100 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . Spectrophotometric Determination of Chlorpromazine – HCl Using Potassium Permanganate Sarmad B. Dikran Alaa K. Mohammed Azhar S. Hammodi Dept. of Chemistry/College of Education for pure science (Ibn Al- Haitham) University of Baghdad Received in: 26 March 2014، Accepted in:21 December 2014 Abstract Twosimple, sensitive,accurate, and precise spectrophotometric methods have been developed for the determination of chlorpromazine – HCl in pure form and pharmaceutical formulation. The first method involved treatment of cited drug with a measured excess of permanganate in acid medium and the unreacted oxidant was measured at 525 nm. The second method involves the reaction of the drug with potassium permanganate in the presence of sodium hydroxide to produce a bluish – green colored manganite which is measurable at 610nm. All the experimental variables affecting the development of the manganite ions were investigatedand conditions were optimized. Working linearity ranges were 5-45 µg.mL-1and 1-20 µg.mL-1 by two methods respectively. The apparent molarabsorptivities are 4.015  103and 18.717 103L.mol-1.cm-1 respectively, with corresponding Sandell’s sensitivityvalues of0.0885and 0.01798µg.cm-2 respectively. The methods have successfully been applied to the determinationof drug in dosage forms. Key Words: Chlorpromazine – HCl , Potassium Permanganate, Spectrophotometry. 101 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . Introduction Chlorpromazine is chemically (10-[3–dimethylaminopropyl] phenothiazine), is a phenothiazine neuroleptic used for the control of psychoses including schizophrenia and mania and it also used in treatment of vomiting and vertigo because of its sedative and extrapyramidal effect.[ 1,2 ] Several methods have been reported for the determination of chlorpromazine in both pharmaceutical dosage forms and biological fluids includes HPLC [ 3,4 ], GC [5,6],voltammetry[7] , potentiometry[8] and spectrophotometry [9-12].The aim of the present work is to develop two simple sensitive and cost–effective spectrophotometric methods forthe determination of chlorpromazine in pure and pharmaceutical formulations. The methods involve using permanganate as oxidimetric reagent for the determination ofchlorpromazinein either acid or alkaline medium. In acidic medium the unreacted permanganate ion is determined at 525 nm, while in alkaline mediumthe bluish–green color of manganate is measured at 610 nm.The validated methods when applied to the determination of chlorpromazine in dosage form yielded results, whichwere in agreement with the label claim. Experimental Instruments A centra 5 GBC–Scientific–Equipment UV-Visible double beam spectrophotometer with 10 mm matched quartz cells was used for all absorbance measurements. Reagentsand Materials All chemicals used were of analytical grade and distilled water was used to prepare all solutions. For Method A Potassium permanganate solution (0.0038M) is prepared by dissolving 0.058gm of KMnO4 in 50 ml of distilled water and treated as mentionedabove, then diluted to the 100mL by using a volumetric flask. Sulfuric acidsolution (2 M) is prepared by mixing 5.50 mL of concentrated sulfuric (sp.gr1.84) with 50 mL of distilled water and diluted to the mark in a 100 mL volumetric flask using distilled water. Standard drug (100 ppm) solution is prepared by dissolving 10 mg of the drug in 10 mL of 0.1 M H2SO4 and diluting to the mark in a100 mL volumetric flask with the same acid. For Method B Potassium permanganate solution (0.0126 M) is prepared by dissolving approximately 0.2 gm of KMnO4 (Riedel–de Haen-Germany ) with 50 mL of water, the solution was boiled for 10 minutes to remove any residual manganate (IV) ions, cooled filtered and diluted to 100 mL and standardized. Sodium hydroxide solution (2 M) is prepared by dissolving 8 gm of NaOH in 100 mL of water. Acetic acid solution (0.5 M) is prepared by mixing 2.85 mL of concentrated acetic acid (sp.gr. 1.052) with distilled water and making the final volume to 100 mL in a volumetric flask. Standard drug (100 ppm) solution is prepared by dissolving 10 mg of the drug in 10 mL of 0.5 M acetic acid and diluted to 100 mL in a volumetric flask with distilled water. 102 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . General Assay Procedures and Calibration Curves Method A Aliquots of the working standard solutionof containing chlorpromazine(50-450 µg) were transferred into a series of 10 mLvolumetric flasks andthe volume was adjusted to 4 mL with distilled water. To each flask, 2mL of2 M H2SO4 was added followed by 1 mLof 0.0038M KMnO4. The flasks were kept aside for 20 minwith occasional shaking before diluting to the mark with distilled water. The absorbance of each solution was measured at 525nm against areagent blank. Method B Aliquots of the working standard solution of chlorpromazine containing(10-200 µg)were transferred into a series of 10 mL volumetric flask and the volume in each flask was adjusted to 7mL with distilled water. To each flask, 2 mL of 2 M NaOH were added followed by 1mL of 0.0126M KMnO4. The flasks were kept aside for 20 min with intermittent shaking before diluting tothe mark withwater. The absorbance was measuredafter complete color formationat 610nm against areagent blank. Procedure for the Assay of the Drug in Pharmaceutical Formulation Tablets An accurately weighed quantity of the mixed content of 10 tablets equivalent to 10 mg of drug was transferred into 100 mL volumetric flask and dissolved with 0.1 M H2SO4, mixed and filtered using a Whatman no 42 filter paper to avoid any suspended or undissolved material before use.The first 10 mL of the filtrate was discarded and the filtered solution was diluted quantitatively with distilled water to obtain suitable concentration for the analysis . Ampoules An accurately measured volume of the mixed contents of 5 ampoules equivalent to 25 mg of chlorpromazine were transferred into 50 mL volumetric flask , and diluted to 50 mL with distilled water.Theworking solutions werefreshly prepared bysubsequent dilutions and analyzed by the above procedure. Results and Discussion Optimization of Experimental Variables The spectrophotometric properties of the colored product as well as the different experimental parameters affecting the color development werecarefully studied.The optimization of experimental conditions is accomplished by sequentially optimizing one variable at a time while keeping all other variables constant, these factors include time required formaximum and stable color development, concentration of potassium permanganate. Method Development Potassium permanganate is a strong oxidizing agent that can react with several organic substances. Recently, permanganate was studied to determine pharmaceutical active compounds[13]in formulation of both in alkaline medium[14] as well as in acidic medium [15]. 103 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . Method A In order to determine the permanganate concentration which would give a reasonable maximum absorbance,preliminary experiments were performed in sulfuric acid medium at 525 nm (Figure 1). Hence different concentrations of chlorpromazine (5.0-45µg.mL-1) were reacted with 1 mL of 0.0038 M KMnO4in acidic medium, and after elapsed the contact time the absorbance of the residual permanganate was measured and related to chlorpromazine concentration. When different concentrations of chlorpromazine react with a fixed amount of KMnO4(60µg) in H2SO4 acid medium the KMnO4 was consumed in proportion to chlorpromazine concentration and a concomitant fall in the concentration of KMnO4 will take place as shown by the decreasing absorbance values at 525 nm with the increase ofchlorpromazineconcentration. The results are representedin Figure 2. Effect of Reaction Time To study the effectof reaction time for maximum color development,the method was fixed by carrying out the experiment using 10 µg.mL-1of chlorpromazine. The maximum intensity of color was obtained after 20 minutes and remained constant up to 45 min at room temperature (Figure 3). Effect of Volume of Sulfuric Acid Different volumes (0.5 – 3.0)mL of 2 M H2SO4 were mixed with 1 mL of chlorpromazine (10 µg), and 1 mL of 0.0038 M KMnO4. It was found that the highest color intensity was attained at (2.0-2.5) mL of 2M H2SO4(Figure 4). Excess volume of H2SO4 hadlittleeffect on the absorbance of chlorpromazine;therefore, 2 mLwas selected forrecommended procedure. The possible reaction between the permanganate and the drug in acidic medium could be represented as in scheme 1. CLP + known excess of KMnO4→ CLP-sulfoxide + unreacted KMnO4 (measured at 525 nm) Scheme 1: Proposed reaction in acidic medium. Method B In this method a green colored manganate ion MnO4-2 was obtained as a result of reduction of KMnO4 by chlorpromazine in the presence of NaOH which showed maximum absorption peak at 610 nm against the reagent blank (Figure 5). Effect of Reaction Time The optimum reaction time for the development of color intensity at ambient temperature (25 ± 2oC) was studied and it was found that complete and maximum intensity was obtained at 20 min and remained constant up to 2 hrs.(Figure 6), therefore, 20 min of reaction time was usedthroughout thedetermination process. Effect of Volume of NaOH The influence of volume of 2M NaOH for green colored manganate ion MnO42- was examined in the range of (1 – 3.5mL).It is apparent from Figure 7, that increasing the volume of NaOH would increase the absorbance of the reaction product upto 2.5 mL; after which furtherincrease in the volume resulted in no change in the absorbance. Thus, 2.5mL of2M NaOH waschosen as an optimum value for maximum absorbance. 104 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . Effect of Volume of Potassium Permanganate The effect of the volume of 0.0126 M of KMnO4 on the intensity of the green colored developed was studied in the range of(0.5-2.5 mL).It is evident from Figure 8 that the maximum absorbance was attained with 1.5mL KMnO4was selected for all the experimental process. The possible reaction between the permanganate and the drug in alkaline medium could be represented as in scheme 2. CLP + Excess of KMnO4 → K2MnO4+ oxidation product of CLP (measured at 610 nm) Scheme 2: Proposed reaction in alkaline medium. Validation of the Proposed Methods Concentration Range and Calibration Graphs Under the optimized experimental conditions, the response measured for each methods at the specified working wavelengths was found to be proportional to the analyte concentrationFigure 9 and Figure 10. The linear regressionequations were derivedby least–squares treatment of the calibration data. (Table 1), summarized the performance data and statisticalparameters for theproposed methods. Sensitivity Sensitivity of the proposed method was evaluated by calculating limit of detection (LOD) and limit of quantification (LOQ). LOD is the lowest detectable concentration of the analyte by the method while LOQ is the minimum quantifiable concentration. LOD andLOQ werecalculated by equations:LOD and LOQ respectively,where s is the standard deviation of a blank and m is the slope of the calibration curve. The results Table (2) indicatingproposed methods are highly sensitive. Precision and Accuracy The competence of the method wasstatistically evaluated by measuring accuracy and precision of the proposed methods. The accuracy was determined by measuring the relative error percentage (E%), and the results are shown in Table (2).The obtained results were satisfactory andindicate that theproposed methods have a good accuracyand precision. Application of the Proposed Methodsto Pharmaceuticaldosage Forms It is evident from the aforementioned resultsthat theproposed methodgave satisfactory results with the investigateddrugs.The results of the application of the recommended procedure aresummarized in Table (3). 105 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . References 1. Marder, S.R.(1998)Antipsychotic Medication, IntheAmerican Psychiatric Press Textbook of Psychopharmacology.Schatzberg AF and Nemeroff CB (eds) American Psychiatric Press. Washington DC.309 -321. 2. Malcolm , L. (1983)Introduction topsychopharmacology Upjohn company, Michigan, 51-61. 3. Tanaka,E.;Nakamura, T.; Terada, M. ; Shinozuka, T.; Hashimoto, C.andKurihara, K. (2007) Simple and Simultaneous determination for12 phenothiazines in human serum by RP –HPLC. Journal of Chromatography B,1-2 (854) 116 -120 . 4. Cruz–Vera, M.; Lucena, R.;Cardenas, S.and Valcarcel, M. (2009) Determination of phenothiazine derivatives in human urine by using ionic–based dynamic liquid– phase microextraction coupled with liquid chromatography, Journal of chromatography B, 1-2( 877) 37-42 . 5. Shen, M. ;Xiang, P.; Shen, H. Wu, B. andHuang, Z. (2002) Detection of antidepressant and antipsychotic drugs in human hair, Forensic Science international,2(126),18 April; 153-161. 6. Pujadas,M.; Pichini,S.;Civit, E. ;Santamarina, E. ; Perez , K. and dela Torre R. (2007). Asimple and reliable procedure for the determination of psychoactive drugs in oral fluid by gas chromatography-mass spectrometry, Journal of pharmaceutical and Biomedical Analysis 2(44) 594 – 601 7. Yongnian, Ni.; Wang,Li.and Serge Kokot,(2001). Voltametric determination of chlorpromazine hydrochloride and promethazine hydrochloride with the use of multivariate calibration,Analytica Chimica Acta. 439 , 159–168 8. Farhadi,K. and Shamsipur,M.(2003)Potentiometric and spectrophotometric determination of phenothiazine derivatives based ontheir titration with 2,3–dichloro- 5,6-dicyano-1,4- Benzoquinone, Acta. Chim slov, 50, 395–407 . 9. Hassouna,M.E.M.;Adawi,A.M.andAli,E.A.(2012)Extractive spectrophotometric determination of chlorpromazine and trifluoperazine hydrochloride in pharmaceutical preparations, Egyptian Journal of Forensic Science,2(2) 62-68 . 10. Basavaiah, K.(2004) Determination of some psychotropic phenothiazine drugs by charge–transfer complexation reaction withchloranilic acid,II Farmaco, 4(59) 315 - 321. 11. Abdul Latif,M.H.(2008) Spectrophotometric determination of chlorpromazine–HCl by Ion–Pair complex formation with Acid–Dyes,Ibn–AL–HaithiamJ. for pure andApplied Sci., 21 (1) . 12. K.Basavaiah,K.;Nagegowda,P.;Prameela,H.C.andSomashekar,B.C.(2005)Sensitive titrimetric andspectrophotometric assaymethods for chlorpromazine with bromated– bromide mixture and two dyes,Indian Journal of Chemical Technology,(12) 25-29 . 13. Askal,H.F.;Abdelmageed,O.H.;Ali,S.M.S.andAbo.ElHamd,M.(2010)Spectrophotomet ric and spectrofluorimetric determination of 1,4-dihydropyridine drugs using potassium permanganate and cerium (IV) ammonium sulphate. Bull. Pharm.Sci.,2( 33). 14. Venkata Shanmukha,J.; Prasanthi,S.; Guravaiah,M.and Bala Sekaran,C.(2010). Application of potassium permanganate to the spectrophotometric determination of oseltamivirphosphate in bulk and Capsules . Asian .Journal of pharmaceuticaland clinical research. 2( 5) . 15. Rahman,N.; Anwar,N.;Kashif,M.; Hoda,N.and,Rahman,H.(2011) Determination of labetalol hydrochloride by kinetic spectrophotometry using potassium permanganate as oxidant;J.Mex.Chem.Soc. 55 (2), 105 – 112 . 106 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . Table No(1): Analytical parameters for the proposed methods Method B Method A Parameter 610 525 λmax (nm) 1 – 20 5 – 45 Concentration range (µg.mL-1) Y=0.0587x+0.0333 Y=0.0113x+0.5328 Regression equation 0.0333 0.5328 Intercept 0.0587 0.0113 Slope 0.9994 0.9992 Correlation coefficient (r) 18.717  103 4.015  103 Molar absorptivity (L.mol-1 .cm-1) 0.0180 0.0885 Sandell’s sensitivity (µg. cm-2) 0.545 -- Limit of detection (LOD) (µg.mL-1) 1.652 -- Limit of quantification (LOQ) (µg.mL-1) Tablet No.(2): Evaluation of accuracy and precision for the determination of chlorpromazine-HCl R .S .D % Relative Error % Conc. (µg.mL-1) Found * Taken 2.016 2.792 1.805 -0.80 -1.515 -1.410 9.920 19.69 39.436 10 20 30 Method A 0.937 0.508 2.792 + 2.42 +1.730 -1.515 5.121 10.038 19.69 5 10 20 Method B *Average of three determinations. 107 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . Tablet No.(3): Assay results of different brands of chlorpromazine using the proposed methods. Method A R.S.D % R .E % Conc.found (µg.mL-1) Conc.taken (µg.mL-1) Found* Amount (mg) Labeled Amount (mg) Sample 2.860 1.730 10.173 10 102.01 100 Largactil OUBARI Syria 1.510 1.125 20.225 20 101.9 100 Largactil S D I/Iraq 1.579 -0.58 9.942 10 23.19 25 Ampoule 25mg/5mL OUBARI Syria Method B 1.810 2.02 5.101 5 101.92 100 Largactil OUBARI Syria 0.690 0.21 10.021 10 100.09 100 Largactil S D I/Iraq 0.965 -0.38 9.962 10 24.69 25 Ampoule 25mg/5mL OUBARI Syria *Average of three determinations. 108 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . Figure No (1): Visible spectrum of (60 µg.mL-1) potassium permanganate in the presence of 2 M H2 SO4. Figure No (2): Effect of chlorpromazine concentration on the absorption spectrum of 60 µg.mL-1 of KMnO4 (A for 5 µg.mL-1, B for 15 µg.mL-1, C for 25 µg.mL-1, D for 35 µg.mL-1, and E for 45 µg.mL-1). 109 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . Figure No.(3): Effect of reaction time Figure No.(4): Effect of H2SO4 volume Figure No.(5): Visible spectrum of manganite (MnO4-) in the presence of2M NaOH. 0.37 0.38 0.39 0.4 0.41 0.42 0.43 0 10 20 30 40 A b so rb a n ce Time (min) 0.38 0.39 0.4 0.41 0.42 0.43 0 0.5 1 1.5 2 2.5 3 3.5 A b so rb a n ce Volume of H2SO4 (mL) 110 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . Figure No.(6): Effect of reaction time and stability species. 0.3 0.35 0.4 0.45 0.5 0.55 0.6 0.65 0.5 1 1.5 2 2.5 3 3.5 4 A b so rb an ce Volume of NaOH (mL) Figure No. (7): Effect of NaOH volume Figure No.(8): Effect of KMnO4 volume. Figure No. (9):Calibration graph of chlorpromazine – HCl under optimum experimental conditions for method A. y = -0.0113x + 0.5332 R² = 0.9985 0 0.1 0.2 0.3 0.4 0.5 0.6 0 10 20 30 40 50 A b so rb a n ce Concentration of chlorpromazine (µg.mL-1) 111 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . Figure No.(10):Calibration graph of chlorpromazine – HCl under optimum experimental conditions for method B. y = 0.0587x + 0.0333 R² = 0.9992 0 0.2 0.4 0.6 0.8 1 1.2 1.4 0 5 10 15 20 25 A b so rb a n ce Concentration of chlorpromazine (µg.mL-1) 112 | Chemistry 2015) عام 1العدد ( 28مجلة إبن الھيثم للعلوم الصرفة و التطبيقية المجلد Ibn Al-Haitham J. for Pure & Appl. Sci. Vol. 28 (1) 2015 . ھيدروكلوريد باستعمال-لتقدير الطيفي لعقار كلوروبرومازينا برمنكنات البوتاسيوم سرمد بھجت ديكران محمدعالء كريم أزھار صادق حمودي )/ جامعة بغدادابن الھيثم(الصرفة قسم الكيمياء/ كلية التربية للعلوم 2014كانون االول 2قبل البحث في :،2014اذار 26استلم البحث في: الخالصة ھيدروكلوريد بصورته النقية وفي - اقترحت طريقتان طيفيتان سھلتان وحساستان ودقيقتان لتقدير الكلوروبرومازين بعض المستحضرات الصيدالنية. تضمنت الطريقة األولى معاملة العقار مع محلول برمنكنات البوتاسيوم في وسط حامضي نانوميتر. أماالطريقة الثانية فتضمنت مفاعلة العقار مع محلول برمنكنات 525وقياس االمتصاص عند الطول الموجي نانوميتر. 610األخضر المزرق عند طول موجي قدره4MnO-البوتاسيوم في وسط قاعدي وقيس امتصاص لون أيون ة قانون بيير درست المتغيرات للوصول إلى الظروف الفضلى للتفاعلين، وأظھرت النتائج التي تم الحصول عليھا مطاوع لكال الطريقتين على التوالي وكانت قيم معامالت مايكروغرام/مل 20- 1.0و 45-5في مديات تراكيز تراوحت بين و 0.0885لتر/مول.سم وقيم حساسية ساندل ھي 318.717x10و34.015x10االمتصاص المولية لكلوروبرومازين في اقراص دوائية وامبوالت من على التوالي.طبقت الطريقتان بنجاح لتقدير ا 2مايكروغرام/سم 0.01798 مناشئ مختلفة. يدروكلوريدھ- المطيافية، برمنكنات البوتاسيوم، كلوروبرومازين الكلمات المفتاحية: