RP-HPLC

This research manuscript describes simple, sensitive, accurate, precise and repeatable reverse phase high performance liquid chromatography method for the estimation of Chlorhexidine gluconate in mouthwash. The sample was analyzed by reverse phase ACE 5 C18 column (150 mm × 4.6 mm i.d, 5 μm particle size) as stationary phase; acetonitrile : methanol : triethyl Amine (0.1 %) PH 3.0 (22: 49: 29, v/v/v) as a mobile phase at a flow rate of 0.8 ml/min. Quantification was achieved with Photo Diode Array detector at 258 nm. The retention time for chlorhexidine gluconate was found to be 2.477 min. The linearity was obtained in the concentration range of 10 -80 µg/ml for chlorhexidine gluconate. The method was successfully applied to mouthwash because no chromatographic interferences from formulation excipients were found. The method retained its accuracy and precision when the standard addition technique was applied.

An isocratic, reversed phase-HPLC method was developed and validated for the quantitative determination of Ofloxacin and Ketorolac Tromethamine in combined-dosage form. A thermo hypersil BDS C18 (250mmx4.6mm, particle size 5µm) column with mobile phase containing water (pH - 2.8 adjusted with ortho phosphoric acid) and methanol in the ratio of 60: 40, v/v was used. The flow rate was 1.0 mL/min, column temperature was 30°C and effluents were monitored at 241 nm. The retention times of ofloxacin and ketorolac tromethamine were 4.671min and 5.751 min respectively. The correlation co-efficient values for both the ofloxacin and Ketorolac tromethamine were found to be 1. The proposed method was validated with respect to linearity, accuracy, precision, specificity, and robustness. Recovery of ofloxacin and Ketorolac tromethamine in formulations was found to be 100% confirms the non-interferences of the excipients in the formulation. Due to its simplicity, rapidness and high precision, the method can be successfully applied to the estimation of Ofloxacin and Ketorolac tromethamine in combined dosage form.

A rapid and stability indicating ion-pair reversed phase high performance liquid chromatographic method was developed for qualitative and quantitative estimation of three oxicam drugs: tenoxicam, meloxicam and lornoxicam. The method was validated according to ICH, FDA and USP guidelines with respect to accuracy, precision, specificity, linearity, solution stability, robustness, sensitivity and system suitability. The method was developed by using an isocratic condition of mobile phase comprising buffer pH 6.5 [tetra butyl ammonium hydroxide (0.008M) and sodium 1-heptane sulfonate (0.003M)] and acetonitrile in a ratio of 65:35 v/v ratio at a flow rate of 1.5 mL/min over C-18 (ODS, 250 x 4.6 mm) column at ambient temperature. The method showed linear response with correlation coefficient (r2) value of 0.999. The recoveries for all drugs were found more than 99% which demonstrated the accuracy of this method. Intraday and inter-day precision studies of the new method were less than the maximum allowable limit (RSD%£ 2.0). Forced degradation studies were carried on to check its stability indicating property. All the drugs gave sharp peaks within 7min with excellent symmetry and high resolution. Therefore, a rapid, sensitive and stability indicating ion pair RP-HPLC  method was developed for simultaneous separation of tenoxicam, meloxicam and lornoxicam in their any combination or in bulk raw materials.

Most of the synthetic drugs which induce side effect during overdose. The natural compounds are playing important role to reducing side effects of the synthetic drugs and also increase the immunity of recipients. The objective of the present study is to investigate the compatibility between Paracetamol as a synthetic compound with Spirulina as a natural compound.  The Paracetamol and Spirulina mixture were prepared in different ratio 1:1 (w/w) (Paracetamol: Spirulina), 1:2 (w/w) (Paracetamol: Spirulina) and 2: 1 (w/w) (Paracetamol: Spirulina). These samples were stored at different temperature (5 ˚C, 15 ˚ C, 30 ˚ C and 40 ˚ C) for one month. For every week samples were taken in the mixtures and the quantification of Paracetamol by using reversed phase liquid chromatography and estimation of chlorophyll A, B, total chlorophyll and total carotenoids in the mixture by using UV spectrophotometer. There was no significant loss of Paracetamol, chlorophyll, and carotenoids in their mixture. Both Paracetamol and Spirulina were stable under different temperature. Therefore it conclude that Paracetamol and Spirulina were compatible when mixed in a ratio of 1:1(w/w), 1:2 (w/w) and 2:1 w/w) 

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.

Cetrizine hydrochloride and Phenylephrine hydrochloride are used in combination in the treatment of allergy. The aim of this project work is to develop selective, accurate, specific and economic RP–HPLC method for simultaneous estimation of Cetrizine hydrochloride and Phenylephrine hydrochloride in Bulk and Tablet dosage form. The reverse phase C18 (250 mm × 4.6 mm i.d) column with 5 µm particle size was used. Acetonitrile: Water was taken as mobile phase was with a flow rate of 1 mL/min and detection was carried out at 222 nm. The retention time of Cetrizine hydrochloride and Phenylephrine hydrochloride was 1.956 min and 4.561 min respectively. The calibration curves were linear (>0.998) in the range of 5-25 μg/ml for Cetrizine hydrochloride and Phenylephrine hydrochloride. The Limit of Detection for Cetrizine hydrochloride and Phenylephrine hydrochloride was found to be 0.26 μg/mL and 0.51μg/mL. The Limit of Quantification was found to be 0.81μg/mL and 1.54 μg/mL respectively for Cetrizine hydrochloride and Phenylephrine hydrochloride. The developed method was simple, selective and precise and can be used for routine analysis of Cetrizine hydrochloride and Phenylephrine hydrochloride in tablet dosage form.

A new, simple, specific, sensitive, rapid, accurate and precise RP-HPLC method was developed for the estimation of Simvastatin in bulk and pharmaceutical formulations. Simvastatin was chromatographed on a hypersil C18 column (250x4.6mm I.D., particle size 5 μm) in a mobile phase consisting of sodium dihydrogen phosphate and Acetonitrile in the ratio 40:60 v/v. The mobile phase was pumped at a flow rate of 1.0 ml/min with detection at 239 nm. The detector response was linear in the concentration of 10-200 μg/ml. The intra and inter day variation was found to be less than 2%. The mean recovery of the drug from the solution was 99.39%. The proposed method is simple, fast, accurate, precise and reproducible hence, it can be applied for routine quality control analysis of Simvastatin in bulk and pharmaceutical formulations.

A simple isocratic RP-HPLC stability indicating method has been developed and subsequently validated for the determination of Irinotecan HCl and its related substance (SN-38) in pharmaceutical dosage forms as per ICH guidelines. The separation achieved on a reversed phase Phenomenex Luna C18 Column (5µ, 250 × 4.60 mm) as a stationary phase and 0.5% trichloro acetic acid: Acetonitrile: Methanol (60: 20: 20 v/v/v) as mobile phase at a flow rate of 1.0 ml/min. The UV detection was performed at 372 nm.  The retention time for Irinotecan HCl and SN-38 was found to be 8.65 and 7.30 min respectively. The detector response was linear in the concentration range of 30-150 µg/ml. The respective linear regression equation being Y= 5233.x + 13299 with R2 = 0.999. The percentage of Irinotecan HCl in pharmaceutical dosage form was found to be 100.5% and the percentage of related substance (SN-38) in formulation was found to be 0.19%. The limit of detection and the limit of quantification were found to be 0.014 µg/ml and 0.045 µg/ml respectively. The results of the study showed that, the proposed RP-HPLC method was simple, rapid, precise, accurate and stability indicating, which can be used for the routine determination of Irinotecan HCl and its related substance (SN-38) in pharmaceutical dosage form.

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.

A simple, accurate, precise and rapid reversed-phase high performance liquid chromatographic (RP-HPLC) method has been developed and subsequently validated for the estimation of Liraglutide in bulk and Parenteral Dosage form. The proposed method is based on the separation of drugs in reversed-phase mode using Waters HPLC 22695 model, Inertsil ODS column (250 x 4.6 mm, 5µm particle size).The optimum mobile phase consisted of methanol: phosphate buffer in the ratio of 85:15 v/v(Phosphate buffer pH 5.5) was selected as a mobile phase, flow rate of 1.0 ml/min and UV detection was set at 246 nm. The retention time was 3.25.The method was validated according to ICH guidelines. It was found to be accurate and reproducible. Linearity was obtained in the concentration range of 20-80 μg/ml . The percentage RSD for precision and accuracy of the method was found to be less than 2%. The proposed method can be successfully applied in the quality control of bulk and pharmaceutical dosage forms.