Validation.

To develop a fast and sensitive  UV spectrophotometric method for the quantitative estimation of Formaldehyde in Rivastigmine tartrate drug substances and validate as per ICH guidelines. The method was based upon the observation, that a characteristic colourresults upon addition of  solution of Pentane-2,4-dione , also known as acetylacetone . In acetic acid and ammonium acetate buffer condition, acetylacetone and formaldehyde react to form dimethyl pyridine. Dimethyl pyridine is slightly yellow and its absorption maximum in aqueous solution is λ 420 nmin Rivastigmine tartrate drug substance.The developed method resulted in Formaldehyde exhibiting linearity in the range 0.975 to 234 µg/g. The Intraday and interday precision is exemplified by relative standard deviation of 0.562 % and 0.757%. Percentage Mean recovery was found to be in the range of 98‐101%, during accuracy studies. The limit of detection (LOD) and limit of quantitiation (LOQ) were found to be 1.3 µg/g and 3.9 µg/g respectively.The present work was aimed to develop a visible spectrophotometric method, which is simple, sensitive, accurate and cost effective to evaluate the quality of the bulk drugs.

A new, simple, accurate, precise and reproducible RP-HPLC method for the simultaneous estimation of Sitagliptin Phosphate Monohydrate and Simvastatin has been developed and validated. Chromatographic separation was achieved using Neosphere C-18 column (4.6 mm 250mm,5mm) in gradient mode with a mobile phase consisting a mixture of acetonitrile:methanol:0.1% orthophosphoric acid in water (70:15:15v/v) at a flow rate of 1ml/min. The eluents were detected at 254 nm using UV detector. The retention time of Sitagliptin Phosphate Monohydrate and Simvastatin were found to be 2.09min and 7.79min respectively. The method was linear over the concentration range of 25-150μg/mL for Sitagliptin Phosphate Monohydrate and 10-60μg/mL for Simvastatin. The % recoveries for Sitagliptin Phosphate Monohydrate and Simvastatin were found to be 98.98-101.06% and 99.89-102.43% respectively. The developed method was validated for parameters like system suitability, specificity, linearity, accuracy, precision, ruggedness and robustness as per International Conference on Harmonization guidelines and the results were found to be within the limits. This validated method can be used for the routine quality control analysis of Sitagliptin Phosphate Monohydrate and Simvastatin in combined dosage form.

Use of herbal medicines is increasing day by day. Herbal medicine has become popular form of healthcare. The consumption of plant based medicines and other botanicals in the west have increased manifold in recent years. Application of modern scientific knowledge coupled with sensitive analytical technique is important for the quality evaluation and standardization of polyherbal formulations. Mucuna pruriens, an important medicinal plant with wide medicinal properties, is frequently used in a large number of traditional herbal preparations. The present study deals with development and validation of RP-HPLC method for the quantitative estimation of L-DOPA from polyherbal formulations. The method involves Chromatographic separation was performed with SHIMADZU HPLC (Model no. LC- 20 AD) equipped with isocratic pump and manual injector (20μl). Spinchrom chromatographic software was used for data acquisition. RP- C18 (250 mm X 4.6 mm X 5 micron) column was used for analysis. Mobile phase comprising of methanol and water in ratio of 20:80 v/v was filtered through 0.45 micron in membrane filter (Millipore) and degassed by sonication; flow rate of 1ml/min was maintained throughout the run. Column effluent was monitored at 280 nm with variable wavelength UV detector. Thus this developed HPLC method could be further used for the determination of L-dopa in the polyherbal formulations.

A simple, precise and sensitive reverse-phase high performance liquid chromatographic method was developed and validated for the simultaneous estimation of Ramipril and Metoprolol tartarate in pharmaceutical formulations. Chromatographic separation was performed on a High performance liquid chromatography equipped with auto sampler and UV detector. Good sensitivity for all analyte was observed with UV detection at wavelength of 218 nm, Separation was performed on a BDS Hypersil C18 (250 X 4.6mm) 5µm, using a mixture of 0.1% Triethylamine buffer pH 3.5 and Acetonitrile in the ratio of (10:90, v/v). The method results in excellent separation with good resolution between the two analytes. The within day variation %RSD values between Ramipril and Metoprolol tartarate were 0.55 and 0.82. The recovery was greater than 98% with %RSD less than 1.00. The method was validated according to ICH guidelines by performing linearity, accuracy, precision, limits of quantitation and selectivity. The results show the method is suitable for its intended use.

Two simple, sensitive and precise HPTLC and RP-HPLC methods were developed for the estimation of berberine from Coscinium fenestratum, and its formulation. For the determination of berberine by HPTLC method, precoated silicagel 60F254 on aluminium sheets and a mobile phase system comprising of n-butanol: glacial acetic acid : water (8:1:1 % v/v/v ) was selected. After development the plate was scanned and quantified at 350 nm. Linearity was found in the concentration range of 10 to 50 ng/spot (r=0.9992). Limit of detection was found to be 5 ng/spot and limit of quantification was found to be 10 ng/spot. In RP-HPLC method, separation was achieved on a Phenomenex, Luna, C18 column (150 x 4.6mm internal diameter, 5µ particle size) using a mobile phase consisting of potassium dihydrogen phosphate (pH - 2.5) (A)  : acetonitrile (B), where B was run in gradient programme (20% for 0.01-20min, 50% for 20-25min, 50% for 25-26min, 20% for 26-30min), at a flow rate of 1ml/min and the elute was monitored at 220nm. The calibration curve was obtained in the range of 100 - 500 µg/mL. The slope, intercept and correlation coefficient values were found to be 57588, 6041and 0.9959, respectively. The method was validated in compliance with ICH guidelines. Low relative standard deviation and good % recovery values of both the methods showed that the developed methods were highly precise, accurate and can be employed for the routine analysis of formulations containing berberine.

A simple, sensitive, precise, accurate and economic Q-absorption ratio method for the quantitative determination of betamethasone sodium phosphate and ofloxacin in bulk and combined dosage form has been developed and validated. Q absorption ratio method uses the ratio of absorbances at two selected wavelengths, one which is iso-absorptive point and other being the λ-max of one of the two components. Betamethasone sodium phosphate and ofloxacin shows an iso-absorptive point at 247nm in methanol, the second wavelength used was 289nm, which is λ-max of ofloxacin. The linearity was obtained in the concentration range of 2-12 ug/ml for both the drugs in distilled water. The proposed method is recommended for routine analysis. The results of analysis have been validated statistically and by recovery studies.

A simple, economic, selective, precise, and accurate High Performance liquid Chromatographic method for the analysis of Furazolidone in bulk drug and pharmaceutical formulations was developed and validated in the present study. The mobile phase consists of a mixture of potassium dihydrogen phosphate and acetonitrile 80:20. And Adjust pH 6.50±0.1 with dilute potassium hydroxide solution. This was found to give a sharp peak of Furazolidone at a retention time of 4.245 min. HPLC analysis of Furazolidone was carried out at a wavelength of 354 nm with a flow rate of 1.0 mL/min. The linear regression analysis data for the calibration curve showed a good linear relationship with a regression coefficient (r2) of 1 in the concentration range of 25 to 150 µg ml-1. The linear regression equation was y =46002x+40407. The developed method was employed with a high degree of precision and accuracy for the analysis of Furazolidone. The developed method was validated for accuracy, precision, robustness, detection and quantification limits as per the ICH guidelines.  The wide linearity range, accuracy, sensitivity, short retention time and composition of the mobile phase indicated that this method is better for the quantification of Furazolidone.

A simple, fast and precise reverse phase, isocratic HPLC method was developed for the separation and quantification of perindopril and indapamide in pharmaceutical dosage form. The quantification was carried out using YMC Column (150 x 4.6mm, 3µ particle size) and mobile phase comprised of ammonium dihydrogen phosphate pH 2.5 and acetonitrile in the ratio of 60:40% v/v and degassed under ultrasonication. The flow rate was 1.0 mL/min and the effluent was monitored at 230 nm. The retention time of perindopril and indapamide were 2.4 and 4.2 min respectively. The method was validated in terms of linearity, precision, accuracy and specificity. Linearity of perindopril and indapamide were in the range of 48 to 112 μg/mL and 15 to 35 μg/mL respectively. The proposed method is suitable for simultaneous determination of perindopril and indapamide in pharmaceutical dosage form. Key words: Perindopril and Indapamide, RP-HPLC, Validation.

The aim of present work was to develop an accurate, precise, reproducible and economical UV spectrophotometric method for estimation of Loratadine. This method was based on area under curve of UV spectrum between 241 to 251 nm and validated as per ICH guideline Q2 (R1). The method has followed linearity in the range of 5-30µg/ml. The value of correlation coefficient was 0.999. Satisfactory values of Percent relative standard deviation for the intra-day and inter-day precision indicated that method is precise. Results of the recovery studies (99.66 % to 99.95 %) showed accuracy of the method. LOD and LOQ were calculated as 0.581 µg/ml and 1.935 µg/ml, respectively. The developed method can be used for routine estimation of Loratadine in bulk and tablet dosage forms.

A simple, rapid, and precise RP-HPLC method for simultaneous analysis of Amlodipine besylate and Glimepiride in bulk and its pharmaceutical formulations has been developed and validated. Amlodipine besylate was separated from Glimepiride by using Grace Smart Altima C8 column (25 cm × 4.6 mm, 5-μm) with a mobile phase consisting of acetonitrile: 20mM phosphate buffer (55:45 (v/v), pH 3.5) a flow rate of 1 mL/min and detection wavelength at 230 nm.  Amlodipine besylate and Glimepiride were eluted with retention times of 5.47 min and 14.17 min respectively. The method was validated for accuracy, precision, linearity, specificity and sensitivity in accordance with ICH (Q2B) guidelines. The results of all the validation parameters were found to be within the acceptable limits. The calibration plots were linear over the concentration ranges from 70-3000ng/mL for Amlodipine besylate and 100-3000ng/mL for glimepiride. The limit of detection and limit of quantification were found to be 19.4ng/mL and 58.8ng/mL for amlodipine besylate, 25.6ng/mL and 76.2ng/mL for glimepiride respectively for both the drugs. From the results it is suggested that the method is simple, reproducible, accurate and precise. The method was successfully applied for the determination of content and the dissolution profile of the combined bilayer tablet dosage form.