RP-HPLC Method For Simultaneous Determination Research Articles

Novel RP-HPLC method for simultaneous determination of dapagliflozin and teneligliptin in tablet formulation and identification of degradation products by LC-MS/MS

Abstract Background Diabetes mellitus affects millions globally, necessitating effective management strategies. Glenmark Pharmaceutical Limited introduced a fixed-dose combination of teneligliptin and dapagliflozin in 2022 to address this need. However, existing methods for their simultaneous detection are limited, lacking forced degradation studies essential for assessing drug stability. Results A stability-indicating RP-HPLC method was developed for the simultaneous determination of dapagliflozin and teneligliptin in pharmaceutical formulations. The separation was efficiently achieved employing a Zorbax Eclipse Plus C18 column (150 mm × 4.6 mm, 5 µm). The mobile phase comprised a mixture of 10 mM ammonium acetate buffer in water, methanol, and acetonitrile in a suitable proportion and delivered at a flow rate of 0.6 mL/min. Detection of the isocratic eluents was performed at 224 nm using a photodiode array (PDA) detector. Validation against various stress conditions, as per ICH guidelines, revealed significant degradation of teneligliptin primarily under acidic, basic, and oxidative stress. Moreover, liquid chromatography tandem mass spectrometry (LC-MS/MS)-based characterization was conducted for the primary degradation products of teneligliptin generated under acidic, basic, and oxidative conditions. Conclusions The developed RP-HPLC method with a stability-indicating approach provides an efficient means for the simultaneous determination of dapagliflozin and teneligliptin in pharmaceutical formulations. The significant degradation was observed under various stress conditions and also successfully separated the degradation products by this method. The proposed method directly applied for the characterization of degradation products and based on LC-MS/MS data the structure of degradation products was generated and also degradation pathways under various stress conditions were predicted. The proposed method ensured accurate and precise results in quality control process in pharmaceutical industry, and adherence to ICH validation guidelines affirms its reliability in assessing the stability of these compounds.

Development and Validation of a RP-HPLC Method for the Simultaneous Determination of Silver Sulfadiazine and Sodium Hyaluronate in the Presence of Methyl and Propyl Paraben in a Pharmaceutical Cream for Burns.

A direct and precise isocratic RP-HPLC method for simultaneous determination of silver sulfadiazine (SSD) and sodium hyaluronate (SH) in the presence of methyl (MP) and propyl parabens (PP) was developed and validated. Agilent chromatograph with X-Select C18 column (250 × 4.6mm2, 5μm) was used. Chromatographic separation was achieved using a mobile phase consisting of acetonitrile and 0.05M phosphate buffer (pH5.0 to which added triethyl amine 0.5ml/L), at a ratio 35: 65v/v. Elution was used at flow rate of 1.0mL/min at ambient temperature with UV detection at 205nm. The retention times for SH, SSD, MP and PP were 1.49, 3.3, 6.7 and 19.5min, respectively. The presented chromatographic method was fully validated in accordance with ICH requirements, it was valid over linearity ranges of (0.80-100.00μg/mL) and (3.20-100.00μg/mL) for SSD and SH, respectively. Acceptable precision and accuracy were obtained for concentrations over the standard curve ranges and the sensitivity of the method, as the limits of detection and quantification for each active ingredient was also determined. The validated method was successfully applied for the quantification of SSD and SH in pharmaceutical cream formulation and the mean recovery %±SD were 100.93 ± 0.985 and 100.05 ± 0.668 for SSD and SH; respectively, indicating satisfactory accuracy of the method.

Utilizing experimental design and desirability function in optimizing RP-HPLC method for simultaneous determination of some skeletal muscle relaxants and analgesics

An experimental design and response surface methodologies using Plackett–Burman and Box-Behnken designs were applied for selecting and optimizing the most appropriate parameters which significantly affect the separation and quantitative estimation of five skeletal muscle relaxants and four analgesic drugs (baclofen, methocarbamol, dantrolene sodium, orphenadrine citrate, cyclobenzaprine hydrochloride, ketoprofen, etoricoxib, ibuprofen, and mefenamic acid) with a relatively short duration of analysis in a single run. For the separation of the nine drugs, an INERTSIL ODS-V3-5 µm C18 column (250 × 4.6 mm I.D.) was used with the optimum mobile phase conditions (45.15 mM ammonium acetate buffer pH 5.56 adjusted with acetic acid, acetonitrile, and methanol in a ratio of 30.5:29.5:40, v/v/v with a flow rate of 1.5 mL/min) and UV-detection at 220 nm. The optimized method was successfully subjected to the validation steps as described in ICH guidelines for linearity, precision, accuracy, robustness, and sensitivity. The optimized and validated method was effectively applied to determine the content of the studied drugs in their pharmaceutical preparations and to expand its applicability to the counterfeit estimation of etoricoxib in different brands of tablet dosage forms.

Whiter and Greener RP-HPLC Method for Simultaneous Determination of Dorzolamide, Brinzolamide, and Timolol Using Isopropanol as a Sustainable Organic Solvent in the Mobile Phase

A sustainable reversed-phase chromatographic method has been developed and validated for the simultaneous determination of three active pharmaceutical ingredients, dorzolamide, brinzolamide, and timolol, used to treat glaucoma. The eco-friendly solvent isopropanol has been used as an organic mobile phase constituent. According to the Hansen space green solvent selection tool, isopropanol has a G score of 6.5, comparable to ethanol, which has a G score of 6.6. The mobile phase consists of isopropanol: aqueous sodium acetate buffer (0.1 M, pH 4.25) in the ratio of 10:90 (v/v). The flow rate was maintained at 1 mL/min. Dorzolamide and brinzolamide were detected at 254 nm, and timolol was detected at 295 nm. A high-purity silica with a polymeric C18 modification column (150 × 4.6 mm, 5 µm particle size) was used for this separation. The three compounds were eluted within 8 min. The method was validated according to ICH guidelines. The calibration curves were linear in the range of 20–70 µg/mL, 40–140 µg/mL, and 20–70 µg/mL for dorzolamide, brinzolamide, and timolol, respectively. The LODs were found to be 1.61 µg/mL, 1.60 µg/mL, and 3.16 µg/mL for dorzolamide, brinzolamide, and timolol, respectively. Good accuracy and precision were obtained for the three compounds. The greenness and whiteness of the method were indicated using the AGREE, ChlorTox, and RGB12 tools.

Development of the RP-HPLC Method for Simultaneous Determination and Quantification of Artemether and Lumefantrine in Fixed-Dose Combination Pharmaceutical Dosage Forms.

Developing countries face enormous challenges with substandard and falsified antimalarial drugs. One specific issue is the lack of a simple, cost-effective, and robust HPLC method to simultaneously determine and quantify the active pharmaceutical ingredients (APIs) in fixed-dose artemether-lumefantrine pharmaceutical dosage forms. The current study developed a novel, simple, sensitive, precise, accurate, and cost-effective RP-HPLC method for the simultaneous determination and quantification of artemether and lumefantrine in pharmaceutical dosage forms. The HPLC analysis was carried out on an Agilent 1260 Infinity Series HPLC system equipped with an ODS Intersil-C8 (150 × 4.6 mm) 5.0 µm column, by isocratic elution. The mobile phase composition consisted of acetonitrile and 0.05% orthophosphoric acid buffer of pH 3.5 in the ratio of 70 : 30 v/v. The analysis was performed at a 1 mL/min flow rate and a column temperature of 25°C. The total run time was 6 minutes. The detection was done with a variable wavelength detector (VWD) at an isosbestic point wavelength (λ) of 210 nm. The developed method was validated according to the ICH guidelines concerning system suitability, specificity, linearity, accuracy, precision, and robustness. The system suitability of the developed method revealed satisfactory theoretical plates and symmetry factors. The method proved to be specific, with no interference of mobile phase or excipients. The calibration plot exhibited linearity over the concentration range of 275-1925 μg/mL with R2 = 0.9992 for artemether and a range of 150-1050 μg/mL with R2 = 0.9985 for lumefantrine. The accuracy of the method, determined by the recovery study, was 99.79-100.16% for artemether and 99.04-99.50% for lumefantrine. The % RSD values for intraday precision were 0.175 and 0.203, while interday precision values were 0.340 and 0.554 for artemether and lumefantrine, respectively. The method demonstrated robustness when subjected to slight modifications in the flow rate, column temperature, and mobile phase composition. The developed analytical method proved satisfactory as per ICH guidelines and hence can be used for the determination and quantification of artemether and lumefantrine in bulk drug and pharmaceutical dosage forms.

Using RP-HPLC Method for Simultaneous Determination of Fructose and Mannitol in Anidulafungin for Injection

Using RP-HPLC Method for Simultaneous Determination of Fructose and Mannitol in Anidulafungin for Injection

Development and Validation of RP-HPLC Method for Simultaneous Determination of Aceclofenac and Piperine in Rat Plasma to Study Pharmacokinetic Parameters

The BCS class II medication aceclofenac is poorly soluble in water and undergoes significant metabolism in human hepatocytes and microsomes. It has a 15% oral bioavailability, which is low. Piperine is a naturally occurring bioenhancer that is used to increase the oral bioavailability of many different medications. In this study, the pharmacokinetics of aceclofenac (20 mg/kg) alone and in combination with piperine (10 mg/kg) in rats were tested. Despite their distinct chemistry and physical properties, in this study, piperine was used to enhance the bioavailability of aceclofenac, followed by the development and validation of HPLC method to determine their simultaneous measurement in rat plasma. Plasma samples were processed by solid phase extraction and subjected to HPLC analysis. The analysis identified retention time for aceclofenac and piperine to be 5.3 and 14.4 minutes respectively. We found the linear concentration range of aceclofenac in rat plasma between 0.1 to 20 μg/mL. The correlation coefficient (R2) determined was 0.9995. Cmax was increased from 23.59 to 48.52 μg/mL. Including piperine with aceclofenac in formulation can improve its oral bioavailability.

Glycerol-Based Green RP-HPLC Method for Simultaneous Determination of Methionine and Paracetamol in Pharmaceutical Tablets

Methionine and paracetamol are combined in dosage forms due to the ability of methionine to compensate the deficiency in glutathione in hepatic patients. In this work, an innovative simple time-saving green RP-HPLC method using glycerol as a solely green mobile phase with water for the first time was developed for the simultaneous determination of methionine and paracetamol in their standard and in Hepamol® tablets. The chromatographic conditions were optimized using factorial design with the aid of Minitab 17® Software. The method was performed on a C18 column at 38 °C, and a mobile phase consisting of glycerol and phosphate buffer (pH 2.4) (40:60, v/v), using diode array detector at 210 nm. The mixture was separated in 5 min. The developed method was validated in accordance with ICH requirement over linearity ranges of 10–90 μg/mL for both drugs, and LODs were 3.33 μg/mL for both methionine and paracetamol. Glycerol has high safety, low UV cut-off point, low flammability, and its viscosity can be manipulated when diluted with water. The procedure was compared to the reported reversed phase liquid chromatography method in the terms of their greenness with the green analytical procedure index and the analytical eco-scale. This work breaks new ground for scientists to use glycerol in greener RP-HPLC applications.

AGREE and ESA for Greenness Assessment of a Novel Validated RP-HPLC Method for Simultaneous Determination of Aspirin, Warfarin and Clopidogrel in Rat Plasma: Application to Pharmacokinetic Study of the Possible Interaction between the Three Drugs.

The green profile of the developed method is assessed and compared with previously reported methods. Percutaneous coronary intervention is a procedure where a catheter is utilized to place a stent in order to facilitate opening of the blood vessels in the heart. Triple antithrombotic therapy includes oral anticoagulation as warfarin and dual antiplatelet therapy (composed of aspirin and clopidogrel bisulfate). The aim of the current study was to evaluate the pharmacokinetic parameters of ASP, WAR and CLP and to investigate the possible interaction between the three drugs upon co-administration in rats. A selective and precise RP-HPLC method was developed for the simultaneous determination of ASP, WAR and CLP in rat plasma. Pharmacokinetic study was conducted in rats that received ASP, WAR and CLP as an application of the developed method. From the statistical evaluation of the pharmacokinetic parameters, it was observed that the co-administration of ASP, WAR and CLP significantly increased the ASP and CLP bioavailability in rats. A significant drug-drug interaction was confirmed in the current study. The elevated Cmax of ASP and CLP upon the co-administration of ASP, WAR and CLP may explain the reported bleeding.

Development and Validation of Stability Indicating RP-HPLC Method for Simultaneous Determination of Chlorthalidone and Amlodipine Besylate in Bulk and in Tablet Dosage Form

Development and Validation of Stability Indicating RP-HPLC Method for Simultaneous Determination of Chlorthalidone and Amlodipine Besylate in Bulk and in Tablet Dosage Form

Development and validation of a RP-HPLC method for simultaneous determination of Ciprofloxacin and Ornidazole in a solid dosage formulation

Development and validation of a RP-HPLC method for simultaneous determination of Ciprofloxacin and Ornidazole in a solid dosage formulation

Development of RP-HPLC Method for Simultaneous Determination of Bilastine and Montelukast by Qbd Approach and Its Validation

This study proposes to develop and validate the RP-HPLC method for Bilastine (BIL) and Montelukast (MKT) byQbD to substantiate the RP-HPLC analysis as per ICH validation guidelines. Quality by Design (QbD) allows the accomplishmentof specific unsurprising quality with a predetermined and wanted determination. The simultaneous estimation of BIL and MKTwas performed with C18 (4.6×250 mm, 5-μm particle size) with an LC-10AD pump and PDA detector. The mobile phaseemployed methanol and ammonium acetate buffer pH-3.6 at 85:15 v/v. The flow rate was maintained at 1.0 ml/min, and BIL andMKT were detected at 249nm and 293 nm by UV detector, respectively. The HPLC method provided linear responses found inthe 200–600 μg/ml range. The correlation coefficient was 0.9995 for BIL and 0.9991 for MKT. The LOD and LOQ for BIL andMKT were found to be 0.493, 1.495 μg/ml, and 0.693, 2.100, respectively. The percentage recovery for BIL was 95.33 to 102.06,and for MKT was 96.31 to 104.05, respectively. Calculated information acquired for both the preliminaries roughly coordinateswith the information given by Design expert programming, showing the chromatographic condition's genuineness. Design-Expertversion 10 ("DX10") software has calculated this calculation, setting a composite design of significant parameters. A newselective, rapid, accurate, precise, and sensitive RP-HPLC method was developed and evaluated for the simultaneousdetermination of Bilastine (BIL) and Montelukast sodium (MKT) in a bulk and pharmaceutical dosage form. This method is usefulin the routine quality analysis of combinations of BIL and MKT in bulk and its tablet formulations.

QbD based RP-HPLC method for simultaneous determination of a emtricitabine, tenofovir diproxil fumarate and efavirenz in tablet dosage form-an application to stability indicating assay

The current studies entail Quality by Design (QbD)-enabled development of a simple, rapid, sensitive and cost-effective high-performance liquid chromatographic method for estimation of emtricitabine (EMT), tenofovir diproxil fumarate (TEN), and efavirenz (EFA) in combined dosage form. Systematic optimization was performed employing Box-Behnken design by selecting the flow rate, buffer molarity, and acetonitrile volume as the critical method parameters (CMPs) identified from screening studies, thus evaluating the critical analytical attributes (CAAs), namely, retention duration, theoretical plates, resolution, and asymmetry factor as the parameters of method robustness. The optimal chromatographic separation was achieved using acetonitrile and phosphate buffer (pH 3.0) 74.1:25.9 v/v as the mobile phase with a flow rate 0.91 mL/min, and UV detection at 260 nm. The method was validated as per the ICH recommended conditions, which revealed high degree of linearity, accuracy, precision, sensitivity and robustness over the existing liquid chromatographic methods of the drug. Studies on forced degradation under acid, basic, oxidative, photolytic, and thermal conditions revealed the drug’s well-resolved peak and the degradation products’ peaks as well.

A validated new RP-HPLC method for simultaneous determination of amoxicillin, ampicillin and cloxacillin in pharmaceutical formulations

AbstractAs per the World Health Organization, 10% of medicines in low- and middle-income nations are of poor quality and pose a huge public health risk. The development and implementation of cost-effective, efficient and quick analytical methods to control the quality of these medicines is one of the immediate strategies to avoid such a situation. Hence, the main goal of this study was to develop and validate a simple, specific and precise new RP–HPLC method for simultaneous analysis of amoxicillin, ampicillin and cloxacillin in pharmaceutical formulations. The chromatographic analysis was achieved using Shodex C18 (250 × 4.6 mm, 5 μm) column with UV detection at 225 nm. The mobile phase was a gradient mixture of 30 mM phosphate buffer, pH 4.0 (mobile phase A) and acetonitrile (mobile phase B). Efficient separation of the three drugs was obtained using the final optimized chromatographic conditions. The developed method was validated for its specificity, linearity, precision, accuracy and robustness as per the ICH guidelines. The validation results showed that the method was specific, linear, precise, accurate and robust for the simultaneous determination of the three drugs. The developed method was applied to determine the content of the three drugs in pharmaceutical formulations. The assay results of the preparations showed that their drug content was within the pharmacopeial limit stipulated for each drug product. It can be concluded that the proposed method is suitable for simultaneous determination of amoxicillin, ampicillin and cloxacillin in pharmaceutical formulations in industries and regulatory laboratories.

Development and Validation of Stability-Indicating RPHplc Method for Simultaneous Determination of Canagliflozin and Metformin in Fixed-Dose Combination

Development and Validation of Stability-Indicating RPHplc Method for Simultaneous Determination of Canagliflozin and Metformin in Fixed-Dose Combination

STABILITY INDICATING ISOCRATIC RP-HPLC AND SECOND DERIVATIVE UV SPECTROSCOPIC METHODS FOR SIMULTANEOUS DETERMINATION OF REMOGLIFLOZIN ETABONATE AND VILDAGLIPTIN HYDROCHLORIDE

Remogliflozin and Vildagliptin are the latest drug combination in order to treat type 2 diabetes. The present work describes a comparative study of 2nd-ordered derivative UV and isocratic stability indicating RP-HPLC methods for simultaneous determination of Remogliflozin and Vildagliptin in the drug and its formulation. The HPLC method was established with a C18 column and the mobile phase with Buffer (pH-6.50): Acetonitrile: Methanol ratio of 55:44:1, at 1.2 ml/min flow and wavelength of 210 nm. The UV spectrophotometric method works on spectrophotometry with 2nd order derivative spectra(2D) using a zero-line cross method. The absorbance determination was performed at 246.0 nm for REM and at 219 .6 nm for VIL. The methods were validated for Linearity, Precision, Recovery, Robustness, and Forced degradation as per ICH-Q2(R1). The linear range was established in the range of 4-80 ppm for REM and 2-40 ppm for VIL for the UV method and 2-200 ppm for REM and 1-100 ppm for VIL for the HPLC method with regression coefficient ≥0.990. The accuracy was 99.37% & 99.62% for UV and 100.98 & 100.65 for the HPLC method for REM and VIL respectively. The proposed methods have been effectively used for the quantification of REM and VIL in API mixture and formulation dosage forms. Both methods are economical, fast, simple, and accurate which can be easily adopted for laboratory use.

Development and validation of a RP-HPLC method for simultaneous determination of Benzydamine hydrochloride and Cetylpyridinium chloride in oral spray solution

Development and validation of a RP-HPLC method for simultaneous determination of Benzydamine hydrochloride and Cetylpyridinium chloride in oral spray solution

Development and validation of an RP-HPLC method for simultaneous determination of curcumin and metronidazole in combined dosage form

Development and validation of an RP-HPLC method for simultaneous determination of curcumin and metronidazole in combined dosage form

Green RP-HPLC method for simultaneous determination of sofosbuvir, ledipasvir, velpatasvir antivirals and beyond in their bulk material and co-formulated products

In the present era, pharmaceutical industry is floated with the development of novel antiviral medications either in single or co-formulated pills. This growth of antiviral drug market is gaining the interest due to economic and therapeutic policies. Therefore, it is a real demand to validate analytical methods that monitor the entire standards considering their quality, safety and efficacy with maximum accuracy and maximum time and cost efficiency.HPLC offers a quick, automated, and highly accurate method to identify certain chemical components in a sample. Medical, forensic, bioavailability, environmental and manufacturing labs use the technique to quantify and separate the chemicals in a multicomponent sample. Consequently, an HPLC method is developed for quantitative analysis of a mixture containing five anti-viral agents namely, sofosbuvir (SF), ledipasvir (LD), velpatasvir (VL), daclatasvir (DC), and valacyclovir (VC) The method based on partitioning between reversed phase and methanol/water mobile phase with gradient elution at 1 mL/min flow rate and UV detection at 260 and 330 nm wavelengths along linear concentration ranges 5.00–80.00, 2.00–50.00, 5.00–50.00, 2.00–80.00 and 5.00–45.00 µg/ mL for SF, LD, VL, DC and VC, respectively.The greenness profile was assessed using Analytical Eco-scale and GAPI tools and found to be in agreement with the green analytical chemistry principles. The proposed method was validated experimentally and statistically and found to be a viable choice for single and multicomponent analysis in multi-tasks laboratory that consider the environmental impact and quality-cost balance with precision RSD% <2.00 % and accuracy ranging from 99.79 to 100.90 %.

Development of RP-HPLC Method for Simultaneous Determination of Nebivolol Hydrochloride, Valsartan and Hydrochlorothiazide in co-administrated Antihypertensive Agents and Human Plasma

Development of RP-HPLC Method for Simultaneous Determination of Nebivolol Hydrochloride, Valsartan and Hydrochlorothiazide in co-administrated Antihypertensive Agents and Human Plasma