Potassium Dihydrogen Phosphate Buffer Research Articles

Simultaneous HPLC determination of remdesivir and dexamethasone in the presence of metformin, sitagliptin, and glimepiride in a synthetic mixture and spiked human plasma

The COVID-19 pandemic has raised many questions regarding the control and therapy of type 2 diabetes and the higher risk of severe disease progression. One of the therapeutic regimens used in moderate and severe cases of COVID-19, endorsed by the World Health Organization, involves the administration of an antiviral medicinal product and a corticosteroid. The present study describes the development of a liquid chromatographic method for the simultaneous separation and quantification of Remdesivir and Dexamethasone in the presence of Metformin, Sitagliptin, and Glimepiride in a synthetic mixture. The developed method also allows determination of Remdesivir, Dexamethasone, and Glimepiride in spiked plasma samples, using Sitagliptin as internal standard. A mixture of acetonitrile and potassium dihydrogen phosphate buffer (pH 3) in a ratio of 45:55 v/v was used as а mobile phase on a C18 column. The recovery percentages from plasma ranged from 85.1 to 108.5%. The developed method can serve in routine quality control and clinical laboratory practice.

Challenges and Strategies to develop RP-HPLC Method of L-Arginine with Polyphenolic compounds

While developing a method of RP-HPLC for a new compound or combination of drugs simultaneously, many challenges are faced by the researcher pertaining to effective elution and sufficient resolution of a mixture of compounds. Many factors come into the picture, which reflects the ultimate elution of chemical compounds. Factors like the physicochemical properties of chemical compounds, the chemical nature of the mobile phase, instrumental factors, and experimental conditions play crucial roles in RP-HPLC method development. This research article discusses the challenges faced while developing a method of L-Arg along with polyphenolic compounds. Both the compounds have contrasting solubility profiles as L-Arg is soluble in water but PIC is soluble in organic solvents. Strategies were used to develop RP-HPLC of this combination with mobile phases like acetonitrile, orthophosphoric acid, methanol, and potassium dihydrogen phosphate buffer of pH 2.6. The present article provides an insightful approach to develop a new RP-HPLC method for a combination of compounds having related physicochemical characteristics.

Isocratic RP-HPLC Method Development, Validation, and Optimization of BCS-II in Bulk and Dosage Form

Background:: Previous studies of dextromethorphan hydrobromide basically worked on simultaneous research with other compounds. So, the development of a novel method using the isocratic elution mode is needed. Objective:: For the detection of dextromethorphan hydrobromide (DXM) in diverse matrices, a straightforward, accurate, and sensitive reversed-phase HPLC technique using a Waters 2487 Dual λ Absorbance detector has been designed and validated. Methods: In this experimental work, utilizing methanol/pH 3.0 potassium dihydrogen phosphate buffer (70:30, v/v) as the mobile phase, the separation was completed in 7 minutes on a C-18 HPLC column (4.6 cm length, 4.6 mm internal diameter; 5 μm particle size) utilizing an isocratic elution mode, flow rate of 1.0 mL/min, and UV-detection at 278 nm. Integration of the chromatography response was carried out using Empower 2.4 software Results:: With an R2 of 0.9987, the current approach showed high linearity for DXM in the 10- 60 ppm range (retention time 4.281 ± 0.505 min). For DXM Hbr, the limits of detection (LOD & LOQ) were 10.633 μg/mL and 32.221μg/mL, respectively. Samples remained stable in the presence of the matrices without any apparent influence. Conclusion:: The novel approach, which used a straightforward liquid/liquid extraction procedure with recovery ranging from 100 ± 10 % performed by two different analytes, was accurate. The precision within and between days was ≤ 2.0% (RSD). The technique was proven to be reliable and repeatable, and it can be utilized with pharmacological (active ingredients, syrups) and also for biological (blood) matrices which can be used in future research work for bioanalytical method development such as pharmacokinetics studies.

Simultaneous analysis of the of levamisole with triclabendazole in pharmaceuticals through developing TLC and HPLC–PDA chromatographic techniques and their greenness assessment using GAPI and AGREE methods

Two simple and rapid chromatographic methods were developed and validated for the analysis of levamisole and triclabendazole simultaneously in pure and pharmaceutical products. The first method is thin-layer chromatography (TLC) with densitometry, and the second method is high-performance liquid chromatography with PDA detection (HPLC–PDA). A Hypersil BDS C18 column with dimensions of 4.6 × 150 mm and a particle size of 5 µm was used in the HPLC–PDA method. An isocratic condition was used to carry out the separation, and the mobile phase was made up of acetonitrile and a 0.03 M potassium dihydrogen phosphate buffer in double-distilled water. The ratio of the mobile phase preparation was 70:30 (v/v), and the flow rate was 1 mL/min. A wavelength of 215 nm was employed for analyte detection. Precoated silica gel 60 F254 aluminium plates were used for the TLC method's separation. Mobile phase was made of ethyl acetate, hexane, methanol, and ammonia (69:15:15:1) for the separation. The detection wavelength selected was 215 nm. According to the International Council for Harmonization (ICH) guidelines, the proposed methods were validated and it was found that the two chromatographic methods are accurate, precise, and linear for both compounds in the range of 3.75–37.5 and 6–60 mg/L for the HPLC method for levamisole and triclabendazole, respectively and in the range of 2–14 µg/spot for the TLC method. The developed methods greenness profile was assessed using AGREE and ComplexGAPI tools.Graphical

DEVELOPMENT OF QUANTITATIVE METHOD OF TULOBUTEROL HYDROCHLORIDE IN RAT PLASMA: VALIDATION AND APPLICATION TO PRECLINICAL PHARMACOKINETICS

Objective: A robust, simple, accurate, rapid, and selective bioanalytical high-performance liquid chromatography (HPLC) method was established and validated to determine the tulobuterol hydrochloride in rat plasma. Methods: The protein precipitation method deproteinated analyte from rat plasma using acetone. The analysis of tulobuterol hydrochloride from rat plasma was accomplished using a mobile phase comprising of methanol: potassium dihydrogen orthophosphate buffer (0.05M; pH 4.0) in 90:10 (v/v) ratio run at 1.0 ml/min flow rate. Separation was carried on BDS hypersil C18 column (4.6 mm × 250 mm; 5 µ) at ambient temperature employing a 996 photodiode array (PDA) detector at 228 nm. Results: The linearity model was exhibited from 100-500 ng/ml with a good correlation of 0.999. Tulobuterol hydrochloride was efficiently separated at a retention time of 7.281 min. The percent recovery rate was between 100.21-100.46 %. The accuracy, precision, robustness, and ruggedness study showed relative standard deviation (%RSD) was within 2% (acceptable limit), and that revealed the method was efficient, precise, reliable, and reproducible. Conclusion: A simple, accurate, suitable method to quantitate tulobuterol hydrochloride in rat plasma was established using HPLC employed with a PDA detector that overcomes the increased cost for analysis. The developed method was successfully validated in rat plasma.

A synergistic chemometric combination for whiteness and greenness assessed HPLC-DAD assay of aqueous extracts of ivy and thyme and potassium sorbate in a syrup formula

A stability-indicating fit-for-purpose method has been developed for co-determination of hederacoside C (HC), thymol (TL), and potassium sorbate (PS) combined in a syrup dosage form. Carvacrol (CL) was separated from thyme extract (THY) with demonstrated identity at high confidence. This method was based on a reversed-phase (RP) high performance liquid chromatographic (HPLC) separation of the cited analytes. The HPLC separation was performed on a RP Zorbax Eclipse XDB-C18 analytical column [250 × 4.6 mm, 5 µm (80 Å)] and SecurityGuard C18 guard column (4 × 3 mm, 5 µm) with gradient elution system of HPLC far UV grade acetonitrile and 20 mM aqueous potassium dihydrogen phosphate buffer adjusted to pH 2.1 as the mobile phase at a flow rate of 2.5 mL min−1 and column oven was set at 50 °C. Quantitation was achieved with the photodiode array detection (DAD) with interferent peak suppression. Target Measurement Uncertainty (TMU) was determined by misclassification rates approach employing 100,000 Monte Carlo simulations (MCS). The bias and imprecision standard deviation were combined as a total analytical error (TAE) and compared via an enhanced approach linked to product specifications, process variation, and shelf-life variability for proving a fit-for-purpose analytical procedure. The robustness of the method was checked using supersaturated design constructed from Plackett-Burman design (PB-SSD) which was statistically analysed via the very recent approach of autovalidation using self-validated ensemble modelling (SVEM). The calibration graphs for each analyte were rectilinear in the range of 42.12–560.00, 3.37–47.15 and 0.51–7.07 µg mL−1 for PS, HC, and TL, respectively. The proposed HPLC-DAD method was successfully applied for the determination of the investigated analytes in a syrup formula. Moreover, the whiteness and greenness of the procedure were assessed via the most recently cited approaches.

Validated Analytical Method for Multicomponent Analysis of Famotidine and Ofloxacin in Bulk drug and Tablet Formulation by using UV-Visible Spectrophotometer and RP-HPLC

A simple, sensitive, accurate, precise, and reproducible UV- spectrophotometric method and RP-HPLC methods were developed and validated for the estimation of Famotidine and Ofloxacin in bulk drug and pharmaceutical formulation. The Linearity regression was detected and shows a good linear relationship; in the concentration range of 10-50µg/mL (R2 >0.9908) for famotidine and 10-50µg/mL (r2>0.9913) for ofloxacin. The UV – spectrophotometric estimation was carried out by the first-order derivative spectrophotometric method and absorbance were recorded at 273 and 280nm. Beers range were found to be 5-50µg/mL, respectively for both drugs while, correlation coefficient r2 > 0.9988 and 0.9941 for famotidine and ofloxacin. The isoabsorptive point was found to be 274nm in HPLC optimized mobile phase composition, potassium dihydrogen orthophosphate: methanol (60:40). Chromatographic condition consisted of mobile phase potassium dihydrogen orthophosphate buffer pH 2.3, methanol (60:40v/v), run time 30 min, C-18 column (ODS Hypersil) and flow rate 0.8mL/minute. The retention time for famotidine and ofloxacin were found to be 2.44 min, 7.99 min. respectively, and detection at λmax 274nm for both drugs (overlain spectra). The UV methods and RP-HPLC showed good reproducibility and recovery with the percent relative standard deviation (RSD) less than 5%. As per ICH guidelines, the developed method was validated for linearity, accuracy, precision, Sandell's sensitivity, and repeatability proving its utility in the estimation of famotidine and ofloxacin in house tablet formulation.

The use of an RP-HPLC-UV method for the analysis of oxcarbazepine in the presence of its preservatives; stability studies and application to human plasma samples

AbstractA simple, sensitive, selective, accurate and precise method was developed and fully validated for determination of oxcarbazepine (OXC) in presence of their preservatives and determination of oxcarbazepine (OXC) in human plasma. A reversed phase liquid chromatography (RP-HPLC) with UV detection techniques were applied for separation and quantification of studied drug OXC. Successful separation of the drug from methyl paraben (M.P.), propyl paraben (P.P.) and potassium sorbate (P.ST.) was achieved on a Kromasil C18 column (5 μm particle size, pore size 300 Å, l × I.D. 250 × 4.6 mm). The mobile phase that contain aqueous 0.05M potassium dihydrogen phosphate buffer (pH 7): acetonitrile, (50: 50, %v/v). The method was linear over concentration ranges 5.0–50 μg mL−1 for OXC. Bioanalytical validation of the developed method was carried out according to US-FDA guidelines and revealed a good linear relations over a range of (5.0–50), (0.5–10), (0.05–0.15), and (1.0–10) μg mL−1 for OXC, M.P, P.P, and P.ST, respectively, with a correlation coefficient (R2) of more than 0.999. Limit of detection (LOD) were 1.15, 0.03, 0.01 and 0.04 μg mL−1 for OXC, M.P, P.P, and P.ST, respectively, Intra and inter-day precisions, calculated as percentage relative standard deviation (% RSD), were lower than 2.0%. The developed method can be applied for routine drug analysis, therapeutic drug monitoring and bioequivalence studies through the analysis of plasma samples taken from blood bank.

Determination of Valbenazine related substances by liquid chromatography method complying with the current regulatory guidelines. Method robustness evaluation by Design of Experiments software

AbstractA simple, specific, rapid, sensitive reversed‐phase high‐performance liquid chromatography method was developed and validated for the determination of Valbenazine, impurities, and its degradation products. The proposed high‐performance liquid chromatography method showed optimum separation in 60 min using potassium dihydrogen phosphate buffer and ACN in the ratio of 90:10 (V/V) as mobile phase‐A whereas ACN and methanol in the ratio of 60:40 (V/V) as mobile phase‐B. YMC‐Pack ODS‐AQ (150 × 4.6 mm, 3.0 μm), column used along with security guard cartridge C18, (4.0 × 3.0 mm) at 1.1 ml/min flow. The injection volume was 10 μl and detection was carried out at 285 nm. The drug was degraded under different stress conditions and the method was validated as per International Conference on Harmonization guidelines on the basis of accuracy, precision, linearity, and robustness. A degradation study indicates that Valbenazine is prone to oxidative degradation. The method was linear over the concentration range of limit of quantitation to 200% for impurities and drug substances. The method was found robust and was studied by the design of experiments. The proposed method is applicable for the determination of the stability of Valibenazine and was successfully used in the quality control of bulk drug manufacturing and pharmaceuticals.

Development and validation of a high-performance liquid chromatography method for the estimation of esomeprazole in bulk and tablet dosage form

Introduction: An accurate, simple, precise, rapid, economic and reproducible reverse-phase high-performance liquid chromatography method was developed and validated for the estimation of Esomeprazole (ESO) in bulk and tablet dosage form. Method: The separation was carried out on Finepak SIL C18T-5 column (250 × 4.6 mm, 5.0 µm i. d.) using potassium dihydrogen phosphate buffer (0.025M): ACN (20:80 v/v) and at a flow rate of 1.0 mL/min. using UV detector at 302 nm with a run time of 10 min. The method was validated for accuracy for linearity, accuracy, precision, limit of detection (LOD), limit of quantification (LOQ) and robustness. Results: The standard calibration curve was linear with R2 = 0.995. LOD and LOQ obtained for esomeprazole were 0.0001 and 0.0004 µg/mL respectively. The method was found robust for possible changes. Results of analysis of other parameters were also tested and validated as per ICH guidelines and recovery studies confirmed the accuracy of the proposed method. validation studies showed that the developed HPLC method is simple, reproducible, rapid, precise and reliable. The high recovery and low relative standard deviation confirm the suitability of the developed method for the determination of esomeprazole in the tablet dosage form. Conclusion: This method may be used as a more convenient and efficient option for the analysis of esomeprazole to establish the quality of the substance during routine analysis with consistent and reproducible results.

Eco-friendly micellar HPLC approach for assessment of two anti-asthmatic breezhalers; Application of central composite design

Short-term breathing problems are brought on by asthma, a chronic lung disease. Mometasone furoate (MOF) and Indacaterol (IDT), are co-formulated alone or with Glycopyrronium (GLY) in the form of inhalation powder to control the symptoms of asthma. A novel green hybrid micellar liquid chromatographic method was developed to analyze the three mentioned medications. Contrary to routine analysis, Quality by Design methodology was applied to maximize the resolution with the fewest number of trials. Fractional factorial design was used to study the impact of three significant variables: sodium dodecyl sulphate (SDS) concentration, methanol %, and the mobile phase pH. Separation of the analytes was optimized by central composite design using a mobile phase consists of (191.49 mM) SDS and (10.00 mM) potassium dihydrogen phosphate buffer: methanol (77.50: 22.50 v/v), pH 3.00 on a reversed phase C18 column, within eight minutes. The flow rate was 1.2 mL/min, with 20 µL injection volume, and UV detection at 210 nm. Green Analytical Procedure Index, and Analytical Greenness Calculator were used for the method’s greenness evaluation, and it was deemed to be an effective green methodology. The developed approach was effectively applied for quantitative determination of the three drugs in bulk powder and two pharmaceutical dosage forms. International Conference of Harmonization requirements were adhered to when validating the approach. Under optimized conditions, calibration curves of the studied drugs were linear over the concentration range of (10.00 – 100.00 µg/mL) with high accuracy results and correlation coefficients of 0.9997 for MOF, 0.9996 for IDT and 0.9996 for GLY. Additionally, a precise approach has been demonstrated by the intra- and inter-day coefficients of variation ≤ 2%. The LOD and LOQ were (2.39 μg/mL) and (7.25 μg/mL) for MOF, (2.67 μg/mL) and (8.08 μg/mL) for IDT, and (2.80 μg/mL) and (8.48 μg/mL) for GLY, respectively.

Residue analysis of 10 aminoglycoside antibiotics in aquatic products by multiwalled carbon nanotubes combined with mixed-mode ion exchange liquid chromatography-tandem mass spectrometry.

An ultra-high-performance liquid chromatography-tandem mass spectrometry method was developed for simultaneous determination of 10 kinds of aminoglycosides in edible parts of aquatic products. The samples were extracted with 10mmol/L potassium dihydrogen phosphate buffer solution, then the pH value of the extract was adjusted to neutral by sodium hydroxide. Half volume of the extract was loaded onto multiwalled carbon nanotubes cartridge. All the target compounds were separated on a mixed-mode ion exchange column and detected by ultra-high-performance liquid chromatography-tandem mass spectrometry with electrospray in the positive ionization mode. Under optimized conditions, this method had a good linearity with a squared correlation coefficient>0.999. For neomycin, the limit of detection and limit of quantification were 5.0μg/kg and 10.0μg/kg, respectively; for hygromycin B and apramycin, values were 2.0μg/kg and 5.0μg/kg, respectively; for the other seven kinds of aminoglycosides, values were 1.0μg/kg and 2.0μg/kg, respectively. The average recoveries presented 75.8%-107.2% with intra- and interday reproducibility ranging between 3.8% and 12.5%. The method was rapid with good separation and sharp peak shapes, had the characteristicsis of high accuracy and good precision, and was suitable for simultaneous determination of 10 kinds of aminoglycosides in aquatic products.

Experimental Design Approach for Development of HPLC Method for Simultaneous Analysis of Triamcinolone, Nystatin, and Gramicidin in Industrial Wastewater

This study used an experimental design approach to optimize an HPLC method for the simultaneous determination of three pharmaceutical residues (triamcinolone, nystatin, and gramicidin) in industrial wastewater samples. The goal of using an experimental design approach was to maximize the method performance through separation enhancement and shortening the time of analysis and/or minimizing the environmental effects through the reduction in wastes and sample treatment. To achieve this goal, two steps were performed: a full factorial screening design for the three chromatographic variables, and optimization design using central composite design to select the optimum conditions that accomplished the highest resolution between adjacent peaks within a minimum run time of less than 5 min. The optimal chromatographic conditions derived from Minitab software using the desirability function were applied. Separation was carried out on a Zorbax C18 column (250 mm × 4.6, 5 μm) with gradient elution of a mobile phase composed of methanol and 0.25 M potassium dihydrogen phosphate buffer (pH 3.6) at different UV detections. For the validation of the developed HPLC method, ICH guidelines were followed, and the obtained results were found to be in compliance with the acceptance criteria. Linearity was over the concentration range of 1.00–25.00 μg/mL for triamcinilone and nystatin and 10.00–50.00 µg/mL for gramicidin. The proposed method was successfully applied to quantify the three studied pharmaceutical compounds in rinsing wastewater samples.

Accurate and Precise RP-HPLC Quantification of Piperacillin and Tazobactum Combination in Dosage Forms: A Quality Control Perspective

A simple, precise and accurate reverse phase high performance liquid chromatography (RP-HPLC) method has been developed and validated for the simultaneous estimation of piperacillin and tazobactum in pharmaceutical formulations. Separation was achieved on a C18 column using a mixture of potassium dihydrogen orthophosphate buffer (pH 3.5) and acetonitrile in the ratio of 60:40 v/v as the mobile phase at a flow rate of 1 ml/min. Detection was carried out at 226 nm. The retention times of piperacillin and tazobactum were about 2.5 and 4.3 min respectively. The method was validated as per ICH guidelines with respect to linearity, accuracy, precision, limit of detection, limit of quantification, robustness and ruggedness. The linearity ranges were found to be 5-15 μg/ml and 10-30 μg/ml for piperacillin and tazobactum respectively with correlation coefficients greater than 0.996. Accuracy, expressed as percentage recovery, was within 98-102%. The method also showed good precision and selectivity for the analysis of the drugs. The limits of detection and quantification for both analytes were adequate. In conclusion, the developed method was found to be suitable for the routine quality control analysis of piperacillin and tazobactum in pharmaceutical dosage forms.

Analytical quality-by-design approach for development and validation of HPLC method for the simultaneous estimation of omarigliptin, metformin, and ezetimibe: application to human plasma and dosage forms

A simple, selective, and sensitive RP-HPLC method was proposed for the simultaneous determination of two co-administered antidiabetic drugs (omarigliptin and metformin) with an anti-hyperlipidemic drug (ezetimibe) in a medicinally-recommended ratio of 2.5:50:1, respectively. The proposed procedure was optimized by adopting a quality-by-design approach. The influence of different factors on chromatographic responses was optimized by applying the two-level full factorial design (25). The optimum chromatographic separation was achieved using Hypersil BDS C18 column at 45 °C, and the mobile phase pumped isocratically composed of methanol: potassium dihydrogen phosphate buffer (6.6 mM; pH 7, 67:33% v/v) at a flow rate of 0.814 mL/min using 235 nm as a detection wavelength. The developed method was capable of separating this novel mixture in less than 8 min. The calibration plots of omarigliptin, metformin, and ezetimibe showed acceptable linearity over the ranges of 0.2-2.0, 0.5–25.0, and 0.1-2.0 µg/mL with quantitation limits of 0.06, 0.50, and 0.06 µg/mL, respectively. The proposed method was successfully applied to determine the studied drugs in their commercial tablets with high % recoveries (96.8-102.92%) and low % RSD values (less than 2%). The applicability of the method was extended to the in-vitro assay of the drugs in spiked human plasma samples with high % recoveries (94.3-105.7%). The suggested method was validated in accordance with ICH guidelines.

Design of Optimized RP-HPLC Method for Quantitative Analysis of Bisoprolol Fumarate in Bulk and Pharmaceutical Dosage Form

In this study an effective method was developed to assay Bisoprolol fumarate in tablets dosage form. The chromatographic separation was achieved on Reprosil pure basic C18 analytical column. A mixture of acetonitrile + Potassium dihydrogen phosphate buffer (0.050 mol L-1) (30:70 V/V), pH 3.5 was used as the mobile phase, effluent flow rate monitored at 1.0 mL/min, and UV detection at 233 nm. In forced degradation studies, the effects of acid, base, oxidation, UV light and temperature which were investigated showed no interference in the peak of drug. The proposed method was validated in terms of specificity, linearity, robustness, precision and accuracy. The method was linear at concentrations ranging from 5µg/mL to 17.5µmg/mL, precise (intra- and inter-day relative standard deviations R.S.D. < 2 %), (r2 = 0.9995).

Comprehensive and Validated Chromatographic Techniques for the Estimation of Lercanidipine Hydrochloride and Atenolol in Bulk and Combined Dosage Form in the Presence of Lercanidipine Degradation Products with LC/MS Characterization.

Two rapid, smart and validated stability indicating HPLC and TLC techniques were developed to determine atenolol (ATE) and lercanidipine HCl (LER) simultaneously in their pharmaceutical formulation. HPLC chromatographic separation was implemented by using Microsorb C18 (250 × 4.6mm, 5μm) column, with mobile phase of acetonitrile and 20mM potassium dihydrogen phosphate buffer pH3.5 adjusted by orthophosphoric acid in the ratio of (65:35, v/v) at a flow rate of 1.2mL/min at 240nm also the injection volume adjusted to be 30μL. These selected conditions effectively separated ATE and LER at a retention time of 2 and 6.7min, respectively, by isocratic elution mode without any interference from the obtained degradation products of LER. The densitometric determination was performed by using precoated silica gel 60F254 aluminum plates and chloroform, methanol and triethylamine (11.3:1.3: 0.3, by volume) as a developing system. The detection wavelength for simultaneous estimation of both drugs was 240nm in the presence of the oxidative product of LER. The RF values for ATE and LER were 0.22 and 0.78, respectively. The calibration curves of both techniques were constructed with linearity ranges of (5-55)μg.mL-1 and (1-55)μg.mL-1 for both ATE and LER, respectively, for HPLC determination. While for TLC, the linearity ranges were (1-4)μg/band and (0.2-1.4)μg/band for ATE and LER, respectively. LER degradation products were characterized using UPLC/MS and the suggested mechanisms and degradation pathways were introduced.

Deduction of the operable design space of RP-HPLC technique for the simultaneous estimation of metformin, pioglitazone, and glimepiride

A reversed-phase RP-HPLC method was developed for the simultaneous determination of metformin hydrochloride (MET), pioglitazone (PIO), and glimepiride (GLM) in their combined dosage forms and spiked human plasma. Quality risk management principles for determining the critical method parameters (CMPs) and fractional factorial design were made to screen CMPs and subsequently, the Box–Behnken design was employed. The analytical Quality by Design (AQbD) paradigm was used to establish the method operable design region (MODR) for the developed method depended on understanding the quality target product profile (QTPP), analytical target profile (ATP), and risk assessment for different factors that affect the method performance to develop an accurate, precise, cost-effective, and environmentally benign method. The separation was carried out using a mobile phase composed of methanol: 0.05 M potassium dihydrogen phosphate buffer pH 3.7 with 0.05% TEA (78:22, v/v). The flow rate was 1.2 mL/min. DAD detector was set at 227 nm. Linagliptin (LIN) was used as an internal standard. The proposed method was validated according to The International Council for Harmonization of Technical Requirements for Pharmaceuticals for Human Use (ICH). The assay results obtained by using the developed method were statistically compared to those obtained by the reported HPLC method, and a satisfying agreement was observed.

HPLC-DAD technique for the quantification of a recently approved anti-diabetic triple combination along with two toxic official impurities: Toxicity confirmation aided by molecular docking application

BackgroundGliflozins and gliptins are two distinct groups of pharmacological drugs that reduce blood glucose levels in individuals with type II diabetes in various ways that may perform their functions harmoniously. Trijardy® tablet, which contains empagliflozin, linagliptin, and metformin, was recently approved. The scientific database does not yet have a method that is sensitive enough to quantify the aforementioned medications in the presence of metformin official toxic impurities melamine and cyanoguanidine. Molecular docking modeling was utilized in this work to further prove the toxicity of melamine.MethodsThe five analytes listed before were quantified using RP-HPLC-diode array detector and a Zorbax® C8 column (4.6 × 250 mm, 5 μm) with isocratic mobile phase composed of acetonitrile and 0.05 M potassium dihydrogen phosphate buffer, which had been treated by ?-phosphoric acid to restore a pH of 4.0 (90:10, v/v) at a flow rate of 1.2 mL/min and the eluted peaks were scanned at 250 nm.ConclusionThe utilization of the simplest isocratic elution mode give the current technique a significant time-and cost-saving benefit. The current method can quantify the triple therapy agents in the presence of each other as well as with two official toxic impurities of metformin in one short analytical run.

Development and Validation of RP-HPLC Method for the simultaneous determination of Nirmatrelvir and Ritonavir in bulk and pharmaceutical formulation

In this study, a rapid and sensitive stability indicating reversed phase HPLC method was developed for quantitation of Nirmatrelvir and Ritonavir simultaneously in bulk and tablet formulation. Nirmatrelvir and ritonavir were separated on a Thermo C18 column with mobile phase containing 0.01M potassium dihydrogen phosphate buffer and acetonitrile (45:55, v/v). The flow rate was 1 mL/min and detection wavelength was 272 nm. Method linearity was established over a range of 75–225 μg/mL for nirmatrelvir and 50–150 μg/mL for ritonavir. Limit of quantification was 0.694μg/mL for nirmatrelvir and 0.820μg/mL for ritonavir. The recovery (%) was 99.96 to 100.45 (Nirmatrelvir) and 100.25 to 101.35 (Ritonavir). The method precisions were 0.11% (Nirmatrelvir) and 0.33% (Ritonavir). Method was suitable to assay nirmatrelvir and ritonavir in tablet formulation (Paxlovid). Stress degradation studies have shown that this method can be implemented to assay nirmatrelvir and ritonavir in the presence of its degradants.