2010) 1( 23مجلة ابن الھیثم للعلوم الصرفة والتطبیقیة المجلد طریقة مباشرة وطریقة غیر مباشرة لتعیین البراسیتامول في مستحضرات هیدروكسي بنزوات الصودیوم-2- امینو-باراصیدالنیة باالزدواج التأكسدي مع ليخالـد فهـد عـ، سرمد بهجت دیكران ،اسماء هاشم شاكر* جامعة تكریت، كلیة التربیة للبنات، قسم الكیمیاء* جامعة بغداد، ابن الهیثم-كلیة التربیة، قسم الكیمیاء الخالصة تعتمد الطریقة على اقترانه مع . ن لتقدیر الباراسیتامول في الوسط المائيان وحساستان سهلتان طیفیتاطریقت تطور بوجود العامل المؤكسد بیریودات الصودیوم في الوسط ) AHB(الصودیوم هیدروكسي بنزوات-2- امینو-كاشف بارا λmax =470لون بني برتقالي ذائب في الماء التي یقاس امتصاصها عند الطول الموجي يذ مركبالحامضي لتكوین مكغم 0.0449، وداللة ساندل تساوي 1-سم.1-مول.لتر 3371وقد بلغت قیمة معامل االمتصاص الموالري . نانومیتر مكغم من الباراسیتامول في حجم 500.0 – 12.5واتبعت الطریقة قانون بیر في مدى من التراكیز یتراوح بین 2-سم. مل.مكغم 20.0 – 0.5مل اي 25محلول نهائي -1 بینما ، % 4.70- 0.26وتراوح االنحراف القیاسي النسبي للطریقة بین مستویات من تراكیز الباراسیتامول ةولخمس% 106 - 95المئویة بین بلغت دقتها المعبر عنها بنسبة االسترجاع أما الطریقة الثانیة فتعتمد على تقدیرالباراسیتامول بصورة غیر مباشرة من خالل إجراء تحلل مائي حامضي را (للباراسیتامول ثم مفاعلة ناتج التحلل ) AHB(ت الصودیوم هیدركسي بنزوا-2- امینو-مع كاشف بارا) امینوفینول–البا إذ یتكون مركب ذو لون بنفسجي مزرق ذائب في الماء یقاس أقصى ، بوجود بیریودات الصودیوم في الوسط القاعدي وداللة 1-سم.1-مول.لتر 11884نانومیتر وبامتصاصیة موالریة مقدارها λmax=580امتصاص لها عند طول موجي سم.مكغم 0.0127ساندل -2 مكغم من الباراسیتامول في حجم نهائي 500.0–12.5قانون بیر بین وكانت حدود تطبیق . ي % 101.60-94.28وبمعدل نسبة اسـترجاع مئویة تراوح بین 1-مل.مكغم 20.0 -0.5مل أي 25 وانحراف قیاسـ ن بنجاح على بعض المستحضرات الصیدالنیة الحاویة على الباراسیتامول تاطبقت الطریق%. 1.1- 0.6نسـبي .مستویات من التراكیز ةالطریقین المباشرة واإلضافة القیاسیة ولثالثوب IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 Direct and Indirect Spectrophotometric Determination of Paracetamol in Pharmaceutical Preparations By Oxidative Coupling With p-Amino-2-hydroxy Sodium Benzoate *A. H. Shakir, S.B. Dikran, K. F. Ali *Department of Chemistry, College of Education for Women, Tikrit University Department of Chemistry, College of Education .Ibn-Al-Haitham, University of Baghdad Abstract New simple and sensitive spectrophotometric methods for the determination of paracetamol in aqueous medium were developed. The first method is based on coupling of paracetamol with p-amino-2-hydroxy sodium benzoate (AHB) in the presence of sodium periodate, as oxidizing agent, to form a brownish-orange compound which shows a λmax at 470 nm. The molar absorptivity (εmax) of the colored product was found to be (3371) l.mole- 1 .cm -1 and Sandel’s index 0.0449 μg.cm -2 . The method follows Beer’s law in the concentration range of 12.5-500.0 μg of paracetamol in a final volume of 25 ml (0.5-20.0) μg.ml-1 with relative standard deviation percent (R.S.D%) ranged between 0.26-4.71% and accuracy, expressed by recovery percent, 95-106% for five levels of parcetamol concentration. The second method is based on indirect spectrophotometric procedure for the determination of paracetamol, after its hydrolysis in acidic medium and the reaction of hydrolyzed product (p-aminophenol) with p-amino-2-hydroxy sodium benzoate in the presence of sodium periodate in alkaline medium. A bluish-violet water soluble compound is formed with λmax =580nm and εmax =11884 l.mole-1.cm-1 and Sandel’s index 0.0127 μg.cm-2. Beer’s law is applicable for concentration range of paracetamol 12.5-500.0 μg per 25 ml volume of solution (0.5-20.0) μg.ml-1 with (R.S.D %) ranged between 0.60-1.10 % and recovery percent 94.28-101.6% for three levels of parcetamol concentration. The proposed methods were successfully applied for the determination of paracetamol in pharmaceutical preparations by both direct and standard addition method and for three levels of concentration. Introduction Paracetamole chemically known as N-(4-hydroxyphenyl)acetamide is used therapeutically as an analgesic-antipyretic agent alone or associated with other drugs[1], it is effective in treating mild to moderate pain such as headache, neuralgia, and pain of musculo-skeletal orgin[2]. IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 Several analytical methods have been reported for the analysis of paracetamol in pharmaceutical or biological fluids, include titrimetric [3-6], fluoremetric[7-11], chromatographic[12-14], spectophotometric[15-19] and other procedures[20-23]. Some of these procedures are not simple for routine analysis and require expensive or sophisticated instruments. Oxidative coupling reactions, which rely on the coupling of two organic compounds in the presence of oxidizing agent under suitable conditions, could be considered as a type among the most important organic reactions which have wide applications in analytical chemistry[24,25]. The present work describes a spectrophotometric procedures for the assay of paracetamol. The first one is based on coupling with p-amino-2-hydroxysodium benzoate in the presence of sodium perchlorate, while the second depends on the oxidative coupling of the acid hydrolyzed product of paracetamol with p-amino-2-hydroxysodium benzoate in alkaline medium. Experimental Apparatus All absorbance measurements were performed by using a Spectronic 21D Uv-visible spectrophotometer, while the spectral measurements were carried out with Centra-5 double- beam spectrophotometer using 1-cm silica match cells. The pH measurements were taken with Jenway pH-meter3310. Reagents All reagents were of analytical grade. 1. Paracetamol stock solution (1000 µg.mL-1) was prepared by dissolving) 0.1gm of pure paracetamol powder (obtained from the state company for drug industries and medical appliance (S.D.I.), Samara-Iraq)in 10 mL of ethanol and diluted to 100 mL with distilled water. Working solutions were freshly prepared by subsequent dilutions. 2. P-amino-2-hydroxy sodium benzoate (AHB) solution (1.0 x 10 -2 M) was prepared by dissolving 0.2111gm of the reagent in distilled water and diluted to 100 mL in a volumetric flask. 3. Sodium periodate solution (1.0 x 10 -1M) was prepared by dissolving 2.1389 gm of the salt in 100 mL of distilled water. 4. Acetate buffer solution (pH = 4) was prepared by dissolving 13.7 gm of sodium acetate in 6 mL of glacial acetic acid and making the volume to 100 mL in a calibrated flask. The pH of the resulted buffer mixture was adjusted to pH = 4 ± 0.2 via the addition of glacial acetic acid using a pH-meter. 5. Solutions of pharmaceutical preparations containing paracetamol: Different pharmaceutical preparations from different sources containing paracetamol were obtained from local market (Table (1)). a. Solution for tablet analysis: 10 tablets were weighted out , grinded and mixed well. A portion of the resulted powder ( containing 0.1 gm of paracetamol) was used for the IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 preparation of the solution as described before. The resulted solution was filtered off and subsequently diluted to obtain working solutions. b. Solution for suppositories analysis: The containing of five suppositories was mixed well and a weight from the resulted mixture containing 0.1 gm of paracetamol was d issolved in 10 mL of ethanol and a little amount of hot distilled water. The solution was then diluted to 100 mL with hot distilled water and the volume was checked after the solution been cooled. Working solutions were prepared after filtration of the prepared stock solution. c. Hydrolyzed paracetamol solution: 25 mL of concentrated hydrochloric solution was added to 150 mL of 1000 µg.mL-1 of stock paracetamol solution in a round bottom flask. The mixture was refluxed for 1hr and after cooling the volume was diluted to 250 mL with distilled water. A 600 µg.mL-1 of the hydrolyzed paracetamol (as p-aminophenol) stock solution was obtained and working solution were freshly prepared after neutralizing ( pH = 7) the required aliquots of stock solution with 20% sodium carbonate solution before dilution with distilled water. d. Solutions of drugs for analysis by direct and standard addition methods: To 60 mL of 1000 µg.mL-1 solution of drug ( tablet or suppositories), which is prepared as described before, 10 mL of concentrated HCl solution (11.8 M) was added and the mixture was refluxed for 1 hr. After cooling the mixture was diluted with distilled water in a 100 mL volumetric flask. Working solutions were freshly prepared for each drug (100 µg.mL-1) after adjusting the pH to 7 with 20% sodium carbonate solution. Recommended Procedures 1.Direct Determination of Paracetamol : a.Calibration curve procedure: To a series of 25-mL volumetric flasks, different volumes (0.25 – 10 mL) of standard 50 µg.mL -1 of paracetamol solution were added followed by the addition of 1.6 mL of 0.1 M of NaIO4 solution, 0.6 mL of AHB reagent solution (1 x 10-2 M) and 2.0 mL of acetate buffer solution. The volume in each flask was diluted with distilled water and the absorbance of the formed colored compound was measured at 470 nm against reagent blank solution after 30 minutes. Figure (3) shows that linear calibration graph was obtained in the range of paractamol solution of (0.5 – 20.0 µg.mL-1) while higher concentrations show negative deviations from Beer's law. b. Standard addition procedure: Standard addition procedure was used for the assay of paracetamol contained in pharmaceutical preparations at three concentration levels (viz. 1, 5 and 10 µg.mL-1) as follows; to a series of f ive 25-mL calibrated flasks, aliquots of the sought drug solution containing (25 or 125 or 250 µg) of paracetamol were transferred followed by the addition of ( 0, 0.5, 1.0, 1.5 and 2.0 mL) of standard paracetamol solution ( 100 µg.mL-1) respectively. The resulted mixtures were then treated as described in the calibration curve procedure and the absorbances were measured at 470 nm after standing for 30 minutes. 2.Indirect Determination of Paracetamol : a.Calibration curve procedure: To a series of 25 mL calibrated flasks, different volumes (0.25 – 10 mL) of 50 µg.mL-1 of the hydrolyzed paracetamol solution were added followed by the IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 addition of of 0.2 mL of NaIO4 solution (0.1 M), 3 mL of AHB solution (1.0 x 10-2 M) and 2 mL of NaOH solution (1.0 M). Dilution was made with distilled water and the absorbance was measured after 20 minutes at λmax = 580 nm against reagent blank. Figure (6 ) shows the calibration curve which is linear in the concentration range of (0.5 – 20 µg.mL-1) of paracetamol. b. Standard addition procedure: The method was applied to cover three concentration levels of paracetamol (hydrolyzed as described before) in the studied druges ( viz. 2.0, 4.0 and 10.0 µg.mL -1 ) applying the following procedure; to a series of five 25 mL calibrated flasks aliquots of the studied drug solution (hydrolyzed as described before) containing (50 or 100 or 250 µg) of paracetamol were transferred, followed by the addition of (0, 1, 2, 3 and 4 mL) of standard hydrolyzed paracetamol solution (50 µg.mL -1) resepectively. The resulted mixtures were treated as described in calibration curve procedure and the absorbances were measured at 580nm after standing for 20 minutes. Results and Discussion 1.Direct Determination of Paracetamol The present work depends on quantitative oxidative-coupling of paracetamol via reaction with 4-amino-2-hydroxy sodium benzoate followed by spectrophotometric determination of the resulted colored compound. Preliminary investigations showed that a brownish-orange product resulted upon treating paracetamol solution with AHB and NaIO4 in acidic medium. The absorbance of the colored product was measured at 470 nm against reagent blank solution and its absorbance was found to be stable after 30 minutes of mixing(Fig. 1 a). A univariate study of the variables affecting the color development of reaction product, namely the amount of reagent for the range of (0.2 – 1.2 mL) of 0.1 M, the amount of oxidizing agent for the range (0.2 – 2.0 mL) of 0.1 M, the pH of the oxidation medium by using different volumes (0.2 – 6.0 mL) of acetate buffer (pH = 4.47), the order of mixing of analyte and reagent solutions and the of reaction temperature (17.5 ºC, 20 ºC, 25 ºC, 30 ºC, 40 ºC), was performed on a 20 µg.mL -1 of the analyte solution. One milliliter of AHB reagent solution was selected as an optimum amount (Fig.1b), and was used subsequently. Among different types and amounts of oxidizing agents which were investigated, 1.6 mL of 0.1 M solution of NaIO4 was found to be the optimum (Fig.1c). Different types of buffer solutions ( acidic and basic) were tested to control the pH of reaction medium. The study showed that using 2 mL of acetate buffer (pH = 4.42) gave the best result (Fig.1d). Table (2) illustrates the results obtained when different orders of mixing of analyte and reagents solutions were followed. The optimum order which gave the highest absorbance was: paracetamol solution(S) + NaIO4 solution (O) + AHB reagent solution (R) + acetate buffer solution (B). A suggested mechanism for the reaction is proposed in which paracetamol oxidized in the presence of NaIO4 to form a benzoquinon immine which in turn couples with AHB through ortho-position to -NH2 group in acidic medium as shown in the following scheme. IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 OH N H CH3 O NaIO4 O NH COONaH2N OH O N COOH NH2 HO pH=4.42 Final Absorption Spectrum Absorption spectrum of the colored product formed under optimum reaction conditions shows a maximnm absorption at 470 nm in contrast to reagent blank Fig (2 ). Calibration Graph Applying the conditions described in the recommended procedure, a linear calibration plot for paracetamol concentration in the range of ( 0.5 – 20 µg.mL-1) against the measured absorbance is obtained (Fig.(3)) with correlation coefficient of 0.9995. The conditional molar absorptivity was found to be 3.371 x 10 3 liter. mole -1 . cm -1 and Sandle's index 0.0449 µg.cm -2 . Interferences To assess the possibility of the analytical application of the proposed method, the effect of the presence of some foreign substances (viz. talk, glucose, lactose, starch) was studied by adding different amounts of the mentioned compounds to 15 µg.mL -1 of paracetamol solution and applying the recommended procedure for color development. Each of the studied compounds was considered not to interfere if its addition causes a relative error less than 5%. It was found the presence of 200 µg.mL -1of each of the compounds is tolerated in the determination of paracetamol. Precision and Accuracy To check the precision and accuracy of the proposed method, three replicate of paracetamol solution were determined under the established conditions at five concentration levels. The results are listed in Table 3. Analytical Applications Five types of commercially available paracetamol containing pharmaceutical preparations were analyzed. On applying the recommended procedure, good recoveries were obtained (Table 4). Comparison with Other Methods To assess the validity of the proposed method which was checked by comparing the results with those obtained by standard methods ( Table 5 ). Moreover, F-test and t-test show that there were no significant difference between the proposed and standard methods. IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 2.Indirect Determination of Paracetamol In this procedure, the acid hydrolysis product of paracetamol (i.e. p-aminophenol) is oxidized by sodium periodate and coupled with AHB in sodium hydroxide medium to y ield a brownish-violetwater soluble dye which absorbs at 580 nm. The experimental conditions for the color producing reaction were optimized by following the same steps mentioned in the direct method starting with arbitrary conditions ( i.e. mixing 1.0 mL of 0.1 M of NaIO4 solution, 1.0 mL of 1 x 10-2M of AHB solution and 1.0 mL of NaOH solution in a volumetric flask containing 300 µg of the analyte) and measuring the absorbance of the colored product after 20 minutes at 580 nm against reagent blank solution(Fig. 4 a). The effect of using different volumes of coupling reagent solution(1.0 x 10-2 M) was investigated, and the results show that 3.0 mL gave the highest absorbance (Fig.4 b ) which was used as a subsequent work. The results obtained from using various volumes (0.1 – 2.0 mL) of 0.1M of NaIO4 (Fig.4 c), indicate that 0.2 mL of the oxidizing reagent was the optimum. The study shows that a stable color product could only be obtained in alkaline medium, therefore, the reaction was carried out in the presence of ammonium buffer (pH range 9 – 12 ) and in the presence of NaOH. Figure (4 d) shows that using 2.0 mL of 1.0 M NaOH solution resulted in obtaining the best results. The order of addition of the reacting solutions shows that the following order : (S) + (O) + (R) + (B) must be followed to obtain the highest absorbanc (Table 6). Finally, the effect of temperature on the color reaction was investigated in at the given values (20 ºC, 25 ºC, 30 ºC, 40 ºC and 50 ºC). The investigation shows that the colored product is not stable at elevated temperatures, therefor, the reaction was carried out at room temperature (i.e. 20 ºC). A mechanism was suggested for the oxidation of the hydrolysis p roduct of paracetamol via NaIO4 and coupling with AHB to form the coloed product in alkaline medium as follows: OH OH NH2 N H CH3 O conc. HCl NaIO4 O NH COONaH2N OH O N COONa NH2 HO NaIO4/NaOH Final Absorption Spectrum The absorption spectrum of the colored product was recorded, for the range (400 – 700 nm), under established optimum conditions against reagent blank solution. Figure (5) shows a spectrum with wavelength of maximum absorption(λmax) at 580. IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 Calibration Graph Employing the recommended procedure, Beer's law is obyed over the range of ( 12.5 – 500 µg.mL-1 ) of paracetamol per 25 mL of solution with correlation coefficient of 0.9956 (Fig. 6). The conditional molar absorptivity for the colored product was found to be 11884 liter. mole- 1.cm-1 and Sandl's index was 0.0127 µg.cm-2. Interferences The presence of 250 µg.mL-1 of foreign substances (talk, lactose and starch) can be tolerated in the analysis of 12 µg.mL-1 of paracetamol solution since it would cause relative error percent less than 5%. Precision and accuracy The precision of the proposed method was calculated in term of R.S.D.% and its accuracy in term of relative error percent for three replicates of solution at three concentration levels (2, 10 and 20 µg.mL-1 ). Satisfactory results were obtained under optimum conditions (Table 7). Analytical application Table ( 8 ) shows the results obtained upon application of the proposed method in the determination of five drugs containing paracetamol. Comparison with other methods Table (9) shows the results obtained by the proposed and other standared methods. Both F- test and t-test were applied and showed that there were no significant differences between the results obtained in comparison with standard methods. References 1. Martindal, "The Complete Drug Reference"(2002)., 33rd edition , Pharmaceutical press, London, 2. " British Pharmacopoeia on –CD-ROM "(2005)., 3rd Ed. , Copyright by System Simulation Ltd., The Stationery Office , London, 3." Indian Pharmacopoeia " (1996)., Thecontroller of Publication , Delhi, 4. Parimoo, P. ;Sethuraman, Amalarj, A. and Seshadari, N. (1989). Indian. Drugs. , 26 : 704 5. Ding, T. ; Wang, j. and Zhongguo Yaovue Zazhi (1989) 24:295-296 ; Anal. Abst. , 52, 2E11 (1990). 6. Mahendra, K. ; Ahmed, S. ; Singh, D. and Shukla, I. (1985)Analyst, 110 : 735 7. Oztunc, A. (1982) Analyst, 107:585 8. Kaito, T. Kazuhiko, S. , Yuji, I. and Tsuguchila, Y. (1974) J. Pharm. Soc. , 94:633 9. Tsunetoshi, k. and Kazunhka, S. (1974).J. Pharm. Soc . , 94:639 10. Kaito, T. ;Kazuhiko, S. and Yuji, I. (1976) Bunski. Kagaku, 25:776. Throgh anal. Abst. , 33:1E53 (1977). 11. Pulgarian, J. A. M. and Bermcgo, I. F. G. (1996) Anal. Chimi. Acta. , 333: 59 12. Gopinath, R.; Rajan, S. ;Meyyanatnan, S. N.; Krishnaveni, N.and Suresh, B. (2007). Indian J. of Pharmaceutical Sciences, 6a(1):137-140, 13. Senthamil Selvan, P.; Gopinath, R.; Saravanan,V. S.; Gopal, N.; Sarvana Kumar, A. and Periyasamy, K. (2007) Asia J. of Chemistry, 19(2): 1004-1010 IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 14. Emre, D.and Ozaltin, N. (2007). J. of Chromatography B: Analytical Technologies in the Biomedical and life Sciences, 847(2):126-132, 15. Lavorante, A. F.;Pires, C. K.and Reis, B. F. (2006).J. of Pharmaceutical and biomedical analysis, 42(4): 423-429, 16. De Los, A.; Oliva, M.;Olsina, R. A.and Masi, A. N. (2005)Talanta, 66(1): 229-235, 17. Burakham, R.; Duanhthong, S.; Patimapornlert, L.; Lenghor, N.; Kasiwad, S. ;Srivichai, L.; Lapanantnoppakhum, S.; Jakmunee, J.and Grudpan, K. (2004) Analytical Sciences, 20(5): 837-840, 18. Abbas, A.; Nahid, S.and Ali, R.Z. (2006)Acta Chim. Slov., 53:357-362 19. Wafaa, S. H. (2008).American Journal of Applied Sciences, 5(8):1005-1012, 20. Walash, M . I. ; Elbrashy , A. M. and Sultan, M. A. (1994) Mikrochim, Acta , 113:113-124 21. Danet, A. F. ; David, V. and David, I. (1998).Rav. Roum. Chim. , 43:811-816 22. Easwaramoorthy , D. ;Ya, Y. C. and Huang, H. J. (2001). Anal. Chim , Acta. 439:95-100 23. Azhagvuel, S.and Sekar, R., (2007) J. of Pharmaceutical and Biomedical Analysis, 43(3):873-878, 24. Russel, J. A. (1944) J. Biol. Chem. , 156:165 25. Lubochinsky, B. and Zalta, J. P. (1954).Bulls. Soc. Chem. Biol. 36: 1363 IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 Fig. (1): Optimization of experimental conditions. Fig. (2): Final absorption spectrum. Fig. (3): Calibration graph of paracetamol IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 Fig. (4): Optimization of experimental conditions. Fig.(5): Final absorption spectrum. Fig. (6): Calibration graph of paracetamol . IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 Table (1): Pharmaceutical preparations used Company Contains Pharmaceutical preparation S.D.I -Iraq 500 mg paracetamol Paracetamol tablets S.D.I -Iraq 350 mg paracetamol 50 mg caffeine 10 mg codeine phosphate Algesic tablets U.K- London 500 mg paracetamol Paramol tablets M.P.K- Syria 450 mg paracetamol 35 mg orphenadrine citrate Kanagesic tablets S.A.R 250mg paracetamol Panatol delta Suppositories Table (2): Effect of order of mixing Table( 3): Precision and accuracy of the method Absorbance Order of addition 0.466 S+O+R+B 0.465 O+S+R+B 0.459 R+S+O+B 0.389 B+S+O+R 0.431 O+R+B+S 0.445 R+O+S+B Taken Conc. (ppm) Found Conc. (ppm) Average Recovery (%) Error (%) R.S.D (%) 0.5 0.4909 0.4507 0.4839 0.4751 95.0202 -4.9798 4.5156 1 1.1261 1.0334 1.0456 1.0684 106.8377 +6.8377 4.7155 5 5.0876 5.1125 5.0904 5.0968 101.9363 +1.9363 0.2681 10 9.6469 9.8640 9.8092 9.7734 97.7341 -2.2659 1.1549 20 20.1984 20.0394 20.7091 20.0863 100.4315 +0.4315 0.2796 IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 Table (4): Analytical application of the direct and standard addition methods. Pharmaceutical preparation Procedure Amount Recovery (%) Certified value (mg) Conc. Of Paracetamol (µg/ml) Found Value(mg) Each assay Average Paracetamol tablets S.D.I –Iraq Calibration curve 500 10 522.8460 104.5690 101.2435 5 509.0640 101.8120 1 486.7421 97.3484 Standard addition 10 507.591 101.5182 99.3506 5 490.675 98.1350 1 492.00 98.4000 Algesic tablets S.D.I –Iraq Calibration curve 350 10 365.5990 104.456 102.0797 5 364.2092 104.059 1 342.0296 97.7226 Standard addition 10 351.583 100.4523 100.2193 5 351.344 100.3840 1 349.376 99.82172 Paramol tablets U.K- London Calibration curve 500 10 485.3933 97.0787 98.7492 5 501.5730 100.314 1 494.2322 98.8464 Standard addition 10 502.4390 100.4878 100.5014 5 502.7851 100.5570 1 502.2971 100.4594 Kanagesic tablets M.P.K- Syria Calibration curve 450 10 453.7079 100.823 100.3779 5 458.1573 101.812 1 443.1236 98.4969 Standard Addition 10 452.1344 100.4743 100.5266 5 453.4599 100.7688 1 451.5152 100.3367 Panatol delta S.A.R Calibration curve 250 10 261.4232 104.569 104.3645 5 260.1498 104.059 1 261.1610 104.464 Standard addition 10 249.2453 99.69812 100.3144 5 250.9978 100.3991 1 252.1151 100.8460 Table (5): Comparison with other methods Recovery % Drug samples Standard method The proposed method 101.000* 100.394 Paracetamol Pure 99.000* 101.243 Paracetamol tablets S.D.I –Iraq 97.970 ** 102.079 Algesic tablts S.D.I –Iraq 100.501 *** 98.749 Paramol tablts U.K- London 100.526*** 100.377 kanagesic tablts M .P.K- Syria 100.314*** 104.364 Panatol delta S.A.R * British Pharmacopoeia B.P, ** S.D.I standard methods , *** Oxidative coupling versus standard addition value. IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 Table (6): Effect of order of mixing Absorbance Order of Addition 1.062 S + O + R + B 1.026 S + R + O + B 0.872 S + B + O + R 0.950 R + O + S + B 0.954 R + S + B + O 0.898 B + S + O + R 1.020 O + S + R + B 0.794 O + R + B + S Table (7): Precision and accuracy of the method. Taken Conc. (ppm) Found Conc. (μg.ml -1 ) Average Recovery (%) Error(%) R.S.D(%) 2 1.8651 1.8899 1.9022 1.8857 94.2863 -5.7137 1.0025 10 10.0582 10.1448 10.2809 10.1613 101.6130 + 1.6130 1.1052 20 19.9839 19.8601 20.1077 19.9839 99.9196 -0.0804 0.6193 IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L. 23 (1) 2010 Table (8): Analytical application of the direct and standard addition methods. Pharmaceutical preparation Procedure Amount Recovery (%) Certified value (mg) Conc. of Paracetamol (µg/ml) Found Value(mg) Each assay Average Paracetamol tablets S.D.I –Iraq Calibration curve 500 10 506.8948 101.3789 99.7617 4 493.2985 98.6597 2 496.2322 99.2464 Standard addition 10 504.8823 100.9764 100.9449 4 504.9708 100.9941 2 504.3217 100.8643 Algesic tablets S.D.I –Iraq Calibration curve 350 10 336.4066 96.1161 99.2741 4 348.4753 99.5643 2 357.4970 102.1421 Standard addition 10 356.9930 101.9980 101.5203 4 354.3256 101.2359 2 354.6450 101.3271 Paramol tablets U.K- London Calibration curve 500 10 486.9397 97.3879 99.3166 4 491.4631 98.2926 2 511.3459 102.2692 Standard addition 10 505.4348 101.1087 101.0055 4 502.8191 100.5638 2 506.7203 101.3440 Kanagesic tablets M.P.K- Syria Calibration curve 450 10 458.2759 101.8391 100.8003 4 448.0397 99.5644 2 454.4884 100.9474 Standard Addition 10 453.9413 100.8758 99.6598 4 450.9009 100.2002 2 440.4731 97.8829 Panatol delta S.A.R Calibration curve 250 10 257.7771 103.1108 103.6618 4 264.0131 105.6052 2 255.6730 102.2692 Standard addition 10 246.2356 98.4942 100.3073 4 252.5189 101.0076 2 253.5550 101.4220 Table( 9): Comparison with other methods. Recovery % Drug samples Standard method The proposed method 101.0000* 99.7617 Paracetamol Pure 99.0000 * 98.6000 Paracetamol tablets S.D.I –Iraq 97.9700 ** 99.2741 Algesic tablts S.D.I –Iraq 100.5014*** 99.3165 Paramol tablts U.K- London 100.5260*** 100.8002 kanagesic tablts M .P.K- Syria 100.3140*** 103.6617 Panatol delta S.A.R * British Pharmacopoeia B.P, ** S.D.I standard methods , *** Oxidative coupling versus standard addition value.