RP-HPLC Method Research Articles

A Robust RP-HPLC Method for Simultaneous Quantification, Validation and Stability Studies on Vildagliptin (VILD), Dapagliflozin (DAPA) and Metformin Hydrochloride (METF) in its Combined Dosage Formulation

Background: Vildagliptin, Dapagliflozin and Metformin Hydrochloride are anti- hyperglycemic agents. Anti-hyperglycemic agents are drugs that stimulate insulin secretion and lower the glucose level in the blood. They were investigated as a medication for Diabetes Mellitus (DM). Various studies have reported the HPLC, HPTLC, LC/MS methods for the estimation of individuals of VILD, DAPA and METF. However, until date stability indicating HPLC analysis method has not been reported for the estimation of VILD, DAPA and METF in combined glucagon secretion in a glucose-dependent manner. Introduction: The objective of present work was to develop and validate the stability of RPHPLC method for the estimation of Vildagliptin (VILD), Dapagliflozin (DAPA) and Metformin Hydrochloride (METF) in combined dosage form. Method: In this research work, High Performance Liquid Chromatography (HPLC) was used for validation in the chromatography. In the HPLC, Methanol and Potassium Dihydrogen phosphate buffer (85:15 v/v) were used as mobile phases. HPLC analysis was performed at 230 nm. The method was validated with different parameters such as linearity, precision, accuracy, specificity, and robustness, limit of detection (LOD) and limit of quantitation (LOQ). The RF values of VILD, DAPA and METF were 6.24 ± 0.05 min, 8.34 ± 0.05 min. and 3.68 ± 0.05 min, respectively. The accuracies of drug recovery were 0.6398 %,0.3680 %, 0.6398 % and over the range of 1–4 μg/ml, 20-80 μg/ml and 15-85 μg/ml of VILD, DAPA and METF, respectively. Results: Degradation products found in stress conditions did not interfere with the detection of VILD, DAPA and METF; therefore, the proposed technique can be considered stable. Significant degradation products of VILD, DAPA and METF were obtained in an acid at METF at 2.05 ± 0.05 min. (MDP-1) and degradation product for oxidation of METF at 2.11 ± 0.05 min. (MDP-2). When alkaline hydrolysis was conducted, photolytic and thermal conditions were applied alone or in combination with METF, no degradation product of VILD, DAPA and METF was found. Conclusion: The developed and validation method was satisfactorily applied for the analysis of pharmaceutical preparations and proved specific and accurate for quality control of the cited drugs in their combined dosage form.

Risk assessment of antibiotic residues in raw cow’s milk in Bangladesh

ABSTRACT This study evaluated the presence of residual levels of commonly utilised antibiotics, including oxytetracycline (tetracyclines), ciprofloxacin, enrofloxacin, and levofloxacin (fluoroquinolones), in milk samples. Fifty raw milk samples were collected from five farms in Keraniganj, Dhaka. A validated RP-HPLC method was applied to detect and quantify antibiotic residues, demonstrating good linearity with coefficients ranging from 0.999 to 1.0 in the concentration range of 1.25–15.00 µg/mL. The study revealed that oxytetracycline was detected in 90% of the samples, followed by levofloxacin (66%), enrofloxacin (64%), and ciprofloxacin (62%). One farm showed the highest antibiotic prevalence, with oxytetracycline in all samples and levofloxacin, enrofloxacin, and ciprofloxacin in 80% of the samples. About 30% of the oxytetracycline-positive samples exceeded the MRL, while none surpassed the MRL for enrofloxacin and ciprofloxacin. Calculated human health risks appeared to be low, but children might face potential risks due to prolonged exposure.

Analytical Quality by Design aided RP-HPLC method for the estimation of Sunset Yellow in commercial food samples employing green ultrasound assisted extraction: Greenness, Blueness and Whiteness evaluation

Analytical Quality by Design aided RP-HPLC method for the estimation of Sunset Yellow in commercial food samples employing green ultrasound assisted extraction: Greenness, Blueness and Whiteness evaluation

Investigating the stability of a cerebral vasodilator drug using chromatographic methods: Evaluation of methods’ practicality and environmental aspects

A vinca alkaloid; vinburnine (VNB) is utilized as an effective vasodilator. As a cyclic amide-containing drug, it is likely susceptible to hydrolytic degradation. This study examined the degradation profile of VNB, findings indicated that VNB undergoes degradation solely in the presence of alkali, generating a carboxylic acid derivative (DEG). The present study aimed to design and apply green TLC-densitometric and RP-HPLC assays for concurrently measuring VNB and its degradation product for the first time. TLC-densitometric assay was carried out on silica gel 60 F254 TLC plates and a developing system of ethyl acetate: methanol: triethylamine (6:4:0.05, by volume) and detection at 230 nm. RP-HPLC method depended on a C8 column and a mixture of methanol: water (95:5, v/v). The rate of flow was 1 mL/min and UV detection at 230 nm. The proposed assays were used for prediction of the degradation behavior of VNB under the mentioned conditions and then applied for quantitation of VNB in its commercially available capsules. Four distinct metric approaches; National Environmental Method Index (NEMI), Analytical Eco-Scale, Green Analytical Procedure Index (GAPI), and Blue Applicability Grade Index (BAGI) were utilized to assess the chromatographic method’s ecological effect. Findings obtained from the provided methodologies were contrasted statistically with the stated HPLC method using Student’s t and F-tests. The analysis revealed that there were no significant differences between them. The established methods were verified in accordance with the recommendations of the International Council for Harmonization (ICH), and all the outcomes were deemed to fall within the permissible limit.

Design of Experiment and green analytical chemistry principles for developing a robust and sustainable Stability-Indicating HPLC method for Bempedoic acid impurity profiling

Bempedoic acid (BEM), a drug used to lower cholesterol levels in patients with atherosclerosis. Controlling its process-related impurities is crucial due to their potential toxicity. This study details the development of a RP-HPLC method for the analysis of four process-related substances in BEM. To ensure robust chromatographic performance meeting six sigma quality standards, the method was optimized using a combined approach of response surface methodology (specifically, an I-optimal design) and tolerance analysis, integrating predefined specifications for critical method attributes such as resolution, tailing factor, and number of theoretical plates into the developmental process. Chromatographic separation was achieved using a C18 column. The mobile phase consisted of 0.1 % phosphoric acid in water and acetonitrile, utilizing a stepped gradient elution profile. The flow rate was maintained at 0.9 mL/min, with UV detection performed at 210 nm. The column temperature was set to 20 °C. The method demonstrated linearity for BEM process related impurities in the range of LOQ to 30 μg mL−1. The method ability to differentiate the drug substance from its degradation products was confirmed through forced degradation study. The environmental sustainability of the developed method was assessed using multiple Green Analytical Chemistry (GAC) tools. The National Environmental Methods Index (NEMI) showed two green-shaded quadrants, and the Green Certificate modified Eco-scale awarded a score of 70. The Complementary Green Analytical Procedure Index (Complex GAPI) confirmed the method environmentally friendly profile. Additionally, The AGREE tool generated a score of 0.77, indicating strong overall greenness, while the AGREEprep tool, focused on sample preparation, yielded a score of 0.54. Other tools such as the Sample Preparation Metric of Sustainability (SPMS) and the HPLC Environmental Assessment Tool (HPLC-EAT) scored 8.42 and 569, respectively, further demonstrating the method green credentials. This robust and environmentally conscious analytical framework offers significant potential for application in quality control of BEM.

Comparative validation of UV-spectrophotometry and RP-HPLC methods for cefixime and moxifloxacin analysis

AimThis study presents the development and validation of UV-spectrophotometry and RP-HPLC methods for the simultaneous quantification of Cefixime Trihydrate (CEFI) and Moxifloxacin Hydrochloride (MOXI) in pharmaceutical formulations. MethodologyTwo UV-spectrophotometric methods, including the absorbance ratio (Q-Absorption) and First Order Derivative Spectroscopy, were developed and validated for their linearity, precision, accuracy, and sensitivity. Additionally, a robust RP-HPLC method using a C18 column and optimized mobile phase was employed for efficient separation and simultaneous estimation of CEFI and MOXI. All methods were validated in accordance with ICH guidelines, with system suitability parameters confirming the reliability of the RP-HPLC method for routine analysis. ResultsThe absorbance ratio and First Order Derivative methods demonstrated low %R.S.D values, high accuracy, and satisfactory sensitivity for both drugs. Similarly, the RP-HPLC method achieved high resolution, precision, and robustness. Statistical analysis through ANOVA revealed no significant differences between the methods in terms of accuracy and precision. The methods were applied to analyze marketed formulations, further confirming their applicability in routine quality control. ConclusionIn conclusion, the validated methods provide accurate, precise, and sensitive techniques for the simultaneous estimation of CEFI and MOXI, making them suitable for pharmaceutical quality control and regulatory compliance.

Stability-Indicating RP-HPLC Method Development and Validation for Simultaneous Estimation of Metformin, Sitagliptin and Dapagliflozin Bulk and in Tablet Dosage Form

Stability-Indicating RP-HPLC Method Development and Validation for Simultaneous Estimation of Metformin, Sitagliptin and Dapagliflozin Bulk and in Tablet Dosage Form

Integrative greenness, blueness and whiteness determination of analytical RP-HPLC method for identifying Imipenem, Cilastatin and Relebactam in combined tablet form followed by forced degradation studies

Most clinically preferred, US FDA and EU approved anti-infective drug, a multi-component formulation of Imipenem (IPM), Cilastatin (CS) and Relebactum (REL). The combination is used in medicating the most complicated, gram-negative bacterial infections. Henceforth, it requires the development of selective, precise, accurate, robust analytical method. Hence the present work comprises RP-HPLC method development for determination of IPM, CS and REL. The detection is achieved by using new and apt conditions i.e., Discovery C18 (4.6 × 150 mm, 5 µm) column, 0.01 N Potassium Dihydrogen Phosphate buffer: Acetonitrile (60:40) as mobile phase. The detection was found at short RT i.e., 2.32, 3.08, and 3.62-min for IPM, CS and REL correspondingly. Good sensitivity was observed, where LOD of 0.11 µg/ml (IPM), 0.14 µg/ml (CS), and 0.06 µg/ml (REL) was obtained. While, LOQ was noticed at 0.33 µg/ml (IPM), 0.43 µg/ml (CS) and 0.18 µg/ml (REL). As a counterpart, other study of forced degradation was also performed for the selected drugs at different extreme conditions. A sustainability evaluation has been done on the proposed method. The creative tools such as AGREE, GAPI, ChlorTox scale in green analytical counterpart were performed providing good score. Another BAGI software was opted as blue principle-evaluator, that resulted in a good score of 85 %. Highlighting the use of the most recent sustainability determination software, RGBfast model in present work with a good score of 74 %. Moreover, white and green evaluation has been performed in the form of comparative mode, for better understanding of the significance of current method.

Development of HPLC-UV method for determination of Nilotinib in spiked human plasma with greenness assessment

One of the approved second-generation tyrosine kinase inhibitors (Nilotinib) shows great efficacy and selectivity in the treatment of patients with chronic or accelerated phase chronic myelogenous leukemia, and in resistant/ intolerant to prior therapy cases. For the analysis of Nilotinib in synthetic mixture and human plasma samples, a new reversed-phase high-performance liquid chromatographic method was developed. Determination was performed successfully using RP–18 column and a mobile phase consisting of 10 mM potassium dihydrogen phosphate buffer (pH = 5.5), methanol, and acetonitrile in the ratio 37:37:26 (v/v/v). Isocratic mode of elution with flow rate 1.2 ml/min and UV detection at 265 nm were applied. The method was validated for linearity in the range 0.5–24.0 µg/ml and best accuracy and precision results were obtained at temperature 40 °C. The percentage recovery of the analyzed drug was in the range 88.0–106.0 %. Moreover, the proposed RP-HPLC method was evaluated in terms of greenness, whiteness and blueness using three newly developed ecological metrics – The Analytical Greenness software, White Analytical Chemistry and Blue Applicability Grade Index. Good results were obtained and the technique was proved effective, specific and suitable for implementation in routine quality control and clinical laboratory practice for therapeutic drug monitoring.

Development and Validation of RP-HPLC Method for Quantification of Resveratrol in a Resveratrol-Solid Lipid Nanoparticles Loaded Microneedles Patch

Development and Validation of RP-HPLC Method for Quantification of Resveratrol in a Resveratrol-Solid Lipid Nanoparticles Loaded Microneedles Patch

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

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

Analytical Method Development and Validation of an RP-HPLC Method for Stability-Indicating Determination of Idelalisib

Analytical Method Development and Validation of an RP-HPLC Method for Stability-Indicating Determination of Idelalisib

Development and Validation of Stability Indicating RP-HPLC Method for the Estimation of Pirtobrutinib and Characterization of its Degradants by LC - MS

Background: Pirtobrutinib is a novel non-covalent BTK inhibitor used to treat adult patients with relapsed/refractory mantle cell lymphoma and B-cell leukemias. The mechanism of action involves binding and inhibition of Bruton's tyrosine kinase (BTK). Objective: The main goal of the current work was to create a selective liquid chromatographic method (RP-HPLC) that is easy to use, accurate and exact for quantifying Pirtobrutinib and its degradation products, thereby seeking insight into the drug’s degradation behaviour. Methods: Using an isocratic mode and a 50:50 mixture of acetonitrile and buffer solution (0.1% orthophosphoric acid) as the mobile phase, a High Performance Liquid Chromatographic System with a PDA detector and an X-Bridge Phenyl column (150 x 4.6mm, 3.5μm) at a flow rate of 1.0 ml/min was able to achieve good chromatographic separation. At 219 nm, the detection was carried out. A retention time of 2.271 minutes was discovered. To ascertain the drug's degrading properties and stability, forced degradation tests were carried out, leading to the development of the RPHPLC method. The chemical structures of the degradation products were clarified and their fragmentation mechanisms were suggested using LC-MS. Results: The suggested approach demonstrated a linearity within the 25-150% (2.5 to 15μg/mL) concentration range, with a correlation coefficient of 0.9999. The precision of the system (measured by % RSD=0.49) and the method (measured by % RSD=0.86) were all within the acceptable limits set by ICH guidelines, with % RSDs less than 1% and less than 2% for system precision and method precision, respectively. The LOD was 0.3μg/mL, and the LOQ was1μg/mL. Pirtobrutinib underwent rigorous tests for forced degradation under the specified conditions outlined in ICH Q1 (R2) guidelines. Pirtobrutinib was primarily broken down in acidic, alkaline, peroxide, and thermal conditions. It was found to remain stable under reduction, photolytic, and hydrolytic conditions. Through LC-MS analysis, the chemical structures of the resulting degradation by-products were identified, along with the proposed pathways of their fragmentation. Conclusion: The current study gives insight into Pirtobrutinib’s degradation behaviour. Pirtobrutinib was stable in reduction, photolytic, and hydrolytic conditions but degraded more readily in acidic, alkaline, peroxide, and thermal environments. The degradation products were characterized as 4-carbamoyl- 5-chloro-3(4-((5-fluoro-2-methoxybenzamido)methyl)phenyl)-4,5-dihydro-1H-pyrazol-5-aminium (acid impurity, DP1), N-(4-(4,5-diamino-1H-pyrazol-3-yl)benzyl)-5-fluoro-2-hydroxybenzamide (alkali impurity, DP2), 5-amino-3-(4-((s-fluro-2-methoxybenzamido)methyl))-4,5dihydro-1H-pyrazol-5-aminium (peroxide impurity, DP3) and N-(4-(4-acetyl-5-amino-4,5-dihydro-1H-pyrazol-3-yl)benzyl)-5-fluoro-2- methoxybenzamide compound with λ1-oxidane (1:1) (thermal impurity, DP4) and their fragmentation pathways were proposed. This study presents the first ever reported method for the quantification of Pirtobrutinib. It ensures precise quantitation of Pirtobrutinib, a new Bruton's Tyrosine Kinase inhibitor, and its degradation products. The method's enhanced sensitivity and regulatory compliance ensure its consistent use in the quantification of Pirtobrutinib.

Analytical Quality by Design Assisted Ecofriendly RP-HPLC Method for the Simultaneous Estimation of Artificial Sweeteners in Commercial Food Samples Utilizing Green Ultrasound Assisted Extraction Technique: Greenness, Blueness and Whiteness Appraisal.

Acesulfame K (E950) and Saccharin Na (E954) are commonly utilized synthetic sweeteners that are added to various processed food products to improve the sweet flavor. It is essential to give priority to environmentally friendly technology while assessing them, as overuse of these sweeteners presents serious health hazards. The main aim of this study was to develop an AQbD aided ecofriendly RP-HPLC technique that can detect both Acesulfame K and Saccharin Na simultaneously incorporating green analytical chemistry (GAC) principles and white analytical chemistry (WAC); using ultrasound-assisted extraction (UAE) technique on commercial food samples. The usage of ethanol was in accordance with eco-friendly ideals due to its ease of use, speed, and lack of environmental impact. Rotatable central composite designs (rCCD) was used for method optimization and a mobile phase consisting of a 50:50 (v/v) mixture of ethanol and 1% aqueous acetic acid (v/v), the separation was carried out on a Zorbax column (SB-C18, 150 × 4.6 mm, 5 µm) with a flow rate of 1 ml/min and a detection wavelength of 217 nm. Acesulfame K had retention duration of 1.134 minutes and Saccharin Na of 2.134 minutes. Acesulfame K and Saccharin Na recovery rates varied among different commercially available food samples, ranging from 65-102% and 75-101%, respectively. At the defined operating point, the developed procedure displays conformity with the previously defined requirements for linearity, accuracy, sensitivity, and repeatability. The most accurate assessments of greenness were produced by the GAPI, AES, and AGREE tools. Results from the Red-Green-Blue 12 (RGB 12) algorithm for whiteness and BAGI for blueness indicate that the method is very practical, cost-effective, and environmentally friendly. The results of this study could pave the way for more eco-friendly and effective AQbD methods to be used in the future for evaluating various sweeteners using green solvents.

BOX-BEHNKEN DESIGN ASSISTED ECO-FRIENDLY RP-HPLC-PDA METHOD FOR THE QUANTIFICATION OF PACLITAXEL: APPLICATION TO EVALUATE THE SOLUBILITY OF PACLITAXEL-CYCLODEXTRIN COMPLEX

Objective: Paclitaxel (PTX) is one of the oldest chemotherapeutic agents for cancer treatment. However, PTX is a class IV drug under the Biopharmaceutical Classification System (BCS), and its oral administration is restricted due to its low bioavailability. Complexing PTX with Beta-Cyclodextrin (β-CD) is an option to overcome the low solubility and bioavailability. This study aims to optimize and develop an RP-HPLC analytical method for quantifying PTX from the fabricated β-CD complex. Methods: The HPLC settings were optimized using Design-of-Experiments (DOE) software. The independent variables for the optimization process were buffer ratio, buffer pH, flow rate, and injection volume. The responses were Retention Time (RT), peak area, Tailing Factor (TF), and number of Theoretical Plates (TP) of PTX. The validated method was then used to measure the % entrapment from the PTX-β-CD complex. Results: The developed and optimized RP-HPLC method was validated as per International Council for Harmonisation (ICH) Q2 (R1) guidelines. The developed method showed linearity R2 = 0.999 with a 0.5-20 µg/ml range. The Limit of Detection (LOD) and Limit of Quantification (LOQ) were 95 and 125 ng/ml, respectively. The accuracy and precision for the developed method came under the acceptance criteria. The developed method was used to evaluate the enhancement of solubility of the prepared PTX-β-CD complex. The method was also used in the evaluation of % drug loading, % drug release and stability of the PTX-β-CD complex. The study clearly showed that the solubility of PTX increased from 0 to 1.14±0.53 µg/ml at pH 1.2 and 0 to 3.18±0.61 µg/ml at pH 6.8, respectively. The PTX-β-CD complex showed 73±3.75% drug release in 120 min at pH 1.2 and 87±3.51% at pH 6.8. The developed RP-HPLC method was found to be eco-friendly as per the Analytical Greenness (AGREE) metric approach and software analysis. Conclusion: An eco-friendly RP-HPLC analytical method was successfully developed and optimized for the quantification of PTX from the PTX-β-CD complex.

Phytochemical Composition and Functional Properties of Brassicaceae Microgreens: Impact of In Vitro Digestion.

The aim of this study was to compare the concentration of phenolic compounds, glucosinolates, proteins, sugars and vitamin C between kohlrabi (Brassica oleracea var. acephala gongylodes), Savoy cabbage (B. oleracea sabauda), Brussels sprouts (B. oleracea gemmifera), cauliflower (B. oleracea botrytis), radish (Raphanus sativus) and garden cress (Lepidium sativum) microgreens for their antioxidant and hypoglycemic potential. In addition, we applied an in vitro-simulated system of human digestion in order to track the bioaccessibility of the selected phenolic representatives, and the stability of the microgreens' antioxidant and hypoglycemic potential in terms of α-amylase and α-glucosidase inhibition after each digestion phase. Using spectrophotometric and RP-HPLC methods with statistical analyses, we found that garden cress had the lowest soluble sugar content, while Savoy cabbage and Brussels sprouts had the highest glucosinolate levels (76.21 ± 4.17 mg SinE/g dm and 77.73 ± 3.33 mg SinE/g dm, respectively). Brussels sprouts were the most effective at inhibiting protein glycation (37.98 ± 2.30% inhibition). A very high positive correlation (r = 0.830) between antiglycation potential and conjugated sinapic acid was recorded. For the first time, the antidiabetic potential of microgreens after in vitro digestion was studied. Kohlrabi microgreens best inhibited α-amylase in both initial and intestinal digestion (60.51 ± 3.65% inhibition and 62.96 ± 3.39% inhibition, respectively), and also showed the strongest inhibition of α-glucosidase post-digestion (19.22 ± 0.08% inhibition). Brussels sprouts, cauliflower, and radish had less stable α-glucosidase than α-amylase inhibitors during digestion. Kohlrabi, Savoy cabbage, and garden cress retained inhibition of both enzymes after digestion. Kohlrabi antioxidant capacity remained unchanged after digestion. The greatest variability was seen in the original samples, while the intestinal phase resulted in the most convergence, indicating that digestion reduced differences between the samples. In conclusion, this study highlights the potential of various microgreens as sources of bioactive compounds with antidiabetic and antiglycation properties. Notably, kohlrabi microgreens demonstrated significant enzyme inhibition after digestion, suggesting their promise in managing carbohydrate metabolism and supporting metabolic health.

Stability indicating RP-HPLC method development and validation for quantification of impurities in gonadotropin-releasing hormone (Elagolix): Robustness study by quality by design.

The purpose of this research was to establish and validate a reverse phase HPLC method for the determination of Elagolix impurities in pharmaceutical dosage form. Mobile phase A, consisting of 10mM sodium dihydrogen phosphate (pH6.0) and acetonitrile in a 95:5v/v ratio, and mobile phase B, containing 85:10:5v/v/v of acetonitrile, Milli-Q water, and methanol, were used to achieve the method's specificity in the analytical column Kromasil 100-C18 (250 mm × 4.6mm, 5μm). The gradient program includes (%B/Time [min]: 36/0, 36/10, 38/15, 85/55, 85/65, 36/67, and 36/75). The flow rate is 0.8mL/min. The overall run duration is 75.0min, the injection volume is 10.0μL, and the detection is at 210 nm in UV. The samples were subjected to hydrolysis, oxidation, and heat conditions in order to facilitate their forced degradation. The procedure was validated and determined with the standards of ICH guidelines. From the LOQ to a concentration level of 200%, the linearity of the technique was ascertained. An accuracy range of LOQ to 150% was established for the method, and the average recovery was acceptable. Design of experiments, part of the quality by design idea, was used to prove the method's reliability.

Green analytical comparison and central composite design optimization for simultaneous estimation of pain management drugs using RP-liquid chromatography

The Central Composite Design method was utilized to validate a precise RP-HPLC method for concurrently determining the quantities of Paracetamol (PC), Diclofenac Sodium (DS), and Eperisone Hydrochloride (EH) in tablet compositions. By employing Design of Experiment (DOE), the experimental parameters were fine-tuned, resulting in an optimized eluent consisting of methanol: water (90:10) with 0.1 % orthophosphoic acid at a eluent velocity of 1 mL/min. The method exhibited exceptional purities: PC (100.83 % ± 0.85), DS (102.01 % ± 0.90), and EH (100.49 % ± 1.29). Regression equations were formulated for PC, DS, and EH as follows: y = 479762x + 151907, y = 2182788x + 2409442, and y = 777144x − 1146334, respectively. The analytical method underwent comprehensive validation, including tests for: Accuracy, Precision, Linearity and Robustness. To assess the method’s environmental impact, several Green Analytical Chemistry (GAC) tools were employed. These tools provided a multifaceted evaluation of the method’s sustainability and eco-friendliness.

A new method development and validation for simultaneous estimation of ramipril and telmisartan by RP-HPLC method in combined dosage form

The present research work was development and validation of new RP-HPLC method for simultaneous estimation of Ramipril and Telmisartan in combined dosage form. In simultaneous RP-HPLC method development, Waters 2695 Separations Module with PDA Detector and column used is C8 SB ZORBAX (150 X 4.6mm) column with 3.5-micron particle size. Injection volume of 10 µL is injected and eluted with the mobile phase selected after optimization was Phosphate buffer and Acetonitrile in the ratio of 70:30 was found to be ideal. The optimized flow rate was at 1.0 mL/min. Detection was carried out at 230 nm. This system produced symmetric peak shape, good resolution and reasonable retention times of Ramipril and Telmisartan were found to be 2.275 and 4.261 minutes respectively. The Ramipril and Telmisartan showed linearity in the range of 20-60 µg/mL and 160-480 µg/mL respectively. The %RSD values for precision was found to be within the acceptable limit, which revealed that the developed method was precise. The developed method was found to be robust and the %RSD value for percentage recovery of Ramipril and Telmisartan was found to be within the acceptance criteria. The results indicate satisfactory accuracy of method for simultaneous estimation of the Ramipril and Telmisartan.

Analytical Method Developement and Validation of Beclomethasone, Clotrimazole, Ofloxacin, Lidocaine by RP-HPLC in Combination Dosage Form

A rapid and precise method (in accordance with ICH guidelines) is required for the quantitative simultaneous determination of drugs in a combined pharmaceutical dosage form. The pursuit of desired quality is a continual challenge for pharmaceutical industries, necessitating a meticulous approach known as validation. Sensitive and specific RP-HPLC method involving UV detection was carried for determination and quantification of various drug in combination dosage form. The aim is "Analytical Method developement and Validation of Beclomethasone, Clotrimazole, Ofloxacin, Lidocaine by RP- HPLC In Combination Dosage Form " as per ICH guidelines. The developed method was validated for various parameters as per ICH guidelines like system suitability, specificity, linearity, system precision, method precision, accuracy, ruggedness and robustness. The separation method was carried out by using a mobile phase A consisting of Potassium dihydrogen phosphate buffer (0.05M): Methanol 45:55 v/v) pH 3.5 ± 0.1 and Mobile phase B comprises Acetonitrile in the ratio of 50:50 v/v. The detection was carried out by using UV detector at 300nm. The column was Agilent C 18 (250X4.6mm) 5µ. The flow rate was selected as 1.5 ml/min. The retention time of Ofloxacin, Lidocaine, Beclomethasone, and Clotrimazole, was found to be 5.67, 9.44, 18.29, and 21.85 respectively.