Spectrophotometric and HPLC Methods for Simultaneous Estimation of Amlodipine Besilate, Losartan Potassium and Hydrochlorothiazide in Tablets

Two UV-spectrophotometric and one reverse phase high performance liquid chromatography methods have been developed for the simultaneous estimation of amlodipine besilate, losartan potassium and hydrochlorothiazide in tablet dosage form. The first UV spectrophotometric method was a determination using the simultaneous equation method at 236.5, 254 and 271 nm over the concentration range 5-25, 10-50 and 5-25 μg/ml for amlodipine besilate, losartan potassium and hydrochlorothiazide, respectively. The second UV method was a determination using the area under curve method at 231.5-241.5, 249-259 and 266-276 nm over the concentration range of 5-25, 5-25 and 10-50 μg/ml for amlodipine besilate, hydrochlorothiazide and losartan potassium, respectively. In reverse phase high performance liquid chromatography analysis is carried out using 0.025 M phosphate buffer (pH 3.7):acetonitrile (57:43 v/v) as the mobile phase and Kromasil C18 (4.6 mm i.d×250 mm) column as stationery phase with detection wavelength of 232 nm linearity was obtained in the concentration range of 2-14, 20-140 and 5-40 μg/ml for amlodipine besilate, losartan potassium and hydrochlorothiazide, respectively. Both UV-spectrophotometric and reverse phase high performance liquid chromatography methods were statistically validated and can be used for analysis of combined dose tablet formulation containing amlodipine besilate, losartan potassium and hydrochlorothiazide.     

Spectrophotometric, spectrofluorimetric, HPLC and CZE determination of mirtazapine in pharmaceutical tablets

Four analytical methods have been developed for the quality control of tablets containing mirtazapine: spectrophotometry, spectrofluorimetry, high performance liquid chromatography (HPLC) and capillary zone electrophoresis (CZE). All the methods only require a simple extraction procedure of mirtazapine from the tablets before analysis. The concentration of mirtazapine in solutions was determined in the linearity range of 5-25 microg/ml at lambda=315 nm for spectrophotometry and at lambda=220 nm for HPLC and CZE. Spectrofluorimetric determinations were achieved at lambda(excitation)=328 nm and lambda(emission)=415 nm in the linearity range of 2-25 ng/ml. All the methods gave similar results and were validated for selectivity, linearity, precision and sensitivity. Spectrometric methods gave slightly higher RSD values (up to 2.54%). The four methods were directly and easily applied to the pharmaceutical preparation with accuracy, resulting from recovery experiments between 99.72% in HPLC and 101.47% in spectrofluorimetry.     

Estimation of Amlodipine Besylate, Valsartan and Hydrochlorothiazide in Bulk Mixture and Tablet by UV Spectrophotometry

A simple, precise, accurate and economic simultaneous UV spectrophotometric method has been developed for the estimation of amlodipine besylate, valsartan and hydrochlorothiazide in combination in bulk mixture and tablet. The estimation was based upon measurement of absorbance at absorbance maxima of 359 nm, 317 nm and 250 nm for amlodipine besylate, hydrochlorothiazide and valsartan in methanol, respectively in bulk mixture and tablet. The Beer Lambert''s law obeyed in the concentration range 5-25 μg/ml, 10-50 μg/ml and 5-25 μg/ml for amlodipine besylate, hydrochlorothiazide and valsartan, respectively. The estimation of bulk mixture and tablet was carried out by simultaneous equation, Q-analysis and area under curve method for estimation of amlodipine besylate and hydrochlorothiazide and standard curve method for estimation of valsartan. The results were found to be in the range of 99.6±1.52% to 102±0.51%. Method was validated with respect to specificity, linearity, range, accuracy, precision, LOD, LOQ, robustness, ruggedness and can be applied for routine analysis of tablet dosage forms.     

Development and Validation of a RP-HPLC Method for Estimation of Montelukast Sodium in Bulk and in Tablet Dosage Form

The present work describes a simple, precise and accurate HPLC method for estimation of montelukast sodium in bulk and in tablet dosage form. The separation was achieved by using octadecylsilane column (C18) and acetonitrile:1 mM sodium acetate adjusted to pH 6.3 with acetic acid in proportion of 90:10 v/v as mobile phase, at a flow rate of 1.5 ml/min. Detection was carried out at 285 nm. The retention time of montelukast sodium was found to be 3.4 min. The limit of detection was found 1.31 µg/ml and limit of quantification 3.97 µg/ml. The accuracy and reliability of the proposed method was ascertained by evaluating various validation parameters like linearity (1-100 µg/ml), precision, accuracy and specificity according to ICH guidelines. The proposed method provides an accurate and precise quality control tool for routine analysis of montelukast sodium in bulk and in tablet dosage form.     

RP-HPLC Method for Simultaneous Estimation of Nitazoxanide and Ofloxacin in Tablets

A reverse phase high performance liquid chromatography method was developed for simultaneous estimation of nitazoxanide and ofloxacin in tablet formulation. The separation and quantification was achieved by Hiq Sil C₁₈V Size 4.6 mm Ø ^*250 mm column in isocratic mode, with mobile phase consisting of acetonitrile-methanol-0.4 M citric acid, (60:30:10, v/v/v). Citric acid used to stabilize nitazoxanide and ofloxacin in mobile phase. The mobile phase was pumped at a rate of 0.6 ml/min and the detection was carried out at 304 nm. The retention time of ofloxacin and nitazoxanide was found to be 3.122 and 5.902 min, respectively. The method was validated for linearity, accuracy, and precision. Linearity for ofloxacin and nitazoxanide were in the range 2-36 μg/ml and 5-90 μg/ml, respectively. The developed method was found to be accurate, precise and selective for simultaneous estimation of ofloxacin and nitazoxanide in tablets.     

Spectrophotometric Estimation of Ketotifen Fumarate from Tablet Formulations

Two simple and sensitive visible spectrophotometric methods have been developed for the quantitative estimation of ketotifen fumarate from its tablet formulation. The developed methods are based on formation of chloroform extractable colored complex of with 2-nitroso- napthol-4- sulphonic acid and rhodizonic acid. The extracted complex of drug with 2-nitroso- napthol-4- sulphonic acid (method-I), showed absorbance maxima at 436.5 nm and with rhodizonic acid (method-II), showed absorbance maxima at 489.5 nm. The linearity range for both the developed methods was observed in the concentration range of 50-250 µg/ml of drug. Results of analysis for both the developed methods were validated statistically and by recovery studies.     

Rapid and sensitive liquid chromatographic method for determination of Paclitaxel from parenteral formulation and nanoparticles

A simple and fast reversed phase liquid chromatographic method was developed for estimation of paclitaxel in commercially available parenteral formulation and nanoparticles. Separations were carried out using mobile phase consisting of acetonitrile and 20 mM potassium dihydrogen phosphate (45:55, v/v) on Lichrocart^® C₁₈ analytical column at a flow rate of 1 ml/min and detection wavelength of 230 nm. The developed method exhibited linearity over an analytical range of 50-2000 ng/ml with regression equation, mean peak area= 137.58 concentration (ng/ml)+1765.94, (R₂ =0.9999). The method demonstrated selectivity with no interfering peaks eluting near the vicinity of drug peak. The method was found to be sensitive with detection and quantification limits of 7.57 ng/ml and 22.94 ng/ml. The method has shown consistent and good recoveries from parenteral formulation (100.06±0.86%) and nanoparticles (100.43±0.91%). The method was successfully employed for the analysis of in vitro release study samples of nanoparticle formulation. The method was also applied for determination of paclitaxel content in various pharmaceutical formulations.     

Simultaneous Determination of Valsartan and Hydrochlorothiazide in Tablets by RP-HPLC

A simple, reproducible and efficient reverse phase high performance liquid chromatographic method was developed for simultaneous determination of valsartan and hydrochlorothiazide in tablets. A column having 200 × 4.6 mm i.d. in isocratic mode with mobile phase containing methanol:acetonitrile:water:isopropylalcohol (22:18:68:2; adjusted to pH 8.0 using triethylamine; v/v) was used. The flow rate was 1.0 ml/min and effluent was monitored at 270 nm. The retention time (min) and linearity range (μg/ml) for valsartan and hydrochlorothiazide were (3.42, 8.43) and (5-150, 78-234), respectively. The developed method was found to be accurate, precise and selective for simultaneous determination of valsartan and hydrochlorothiazide in tablets.     

Development and Validation of RP-HPLC Method for Simultaneous Estimation of Atorvastatin Calcium and Fenofibrate in Tablet Dosage Forms

A reverse phase high performance liquid chromatographic method was developed for the simultaneous estimation of atorvastatin calcium and fenofibrate in tablet formulation. The separation was achieved by Luna C18 column and methanol:acetate buffer pH 3.7 (82:18 v/v) as mobile phase, at a flow rate of 1.5 ml/min. Detection was carried out at 248 nm. Retention time of atorvastatin calcium and fenofibrate was found to be 3.02+0.1 and 9.05+0.2 min, respectively. The method has been validated for linearity, accuracy and precision. Linearity for atorvastatin calcium and Fenofibrate were in the range of 1-5 μg/ml and 16-80 μg/ml, respectively. The mean recoveries obtained for Atorvastatin calcium and fenofibrate were 101.76% and 100.06%, respectively. Developed method was found to be accurate, precise, selective and rapid for simultaneous estimation of atorvastatin calcium and fenofibrate in tablets.     

A validated capillary electrophoresis method for simultaneous determination of ezetimibe and atorvastatin in pharmaceutical formulations

A simple, precise, and sensitive capillary electrophoresis technique coupled with a diode array detector has been developed for the separation and simultaneous determination of ezetimibe and atorvastatin in pharmaceutical formulations. Separation of both ezetimibe and atorvastatin was achieved utilizing fused silica capillary (58 cm × 75 μm ID) and background electrolyte solution that consisted of phosphate buffer (2.5 mM, pH 6.7): methanol (70:30 v/v). The proposed method was validated by testing its specificity, linearity, precision, accuracy, recovery, and detection limit/quantitation limit values. The method was linear over the range 2.5–50 μg/ml for ezetimibe (r = 0.9992) and 1–100 μg/ml for atorvastatin (r = 0.9999). Within-day and between-day RSD for ezetimibe and atorvastatin were ⩽5.6% and ⩽2.9%, respectively. The detection limit was 0.07 μg/ml for ezetimibe and 0.06 μg/ml for atorvastatin. The validated method was successfully employed for the determination of ezetimibe and atorvastatin in tablets with no interfering peaks from common pharmaceutical excipients. The percentage recoveries of the two drugs from their tablets were 99.80 ± 1.76 and 100.19 ± 1.83, respectively.     

Ion-Pairing Reverse-Phase High Performance Liquid Chromatography Method for Simultaneous Estimation of Atenolol and Indapamide in Bulk and Combined Dosage Form

A sensitive, accurate, precise and validated ion-pairing reverse-phase liquid chromatographic method for the quantitative determination of atenolol and indapamide in bulk and tablet dosage form was developed. The proposed ion-pairing reverse-phase high performance liquid chromatography method utilises C₁₈ column with 5 μm, 150×4.6 mm i.d. column and mobile phase consisting of 0.1% w/v solution of octane sulphonic acid, sodium salt and methanol (55:45 v/v), (pH 2.8) and ultraviolet detection at 235 nm. A linearity range of 1-250 μg/ml and 1-25 μg/ml for atenolol and indapamide, respectively, was obtained. The mean recoveries are 100.48 and 99.82% for atenolol and indapamide, respectively. The method was validated as per International Conference on Harmonization guidelines.     

Simultaneous Spectrophotometric Estimation of Haloperidol and Trihexyphenidyl in Tablets

The combination of haloperidol and trihexyphenidyl is a dosage form to be used as antidyskinetic agent. Literature revealed that there is no single method for the simultaneous estimation of these drugs in tablet dosage form, which prompted us to develop a simple, rapid, accurate, economical and sensitive spectrophotometric method. The simultaneous estimation method is based on the principle of additivity of absorbance, for the determination of haloperidol and trihexyphenidyl in tablet formulation. The absorption maxima of the drugs were found to be at 245.0 nm and 206.0 nm respectively for haloperidol and trihexyphenidyl in methanol and 0.1N HCl (90:10). The obeyance of Beer Lambert’s law was observed in the concentration range of 2.5-12.5 µg/ml for haloperidol and 1.0-5.0 µg/ml for trihexyphenidyl. The accuracy and reproducibility of the proposed method was statistically validated by recovery studies.     

RP-HPLC Method for the Determination of Losartan Potassium and Ramipril in Combined Dosage Form

A simple, specific and accurate reverse phase liquid chromatographic method was developed for the simultaneous determination of losartan potassium and ramipril in table dosage forms. A hypersil ODS C18, 4.6×250 mm, 5 μm column in isocratic mode, with mobile phase acetonitrile:methanol:10 mM tetra butyl ammonium hydrogen sulphate in water in the ratio of 30:30:40% v/v/v was used. The flow rate was 1.0 ml/min and effluent was monitored at 210 nm. The retention times of losartan potassium and ramipril were 4.7 and 3.3 min, respectively. The linearity range for losartan potassium and ramipril were in the range of 0.04-100 μg/ml and 0.2-300 μg/ml, respectively. The proposed method was also validated and successfully applied to the estimation of losartan potassium and ramipril in combined tablet formulations.     

Simultaneous Spectrophotometric Determination of Drotaverine Hydrochloride and Paracetamol in Tablet

Two simple, accurate and reproducible spectrophotometric methods; Q analysis and first order derivative method have been described for the simultaneous estimation of drotaverine hydrochloride and paracetamol in combined tablet dosage form. Absorption maxima of drotaverine hydrochloride and paracetamol in distilled water were found to be 303.5 nm and 243.5 nm respectively. Beer''s law was obeyed in the concentration range 5-50 μg/ml for drotaverine and 5-60 μg/ml for paracetamol. In Q analysis method, two wavelengths were selected at isobestic point (277 nm) and λ_max of paracetamol (243.5 nm). In first order derivative method, zero crossing point for drotaverine hydrochloride and paracetamol were selected at 303.5 nm and 243.5 nm, respectively. The results of two methods were validated statistically and recovery studies were found to be satisfactory.     

Simultaneous Derivative Spectrophotometric Analysis of Doxylamine Succinate, Pyridoxine Hydrochloride and Folic Acid in Combined Dosage Forms

Two UV spectrophotometric methods have been developed, based on first derivative spectrophotometry for simultaneous estimation of doxylamine succinate, pyridoxine hydrochloride, and folic acid in tablet formulations. In method I, the concentrations of these drugs were determined by using linear regression equation. Method II is also based on first derivative spectrophotometry however simultaneous equations (Vierdot''s method) were derived on derivative spectra. The first derivative amplitudes at 270.0, 332.8 and 309.2 nm were utilized for simultaneous estimation of these drugs respectively by both methods. In both the methods, linearity was obtained in the concentration range 2.5-50 μg/ml, 1-40 μg/ml and 1-30 μg/ml for doxylamine succinate, pyridoxine hydrochloride, and folic acid respectively. The developed methods show best results in terms of linearity, accuracy, precision, LOD, LOQ and ruggedness for standard laboratory mixtures of pure drugs and marketed formulations. The common excipients and additives did not interfere in their determinations.     

Stability-indicating Reversed-phase Liquid Chromatographic Method for Simultaneous Determination of Losartan Potassium and Ramipril in Tablets

A stability-indicating reversed-phase liquid chromatographic method has been developed and validated for simultaneous determination of losartan potassium and ramipril. Separations were achieved using a C₁₈ column with mobile phase consisting of acetonitrile and (0.2% v/v, pH 2.5) aqueous trifluoroacetic acid (45:55, v/v) in isocratic mode at 1 ml/min flow rate. Column effluent was monitored at 210 nm using a UV detector. The method was validated for selectivity, linearity, accuracy, precision, sensitivity and robustness. Novel microwave-assisted forced degradation technique was employed for evaluation of selectivity. The method demonstrated excellent linearity for losartan potassium and ramipril with regression coefficients of 0.9999 and 0.9998, respectively. The linearity range was found to be 62.5-5000 ng/ml and 125-10,000 ng/ml with the mean percentage recoveries of 100.36% (±2.27) and 100.16% (±3.33) for losartan potassium and ramipril, respectively. In a robustness study, a full factorial design revealed that the analytical response remains unaffected by small variations in the critical chromatographic factors. The method was found to be sensitive with quantification limits of 44.30 and 79.93 ng/ml for losartan potassium and ramipril. The method was successfully employed for the determination of losartan potassium and ramipril in commercially available and in-house prepared tablets.     

Development and Validation of a HPLC Method for the Determination of Cyclosporine A in New Bioadhesive Nanoparticles for Oral Administration

A simple and reliable high performance liquid chromatography method was developed and validated for the rapid determination of cyclosporine A in new pharmaceutical dosage forms based on the use of poly (methylvinylether-co-maleic anhydride) nanoparticles. The chromatographic separation was achieved using Ultrabase C₁₈ column (250×4.6 mm, 5 μm), which was kept at 75°. The gradient mobile phase consisted of acetonitrile and water with a flow rate of 1 ml/min. The effluent was monitored at 205 nm using diode array detector. The method exhibited linearity over the assayed concentration range (22-250 μg/ml) and demonstrated good intraday and interday precision and accuracy (relative standard deviations were less than 6.5% and the deviation from theoretical values is below 5.5%). The detection limit was 1.36 μg/ml. This method was also applied for quantitative analysis of cyclosporine A released from poly (methylvinylether-co-maleic anhydride) nanoparticles.     

Application of HPLC for the simultaneous determination of aceclofenac, paracetamol and tramadol hydrochloride in pharmaceutical dosage form

A simple, precise and accurate reversed-phase liquid chromatographic method has been developed for the simultaneous estimation of aceclofenac (ACF), paracetamol (PCM) and tramadol hydrochloride (TRM) in pharmaceutical dosage form. The chromatographic separation was achieved on a HiQ-Sil™ HS C18 column (250×4.6 mm i.d., 5 μm particle size), kromatek analytical column at ambient temperature. The mobile phase consisted of 40: 60 (v/v); phosphate buffer (pH 6.0): methanol. The flow rate was set to 1.0 mL min⁻¹ and UV detection was carried out at 270 nm. The retention time (t_R) for ACF, PCM and TRM were found to be 14.567 ± 0.02, 3.133 ± 0.01 and 7.858 ± 0.02 min, respectively. The validation of the proposed method was carried out for linearity, precision, robustness, limit of detection, limit of quantitation, speci city, accuracy and system suitability. The linear dynamic ranges were from 40–160 μg mL⁻¹ for ACF, 130–520 μg mL⁻¹ for PCM and 15–60 μg mL⁻¹ for TRM. The developed method can be used for routine quality control analysis of titled drugs in pharmaceutical dosage form.     

Development and Validation of a Reversed-Phase High-performance Liquid Chromatography Method for Determination of Desmopressin in Chitosan Nanoparticles

A simple isocratic reversed-phase high performance liquid chromatographic method was developed for determination of released desmopressin from chitosan nanoparticles in the in vitro media. The chromatographic separation was achieved with acetonitrile/water (25:75, v/v), in which water contained 0.1% v/v trifluoroacetic acid with pH=2.5 as mobile phase, a Chromolith^® Performance RP-18e column (150×4.6 mm; 5 μm) kept at 40° and ultraviolet detection at 220 nm. The compound was eluted isocritically at a constant flow rate of 1.6 ml/min. The method was validated according to the International Conference on Harmonisation guidelines. The validation characteristics included accuracy, precision, linearity rang, selectivity, limit of detection, limit of quantitation and robustness. The calibration curve was linear (r>0.9999) over the concentration rang 0.5-100 μg/ml. The limit of detection and limit of quantitation in the release media were 0.05 and 0.5 μg/ml, respectively. The proposed method had an accuracy of and intra- and inter-day precision <4.2. Furthermore, to evaluate the performance of the proposed method, it was used in the analysis of desmopressin level in real samples containing chitosan nanoparticles in the in vitro media.     

Rapid and Sensitive Reverse-phase High-performance Liquid Chromatography Method for Estimation of Ketorolac in Pharmaceuticals Using Weighted Regression

A reliable, rapid and sensitive isocratic reverse phase high-performance liquid chromatography method has been developed and validated for assay of ketorolac tromethamine in tablets and ophthalmic dosage forms using diclofenac sodium as an internal standard. An isocratic separation of ketorolac tromethamine was achieved on Oyster BDS (150×4.6 mm i.d., 5 μm particle size) column using mobile phase of methanol:acetonitrile:sodium dihydrogen phosphate (20 mM; pH 5.5) (50:10:40, %v/v) at a flow rate of 1.0 ml/min. The eluents were monitored at 322 nm for ketorolac and at 282 nm for diclofenac sodium with a photodiode array detector. The retention times of ketorolac and diclofenac sodium were found to be 1.9 min and 4.6 min, respectively. Response was a linear function of drug concentration in the range of 0.01-15 μg/ml (R²=0.994; linear regression model using weighing factor 1/x²) with a limit of detection and quantification of 0.002 μg/ml and 0.007 μg/ml, respectively. The % recovery and % relative standard deviation values indicated the method was accurate and precise.