Pharmaceutical Methods

VOL 1 NUMBER 1 (2010)

• Welcome to Pharmaceutical Methods
  Ambrose Furey
  
  Pharmaceutical Methods 2010 1(1):1-1
  
  
  DOI: 10.4103/2229-4708.72221
  
• Bioanalysis in drug discovery and development
  Saurabh Pandey, Preeti Pandey, Gaurav Tiwari, Ruchi Tiwari
  
  Pharmaceutical Methods 2010 1(1):14-24
  
  Recent years have witnessed the introduction of several high-quality review articles into the literature covering various scientific and technical aspects of bioanalysis. Now it is widely accepted that bioanalysis is an integral part of the pharmacokinetic/pharmacodynamic characterization of a novel chemical entity from the time of its discovery and during various stages of drug development, leading to its market authorization. In this compilation, the important bioanalytical parameters and its application to drug discovery and development approaches are discussed, which will help in the development of safe and more efficacious drugs with reduced development time and cost. It is intended to give some general thoughts in this area which will form basis of a general framework as to how one would approach bioanalysis from inception (i.e., discovery of a lead molecule) and progressing through various stages of drug development.
  DOI: 10.4103/2229-4708.72223
  
• Introduction to hyphenated techniques and their applications in pharmacy
  Kalpesh N Patel, Jayvadan K Patel, Manish P Patel, Ganesh C Rajput, Hitesh A Patel
  
  Pharmaceutical Methods 2010 1(1):2-13
  
  The hyphenated technique is developed from the coupling of a separation technique and an on-line spectroscopic detection technology. The remarkable improvements in hyphenated analytical methods over the last two decades have significantly broadened their applications in the analysis of biomaterials, especially natural products. In this article, recent advances in the applications of various hyphenated techniques, e.g., GC-MS, LC-MS, LC-FTIR, LC-NMR, CE-MS, etc. in the context of pre-isolation analyses of crude extracts or fraction from various natural sources, isolation and on-line detection of natural products, chemotaxonomic studies, chemical fingerprinting, quality control of herbal products, dereplication of natural products, and metabolomic studies are discussed with appropriate examples.
  DOI: 10.4103/2229-4708.72222
  
• Bioanalytical method validation: An updated review
  Gaurav Tiwari, Ruchi Tiwari
  
  Pharmaceutical Methods 2010 1(1):25-38
  
  The development of sound bioanalytical method(s) is of paramount importance during the process of drug discovery and development, culminating in a marketing approval. The objective of this paper is to review the sample preparation of drug in biological matrix and to provide practical approaches for determining selectivity, specificity, limit of detection, lower limit of quantitation, linearity, range, accuracy, precision, recovery, stability, ruggedness, and robustness of liquid chromatographic methods to support pharmacokinetic (PK), toxicokinetic, bioavailability, and bioequivalence studies. Bioanalysis, employed for the quantitative determination of drugs and their metabolites in biological fluids, plays a significant role in the evaluation and interpretation of bioequivalence, PK, and toxicokinetic studies. Selective and sensitive analytical methods for quantitative evaluation of drugs and their metabolites are critical for the successful conduct of pre-clinical and/or biopharmaceutics and clinical pharmacology studies.
  DOI: 10.4103/2229-4708.72226
  
• A validated stability indicating HPTLC method for simultaneous estimation of irbesartan and hydrochlorothiazide
  Amol S Khodke, Laxman V Potale, Mrinalini C Damle, Kailash G Bothara
  
  Pharmaceutical Methods 2010 1(1):39-43
  
  Introduction: Irbesartan, a diazaspiro angiotensin II blocker, is marketed in combination with Hydrochlorothiazide, which is a diuretic acting on distal convoluted tubule; for synergistic anti-hypertensive action. The present study deals with development and validation of a stability indicating HPTLC method for simultaneous estimation of Irbesartan and Hydrochlorothiazide using TLC plates precoated with Silica gel 60F254 and the mobile phase comprising Acetonitrile: Chloroform in the ratio of 5:6 v/v. Irbesartan and Hydrochlorothiazide were well resolved with Rf 0.27 ± 0.03 and 0.45 ± 0.03, respectively. Wavelength selected for the quantization was 270 nm. Inherent stability of these drugs was studied by exposing both drugs to various stress conditions as per ICH guidelines viz. Dry heat, oxidative, photolysis (UV and cool white fluorescent light) and hydrolytic conditions under different pH values. Results: Both the drugs were not degraded under dry heat and photolytic conditions, but showed degradation under hydrolytic condition. The degraded products of Irbesartan and hydrochlorothiazide were well resolved from the individual bulk drug response. Conclusion: The developed method is found to be simple, specific, precise and stability indicating. The specificity of the method was confirmed by peak purity profile of the resolved peaks.
  DOI: 10.4103/2229-4708.72229
  
• A simple and precise method for quantitative analysis of lumefantrine by planar chromatography
  Purnima Hamrapurkar, Mitesh Phale, Sandeep Pawar, Priti Patil, Mittal Gandhi
  
  Pharmaceutical Methods 2010 1(1):44-48
  
  A simple, precise and sensitive high performance thin layer chromatographic (HPTLC) method has been developed and validated for drug of choice Lumefantrine in treatment of malaria (P. falciparum). Silica gel 60 F254 HPTLC precoated plates were used for quantitative analytical purpose. Methanol water 9.5 + 0.5 (v/v) was used as the solvent system. Densitometric scanning was carried out with deuterium lamp set at detection wavelength of 266 nm. The response to lumefantrine concentration was linear in the concentration range of 1.25-12.50 μg/ml. The suitability of the method developed and validated was in accordance with the requirements of the ICH guidelines (Q2B). Thus the validated method can be further applied to quantitative analysis of lumefantrine in commercial pharmaceutical dosage form. The proposed method is simple, sensitive, precise and accurate, confirming its pharmaceutical application in routine quality control analysis.
  DOI: 10.4103/2229-4708.72230
  
• Spectrofluorimetric estimation of salbutamol sulphate in different dosage forms by formation of inclusion complex with β-cyclodextrin
  Harshit Narmadashankar Pandya, Hiren Harshadlal Berawala, Deepak Mohanlal Khatri, Priti Jignesh Mehta
  
  Pharmaceutical Methods 2010 1(1):49-53
  
  A simple, precise, reproducible and accurate spectrofluorimetric method for estimation of Salbutamol sulphate (SAL) in bulk drug and various dosage forms has been developed. This method is based on formation of inclusion complex of SAL in β-cyclodextrin (BCD) which gives fluorescence at excitation wavelength of 279.6 nm and emission wavelength of 609.8 nm in water. Formation of inclusion complex of drug with BCD enhances fluorescence intensity of drug leads to increased sensitivity. The developed method was validated according to ICH guidelines with respect to accuracy, precision, linearity, limit of detection, limit of quantification. Linearity was observed in the range of 4-20 μg/ml with correlation coefficient of 0.9982. The simplicity of the method permitted rapid analysis suitable for routine control. The developed method was successfully applied for the estimation of SAL in different marketed dosage forms like tablets, syrup and aerosol.
  DOI: 10.4103/2229-4708.72231
  
• A validated HPTLC method for estimation of moxifloxacin hydrochloride in tablets
  Vandana Dhillon, Alok Kumar Chaudhary
  
  Pharmaceutical Methods 2010 1(1):54-56
  
  A simple HPTLC method having high accuracy, precision and reproducibility was developed for the routine estimation of moxifloxacin hydrochloride in the tablets available in market and was validated for various parameters according to ICH guidelines. moxifloxacin hydrochloride was estimated at 292 nm by densitometry using Silica gel 60 F 254 as stationary phase and a premix of methylene chloride: methanol: strong ammonia solution and acetonitrile (10:10:5:10) as mobile phase. Method was found linear in a range of 9-54 nanograms with a correlation coefficient >0.99. The regression equation was: AUC = 65.57 × (Amount in nanograms) + 163 (r 2 = 0.9908).
  DOI: 10.4103/2229-4708.72232
  
• Simple spectrophotometric methods for estimation of aceclofenac from bulk and formulations
  A Bose, PP Dash, MK Sahoo
  
  Pharmaceutical Methods 2010 1(1):57-60
  
  Two simple, precise and accurate visible spectrophotometric methods were developed for the estimation of Aceclofenac in bulk drug and in pharmaceutical formulations. The proposed methods were indirect and based on determination of aceclofenac after its reaction with either (p-dimethylaminocinnamaldehyde or 3-Methyl-2-benzothiazolinone hydrazine hydrochloride and measuring the chromogen at the λmax by 658 and 592, respectively. Beers law obeyed in the concentration range of 1-200 μg/ml for method A and 1-100 μg/ml for method B. The accuracy of the methods was determined by recovery studies. The methods showed good reproducibility and recovery with relative standard deviation (in %) less than 2. The methods were found to be simple, economical, accurate and reproducible and can be used for routine analysis of Aceclofenac in bulk drug and in pharmaceutical formulations.
  DOI: 10.4103/2229-4708.72233
  
• A validated method for development of atovaquone as API and tablet dosage forms by UV spectroscopy
  Kalpesh N Patel, Jayvadan K Patel, Manish P Patel, Ganesh C Rajput
  
  Pharmaceutical Methods 2010 1(1):61-64
  
  A simple new spectrophotometric method has been developed for estimation of Atovaquone in bulk and tablet dosage form. Atovaquone is estimated to be 251 nm in methanol. The Beer's law is obeyed in the concentration range of 1-10 μg/mL of the drug. The slope and intercept values are 0.111 and 0.012, respectively. Results of analysis of this method have been validated statically and by recovery studies. The method is applied to the marketed tablet formulation. A result of the analysis of tablet formulation, given as a percentage of label claim ± standard deviation, is 99.14 ± 0.66. The precision and accuracy has been examined by performing recovery studies and found to be 100.09 ± 1.14. The developed method is simple, sensitive, and reproducible, and can be used for the routine analysis of Atovaquone in bulk and tablet dosage form.
  DOI: 10.4103/2229-4708.72234
  
• Plant metobolomics - A novel method in phytochemical analysis
  KK Ahmed Mueen, Hafez Madkor, Mahesh Attimarad, Tahir M Khan
  
  Pharmaceutical Methods 2010 1(1):65-66
  
  
  DOI: 10.4103/2229-4708.72235
  

  VOL 2 NUMBER 1 (2011)

• Method validation: A complex concept
  Ambrose Furey
  
  Pharmaceutical Methods 2011 2(1):1-2
  
  
  DOI: 10.4103/2229-4708.81081
  
• A developed and validated stability-indicating reverse-phase high performance liquid chromatographic method for determination of cefdinir in the presence of its degradation products as per International Conference on Harmonization guidelines
  Purnima Hamrapurkar, Priti Patil, Mitesh Phale, Mital Gandhi, Sandeep Pawar
  
  Pharmaceutical Methods 2011 2(1):15-20
  
  The present article deals with the development and validation of a stability-indicating, reverse-phase high performance liquid chromatographic (RP-HPLC) method, for the determination of cefdinir on a Waters RP Spherisorb C-18 column (250 mm × 4.6 mm, 5 μm). A mobile phase consisting of water (pH adjusted to 3.0 with orthophosphoric acid) : acetonitrile : methanol 13:5:2 (v/v/v) was used. The flow rate was 1 mL min -1 . The separation was performed at room temperature. Detection was carried out at 286 nm, using a PDA detector. The developed method was statistically validated for the linearity, accuracy, specificity, Limit of Detection (LOD), and Limit of Quantitation (LOQ). The specificity of the method was ascertained by forced degradation studies, by acid and alkali degradation, oxidation, photolysis, and heat degradation. The degraded products were well-separated from the analyte, with significant differences in their retention time values. The Beer Law was obeyed over a concentration range of 0.05 - 15.00 μg mL -1 and the correlation coefficient was 0.999.
  DOI: 10.4103/2229-4708.81085
  
• Spectrophotometric estimation of solifenacin succinate in tablet formulations
  Lokesh Singh, Sanju Nanda
  
  Pharmaceutical Methods 2011 2(1):21-24
  
  Aim: The aim of this study is to develop a simple, sensitive, rapid, accurate, and precise spectrophotometric method for the estimation of solifenacin succinate in tablet dosage forms. Materials and Methods: For methods I and II, in a series of 10 ml volumetric flasks, aliquots of standard drug solution (100 μg/ml) in double distilled water were transferred and diluted with the same so as to give several dilutions in the concentration ranges of 10 - 60 μg/ml and 10 - 60 μg/ml, respectively, of solifenacin succinate. To 5 ml of each dilution taken in a separating funnel, (5 ml of bromo thymol blue for method I and 5 ml of bromo phenol blue for method II) reagent and 5 ml of chloroform were added. The reaction mixture was shaken gently for five minutes and allowed to stand so as to separate the aqueous and chloroform layers. The absorbance maxima were measured at 415.6 nm and 412 nm for methods I and II, respectively. Results: The recovery studies were found close to 100%, which indicates the accuracy and precision of the proposed methods. Statistical analysis was carried out, the results of which were found to be satisfactory. Standard deviation values were found to be low and that indicated the reproducibility of the proposed methods. Conclusion: The results indicated that both methods could be used for the routine estimation of solifenacin succinate from tablet formulations.
  DOI: 10.4103/2229-4708.81086
  
• Isolation and determination of deoxynivalenol by reversed-phase high-pressure liquid chromatography
  Vikas Kumar Gupta, Pronobesh Chattopadhyay, Mohan Ch Kalita, Asshwani Kumar Chaurasia, Hemanta Kumar Gogoi, Lokendra Singh
  
  Pharmaceutical Methods 2011 2(1):25-29
  
  Deoxynivalenol (DON) is a mycotoxin produced by food contamination. It is a pharmacologically active compound that acts on the serotonin receptor, leading to several neuroendocrine and hematological disorders. In this article we describe a simple, accurate, and sensitive method for the quantification of DON. DON was quantified using a Phenomenex® ODS analytical C18 column (150 mm × 46 mm, 5 μm) with a mobile phase composed of mixture of water-acetonitrile-methanol (5:4:1, v/v/v) at a flow rate of 1.5 ml/min and at 254 nm in an ultraviolet (UV) detector The method has been validated with isolated samples of DON and provides a tool for the control of substandard and counterfeit commercial food products.
  DOI: 10.4103/2229-4708.81087
  
• Recent updates on codeine
  Monika Bhandari, Anil Bhandari, Aakanksha Bhandari
  
  Pharmaceutical Methods 2011 2(1):3-8
  
  Alleviation of pain is a major objective in medicine to increase the quality of life. Analgesics are agents that relieve pain by elevating the pain threshold without disturbing consciousness or altering other sensory modalities. Opium is an isoquinoline alkaloid obtained from poppy plant Papaver somniferum (Papaveraceae). Codeine is an alkaloid prepared from opium or morphine by methylation. Codeine is used as a central analgesic, sedative, hypontic, antinonciceptive, antiperistaltic, and is also recommended in tuberculosis and insomnia due to incessant coughing. The literature information relate mostly to the determination of codeine active components using Gas chromatography (GC), Capillary electrophoresis, Thin layer chromatography, High-performance thin layer chromatography, UV-Vis Spectrophotometry, High-performance liquid chromatography and GC in combination with Mass spectroscopy. This contribution provides a comprehensive review of its analytical and pharmacologic profile of codeine.
  DOI: 10.4103/2229-4708.81082
  
• Stress degradation studies and development of a validated stability-indicating-assay-method for determination of diacerein in presence of degradation products
  Purnima Hamrapurkar, Priti Patil, Masti Desai, Mitesh Phale, Sandeep Pawar
  
  Pharmaceutical Methods 2011 2(1):30-35
  
  Background: To understand the degradation behavior of diacerein and to develop a simple, rapid, sensitive, and validated RP-HPLC method for the determination of diacerein, in the presence of its degradation products. Materials and Methods: An accurate, sensitive, precise, rapid, and isocratic reversed-phase high-performance liquid chromatography (RP-HPLC) method, equipped with a photo-diode array (PDA) detector for analysis of diacerein in the bulk drug has been developed and validated. The best separation was achieved on a 250 mm × 4.6 mm i.d., 5-μm particle, RP C18 column with 50 : 50 (v/v) of water (pH adjusted to 2.9 with orthophosphoric acid) : acetonitrile as the mobile phase, at a flow rate of 1.0 ml/minute. The detection wavelength was set at 257 nm. Results: The response was a linear function of concentration over the range of 0.50 - 20 μg/ml (r = 0.999) and the limits of detection and quantitation were 0.1 μg/ml and 0.50 μg/ml, respectively. The method was validated in accordance with the International Conference on Harmonization (ICH) guidelines. The drug was subjected to oxidative, hydrolytic, photolytic, and thermal stress. The drug decomposed under alkaline hydrolytic stress conditions and also on thermal degradation and photolysis. It was stable on acid hydrolysis and oxidation. The degradation products produced as a result of this stress did not interfere with the detection of diacerein, and the assay could thus be regarded as stability-indicating. Conclusion: The method was suitable for application in the analysis of formulations of diacerein in quality-control laboratories, because it was simple and rapid, with good accuracy and precision.
  DOI: 10.4103/2229-4708.81088
  
• Development of the UV spectrophotometric method of Olmesartan medoxomil in bulk drug and pharmaceutical formulation and stress degradation studies
  Jaydeep Patel, Garala Kevin, Anjali Patel, Mihir Raval, Navin Sheth
  
  Pharmaceutical Methods 2011 2(1):36-41
  
  A simple, sensitive, specific, spectrophotometric method was developed for the detection of Olmesartan medoxomil (OLM) in bulk and pharmaceutical formulations. The optimum conditions for the analysis of the drug were established. OLM was subjected to stress degradation under different conditions recommended by the International Conference on Harmonization (ICH). The samples so generated were used for degradation studies using the developed method. The λmax of the OLM was found to be 257 nm. The method exhibited high sensitivity, with linearity, in the 2 to 20 μg/ml range. The lower limit of detection and the limit of quantification were found to be 1.012 μg/ml and 3.036 μg/ml, respectively. All the calibration curves demonstrated a linear relationship between the absorbance and concentration, with the correlation coefficient higher than 0.99. The regression equation of the curve was Y = 0.0579x + 0.0006. The precision of the method was found to be 40.043 ± 0.067 against the label claim of 40 mg. The percentage recovery was found to be 101.32 ± 0.452. The sample solution was stable for up to two hours. Hence, it could be concluded that the proposed method would be suitable for the analysis of OLM in bulk and pharmaceutical formulations.
  DOI: 10.4103/2229-4708.81092
  
• Development and validation of reversed phase-high-performance liquid chromatography method for determination of paracetamol and lornoxicam in tablet dosage form
  Deepak Kumar Jain, Pratibha Patel, HS Chandel, Abhay Kushwaha, Nilesh Jain
  
  Pharmaceutical Methods 2011 2(1):42-46
  
  A simple, precise, reliable, rapid and reproducible reversed phase-high-performance liquid chromatography method was developed and validated for the simultaneous estimation of Paracetamol (PCM) and Lornoxicam (LOX) present in tablet dosage forms. Chromatographic separation achieved isocratically on Luna C 18 column (5 μm, 150 × 4.60 mm) and methanol/phosphate buffer (60:40, v/v, pH 7.0) as mobile phase, at a flow rate of 1 ml/min. Detection was carried out at 260 nm. Parameters such as linearity, precision, accuracy, recovery, specificity and ruggedness are studied as reported in the ICH guidelines. The retention times for PCM and LOX was found to be 2.06±0.013and 4.38±0.07 min, respectively. Linearity for PCM and LOX was in the range of 10-50 mg/ml and 8-40 mg/ml, respectively. The mean recoveries obtained for LOX and PCM were 100± 0.16 and 99.50± 0.43%, respectively, and relative standard deviation (RSD) was less than 2. The correlation coefficients for all components are close to 1. The RSDs for three replicate measurements in three concentrations of samples in tablets are always less than 2%. Developed method was found to be accurate, precise, selective and rapid for simultaneous estimation of PCM and LOX in tablets.
  DOI: 10.4103/2229-4708.81095
  
• Spectrophotometric simultaneous determination of Tenofovir disoproxil fumarate and Emtricitabine in combined tablet dosage form by ratio derivative, first order derivative and absorbance corrected methods and its application to dissolution study
  Vishnu P Choudhari, Snehal Ingale, Sacchidanand R Gite, Dipali D Tajane, Vikram G Modak, Archana Ambekar
  
  Pharmaceutical Methods 2011 2(1):47-52
  
  Three simple, economical, precise, and accurate methods are described for the simultaneous determination of Tenofovir disoproxil fumarate (TE) and Emtricitabine (EM) in combined tablet dosage form. The first method is ratio derivative spectra, second is first-order derivative spectrophotometry and third is absorption corrected method. The amplitudes at 271.07 and 302.17 nm in the ratio derivative method, 224.38 and 306.88 nm in the first order derivative method were selected to determine Tenofovir disoproxil fumarate (TE) and Emtricitabine (EM), respectively, in combined formulation. Beer's law is obeyed in the concentration range of 3-21 μg/ml for TE and 2-14 μg/ml for EM for first two methods and range for third method was 6-30 μg/ml of TE and 4-20 μg/ml of EM. The percent assay for commercial formulation was found to be in the range 98.91%-101.72% for both the analytes by the proposed three methods. Absorption corrected method was successfully applied to carry out dissolution study of commercial tablet formulation by using USP II dissolution test apparatus. The methods were validated with respect to linearity, precision, and accuracy. Recoveries by proposed methods were found in the range of 99.06 %-101.34 % for both the analytes.
  DOI: 10.4103/2229-4708.81096
  
• Spectrophotometric estimation of tamsulosin hydrochloride by acid-dye method
  Alankar Shrivastava, Prachi Saxena, Vipin B Gupta
  
  Pharmaceutical Methods 2011 2(1):53-60
  
  A new spectrophotometric method for the estimation of tamsulosin hydrochloride in pharmaceutical dosage forms has been developed and validated. The method is based on reaction between drug and bromophenol blue and complex was measured at 421 nm. The slope, intercept and correlation coefficient was found to be 0.054, -0.020 and 0.999, respectively. Method was validated in terms of specificity, linearity, range, precision and accuracy. The developed method can be used to determine drug in both tablet and capsule formulations. Reaction was optimized using three parameters i.e., concentration of the dye, pH of the buffer, volume of the buffer and shaking time. Maximum stability of the chromophore was achieved by using pH 2 and 2 ml volume of buffer. Shaking time kept was 2 min and concentration of the dye used was 2 ml of 0.05% w/v solution. Method was validated in terms of linearity, precision, range, accuracy, LOD and LOQ and stochiometry of the method was also established using Mole ratio and Job's method of continuous variation. The dye benzonoid form (blue color) of dye ionized into quinonoid form (purple color) in presence of buffer and reacts with protonated form of drug in 1:1 ratio and forms an ion-pair complex (yellow color).
  DOI: 10.4103/2229-4708.81089
  
• Simultaneous determination of paracetamol and lornoxicam by RP-HPLC in bulk and tablet formulation
  Mahesh Attimarad
  
  Pharmaceutical Methods 2011 2(1):61-66
  
  Aim: The objective of the study was to develop simple RP-HPLC method for the simultaneous determination of paracetamol and lornoxicam without prior separation. Materials and Methods: In this method, Kromasil C8 (250 mm, 4.6 mm, 5 μm) column was used. The mobile phase used was methanol:phosphate buffer (60:40, v/v, pH 6.4), at flow rate of 1 ml min -1 . UV detection was monitored at 302 nm. Results: Calibration graphs were established in the range of 1-150 μg ml -1 and 0.5-100 μg ml -1 for paracetamol and lornoxicam, respectively. The average retention time for paracetamol and lornoxicam was found to be 3.15 ± 0.03 min and 5.25 ± 0.06 min, respectively. The detection limit and quantitation limit for paracetamol are 0.19 μg ml -1 and 0.59 μg ml -1 and for lornoxicam 0.10 μg ml -1 and 0.31 μg mL -1 , respectively. The intraday and interday precision expressed as percent relative standard deviation were below 2%. The mean recovery of paracetamol and lornoxicam was found to be in the range of 99.03-101.2%. Conclusion: The validated HPLC method was found to be rapid, precise and accurate and can be readily utilized for analysis of paracetamol and lornoxicam in bulk and in pharmaceutical formulations.
  DOI: 10.4103/2229-4708.81084
  
• Simultaneous estimation of amlodipine besylate and nebivolol hydrochloride in tablet dosage forms by reverse phase-high-performance liquid chromatographic using ultraviolet detection
  Deepak Sharma, Anurekha Jain, Alankar Shrivastava
  
  Pharmaceutical Methods 2011 2(1):9-14
  
  Background: The present study aimed to develop and validate the simultaneous estimation of amlodipine and nebivolol in tablet dosage forms. Materials and Methods: An isocratic reversed phase high-performance liquid chromatographic (HPLC) method with ultraviolet detection at 268 nm has been developed for the determination of amlodipine besylate (ADB) and nebivolol hydrochloride in dosage formulation. Results: Good chromatographic separation was achieved by using a stainless steel analytical column, the Lichrospher ODS RP-18 column (250 × 4 mm), particle size 5 μm. The system was operated at ambient temperature (25 ± 2°C) using a mobile phase consisting of acetonitrile (ACN) and a phosphate buffer (pH 3.0), mixed in a ratio of 40 : 60 at a flow rate of 0.8 ml/minute. The slope, intercept, and correlation coefficient were found to be 8818.2, - 18159, and 0.9993 for amlodipine and 9048.7, 108595, and 0.9998 for nebivolol, respectively. The proposed method was validated for its specificity, linearity, accuracy, and precision. Conclusion: The method was found to be suitable for the quality control of amlodipine besylate and nebivolol hydrochloride simultaneously in a bulk drug as well as in a formulation.
  DOI: 10.4103/2229-4708.81083
  

  VOL 3 NUMBER 2 (2012)

• Development and validation of a method for simultaneous estimation of ofloxacin and ornidazole in different dissolution media
  Dasharath M Patel, Jignesh A Soneji, Parth B Patel, Chhagan N Patel
  
  Pharmaceutical Methods 2012 3(2):102-105
  
  Introduction: Ofloxacin and ornidazole in a combined tablet dosage form is available in the market. This combination has gained increasing acceptance in diarrhea caused due to bacterial and protozoal infections. Ofloxacin and ornidazole are also combined in the capsule dosage form to modify its release pattern in different studies. Spectrophotometric and HPTLC methods have been reported for their simultaneous estimation in the tablet dosage form in specific solvents. This paper presents a simple, accurate, and reproducible spectrophotometric method for simultaneous estimation of ofloxacin and ornidazole in the tablet dosage form in different dissolution media. The reported method is helpful in determination of ofloxacin and ornidazole during a dissolution study. Materials and Methods: A simple, sensitive, accurate, and economical spectrophotometric method based on the simultaneous equation was developed for the estimation of ornidazole and ofloxacin simultaneously in the tablet or capsule dosage form in different dissolution media at different pH values. Results: Ofloxacin showed absorption maxima at 294 nm in 0.1 N HCl and at 287 nm in phosphate buffer pH 6.8 and phosphate buffer pH 7.4 while ornidazole showed absorption maxima at 277 nm in 0.1 N HCl and at 319 nm in two buffers, respectively. The linearity was obtained in the concentration range of 1-8 μg/ ml for ofloxacin and 4-26 μg/ml for ornidazole. Discussion: The concentrations of the drugs were determined by the simultaneous equation method. The results of analysis have been validated statistically and by recovery studies.
  DOI: 10.4103/2229-4708.103888
  
• Absorbance correction method for estimation of telmisartan and metoprolol succinate in combined tablet dosage forms
  Komal Patel, Amit Patel, Jayant Dave, Chaganbhai Patel
  
  Pharmaceutical Methods 2012 3(2):106-111
  
  Aim and Background: The present manuscript describes simple, sensitive, rapid, accurate, precise and economical spectrophotometric method for the simultaneous determination of telmisartan and metoprolol succinate in combined tablet dosage form. Materials and Methods: The method is based on the absorbance correction equations for analysis of both the drugs using methanol as solvent. Telmisartan has absorbance maxima at 296 nm and metoprolol succinate has absorbance maxima at 223 nm in methanol. The linearity was obtained in the concentration range of 2-16 μg/ ml and 3-24 μg/ml for telmisartan and metoprolol succinate, respectively. The concentrations of the drugs were determined by using absorbance correction method at both the wavelengths. The method was successfully applied to pharmaceutical dosage form because no interference from the tablet excipients was found. The suitability of this method for the quantitative determination of telmisartan and metoprolol succinate was proved by validation. The proposed method was found to be simple and sensitive for the quality control application of telmisartan and metoprolol succinate in pharmaceutical dosage form. Result: The result of analysis has been validated statistically and by recovery studies. Recoveries were found in the range of 98.08-100.55% of telmisartan and 98.41-101.87% of metoprolol succinate.
  DOI: 10.4103/2229-4708.103891
  
• Validated spectrophotometric methods for the simultaneous determination of telmisartan and atorvastatin in bulk and tablets
  Kaliappan Ilango, Pushpangadhan S Shiji Kumar
  
  Pharmaceutical Methods 2012 3(2):112-116
  
  Aim: Three simple, accurate, and reproducible spectrophotometric methods have been developed and validated for simultaneous estimation of telmisartan (TELM) atorvastatin (ATV) in combined tablet dosage form. Materials and Methods: The first method is based on first-order derivative spectroscopy. The sampling wavelengths were 223 nm (zero crossing of TELM) where ATV showed considerable absorbance and 272 nm (zero crossing of ATV) where TELM showed considerable absorbance. The second method Q-analysis (absorbance ratio), involves formation of Q-absorbance equation using respective absorptivity values at 280.9 nm (isobestic point) and 296.0 nm (λmax of TELM). The third method involves determination using multicomponent mode method; sampling wavelengths selected were 296.0 and 246.9 nm. Results : TELM and ATV followed linearity in the concentration range of 5-40 and 4-32 μg/ml for method I, 5-30 μg/ml and 2-24 μg/ml for method II and III, respectively. Mean recoveries for all three methods were found satisfactory. All methods were validated according to International Conference on Harmonization Q2B guidelines. Conclusion: The developed methods are simple, precise, rugged, and economical. The utility of methods has been demonstrated by analysis of commercially available tablet dosage form.
  DOI: 10.4103/2229-4708.103892
  
• Development and validation of spectrophotometric method of cefpodoxime proxetil using hydrotropic solubilizing agents
  Geet Asnani, Kiran Jadhav, Dinesh Dhamecha, Ashwini Sankh, Mrityunjaya Patil
  
  Pharmaceutical Methods 2012 3(2):117-120
  
  Purpose: To develop and validate specific and accurate UV spectrophotometric method of cefpodoxime proxetil by using different hydrotropic solubilizing agents. Materials and Methods: The present study deals with spectrophotometric analysis of cefpodoxime proxetil by utilizing 4 different hydrotropic agents such as ammonium acetate (6 M), sodium citrate (1.25 M), sodium gycinate (1 M), sodium chloride (1 M), and urea (1 M). Results : From different hydrotropic agents, urea showed best aqueous solubility of cefpodoxime proxetil. The linearity was observed in the concentration range of 10-120 μg/ml. The method was validated and found to be precise. Accuracy (percent recovery) for cefpodoxime proxetil was found to be 99.82 ± 0.106. Conclusion: Urea as hydrotropic agent showed best aqueous solubility of cefpodoxime proxetil, which can be used as solubilizing agent. The proposed method is new, simple, safe, eco-friendly, economic, accurate, and cost-effective and can be successfully employed in routine analysis.
  DOI: 10.4103/2229-4708.103893
  
• RP-UPLC method development and validation for the simultaneous estimation of ibuprofen and famotidine in pharmaceutical dosage form
  Yarram Ramakoti Reddy, Kakumani Kishore Kumar, MRP Reddy, K Mukkanti
  
  Pharmaceutical Methods 2012 3(2):57-61
  
  Aim and Backrgound: A stability-indicating LC method was developed for the simultaneous determination of Ibuprofen and Famotidine in pharmaceutical dosage forms. Materials and Methods: The chromatographic separation was achieved on Acquity UPLC BEH C-18,50 mm x 2.1 mm and 1.7 μm column with gradient elution. The mobile phase A contains a mixture of 50 mM sodium acetate buffer (pH 5.5): methanol (85:15, v/v), and the mobile phase B contains a mixture of 50 mM sodium acetate buffer (pH 5.5): methanol (25:75, v/v). The flow rate was 0.3 mL min -1 , and the detection wavelength was 260 nm. Results: The limit of detection for Ibuprofen and Famotidine was 1.6 and 1.2 μg mL -1 , respectively. The limit of quantification (LOQ) for Ibuprofen and Famotidine was 5.1 and 4.3 μg mL -1 , respectively. Conclusion: This method was validated for accuracy, precision, and linearity. The method was also found to be stability indicating.
  DOI: 10.4103/2229-4708.103873
  
• Estimation of sertraline by chromatographic (HPLC-UV273 nm ) technique under hydrolytic stress conditions
  Md Akhlaquer Rahman, Zeenat Iqbal, Mohd. Aamir Mirza, Arshad Hussain
  
  Pharmaceutical Methods 2012 3(2):62-67
  
  Purpose: In this paper, simple, specific and accurate RP-HPLC method was developed in order to study decomposition of sertraline (SRT) under the hydrolytic stress conditions (acid, neutral, alkaline and oxidative). Materials and Methods: The best separation of SRT and its degradation products were achieved on reverse phase LiChoCART with Purospher (RP-18e) column. The mobile phase was composed of methanol/water (75:25, v/v). The detection wavelength was 273 nm. The method was validated and response was found to be linear in the drug concentration range of 10-200 μg ml−1 with correlation coefficient of 0.998. Results: The RSD values for intra- and inter-day precision were < 0.65 and < 0.72%, respectively. Employing RP-HPLC method, degradation products were detected in the exposed samples. Conclusion: It was found that the susceptibility of SRT to hydrolytic decomposition increased in following manner: Neutral condition < alkaline condition < acid condition < oxidative condition.
  DOI: 10.4103/2229-4708.103874
  
• Comparison of UV spectrophotometry and high performance liquid chromatography methods for the determination of repaglinide in tablets
  Seema M Dhole, Pramod B Khedekar, Nikhil D Amnerkar
  
  Pharmaceutical Methods 2012 3(2):68-72
  
  Background: Repaglinide is a miglitinide class of antidiabetic drug used for the treatment of type 2 diabetes mellitus. A fast and reliable method for the determination of repaglinide was highly desirable to support formulation screening and quality control. Objective: UV spectrophotometric and reversed-phase high performance liquid chromatography (RP-HPLC) methods were developed for determination of repaglinide in the tablet dosage form. Materials and Methods: The UV spectrum recorded between 200 400 nm using methanol as solvent and the wavelength 241 nm was selected for the determination of repaglinide. RP-HPLC analysis was carried out using Agilent TC-C18 (2) column and mobile phase composed of methanol and water (80:20 v/v, pH adjusted to 3.5 with orthophosphoric acid) at a flow rate of 1.0 ml/min. Parameters such as linearity, precision, accuracy, recovery, specificity and ruggedness are studied as reported in the International Conference on Harmonization (ICH) guidelines. Results: The developed methods illustrated excellent linearity (r 2 > 0.999) in the concentration range of 5-30 μg/ml and 5-50 μg/ml for UV spectrophotometric and HPLC methods, respectively. Precision (%R.S.D < 1.50) and mean recoveries were found in the range of 99.63-100.45% for UV spectrophotometric method and 99.71-100.25% for HPLC method which shows accuracy of the methods. Conclusion: The developed methods were found to be reliable, simple, fast, accurate and successfully used for the quality control of repaglinide as a bulk drug and in pharmaceutical formulations.
  DOI: 10.4103/2229-4708.103875
  
• RP-HPLC method for simultaneous estimation of tenofovir disoproxil fumarate, lamivudine, and efavirenz in combined tablet dosage form
  Dhara S Bhavsar, Bhavini N Patel, Chhaganbhai N Patel
  
  Pharmaceutical Methods 2012 3(2):73-78
  
  Background: A simple, precise, accurate, and rapid reverse phase-high performance liquid chromatography (RP-HPLC) method with UV-Visible detector has been developed and subsequently validated for the simultaneous determination of tenofovir disoproxil fumarate (TDF), lamivudine (LAMI), and efavirenz (EFV) in their combined tablet dosage form. Materials and Methods: The separation was based on the use of a Kromasil C 18 analytical column (150 × 4.6 mm, i.d., 5 μm). The mobile phase consisted of a mixture of 70 volumes of methanol and 30 volumes of 10 mM phosphate buffer (pH 5.0). The separation was carried out at 40°C temperature with a flow rate of 1 ml/min. Results: Quantitation was achieved with UV detection at 254 nm, with linear calibration curves at concentration ranges of 1-6 μg/ml for TDF and LAMI and 2-12 μg/ml for EFV. The recoveries obtained were 99.46-101.36% for LAMI, 99.57-101.42% for TDF, and 99.96-100.87 for EFV. Conclusion: The method was validated according to International conference of harmonisation guidelines in terms of accuracy, precision, specificity, robustness, limits of detection and quantitation, and other aspects of analytical validation.
  DOI: 10.4103/2229-4708.103876
  
• A validated RP-HPLC-UV method for quantitative determination of puerarin in Pueraria tuberosa DC tuber extract
  Amal K Maji, Niladri Maity, Pratim Banerji, Debdulal Banerjee
  
  Pharmaceutical Methods 2012 3(2):79-83
  
  Background: Pueraria tuberosa (Fabaceae) is a well-known medicinal herbs used in Indian traditional medicines. The puerarin is one of the most important bioactive constituent found in the tubers of this plant. Quantitative estimation of bioactive molecules is essential for the purpose of quality control and dose determination of herbal medicines. The study was designed to develop a validated reversed phase high-performance liquid chromatography (RP-HPLC) method for the quantification of puerarin in the tuber extract of P. tuberosa. Materials and Methods: The RP-HPLC system with Luna C18 (2) 100 Å, 250 × 4.6 mm column was used in this study. The analysis was performed using the mobile phase: 0.1% acetic acid in acetonitrile and 0.1% acetic acid in water (90:10, v/v) under column temperature 25°C. The detection wavelength was set at 254 nm with a flow rate of 1 ml/min. The method validation was performed according to the guidelines of International Conference on Harmonization. Results: The puerarin content of P. tuberosa extract was found to be 9.28 ±0.09%. The calibration curve showed good linearity relationship in the range of 200-1000μg/ml (r 2>0.99). The LOD and LOQ were 57.12 and 181.26μg/ml, respectively and the average recovery of puerarin was 99.73% ±1.02%. The evaluation of system suitability, precision, robustness and ruggedness parameters were also found to produce satisfactory results. Conclusions: The developed method is very simple and rapid with excellent specificity, accuracy and precision which can be useful for the routine analysis and quantitative estimation of puerarin in plant extracts and formulations.
  DOI: 10.4103/2229-4708.103879
  
• Development and validation of a stability indicating RP-HPLC method for simultaneous estimation of Olmesartan Medoxomil and Metoprolol Succinate in pharmaceutical dosage form
  Nirmal M Thakker, Haresh B Panchal, Dinesh R Rakholiya, R Murugan, Vishnu P Choudhari, Bhanudas S Kuchekar
  
  Pharmaceutical Methods 2012 3(2):84-89
  
  Aim and Backrgound: A simple, rapid, precise and isocratic RP-HPLC (Reverse Phase High Performance Liquid Chromatography) method is aimed to develop for the simultaneous estimation of Olmesartan Medoxomil and Metoprolol Succinate in bulk drug and pharmaceutical dosage form. Materials and Methods: The quantification is carried out using YMC-Pack CN (250 × 4.6 mm, 5.0 μm) column and the mobile phase comprises of 0.05% Trifluoro acetic acid (TFA) and Acetonitrile (ACN) (70:30 v/v). The flow rate is 1.0 ml/min. The eluent is monitored at 220 nm. The retention times of Olmesartan Medoxomil and Metoprolol Succinate are 7.9 min and 4.1 min respectively. The method is validated in terms of linearity, precision, accuracy, specificity, limit of detection and limit of quantitation. Results: Linearity and percentage recoveries of both Olmesartan Medoxomil and Metoprolol Succinate are in the range of 5-35 μg/ml and 100 ± 2%, respectively. The stress testing of both the drugs individually and their mixture is carried out under acidic, alkaline, oxidation, photo-stability and thermal degradation (dry heat and wet heat) conditions and its degradation products are well resolved from the analyte peaks. Conclusion: This method was successfully validated for accuracy, precision, and linearity.
  DOI: 10.4103/2229-4708.103880
  
• Development and validation of HPTLC method for the determination of mycophenolate mofetil in bulk and pharmaceutical formulation
  S Kathirvel, K Rajendra Prasad, K Madhu Babu
  
  Pharmaceutical Methods 2012 3(2):90-93
  
  Aim: Described in this manuscript is the first ever reported, new, simple, high-performance thin-layer chromatographic method for the determination of mycophenolate mofetil in bulk and tablet dosage form. Materials and Methods: The drug was separated on aluminum plates precoated with silica gel 60 F 254 with toluene, acetone, and methanol in the ratio of 6:2:2 (v/v/v) as the mobile phase. Quantitative analysis was performed by densitometric scanning at 254 nm. Results: The method was validated for linearity, accuracy, precision, and robustness. The calibration plot was linear in the range of 100-500 ng band -1 for mycophenolate mofetil. The method was successfully applied to the analysis of the drug in a pharmaceutical dosage form.
  DOI: 10.4103/2229-4708.103882
  
• Validation of 1-methyl-2-phenylindole method for estimating lipid peroxidation in the third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ)Bg 9
  Yasir Hasan Siddique, Smita Jyoti, Falaq Naz, Mohammad Afzal
  
  Pharmaceutical Methods 2012 3(2):94-97
  
  Background: A method using 1-methyl-2-phenylindole was developed for the estimation of lipid peroxidation in third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ)Bg9 . The method is specific for the estimation of malonaldehyde. Materials and Methods: The larvae were exposed to 0.0025, 0.025, 0.050, and 0.100 μl/ml of cyclophosphamide for 24 and 48 h. The homogenate was prepared of the larvae tissue explant and the absorbance was noted at 586 nm. Results: A significant dose-dependent increase in the mean absorbance values was observed for both 24 and 48 h of exposure as compared to the untreated group. Conclusions: On the basis of results obtained, it is suggested that the present method is more precise, accurate, and robust for the estimation of lipid peroxidation in the third instar larvae of transgenic D. melanogaster (hsp70-lacZ)Bg9.
  DOI: 10.4103/2229-4708.103883
  
• Development and validation of spectrophotometric method for simultaneous estimation of paracetamol and lornoxicam in different dissolution media
  Dasharath M Patel, Bhavesh M Sardhara, Diglesh H Thumbadiya, Chhagan N Patel
  
  Pharmaceutical Methods 2012 3(2):98-101
  
  Background: Paracetamol and lornoxicam in combined tablet dosage form are available in the market. This combination is used to treat inflammatory diseases of the joints, osteoarthritis and sciatica. Spectrophotometric and high performance liquid chromatography (HPLC) methods have been reported for their simultaneous estimation in tablet dosage form in specific solvent. This paper presents simple, accurate and reproducible spectrophotometric method for simultaneous determination of paracetamol and lornoxicam in tablet dosage form in different dissolution media. The reported method is helpful in determination of paracetamol and lornoxicam during dissolution study. Materials and Methods: Simple, sensitive, accurate and economical spectrophotometric method based on an absorption correction equation was developed for the estimation of paracetamol and lornoxicam simultaneously in tablet dosage form in different dissolution media at different pH. Results: Paracetamol showed absorption maxima at 243 nm in 0.1N HCland phosphate buffer pH 6.8, while lornoxicam showed absorption maxima at 374 nm in 0.1N HCland phosphate buffer pH 6.8. The linearity was obtained in the concentration range of 4-12 μg/ml for paracetamol and 4-16 μg/ ml for lornoxicam. Discussion: The concentrations of the drugs were determined by an absorption correction equation method. The results of analysis have been validated statistically by recovery studies.
  DOI: 10.4103/2229-4708.103885