Simplified and Rapid Determination of Primaquine and 5,6-Orthoquinone Primaquine by UHPLC-MS/MS: Its Application to a Pharmacokinetic Study

The method for the determination of primaquine (PQ) and 5,6-orthoquinone primaquine (5,6-PQ), the representative marker for PQ active metabolites, via CYP2D6 in human plasma and urine has been validated. All samples were extracted using acetonitrile for protein precipitation and analyzed using the ultra-high-performance liquid chromatography–tandem mass spectrometry (UHPLC-MS/MS) system. Chromatography separation was carried out using a Hypersil GOLDTM aQ C18 column (100 × 2.1 mm, particle size 1.9 μm) with a C18 guard column (4 × 3 mm) flowed with an isocratic mode of methanol, water, and acetonitrile in an optimal ratio at 0.4 mL/min. The retention times of 5,6-PQ and PQ in plasma and urine were 0.8 and 1.6 min, respectively. The method was validated according to the guideline. The linearity of the analytes was in the range of 25–1500 ng/mL. The matrix effect of PQ and 5,6-PQ ranged from 100% to 116% and from 87% to 104% for plasma, and from 87% to 89% and from 86% to 87% for urine, respectively. The recovery of PQ and 5,6-PQ ranged from 78% to 95% and form 80% to 98% for plasma, and from 102% to from 112% to 97% to 109% for urine, respectively. The accuracy and precision of PQ and 5,6-PQ in plasma and urine were within the acceptance criteria. The samples should be kept in the freezer (−80 °C) and analyzed within 7 days due to the metabolite stability. This validated UHPLC-MS/MS method was beneficial for a pharmacokinetic study in subjects receiving PQ.

DEVELOPMENT OF LIQUID CHROMATOGRAPHIC METHOD FOR ESTIMATION OF ACAMPROSATE CALCIUM AND BACLOFEN COMBINATION USED IN TREATMENT OF NEUROLOGICAL DISORDERS

A specific, accurate, precise, and reproducible liquid chromatographic method has been developed and validated for the estimation of acamprosate calcium and baclofen in combination. The separation was achieved using stationary phase Phenomenex C18 column (150 mm× 4.6 mm.) in isocratic mode, with mobile phase containing 0.05 M potassium dihydrogen orthophosphate buffer (pH 7) : acetonitrile (10:90 V/V), at a flow rate of 1.0 mL/min and effluents were monitored at 210 nm. The retention time of acamprosate calcium and baclofen were found to be 1.9 min and 5.3 min, respectively. The linearity for acamprosate calcium and baclofen were in the range of 2-64 µg/mL and 1.2- 38.4 µg/mL, respectively. The method was validated as per ICH guideline. The recoveries of Acamprosate calcium and baclofen were found in the range of 98.90 - 100.13 % and 98.60 -100.02 %, respectively. The method was successfully applied for the determination of both the drugs in combination.

Validated RP-HPLC Method for Simultaneous Estimation of Domperidone and Naproxen in Bulk Drug and Formulations

A new , precise , accurate and reproducible RP – HPLC method for simultaneous estimation of bulk and pharmaceutical formulations . Separation of Naproxen and Domperidone was successfully achieved. THERMO HYPERSIL , C 18 , 250mm X 4.6mm , 5μm , or equivalent in an isocratic mode utilizing Acetonitrile : Methanol : Water (20: 60:20 ) at a flow rate of 1.0 mL/ min and elute was monitored at 284nm , with a retention time of 2.341 and 5.225 minutes for Naproxen and Domperidone respectively. The method was validated and the response was found to be linear in the drug concentration range of 1μg /mL to 6μg/mL. The values of correlation coefficient was found to be 0.988 for Naproxen and 0.998 for Domperidone respectively. The LOD and LOQ for Naproxen was found to be 0.56, 1.71 respectively . The LOD and LOQ for Domperidone was found to be 1.49 , 4.5 respectively. This method was found to be good percentage recovery for Naproxen and Domperidone were found to be 98% and 101.6% respectively indicates that the proposed method was highly accurate. The specificity of method shows good correlation between retention times of standard with the sample. The method was extensively validated according to ICH guidelines for Linearity , Accuracy , Precision , Specificity , and Robustness .

High Response Bio-Analytical Validation approach of Nadolol and Bendroϑlumethiazide by LC-MS/MS on Rat plasma

A very strong responsive and straight forward the LC-MS / MS assay was developed to and witnessed for that gradation in Nadolol and Bendroflumethiazide in rat plasma. The chromatographic conditions involve isocratic mode using Waters symmetry C18 (150x4.6 mm, 3.5 microns) column. A 0.1 per cent Smartphone process OPA (orthophosphoric Acid) Acetonitrile and in 60:40 is employed, and therefore the detection was administered during +ve mode of electrospray ionisation by using MS. The valid method had validated in the linear range of 8-160 ng/ml Nadolol and 1-20 ng/ml bendroflumethiazide, the precise values considered to be intraday and interday within the acceptable limit. Here these drugs are extracted from the rat plasma by using the liquid-liquid extraction. And these drugs are found stable. Through freeze Thaw, sampler app, vehicle sampler and top of the bench for the future studies. Form of liquid chromatography-tandem mass spectrometry and checked in compliance with guidelines for quantification of food and drug administration Nadolol and Bendroflumethiazide plasma in rats using D6-Nadolol and D6-Bendroflumethiazide as internal standards utilising LC-MS incorporated with quadrupole spectrometer by using electrospray ionisation technique. The target of this analysis is to be done work out the appropriateness of this approach to Nadolol and Bendroflumethiazide Applying Nadolol and bendroflumethiazide and their internal requirements at various quantification stages and retaining different parameters such as instrument durability, precision and accuracy, sample preparation techniques, instrument synchronisation, recovery and matrix effect.

Development and Validation of Teneligliptin Stereoisomers by HPLC Using Cellulose Based Immobilized Polysaccharide Chiral Stationary Phase

Background:: There is no single chiral method for the quantitation of teneligliptin stereoisomers by high performance liquid chromatography (HPLC). Hence, there is a need for the quantification of teneligliptin (TNGP) and its stereoisomers. Objective:: The main aim of the research work is to develop a novel simple, selective, precise and accurate HPLC method for separation of TNGP and its stereoisomers. Methods:: Different screening trials were executed by changing the mobile phase compositions to normal phase and polar mode and also by utilizing the different immobilized polysaccharide chiral columns like CHIRALPAK IA, IC, ID, IE, IF and IG. All the stereoisomers were eluted with high resolution, on CHIRALPAK IC-3 (4.6×250 mm), 3 μm chiral stationary phase (CSP) with flow rate of 0.7 mL/min. The chromatographic system was processed with isocratic mode comprising ethanol: acetonitrile: ethanolamine in the proportion of 90:10:0.1% v/v/v with column oven temperature of 15 °C and detection wavelength of 250 nm. Results:: The limit of detection (LOD) and limit of quantification (LOQ) values of TNGP(API), R,S-isomer, S,R-isomer and R,R-isomer were found to be 0.036/0.11, 0.029/0.09, 0.038/0.011 and 0.020/0.06 μg/mL, respectively. The method was found to be precise, accurate and linear (R2 > 0.999). Conclusion:: The developed method also successfully applied for the quantification of bulk drug without any interference with the extraneous components. Hence, the method can be utilized successfully in the pharmaceutical organizations for the separation and quantification of TNGP isomers.

DEVELOPMENT AND VALIDATION OF RP-HPLC METHOD FOR ESTIMATION OF EDOXABAN TOSYLATE IN TABLET DOSAGE FORMS

A rapid, simple and precise reversed phase high performance liquid chromatography (RP-HPLC) method was developed for the estimation of edoxabantosylate in tablets. The quantification was carried out using a Phenomenex C-18 column (250×4.6 mm i.d., 5µm particle size) in isocratic mode with mobile phase comprising of ammonium acetate buffer andacetonitrile in the ratio of 50:50 (V/V) at a flow rate 1 mL/min. The eluent was monitored at 240 nm. The retention time of the drug was 3.486 min. The calibration curve was linear in the concentration range of 5-25 µg/mL and per cent recovery ranged from 98.25-101.6.The developed method was validated as per ICH guidelines and the results obtained were satisfactory.The method can be applied for routine quality control analysis of edoxabantosylate in tablet dosage forms.

A Simple RP-HPLC Method to Simultaneously Assay the Contents of Lamivudine, Tenofovir, and Nevirapine in Fixed Dose Combined Oral Antiviral Medicines

An accurate and rapid reverse HPLC method has been developed and validated for the simultaneous quantification of lamivudine, nevirapine, and tenofovir disoproxil fumarate. Suitable separation was achieved on Phenomenex Synergi C18 (250 × 4.6 mm, 4 μm) using mobile phase, methanol (50%): ammonium acetate buffer (adjusted to pH 2.80) (40%): acetonitrile (10%) in an isocratic mode. The drugs were detected at 270 nm with a flow rate of 1.0 ml/min, and the retention times were found to be 3.26, 5.42, and 7.55 minutes for lamivudine, nevirapine, and tenofovir disoproxil fumarate, respectively. The developed method was validated per ICH guidelines. Good linearity was obtained within the concentration ranges of 10–59 µg/ml, 7–42 µg/ml, and 15–90 µg/ml with a correlation coefficient of not less than 0.990. The % RSD values for precision (intraday and interday) and accuracy studies were found to be less than 2%. The results obtained from quantitative analysis conform to USP content requirements for marketed tablet dosage forms, RICOVIR-LN, and tenofovir disoproxil fumarate/lamivudine tablets. The method is therefore useful for routine quality control of antiretroviral tablet dosage forms containing tenofovir disoproxil fumarate, lamivudine, and nevirapine.

Developing an isotope dilution UHPLC–MS/MS method to quantify linezolid in human plasma: application to therapeutic drug monitoring

Aim: To optimize clinical efficacy and reduce the drug-exposure-related toxicity of linezolid, whose concentrations show wide inter-variabilities, a simple and reliable quantitative assay for therapeutic drug monitoring is necessary. Results: A UHPLC–MS/MS assay has been established for determination of linezolid in human plasma and fully validated according to the US FDA guidelines. After a simple, isotope-dilluted precipitation with methanol, the analytes were separated by a straightforward isocratic mode and the MS/MS was conducted under the ESI+ mode fitted with SRM. The calibration curves proved acceptable linearity in the range of 0.1–30.0 µg/ml. Conclusion: The present assay is currently used in routine clinical practice, being applied to therapeutic drug monitoring and helps to optimize individual dosing regimens and manage adverse effects in ICU patients.

Efficient Validated HPLC/UV Method for Determination of Valsartan and Atenolol in Dosage Form And In Vitro Dissolution Studies

The main purpose of this study was to develop and validate an efficient HPLC/UV method for determination of valsartan and atenolol and to introduce the dissolution profiles of tablets; The resolution of peaks was best achieved with Zorbax C8 (4.6 mm i.d. X 150 mm, 5 μm) column. Samples were chromatographed in a isocratic mode (methanol and 25 mM solution potassium dihydrogen phosphate pH 7.3 (55:45, V/V)), pumped with 1.0 mL/min at 40 °C set temperature of column oven, with UV detector set to 225 nm wavelength; The total chromatographic run time was 6 minutes. The retention time of valsartan is 1.753 min, atenolol – 3.064 min. Linearity was examined and proven at different concentration levels in the range of working concentration of valsartan ( 0.16-0.96 mg/mL) and atenolol (0.2–1.2 mg/mL). The high value of recoveries obtained for valsartan and atenolol indicates that the proposed method was found to be accurate. In all three dissolution media the releases of valsartan and atenolol are more than 85% in 15 min A rapid, simple, accurate, selective, and sensitive method was developed for the determination of valsartan and atenolol in dosage forms. The method was strictly validated according to the ICH guidelines. Acquired results demonstrate that proposed strategy can be effortlessly and advantageously applied for routine quality control of drugs and in vitro dissolution study.

Development of an Efficient Analytical Method for the Extraction and Analysis of Biocide Contents from the Textile Test Specimens on LC-DAD

Biocides are frequently used in the manufacturing of textiles that are in direct contact with human skin. Recently regulated biocides do not have validated methods for testing; so, their presence cannot be estimated in the consumer products. Hence a rapid method was developed for the separation and quantitative analysis of biocide contents (2-methyl-4-isothaizolin (MIT), 5-chloro-2-methyl-4-isothaizolin-3-one (CIT), 2-octo-4-isothaizolin-3-one (OIT), and 5-chloro-2-(2,4-dichlorophenxy) phenol (triclosan)) from the textile test specimens. Test specimens were extracted with methanolic sonication and purified by centrifugation and filtration. Biocide contents were separated at C18 column with 0.4% acetic acid: methanol (1 : 1 v/v) under isocratic mode and detected at 280 nm wavelength. Pretreatment factors such as extraction solvent, extraction method, dilution ratio, and extraction time were optimized initially and plotted calibration curve showed regression (r2 ≥ 0.9995) in the range of 1.0–5.0 mg L−1. Recoveries were between 95% and 108% with the relative standard deviation ≤ 4%. Limits of detection (LODs) were between 0.06 mg L−1 and 0.12 mg L−1 and limits of quantification (LOQs) were between 0.21 mg L−1 and 0.38 mg L−1. From the results, conclusion was made that the method can achieve the purpose of quantitative detection and the analysis of real test specimens verified the reliability of this method.