Development Of HPLC Method Research Articles

Systematic Development of a Gradient Elution HPLC Method for the Analysis of Voxelotor and Its Structurally Related Substances Applying Analytical Quality by Design Approach

Systematic Development of a Gradient Elution HPLC Method for the Analysis of Voxelotor and Its Structurally Related Substances Applying Analytical Quality by Design Approach

Sensitive HPLC Method to Support Pre-formulation Studies and to Determine Critical Quality Attributes of Therapeutic Contact Lenses Containing Dorzolamide Hydrochloride

Background: Dorzolamide hydrochloride (DRZ) is a carbonic anhydrase inhibitor used to treat glaucoma and ocular hypertension. Drug-eluting contact lenses, such as Acuvue Theravision ™ with Ketotifen, offer improved drug delivery and reduced side effects compared to eye drops. Drug-loaded nanoparticle-loaded contact lenses can sustain drug release and enhance comfort for extended wear. Objective: High buffer concentration and low pH increase the risk of damage to silica-bonded columns. Therapeutic contact lenses face challenges related to critical lens parameters, including the estimation of drug incorporation and release due to interference of lens matrix leaching. There is currently no analytical method available for estimating DRZ in contact lenses. Method: The HPLC method, which was developed and validated using ICH Q2 (R1) criteria, used a C18 column (250 mm × 4.6 mm, 5 μm) as a stationary phase and methanol:water (70:30 v/v) as the mobile phase. The detecting wavelength was 253 nm. Moreover, to support the efficiency of the developed method, the marketed formulation of DRZ eye drops, drug purity, and loading in contact lenses were analysed. The method was also employed to determine the Critical Quality Attributes (CQAs) of therapeutic contact lenses and drug release and drug leaching during the sterilization process. Result: The developed HPLC method shows Rt for DRZ at 2.881 minutes with good linearity (r2 > 0.998) between 2-32μg/mL, precision (RSD < 2%), accuracy (Recovery > 99.5%), sensitivity, and specificity for quantifying DRZ in marketed formulations and therapeutic contact lenses. The developed method is devoid of any buffer or modifier in the mobile phase, making it safer for the stationary phase. This method mitigates the interference of lens matrix leaching, which induces an overestimation of DRZ. All the result for therapeutic contact lenses was found to be closely aligned with theoretically expected results, confirming the reliability of the developed HPLC method for therapeutic contact lenses. Conclusion: This method is specific, accurate, and precise for quantifying DRZ in commercial formulations and newly developed therapeutic contact lenses. It effectively evaluates the critical quality attributes of these lenses, demonstrating their reliability for assessing their performance and ensuring quality in therapeutic applications.

Possibilities and limitations of computer assisted chiral HPLC method development for ozanimod on polysaccharide based chiral stationary phases.

In this study, a direct HPLC method was developed to determine the enantiomeric purity of the immunomodulatory drug, ozanimod. A systematic method development process was followed, incorporating risk assessment, identification of critical analytical procedure parameters, initial screening of stationary phases, and software-assisted optimization of method parameters. Eight different polysaccharide-based chiral columns were selected to assess chiral separation of enantiomers under polar organic elution mode. The most promising results were obtained using a methanol:2-propanol mixture on the amylose-based Chiralpak AD column. Following this, systematic modeling was conducted using DryLab software to optimize method conditions, including isocratic eluent composition, temperature, and flow rate. Baseline separation was achieved within fifteen minutes using the optimized parameters: Chiralpak AD column thermostated at 10°C, and a mobile phase of methanol:2-propanol: diethylamine, 70:30:0.1 (v/v/v %), delivered at a flow rate of 0.8 mL/min. The developed method was validated according to current guidelines and in silico robustness testing was conducted to determine tolerance limits for critical separation parameters and their impact on enantioresolution. Our findings demonstrate the utility of DryLab, typically employed for reversed-phase achiral separations, in optimizing chiral methods even in polar organic mode. Limitations of the selected approach the development of chiral separation methods are also highlighted.

Formulation development, characterization, and evaluation of sorafenib-loaded PLGA-chitosan nanoparticles.

The basic purpose of this work was to develop environmentally friendly, biodegradable, and biocompatible polymeric nanoparticles of sorafenib that can effectively release the desired drug in a customized and controlled manner for targeting hepatocellular carcinoma. The solvent evaporation technique was employed for the synthesis of sorafenib-loaded PLGA-chitosan nanoparticles, followed by various experimental specifications and compatibility studies using poloxamer 407 as the stabilizer. The best nanoparticles thus synthesized were selected to be used for cytotoxicity investigations through in vitro and in vivo assessments. For the in vitro drug release tests, the dialysis bag diffusion technique was used. For both chitosan nanoparticles and PLGA loaded with sorafenib, a biphasic release pattern was found, exhibiting a protracted release lasting 10 days after a 24-h burst release. As experimental animals, rabbits were utilized to evaluate different in vivo pharmacokinetic properties of the selected formulations. Plasma samples were extracted with acetonitrile and analyzed through the developed HPLC method. Pharmacokinetic parameters such as AUC0-t, Cmax MRT, Vd, and half-life (t1/2) were enhanced significantly (p ≤ 0.001), while clearance was considerably decreased (p ≤ 0.001) for the chosen synthesized nanoparticles in contrast to the commercially accessible sorafenib formulation (Nexavar®). The cytotoxicity of the reference drug and sorafenib-loaded PLGA and chitosan nanoparticles was calculated by performing an MTT assay against HepG2 cell lines. The developed polymeric sorafenib nanoformulations possess the appropriate physicochemical properties, better targeting, surface morphology, and prolonged release kinetics. The pharmacokinetic parameters were improved significantly when the results were compared with commercially available sorafenib formulations.

Development and validation of a stability-indicating HPLC method for assay of tonabersat in pharmaceutical formulations

A stability-indicating reversed-phase high-performance liquid chromatography (RP-HPLC) method was developed to assay tonabersat and assess its stability in pharmaceutical formulations. Chromatographic separation was achieved using a Kinetex® C18 column (2.6 µm, 150 x 3 mm, 100 Å) at 50 °C, with a 20 µL injection volume. A linear gradient of acetonitrile in water (5 – 33.5 %) was applied for 1 min, followed by a gradual increase to 100 % over 26 min at a flow rate of 0.5 mL/min. Tonabersat and its degradation products were detected at 275 nm and 210 nm, respectively. The optimized method was used to evaluate the stability of tonabersat in lipid-based pharmaceutical formulations at 5 ± 3 °C, 25 ± 2°C/60 ± 5 % RH, and 40 ± 2 °C/75 ± 5 % RH over 3 months. The method was validated as per ICH guidelines and demonstrated linearity in the range of 5 – 200 µg/mL (R2 = 0.99994) with good accuracy (98.25 – 101.58 % recovery) and precision (% RSD < 2.5 %). The limits of detection and quantitation were 0.8 µg/mL and 5 µg/mL, respectively. Forced degradation studies showed significant degradation on exposure to alkaline (90.33 ± 0.80 %), acidic (70.60 ± 1.57 %), and oxidative stress (33.95 ± 0.69 %) at 70 °C, but no degradation was observed on exposure to thermal or photolytic stress. No chemical degradation was observed in either formulation on storage. Thus, the method was sensitive, specific, and suitable for stability testing of tonabersat in pharmaceutical formulations.

Stability-Indicating HPLC Method Development and Validation for the Simultaneous Determination of Vildagliptin and Dapagliflozin in Pharmaceutical Dosage Form.

Vildagliptin (VD) and Dapagliflozin (DP) were determined simultaneously by RP-HPLC using an ultraviolet (UV) detector, a Hypersil Gold C18 (250 × 4.6 mm) column, 5 µm and a mobile phase of acetonitrile: water (adjusted with O-Phosphoric acid to pH 3) in ratio 40: 60% v/v. The estimation wavelength was chosen to be 213 nm. VD and DP were shown to have retention times of 2.9 and 8.3 minutes, respectively. Q2 (R1) ICH guidelines were followed during the validation process. VD and DP correlation coefficient R2 values are 0.9993 and 0.999, respectively. While DP’s linearity range is 1 to 6 µg/mL, VD’s is 10 to 60 µg/mL. The precision and accuracy studies show %relative standard deviation (%RSD) below 2%. Recovery% was assessed to meet the ICH Q2 (R1) guidelines criteria. The presented research on forced degradation showed stability indicating studies. The findings also demonstrated that the suggested method is suitable for determining VD and DP precisely and accurately.

Method Development for Analysis of Cyclodextrin Inclusion Complexes with HPLC

Cyclodextrin inclusion complexes have been widely used in the pharmaceutical industry to improve drug solubility and bioavailability. Analysis of these complexes using HPLC requires the development of appropriate and validated methods. To analyze and compare various HPLC methods used for the analysis of cyclodextrin inclusion complexes, and to identify challenges and trends in method development. A literature review was conducted of articles published in the last 10 years, focusing on HPLC methods for the analysis of cyclodextrin inclusion complexes. Parameters compared included column type, mobile phase composition, flow rate, and detection method. The majority of studies used C18 columns with varying lengths and particle sizes. The composition of the mobile phase varied, reflecting the diversity of properties of the complexes analyzed. The flow rate was generally 1.0 mL/min, with the exception of the HPLC/MS method. Detection methods included UV-Vis, PDA, fluorescence, and mass spectrometry. The main challenges included the dynamic equilibrium of the complexes and stability during analysis HPLC method development for the analysis of cyclodextrin inclusion complexes requires optimization of various parameters to overcome specific challenges. Future trends are towards the use of advanced technologies such as UHPLC and high-resolution mass detection.

HPLC-DAD method validation for quantification of dehydroabietic acid and abietic acid in oral spray containing Pinus merkusii heartwood extract and its antibacterial effects on clinically isolated Streptococcus mutans

AbstractPinus merkusii Jungh &amp; De Vries. has become increasingly gathered more attention from researchers because the plant has a range of folk medicinal uses. Heartwood plant is the major source of dehydroabietic acid (DHAA) and abietic acid (AA), which possesses several medicinal properties, such as antiviral, antimicrobial, antiobesity and anti-inflammatory. The research proposed herein a low-cost, fast, specific, uncomplicated, sensitive, precise reverse-phase high-performance liquid chromatography (RP-HPLC). This method was conducted and validated for evaluating an amount of DHAA and AA in ethanol extract and oral spray containing P. merkusii heartwood extract. Additionally, stability and antimicrobial activities against clinically isolated Streptococcus mutans of the oral spray were determined. The separation was achieved on Pursuit 200Å PFP column, 150 × 4.6 mm, particles of 3 µm with a flow rate of 1.0 mL min−1. Methanol and water (70:30 v/v) were used as eluent with an isocratic mode and sample analysis volume was set at 10 µL, at a detection wavelength of 210 and 245 nm. The developed HPLC method for analysis of DHAA and AA showed good linearity with correlation coefficients equal to 1. Moreover, other validation parameters, comprised of accuracy, precision, specificity and detection and quantitation limits of this method displayed excellent reliability, validity and sensitivity. This method could be an interesting alternative for quantitative measurement of P. merkusii heartwood extract, oral spray formulation and other P. merkusii heartwood extract preparations. The result from antibacterial tests suggested that the oral spray containing P. merkusii heartwood extract is able to inhibit the oral pathogens causing dental caries. The oral spray decreased S. mutans population size by about 0.5–2 Log CFU mL−1 at 1–4 h and complete elimination of all bacteria strains within 24 h. This study provides validity for using P. merkusii heartwood extract as an alternative for preventing and treating oral infectious diseases.

Development and Validation of an ICH-Compliant Optimized RP-HPLC Method for Quantitative Analysis of Favipiravir

Favipiravir (FAV) has emerged as a promising antiviral agent. It is particularly effective against influenza and other RNA virus infections. The aim and objective of the present study was developing and optimizing chromatographic conditions for faster analysis of FAV and demonstrating its applicability for tablet assay. The chromatographic separation was obtained using mobile phase with 90:10 % v/v ratio of ammonium acetate buffer (pH 4.5) and methanol on a Waters C18 column (250 x 4.6 mm, 5 µ). A flow rate of 1.4 mL/min was optimized. Chromatographic detection was perform at a wavelength of 323 nm. The FAV retention time was 6.46 min, indicating the efficiency and speed of the method. The developed method was rigorously validated according to ICH guidelines. A strong linear correlation (r2 = 0.9995) was established in the 5–60 μg/mL range, indicating adequacy for quantitative analysis. The method demonstrated high accuracy, with FAV recovery ranging from 98.77 % to 100.89 %. The % RSD results of less than 2% for intermediate precision and repeatability showed that the method exhibited high precision. The developed HPLC method reduces the Rt. Notably, this method complies with regulatory standards, establishing it as a valuable tool for quality assurance, pharmacological evaluation, and clinical monitoring of FAV. Thus, this validated RP-HPLC method provides a robust and sensitive method for routine FAV quantification, with high accuracy, precision, and compliance results, supporting its potential use in the clinical pharmaceutical industry.

Development and Application of HPLC and UFLC Methods for the Analysis of Cardiovascular Drugs

Separation and identification of amiloride, metoprolol, hydrochlorothiazide, Carvedilol and amlodipine, in human plasma using the presented SPE and HPLC procedures was selective, efficient, robust, economical, environmentally friendly, and repeatable. Because plasma samples showed no evidence of a secondary peak, it was determined that the claimed drug-drug interaction between cardiovascular medications did not occur. Furthermore, no new peak was generated, indicating no metabolic product. This is the first report on isolating and identifying these nine cardiovascular medicines. Thus, SPE and HPLC techniques may be used to investigate these 5 cardiovascular medications. Successful monitoring of these medications in human plasma was achieved using the established SPE and HPLC technologies. The results of this inquiry were contrasted with those that had already been released. In comparison to other works in the literature, it was determined that the current study had the best complete baseline separation and lowest detection limit. The retention, separation, and resolution factors were discovered to have ranges of 0.07-9.14, 1.44-4.21, and 2.15-18.66, respectively. The SPE and UFLC techniques created and verified for this purpose were used to examine human plasma samples for the presence of cardiovascular drugs. Peak retention, separation, resolution, and symmetry all matched the reference samples' values. The results for symmetry, separation, and resolution were all in agreement with the reference samples. The methods used to separate and identify amiloride, metoprolol, hydrochlorothiazide; Carvedilol and Amlodipine in human plasma were found to be selective, effective, hardy, affordable, environmentally friendly, and reproducible, according to the authors. The selectivity of the SPE technique was validated by the absence of any secondary signal in the plasma samples. Additionally, there was no evidence of these drugs degrading in the plasma sample. This is the initial report on the simultaneous isolation and identification of these eight medications. The well-known SPE and UFLC techniques were successfully used to monitor these drugs in human plasma. Therefore, these drugs can be detected in any plasma sample using SPE or UFLC methods

Development and Application of HPLC and UFLC Methods for the Analysis of Anti Histaminic Drugs

Research focused on medications used to treat allergic reactions. The results of this inquiry were contrasted with those that had already been released. In comparison to other works in the literature, it was determined that the current study had the best complete baseline separation and lowest detection limit. The retention, separation, and resolution factors were discovered to have ranges of 0.07-9.14, 1.44-4.21, and 2.15-18.66, respectively. The SPE and UFLC techniques created and verified for this purpose were used to examine human plasma samples for the presence of antihistaminic drugs. Peak retention, separation, resolution, and symmetry all matched the reference samples' values. The results for symmetry, separation, and resolution were all in agreement with the reference samples. The methods used to separate and identify Phenylephrine, cetirizine, loratadine, montelukast, and Ebastine in human plasma were found to be selective, effective, hardy, affordable, environmentally friendly, and reproducible, according to the authors. The selectivity of the SPE technique was validated by the absence of any secondary signal in the plasma samples. Additionally, there was no evidence of these drugs degrading in the plasma sample. This is the initial report on the simultaneous isolation and identification of these eight medications. The well-known SPE and UFLC techniques were successfully used to monitor these drugs in human plasma. Therefore, these drugs can be detected in any plasma sample using SPE or UFLC methods.

Simultaneous spectrophotometric and chromatographic determination of Ketorolac and Dexamethasone: Overall assessment regarding environmental Impact, Practicality, and functionality principles

Determination of a drug combination that contains a minor component that is also a poorly UV absorbing analyte represents greet challenges in drug analysis. A fixed-dose ophthalmic combination of ketorolac tromethamine (KET) and dexamethasone sodium phosphate (DEX) (in the ratio of 5:1) is used to relieve post-surgical inflammation and seasonal allergic conjunctivitis due to their analgesic and anti-inflammatory activity. Five spectrophotometric methods and a reversed phase chromatographic separation method were developed and validated for simultaneous determination of both drugs in lab-prepared eye drops. Direct spectrophotometric determination of KET (the major, highly UV absorbing analyte) was achieved from the zero-order UV absorption spectra at 323 nm while DEX (which is the minor, poorly UV absorbing analyte) was quantified after applying several spectrophotometric methods. Either fundamental zero-order spectra or ratio spectra were used, and measurements were done at either single or dual wavelengths. The developed methods were: isosbestic point method, absorption factor method, dual wavelength method, ratio difference UV spectrophotometric method, and simultaneous equation method. The developed spectrophotometric methods determine KET in the linearity range of 2.0–12.0 μg/mL and DEX in the range of 5.0 – 35.0 μg/mL. For accurate spectrophotometric determination of DEX in the ratio of the dosage form, multiple standard addition method was applied. HPLC technique was also applied for separation and quantification of both drugs. Chromatographic separation was attained by using Hypersil ODS-C18 (150 x 4.6 mm, 5 μm) column. The mobile phase was composed of 50 % methanol: 50 % phosphate buffer (pH 2.5). The column compartment was maintained at 25 °C. Flow rate was adjusted at 1.2 mL/min and detection was performed by DAD at 254 nm. The linearity of HPLC method was 5.0–100.0 μg/mL, and 5.0–50.0 μg/mL for KET and DEX, respectively. Simultaneous determination of both drugs was applied to lab-prepared eye drops that contain also benzalkonium chloride as inactive ingredient. The developed methods determined the cited drugs with good and acceptable % recovery. The results of the developed methods and a reported HPLC method were compared using one-way ANOVA test, F-test and student’s t-test at confidence level 95 %, and they show no significant statistical difference. Assessment of greenness of the developed methods in comparison with reported HPLC method was performed using different tools, including analytical eco-scale, GAPI and AGREE. HPLC-EAT was also performed to ensure greenness of developed HPLC method. Moreover, assessment of practicality, applicability, and functionality of developed and reported methods was evaluated by BAGI metric tool and WAC calculator, respectively. The developed methods allow simple, rapid, and accurate analysis of quality control samples that contain KET and DEX.

Green Analytical Approach for HPLC Method Development for Quantification of Sorafenib and Its Pharmacopeia Impurities: LC–MS/MS Characterization and Toxicity Prediction of Stress Degradation Products

ABSTRACTThis research presents the development and validation of chromatographic method for analyzing sorafenib and its pharmacopeial impurities, with a focus on stability studies and degradation product (DP) characterization. Initial method optimization involved exploring various column and buffer combinations, ultimately achieving optimal separation and peak symmetry using an ODS‐AQ YMC (150 mm) column with 0.6 mL/min gradient flow of 10 mM ammonium formate buffer adjusted to pH 3.4 with formic acid as solvent A, and ethanol as solvent B as mobile phase and 246 nm wavelength. Method exhibits calibration curve linear in 50–300 µg/mL for sorafenib and 0.050–0.30 µg/mL for impurities with a detection limit of 0.015 µg/mL for impurities. A structural elucidation of DPs was performed using LC–MS/MS, providing valuable insights into their molecular compositions, and was characterized as 4‐[4‐(carboxyamino)phenoxy]pyridine‐2‐carboxylic acid (DP 1) and 4‐(4‐aminophenoxy)pyridine‐2‐carboxamide (DP 2). Using AGREE and GAPI metrics, evaluation highlighted method sustainability through ethanol–water solvents and shorter column to reduce energy consumption. Toxicity assessments revealed differences in environmental impact and toxicological profiles of DPs, emphasizing importance of managing safety considerations for sorafenib and its DPs. This research offers novel insights into sorafenib analysis by addressing pharmacopeial impurities, characterizing DPs, and evaluating method sustainability and safety.

Stability‐Indicating HPLC Method Development and Validation for the Quantification of Tofacitinib Citrate and Its Related Substances Using Hydrophilic Liquid Interaction Chromatography

ABSTRACTA new stability‐indicating RPHPLC method with short run time has been developed and validated for tofacitinib citrate and its related substances. The novel HPLC method integrates hydrophilic interaction liquid chromatography (HILIC) technology as the stationary phase and employs a mobile phase composed of phosphate buffer (pH 7.0) and acetonitrile (45:55, %v/v) at a flow rate of 0.5 mL/min under isocratic elution. Analytes were monitored via a UV detector at 210 nm and with the column oven temperature at 30°C for a 20‐min analysis. Precision (%RSD) for Impurity‐A and Impurity‐B and tofacitinib met specifications at 2.4%, 0.8%, and 0.0%, respectively. Accuracy ranged from 86% to 100% for impurities, with LOD at 0.03% and LOQ at 0.05%–0.06%. Correlation coefficients exceeded 0.999 for impurities and tofacitinib citrate. Solution stability was confirmed for 24 h at room temperature. The method range was extended from LOQ to ∼1.5% for impurities and LOQ to ∼150% for tofacitinib citrate. The method's stability was evaluated under acid and base hydrolysis, oxidative and water hydrolysis, and thermal and photolytic degradation. Introducing HILIC as the stationary phase for this work proved effective, eliminating the need of ion pair reagents, reducing analysis time, and ensuring consistent results.

Advanced stability-indicating gradient elution reverse phase-high performance liquid chromatography method development and validation for simultaneous estimation of rutin and curcumin using an analytical quality by design approach

This research delves into a systematic Analytical Quality by Design (AQbD) approach to develop and validate a stability-indicating robust RP-HPLC method for the simultaneous determination of Rutin and Curcumin, compounds known for their anti-inflammatory, antioxidant, and wound-healing properties. Utilizing QbD principles, critical method parameters (CMPs) were identified and screened using design of experiments (DoE) and optimized via response surface methodology (RSM) with Central Composite Design (CCD). The optimized chromatographic separation used a Phenomenex Luna C18 column with gradient elution, using methanol and 0.1% formic acid, a flow rate of 1 ml/min, and detection at 345 nm (isosbestic point). The retention times were 5.63 mins for Rutin and 17.01 mins for Curcumin. The stability indication of the developed HPLC method was performed, and validation parameters were meticulously conducted following the ICH Q2(R1) guidelines, demonstrating linearity over 2-10 μg/ml for Rutin and 5-25 μg/ml for Curcumin, with correlation coefficients of 0.9999 and 0.9998, respectively. Precision studies showed %RSD within acceptable limits, and accuracies ranged between 97-100%. This validated HPLC method offers a reliable, sensitive approach for simultaneous quantifying Rutin and Curcumin, suitable for routine analysis in pharmaceutical laboratories and quality control.

Development and Validation of New HPLC Method for the Determination of Imidazolidinyl Urea in Topical Formulation

Introduction: In this study, a new, simple, selective, and fast liquid chromatographic method has been developed and validated for the determination of imidazolidinyl urea used as an anti-microbial agent in topical formulation. Methods: The developed HPLC method using a diode array detector for the determination of imidazolidinyl urea was applied to the topical formulation. Imidazolidinyl urea in the sample was analyzed in the cyano column (250 x 4,6 mm, 5 μm i.d.) under chromatographic conditions, where the flow rate was determined as 1.0 mL/min. The column oven was 40.0°C, and imidazolidinyl urea was detected at 210 nm. Isocratic application of acetonitrile-water (25:75, v/v) was used as the mobile phase system. The validation of the developed method was performed according to the International Conference on Harmonisation guidelines Q2 (R1). Results: The linearity range of the imidazolidinyl urea was 0.050-0.150 mg/mL, and the limits of detection and quantification were calculated to be 62.5x10-6 mg/mL and 125x10-6 mg/mL, respectively. Assay recovery and precision of imidazolidinyl urea from topical formulation at 0.050, 0.100, and 0.125 mg/mL concentrations were evaluated. The mean recoveries for imidazolidinyl urea in the topical formulation were calculated as 98.857-104.560%. Conclusion: The validated method was successfully applied to the determination of imidazolidinyl urea in a topical formulation. The proposed method is reproducible and reliable and can be used safely for routine analysis.

AQbD-guided development and validation of an innovative extraction procedure and stability-indicating RP-HPLC method for quantification of posaconazole in tablet formulation

The objective of this work is to develop a stability-indicating HPLC method for the quantification of Posaconazole (PCZ) in tablet formulation using an Analytical Quality by Design (AQbD) approach. The development process involved the Design of Experiments (DOE) utilizing distinctive constraints mixture design for mobile phase ratio optimization and a 2-level factorial design for selection of extraction diluent compositions. Key responses measured included % assay and system suitability parameters. Method operable design regions (MODR) were determined, and final optimum conditions were selected. Forced degradation studies were conducted to assess method stability. The optimized HPLC method employed a Zorbax C18 column with a mobile phase consisting of pH 3.5 10mM phosphate buffer, acetonitrile, and methanol in a ratio of 30:53:17 % v/v/v. The method demonstrated stability-indicating capabilities, with PCZ degradation observed in acidic and oxidative environments, while remaining stable in alkali. Peak purity analysis from Empower software confirmed the absence of interaction with degradants. Validation according to ICH Q2 (R2) guidelines showed precision, linearity over the range of 0.25 µg/mL to 376 µg/mL, and accuracy demonstrated through recovery studies from 50 to 150%. The developed HPLC method utilizing AQbD approach is specific, robust, precise, and accurate for the quantification of PCZ in tablet formulations, thus suitable for routine analysis.

HPLC Method Development and Validation for Simultaneous Estimation of Tramadol and Piroxicam

HPLC Method Development and Validation for Simultaneous Estimation of Tramadol and Piroxicam

QBD approach in HPLC method development and validation of furazolidone

QbD principles for establishment of a HPLC method for Furazolidone with improved robustness and performance. On the basis of the initial prioritization and factor screening studies, the highly influential factors were identified and subsequently optimized for improving the method robustness. The response surface mapping particularly facilitated improved understanding of the factor–response relationship and interactions associated with them. Extensive validation studies further ensured high degree of method robustness with extreme variation in the key variables influencing the method performance. In addition, the method showed improved sensitivity for the Furazolidone much beyond the values reported in the literature.

Development and Validation of HPLC Method for the Quantification of Impending Genotoxic Impurities in Dapson Drug Substances

Development and Validation of HPLC Method for the Quantification of Impending Genotoxic Impurities in Dapson Drug Substances