Ion Association Methods Development for the Assay of Venlafaxine Hydrochloride Using Two Acidic Dyes
K. Raghubabu1*, L. Santhi Swarup1, B. Kalyana Ramu2, G. Rupakumari, M. Narayanarao3 and C. Ramdas3
1Department of Engineering Chemistry, AU College of Engineering (A), Andhra University, Visakhapatnam -530003 Andhra Pradesh (India)
2Department of Chemistry, Maharajah’s College (Aided and Autonomous), Vizianagaram-535002 (AP) India.
3Tychy Industries, R and D Division, Hyderabad-AP (India)
Corresponding author: drraghualways@yahoo.co.in
ABSTRACT:
Two simple, rapid and sensitive extractive visible spectrophotometric methods (A and B) for the assay of venlafaxine hydrochloride in pure and solid dosage forms based on the formation of colored chloroform soluble ion - association associates under specified experimental conditions are described. Two dyes namely acidic dyes Naphthalene Blue 12BR (NB12BR, method A) and Bromo Cresol Purple (BCP, method B) are utilized. The extracts of the ion-associates exhibit absorption maxima at 620.5 nm and 430 nm for methods A and B respectively. Regression analysis of Beer-Lambert plots showed good correlation in the concentration ranges (4.0 -12.0) µg/ml for method A, (8.0-40.0) µg/ml for method B respectively. The proposed methods are applied to commercial available tablets and the results are statistically compared with those obtained by the UV reference method and validated by recovery studies. The results are found satisfactory and reproducible. These methods are applied successfully for the estimation of the venlafaxine hydrochloride in the presence of other ingredients that are usually present in dosage forms. These methods offer the advantages of rapidity, simplicity and sensitivity and normal cost and can be easily applied to resource-poor settings without the need for expensive instrumentation and reagents.
KEYWORDS: Anti-depressant, Extraction methods, NB12BR, BCP, Statistical analysis, Tablets.
Venlafaxine hydrochloride (VX) is a bicyclic third generation antidepressant chiral compound of phenethyl amine type with a novel chemical structure 1, 2 (Fig.1).
It is designated (R/S)-1-[2-(dimethyl amino)-1-(4-methoxy phenyl) ethyl] cyclohexanol hydrochloride salt and is usually categorized as a serotonin-nor epinephrine reuptake inhibitor (SNRI) but it has been referred to as a serotonin-norepinephrine–dopamine reuptake inhibitor. It weakly inhibits the reuptake of dopamine. VX is well absorbed, with peak plasma concentrations occurring approximately 2 hours after dosing. It is extensively metabolized, to O-desmethyl venlafaxine, the only major active metabolite. It has the empirical formula of C17H27NO2.HCl. Its molecular weight is 313.87. The drug is official in BP 3 and suggests potentiometric method for the determination of VX in bulk and tablet formulations.
Fig. 1: Chemical structure of Venlafaxine hydrochloride
Various methods have been reported for the estimation of venlafaxine hydrochloride in biological matrices such as plasma, which include the use of liquid chromatography (LC) with UV detection4, LC with electro spray ionization mass spectrometry5, LC with coulometric detection6, LC with fluorimetric detection7,8, LC with diode array detection9,10, gas chromatography-mass spectrometry (GC–MS) 11, LC-MS12, LC–MS–MS13,14, and for the estimation in serum by using LC15, HPLC methods16-22, HPLC-MS/ESI 23, voltammetry24,Capillary Electrophoresis25,26, Flow injection analysis27. Stability indicating methods have also been reported for its in vitro determination in gastric and intestinal fluids 28 and pharmaceutical formulations 29, few UV 30-33 and Visible Spectrophotometry 34-38. However analytical important functional groups in VX have not been exploited properly in developing visible spectrophotometric methods and most of the previous methods involve sophisticated equipments which are costly and pose problems of maintenance.
As the extraction spectrophotometric procedures are popular for their sensitivity and selectivity in the assay of drugs, the extractive spectrophotometric technique 39 was therefore, utilized in the present work for the estimation of VX. The present paper describes two simple and sensitive extraction visible spectrophotometric methods for the determination of VX, based on its tendency to form chloroform extractable ion-associates with acidic dyes NB12BR (CI No.20500) belonging to azo category dye (method A) and BCP belonging to tri aryl methane category dye (CB No.1347717) (method B) under experimental conditions by exploiting the basic nature (tertiary nitrogen) of the drug molecule.
MATERIALS AND METHODS:
Apparatus and Chemicals:
A Systronics UV/Visible spectrophotometer model-2203 with 10mm matched quartz cells was used for all spectral measurements. Systronics model-362 pH meter was used for all the pH measurements. A pure drug sample of VX was provided as a gift sample by Tychy industries, Hyderabad AP (India). Capsules and tablets purchased from local market. All the chemicals used were of analytical grade. Aqueous solution of NB12BR (BDH,0.2%, 3.14x10-3M prepared by dissolving 200mg of NB12BR in 100ml distilled water and subsequently washed with chloroform to remove chloroform soluble impurities), pH 1.5 buffer solution (prepared mixing 289ml of 0.1M glycine solution with 711ml of 0.1M HCl and the pH of the solution was adjusted to 1.5), Aqueous solution of BCP (Loba, 0.1%, 1.433x10-3M prepared by dissolving 100mg of BCP in 100ml distilled water and subsequently washed with chloroform to remove chloroform soluble impurities), pH 3.5 buffer solution (prepared mixing 50ml of 0.2M Potassium hydrogen phthalate, 15.7 ml of 0.1M HCl and 134.3 ml of distilled water and the pH of the solution was adjusted to 3.5) were prepared.
Preparation of Standard stock solution:
The standard stock solution (1mg/ml) of VX was prepared by dissolving 100mg of VX initially in 10ml of 0.1M sodium hydroxide and followed by dilution to 100 ml with distilled water. The working standard solution of VX (100µg/ml- method A and 200µg/ml –method B) was obtained by appropriately diluting the standard stock solution with the same solvent.
Sample solution:
About 10 tablets or capsules were pulverized and the powder equivalent to 100mg of VX was weighed, dispersed in 25ml of IPA, sonicated for 15 minutes and filtered through Whatman filter paper No 41.The filtrate was evaporated to dryness and the residue was dissolved as under standard solution preparations.
Assay: Aliquots of the standard VX solution [1.0-3.0ml, 100µg/ml (method A) and 1.0-5.0ml, 200µg/ml (method B)] were placed in a series 125ml separating funnels. Then 5.0ml of pH 1.5 buffer and 2.0ml of 3.17x10-3M NB12BR solution (method A) or 6.0ml of pH3.5buffer solution and 2.0ml of 1.433x10-3M BCP solution (method B) were added. The total volume of aqueous phase in each separating funnel was adjusted to 15.0ml (method A) or 10.0ml (method B) with distilled water. Then 10.0ml (method A) or 15.0ml (method B) of chloroform was added to each separating funnel and the contents were shaken for 2 minutes. The two phases were allowed to separate. The absorbance of the separated chloroform layer were measured at 620.5 nm (method A) or 430 nm (method B) (Fig. 2and4 showing absorption spectra) against a reagent blank within the stability period (5 minutes to 1hour). The amount of drug was computed from its calibration graph (Fig 3and5 Beer’s law plot).
Fig.2: Absorption spectra of VX-NB12BR
Fig.3: Beer’s law plot of VX-NB12BR
Fig.4: Absorption spectra of VX-BCP
Fig.5: Beer’s Law plot of VX-BCP
RESULTS AND DISCUSSIONS:
Optimum operating conditions used in the procedure were established by adopting variation of one variable at a time (OVAT) method. The effect of various parameters such as time, volume and strength of NB12BR, BCP reagents, pH buffer solutions and solvent for final dilution of the colored species were studied. The water immiscible solvents tested for the extraction of colored complex into organic phase include chloro benzene, dichloromethane, carbon tetra chloride, benzene, n-butanol or chloroform. Chloroform was preferred for its selective extraction of colored drug -dye complex into organic layer from the aqueous phase. The stoichiometric ratio of the dye-drug was determined by the slope ratio method and was found to be 1:2 for methods (A and B). The optical characteristics such as Beer’s law limit, Sandell‘s sensitivity, molar absorptivity, percent relative standard deviation, (calculated from the six measurements containing 3/4th of the amount of the upper Beer’s law limits). Regression characteristics like standard deviation of slope (Sb), standard deviation of intercept (Sa), standard error of estimation (Se) and % range of error (0.05 and 0.01 confidence limits) were calculated and the results are summarized in Table-1.
Table - 1 Optical characteristics, precision and accuracy of the proposed methods
|
Parameters |
Method A |
Method B |
|
λmax(nm) |
620.5 |
430 |
|
Beer’s law limit(µg/ml) |
4-12 |
8-40 |
|
Sandell’s sensitivity (µg/cm2/0.001 abs. unit) |
0.000911681 |
0.003047619 |
|
Molar absorptivity (Litre/mole/cm) |
344276.1563 |
102988.5938 |
|
Regression equation (Y)*= a +b x |
|
|
|
Intercept (a) |
-0.111 |
-0.005 |
|
Slope(b) |
0.058 |
0.013 |
|
%RSD |
1.797 |
1.419 |
|
% Range of errors(95% Confidence limits) 0.05 significance level 0.01 significance level |
1.886 |
1.489 |
|
2.958 |
2.336 |
*Y = a + b x, where Y is the absorbance and x is the concentration of VX in µg/ml
Commercial formulations containing VX were successfully analyzed by the proposed methods. The values obtained by the proposed and reference methods for formulations were compared statistically by the t-and f-test and found not to differ significantly. As an additional demonstration of accuracy, recovery experiments were performed by adding a fixed amount of the drug to the pre analyzed formulations at three different concentration levels. These results are summarized in Table-2.
Table-2 Analysis of VX in pharmaceutical formulations
|
Met hod |
*Formu lations |
Labeled Amount (mg) |
Found by Proposed Methods |
Found by Reference Method ± SD |
#% Recovery by Proposed Method ± SD |
||
|
**Amount found ± SD |
t |
F |
|||||
|
A |
Tablet-1 |
37.5 |
36.99 ± 0.548 |
1.28 |
2.50 |
36.13±0.87 |
98.62 ± 1.498 |
|
Tablet-2 |
75 |
72.29 ± 0.951 |
1.49 |
4.43 |
71.85±0.45 |
96.39 ± 1.27 |
|
|
B |
Tablet-1 |
37.5 |
36.89±0.466 |
1.005 |
3.46 |
36.13±0.87 |
98.39±1.24 |
|
|
Tablet-2 |
75 |
72.44±0.950 |
2.03 |
4.42 |
71.85±0.45 |
96.58±1.267 |
* Tablet- 1 and Tablet-2 from two different companies. (Tablet-1: Venlor-XR capsules of Cipla (India), Tablet- 2: Ventab XL tablets of Intas pharmaceuticals (India).
**Average ± Standard deviation of six determinations, the t- and f-values refer to comparison of the proposed method with UV reference method. Theoretical values at 95% confidence limits t =2.57 and F = 5.05.
# Recovery of 10mg added to the pre analyzed sample (average of three determinations).
Reference method (reported UV method) using double distilled water (λ max=224nm).
Fig.6: Scheme of reactions
Chemistry of colored species:
The positively charged aliphatic tertiary nitrogen of VX molecule in acid medium is expected to attract the negatively charged part of the acidic dyes NB12BR or BCP and form an ion pair held together through electrostatic attraction. Based on the analogy, the structure of ion association complex in these methods is shown in the scheme (Fig-6).
CONCLUSION:
A significant advantage of an extraction spectrophotometric determination is that it can be applied to the determination of individual compounds in a multi component mixture. This aspect of spectrophotometric analysis is of major interest in analytical chemistry, since, it offers distinct possibilities in assay of a particular component in a complex dosage formulation. In the present study, VX was determined successfully as pure compound as well as a component in representative dosage formulations. The proposed methods applicable for the assay of drug and the advantage of wider range under Beer’s law limits. The proposed extractive visible spectrophotometric methods are validated as per ICH guide lines and possess reasonable precision, accuracy, simple, sensitive and can be used as alternative methods to the reported ones for the routine determination of VX depending on the need and situation.
ACKNOWLEDGEMENT:
The authors are very much thankful to the m/s Tychy Industries for providing gift sample of the drug and also thanks to University authorities for providing necessary facilities in this work.
REFERENCES:
1. Reynolds JE.F, Eds., In; Martindale, the Extra pharmacopoeia, 31st ed., The Pharmaceutical society, London, 1996: 337.
2. Budavari S, Eds; The Merck Index, 12 th ed. Merck and Co White house Station, NJ, 1996: 1695.
3. British Pharmacopoeia, 2007, volume II: 6317.
4. Raut BB, Kolte BL, Deo AA, Bagool MA, Shinde DB, A rapid and sensitive high performance liquid chromatographic method for determination of venlafaxine and o-desmethyl venlafaxine in human plasma with UV detection. J Liq. Chromatogr Technol., 26; 2003: 1297–1313.
5. Juan H, Zhiling Z, Huande L, Simultaneous determination of fluoxetine, citalopram, paroxetine, venlafaxine in plasma by high performance liquid chromatography-electrospray ionization mass spectrometry (HPLC-MS/ESI)J Chromatogr B Analyt Technol Biomed Life Sci., 820;2005:33–39.
6. Clement EM, Odontiadis J, Franklin M, Simultaneous measurement of venlafaxine and its major metabolite, O-desmethylvenlafaxine, in human plasma by high-performance liquid chromatography with coulometric detection and utilisation of solid-phase extraction. J Chromatogr B Biomed Sci. Appl., 705; 1998:303–308.
7. Vu RL, Helmeste D, Albers L, Reist C, Rapid determination of venlafaxine and O-desmethylvenlafaxine in human plasma by high-performance liquid chromatography with fluorimetric detection J Chromatogr B Biomed Sci Appl., 703;1997: 195–201
8. Li HD, Diang DR, Zhang BK, Yuan HY, Studies on determination of new drug venlafaxine in human plasma by HPLC with fluorimetric detection. Yaowu Fenxi Zazhi, 21; 2001:240-242.
9. Duverneuil C, de la Grandmaison GL, de Mazancourt P, Alvarez JC, A high-performance liquid chromatography method with photodiode-array UV detection for therapeutic drug monitoring of the nontricyclic antidepressant drugs. Ther Drug Monit, 25; 2003:567–73.
10. Titier K, Castaing N, Scotto-Gomez E, Molimard M, HPLC method with diode array detection for identification and quantification of the eight new antidepressants and five of their active metabolites in plasma after over dose. Ther Drug Monit, 25; 2003: 581-587.
11. Wille SM, Maudens KE, Van Peteghem CH, Lambert WE, Development of a solid phase extraction for 13 ‘new’ generation antidepressants and their active metabolites for gas chromatographic-mass spectrometric analysis. J Chromatogr A. 1098; 2005:19–29.
12. Wei z, Bing-Ren X, Cai.Yun W, LC-MS method for the determination of venlafaxine in human plasma and application to a pharmacokinetic study. Biomed Chromatography, 21(3); 2007: 266-272.
13. Shah GR, Thaker BT, Surathi KR and Parabia MH, Simultaneous determination of venlafaxine and its main active metabolite o-desmethyl venlafaxine in rat plasma by LC-MS/MS. Analytical Sciences, 25; 2009:1207-1210.
14. Bhatt J, Jangid A, Venkatesh G, Subbaiah G, Singh S, Liquid chromatography-tandem mass spectrometry (LC-MS-MS) method for simultaneous determination of venlafaxine and its active metabolite O -desmethyl venlafaxine in human plasma. J Chromatogr B Analyt Technol Biomed Life Sci., 829; 2005:75–81.
15. Frahnert C, Rao ML, Grasmäder K. Analysis of eighteen antidepressants, four atypical antipsychotics and active metabolites in serum by liquid chromatography: A simple tool for therapeutic drug monitoring. J Chromatogr B Analyt Technol Biomed Life Sci., 794; 2003: 35–47.
16. Baldania SL, Bhatt KK, Mehta RS, Shah DA and Tejal R. Gandhi, RP-HPLC estimation of venlafaxine in tablet dosage forms Indian Journal of Pharmaceutical Sciences, 70(1); 2008: 124-128.
17. Vidyavathi M, Krishna DR, Prasad KVSRG and Vidyasagar J, Rapid HPLC determination of venlafaxine in microbial Bio transformation studies. Current Trends in Biotechnology and Pharmacy, 3(1); 2009: 64-70.
18. Wachgler R, Moll W, Konig P and Conca A, Quantification of venlafaxine and O-dimethyl venlafaxine in human serum using HPLC analysis. Int. J. Clin Pharmacol Ther., 42; 2004:724-728.
19. Tournel G, Handret N, Hedouin VD, Eveaux M, Gosset D and Lhermitte M, HPLC method to screen and quantitate seven selective serotonine reuptake inhibitors in human serum J. Chromatography B, 761;2001 147-158.
20. Matoga M, Derhourca F, Titier K, Dumore F and Jarry C Rapid HPLC measurement of venlafaxine and o-desmethyl venlafaxine in human plasma. J. Chromatography B Biomed Sci., 760; 2001:213-218.
21. Maryam Hosseini, Application of UV spectrophotometry and HPLC for determination of venlafaxine and its four related substances in pharmaceutical dosage forms.Turk J. Pharm. Sci., 8(2);2011: 91-104.
22. Yang X, Jim M, Zhou J, Xiao Li S, Wen AD, Determination of venlafaxine in human plasma by HPLC sing could point extraction and spectroflurometric detection J. Chromatogr. B, 872; 2008: 38-42.
23. Juan H, Zhiling Z, Huande L, Simultaneous determination of fluoxetune, citalopram, Paroxetine, venlafaxine in plasma by HPLC-Electron spray ionization mass spectrophotometry J Chromatogra Bio Sci., Appl. 820;2005: 33-39.
24. LF-Limba, Jose Debbie V Loo, Cristina Delerue-Matos, Aida S Roque da Sila, electrochemical behavior of venlafaxine and its determination in pharmaceutical products using square wave voltammetry II Farmco, 54(3);1999: 145-148.
25. Nicoleta Carmen Purdel, Danbalalau Mihaela Ilie, Corina Cristina Arama, Determination of venlafaxine toxic levels from human serum by non-aqueous capillary electrophoresis. Farmacia, 58(1); 2010:62-69.
26. Cherkaoui S, Rudaz S, Venthey J, Non-aqueous capillary electrophoresis-mass spectrophotometry for separation of venlafaxine and its phase I metabolites Journal of Electrophoresis, 22; 2001: 491-496.
27. Paraskevas D, Verdoukas A, Demetrius G, Development and validation of a flow injection assay for dissolution studies of the antidepressant drug venlafaxine. Analytical Sciences, 21; 2005: 1515.
28. Asafu-Adjaye EB, Faustino PJ, Tawakkul MA, Anderson LW, Yu LX Kwon H, et al. Validation and application of a stability-indicating HPLC method for the invitro determination of gastric and intestinal stability of venlafaxine.J Pharm Biomed Anal., 43;2007: 1854–1859.
29. Makhija SN, Vavia PR, Stability indicating LC method for the estimation of venlafaxine in pharmaceutical formulations. J Pharm Biomed Anal., 28; 2002: 1055–1059.
30. Patel B, Patel J, Banwait H, Darji N, Patel D, Patel AGB, Patel M, Analytical spectrophotometric method development and validation of venlafaxine hydrochloride in pharmaceutical solid dosage forms using UV spectrophotometry Journal of Institutional Pharmacy and Life Sciences, 1(1);2011: 153-163.
31. Sundaraganapathy R, Jambulingam M, Ananda Thangadurai, Subasini U, Development and validation of UV soectrophotometric method for the determination of venlafaxine in bulk and solid dosage forms. Int. J. Pharm and Ind. Res., 1(1); 2011: 28-31.
32. Basaveswara Rao MV, Reddy BCK, Srinivasa Rao T, Prasanthi V, Estimation of venlafaxine in commercial dosage forms using simple and convenient spectrophotometric method. RASAYAN J. Chem., 2(2); 2009: 276-279.
33. Vimal D Shirvi, Vijaya kumar G, Channabasavaraj KP, Third order derivative spectrophotometric estimation of venlafaxine in bulk and pharmaceutical formulations. International Journal of Pharm Tech Research, 2(1); 2010: 700-703.
34. Pillai S, Singhavi I, Spectrophotometric methods for estimation of venlafaxine from tablet formulation. The Indian Pharmacist, 5; 2006: 75-76.
35. Onal A, Kepekci SE, Cetin SM, Erturk S, Spectrophotometric determination of certain antidepressants in pharmaceutical preparations. Journal of AOAC International, 89(4); 2006: 966-971.
36. Rajasekaran A, Arulkumaran M, Kannanraj S, Spectrophotometric determination of venlafaxine with folin ciocalteu reagent. Indian Journal of Pharmaceutical Sciences, 66(1); 2004: 101-103.
37. Padmanabha Reddy, Y and Prabhakar G, Assay of venlafaxine in bulk samples and pharmaceutical formulations by extractive spectrophotometry. International Journal of Chemical Sciences, 4(1); 2006: 151-156.
38. Appala Raju S, Sarasambi PS and Karadi AB, Extractive spectrophotometric determination of venlafaxine hydrochloride, Asian Journal of chemistry, 14(1); 2002: 467.
39. Beckett A.H. and Stenlake J.B, Practical Pharmaceutical chemistry, Vol. II, IV edition, CBS Publishers, New Delhi, 1997: 304.
Received on 22.11.2011 Modified on 14.12.2011
Accepted on 30.12.2011 © AJRC All right reserved
Asian J. Research Chem. 5(1): January 2012; Page 74-7824-27