Development, validation, and simultaneous estimation of netilmicin and benzyl alcohol in netilmicin sulfate injections by RP-HPLC method

The fixed-dose combination of vildagliptin (VDG) and dapagliflozin (DGZ) is used in the treatment of type 2 diabetes mellitus. According to the literature survey, RP-HPLC and HPTLC methods have been reported for routine analysis of VDG and DGZ. These chromatographic methods have been developed using potentially neurotoxic and teratogenic solvents, which are unsafe for human and aquatic animal life and hazardous to the environment. These types of organic solvents shall be replaced or reduced during chromatographic analysis of drugs for the safety of human and aquatic animal life and the protection of the environment. The novel white analytical chemistry (WAC) approach has been introduced, which emphasizes robust, green, user-friendly, economical, and rapid analysis of drug samples. Hence, the WAC-based RP-HPLC method has been developed for the estimation of VDG and DGZ using lower toxic and economical solvents. The development of the RP-HPLC method includes the implementation of the analytical quality by design approach using principles of design of experiments to reduce organic waste generation and regulatory compliance of analytical method. The central composite design was applied for response surface modeling (RSM) and optimization of the RP-HPLC method. The method validation was carried out according to ICH Q2 (R1) guidelines. The fixed-dose combinations of VDG and DGZ were assayed, and results were found in compliance with their labeled claim. The published and proposed RP-HPLC methods were assessed for chromatographic analysis of VDG and DGZ using the Red-Green-Blue (RGB) model, AGREE calculator, Eco-Scale Assessment tool, GAPI software, and NEMI standards. The proposed method was found to be robust, green, economical, and user-friendly for chromatographic analysis of VDG and DGZ. The proposed method can be an economical and eco-friendly analytical tool in the pharmaceutical industry for quality control and routine analysis of fixed-dose combinations of VDG and DGZ. Hybrid principles of WAC and analytical quality by design to RP-HPLC method for simultaneous estimation of VDG and DGZ in their fixed-dose combinations.

The present study narrates the developed and validated simple, reliable, sensitive, precise and accurate Spectrophotometric and RP-HPLC methods for the simultaneous estimation of Fenbendazole and Niclosamide in pharmaceutical dosage form. In the first order derivative method 0.1 N methanolic HCl was used as diluent. The zero crossing point wavelengths selected for the analysis were 226 nm and 317 nm for Fenbendazole and Niclosamide, respectively and RP – HPLC method has been developed using 1% methanolic HCl as diluent. Separations of drugs were achieved on L1 C18 100 A 0 column (250 x 4.6 mm, 5 μ) using 2 gm potassium dihydrogen phosphate and acetonitrile (70:30, v/v) as mobile phase with flow rate 1.0 mL/min. The detection wavelength was 290 nm. Validation of developed methods was done according to ICH Q2 (R1) guideline. Calibration curve was linear over the concentration range of 3-9μg/mL (Fenbendazole) and 10-30 μg/mL (Niclosamide) for spectrophotometric method and 24 - 39 μg/mL (Fenbendazole) and 80 – 130 μg/mL (Niclosamide) for RP – HPLC method. The developed RP-HPLC and derivative spectrophotometric method were successfully applied for the quantitative determination of cited drugs in pharmaceutical dosage form. The correlation coefficients (r 2 ) value greater than 0.995. Accuracy of methods were determined by recovery studies and it was found to be 98 to 102 %. The % RSD values for all the validation parameters were less than 2.0 % for both the methods. The developed UV and RP-HPLC methods were compared by t - test and it was found that tstat value was less than tcritical value for all. Hence there was no significant difference between the developed methods.

Reversed-phase high-performance liquid chromatography (RP-HPLC) and thin-layer chromatography (TLC) methods have been developed and validated for simultaneous estimation of tamsulosin hydrochloride and finasteride in bulk drug and in combined dosage forms. RP-HPLC separation was achieved on a Phenomenex C18 column using methanol/0.02 mol L-1 ammonium acetate buffer/triethylamine (79.9 + 20 + 0.1, V/V/V) (pH 9.2) as mobile phase. TLC separation was achieved on an aluminium-backed layer of silica gel 60 F254 using toluene/methanol/triethylamine (9 + 1.5 + 1, V/V/V) as eluent. Quantification was achieved with photodiode array (PDA) detection at 235 nm over the concentration range 0.5-16 and 1-50 μg mL-1 with mean recovery of 99.8 ± 0.9 and 100.0 ± 0.8% for tamsulosin hydrochloride and finasteride, respectively, by the RP-HPLC method. Quantification was achieved with UV detection at 270 nm over the concentration range 100-2000 ng per spot and 250-5000 ng per spot with mean recovery of 98.9 ± 0.9 and 99.6 ± 0.7 % for tamsulosin hydrochloride and finasteride, respectively, by the TLC method. Both methods are simple, precise, accurate and sensitive and are applicable to the simultaneous determination of tamsulosin hydrochloride and finasteride in bulk drug and in combined dosage forms.

Background:Cinnarizine, an antihistaminic drug, is commonly formulated in combination with domperidone and with paracetamol for treatment and prevention of motion sickness and migraine.Objective:The aim of this work was to develop new, simple, precise and selective chromatographic methods (RP-HPLC and TLC-densitometric methods) for the determination of these drugs. These methods can be used as analytical tools in the routine examination in quality control laboratories.Methods:The first method was RP-HPLC method, the separation was carried out on an Inertsil® ODS- 3V C18 column (250 mm × 4.6 mm, 5 µm) using a mobile phase composed of methanol: acetonitrile (45: 55, v/v) at a flow rate of 1 ml/min. The detection was carried out at 220 nm. The second method was a TLC-densitometric method where the studied components were separated using a developing system composed of toluene: ethyl acetate: methanol: triethylamine (5: 4.3: 0.7: 0.5, v/v/v/v) on TLC silica gel 60 F254 plates, followed by densitometric scanning at 270 nm.Results:In RP-HPLC method, the peaks were sharp and well separated, the retention times were 5.25, 3.48 and 2.78 min, for cinnarizine, domperidone and paracetamol, respectively. Linearity was obtained over the concentration ranges 1-22, 0.75-16.5 and 25-550 µg/ml, for cinnarizine, domperidone and paracetamol, respectively. In TLC-densitometric method, good separation of spots and linear relationships were achieved over the concentration ranges of 0.2-2, 0.15-1.5 and 5-50 µg/spot, for cinnarizine, domperidone and paracetamol, respectively. Method validation was conducted according to ICH guidelines in terms of linearity, accuracy, selectivity, precision and robustness.Conclusion:The developed methods were applied for the determination of the cited drugs in tablets containing binary drug mixtures. The methods are simple and precise and can be used for routine analysis of the labelled drugs in combined dosage forms in quality control laboratories.

Development and validation of RP-HPLC and UV-spectrophotometric methods for rapid simultaneous estimation of amlodipine and benazepril in pure and fixed dose combination

Eco-friendly based HPLC and UV-spectrophotometric methods for simultaneous estimation of Efonidipine hydrochloride Ethanolate and Chlorthalidone in their dosage form

The current investigation was pointed at developing and progressively validating novel, simple, responsive and stable RP-HPLC method for the measurement of active pharmaceutical ingredients of Nirmatrelvir and Ritonavir and their related substances.RP-HPLC methodology was used for the quantitative determination of Nirmatrelvir and Ritonavir.Sunfire C18 Column, (5 µm, 4.6 mm X 250 mm) using mobile phase of 0.01N KH2PO4: Acetonitrile: Methanol (80:20:10 v/v) with flow rate of 1 ml/min (Detection wavelength 238 nm) was used for the present study.Using the impurity-spiked solution, the chromatographic approach was streamlined.The proposed method to be fast, simple, feasible and affordable in RS condition.During stability tests, it can be used for routine analysis of production samples and to verify the quality of drug samples during stability studies.

Purpose : The study was performed with the aim to design a Simple, precise and economical RP-HPLC method have been developed for simultaneous estimation of amitriptyline HCl and chlordiazepoxide in pharmaceutical formulation. Approach : RP-HPLC was developed using WATERS C18 250 mm × 4.6 mm (5 μm) column, utilizing a mobile phase of methanol: acetonitrile: 0.2 M orthophosphoric acid, pH 4.5 (adjusted with triethylamine) 56:24:20 ml (with a flow rate of 1ml/min) and U.V detection was carried out at 240nm. Optimization of this method was carried out as a function of mobile phase composition, pH and flow rate. Peak purity was assessed by spectral comparison method. Findings : RP-HPLC method shows linearity in the range of for 0.05-1.6 μg/ml and 0.01-0.32μg/ml amitriptyline HCl and chlordiazepoxide respectively. The LOD and LOQ values for amitriptyline HCl were 0.0250μg/ml and 0.0759 μg/ml and 0.0045 μg/ml and 0.0135 μg/ml for chlordiazepoxide. The results of % purity was found to be 99.97 ± 0.58 % w/w and 99.29 ± 0.59 % w/w for HPLC method and 98.87 ± 0.03 % w/w and 98.81 ± 1.04 % w/w for first derivative spectrophotometric method for amitriptyline HCl and chlordiazepoxide respectively. Conclusion : Results of descriptive statistics studies conclude that RP-HPLC method found to be more sensitive and convenient than first derivative spectrophotometric method.

Objective: The present work was undertaken with the aim to develop and validate a rapid and consistent RP-HPLC method in which the peaks will be appear with short period of time as per ICH Guidelines. The proposed method was simple, fast, accurate and precise method for the Quantification of drug in the dosage form, bulk drug as well as for routine analysis in Quality control. Method: RP-HPLC method was developed and validated for simultaneous estimation of Rosuvastatin and Fenofibrate in bulk drug and in combined dosage forms. The HPLC separation was achieved on a XTerra C18 (4.6 X 150mm, 5?m, Make: Thermosil) or equivalent in an Isocratic Mode. The mobile phase was composed of Phosphate Buffer (25%) whose pH was adjusted to 3.0 by using Ortho Phosphoric Acid and Methanol (75%) [HPLC Grade] Results: The flow rate was monitored at 1.0ml per min. The wavelength was selected for the detection was 254 nm. The run time was 7min. The retention time found for the drugs Rosuvastatin and Fenofibrate were 1.997 min., 3.238 min. and 4.042 min. respectively. The % recovery was found to be 98.46% - 101.79% for the drug Rosuvastatin. The % recovery was found to be 98.04% - 101.79% for the drug Fenofibrate. The linearity was established in the range of 80 to 120ppm for the drug Rosuvastatin and 2.4 to3.6ppm for the drug Fenofibrate. The LOD for the drugs Rosuvastatin and Fenofibrate were found to be 0.07µg/ml, 0.07µg/ml and 0.006µg/ml respectively. The LOQ for the drugs Rosuvastatin and Fenofibrate were found to be 0.23µg/ml, 0.24µg/ml and 0.02µg/ml respectively. Conclusion: The proposed method was adequate sensitive, reproducible, and specific for the determination of Rosuvastatin and Fenofibrate in bulk as well as in Tablet dosage form. The validation of method was carried out utilizing ICH-guidelines. The described RP-HPLC method was successfully employed for the analysis of pharmaceutical formulations containing combined dosage form. Overall the proposed method was found to be suitable and accurate for the Quantitative determination of the drug in Tablet dosage form. The method was simple, precise, accurate and sensitive and applicable for the simultaneous determination of Rosuvastatin and Fenofibrate in bulk drug and in combined dosage forms.

In present work, chemometric-assisted UV spectrophotometric methods as well as RP-HPLC method were developed for the simultaneous estimation of Ofloxacin and Tinidazole in their combined pharmaceutical dosage form. The two chemometric methods i.e. principle component regression (PCR) and partial least square regression (PLS) were successfully applied to quantify each drug in mixture using UV absorption spectra in range of 280 to 320nm at ∆λ of 0.5nm. Chemometric model development was done using 24 binary mixture solutions and 12 solutions were used for validation of model. The chemometric-assisted analysis does not require any prior separation step. In addition, RP-HPLC method was also developed using THERMOSIL C18 column with a mobile phase consisting ofAcetonitrile: Phosphate Buffer (85:15% v/v), flow rate of 1 ml/min and quantification was achieved using UV detector at 300 nm. The methods were successfully applied for the simultaneous determination of these drugs in synthetic mixture. The results obtained for analysis by PCR and PLS methods were compared with RP-HPLC method and a good agreement was found.

Benzyl Nicotine (BN) and etofenamate (ETO), when combined with methylparaben (MP) or benzyl alcohol (BA), are highly effective for treating muscle and joint pain. This study presents a newly developed and verified RP-HPLC method for measuring ETO and BN in the presence of MP or BA in their dosage forms. The approach utilizes eight environmentally friendly tools and the BAGI white tool to assess their eco-friendliness and effectiveness. The method uses a photodiode array PDA detector at 254 nm and symmetry column (C18, 150 mm × 4.6 mm, 5μm) in an isocratic process with a mobile phase of 0.1% orthophosphoric acid and methanol in a (45:55, v/v) adjusted pH 6.5 with 0.2% triethylamine solution at a flow rate of 1.5 mL/min, separation was accomplished in RP-HPLC. The calibration curves exhibited linearity across the concentration levels specified in the approach. Therefore, the procedure has been effectively applied in laboratory evaluation for medications in the presence of MP or BA, showcasing the exceptional selectivity of this strategy. The recommended approach was executed using the Six Sigma methodology, resulting in a Cpk value greater than 1.33. The procedures were successfully validated based on the ICH standards.

Background Sulfadiazine and Pyrimethamine is a combination drug of choice to treat malaria and used in those living in or will be travelling to an area where there is chance of getting malaria. Objective The main objective of the study was to validate the simultaneous estimation and the method established with resisting small changes in flow rate and ratio of organic compounds and proposed method ensures its use in routine quality control analysis of pharmaceutical formulation. MethodologyA HPLC AllianceWater 2695 with UVVIS DetectorPDA detector UV lab India UV 3000 series and Zodiac C18 250 mm times 4.6 mm times 5 microm column were used. A new method was established for simultaneous estimation of Sulfadiazine and Pyrimethamine by RP-HPLC method. The chromatographic conditions were successfully developed for the separation of Sulfadiazine and Pyrimethamine by using Zodiac sil C18 column 4.6times150 mm 5micro flow rate was 1mLmin mobile phase ratio was 7030 vv methanolphosphate buffer KH2 PO4 and K2 HPO4 phosphate pH 3 pH was adjusted with orthophosphoric acid detection wavelength was 240 nm. Results The results were in good agreement with those obtained with official HPLC with maximum absorption of 240 nm by preparing mobile phase 7030 methanolphosphate buffer with flow rate 1 mLmin and was run for 10 minutes by selecting column Zodiac silica RP C18 4.5times100 mm 3.0 microm of ambient temperature. All the results were obtained with good precision accuracy and robustness as per ICH guidelines. Conclusion It can be concluded that the proposed RPHPLC method was accurate precise sensitive specific robust and reproducible for the simultaneous analysis of Sulfadiazine and Pyrimethamine with less tailing factor and also economical. Zodiac sil C18 column 4.6times150mm5micro flow rate was 1mLmin mobile phase ratio was 7030 vv methanol phosphate buffer KH2 PO4 and K2 HPO4 phosphate pH 3 pH was adjusted with orthophosphoric acid detection wavelength was 240 nm.

For the quantitative determination of Fulvestrant, Benzyl alcohol, and Benzyl benzoate in Fulvestrant injection, an original RP-HPLC approach was developed. The gradient method was developed using HPLC and a Phenomenex Luna C8, 150 × 4.6 mm, i.d 3.0 μm particle size column with a gradient programme of mobile phases A and B. With a flow rate of 1.5 mL/minute, injection volume of 10 μL, and column temperature of 35°C, UV wavelength detection at 254 nm for Benzyl alcohol and Benzoyl Benzoate and 280 nm for Fulvestrant, mobile phase-A consists of DI water and mobile phase-B consists of Acetonitrile. The current study describes a single HPLC method for developing a Fulvestrant (Active), Benzyl alcohol (Cosolvent), and Benzyl Benzoate (Cosolvent) assay for Fulvestrant injection. The assay method was determined to be suitable for quantifying three components in the pharmaceutical product and was verified according to ICH guidelines.

A rapid, sensitive, and accurate stability indicating HPLC method was developed for the simultaneous determination of triamcinolone acetonide and benzyl alcohol in pure form, degradation products and pharmaceutical preparation. The separation was carried out on RP BDS Hypersil® C18 column (150 x 4.60 mm, 5μm) using an isocratic mobile phase composed of acetonitrile: 0.05 M phosphate buffer PH 3.50 (55 : 45). Both benzyl alcohol and triamcinolone acetonide quickly eluted at 1.67 min and 3.42 min, respectively, with a flow rate of 1.50 mL /min and UV detection at 254 nm. The linearity was in the range of 1 – 50 µg/mL for triamcinolone acetonide and 2 - 10 µg/mL for benzyl alcohol. The method has been validated for linearity, accuracy, precision, specificity, limit of detection, limit of quantification, robustness, and ruggedness as per the ICH guidelines. Finally, the method was compared statistically with reference methods indicating that there is no significant difference between them in respect of precision and accuracy.

The current work aims to create a new, simple, rapid, quick, accurate, efficient, and reproducible RP-HPLC method for the simultaneous analysis of Lobiglitazone and Glimperide. In compliance with ICH criteria, the developed approach will undergo validation. Through chromatographic condition optimisation, the RP-HPLC method will be used to establish the analytical method for the simultaneous quantification of Glimperide and Lobiglitazone. For a number of factors included in ICH standards, Q2, the developed approach has been validated in accordance with ICH guidelines (R1). According to a survey of the literature, no analytical technique has been published for the simultaneous measurement of glipride and lobiglitazone using RP-HPLC. The documented analytical techniques for chemicals, either alone or in conjunction with other dosage forms, are spectrophotometer, HPLC, and HPTLC. As a result, it was believed that novel analytical techniques were required for the simultaneous measurement of glimeride as well as lobiglitazone in pharmaceutical dosage forms.