Liquid Chromatographic Method For Determination Research Articles

Development of a stability indicating high-performance liquid chromatography method for determination of cenobamate: study of basic degradation kinetics

This study presents a stability indicating high-performance liquid chromatography HPLC method for the determination of cenobamate (CNB) in presence of its main impurity (CNB H-impurity) and degradation products. The chromatographic separation was carried out on a Thermo BDS Hypersil-C18 column (150 × 4.6 mm; 5 μm) with a mobile phase consisting of a 50:50 (%v/v) ratio of methanol and purified water. The flow rate was maintained at 1.0 mL. min− 1. CNB was detected at 210 nm using a PDA detector. The column temperature was held at 40 °C.The retention time of the drug was found to be 3.2 min. Furthermore, the study investigates the degradation behavior of CNB under various stress conditions, including acidic, basic, oxidative, and light-induced degradation. The results indicate that CNB is particularly susceptible to basic degradation. Consequently, a comprehensive study of the basic degradation kinetics was conducted. The method was also successfully applied for the determination of CNB in its dosage form. The results also show that there is no co-elution from degradation products or excipients as indicated by the mass balance and peak purity values confirming the specificity of the proposed method and its applicability for routine analysis of CNB.

Micellar Liquid Chromatographic Method for Determination of Moxifloxacin and its Impurities

A selective eco-friendly micellar HPLC method was developed for investigation of moxifloxacin and related compounds in the presence of its degradation products. Central composite design was used to optimize the experimental conditions. The proposed method is based on isocratic elution on a C18 column using 92.5% (v/v) biodegradable aqueous mobile phase containing 0.01 M sodium dihydrogen phosphate, 0.15 M sodium dodecyl sulfate (SDS) and 0.5% triethylamine (v/v) with a pH of 3.5 and 7.5% isopropanol (v/v) as eco- friendly organic solvent. The flow rate and injection volume were 0.6 ml/min and 5 μl, respectively. Experiments were performed at a temperature of 60 °C and detection was performed at 295 nm. The optimized method was validated. The method was found to be suitable for the quantification of moxifloxacin and its related compounds in moxifloxacin drug substance. The Green Analytical Procedure Index (GAPI) proves the superiority of the developed method against other reported methods.

Highly selective and sensitive high performance liquid chromatography method for determination of fumonisins from feed samples with MIL-101 based dispersed micro-solid-phase extraction and precolumn fluorescence derivatization

A selective and rapid high-performance liquid chromatography with fluorescence detection (HPLC-FLD) method for determining fumonisins was established and validated, which based on dispersed microsolid-phase extraction (D-μ-SPE) and precolumn derivatization. The MIL-101 was prepared as sorbents to selectively enrich two fumonisins from feed samples. The obtained extracts were labeled by a novel derivatization reagent 2,5-dioxopyrrolidin-1-yl 4-(1-methyl-1H-phenanthro[9,10-d]imidazol-2-yl)benzoate (MPIBA-Su). There was no fluorescence from fumonisins itself, while their derivatives exhibited considerable fluorescence with a maximum excitation at λex 255 nm and a maximum emission at λem 457 nm. Key factors affecting extraction efficiency and derivatization efficiency were fully optimized. Excellent linearity was achieved with correlation coefficients > 0.9935 under optimal conditions. The detection limits (LODs) were between 2.4 and 3.5 ng·mL−1, and the quantitation limits (LOQs) were between 8.1 and 11.7 ng·mL−1. With recoveries in the range of 83.8% − 94.3%, the established method has successfully been applied to the quantitative analysis of fumonisins in feed samples.

Development and validation of high-performance liquid chromatography method for determination of clarithromycin in pharmaceutical tablets.

Clarithromycin is a very important macrolide antibiotic used to treat bacterial infections in human and veterinary medicine. This study reports the development and validation of cost-effective, simple, precise, accurate, and robust high-performance liquid chromatography (HPLC) for the determination of clarithromycin (CLA) in tablets. Reversed-phase chromatography was conducted using a standard column at 55°C with ultraviolet detection at 215nm. A mobile phase consisting of acetonitrile -2-methyl-2-propanol -potassium phosphate buffer was used at a flow rate of 1.0mL/min. The proposed method displayed good linearity, precision, accuracy, robustness, and specificity. The present HPLC was compared with capillary electrophoresis and bioassay methods and the results indicated that there was no significant difference between these methods. Moreover, the obtained results demonstrated the validity of the isocratic HPLC, which allows reliable quantitation of CLA in pharmaceutical samples. Thus, it can be used as a substitute alternative methodology for the routine quality control of this medicine, in situations where other methods are less accessible in the laboratory.

GREEN PROCEDURE INDEX ASSESSMENT OF THE NOVEL STABILITY-INDICATING RP-HPLC METHOD FOR THE DETERMINATION OF CAPTOPRIL FROM PHARMACEUTICAL DOSAGE FORM

Objective: In this study, it was aimed to develop a novel reverse-phase liquid chromatography method for the ultra-sensitive determination of the antihypertensive drug captopril, using paracetamol, which is the common pain killer, as the internal standard. Optimization of all experimental conditions including composition of mobile phase, flow rate, and column temperature was carried out step by step, and the method validity of the developed method was examined according to international validation guidelines. Calibration range, linearity, the limit of determination, the limit of quantification, robustness, accuracy from commercial tablet samples, and method stability were examined in detail. In addition, the greenness profile for the developed method was assessed with the Green Analytical Procedure Index and Analytical Greenness Calculator techniques, which are frequently used in the literature. Material and Method: The chromatographic method was conducted with an XBridge C18 column (25 cm x 4.6 mm ID; 5 µm) packed with fully porous silica materials. All analyses were performed isocratically with a mobile phase containing acetonitrile:5 mM, pH 7.0 ammonium acetate solution (50:50, v/v) at a flow rate of 1.5 ml min-1. The injection volume was 5 μl, and the column was kept at 25°C in a column oven. The column eluate was monitored at 220 nm. Under optimized conditions, retention times of captopril, and paracetamol were approximately 1.59, and 2.0 min, respectively. Result and Discussion: This study described a fully validated, simple, sensitive, accurate, linear, precise, and reproducible reversed-phase liquid chromatography method for the determination of captopril in tablet samples. Under optimal experimental conditions, the linear range was found in the range of 0.5-200 µg ml-1 and the correlation coefficient was greater than 0.99. Method precision was acceptable, with coefficients of variation between 0.05% and 0.61%. In addition, as a result of the recovery studies carried out on the tablet samples, the accuracy was found to be within satisfactory limits between 99.45% and 102.55%. Moreover, the greenness profile of the developed method also showed that the method is environmentally friendly.

Development and Validation of Stability Indicating High Performance Liquid Chromatography Method for Determination of Leflunomide

Development and Validation of Stability Indicating High Performance Liquid Chromatography Method for Determination of Leflunomide

Green micellar stability-indicating high-performance liquid chromatography method for determination of rupatadine fumarate in the presence of its main impurity desloratadine: Oxidative degradation kinetics study.

A green micellar stability-indicating high-performance liquid chromatography method was developed for rupatadine fumarate determination in existence with its main impurity desloratadine. Separation was attained using Hypersil ODS column (150×4.6mm, 5μm), the micellar mobile phase consisted of 0.13M sodium dodecyl sulfate, 0.1M disodium hydrogen phosphate adjusted by phosphoric acid to pH 2.8 and 10% n-butanol. The column was maintained at 45◦ C and detection was carried out at 267nm. A linear response was achieved over the range of 2-160μg/ml for rupatadine and 0.4-8μg/ml for desloratadine. The method was applied for rupatadine determination in alergoliber tablets and alergoliber syrup without the interference of methyl paraben and propyl paraben present as main excipients. Rupatadine fumarate revealed pronounced susceptibility to oxidation; further study of oxidative degradation kinetics was carried out. Rupatadine was found to follow pseudo-first-order kinetics when exposed to 10% H2 O2 at 60 and 80°C and the activation energy was found to be 15.69Kcal/mol. At a lower temperature (40°C), degradation kinetics regression was best fitted as a polynomial quadratic relationship, thus rupatadine oxidation at a lower temperature tends to adopt a second-order kinetics rate. Oxidative degradation product structure was revealed using infrared and found to be rupatadine N-oxide at all temperature values.

Development and Validation of Stability Indicating High Performance Liquid Chromatography Method for Determination of Molnupiravir in Capsule Dosage Form

Development and Validation of Stability Indicating High Performance Liquid Chromatography Method for Determination of Molnupiravir in Capsule Dosage Form

Box–Behnken Design Based Development of UV-Reversed Phase High Performance Liquid Chromatographic Method for Determination of Ascorbic Acid in Tablet Formulations

A simple, sensitive, accurate and inexpensive UV-reversed-phase high-performance liquid chromatographic method was developed for the determination of ascorbic acid in tablet formulations. The method was based on the separation of ascorbic acid using a mobile phase of an acetonitrile-NaH2PO4-H3PO4 buffer solution (pH = 3) (5:95 v/v) with a UV detection wavelength of 245 nm and a flow rate of 0.8 mL min−1 at ambient column temperature. The variables of the proposed method, such as acetonitrile fraction (%), flow rate (mL min−1) and column temperature (°C), were optimized on the peak area by response surface methodology via the Box–Behnken design. The mobile phase was passed isocratically, and the separation of ascorbic acid was performed at the retention time of 4.1 min. A calibration graph was obtained and found to be linear in the concentration range of 10–180 µg mL−1. The method suitability was assessed and an asymmetry factor of 1.15 was obtained. The proposed method was successfully applied for the determination of ascorbic acid in tablet formulations and statistically compared with the results of the reference method. The performance of the proposed method was excellent and in agreement with the reference method. The recovery percentage of the proposed and reference methods was in the range of 99.98–100.04% and showed compliance (100 ± 2%) with regulatory guidelines.

Method development and validation of spray content uniformity for ipratropium bromide and budesonide in nasal spray

Ipratropium bromide and Budesonide is combined Pharmaceutical Nasal dosage form. The label claim of this combined dosage form is 21 mcg of Ipratropium Bromide and64 mcg of Budesonide per Spray. It is for treatment of allergic rhinitis. Spray content uniformity is in-vitro investigational test which provides idea of uniformity of product throughout product life cycle. The present study aimed to validate High Performance Liquid Chromatography method for determination of spray content uniformity of these analytes. This validation study performed as per guidelines of International Conference on Harmonization and covers Precision, Linearity, Accuracy, Robustness, Ruggedness, stability of analytical solution and Specificity. The reversed phase chromatographic method uses a mobile phase prepared by mixing Buffer containing Sodium dihydrogen orthophosphate with Acetonitrile, Zorb ax CN150 mm column, 1.5 ml per min flow rate, 220 nm wavelength, 25°C column oven and 50 μL injection volume. The method found linear over the range of 50% to 150% of target concentration with correlation coefficient of 0.9997 for Ipratropium bromide and 0.9999 for Budesonide. The recoveries obtained were 103.1% - 103.8% for Ipratropium bromide and 97.0% - 100.1% for Budesonide. %Related Standard Deviation found well within the acceptance criteria for precision. No interference found at the retention time of analyte peaks in blank, placebo and degradation conditions describe specificity of method. The method showed a successful application for determination of Ipratropium bromide and Budesonide in Nasal spray pharmaceutical formulation.

Development of stability indicating reversed-phase high-performance liquid chromatography method for determination of elobixibat in pure form and laboratory prepared tablets: Application to dissolution study.

A simple stability-indicating reversed-phase high-performance liquid chromatography method has been developed for determination of elobixibat in presence of its potential impurities and degradation products. The chromatographic separation was carried on a Thermo scientific Base Deactivated Silica BDS Hypersil-C18 (150 × 4.6mm; 5μm) column using a mobile phase of acetonitrile and phosphate buffer (25mM, pH 2.5) in a ratio of (70:30, v/v). The experimental conditions were accurately investigated, and the method was validated according to ICH guidelines Q2 (R1). The drug was subjected to various stress conditions including acidic, basic, oxidative, and photolytic conditions. The method successfully separates the drug from the three reported impurities and different degradants. The method was also successfully applied for the determination of elobixibat in laboratory prepared tablets (5.0mg). Analysis shows no interference from excipients and degradation products. The method was also applied for performing in vitro dissolution testing of elobixibat laboratory prepared tablets. Since elobixibat is recently introduced into the market, there are no previous stability studies and no reported analytical methods for its determination. Thus, this study presents a validated and selective method that can be effectively employed in routine quality control studies.

Neuroprotection by resveratrol: A review on brain delivery strategies for Alzheimer’s and Parkinson’s disease

Development and Validation of a New Simple Liquid Chromatographic Method for Determination of Coumarin and its 7-Hydroxy Metabolite as a Marker of Cytochrome P450 2A6 in RatsSundeep Bath, Wael Ebied, Hytham Ahmed, Fawzy Elbarbry

A novel thiolysis-high-performance liquid chromatography method for the determination of proanthocyanidins in grape seeds.

A novel thiolysis-high-performance liquid chromatography method for the quantitative determination of total proanthocyanidins and the mean degree of polymerization in grape seeds has been developed. Following thiolysis with formic acid and benzyl mercaptan, reaction products were separated and purified. Three proanthocyanidin monomers and three derivatives were obtained and their structures were identified by liquid chromatography-mass spectrometry, FTIR spectroscopy, and NMR spectroscopy. A decomposition model of the thiolysis products and a correction formula for proanthocyanidins concentration were established. This thiolysis-high-performance liquid chromatography method displayed good calibration linearity (R2 >0.999 over the concentration range 0.01 to 10mg/mL), excellent accuracy (recoveries of 97.9-99.6%), and precision (repeatability relative standard deviations of 0.45-0.75%). This method is suitable for the quantitative analysis of proanthocyanidins in grape seed products.

Development and validation of liquid chromatography method for determination of Ibrutinib in finished dosage forms using quality by design approach

AbstractA simple, precise, linear, robust, accurate, and stability‐indicating liquid chromatography method was developed and validated to determine Ibrutinib in finished solid dosage forms by reversed‐phase high‐performance liquid chromatography/photodiode array detector. Separation was achieved using a Luna C18 150 mm × 4.6 mm × 3.0 μm with a suitable mobile phase. The mobile phase: A consists of a 10 mM phosphate buffer concentration and 1 mL triethylamine, adjusted to pH 5.6 with a diluted orthophosphoric acid solution and acetonitrile in the ratio of 95:5 v/v to enhance the results. Moreover, mobile phase ‐ B contains 85% of acetonitrile. The optimized chromatographic conditions such as flow rate 1.0 mL/min, injection volume 10 μL, column temperature 40°C, and wavelength 258 nm. The proposed method is significant because it was developed by the Quality by Design approach. All impurities were identified, such as degradation and placebo impurities without interference at principle peak retention time. After development, it has been validated as per current regulatory guidelines. Obtained recovery between 98 and 102% at 50–200% level; linearity found (r2) = 0.9999 from 20 to 200% level; precision results achieved <2.0% relative standard deviation; and the robustness study results are found satisfactory. It's beneficial.

Determination of Alpha-Lipoic Acid in a Nutritional Supplement Using High Performance Liquid Chromatography

Alpha lipoic acid has the ability to react and neutralize reactive oxygen species (ROS) such as superoxide radicals, simple oxygen, hydroxyl radicals, hypochlorous acid and peroxyl radicals. A rapid high-performance liquid chromatographic method for determination of lipoic acid in a nutritional supplement was developed. The method involved sample preparation and the mobile phase comprised of 50 mM disodium hydrogen phosphate buffer (pH 2.5 adjusted with 1 M H3PO4): acetonitrile in the ratio of 50:50. The separation was done using a C18 column (150 mm) and detection was carried out using UV detection at 201 nm. The assay was found to be linear in the range of 1.56 - 50 μg/mL with the correlation coefficient of 0.9997. Method precision was determined while LOD was 0.05 μg/mL and LOQ 0.15 μg/mL. The chromatographic peak LA retention time was 6 min.

Development of a rapid and reliable high-performance liquid chromatography method for determination of water-soluble vitamins in veterinary feed premix.

Background and Aim:Determination of trace amounts of vitamins in multi-component feed premix is a troublesome analytical procedure. In this study, a simple and rapid high-performance liquid chromatography (HPLC) method was developed and validated for the concurrent detection and quantitation of four water-soluble vitamins such as thiamine, riboflavin, pyridoxine, and cyanocobalamin in veterinary feed premixes.Materials and Methods:The chromatographic separation of the vitamins was carried out at 35°C temperature on a reversed-phase C18 column using a gradient pump mode. Mobile phase constituents were solvent (a): 25 mM Potassium dihydrogen phosphate and 5 mM sodium hexanesulfonate in deionized water having pH-4.0 and solvent and (b) 5 mM sodium hexanesulfonate in methanol. Detection was performed with HPLC ultraviolet/visible detection set at 278 and 361 nm wavelength in two different channels. The flow rate was 1.2 mL/min and the total run time was 25 min.Results:The method was validated according to the International Conference on Harmonization and Food and Drug Administration guidelines and acceptance criteria for system suitability, precision, linearity, and recovery were met in all cases. The relative standard deviation for system suitability and precision was <2% for all vitamins. The linearity of the calibration curves was excellent (R2>0.999) at concentration of 5, 10, 15, 20, 25, and 30 μg/mL for all vitamins. The limits of detection values were 0.0125, 0.0017, 0.0064, and 0.0065 μg/mL for thiamine, riboflavin, pyridoxine, and cyanocobalamin, respectively, and the limits of quantification values were 0.0378, 0.0051, 0.0213, and 0.0198 μg/mL for thiamine, riboflavin, pyridoxine, and cyanocobalamin, respectively. The recovery percentages ranged from 88% to 115%.Conclusion:The overall parameters of the proposed method met the validation criteria and this method could be a highly desirable technique for routine analysis of water-soluble vitamins in veterinary feed premix.

Simple, Accurate and Efficient High Performance Liquid Chromatographic Method for Determination of Vandetanib in Bulk and in Pharmaceutical Forms

A simple, selective, linear, precise and accurate isocratic RP-HPLC method was developed and validated for rapid assay of Vandetanib, an anticancer drug, in both bulk and tablet dosage form. Elution at a flow rate of 1ml/min was employed on a symmetry C18 column at ambient temperature. The mobile phase consisted of acetonitrile, water and orthophosphoric acid in the ratio of 90:08:02 (v/v/v). Linearity was observed in concentration range of 50-200 ppm. The retention time for Vandetanib was 3.326 min. The method was validated as per the ICH guidelines. The proposed method can be successfully applied for the estimation of Vandetanib in pharmaceutical dosage forms. Moreover the detection alone was also verified through LC-MS of the Vandetanib drug using ESI method which provides future scope for study of this drug using LC-MS method also.

Design of experiments in optimization and validation of hydrophilic interaction liquid chromatography method for determination of amlodipine besylate and its impurities

Abstract A new and rapid hydrophilic interaction liquid chromatographic method has been developed for the quantitative analysis of amlodipine besylate and its specific impurities (D, E, and F). For development of this method, a systematic approach which includes Design of Experiments methodology was applied. For the method optimization, Box–Behnken design and specific way Derringer's desirability function were applied. They provided identification of the optimal chromatographic conditions on the basis of obtained mathematical models and graphical procedures (three D graphs). The optimal chromatographic conditions were the analytical column ZORBAX NH2 (250 × 4.6 mm, 5 µm particle size); mobile phase consisted of acetonitrile-water phase (50 mM ammonium acetate, pH adjusted to 4.0 with glacial acetic acid) (90.5:9.5 v/v); column temperature 30 °C, mobile phase flow rate 1 mL min−1, wavelength of detection 230 nm. As other validation parameters were also found to be suitable, the possibility to apply the proposed method for the determination of amlodipine besylate and its impurities in any laboratory under different circumstances has been proven.

A liquid chromatographic method for determination of acetamiprid and buprofezin residues and their dissipation kinetics in paddy matrices and soil.

The present study was conducted to investigate the residue status of two insecticides (acetamiprid and buprofezin) and their dissipation kinetics in three matrices viz. paddy grain, straw, and soil. The extraction procedure for residues of these two insecticides was executed using acetonitrile solvent. The analytical method was validated, which showed good linearity with the limit of quantification (LOQ) value of 0.01 and 0.02 mg kg-1 for acetamiprid and buprofezin, respectively. The recovery range was 79.67-98.33 % concerning all the matrices in both the insecticides. Acetamiprid (20% SP) and Buprofezin (25% SC) were applied separately in the paddy field in two doses: single dose (recommended dose) and double dose along with untreated control throughout the experiment. Residue analysis of these two insecticides in paddy (grain and straw) and soil was accomplished employing high-performance liquid chromatography (HPLC) with ultraviolet (UV) detector and confirmed by ultra-performance liquid chromatography (UPLC) coupled with mass spectrometry (UPLC-MS/MS). The dissipation data showed that acetamiprid exhibited higher dissipation in comparison with buprofezin. However, their persistence was found slightly higher in soil. The dissipation dynamics in the rice and soil were discussed with biological half-lives of both the insecticides. Consumer risk assessment study was also made considering its fate to the consumers.

High performance liquid chromatographic method for determination of active components in lithospermum oil and its application to process optimization of lithospermum oil prepared by supercritical fluid extraction

紫草提取制备成的紫草油能够预防及治疗婴儿尿布疹、皮肤溃烂、湿疹等多种皮肤疾患,临床应用非常广泛,超临界流体萃取是紫草有效成分提取的优选方法。该文建立了紫草油有效成分的高效液相色谱(HPLC)测定方法,并以紫草油所含的有效成分含量为评价指标,采用三因素三水平正交试验法对紫草超临界流体萃取制备过程中的几个重要因素(萃取压力、萃取温度和CO2流量)进行考察,确定了紫草超临界流体萃取的最佳工艺流程。所建立的HPLC方法如下:Diamonsil C18色谱柱(250 mm×4.6 mm, 5 μm),流动相为含0. 1%甲酸的乙腈-含5 mmol/L甲酸铵的0.1%甲酸水溶液(75:25, v/v),等度洗脱,流速为1 mL/min;进样量为15 μL;柱温为室温;二极管阵列检测器(PAD)检测波长为275 nm。该法可在30 min内同时测定紫草油中的有效成分紫草素、乙酰紫草素、β-乙酰氧基异戊酰阿卡宁、异丁酰紫草素、β,β-二甲基丙烯酰紫草素和2-甲基丁酰基紫草素的含量,方法精密度、准确度、重复性好。在萃取压力23 MPa、萃取温度40 ℃、CO2流量27 L/h这一优化工艺流程下得到的紫草油有效成分含量最高。所建立的HPLC-PAD法简便可行、准确可靠,可用于超临界流体萃取制备紫草油的工艺过程优化和质量控制,也可为紫草及其制剂的质量评价提供参考。优化后的工艺条件能够满足紫草油制备需求,也符合生产实际需求。