Method Development for Analysis of Aspirin Tablets Kenneth W. Sheet, Jr. Kent State University, Kent, OH 44242 The experiments often performed in undergraduate analytical courses consist of analyzing a homogeneous, readily soluble powder according to cookbook directions. In a real industrial method development situation, the analytical protocol iroften far more c&nplev and may include interference studies, optimizing ofconditions, etc. A large number of universities &e now usine the latter Dart of ouantitative analysis for introductory instrumental analysis, which is an ideal place to introduce the development of an analytical method. We have developed a lab experiment that satisfies this need with simple instrumentation and straightforward chemical reactions:~eplan to use the experiment a t the end of the auantitative analvsis course next year. A large number of methods for the determination of aspirin in tablets have been proposed. Titrimetric methods are fast and inexpensive hut are less suited for the demonstration of interference, condition optimization, etc. (1-4). Direct UV absorption methods (5) are subject to interference from other drugs, e.g., caffeine or acetaminophen and are used to demonstrate multicomponent analysis. Many of the interferences can be eliminated by the spectrofluorometric analyses (6, 7), which require a more sophisticated instrument and in many cases require organic solvent matrices. ic The visible s ~ e c t r o ~ h o t o m e t rassav (8.9)used in this experiment can be performed by using commonly available calorimeters without the need for the elaborate separations procedures required for the direct UV analysis (10).The colorimetric tablet analysis also avoids the reported queasiness of some students when dealing with biological specimens such as blood and urine (11).
Standard Preparation for Callbration Curve Salicylic Acid 1mLI
Modified Trinder Water lmLl
Reaaent fmLl
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Experlmerdal Deldls For the sake of hrevitv. .. the introduction to the e x ~ e r i ment and discussion questions have been omitted.' In order to standardize the results, the ferric-chromophore-forming reagent was provided to the studenw in an oxford pipettor. The small volumes used in the interference studies and hydrolysis study were measured using Eppendorf pipets. (These devices allow introduction of commonly employed volumetric dispensing devices shortly after a semesctr's usage of the traditional analytiral \,olumetric equipment, e.g., $pets, burets.) Groups of two (or three) students are required for completion of the experiment in a single threehour laboratory period. A number of steps in the procedure are intentionally vague and require a judgement by the students prior t o continuing with subsequent parts of the experiment. Calibration Curve Preparation (1) Weigh 300.0 mg of salicyclicacid into a 250-mL volumetric flask
and add 10 mL of NaOH solution (0.25 M). Once dissolved, dilute to volume with water. (2) Prepare (by 10-mL buret) the solutions given in the table in 25x 150-mm test tubes (requires three burets). Once the Trinder reagent has been added, the solutions are stable for at least onehalf hour. (The modified Trinder's reagent is prepared by dissolving 4.0 g Fe(NOJ3 . 9H10 in 12 mL of 12 M HCI then diluting to 1.0 L.)
'
Coples of the complete experiment are available fromthe author, including photocopies of the pertinent pages from the USP-NF.
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Journal of Chemical Education
(3) Measure the absorbance (at 540 nm) in l-cm matched cells using a Soectronic 20 (or similar soectroohotometer). . Preoare . a calibration curve of ahsurhance vs. salicylnte conrrntratim for subsrqucnt parts ro theexperiment. Interference Studies (1) Weigh 150 mg of each of the following ingredients into separate 50-mL beakers (or test tubes): aspirin; salicylamide (no longer an official drug because of its questionable efficacy); caffeine (often included to keep one from becoming drowsy while on medication), and acetaminophen (active ingredient in Tylenol and many other non-aspirin-containingpharmaceuticals). (2) Dissolve each compound in 5 mL methanol (caffeine may be only partially soluble if the citrate is used). (3) Add 5 mL of 0.25 M NaOH to the salicylamide, caffeine, and acetaminophen standards. Mix, and let stand for 5 min. (4) To the aspirin standard add 5 mL ofwater, mix, and immediately transfer 100 pL to a test tube. Quickly add 10 mL of Trinder reagent, and record the absorbance by using the spectrophotometer. (5) Take 100r L of standard solutions from step 3 above, add 10mL of Trinder reagent, and measure the absorbance. Hydrolysis of Aspirin to Salicylic Acid Read all steps carefully before proceeding, as this experiment is over in several minutes. (1) Weigh exactly 300 mg of aspirin into a 50-mL beaker, and dissolve in 10 mL methanol. (2) Add 10 mL of 0.25 M NaOH, and start a timer. This solution should be mixed vigorously with a magnetic stirrer. (3) After 30 s withdraw 250 r L of this standard, add 10 mL Trinder reagent, and mix thoroughly. Note the time when the Trinder reagent is added, as.the reaction will continue until that time. Record the lapsed time when the Trinder reagent is added and not when the sample is withdrawn. (4) Continue to withdraw 250-pL samples at 1-min intervals for 10 min. Treat the samples as in step 3. (5) Once all samples have been collected and treated, measure the sbsorbances as quickly as possible. (6) Plot the amount of salicylate formed 7s. reaction time on graph paper prior to tablet analysis. Tablet Assay for Aspirin Content (1) Weigh an aspirin tablet into a 50-mL beaker. (Be sure to record
brand, aspirin content, other active ingredients, lot number, etc.) (2) Add 10 mL of methanol, and crush the tablet with a stirring rod (a few insoluble excipients will remain). Add 10 mL of 0.25 M NaOH, and let stand until the aspirin has been completely
3
6
I
0
TIME lminl Wdrolysis of aspirin to salicylb acid: (a)wilh 0.5 M NaOH. (b) wilh 0.25 M NaOH. and (c) with 0.20 M NaOH. All experimental data points belong with cuwe a.
hydrolyzed to salicylic acid. This time is determined from the plot (step 6) of the hydrolysis experiment. (3) Quantitativelytransfer the tablet solution toa 250-mLvolumetrie flask,and dilute to volume with water. Filter 50 mL of the solution through coarse filter paper. (4) Take several (at least three) 2.5-mL aliquot8 of the filtrate into test tuhes, add 10 mL Trinder reagent, and read the absorbance. ( 5 ) Calculate the mass of aspirin in the tablet and percentage of aspirin in the tablet. Discussion The calibration curves prepared by most students for this experiment were in the form of absorhance vs. volume of salicvlic acid standard. A few d o t t e d absorbance vs. molar conc&tration. The average caiihrat ion curve for the class ( 5 mouos of Dartnerd oroduced a molar absorptivity of 305 LM'I ~ m a-t 540 ~ n& The interference studies produce a large number of ohservations that are useful for other parts o f t h e method development. Acetaminophen, caffeine, and salicylate-free aspirin do not react with the ferric reagent and therefore do not interfere. The aspirin result is useful in establishing the need for a hvdrolvsis sten. Furthermore. this data ~ o i nconfirms t "~~ t h a t no salicylate & present at time zero in i h e hydrolysis study. I t should be apparent that salicylic acid, if present, in the aspirin tablets would also interfere. Although salicylamide is not anofficial drug, it has beenused in multi-ingredient pharmaceuticals. Salicylamide undergoes the same chromo~hore-formingreaction as salierlate (12) and, if Present in pharmaceut~al,will interfere with the assay. Obtaining data during the hydrolysis reaction requires a firm understanding of the procedure prior to the start because the reaction will be finished within the first few mint utes. The rate a t which hvdrolvsis occurs is d e ~ e n d e nuDon theconcentration of addid N ~ O Has shown inihe figure..~n initial concentration of 0.5 Xl causes the reaction to proceed too rapidly to obtain adequate absorhance measurements as indicated bv the scatter in the date (all of which helong to curve a of the figure). Immediate supervision of the students is required during the hydrolysis portion of the experiment. . . An optional experiment(two shorter lah periods or a longer lab period) consists of having the students establish the curve for 0.20 M NaOH. which illustrates the need for a
~.
a
prolonged hydrolysis during tablet analysis at lower hydroxide concentrations. Curve b indicates that the minimum hydrolysis time necessary for the conversion of aspirin to a chromophore-forming species. This data is not given in the assay step of the procedure; therefore, the student must complete the hydrolysis graph and consult with the instructor regarding its interpretation. Only one brand of aspirin tablets was assayed (Bayer, adult strength), and each student was provided a single tablet from a fresh bottle. The results of the analysis ranged from 320 to 330 me a s ~ i r i nner tablet (325 me accordine to manufacturer labefi indicating that thetabletsmet USP:NF s~ecifications,i.e., =SOc of labeled asnirin content. T v ~ i c a l ranges reported fo; assay of the same tablet were +4 m i from the value the students reported. The r e ~ o r t e da s ~ i r i ncontent was 80% for all tablets. The same bittle was iater analyzed spectrofluorimetrically (7)by the same class with similar results. The USP-NF gives specifications for salicylic acid contamination with resect to aspirin content for tablets (0.75% for tablets and 0.3% for cansulesL I t should be ohvious to the students that this method does not distinguish between a s ~ i r i nand salicvlic acid in the orieinaltablet: however, from'the fluoresceht assay of the &me tablet's, the salicylic acid content was typically determined to be less than 0.1% of the aspirin content. Conclusion The simulated method development DroDosed in this experiment is safe and contains inexpensi;,e, kasily disposable reagents. Results ohtained by the method are both fast and accurate. The experiment demonstrates the need to (1) convert the analyte (aspirin) into a compound suitable for reaction with ferric reagent (salicylicacid), (2) optimize reaction time (and concentration of hydrdysis reagent-optional!, and (3) evaluate potential interferences. This experiment complements a previous experiment where the students determine the metal-to-lieand ratio and the value of the formation constant for the fekc-salicylate complex (13). Acknowledgment The author wishes to compliment the freshman honors chemistry students, Sheryl Tucker, Charlene Keane, and Rebecca Robinson, who did an exceptional job in piloting this experiment, which established the majority of the concentrations, etc., necessary to make this an effective experiment. Llbrature Cited 1. Bmun,R.D.J. Chom.Edue. 1985.62,811-812. 2 Shen, S. Y.; Cllman, A. J J.Ckem Educ. lW.42.540. 3. Prwtor. J . S.:Roberts, J. E. J Chom. Edur. 1961.38, nl. 4. Kolthoff, I. M.: Stenger, V. A. Volumetric Analysis. 2nd ed.;Interacienee:New York. 194l; Voi. 2, p 233. 5. Jones.M.;Thatcher,R.L.AnoLChom. 1951,23,957-9M) 6. Fiigen. R. A,: Plude. J. L.:Seih. W. R.J. Chsm. Educ. 1379.56.658. 7. St7eet.K. W.;SEhenk.G.H. J.Phorm.Sci. 1981,70,641446. 8. Tim. N. W. Fundamentals of Clinieol Chemistv; Saundors: Philadelphia, 101(1;p 882.
9. Sunshine, I. Methodology for Anolyticol Toricology; CRC: Clewland. OH, 1975: pp
1. Chcm. Edue. 1913,50,21P215.
Volume 65 Number 10 October 1988
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