disturb it or suck any of it into the pipette. Add 2 - 3 drops of phenolphthalein and titrate with the hydrochloric acid until a faint pink colour persists. Repeat the titration 2 more times. Your results should agree within 0.5%. Calculate the initial quantity of NaOH (in 25 mL), the amount of NaOH in excess and determine the quantity of acetyl salicylic acid in each tablet. Calculate the ASA as a %wt.

Analysis of aspirin powder 1. A number of aspirin tablets have been crushed and ground into a homogenous mixture. You will be given 1 portion of this powder to analyze for the concentration of aspirin. 2. Precisely weigh an amount of aspirin powder roughly equal to the weight of an aspirin tablet. 3. Analyze the ASA as described above. 4. Calculate the amount of ASA as a %wt concentration. KEEP YOUR HCl SOLUTION FOR EXPERIMENT 2 Calculate the concentration of ASA (% wt) in 3 separate tablets using the data obtained above. This will give you data for each tablet, and along with the mass ASA per tablet, will allow you to calculate statistics for the tablet-to-tablet variation. Hand your data into the TA at the end of the lab. Solution concentrations (NaOH and HCl) should be as molarities, and ASA concentrations as %weight.

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Formal Report for Experiment 2

This is the only lab for which you will be required to complete a formal report. Formal reports should be typed and should contain all of the information requested below. Title Page: 1) Experiment Title 2) Your name 3) Student ID # 4) Date 5) Your TA Abstract (3 points): A brief (5 to 10 lines) description of the experiment, your results, and the general importance of these findings. Introduction (10 points): 1) A discussion of the background and motivation behind the experiment. Why is the experiment being done? Why might the results be interesting to the general public? 2) A general description of the method used to accomplish the goal of the experiment. What experimental method did you use to determine the concentration of ASA in aspirin? What are the theoretical concepts behind this method? (Note: You should only give a general discussion of the procedural methods here, not a vast amount of procedural detail. The point of the introduction is to give the reader a general idea of the methodology. If they want experimental details, they can read that in the experimental section of your report.) 3) Advantages and Disadvantages of the experimental method used. What problems were encountered that were inherent to the experimental technique and could cause inaccuracy in the results? Would you recommend this experimental method as an accurate method to use, or would it be desirable to find a different method for accomplishing this goal? Experimental (7 points): 1) A detailed description of the experimental procedure. You may refer to the lab manual rather than rewriting the procedure. However, if you deviated in any way from the procedure described in the manual, you should explain this here. 2) Any measurements that were made in the process of preparing (as opposed to analyzing) your samples. (e.g. masses of potassium hydrogen phthalate, aspirin tablets, aspirin powder). All measurements should include units and uncertainty (e.g. +/- 0.0002 g). You want your reader to

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be able to exactly reproduce your experiment using the same masses and volumes of reagents that you used. Results and Discussion (30 points): 1) Data: Any measurements that were made in the process of analyzing (as opposed to preparing) your samples. (e.g. volumes of titrant delivered to reach the equivalence points in each of your titrations). 2) Results: Values that are calculated from your data and from the group data. Concentration of potassium hydrogen phthalate Concentration of NaOH in standard Concentration of HCl in standard Concentration of ASA in tablets (both % wt and mg/tablet) show results of each of your individual trials as well as your mean and standard deviation. show the mean and standard dev from the group data Concentration of ASA in powder show results of each of your individual trials as well as your mean and standard deviation. show the mean and standard dev from the group data Results of all t-tests Shewhart Chart Results should include units and measures of uncertainty. (Note: You do not need to show calculations in this section, put please show them at the end of the report.... see below). All data and results should be included neatly in well-labeled tables! Tables should include captions so that it is easy for the reader to determine what information is in the table. Every column of data should have a clear heading. Include units and uncertainties in your data and results 3) All data and results should be explained and discussed with clear reference to the table in which those data or results may be found. In your discussion please be sure to explain and discuss each of the following: Your results for mg ASA/tablet, %wt ASA/tablet, %wt ASA in powder Comparison of your results to the group results. If your results are very different from the group results, explain why. (What were your sources of random error or systematic error?) The standard deviations in your results (e.g. are they too large, very small? If your standard deviations are large, what might be a reason for this?) The results of each of the t-tests: What do these results mean? What is the value in doing each of these t-tests? The Shewhart chart: Can any data points be eliminated based on your Shewhart chart analysis? Why is a Shewhart chart useful for an analytical chemist?

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If there is any question in your mind about the accuracy of some data, you should mention this and explain the source of your concern. (For example, insolubility of tablets.) Remember, you should assume that your reader is not familiar with your experiment and you need to explain everything to him or her. Conclusions (5 points): A short paragraph about your final finding - provide a summary of the objectives and outcome of the experiment. What can be concluded from you Shewhart Chart? Calculations (45 marks): The calculations may be typed or hand-written, but please make them neat and easy to follow. Each calculation should be clearly labeled with a heading so the TA knows what you are calculating. Any variables should be explained. A. Calculations From YOUR data: Show sample calculations for each of the following: 1) 2) 3) 4) 5) 6) Concentration of potassium hydrogen phthalate Concentration of NaOH Concentration of HCl Concentration of ASA in tablets (both % wt and mg/tablet) Concentration of ASA in powder One example to show how you calculated the mean concentration of ASA (mg/tablet) from your three trials of one tablet. 7) One example to show how you calculated the standard deviation in the concentration of ASA (mg/tablet) from your three trials of one tablet. A. Calculations from the GROUP data: The following calculations should be done on Microsoft Excel spreadsheet. Please print your completed spreadsheet, highlight the results so it easy for your TA to find them and attach it to your report. A sample spreadsheet is shown on the next page. (Formulas for these calculations can be found on p. 10-15 of the lab manual) 1) 2) 3) 4) 5) 6) 7) 8) Group mean for mg ASA/tablet Group mean for %wt ASA in tablet Group mean for %wt ASA in powder Standard Deviations in all 3 of the above values t-test between your mean mg/tablet and the manufacturers mg/tablet (325 mg/tablet) t-test between the group mean mg/tablet and the manufacturers mg/tablet (325 mg/tablet) t-test between the group mean %wt for powder and the group mean %wt for tablets A Shewhart chart (p. 15 of statistical analysis) comparing group data of mass ASA per tablet to the manufacturers mg/tablet (325 mg/tablet).

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In the calculations section of your report, please provide each of the formulas used for calculations 1-8 above exactly is it was input into your spreadsheet. Clearly label each formula and its location in the spreadsheet. For example, for the group mean mg/tablet reported in cell D11 in the spreadsheet below, the calculations section should include the following: Group mean for mg ASA/tablet (cell D11): AVERAGE(B11:B28) Sample Spreadsheet:
A 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 Data for Powder Mean 90.19 Standard Deviation 1.5 N 18 Individual Tablets (group) Sample No 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 Mg/tab 325.23 326.36 319.99 324.56 324.23 325.69 323.56 328.96 325.12 324.13 323.16 325.12 324.56 326.66 321.68 324.55 326.59 325.99 %wt Mean(mg) 90.12 90.15 90.25 91.23 90.26 90.56 Mean(%wt ) 90.78 90.51 90.98 90.46 90.89 90.26 90.19 90.88 90.5 90.55 90.23 90.36 90.55 t calc (comparison of 2 means) 0.036 t calc (comparison of mean with true value) 0.479 (group data) t calc (comparison of mean with true value) 1.039 (single analyst data) N 18 Standard Deviation 0.32 T tests 95 % t value 4.303 2.110 (N = 3) (N = 18) N 18 324.79 Standard Deviation 1.99 Action Line Lower 319.30 Upper 330.70 Individual Tablets (single analyst) SampleNo 1 2 3 B C D E F G H I Control Chart data mg/tab 325.23 326.36 319.99 N 3 Warning line Lower 321.20 Upper 328.80 Mean 323.86 Standard Deviation 3.40 Target Value 325 Standard Deviation 1.9

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EXPERIMENT 3: THE DETERMINATION OF SODIUM CARBONATE IN SODA ASH.

Suggested Reading: Exploring Chemical Analysis, 5th Ed. Daniel C. Harris, Chapter 10.

PRELAB QUESTIONS 1. Exactly 50.00 mL of an HCl solution required 24.75 mL of 0.02043M Ba(OH)2 to reach the end point. Calculate the molarity of the HCl. 2. pKa of H2CO3 and HCO3- are respectively 6.4 and 10.3. Make a rough sketch of the titration curve, which should be obtained upon titration of Na2CO3 with HCl. 3. Explain the effect of boiling the solution just before the end point is reached in a titration of carbonate. (Harris pp. 221) 4. Calculate the amount of 0.1000M HCl used to titrate 0.2124g Na2CO3. 5. Why is it necessary to titrate the solution until the colour changes from blue to green, and not from blue to yellow?

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EXPERIMENT 3: THE DETERMINATION OF SODIUM CARBONATE IN SODA ASH

Titrimetric methods of analysis are an accurate and relatively simple way of quantitatively determining acids and bases. Most reagents required are cheap and easily available in very pure forms. Titrating accurately requires some sophisticated techniques, and attention to detail in order to achieve the precision and accuracy required. In this laboratory you are titrating a weak base, sodium carbonate in soda ash, a mixture of sodium oxide and sodium carbonate. Soda ash is an inexpensive substance widely used as a base in industry. The basic constituent is carbonate which reacts thus: CO32- + H+ HCO3- + H+ H2CO3 HCO3H2CO3 CO2 + H2O Kb1 = 2.13 x l0-4 Kb2 = 6.60 x 10-8

Carbonate is thus a weak base, which, upon titration with acid, gives a titration curve with two points of inflection. At the second equivalence point, the system consists of NaCl and H2CO3 and the pH is determined by the latter substance. A 0.05 M solution of H2CO3 would have a pH of 3.8. It is evident that a sharp drop of pH does not occur at the end-point as in the titration of a strong base. The carbonic acid dissociates to form CO2 molecules, which are fairly volatile. If the solution is boiled for a short time just before the end-point is reached, the CO2 is lost immediately upon its formation and only the small quantity of untitrated NaHCO3 remains in solution along with NaCl. Calculating pH of solutions containing amphiprotic salts (salts that have both acidic and basic properties, such as NaHCO3) is quite complicated. However, we can make an approximation: If NaHA is the only species that contributes significantly to the pH of the solution (i.e. concentrations of other species are negligible), then the [H+] can be calculated as follows:

[H + ] =

K a 2 [ NaHA] + KW 1 + [ NaHA] / K a1

where Ka1, and Ka2 are the acid dissociation constants of H2A and NaHA. This formula can be used to calculate the pH at the second equivalence point in the titration of carbonate anion with HCl, but we wont need it for this lab. A sharp drop in pH can occur when passing through the end-point and the use of bromocresol green indicator makes an accurate titration possible. What is the pH range over which this indicator changes color? What colors are observed?

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Experimental Procedure A. Preparation and Standardization of 0.1 M Hydrochloric Acid 1. Add 8.6 mL of concentrated hydrochloric acid (11.6 M) to 500 mL (use a measuring cylinder) water in a storage bottle and make up to 1 litre by adding approximately 490 mL water. Concentrated acid should always be added to water, never the other way around. 2. Mix well by inverting the sealed bottle several times. 3. The acid is standardized against pure Na2CO3 (MW 105.989 g/mol). Dry the Na2CO3 for 1 h at 110-140oC. Place the open sample in a beaker and cover with a watch glass before placing the sample in the oven. Store the dry sample in a desiccator when it is not in use. The primary standard (Analar grade) is certified to have an assay of 99.9-100.1%. 4. Determine the indicator correction (blank analysis) by titrating 100 mL of 0.05 M NaCl. Prepare this blank sample by diluting 10 mL of 0.5 M NaCl solution with distilled water. 5. Rinse your calibrated burette with a little HCl, and allow a little HCl to pass through the stopcock and tip. Discard this HCl. 6. Fill the burette with HCl and take an initial reading to 2 decimal places. Dont forget to remove your funnel first. 7. Add 3 drops bromocresol green to the NaCl solution, and titrate with HCl until the solution changes to green. (This may only take a couple of drops, so be careful.) 8. Make a note of your readings in a table in your lab notebook similar to the one shown below. This table shows the minimum number of readings you should record in your lab note book, but you may want to add more for convenience eg Initial and final burette readings. DO NOT WRITE ANYTHING IN THIS LAB MANUAL.
Mass Na2CO3 /g Moles Na2CO3 /mol Vol. HCl /mL Conc. HCl /M

9. Accurately weigh (precision balance) a 0.20 to 0.25 g sample of sodium carbonate into an Erlenmeyer flask. 10.Dissolve in about 50 mL of distilled water. The volume of water added does not need to be accurately measured, since it will not affect the number of moles of base present. 11.Add 3 drops of bromocresol green. 12. Titrate with HCl until the solution changes from blue to green. (Calculate your approximate titre before you do this, so that you have a rough idea of how much HCl you will be adding.) A piece of white paper under the conical flask will allow for easier determination of the end point. If your solution goes yellow, you have over titrated. 13.Boil the solution gently for 2 to 3 min, until the solution turns blue again. If your solution remains green, you have over titrated. (Boiling and reacting solutions throw a fine spray into the air and losses may result. A beaker must be covered with a watch glass (concave side up) when the contents are reacting or boiling. When removing the watch glass, its surface should be rinsed into the beaker with the aid of a wash bottle. The watch glass should be placed, convex side up, on a clean surface if it is to be subsequently returned to its place on the beaker. The contents of an Erlenmeyer flask may be boiled briefly without danger of appreciable loss, but if a solution is to be boiled for several minutes, a powder funnel should be inserted, neck downwards, into the mouth of the flask. Solutions should not be allowed to boil to dryness, unless this is specifically called for, and then it can only be done with care to avoid cracking the glass. Solutions that boil dry sometimes fail

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to respond properly to the analytical procedure. If a dense liquid such a sulfuric acid is added to a solution to be heated, mix the solution well before heating. Failure to do so can result in the solution "bumping". If a solution is to be stirred, use a fire-polished glass rod and avoid knocking it against the walls of the beaker. Scratches may otherwise be formed, and scratches can trap solids, particularly precipitates. When removing the glass rod, it should be rinsed into the beaker with the aid of a wash bottle or you may loose some of your analyte. Never transfer a glass rod from one solution to another without rinsing it off first. Be careful to put glass rods on a clean surface if you intend using them to stir the solution again. ) 14.Cool to near room temperature and complete the titration to a green colour. 15.Take a final reading from the burette to 2 decimal places. Your titre should always be approximately 30 mL to avoid error. If it is much less than this, then you need to increase the mass of the sample you are titrating in subsequent steps. 16.Repeat steps 8 -14 for at least two other samples. 17.Calculate the concentration of the HCl solution in mol L-1. (Remember that 2H+ reacts with 1CO32-.) Results should agree to within a few ppt (parts per thousand). If results are not within a few ppt of each other, then perform another titration. (Be sure to apply burette corrections to your volume as determined in by the calibration experiment.) B. Titration of "unknown" samples with bromocresol green indicator. 1. Dry your unknown sample for 2 h at 110 - l 40oC. 2. Set up a table in your laboratory notebook for your data. A sample is given below:
Mass unknown /g Vol. HCl /mL Moles Na2CO3 /mol % Na2CO3

3. Use a high precession balance to accurately weigh three 0.4 - 0.6g portions of the soda ash sample into Erlenmeyer flasks. 4. Dissolve in about 50 mL of distilled water and add three drops of bromocresol green solution. 5. Titrate with standard HC1 until the appearance of a green colour. 6. Boil the sample for 2-3 min. 7. Cool to room temperature and continue the titration until the reappearance of the green color. 8. Calculate the percentage of Na2CO3 in the sample by taking the mean of at least three trials, and report your determination. If your results do not agree within a few ppt, then repeat the procedure for a fourth titration. Suitable places to stop this experiment: 1. After making the HCl solution. 2. After standardizing the HCl solution

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EXPERIMENT 4: THE DETERMINATION OF CHLORIDE BY VOLHARD'S METHOD

Suggested Reading: Exploring Chemical Analysis, 5th Ed. Daniel C. Harris, Chapter 6, pg.136-137.

PRELAB QUESTIONS 1. What is the effect of sunlight falling on the titration vessel? 2. In the Volhard method, proper functioning of the indicator requires that the sample solution be acidic. Why? 3. Which dissolved ions are present in significant amounts in the titrated solution: (a) during titration of the excess silver? (b) after adding an excess of KSCN? 4. Diethyl ether is effective at preventing fading of the end-point. Explain why fading takes place and why diethyl ether is effective at preventing it. 5. What would be the consequence of adding significantly more than 2 mL of the ferric ammonium sulfate indicator? 6. Determine the %(w/w) of Cl in a solid sample. The sample (0.3721g) was treated with 50.00 mL of 0.02051M AgNO3 after dissolving it in H2O. The excess Ag+ was titrated with 24.21 mL of 0.01997 M KSCN.

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EXPERIMENT 4: THE DETERMINATION OF CHLORIDE BY THE VOLHARD'S METHOD

Objective: Determine the amount of Cl- in a sample using Volhards Procedure. Volhard's procedure for the determination of chloride or bromide is an example of a precipitation titration. Many precipitation titrations make use of a standard silver nitrate solution to effect such reactions as the following: Ag+ (aq) + Cl- (aq) " AgCl (s) (1) 3 Ag+ (aq) + AsO43- (aq) " Ag3AsO4 (s) (2) In the present method, the chloride ion is quantitatively precipitated from acidic solution by adding an excess of silver nitrate solution (as in chemical equation 1 above). The nitrate anion is a spectator ion. The amount of excess Ag+ remaining after Cl- has been consumed is then determined by precipitating the remaining silver ion with potassium thiocyanate. The equation for which is: Ag+ (aq) + SCN- (aq) " AgSCN (s) (3)

Iron(III) ion serves as the indicator, imparting a colour to the solution with the first slight excess of thiocyanate: Fe3+ + SCN- " FeSCN2+ (red) Only a trace of this is to be formed at the end-point, which imparts a pale buff (beige/pale orange) colour to the solution. The use of a standardized solution of potassium thiocyanate enables one to calculate the number of moles (or millimoles) of silver ion precipitated as AgSCN and hence, by difference, the number of moles of silver ion initially precipitated as AgCl. This is, of course, also the number of moles of chloride in the portion of sample taken for titration. # moles KSCN added = # moles excess Ag+ # moles Cl- = (# moles Ag+ in AgNO3) (# moles excess Ag+) Experimental procedure 1. You are to obtain a sample of chloride-containing salt from an instructor. Immediately record the code number in your laboratory notebook. 2. Obtain also 1.7-1.8 g of AgNO3 (MW 169.873 g/mol) crystals - no more as this compound is very expensive ($1500 per 500 g!!). Protect this compound from light to prevent it from decomposing.

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A. Preparation of a standard 0.02 M Silver Nitrate solution 1. Use an electronic balance to weigh approximately 1.7 - 1.8g of primary standard grade silver nitrate into a clean, dry weighing bottle. Heat the bottle and its contents for 1 hr, but not longer, at 110oC. Ensure the oven temperature does not rise above 110oC. Store the cool silver nitrate in a desiccator. 2. Use a high precision balance to accurately weight 1.7g of the dry silver nitrate. 3. Carefully transfer the solid into a powder funnel held in the neck of a 500 mL volumetric flask. 4. Wash the crystals into the volumetric flask with distilled water. Remove the funnel, half-fill the flask with water and swirl until the crystals are completely dissolved. Dilute to the calibration mark and mix the solution thoroughly by inverting and shaking. 5. Transfer the solution into a clean brown storage bottle. Begin by rinsing the bottle with three small portions of the solution and discarding these; then transfer the bulk. When not actually in use, the silver nitrate solution should be stored in a dark place. B. Preparation and standardization of 0.02 M Potassium Thiocyanate solution 1. Prepare the solution by dissolving approximately 0.97 g of KSCN (MW 97.182 g/mol) (weigh accurately) in 500 mL of distilled water in a volumetric flask. Mix well. 2. Use your standardized 25 mL pipette to transfer an aliquot of standard silver nitrate solution into each of three 250 mL Erlenmeyer flasks and dilute to approximately 50 mL. 3. Add 2 mL of concentrated HNO31 and 2 mL of ferric alum (Ferric ammonium sulfate) indicator solution. 4. Titrate each aliquot with KSCN to a faint but definite, persistent pale buff colour2 of FeSCN2+ in the solution. If the solution is orange, you have over titrated. 5. Calculate the molarity of the KSCN solution. Results from the various titrations should show an agreement to within a few ppt (parts per thousand); if this precision has not been attained, check to see if your volumetric ware is perfectly clean and drains well, and then perform further standardization titrations. C. The Determination of chloride in an "unknown" sample 1. 2. 3. 4. 5. 6. 7. 8.
1 2

Dry the sample at 110oC in a weighing bottle for 1 hr. Transfer 0.35-0.4 g (weigh accurately) of the sample into a clean 250 mL volumetric flask. Dissolve the sample in distilled water and make up to the calibration mark. Mix well. Use your 25 mL pipette to transfer aliquots of the sample solution to three 250 mL Erlenmeyer flasks. Add 2 mL concentrated HNO3 and twice 25 mL (pipette) of standard silver nitrate solution to each aliquot. Also add 2 mL ferric alum solution and 5 mL of diethylether to each aliquot. While shaking vigorously, titrate the excess silver with standard KSCN solution until the color of FeSCN2+ persists for 1 minute or longer. The colour will fade slowly. Perform analysis on aliquots of sample until three consecutive titrations agree to within 0.5%.

If the nitric acid is yellow it should be boiled until it is colorless. Titrate always against a pure white surface, e.g. white paper. Bright daylight will cause precipitates of AgCl to become discolored (mauve)

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9. Calculate the mass of Cl- in each aliquot and use this to calculate the percent mass of Cl- in the unknown sample. (Dont forget to take into account the factor of 10 resulting from using only 25 mL of your 250 mL sample for the titration.) 10.Average your results (at least three trials) and calculate the standard deviation. It is customary to perform a "blank" titration with each analysis. The Volhard method, an example of a back titration method, does not make use of a blank. Explain why in your discussion. Suitable places to stop: 1) After preparation and standardization of AgNO3.

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Experiment 5: An investigation of interferences in the determination of Ca2+ in water by Flame Atomic Absorption Spectrophotometry
Suggested Reading: Exploring Chemical Analysis, 5th Ed. Daniel C. Harris, Chapter 20. Prelab Questions: 1. What type of flames do you know? Describe the differences between them as related to (a) temperature achieved in the flame (b) interferences. 2. What are the main sources of matrix interferences in FAAS. 3. Write the chemical reactions for (a) the interference of phosphates in FAAS determination of calcium in FAAS (b) the effect of EDTA as a releasing agent. 4. Can the standard addition method be used to minimize matrix effects?

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Experiment 5: An investigation of interferences in the determination of Ca2+ in water by Flame Atomic Absorption Spectrophotometry
Method: Flame Atomic Absorption Spectrophotometry (FAAS) Objective: Investigation of interferences and determination of Ca2+ in river water samples by FAAS General information Optical spectrometric methods are those methods where the intensity of absorption or emission of light by a species is measured at specific wavelengths as a means of determining chemical identity or concentrations of species. Atomic spectrometry is an effective tool for determining ppm to ppt levels of analytes by measuring absorption or emission of individual atoms. The first step in an atomic spectroscopy experiment is to break down the analyte into individual atoms (atomization). The atomized mixture is then irradiated at a very specific wavelength, which is ideally absorbed only by the atoms of interest. The amount of radiation absorbed is proportional to the number of atoms in the light pathway, which is, in turn, proportional to the concentration of the analyte in the sample. Thus, by measuring the atomic absorption of a sample, the concentration of the analyte in the sample can be determined. The process by which individual atoms absorb light is depicted below. An electron in the ground state of an individual atom is excited to a higher energy level by absorbing the incoming radiation. The wavelength of radiation that is absorbed is related to the energy difference between the ground state energy and excited state energy of the atom by the equation #E = !/$, where ! is Plancks constant. Because only very specific energy levels are available for the electrons of a given atom, only very specific wavelengths of light can be absorbed by the atom. Thus, atoms of different elements will have distinguishably different atomic absorption spectra.

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Excited State Atom

E1 "E = h /!

Incoming Radiation Intensity = I0 Wavelength = ! E0 Ground State Atom

Outgoing (unabsorbed) Radiation Intensity = It

It = Io exp(k * concentration) absorbance = ln (Io / It) = k * concentration k = constant

Measuring the concentration of an absorbing species in a sample is accomplished by applying the Beer-Lambert Law. A= abc where a- absorption coefficient b- path length c- concentration of absorbing species Applying the Beer-Lambert law directly in AA spectrometry is difficult due to variations in the atomization efficiency from the sample matrix, and nonuniformity of concentration and path length of analyte atoms (in graphite furnace AA). Concentration measurements are usually determined from a working curve after calibrating the instrument with standards of known concentration. The main parts of an AAS instruments are: Light source: In AAS the light source is usually a hollow-cathode lamp of the element that is being measured. Hollow-cathode lamps are a type of discharge lamp that produce a narrow emission spectrum from a particular atomic species. They get their name from the cup-shaped cathode, which is made from the element of interest. When a high voltage is applied between the anode and the

46

cathode, rare gas atoms are ionized and the positive ions are accelerated into the cathode resulting in metal atoms being sputtered into the gas phase. Collisions with gas atoms or electrons excite the metal atoms to higher energy levels, which decay to lower levels by emitting light at particular wavelengths characteristic of the metal. Atomizer: This is the part of the instrument where the atomization occurs. There are two major types of atomizers: Flame and Graphite Furnace. Monochromator: This is the system of prisms, diffraction gratings and mirrors, which focuses only the wavelength of interest on the exit slit. By placing the monochromator after the atomizer, only one wavelength from a hollow cathode lamp is selected and the selectivity is improved as most stray emissions are eliminated i.e. the absorption line is isolated from background light due to interferences. Photon Detectors: The final element of instrumentation is the detector used to measure the intensity of the incident radiation from the monochromator. The photo multiplier tube (PMT) is a photodynode encased in a high vacuum glass envelope. Impact of light photons onto the PMT causes electrons to be ejected from the PMT by photoelectric effect. The selection of the proper PMT is crucial to certain analyses as no one tube is efficient across the entire spectral range. Consult an analytical textbook for a schematic representation of the instrumentation of an absorption spectrometer. Most of the problems in AAS measurements are related to different types of interferences. There are two main types of interferences in AAS: Spectral Interferences and Chemical Interferences. Spectral Interferences occur when the absorption or emission of an interfering species either overlaps or lies very close to the analyte absorption wavelength. This species may be either molecules with very broad absorption bands, atoms of other elements (not very often) or products formed during other steps of measurements that scatter the radiation. Chemical interferences occur when a chemical reaction occurs in the solution or atomization region that affects the analysis. There are two types of chemical interferences: Dissociation Equilibrium and Ionization Interferences. Dissociation interference happens when the atoms of the analyte are bound in highly refractory (high boiling), or highly stable compounds (such as oxides) that cannot be dissociated in the atomizer. This interference, which is quite common in AAS measurements can be corrected by using matrix modifiers. Matrix modifiers can either release the analyte from the refractory compound or they can bind to the analyte, thereby allowing higher ashing temperatures while minimizing loss of the analyte. Ionization interference is characteristic of the alkali metals due to their low ionization potential. The thermal energy in the atomizer is sufficient to cause a significant degree of ionization of elements. This causes a reduction in the observed atomic absorbance since ions exhibit a different absorption spectrum than atoms. This interference can be minimized by using lower atomization temperatures (the sensitivity will be lower) or by using an excess of a salt (ion suppressor) that is more readily ionized than the analyte. In the latter method, the high concentration of electrons generated by the ion suppressor will make ionization of the other atoms less facile.

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Useful references Exploring Chemical Analysis, 5th Ed. Daniel C. Harris, Chapter 20, pg. 435-450. In FAAS the atomizer is a flame, which can provide temperatures up to 3100oC. The most common flame is air-acetylene flame with temperatures of 1700 to 2400oC. For more refractory compounds, the NO2- acetylene flame is used. This flame can produce temperatures of 2500 to 3100oC. In flame atomization, a solution of the sample is sprayed into a flame by means of a nebulizer, which converts the sample solution in a mist of tiny droplets. Combustion of the sample occurs in the flame to form a mixture of atoms, ions and molecules. This mixture is irradiated (hollow-cathode lamp) at a wavelength absorbed by the atom being analyzed and the absorbance is measured. Flame AA uses a slot type burner to increase the path length, and therefore to increase the total absorbance. In this experiment you will study the interferences involved in the FAAS determination of Ca2+ in river water. In natural waters Ca2+ ions are very often found in the form of phosphates and sulfates, which are very stable in the flame temperatures. They either cannot dissociate or the product of their dissociation is CaO, which cannot be atomized in the air-acetylene flame. This dissociation interference can be avoided by adding a releasing agent such as EDTA, which forms a calciumEDTA complex that can be readily dissociated. When the EDTA binds to the phosphate or sulfate, Ca2+ is released to form less refractory compounds. An alternative to using releasing agents is to use higher temperature flames where all of the calcium compounds can be dissociated and atomized. Experimental Instrument: A Varian AA 240 instrument will be used with air-acetylene flame as the atomizer. Solutions: Stock Solutions (to prepare) Calcium solution: 100 ml of 1000 ppm Ca2+ solution starting from Ca(NO3)24H2O reagent Phosphate solution (for interference study) 100 ml of 0.1 M NaH2PO4 solution, starting from NaH2PO4H2O reagent Releasing Agent: 100 ml of 0.2 M EDTA solution

Working solutions Week 1 1. Blank: Distilled deionized water (DDW). 2. Interference of phosphates: In six 100 ml volumetric flasks add 1 ml of Ca2+ stock solution and 0.0, 0.2, 0.4, 0.6, 0.8 and 1 ml of H2PO4- stock solution. Dilute with DDW to the 100 ml mark.

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3. Correction of interference In six 100 ml volumetric flasks add 1 ml of Ca2+solution, 0.6 ml of H2PO4- stock solution and 0, 1, 2, 3, 4, and 5 ml EDTA stock solution. Dilute with DDW to the 100 ml mark. Week 2 4. Determination of calcium in river water: (i) External Calibration: a. Prepare a series of calibration solutions of Ca2+ with concentrations 0, 2, 4, 6, 8, 10 ppm. Add 5 ml EDTA stock solution and dilute to the 100 ml mark. b. Put 50 ml sample solution (unknown from your TA) in each of two 100 ml volumetric flassk. Add 5 ml EDTA stock solution in each one of them and dilute to the 100 ml mark. Upon analysis, measurement you will decide if the sample needs any further dilution in order to keep the absorbance within the calibration range (e.g. 10 ml of sample + 5 ml EDTA diluted to 100 ml ), which should be below the maximum concentration used for calibration (10 ppm), yet not diluted too low. (ii) By Standard Addition: c. To each of five 100ml volumetric flasks add 50 ml (or the volume used in step b above with appropriate dilution) of sample, and a specific (but different) concentration of standard. Dilute to the 100 ml mark. (Note: You have already determined in step b above, the volume of sample that is necessary to be within the calibration range, i.e. below 10 ppm. The series of added concentrations in the final solution can be 0, 2, 4, 6, 8 ppm, thus the total concentration can be as high as 18 ppm). d. To each of five 100ml volumetric flasks add 50 ml (or the appropriate volume used in step c above) of sample, and a specific (but different) concentration of standard (same series of concentrations as above). Add 5ml EDTA solution and dilute to the 100 ml mark. Method You will be using two different methods set up on the instruments for the two weeks, based on the method templates; 4116 Ca method week1, and 4116 Ca method week2. The only difference between the two is that the number of calibration standards is different. Your TA will assist you in running these samples using the Flame Atomic Absorption Spectrometer Week 1 Go to the analyze tab, click on start

49

Aspirate the blank( deionized water only) and set the instrument for zero absorbance. Measure the blank and the calibration standard (use the solution which you prepared using 1ml of Ca and 0 ml of phosphate diluted to 100 ml, as the standard). Aspirate and measure the absorbance of the solutions containing Ca2+ with increasing amounts of phosphate starting with the one with no phosphate. Measure the samples containing increasing amounts of La3+ starting with the one with no lanthanum at all. Computer will store all the measured absorbance and concentration values in the worksheet file. For exporting the data for post processing using EXCEL, go to options, and reports , and save your data as a *.PRN file which can be accessed by EXCEL. Turn off the instrument and close the gas valves. GRAPHS: Draw the following graphs and comment on your results. ! Abs. vs. [H2PO4-] for solutions containing the same concentration of calcium ! Abs. vs. [EDTA] for solutions containing the same calcium and phosphate concentrations: Week 2 Open a new worksheet from the template 4116 Ca method week2 and click on the tab develop, and Edit method... Go through each tab and make sure the parameters selected agree with those on the table at the end. Save your worksheet for week2 under a unique name. Set the instrument to Zero by aspirating the blank. Go to analyze and click start Measure the calibration standards starting from the most diluted one. Repeat this at least three times for each sample. Remix the solution between measurements. Go to sequence and edit the list of samples to include six blank measurements (for estimating limit of detection), followed by the sample solutions. Measure the two series of standard addition samples. Plot the calibration curve for calcium determination, using the different methods of calibration. Calculate the concentration of the sample in the given sample in ppm by the external calibration and the two series of internal calibrations (standard addition) methods. Compare the results obtained for the samples with and without addition of EDTA salt. Report Questions: 5. Comments on the sensitivity of the two calibration modes. 6. Comment on the linearity of the calibration plots. How would you minimize errors resulting from the non-linearity of the calibration.

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Experiment 6. Determination of iron in multi-vitamin tablets by colorimetry and verification of their consistency between two production batches.
Suggested Reading: Exploring Chemical Analysis, 5th Ed. Daniel C. Harris, Chapters 4, 18, 19. PRELAB QUESTIONS 1. What is the function of ascorbic acid in this experiment? 2. Briefly outline how would you use this method to determine relative concentration of different species of Fe (i.e. Fe(ii) and Fe(iii) oxidation states in the same solution ), for example in lake water to monitor red-ox environments? 3. What is a t-test and how will you use it to analyze the concentration of iron in your samples?

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! Experiment 6: Determination of iron in multi-vitamin tablets by colorimetry and verification of their consistency between two production batches Objectives
Investigation of the suitability of acid extraction of iron from multi-vitamin tablets followed by colorimetry of Fe(II)-phenanthroline complex for its determination. Application of the method for statistical verification of any significant difference between two batches of tablets. General Information Iron deficiency is considered to be one of the most widespread health problems in the modern world affecting a large proportion of the worlds population (UNICEF/WHO; http://www.micronutrient.org/english/view.asp?x=579). Its most severe form, iron deficiency anaemia, is reported to have a higher overall cost to society than any other disease except tuberculosis. Improving the intake of iro n through fortified food and supplements are among the most effective means of reducing the prevalence of iron deficiency. Measurement of the content of iron in such materials is important. Although sophisticated and highly sensitive analytical techniques can be used for accurate determination of iron, such techniques can be too expensive for many small laboratories, particularly for those in developing countries where the alternatives, such as simple colorimetric techniques, can be quite beneficial. In the manufacturing environment of pharmaceutical products, verification of consistency from one batch to the other is an important aspect of the quality control process. 1,10-Phenanthroline is a heterocyclic bidentate ligand. With ferrous ions in solution it forms the complex [Fe(phen)3]2+ which can be used for the photometric determination of Fe(II). The complex, called "ferroin" has a deep red colour. 1,10-Phenanthroline

Dissolution of iron present in a tablet and reaction of Fe with phenanthroline followed by UV-VIS Spectrophotometry at an appropriate wavelength(s), can be used for its quantification. According to Beer-Lambert Law, the absorbance, A is linearly related to the concentration. A=! bc Where ! is the absorptivity (at unit concentration), b is the path length and c is the concentration.

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Statistical verification of the difference between two batches (Comparison of two experimental means) For verification of consistency, you will be making repeat analysis of tablets from two different batches applying a t-test. In statistical terms, the significance of the difference between two experimental means is verified by testing the null hypothesis (significance testing). Assume that we have two sample means, x1 and x2 with the standard deviations s1 and s2, respectively. We can calculate the pooled standard deviation, s using the following formula.

(n1 ! 1) s1 + (n2 ! 1) s 2 (n1 + n2 ! 2)

Where, n1 and n2 are the number of measurements (tablets) used to calculate x1 and x2 , respectively. The number of degrees of freedom would be (n1 + n2 -2). In order to decide whether the difference between two sample means x1 and x2 is significant, that is to test the null hypothesis, H0: 1 = 2, the statistic t can be calculated using the formula,

= s

( x1 ! x2 ) 1 1 + n1 n2

If the calculated value of t is greater than the critical t value from the table (you may refer to the attached table) corresponding to a chosen level of confidence and the degrees of freedom, null hypothesis is rejected (i.e. the difference is considered significant at that confidence level).

Experimental (Caution: Handling of concentrated acid should be done in the fume hood with proper safety precautions). Week 1: Determination of optimum wavelength for absorbance of phenanthroline Fe complex. 1. Prepare the 250 ml of calibration standard and 250 ml of blank solution according to Table 1. 2. Set the wavelength at 400 nm and zero instrument with sample compartment empty (left most dial). Insert cell containing blank solution, and set % Transmission to 100 %. 3. Measure absorbance / transmittance of standard in 20 nm increments over range of 400 to 600 nm. 4. You must re-zero instrument and rinse with a small volume of solution to be measured every time you change the wavelength. 5. Determine the wavelength in this range which resulted in the maximum absorption. 6. From this wavelength, measure absorbance at +/- 15 nm in 5 nm increments to fine-tune the maximum wavelength

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7. Set the wavelength to that of maximum absorbance; you will conduct all Week II experiments at this wavelength. Table 1 - Preparation of calibration standard for determination of optimum wavelength Calib. Standard 1000 ppm 10% Fe Stock Ascorbic solution acid (ml) (ml) 0.000 0.75 12.50 12.50 5M acetic /sodium acetate buffer (ml) 75.00 75.00 0.1% phenanthroline (ml) 75.00 75.00 Total Volume (ml) 250 250

Blank Standard

Week II: Quantitative analysis of Fe in vitamin tablets Part I: Iron Tablets Extraction/dissolution of Fe from the tablets is done by dissolving tablets in acid (a mixture of concentrated HNO3 and HCl) . 1. Clean plastic tubes (50 ml), small plastic vials and the caps with detergent, then with 1% HCl, and finally with distilled water (1 per student + 1 blank per group) 2. Obtain multivitamin tablets (1 per student) 3. Label and weigh the 50 ml tubes (first empty, then with one tablet). 4. Add 2 ml of conc. HNO3 and 4 ml of conc. HCl to each tube including the blanks. Note time of addition. 5. After 1 hour (actual time noted) carefully swirl the content and add approximately 20-30 ml of distilled water (in the fume hood). 6. Outside the fume hood, add accurately (on the weighing scale) distilled water until the total amount of liquid (including acid) that has been added amounts to 50.0g (Note: you already know the mass of empty tube and the tablet) 7. Close the tube with the screw cap, mix the solution gently and let it stand for 15 min for the un-dissolved material to settle down to the bottom. 8. Accurately transfer 300 micro-litres (using a micro piptette) from the top portion of each solution (try to avoid withdrawing particulate matter) into a 25 ml volumetric flask. 9. Accurately add the reagents (distilled water, ascorbic acid, acetate buffer, phenanthroline solution) according to the amounts shown in Table 2, and gently mix them. Make sure that the reagents do not get contaminated with the sample solutions through pipette tips. 10. After preparation of solutions of tablets and the blanks measure the absorbance at the wavelength determined week 1. Rinse the sample cuvette with 1%HCl in between samples. Sample (ml) 0.300 Table 2: Preparation of iron tablet samples 10% 5M 0.1% Ascorbic acetic/sodium phenanthroline acid (ml) acetate buffer (ml) (ml) 1.3 7.5 7.5 Total Volume (ml) 25.00

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Part II: Calibration Curve (to be completed while tablets are digesting). 1. 2. 3. 4. Prepare calibration standards according to Table 3. Set the wavelength to that of maximum absorbance as determined Week 1. Measure absorbance of calibration standards Plot calibration curve and determine line of best fit. Calib. Standard Table 3 - Preparation of calibration standards 1000 10% 5M acetic 0.1% Total ppm Fe Ascorbic /sodium phenan- Volume Stock acid (ml) acetate throline (ml) solution buffer (ml) (ml) (ml) 0.000 1.3 7.5 7.5 25.00 0.030 1.3 7.5 7.5 25.00 0.060 1.3 7.5 7.5 25.00 0.090 1.3 7.5 7.5 25.00 0.120 1.3 7.5 7.5 25.00 0.150 1.3 7.5 7.5 25.00 Final concn (ppm) ? ? ? ? ? ?

Blank Standard1 Standard2 Standard3 Standard4 Standard5 Questions:

1. Prepare the calibration curve and comment on the linearity. Calculate the concentrations using linear and a quadratic fit to the data. 2. Calculate the amount of Fe in each tablet, average and standard deviation in each group. Compare the measured level with the limit of detection. Is the method that you developed sensitive enough for the purpose? 3. Do you find a significant difference between the two batches at 90%, 95% and 99% confidence levels? 4. If you were to repeat the experiment, name one step you would take to improve the confidence of your conclusion about the consistency between two batches.

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Locker check out Please circle your course number. (2353, 2753, 2354, 2754) Student (1)____________________________________________ Student (2)____________________________________________ Locker # _____________________________________________ Demonstrator _________________________________________
DRAWER WIRE GAUGE CLAMP HOLDER SUPPORT RING RUBBER TUBING BUNSEN BURNER CRUCIBLE TONGS IN 2 2 2 2 2 2 WATCH GLASS LG. WATCH GLASS SM. WASH BOTTLE 4 6 2 OUT FILTER FUNNEL POWDER FUNNEL SINTERED GLASS FUNNEL IN 2 4 2 OUT PIPET BULB 25 ML VOLUMETRIC PIPET 1ML VOLUMETRIC PIPET 5 ML GRADUATED PIPET 5 ML GLASS ROD WITH POLICEMAN SPATULA / SCOOPULA T- JOINT SUCTION ADAPTER IN 2 2 1 1 2 2/ea 2 1 OUT

BOTTOM SHELF SUCTION FLASK 500 ML. ERLENMEYER FLASK 50 ML ERLENMEYER FLASK 250 ML ERLENMEYER FLASK 500 ML VOLUMETRIC PIPET 10 ML VOLUMETRIC PIPET 25 ML VOLUMETRIC FLASK 100 ML VOLUMETRIC FLASK 250 ML VOLUMETRIC FLASK 500 ML DESICCATOR

IN 2 2 8 4 2 2 8 4 2 1

OUT

TOP SHELF

IN

OUT

UNIVERSAL CLAMP BOTTLE 1000 ML BOTTLE 1000 ML NALGEN GRADUATED CYLINDER 10 ML GRADUATED CYLINDER 50 ML GRADUATED CYLINDER 100 ML BEAKER 50 ML BEAKER 150 ML BEAKER 250 ML BEAKER 400 ML BEAKER 600 ML BEAKER 1000 ML

2 8 4 2 2 2 8 2 4 6 2 2

Note the two 50 ml burets in the door be careful closing door. You must show your locker to the demonstrator before leaving.