Spectrophotometric Methods Based on Charge Transfer Complexation Reactions for the Determination of Saxagliptin in Bulk and Pharmaceutical Preparation

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1 Interntionl journl of Biomedicl science ORIGINAL ARTICLE Spectrophotometric Methods Bsed on Chrge Trnsfer Complextion Rections for the Determintion of Sxgliptin in Bulk nd Phrmceuticl Preprtion Rmzi I. El-Bgry, Eh F. Elkdy, Bssm M. Ayou Deprtment of Phrmceuticl Chemistry, Fculty of Phrmcy, Ciro University, Ksr El-Aini St., Ciro 11562, Egypt Astrct Simple, ccurte nd precise spectrophotometric methods hve een developed for the determintion of sxgliptin in ulk nd dosge forms. The proposed methods re sed on the chrge trnsfer complexes of sxgliptin with 2,3-dichloro-5,6-dicyno-1,4-enzoquinone (DDQ) nd 7,7,8,8-tetrcynoquinodimethne (TCNQ). All the vriles were studied to optimize the rections conditions. Beer s lw ws oeyed in the concentrtion rnges of μg/ml nd μg/ml with DDQ nd TCNQ, respectively. The developed methods were vlidted nd proved to e precise nd ccurte for the qulity control of the sxgliptin in its phrmceuticl dosge form. (Int J Biomed Sci 2012; 8 (3): ) Keywords: sxgliptin; spectrophotometry; chrge trnsfer rections; phrmceuticl preprtion INTRODUCTION Sxgliptin (SXG), (1S, 3S, 5S)-2-[(2S)-2-mino-2-(3- hydroxy-1-dmntyl) cetyl]-2-zicyclo [3.1.0] hexne- 3-cronitrile (Fig. 1) is new orl hypoglycemic drug of the new dipeptidyl peptidse-4 (DPP-4) inhiitor clss of drugs (1). Sxgliptin is recently pproved for the tretment of type 2 dietes mellitus (2). DPP-4 inhiitors represent new therpeutic pproch to the tretment of type 2 dietes tht functions to stimulte glucose-dependent insulin relese nd reduce glucgons levels. This is done through inhiition of the inctivtion of incretins, prticulrly glucgon-like peptide-1 (GLP-1) nd gstric inhiitory polypeptide (GIP), therey improving glycemic control (3). Literture survey revels tht the drug hs een estimted y only one LC-MS/MS (4) nd nother spectro- H 2 N O N Corresponding uthor: Bssm M. Ayou, Deprtment of Phrmceuticl Chemistry, Fculty of Phrmcy, Ciro University, Ksr El-Aini St., Ciro 11562, Egypt. Tel: ; fx: ; E-mil: drssmchemistry@hotmil.com. Received July 5, 2012; Accepted July 23, 2012 Copyright: 2012 Rmzi I. El-Bgry et l. This is n open-ccess rticle distriuted under the terms of the Cretive Commons Attriution License ( which permits unrestricted use, distriution, nd reproduction in ny medium, provided the originl uthor nd source re credited. N HO H H Figure 1. Chemicl structure of sxgliptin. 204

2 photometric method in which sxgliptin ws estimted t 208 nm in methnol. Linerity rnge ws found to e 5-40 μg/ml. The LOD nd LOQ were found to e 0.06 μg/ml nd 0.18 μg/ml respectively (5). The im of the work is to present the first chrge trnsfer complextion methods for the determintion of SXG in ulk nd phrmceuticl formultions. Chrge trnsfer rections hve een widely used for the determintion of electron donting compounds through interction with π-cceptors. Among the electron cceptors mostly used in literture re 2,3- dichloro-5,6- dicyno-1,4- enzoquinone (DDQ) (6-9) nd 7,7,8,8-tetrcynoquinodimethne (TCNQ) (10-13). EXPERIMENTAL Instrumenttion A Jenwy 6800 doule em ultrviolet/visile spectrometer connected to n IBM comptile computer with 1-cm qurtz cell nd supported with Jenwy flight deck softwre ws used. Regents nd reference smples DDQ nd TCNQ were supplied from Sigm Aldrich Chemie GmH (Steinheim, Germny). Freshly prepred solutions were prepred (DDQ solution M in methnol), (TCNQ solution M in methnol). Phrmceuticl grde SXG certified to contin 99.85% nd Onglyz tlets nominlly contining 5 mg of SXG per tlet were kindly supplied y Bristol-Myers Squi/ AstrZenec EEIG (United Kingdom). Stndrd stock solutions of SXG (1 mg/ml) ws prepred y dissolving 100 mg of SXG in methnol in 100 ml volumetric flsk nd completing to volume with methnol. All the solvents used were of nlyticl grde. Generl procedures nd clirtion grphs Method using DDQ. Aliquots of SXG contining (0.5-3 mg) were trnsferred into set of 10 ml volumetric flsks, treted with 1 ml DDQ solution nd llowed to stnd for 40 min t room temperture (20-25 C) nd diluted to volume with methnol. The sornce ws mesured t 461 nm ginst regent lnk. Method using TCNQ. Aliquots of SXG contining ( mg) were trnsferred into set of 10 ml volumetric flsks, treted with 1 ml TCNQ solution nd llowed to stnd for 30 min t room temperture (20-25 C) nd diluted to volume with methnol. The sornce ws mesured t 838 nm ginst regent lnk. Procedure for the ssy of the tlets Twenty tlets of were weighed nd the cots were removed y crefully ruing with clen tissue wetted with using methnol. An ccurtely weighed mount of the finely powdered tlets equivlent to 100 mg ws dissolved in methnol in 100 ml volumetric flsk, sonicted for 30 minutes nd completed to volume with methnol. The solution ws then filtered, followed y seril dilution to required concentrtions. The procedure ws continued s mentioned under generl procedures nd clirtion grphs. Effect of the mount of the regent Aliquots of SXG ( M) stock solutions were introduced into set of 10 ml volumetric flsks. Different liquots of (DDQ nd TCNQ) were dded to ech flsk to otin different drug/ regent molr rtios in n incresing order, nd then the procedure ws continued s mentioned under generl procedures nd clirtion grphs. Stoichiometric reltionship Jo s method of continuous vrition ws employed, etween stndrd solutions of M of SXG) with the two regents (DDQ with concentrtion M nd TCNQ with concentrtion M). A series of solutions ws prepred in which the totl volume of the drug nd the regent ws kept t 5 ml. The method ws continued s mentioned under the generl procedures for the clirtion grphs. RESULTS AND DISCUSSION Formtion of the chrge trnsfer complexes The chrge trnsfer regents pplied in this work re DDQ nd TCNQ. DDQ is n electron deficient molecule due to the electron withdrwing effect of the two cyno nd the two chloro groups. The rection etween DDQ s π-cceptor with severl drugs (6-9) to give complex which dissocites in polr solvents to highly colored rdicl nion hs een reported. Likewise, TCNQ is well known π-cceptor (10-13); this chrcter is derived from the high electron ffinity of polyene system conferred y the electron withdrwing effect of the four cyno groups nd lso from the plnrity nd high symmetry of TCNQ. Optimiztion of the rections conditions Different prmeters ffecting the rections such s mount of the regent, rection time nd stility of the color hve een investigted. λ mx of mesurements of SXG Int J Biomed Sci vol. 8 no. 3 Septemer

3 with the two regents re shown in Tle 1. The sorption spectr of the rection products of SXG with the two regents re shown in Figures 2-3. Rte of complex formtion. A study of the effect of time reveled tht the mximum color intensity ws ttined t lest fter 30 min with DDQ method nd t lest fter 20 min with TCNQ method s shown in Figure 4. Stility of the formed complex. A study of the stility of the formed complex reveled tht the rection product remined stle for further 50 min with DDQ method nd for further 60 min with TCNQ method s shown in Figure c nm Figure 2. Asorption spectr of the colored product of sxgliptin ( M) nd DDQ ( M) in methnol (), sxgliptin ( M) in methnol () nd DDQ ( M) in methnol (c) c 0.5 nm Figure 3. Asorption spectr of the colored product of TCNQ ( M) with sxgliptin ( M) in methnol (), sxgliptin ( M) in methnol (), TCNQ ( M) in methnol (c). Tle 1. Results otined y the proposed methods for the determintion of sxgliptin using DDQ nd TCNQ Item DDQ method TCNQ method Solvent Methnol Methnol Time of rection 40 minutes 30 minutes Stility of the color 50 minutes 60 minutes λ mx of mesurements 461 nm 838 nm Oedience of Beer s lw μg/ml μg/ml Regression eqution A 486 = Conc (μg/ml) A 838 = Conc.(μg/ml) Regression coefficient (r 2 ) LOD μg/ml LOQ μg/ml S S Confidence limit of the slope ± ± Confidence limit of the intercept ± ± Stndrd error of the estimtion Results Drug in ulk ± ± 1.32 Drug in dosge form ± ± 1.13 Drug dded ± ±

4 Effect of the mount of regent. Study of the mount of the regent reveled tht the mount of the regent ws optimized to e four times the mount of SXG in the chrge trnsfer rection with oth DDQ nd TCNQ s shown in Figure 6. Effect of temperture. Study of the effect of temperture reveled tht the rection ws optimized t room temperture in the chrge trnsfer rection with oth DDQ nd TCNQ. Stoichiometric reltionship Jo s method of continuous vrition reveled molr rtio of of (1:1) drug: regent ws otined with the two regents s shown in Figure 7. This my e ttriuted to the vilility of one center s n electron donting group which is the primry mine while the mide nitrogen nd cyno nitrogen re not electron donors. Quntifiction, ccurcy nd precision Stndrd clirtion curves were prepred y seprtely prepring series of different concentrtions of SXG nd pplying the suggested procedures with DDQ nd TCNQ. Beer s lw ws vlid within microgrm concentrtion rnge of SXG (Tle 1). The linerity of the clirtion curves were vlidted y the high vlue of correltion coefficients. The nlyticl dt of the clirtion curves including stndrd devitions for the slope nd intercept (S, S ) re summrized in Tle 1. The regression equtions of these clirtion grphs were utilized for determintion of concentrtions of the cited drug in ulk nd tlets. The results otined were of good ccurcy nd precision. The pplicility of the procedures for estimtion of tlets ws vlidted using stndrd ddition technique s check for ccurcy (Tle 1). Figure 6. Effect of the mount of the regent in the determintion of sxgliptin with DDQ () nd TCNQ (). Figure 4. Effect of the rection time in the determintion of sxgliptin with DDQ () nd TCNQ (). Figure 5. Effect of stility of the formed chrge trnsfer complex in the determintion of sxgliptin with DDQ () nd TCNQ (). Figure 7. Continuous vrition plot of the rection products of sxgliptin with DDQ ( ) nd TCNQ (----). Int J Biomed Sci vol. 8 no. 3 Septemer

5 Tle 2. Sttisticl comprison etween the results of the proposed spectrophotometric methods nd the reference method for the determintion of sxgliptin Sttisticl Term Reference Method DDQ Method TCNQ Method Men S.D.± S.E. ± %RSD n V t ( 2.306) Figures in prentheses re the theoreticl t nd F vlues t (p=0.05). No significnt difference etween groups of sitgliptin y using one wy ANOVA with F equls 0.10 nd p equls 0.91; Reference method: liquots of stndrd solutions in distilled wter contining 5 40 μg/ml were mesured t 208 nm using wter s lnk (4). Sttisticl nlysis A sttisticl nlysis of the results otined y the proposed methods for the determintion of SXG ws crried out y SPSS sttisticl pckge version 11. The significnt difference etween groups including the reference method were tested y one wy ANOVA (F-test) t p=0.05 s shown in Tle 2. The test scertined tht there ws no significnt difference mong the methods. CONCLUSION The proposed spectrophotometric methods hve the dvntges of simplicity, precision, ccurcy nd convenience for the quntittion of SXG. Hence, the proposed methods cn e used for the qulity control of the cited drug nd cn e extended for routine nlysis of SXG in ulk nd dosge forms. REFERENCES 1. Augeri D, Rol J, Beteenner D, et l. J. Med. Chem. 2005; 48: Kuls K, Edelmn S. Core Evidence. 2010; 5: Nirogi R, Kndikere V, Mudigond K, et l. Biomed. Chromtogr. 2008; 22: Fur A, Khnn A, Vys V, et l. Drug Met. Dispos. 209; 37: Klichelvi R, Jychndrn E. Int. J. Phrm. Phrm. Sci. 2011; 3: Nour E, Alqrdwi S, Mostf A, et l. J. Mol. Struct. 2010; 980: Fkhroo A, Bzzi H, Mostf A, et l. Spectrochim Act. A Mol. Biomol. Spectrosc. 2010; 75: Shhd L, Mostf A, Nour E, et l. J. Mol. Struct. 2009; 933: Zyed M, Khlil S, El-qudy H. Spectrochim Act. A Mol. Biomol. Spectrosc. 2005; 62: Mostf A, Bzzi H. J. Mol Struct. 2010; 983: Mohmed G, El-Dien F, Mohmed N. Spectrochim Act. A Mol. Biomol. Spectrosc. 207; 68: El-Dien F, Mohmed G, Frg E. Spectrochim Act. A Mol. Biomol. Spectrosc. 2006; 64: Adel-Hmid M, Auirjeie M. Tlnt. 1988; 35: