Analytical Quality By Design Research Articles

Development and Validation of a Stability Indicating Analytical Method for the Estimation of Gliclazide and Sitagliptin in Bulk Drugs and Tablet Dosage Forms by RP-UPLC: An Application of the Quality-by-Design Approach

Sitagliptin and gliclazide are the active ingredients in OpdualagTM. A highly selective DPP-4 inhibitor, sitagliptin is thought to work in type 2 diabetics by delaying the inactivation of incretin hormones, which increases their concentration and lengthens their duration of effect. Sulfonyl urea mostly enhances insulin secretion from pancreatic β-cells by inhibiting ATP-sensitive K+ channels, leading to depolarisation and Ca2+ influx. Reduced serum glucagon levels and enhanced insulin binding to receptors and target tissues are the mechanisms by which it exerts its effects. Objective: This study aims to create and evaluate an RP-UPLC technique for the detection of sitagliptin and gliclazide in pharmaceutical products by applying AQbD principles. Method: Findings By utilising the Design-expert® programme, we were able to execute a central composite design (CCD) consisting of three components arranged in five distinct layers, which greatly improved the chromatographic conditions. One method that oversees the whole analytical procedure life cycle—from method design and optimisation to validation and continual improvement—is Analytical Quality by Design (AQbD). Results: The retention times for sitagliptin were 1.269 minutes and for gliclazide they were 1.572 minutes. The limits of detection (LOD) and quantitation (LOQ) for Sitagliptin and Gliclazide were 0.19 µg/mL, 0.56 µg/mL, 0.28 µg/mL, and 0.86 µg/mL respectively. The recovery percentages for sitagliptin and gliclazide were found to be 99.15% to 100.54% and 99.93% to 100.58%, respectively. Both drugs were found to be susceptible to oxidative and photolytic breakdown in the forced degradation studies. Conclusions A novel RP-UPLC method has been developed for the quantitative detection of sitagliptin and gliclazide in pharmaceutical formulations, utilising the AQbD-based methodology. This approach is straightforward, rapid, precise, particular, and stable-indicating.

An Estimation of Baricitinib by AQbD-driven UV Spectrophotometry Development and Validation Process

Background: Baricitinib (BCTB) is a novel Janus Kinase (JAK) 1 and 2 inhibitor used in the therapy of rheumatoid arthritis, approved by the “Food and Drug Administration” in 2018. It has significant dose-dependent effectiveness and severe side effects. Thus, it is crucial to figure out its concentration in developed dosage forms. The literature search revealed that there has only been one UV spectroscopy technique documented up to this point. Methanol was chosen as the detection medium in this approach, which is not comparable with plasma or serum. As a result, the preliminary research suggested developing a UV spectroscopic approach that can estimate BCTB concentration and compare it to its concentration in the plasma or serum. Thus, in the proposed method, 7.4 pH phosphate buffer was selected as a mobile phase. Aim: Using the Analytical Quality by Design (AQbD) methodology, a simple, robust spectrophotometric method for the detection of BCTB in API form and Niosomes drug delivery system is designed and assessed. Methods: In the AQbD approach, a face-centered CCD design of Design Expert 13 software was used to evaluate two critical method variables: scanning speed and sampling interval. The design space suitability was confirmed by standard error and overlay plots. The 2-D contour and 3-D response surface plots were used to forecast the relationship between the response variable and predictor variables. Results: Baricitinib displays an absorption maximum at 249.40 nm in saline phosphate buffer pH 7.4. The distinguished linearity of the method was obtained over a concentration of 5–30 µg/ml with a correlation coefficient (R2 ) value of 0.998. BCTB % assay was found to be near 99 %. Intraday and Interday precision were found to have % RSDs of 0.067–0.488 and 0.146–0.942, respectively. Conclusion: The established spectrophotometric technique was observed to be precise as per ICH revised guidelines ICH Q2 (R1) and Q14 for analytical method validation. Our findings are instructional for the future design and development of safe and reliable therapeutic dosage forms of BCTB for rheumatoid arthritis.

Green RP-HPLC method for the estimation of carfilzomib in bulk, protein nanocarriers and human plasma: Application of chemometrics and Monte-Carlo simulations

Carfilzomib is a tetrapeptide epoxyketone that has shown potential clinical outcomes in the treatment of multiple myeloma. However, inaccuracies in quantifying such peptide drug products have arisen due to poor stability, low solubility, time-consuming techniques, complex physicochemical properties, and use of non-green solvents with less recyclability. This provides a substantial urge to develop an ecological and sensitive analytical method for quantifying peptide drugs from matrix formulation and biological samples in early as well as lateral stages of product development in pharma industries. As a result, the study aimed to develop a robust ecological method for estimation of carfilzomib via Green RP-HPLC using analytical quality by design (AQbD) paradigms with specific application in protein nanoparticles and biological matrix. Initially, an appropriate wavelength for quantification of carfilzomib was chosen using principal component analysis (PCA) as a chemometric tool.Risk assessment followed by factor screening studies using 8-factor Placket-Burman Design aided in earmarking critical method parameters (CMPs) affecting critical analytical attributes (CAAs). Further, Central Composite Design (CCD) was employed for design space optimisation to demarcate optimum chromatographic conditions, which were corroborated for robustness using Monte-Carlo simulations. The method was validated as per ICH Q2 (R1), followed by quantifying the greenness of the method using Green Assessment tools. The method optimisation resulted in the optimal chromatographic conditions using Green RP-HPLC. The chromatographic system was equipped with a Phenomenex Aeris Peptide-XC C18 column (150 × 4.6 mm × 5 µm), and the mobile phase was composed of isopropanol:methanol:0.1 M PBS (pH 5.5 adjusted using 0.1 % formic acid) (35:45:20v/v), with a 1 ml/min flow rate at a 210 nm ʎmax. The optimised chromatographic conditions resulted in a short retention time (RT) of 4.95 mins, 0.87 tailing factor (TF), 4,875,122 peak area (PA), and 8995 theoretical plate count (TPC). The method demonstrated linearity in a wide range of concentrations (0.1–20 µg/ml) with a correlational coefficient of 0.997 and < 2 % RSD. The method unearthed a high precision rate with more than 95 % of drug recovery in protein nanoparticles and human plasma, thereby confirming the accuracy and sensitivity of the developed method. Chemometrics and Monte-Carlo simulations ratified the robustness and sensitivity of the developed analytical method of Carfilzomib with established greenness and a high degree of practical utility in protein-based nano formulations and human plasma matrix for life cycle product development.

AQbD-guided stability indicating HPLC method for azelnidipine and chlorthalidone fixed-dose combination tablet: a green approach

This study developed a stability-indicating RP-HPLC method for quantifying azelnidipine and chlorthalidone in fixed-dose formulations using Analytical Quality by Design (AQbD) principles. A Plackett-Burman design was used for factor screening, followed by risk assessment and optimization using a central composite design to evaluate the effects of the organic phase percentage, flow rate, and modifier concentration. Acetonitrile percentage was identified as the most critical factor. The optimized method employed a Reliant™ C18 Waters column with a mobile phase of 0.1% formic acid and acetonitrile (62:38% v/v), with chlorthalidone and azelnidipine eluting at 4.1 and 13.7 minutes, respectively. Validation showed the method's robustness, accuracy, and precision. Forced degradation studies confirmed its reliability for tablet formulation analysis, and an Analytical Greenness score of 0.55 highlighted its environmental sustainability. This method is efficient, simple, and well-suited for routine quality control in the pharmaceutical industry.

International Approaches to the Development, Validation, and Change Management of Analytical Procedures (Review)

INTRODUCTION. In 2023, the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) published a new Guideline on Analytical Procedure Development (ICH Q14) and a revised version of the Guideline on Validation of Analytical Procedures (ICH Q2(R2)). Consequently, there is a need for a considerable revision of the approach to the development and validation of analytical procedures that is currently used in the Eurasian Economic Union (EAEU). A revision is also needed for the processes for evaluating and introducing variations to the analytical procedures described in medicinal product registration dossiers.AIM. This review aimed to analyse the significant changes made to international approaches to the development of analytical procedures, as well as to study the advantages and disadvantages of these approaches for pharmaceutical manufacturers and regulatory agencies in the EAEU.DISCUSSION. This review covers the key provisions and practical aspects of the enhanced approaches to the development of analytical procedures introduced by the ICH Q14 guideline. In particular, the review addresses the concepts of the analytical procedure life cycle (APLC) and the modified analytical qualityby-design (AQbD) approach; the development of the analytical target profile (ATP); analytical quality risk management; planning of the design of experiments (DoE) and the analytical procedure control strategy; and the validation, subsequent verification, transfer, and change management of analytical procedures. Additionally, the review describes the ICH Q2(R2) updates that accompany this new regulatory paradigm.CONCLUSIONS. The above guidelines fill the existing gap in recommendations for the development of analytical procedures. The use of the APLC and AQbD concepts provides both pharmaceutical companies and regulatory authorities with flexible approaches that are applicable to analytical procedures both during the development phase and once they have been implemented. Effective implementation of these international approaches in the Russian pharmaceutical industry and regulatory system requires a broad discussion between pharmaceutical industry professionals and regulatory agency experts, possibly, as part of a pilot project. After that, there will be a necessity to provide training for specialists involved in the development of analytical procedures and to amend the EAEU Rules for Marketing Authorisation and Expert Assessment of Medicinal Products for Human Use.

A Review on Advancement in Analytical Quality by Design (AQbD)

Analytical Quality by Design (AQbD) represents a transformative methodology in pharmaceutical development, anchored in a systematic, risk-based, and data-driven framework. This approach optimizes analytical methods, fostering heightened product quality, efficient regulatory compliance, and informed decision-making. The industry's increasing acceptance of AQbD principles signifies a paradigm shift towards enhanced efficiency, sustainability, and global harmonization. This review comprehensively explores AQbD principles, regulatory perspectives, and its applications, particularly in analytical method development, including high-performance liquid chromatography (HPLC) and high-performance thin-layer chromatography (HPTLC). Emphasis is placed on the symbiotic relationship between AQbD and analytical method validation (AMV), elucidating their collective role in ensuring reliable and accurate analytical results. Integrating AQbD in method transfer, automation, and control strategies underscores its pivotal role in achieving robust, efficient, and compliant analytical processes. The review delves into lifecycle management and continuous improvement, coupled with AQbD principles, ensuring sustained method reliability throughout the pharmaceutical product lifecycle. AQbD's significant contribution to pharmaceutical lifecycle management, optimization, and change control is explored, emphasizing its systematic, data-driven, and risk-based approach to method development, validation, and ongoing enhancement. This review illuminates AQbD's transformative impact on pharmaceutical analysis, aligning with industry trends toward quality, efficiency, and regulatory compliance.

Particle size by design: Standardizing measurements for complex topical drug product assessment

The physicochemical and biopharmaceutical properties of drug substances and dosage forms can be significantly influenced by particle size. However, the diversity of equivalent particle diameters generated by different methods poses a fundamental challenge in particle size analysis. This study aimed to develop an Analytical Quality by Design (AQbD) approach to accurately assess the particle size of a complex formulation – clobetasol propionate (CP) 0.5 mg/g cream – through automated microscopy (AM) and laser light diffraction (LD). Additionally, Raman spectroscopy was utilized to determine the chemical composition of the formulation particles. In the AQbD approach, prior knowledge was considered for the construction of the Ishikawa diagram and estimate failure mode and effects analysis (FMEA). The methods were developed following the ICH Q8-Q10, and ICH Q14 guidelines, and validated according to ICH Q2, ISO 13320:2020, and EP2.9.31./USP<429>. Results indicate that a trade-off between the techniques must be established for a particle size by design: while LD offers higher throughput and more precise values at the expense of peak resolution and broadening, AM has higher variability but more reliable information in terms of size and shape analysis. The validated methods successfully demonstrated the implementation of an AQbD method in the definition of particle size methods.

Analytical Quality by Design (AQbD) Principles in Development and Validation of Stability‐Indicating RP‐HPLC Method for Simultaneous Estimation of Diltiazem HCl and Eugenol in Nanoethosomes

ABSTRACTThe study developed and validated a robust reverse‐phase high‐performance liquid chromatography (RP‐HPLC) method for a simultaneous estimation of Diltiazem HCl and Eugenol in nanoethosomes using analytical quality by design (AQbD) principles. The optimized method utilized a mobile phase of methanol and 0.1% Orthophosphoric acid (OPA) in a 50:50 % v/v ratio, with a flow rate of 1.0 mL/min and isocratic elution on a Phenomenex Luna C‐18 column (5 µm, 150 mm × 4.6 mm) at 30°C. This setup resulted in retention times of 6.838 min for Diltiazem HCl and 23.135 min for Eugenol. Validation followed International Council for Harmonization (ICH) Q2 (R1) guidelines, demonstrating excellent linearity with correlation coefficients of 0.9982 for Diltiazem HCl and 0.9991 for Eugenol across the 5–25 µg/mL range. The limits of detection (LOD) were 1.341 µg/mL for Diltiazem HCl and 0.960 µg/mL for Eugenol, with limits of quantification (LOQ) at 4.065 and 2.912 µg/mL, respectively. Stability testing under acidic, alkaline, oxidative, thermal, and photolytic conditions adhered to ICH Q1A (R2) and Q1B guidelines. This AQbD‐based method is effective for routine analysis of Diltiazem HCl and Eugenol in bulk and pharmaceutical dosage forms, fully complying with ICH standards.

Robust analytical method for the enantiomeric separation of lurasidone hydrochloride: Integrating analytical quality by design and green chemistry principles

Schizophrenia, affecting approximately 1% of the global population, necessitates effective treatments like Lurasidone (LUR). However, the accurate separation of LUR from its enantiomeric impurities remains a significant challenge in pharmaceutical analysis. This study addresses this issue by developing and validating an optimized HPLC method for the simultaneous determination of LUR and its enantiomeric impurity using Analytical Quality by Design (AQbD) principles. The separation was achieved on a Chiralcel OD-H column with isocratic elution employing a hexane-ethanol mixture (92:8, v/v) at a flow rate of 0.9 mL/min, UV detection at 215 nm, and a column temperature of 32 °C. The method was validated using accuracy profiles. Environmental sustainability was assessed using various Green Analytical Chemistry (GAC) metrics, demonstrating favorable greenness attributes. This methodological framework not only enhances pharmaceutical quality control but also promotes sustainability in analytical practices, supporting its application in drug manufacturing and regulatory compliance.

Development of a QAMS Analysis Method for Industrial Lanolin Alcohol Based on the Concept of Analytical Quality by Design

The Analytical Quality by Design (AQbD) concept was adopted to establish a quantitative analysis of multi-components with a single marker (QAMS) method for industrial lanolin alcohol, targeting cholesterol, lanosterol, and 24,25-dihydrolanosterol. The potential critical method parameters (CMPs) were identified as column temperature, flow rate, and gradient. Definitive screening design and statistical modeling were employed to optimize the gradient conditions of the mobile phase, column temperature, and flow rate. The Method Operable Design Region (MODR) was determined using a risk-based quantification approach. The robustness was assessed using a Plackett–Burman experimental design, followed by methodological validation. Optimal analytical conditions were as follows: acetonitrile (B)—water (A) mobile phase system; flow rate of 1.58 mL/min; detection wavelength of 205 nm; injection volume of 10 µL; and column temperature of 37 °C. A gradient elution program was implemented as follows: 0–19.0 min, 90.5% B; 19.0–25.0 min, 90.5–100% B; and 25.0–55.0 min, 100% B. Cholesterol served as an internal standard for quantifying lanosterol and 24,25-dihydrolanosterol, with relative correction factors of 0.4227 and 0.8228, respectively. This analytical method utilized only the cholesterol reference substance as an internal standard to quantify the content of cholesterol, lanosterol, and 24,25-dihydrolanosterol in industrial lanolin alcohol. It reduced the testing costs and enhanced efficiency, making it potentially suitable for widespread adoption in lanolin alcohol processing industries.

Stability-indicating green HPLC method for fixed-dose tablets containing remogliflozin etabonate and teneligliptin: an AQbD approach

Background In June 2021, the Central Drug Standards Control Organization approved a fixed-dose combination tablet containing remogliflozin etabonate (100 mg) and teneligliptin (10 mg) to manage type II diabetes. Objective This study aims to develop a stability-indicating RP-HPLC method for quantifying remogliflozin etabonate and teneligliptin in tablet formulations via analytical quality by design (AQbD) principles. Methods Risk assessment, Plackett–Burman design, and central composite design were employed to understand the impact of independent variables on critical analytical attributes. The stationary phase was a HyperClone BDS C18 column, and the mobile phase consisted of acetonitrile and phosphate buffer (20 mM, pH 5) at a 45:55% (v/v) ratio. Results The method, validated per ICH Q2 (R1), resulted in retention times of 3.395 and 12.308 min for teneligliptin and remogliflozin etabonate, respectively. Forced degradation studies confirmed robustness, with clear peak separation and no interference from degradation products. The AGREE score of 0.65 supports its green applicability for tablet analysis in quality control. Conclusion The AQbD-assisted RP-HPLC method developed in this study offers environmental friendliness, efficient separation with well-defined peaks, and simple mobile phase combination.

Analytical quality by design-based stability-indicating high performance liquid chromatography method for the estimation of evogliptin tartrate in bulk and tablet dosage form.

The present study utilized Analytical Quality by Design (AQbD) approach to develop a stability-indicating high-performance liquid chromatography (HPLC) method for estimating evogliptin tartrate using design expert software. The key parameters were methodically optimized, contours were plotted, and stability was evaluated using various forced degradation conditions. Using an Agilent HPLC system with a photo diode array (PDA) detector along with Fortis C18 column (250 × 4.6mm, 5μm) effectively separated the drug from its degradants. The mobile phase used was methanol: water (pH adjusted to 3.0, 76:24; v/v) at 0.8mL/min flow rate. Evogliptin was eluted at 2.98 min, at a detection wavelength of 267 nm. The proposed method was found to be specific, precise, linear and robust. The drug was sensitive to acidic, basic, oxidative, thermal, and photodegradation resolving six degradation products. Thus, the developed AQbD-based stability-indicating HPLC method is applicable in analyzing evogliptin in bulk, tablet dosage form and stability samples.

Integrated Application of Risk Management Techniques in Developing an Analysis Method for Traditional Chinese Medicine: A Case Study of a Percolation Solution for Xiaochaihu Capsules

Risk management should run through the entire process of method development, utilization, and maintenance. Based on the analytical quality by design (AQbD) concept, various integrated risk management techniques were used in this study to develop an analysis method for the percolation solution of Xiaochaihu capsules. During the development of the analysis method, risk assessment was conducted using an Ishikawa diagram and failure mode effects analysis, followed by method optimization using experimental design. The probability of nonconformance calculated via an exhaustive Monte Carlo method quantitatively characterized the risk magnitude of method parameter failures, leading to the establishment of a operable design region method based on risk magnitude. Validation experiments and robustness tests of the data were utilized for model refinement and initial risk review. Methodological validation of the developed method was performed, and control strategies for the analysis method were presented through a decision tree. Stability experiments demonstrated that the samples remained stable at 4 °C for 24 h. The average recovery rate fell between 98.8% and 105%, with relative standard deviations ranging from 2.73% to 4.48%. The results showed that the established analysis method exhibited robustness. This analysis method can simultaneously determine the contents of uridine, adenine, 5-hydroxymethylfurfural, and guanosine. This method can also be employed for process control during percolation. This study integrated various risk management techniques to develop and maintain the analysis method, and this approach can potentially be extended to other analytical methods.

Chemometrics‐Empowered Optimization of a Reliable Reversed Phase–High Performance Liquid Chromatography Method for the Simultaneous Estimation of Erythromycin and Adapalene in Pharmaceutical Nanoformulation

ABSTRACTAn innovative, simple, economical, and sensitive reversed phase–high performance liquid chromatography (RP–HPLC) chromatographic method for simultaneous analysis of erythromycin (ERY) and adapalene (ADP) in bulk and pharmaceutical nanoformulation was developed by analytical quality‐by‐design (AQbD) avenue. According to published research, there is no strategy that integrates RP–HPLC with AQbD for estimating ERY and ADP. The analysis was carried out with Box–Behnken design, which reduced trial runs and design points. Acetonitrile: orthophosphoric acid (OPA) (0.1% triethanolamine) in pH 6.2 water was used as the mobile phase. The ratio was set to 35:65 (v/v) at 0.8 mL/min flow rate to detect the analytes at 231 nm. As per the guidelines in International Conference on Harmonization Q1A (R2), the developed approach was validated. ERY and ADP regression coefficients (r2) were found to be 0.9998 and 0.9999, respectively, over 5–25 µg/mL range. Validation parameters comprising linearity, sensitivity, system suitability, precision, robustness, and accuracy were performed. To assess stability, the drugs were subjected to stress conditions (hydrolytic, oxidative, thermal, and photolytic), and drugs were most vulnerable to degrade in alkaline and oxidative conditions. A new technique successfully analyzed novel drug carriers loaded with ERY and ADP.

Analytical Procedure Development and Proposed Established Conditions: A Case Study of a Mass Spectrometry Based NDSRI Analytical Procedure

With the finalization of the ICH Q14 Analytical Procedure Development guideline, how to apply enhanced approaches (such as analytical quality by design (AQbD)) to develop an analytical procedure, and to propose Established Conditions (ECs) and corresponding reporting categories, is increasingly being discussed. To gain practical experience in applying an enhanced approach for method development and identifying ECs, we developed, validated, and implemented an analytical procedure for a nitrosamine drug substance-related impurity (NDSRI). Here, as an example of the application of Q12 Lifecycle Management guideline principles in regards to analytical procedures, we briefly elaborate how: 1) the principles documented in the ICH Q14 guideline for analytical procedure development were applied, with the focus on identifying an Analytical Target Profile (ATP), knowledge management and risk assessment; 2) analytical procedure robustness according to the recommendations in ICH Q2(R2) Validation of Analytical Procedure guideline and Q14, were evaluated; and 3) mass spectrometry ECs and associated proposed reporting categories were proposed.

Green analytical chemistry and quality by design: A combined approach towards simultaneous determination of Letrozole with its co-administered Zoledronic Acid for cancer patients

Nowadays, breast cancer is the most affecting and globally diagnosed malignancy among women, yet Letrozole (LTZ) was considered the first-line treatment as hormonal anticancer drug. Unfortunately, LTZ develops osteoporosis as a main side effect which was overcome by using the co-administered; Zoledronic Acid (ZDA). Thus, there was a crucial need for this simultaneous quantification innovation, especially there were no any previously reported methods regarding both drugs together. In this study, an integrated framework was conducted between the experimental analytical quality-by-design (AQbD) approach and green analytical chemistry (GAC), emerging sensitive and robust RP-HPLC method. Box-Behnken Design was the developed model for optimizing an isocratic chromatographic separation on C18 Equisil® ODS (4.6 × 250 mm, 5.0 μm) column at ambient temperature, using the mobile phase of 0.1 % aqueous trifluroacetic acid (pH 2.8): acetonitrile (54.5:45.5, v/v), at 1.0 mL/min flow rate with PDA detection at 254.0 nm and 210.0 nm for LTZ and ZDA, respectively. Model statistical and residual plots analysis was significant and normally distributed. Method was fully validated as per ICH guidelines, where good linearity was 0.20–10.00 µg/mL for both drugs in presence of Tadalafil (TDF) as an internal standard, obtaining adequate correlation coefficients (r) values. Calculated LOD results were 0.058 and 0.040 µg/mL while calculated LOQ results were 0.175 and 0.122 µg/mL for LTZ and ZDA, respectively. The proposed method was effectively applied on bulk, pharmaceutical dosage forms, and spiked human plasma. Statistical comparison of the anticipated results with the reported ones was performed. Greenness assessment was evaluated using Green Analytical Procedure Index (GAPI) and Analytical Greenness (AGREE) tools; where superiority results were achieved relative to other reported methods. Finally, an EVG method evaluation tool was assessed, and the attained results were represented through its radar chart.

Box-Behnken design in optimization of the green liquid chromatographic method for the quantification of Afatinib in drug product: AQbD approach

The aim of this research work was to quantitatively determine the tyrosine kinase inhibitor, Afatinib, in tablets using an eco-friendly chromatographic method. The development of a green liquid chromatographic method using analytical quality-by-design (AQbD) approach principles to estimate Afatinib via Box-Behnken design. A simple and effective green liquid chromatographic method was developed and validated. The analysis was done on the Zodiac C18 column by isocratic elution mode. Mobile phase composed of pH 3.0 potassium dihydrogen phosphate buffer and ethanol in the ratio of 65:35%V/V. The flow rate was set to 0.6 mL/min. with an injection volume of 10 µL and at detection wavelength of 254 nm. The optimized method was found to be specific, linear, precise, accurate, and robust. Established linearity in the range of 24.6 to 73.9 µg/mL. The developed method exhibited excellent greenness. Based on the greenness assessment tools, the analytical eco-score value of 94 and the pictograms of NEMI, Modified NEMI, GAPI, Complex GAPI, AGREE, and AGREE prep expressed the method’s greenness. Hence, the AQbD approach based green method and the validation data indicate that the method can be employed in routine analysis for the determination of Afatinib in pharmaceutical dosage form in quality control laboratory use.

Analytical quality by design (AQbD) based optimization of RP-UPLC method for determination of nivolumab and relatlimab in bulk and pharmaceutical dosage forms

BackgroundThe Analytical Quality by Design (AQbD) methodology extends the application of Quality by Design (QbD) principles to the management of the analytical procedure life cycle, encompassing method creation, optimization, validation, and continuous improvement. AQbD assists in creating analytical procedures that are robust, reliable, precise, and cost-efficient. Opdualag™ is a combination of Nivolumab and Relatlimab, which are antibodies that block programmed death receptor-1 (PD-1) and lymphocyte-activation gene 3 (LAG-3) receptors, used to treat advanced melanoma. This work aims to develop and validate a reversed-phase ultra-performance liquid chromatography (RP-UPLC) method using AQbD principles to determine NLB and RTB in pharmaceutical products.ResultsA central composite design (CCD) comprising three factors arranged in five distinct levels was implemented via Design-expert® software to optimize the chromatographic conditions. A mathematical model was constructed and the effects of three independent factors namely flow rate (X1), percentage of methanol in the mobile phase (X2), and temperature (X3) on responses including retention time (Y1–2), resolution factor (Y3), theoretical plates (Y4–5), and tailing factor (Y6–7) were investigated. The software determined the optimal chromatographic conditions for the separation of NLB and RTB, which were as follows: 32.80% methanol in the mobile phase, 0.272 mL/min flow rate, 29.42 °C column temperature, and 260 nm UV detection. The retention time for NLB and RTB were 1.46 and 1.88 min, respectively. The method exhibited linearity across the concentration ranges of 4–24 µg/mL for RTB and 12–72 µg/mL for NLB. The limits of detection (LOD) and limit of quantification (LOQ) for NLB and RTB, respectively, were 0.89 µg/mL, 2.69 µg/mL and 0.15 µg/mL and 0.46 µg/mL. The percentage relative standard deviation (%RSD) of intraday and interday precision for NLB and RTB was below 2. The recovery percentages for NLB and RTB were determined to be 99.57–100.43% and 99.59–100.61%, respectively. Both drugs were found to be susceptible to oxidative and photolytic degradation in forced degradation studies.ConclusionsEmploying the AQbD-based methodology, a straightforward, fast, accurate, precise, specific, and stability-indicating RP-UPLC method has been established for the quantitative analysis of NLB and its RTB in pharmaceutical formulations.

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.

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.