IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL. 24 (1) 2011 Univariate and Simplex Optimization for The Analysis o f Diphenhydramine-HCl Through Ion-pair Formation Received in : 4 May 2010 Accepte d in : 3 June 2010 A. K. Mohamme d, S. B. Dik ran, M. Abdul Sattar Alubydi Departme nt of Chemistry, College of Education Ibn Al-Haitham, Unive rsity of Baghdad Abstract A Simp le, rapid and sensitive extractive and sp ectrop hotometric method has been described for the analysis of dip henhy ldramine –HCl (DPH) in p ure form and in p harmaceutical formulations. The method is based on the formation of chloroform soluble ion-pair comp lex with Bromophenol b lue(BPB) in a phthalate buffer at p H 3.0.T he extracted comp lex shows maximu m absorbance at 410 nm. B eer ' s law is obeyed in the concentration range 0.2-25.0 µg. ml -1 . The molar absorp tivity and Sandell's sensitivity for the sy stem bein g 2.416x10 4 L.mol -1 .cm -1 and 0.012µg. cm -2 , resp ectively. The limit of detection was found to be 0.155 µg.ml -1 . The p rop osed method was successfully app lied to the determination of the dru g in p harmaceutical formulation and satisfactory results were obtained. Key words: The Analy sis of Dip henhy dramine-HCl Through Ion-p air Formation, Univariate and Simp lex Op timization Introduction Dip henhy dramine, is chemically known as 2-(diphiny lmethoxy )-N-N-dimethy lamine hy drochloride (DPH), it is an antihistaminic sp ecies in the H1-receptor antagonist that can be used as antiallergic, antiemetic and antitussive drugs found in many p harmaceutical p rep aration.[1]. Sever al analy tical methods were p reviously p rop osed for determining DPH in p harmaceutical sa mples includin g titrimetry [2], fluorimetry [3], HPLC[4], HPTLC[5], cap illary electrop horesis[6], gas chromatography [7] and electrochemical analysis[8]. UV-visible sp ectrop hotometry is still considered to be the most convenient method for the assay of different classes of drugs in p ure, p harmaceutical formulation and in biolo gical samples, because of its simplicity and reasonable sensitivity with significant economical, advantages. Sever al sp ectrop hotometric methods have been reported for the determination of DPH in bulk material and dosage forms. San Andre ’ s [9] have descr ibed a sp ectrop hotometric method based on measuring the critical micelle concentration of mixed sodium dodecy l sulfate-antihistamine aggregates. In another extractive sp ectrop hotometrice method [10] the drug was p recipitated with reineckate and the ion-pair comp lex was extracted with nitrobenzene and measure ment at 520nm. The drug has also been determined IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL. 24 (1) 2011 sp ectrop hotometrically via the formation of an-ion-p air comp lex with Bromocresol-Green at p H 3.0 [11]. On the other hand, Bromop henol blue which is an an ionic dye has be en used as a p recipitating reagent to form an ion-p air comp lex as well as in other flow-injection p rocedures with a turbimetric determination [12], here, the reaction between DPH and Bromophenol blue takes p lace on-lin e and no liquid extraction is requ ired. In exp erimental chemistry , the optimization of technical sy stem is the p rocess of the adjust ing the control variables to find the levels that achieve the best op timization .Usually, many conflicting resp onses must be optimized simultaneously . In lack of systematic approaches the optimization is done by trial and error, or by changing one control variable at a time while hold ing the rest constant, such methods require a lot of e xp eriments t o be carried out. Simp lex op timization of exp erimental p arameters was first introduced by sp indly [13],and then modified by Nedler [14]and Albery [15]. A simp lex is a geometric figur e in which there are n+1 vertices, where (n) r epresents the number of variables [16], the method found a lot of app lications in field of analytical chemist ry [17,18], because it offers the cap ability of op timizing several factors simultaneously depending on a st atist ical design search to find the maxima or minima of resp onse, by rejecting the p oint p roducing the worst resp onse and a replacement of it by the new p oint which is obt ained statistically. The p resent work describes the utility of Bromophenol blue (BPB) reagent for sp ectrop hotometric determination of Dip henhydramine-HCL in p ure form as well as in the dosage form. In addition, the op timizations of chemical dependent variables of affectin g absorbance h ave been st udied by using modified simplex method (M SM ) via comp uter p rogram. Experime ntal Apparatus A Cintra 5 GBC.Scintific Equip ment sp ectrop hotometer with 1 cm quartz cells were used for absorbance measurements. p H-meter DW-9421 from Philips instrument, a Sartorius B L 210S balance, and a Pentium 4 computer (DELL) was used for data p rocessing. Material and Re agents All of the used Chemicals were of analy tical reagent grade unless otherwise is mentioned. Dip henhydramine - HCl was kindly p rovided by the State Company for Drug Indust ries and M edical Ap p liances, Samara-Iraq ( SDI). Bromophenol blue(BPB) (Aldrich), 0.1% (w/v) solution was p rep ared by dissolving 0.1 g of the dye in 5 ml of methanol and then the solution was diluted to a final volume of 100 ml with distilled water. Working solutions were freshly p repared by subsequent dilutions. Hy drochloric acid (Aldrich), ~ 0.1 M , a 0.85 ml of concentrated hydrochloric acid (sp .gr. 1.18, 37%) was added to 50 ml distilled water and diluted to t he mark in a 100 ml calibrated flask . Sodium hy droxide (Fluka), ~ 0.1 M , was p rep ared by dissolving 0.40 g of sodium hy droxide in 25 ml dist illed water and was diluted to 100 m l in volumetric flask with dist illed water. Pht halate buffer solution, 0.1M , was p rep ared by dissolving 4.08 g of p otassium hy drogen p hthalate (M ERCK) in 25 ml distilled water and was diluted to 200 ml in volumetric flask with dist illed water. The p H was adjust ed to 3.0 by using few drop s of 0.1M HCl and\or 0.1M NaOH {19]. Standard dru gs solution, Dip henhydramine st ock solution (250 µ g.ml -1 ), was p repared by dissolving 25 mg of the drug in 5ml distilled water and was diluted to 100 ml in a volumetric flask with dist illed water . Workin g solutions were freshly p rep ared by subsequent dilutions. IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL. 24 (1) 2011 General Procedure Assay procedure for pure Di phenhydramine 1 ml aliquots of Dip henhy dramine st andard solution containing (1-125 μg) wer e transferred into a series of 50 ml sep arating funnels. To each funnel 2 ml of p hthalate buffer of p H 3.0 and 1 ml of 0.05% BPB reagent solutions were added. The separating funn els were shaken with 5 ml chloroform for one minute. The two p hases were then allowed for clear sep aration and the absorbance of the yellow colored or ganic p hase was measured at 410 nm against a reagent blank p repared similar ly without addition of Dip henhy dramine. The calibr ation gr aph was constructed by p lotting the measured absorbance of the or ganic p hase against the drug concentration. Anal ysis of pharmaceuti cal formulation i. In table ts: Ten tablets were weighed and grounded into a f ine p owder. An amount equiv alent to 25mg of the cited dru g was weighed accurately and transferred into 25ml beak er and then was dissolved in 5ml of distilled water and was diluted to 100ml in a volumetric flask. The resulted solution was then filtered through a Whatman f ilter p aper No. 41 to avoid any suspended material b efore use. Workin g solutions were fr eshly p repared by subsequent dilution with dist illed water and analyzed by the recommend ed procedure. ii. In syrup : An accurately volume of the mixed Allermine sy rup equivalent to 100mg of dru g b ase was quantitatively transferred into a 100ml standard flask and diluted up to the mark with distilled water. An aliquot of the solution was transferred into a sep arating funnel and treated as described for tablets. Results and Discussion Extractive sp ectrop hotometric p rocedures are p op ular for their sensitivity in the assay of drugs and h ence, ion pair extractive sp ectrop hotometry has received considerable att ention for the quantitative determination of many p harmaceutical co mpounds [20, 21]. Prelimin ary invest igations revealed that DPH reacted with BPB in acidic buffer to y ield chloroform-soluble ion-pair comp lex exhibiting an absorp tion maximum at 410 nm against the reagent blank (Fig1). Under exp erimental conditions the corresp onding reagent blank showed a negligible absorbance. Optimization of experimental variable s:- Uni variable method: The exp erimental v ariables affecting the develop ment and st ability of the ion-pair comp lex were achiev ed through a numb er of preliminary exp eriments; these variables include p H, reaction time, reagent concentration, order of addition, shaking time and the ty p e of organic solvent used for extraction. For this reason, a variable was modif ied while maintainin g the other variables at their constant values, t hen by maintainin g that variable at its optimized values, another was modified; all var iable were op timized via this method. Effect of pH: In order to establish the optimum pH range, diphenhydramine-HCl solutions was mixed sep arately with sp ecified volume of BPB. T he pH was then adjusted to a valu e between (2.0- 5.5) with few drop s of or 0.1M NaOH or 0.1M HCl. It was noticed that maximu m color intensity and const ant absorbance was found at p H 3.0 (Fig 2). Low absorbance valu es were IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL. 24 (1) 2011 observed in solution with hi gher or lower p H t han the optimum value. Hence, a pH of 3.0 was used in all the subsequent e xp erimental work. Effect of reaction time: The optimum reaction time was determined by followin g the color develop ment at ambient temp erature (25 ± 2 °C). It was found that the reaction was instantaneous hence, the p roduct attained maximum and constant absorbance immediately after the diphenhy dramine- HCl has been reacted with BPB and the color obtained remained strictly unaltered for 24hr. Effect of reagent concentration: The influence of excess reagent concentration on the absorbance of the comp lex is illustrated in (Fig 3). 0.05% solution of BPB was found to develop the color and reach its maximu m intensity , after which no more increase in absorbance was obtained; therefore 0.05% solution was used. Effect of shaking time: The optimum shaking time for the comp lete extraction of the ion-pair comp lex with chloroform was st udied for the p eriod of 1-8 minutes (Table 1). It was found that the minimu m shaking time for comp lete extraction was 1 minute at room temp erature (i.e. 25 ±2°C). Effect of the extracting sol vent: Sever al or ganic solv ents, namely toluene, carbon tetrachlor ide, benzene, 1,2-dich loro ethane in addition to chloroform, were examined for their ability to extract the drug-dy e ion- p air. The latter was found to be the most suitable solvent in terms of extraction efficiency (Table 2). On the other hand, it was observed that only a single extraction with 5ml p ortion of chloroform was adequ ate to achieve a quantitative recovery of the complex. Effect of the order of addition: The effect of order of add ition of the reactants was also st udied. It was found t hat p utting reagent, buffer and finally the cited drug gave the best results instead of any other orders of addition S toichiometry of the complex To establish mole ratio between diphenhy dramine-HCL and BPB, Job’s method of continuous variation has been used (Fi g 4), the results showed that an ion-p air comp lex with a ratio of 1 :1 was formed between the dru g and BPB through an electrostatic att raction between the positive p rotonated dip henhydramine-HCl and the anion of BPB [22, 23] . The formation of the ion-p air comp lex can be represented as in following sch eme: IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL. 24 (1) 2011 S O O HO OH O Br Br Br Br +HCl Diphenhydramine.HCl Bromophenol Blue Ion-pair Complex + HCl O N O N H S O O HO O Br Br Br Br o Si mplex me thod Simp lex method used to confirm the op timum conditions which were obtain ed by the univariate p rocedure. Three major p arameters (p H, reagent concentration, and shaking time) were optimized by the simp lex p rocedure, while t he other minor p arameters were obtained by the univaiate method. To set simplex p rogram for the three st udied v ariables, four arbitrary exp erimental conditions should be chosen. The values of these p arameters were selected within sp ecified boundaries for each at which they affected the measured absorp tion signal of colored products. The absorbencies of these four exp eriments were measured and the results were feed to the simp lex p rogram. Points (1 t o 4) in (Table 4) represent t he first four exp eriment cycle with their measured absorben cies. The simp lex p rogram starts to reflect the worst p oint through the centroid of other p oint to obtain a new p oint 5. An exp eriment was t hen p erformed utilizing the variable setting as a reflected p oint 4, the latter was rejected and replaceed by p oint 5. A measured absorp tion signal was feed again to the p rogram and the p rocess is repeated successively until op timum conditions were decided by the program and were similar to those obtained by the univariate method. Calibration graph Emp loying the exp erimental cond itions, linear calibration graph for diphenhydramine is obtained (fig 5), which shows t hat Beer’s law was obey ed in the concentration range of 0.2- 25.0 µg.ml -1 S pectral characteristics of the propose d method Accordin g to the optimum exp erimental conditions of t he prop osed method, the regression p lot shows a linear dependence of absorbance sign als on the concentrations of the studied drug in the range given in Table 4. The regression equation, correlation coefficient, molar IBN AL- HAITHAM J. FO R PURE & APPL. SCI. VO L.24 (1) 2011 absorptivity, detection limit and Sandell's sensitivity in addition to other parameter are given in (Table 5). Accura cy and precisi on The accuracy of the prop osed method was confirmed by analyzing three replicate an aly ses of three different amounts of drug (with Beer’s law) by calculating the relative error p ercentage (Table 6). The results indicated good accuracy of the method for Dip henhy dramine-HCl. The p recision was d etermined in each case by calcu lating the p ercentage relative standard deviation (RSD %) for t hree determinations at each of the studied concentration level and were found to be in the ran ge 1.067 – 1.652 %. Inte rference studies The effect of common excip ients that often accomp any the studied drug in v arious p harmaceutical tablets were tested for p ossible interference in the assay . The results showed that no interference were found in the p resence of 500 µ g of the st udied excip ients (st arch, Lactose, sucrose, magn esium st earat and sodiu m alginate) in the determin ation of diphenhydramine-HCl. Anal ysis of dosage forms The obtained satisfactory validation results made the p rop osed p rocedures suitable for the routine quality control analysis of diphenhy dramine in commercial tablets. The results, p resented in (Table 7), reveal that the values obtained for RSD% and R. E.% , indicate high accuracy and precision for the p rop osed method. Re ferences 1.Reynohds, JEF. (1993). “M artindale the Extra pharmacop oeia”, 30 the ed., p harmaceutical p ress, London, 937. 2.Basavaiah, K. and Ch aran, IL, V. S.( 2002). Titrimetric and sp ectrop hotometric assay of some antihist amines t hrough the determination of the chloride of their hydrochlorides Farmaco, 57:9. 3.Wong, C. ; Fan, G.; Lin, M .; Chen, Y. ; Zhao,W. and Wu,Y. (2007). Development and validation of a liquid chromato graphy /tandem mass sp ectrometry assay for the simultaneous determination of D-amp hetamine and dip henhydramine in bea gle do g p lasma and its application to a p harmacokinetic st udy ”. J. Chramatogr. B, 1 (1-2):854- 848. 4.Lou, H.; Zhuo, K . and Chen, J . (2004). Selective Determination of Diphenhydramine in Compound Pharmaceutical Contain ing Ephedrine by Flow-Injection Electrochemilu minescence” Selective Determination of Dip henhydramine in Comp ound Pharmaceutical Containin g Ephedrine by Flow-Injection Electrochemiluminescence” China. p harmacist, 7: 530. 5.Ahrens, B .; Blankenhorn, D .and Sp angenberg, B. ( 2002) Advanced f ibre op tical scannin g in thin-lay er chromatogr aphy for drug identification. J. Chromatogr. , 772, 11. 6.Dong, Y. ; Chen, X. ; Chen, Y. and Hu. Z.,(2005). Seperation and determination of Pseudoep hedrine, dextromethorp han, dip henhydramine and chlorpheniramine in cold medicines by nonaqueous capillary electrop horesis. J- Pharm. Biomed Anal., 39, 285. IBN AL- HAITHAM J. FO R PURE & APPL. SCI. VO L.24 (1) 2011 7.Raj, S. V. ; Kapadia, S. U. and Argekat, A. P. , (1998). Simultaneous determination of pseudoephedrine hydrochloride and diphenhydramine hydrochloride in cough syrup by gas chromatography (GC), Talanta.46:221. 8.Daneshgar, P. ; Norouzi, P.; Ganja, M . and Doust y , F. (2009) A Dy sp rosium Nanowire M odified Carbon Past e Electrode for Determination of Nanomp lar Level of Dip henhydramin by Continuous Square Wave Voltammetry in Flow Injection Sy st em . J. Int Electro hem. Sci, 4: 444. 9.San Andres, M . P. ; Sicilia, D. ; Rubio, S. and Perez ,D. – Bendito. (1998), Determination of antihistamines based on the formation of mixed aggregates with surfactants". Analy st , 123:1079. 10.EL– Shahat, M . F. ; Abdel Bada, M . M . and Daifullah, A. (1992) J. Chem. Technol. Biotechnol, Sp ectrop hotometric determination of ephedrine HCl, cinchonine HCl, chlorp heniramine maleate, atrop ine sulp hate and dip henhydramine HCl by solvent extraction of reineckate comp lexes. 24:175. 11.Tipp arat, P. ; Lap anantnop p akhum, S. ; Jakmunce, J. and Grudp an, K. (2002) Sp ectrop hotometric determination of diphenhydramine hy drochloride in pharmaceutical p reparations and biological fluids via ion-pair formation J. Pharm. Biomed. Anal. 30:105. 12.M artinez Calatayud, J. ; Sanchez sanpedro, A. and Navasquillo Sarrion. S., (1990) Determination of dip henhydramine hy drochloride by flow injection with Bromop henol Blue and turbidimetric measurement. Analyst , 115: 855. 13.sp endely, W.;Hext,G.R. and Himuswort h, F.R. ,( 1962). Sequential Ap p lication of Simp lex Designs in Op timisation and Evolutionary Op eration. Technometrics, 4: 441-462. 14.Nelder,J. A. and M ead, R. A.,(1965). A Simp lex M ethod for Function M inimization. Computer Journal, 7: 308-313. 15. Albery ,E. R. and Gustavsson ,A. G.(1982) Design and Evaluation of M odified Simp lex Analytica Chemica Acta. Vol. 144: 39-53. 16.Walters, F. H. ; Parker, L. R. ; M organ, S. L. and Deming, S. N. ( 1991). “ Sequentiol Simp lex Op timization”, CRC Press, Florida, 44. 17.M omenbeik, F.; M omeniz Z . and Kharasani, J. H. (2005). Sep aration and determination of Vitamins E and A in multivitamin sy rup using micellar liquid chromatography and simplex op timization Journal of Pharmaceutical and Biomedical AnalysisVol, 37( 2): 383-387. 18.Pulgarin, J. A.; M olina, A . and A lanon Pardo, M . T. ( 2002). ”The use of modified Simp lex M ethod t o Op timise the room temperature Phosp horescence Variables in the Determination of Antihyp ertensive Drug” ,Talanta, 57:795-805. 19.Purachat, B.; Liawruangrath, S.; Sooksamiti, P. and Buddhasukh, D. (2001). Univariable and Simp lex Op timization for the Flow-injection Sp ectrop hotometric Determination of Cop er Using Nitro So-R Salt as Complexing Agent, Analy tical Sciences, 17: 443-447. 20.Basavaiah, K. and Shakunthala, V.(2004). Ion-p air Complexometric Determination of Cy p roheptadine Hy drochloride Using Bromophenol Blue Charan. Science Asia 30:163-170 21.Julic, M . ; Cardso, S. C. (2005). J. Pharm. Biomed. Anal, 37: 639. 22.Basavaiah, K. ; Handrashear, H. C. ; Chikkaswamy , B.,( 2007). Sp ectrop hotometric determination of p efloxacin mesy late in p harmaceuticals Acta. Pharm. 57:221-230. 23. Elham Taha, A. ; Susan Soliman, M .; Hisha Abdellatef, E. and M agda Ay ad M .,(2002) Colorimetric method for the determination of some tricyclic antidepressant drugs in their p ure and dosage. M icrochim. Acta. 140: 175 IBN AL- HAITHAM J. FO R PURE & APPL. SCI. VO L.24 (1) 2011 Table (1): Effect of shaking time on e xtraction of 5µg.ml -1 Diphe nhydramine , 0.05% BPB, pH 3.0. Time (min.) Absorbance 1 0.353 2 0.252 3 0.255 4 0.268 5 0.272 6 0.281 7 0.260 8 0.242 Table (2): The effect of the extraction solvent of 5µg.ml -1 Diphe nhydramine Organic p hase Absorbance of Drug-BPB ion p air comp lex Chloroform 0.353 1,2-Dichloroethane 0.272 Benzene 0.288 Dichloromethane 0.313 Toluene 0.097 Table (3): Boundary conditi ons for the studied variable . Variable Range p H 2.00 - 5.50 Reagent Conc. (%) 0.01 - 0.07 Shakin g time (min.) 1.00 - 8.00 IBN AL- HAITHAM J. FO R PURE & APPL. SCI. VO L.24 (1) 2011 Table (4): Multivariate experiments (si mplex) for determination of Diphenhydramine Table (5): Anal ytical characteristics for the propose d. Parameter BPB p rop osed method Linear r ange (µg.ml -1 ) 0.2 – 25.0 Regr ession equation A = 0.0828[ Dip h. µg.ml -1 ] + 2.28x10 -6 Slope (L. mg -1 .cm -1 ) 0.0828 Intercept 2.28 x 10 -6 M olar absorp tivity (L.mol -1 .cm -1 ) 2.416x10 4 Correlation Coefficient 0.9991 Detection Limit (µg. ml -1 ) 0.155 Sandell ' s Sensitivity (µg.cm -2 ) 0.0128 Exp . No. p H Reagent Conc.(%) M ixing Time(min.) Abs. 1 2.5 0.04 2 0.3057 2 2.7 0.05 3 0.3071 3 3.0 0.05 1 0.3535 4 2.5 0.07 6 0.2366 5 3.0 0.03 1 0.2557 6 2.5 0.06 4 0.3280 7 3.0 0.06 2 0..2856 8 2.5 0.05 2 0.2299 9 3.0 0.05 1 0.1935 10 2.5 0.05 2 0.2299 11 3.0 0.04 1 0.2241 12 2.5 0.05 3 0.2484 13 3.5 0.05 1 0.2290 14 2.0 0.06 6 0.1787 15 3.0 0.05 2 0.2008 IBN AL- HAITHAM J . FO R PURE & APPL. SCI. VOL.24 (1) 2011 Table (6): Evalu ati on of accu racy an d pre cision of the propose d me thod Concentration of drug taken (µg.ml -1 ) Concentration of drug found (µg.ml -1 ) Relative Error,% RSD , % 5 4.92 -1.60 1.067 15 15.06 +0.40 1.652 25 25.12 +0.48 1.068 Table(7):S pectrophotome tric determination of Diphenhydramine in pharmaceutical compounds using Ion-pair formation Amount of drug (µg) Concentration (µg.ml -1 ) R. E.% RSD% Labeled Found Taken Found 25 24.48 10 9.80 -2.00 0.661 20 19.62 -1.90 0.721 10 10.04 10 10.05 +0.50 0.409 20 20.09 +0.45 0.387 Fig.(1): Absor ption spectrum of 4µg.ml -1 Di phenhydramine-BPB ion-pair complex IBN AL- HAITHAM J. FO R PURE & APPL. SCI. VO L.24 (1) 2011 Fig.(2): Effect of pH on the absorbance of 5µg.ml -1 Di phenylhydramine,0.05% BPB. Fig.( 3): Effect of reagent concentration on the absorbance of 5µg.ml -1 Di phenhydramine at pH 3.0. Fig.(4): Job's me thods of 3.4270 x10 -5 M (10 µg.ml -1 ) Di phenhydramine with 1.492*10 -3 M BPB. Fig.(5): Calibration graph of Diphenhydramine under optimum experimental conditions. 2011) 1( 24المجلد مجلة ابن الھیثم للعلوم الصرفة والتطبیقیة دراسة الیجاد الظروف المثلى بطریقتي المتغیرات االحادیة و السمبلكس واستخدامها لتحلیل ثنائي فینهیدرامین هیدروكلورید خالل تكوین زوج ایوني 2010 یارآ 4 : استلم البحث في 2010حزیران 3 :قبل البحث في عالء كریم محمد، سرمد بهجت دیكران، مها عبدالستار العبیدي قسم الكیمیاء، كلیة التربیة ابن الهیثم، جامعة بغداد الخالصة –تم في هذا البث وصف طریقة استخالص وتقدیر طیفیة سهلة وسریعة وحساسة لتحلیل ثنائي فنهایدرامین ایوني مع كاشف -تعتمد الطریقة على تكوین معقد زوج. ض المستحضرات الصیدالنیةعهیدروكلورید النقي وفي ب مستخلص . قابل للذوبان في الكلوروفورم) pH = 3.0(الفثاالت دارئ بروموفینول االزرق في وسط لقد اظهر المعقد ال 25.0 -0.2(اوح بین نانومتر وكان مطاوعا لقانون بیر لمدى خطي یتر 410اعظم امتصاص عند طول موجي مقداره µg.ml -1 cm(وكانت قیم معامل االمتصاص المولي وحساسیة ساندل لهذا النظام هي) -1 2.416x10 4 L.mo l -1 و ) . )0.012 µg.cm -2 µg.ml 0.155(على التوالي، بینما كان حد الكشف مساویا الى ) -1 لقد امكن تطبیق هذه الطریقة ). .تم الحصول على نتائج مرضیة اذالصیدالنیة، بنجاح لتقدیر الدواء في بعض المستحضرات ثنائي فینهیدرامین هیدروكلورید، السمبلكس،المتغیرات األحادیة :الكلمات المفتاحیة