Development and Validation of RP-HPLC Method for Simultaneous Determination of Decitabine and Repaglinide: Application to Solid Lipid Nanoparticle Formulations with In Vitro Neurotoxicity Assessment

The present study aimed to develop a validated RP-HPLC method for the simultaneous determination of timolol maleate (TM), moxifloxacin hydrochloride (MOXI), diclofenac sodium (DS) and dexamethasone (DEXA) in human plasma, bovine aqueous humor and pharmaceutical preparations. The chromatographic separation was studied using the C18 column. The chromatographic conditions, such as composition, pH, the flow rate of mobile phase, the temperature of column, wavelength of absorption and injection volume of the sample, were studied. The method was validated to confirm specificity, linearity and accuracy in accordance with an International Conference on Harmonization guidelines. The optimum conditions for separation included mobile phase 0.05M monobasic phosphate buffer: acetonitrile (65:35v/v), pH of buffer adjusted to 6.2 and the flow rate of 1mL/minute. The optimum temperature of the column was found to be 35°C, absorption wavelength 265nm and injection volume 50μL. The baseline separation of all four drugs with good sensitivity, resolution, and a less than 15 min run time was achieved. The retention time of TM, MOXI, DS and DEXA were 4.3,5.7,9.9 and 13.5 minutes respectively. The limit of detection (LOD) values were 6.2, 4.8, 0.8 and 1.2 ng/mL for TM, MOXI, DS and DEXA, respectively, whereas their respective limit of quantification (LOQ) values was: were 42.6, 26.8, 5.6 and 6.2 ng/mL. The coefficient of variation for intra-day and inter-day were in the range of 0.32-1.57 and 1.29-3.07%, respectively. The method was found to be sensitive, precise and accurate in human plasma and bovine aqueous humor and can be applied for the quantification of these compounds in plasma, aqueous humor and pharmaceuticals.

The paper reports an efficient and rapid fast LC method for estimation of diclofenac potassium and its impurities. The main aim of the study is to develop and validate a simple, rapid, accurate, sensitive, less time consuming and less expensive method by using RP-HPLC with UV detector. The chromatographic separation of diclofenac potassium and its impurities was carried out by using 150 × 4.6 mm, i.d., 5 μm C-18 column with prepared mobile phase-A consisting 800:200 (v/v) of 0.01 M ammonium acetate adjusted pH 5.3 with acetic acid and acetonitrile and mobile phase-B consisting 200:800 (v/v) of 0.01 M ammonium acetate adjusted pH 5.3 with acetic acid and acetonitrile. The wavelength for detection of were made at 280.0 nm using UV detector. The flow rate of 1 mL/min. The method was able to detect and separate impurities and diclofenac potassium. The system suitability parameters were found within the limits. The coefficient of correlation for diclofenac potassium and its impurities was found not less than 0.998. The %recovery was found within the ICH limits. The LOD and LOQ values from study demonstrate method is sensitive. The method was validated for linearity, precision, accuracy, specificity, intermediate precision, ruggedness, robustness, stability and suitability.

Recently, a new formulation containing metformin HCl (MFH) and remogliflozin etabonate (RGE) has been approved for the management of diabetes mellitus. However, only one analytical method has been reported for the simultaneous determination of both the analytes. Therefore, the current study was designed to develop simple UV derivative spectroscopic and rapid RP-HPLC methods for simultaneous determination of MFH and RGE. The chromatographic separation of MFH and RGE was performed using a monolithic C18 column with an optimized chromatographic conditions carried out by full factorial Box–Behnken design model. The spectroscopic technique was based on the determination of peak amplitude of second-order derivative UV spectra at zero crossings. Further, both the methods were validated and compared statistically using Student’s-t-test and F-test, and employed for the concurrent estimation of MFH and RGE in laboratory mixed solutions and formulations. Perturbation plots and response surface models showed the effect of chromatographic parameters and the final chromatographic condition was selected from 47 solutions suggested by the desirability function. Further, UV spectroscopic and HPLC procedures showed good linearity in the range of 1–24 µg/mL and 2–150 µg/mL for RGE and 2–30 µg/mL and 5–200 µg/mL for MFH, respectively. The average percent assay was found to be 99.51% and 99.80% for MFH and 99.60% and 100.07% for RGE by spectroscopic and HPLC methods, respectively. The proposed methods were simple, accurate, precise, and rapid. Therefore, they can be used for regular quality control of MFH and RGE formulations and dissolution studies as well.

In this article we describe development and validation of stability indicating, accurate, specific, precise and simple Ion-pairing RP-HPLC method for simultaneous determination of paracetamol and cetirizine HCl along with preservatives i.e. propylparaben, and methylparaben in pharmaceutical dosage forms of oral solution and in serum. Acetonitrile: Buffer: Sulfuric Acid (45:55:0.3 v/v/v) was the mobile phase at flow rate 1.0 mL min-1 using a Hibar® Lichrosorb® C18 column and monitored at wavelength of 230nm. The averages of absolute and relative recoveries were found to be 99.3%, 99.5%, 99.8% and 98.7% with correlation coefficient of 0.9977, 0.9998, 0.9984, and 0.9997 for cetirizine HCl, paracetamol, methylparaben and Propylparaben respectively. The limit of quantification and limit of detection were in range of 0.3 to 2.7 ng mL-1 and 0.1 to 0.8 ng mL-1 respectively. Under stress conditions of acidic, basic, oxidative, and thermal degradation, maximum degradation was observed in basic and oxidative stress where a significant impact was observed while all drugs were found almost stable in the other conditions. The developed method was validated in accordance with ICH and AOAC guidelines. The proposed method was successfully applied to quantify amount of paracetamol, cetirizine HCl and two most common microbial preservatives in bulk, dosage form and physiological fluid.

A new, simple, reliable, fast, sensitive and economical RP-HPLC method was developed and validated for simultaneous estimation of two fixed-dose combinations frequently prescribed in coexisted chronic diseases such as diabetes (GLB) and hypertension (ATN) in bulk for the first time. The mobile phase used for the chromatographic runs consisted of 0.01N potassium dihydrogen ortho phosphate (pH 4.8) and acetonitrile (55:45, v/v). The separation was achieved on column (BDS C18 250 x 2.1mm, 1.6m) using isocratic mode. Drug peaks were well separated and were detected by a UV detector at 235.0 nm. The method was linear at the concentration range 2.5-15µg/ml for Glibenclamide (GLB) and 6.25-37.5µg/ml for Atenolol (ATN), respectively. The method has been validated according to ICH guidelines with respect to system suitability, specificity, precision, accuracy and robustness. The method was validated for system suitability, linearity, accuracy, precision, detection, quantification limits and robustness and was found it is acceptable in the range of 2.5–15 µg/ml for GLB and 6.25–37.5 µg/ml for ATN. The LOD and LOQ of GLB was found to be 0.48 µg/ml and 1.47µg/ml and for ATN was found to be 0.72µg/ml and 2.20 µg/ml, respectively. The method was applied to drug interaction studies of GLB with ATN to illustrate the scope and application of the methods to manage two different therapeutic classes of drugs, as they may co-administered in concurrent diseases.

Purpose: To develop a stability indicating RP-HPLC method for a combination drug product containing a high dose of paracetamol (PR) and low doses of domperidone (DM) and tramadol HCL (TR).Methods: The analytes are well separated by a reverse phase column and an isocratic mobile phase consisting of 0.1 %v/v trifluoroacetic acid: acetonitrile: methanol in the ratio 70:25:5 (v/v) with a flow rate of 1.0 mL/min. The effluent was monitored at 272 nm. The drug products were subjected to stress conditions of acid, base, peroxide, thermal and photolytic degradation and peak homogeneity of PR, TR and DM were obtained using photo diode array detector.Results: The degradation products were well resolved from PR, TR and DM peaks, thus indicating the stability-indicating nature of the method. The assay was linear from 165 – 495 μg/mL for PR, 18.75 – 56.25 μg/mL for TR, and 5 – 15 μg/mL for DM. Although the tablet contained high and low doses of the drugs, the intra- and inter-day variations were < 2.0 %.Conclusion: The proposed method was validated according to the ICH guidelines and proved suitable for stability and homogeneity testing, as well as for quality control of the combined drugs in pharmaceutical preparations.Keywords: HPLC, Isocratic, Peak purity, Simultaneous determination, Paracetamol, Tramadol, Domperidone

Impurity profiling of active pharmaceutical ingredients is a crucial step in assessing their quality. Moreover, the chemical nature of nimodipine makes it susceptible easily to acidic and alkaline hydrolysis. On the other hand, nimodipine is co-formulated with citicoline sodium pharmaceutically as tablets to treat cerebral ischemia. In this study, three degradation products of nimodipine were synthesized, isolated and characterized with aid of FTIR spectroscopy, 1H-NMR as well as LC-MS/MS after exposing to drastic acidic and alkaline conditions. Subsequently, a simple, selective and valid RP-HPLC method was developed for simultaneous estimation of nimodipine and citicoline in the presence of nimodipine acid and alkaline degradation products in bulk and tablets. Chromatographic separation was achieved using an isocratic mobile phase consisting of acetonitrile: 0.02 M KH2PO4 (containing 0.2% v/v, triethylamine and adjusted to pH 3.0 with orthophosphoric acid) (70:30, v/v) at a flow rate 1.0 mL min-1 at ambient temperature (25 ºC) on a Eurospher II C18 (250 mm × 4.6 mm, 5 µm) column with UV detection at 270 nm for citicoline and 235 nm for nimodipine and its acidic and alkaline hydrolytic products. Linearity, accuracy and precision were found to be acceptable over a concentration range of (4.5–120 μg mL-1) for nimodipine and (15–400 μg mL-1) for citicoline. The proposed method could be successfully applied for the routine analysis of the studied drugs in their pharmaceutical preparation in the presence of nimodipine common degradation products without any preliminary separation step.

Purpose: To develop and validate RP-HPLC method for simultaneous determination of four active ingredients: picroside I (PSI), picroside II (PSII), phyllanthin (PHY) and boeravinone-B (BVB) in a polyherbal hepatoprotective tablet formulation.Methods: The study was carried out using Waters X-Bridge, C18 (250 mm x 4.6 mm, 5 μm) column with mobile phase consisting of 5 mM ammonium acetate in 10 % methanol and acetonitrile, with gradient programme at dual wavelengths of 220 nm and 274 nm and flow speed of 1 mL.min-1. The procedure was validated with respect to specificity, linearity, precision, accuracy, system suitability, limit of detection (LOD) and of quantification (LOQ), and robustness in line with ICH specifications.Results: The method was linear within the concentration range of 25 to 200 %, and the values of correlation coefficients (R2) were > 0.999. Intra-day and inter-day RSDs of PAs and RTs were < 5.0 %, with recovery in the range of 100.0 - 106.0 %.Conclusion: The four active ingredients have with good resolution with regard to the method used. The method is rapid, simple, highly selective, sensitive and cost effective, which make it an efficient method for quality assurance.

A simple and robust high-performance liquid chromatography (HPLC) method is described for the assay for levetiracetam (LTC), methyl paraben (MHB), and propyl paraben (PHB) either in their pure form or in commercial Levepsy® syrup. The method is selective and stability indicating and all chromatographic conditions were studied to obtain adequate separation of LTC, MHB, and PHB from their degradation products and from excipients. The HPLC separation was carried out on a RP C18 Hypersil BDS analytical column (150 mm × 4.6 mm ID) using gradient elution system. The mobile phase flow rate was 1.5 mLmin−1 and the column temperature was kept at 40°C. Complete separation of the studied components was obtained within a cycle time of 8 min. LTC, MHB, and PHB were eluted at 1.56, 5.86, and 7.85 min, respectively. Detection was carried out at 240 nm using a dual wavelength detector. The method has been validated for linearity, accuracy, precision, specificity, limit of detection, limit of quantitation, robustness, and ruggedness. The proposed method was successfully applied for the determination of LTC in the presence of parabens in Levepsy syrup.

Background: Curcumin and quercetin are the two important flavonoids that are being used as food ingredients across the globe. They also possess many pharmacological activities such as anti...

A Rapid Resolution Reversed Phase High-Performance Liquid Chromatography (RR RP-HPLC) method has been developed and validated for the simultaneous determination of atorvastatin and seven related compounds specified as impurities. Experimental design was used during method optimization (full factorial 32 design) and robustness testing (central composite design). Chromatography was performed with mobile phase containing phosphate buffer pH 3.5 and a mixture of 10% (v/v) tetrahydrofuran in acetonitrile as organic modifier. A Zorbax Eclipse XDB C18 Rapid Resolution HT 4.6 mm × 50 mm, 1.8 µm particle size column was used. The developed method allowed determination of Atorvastatin Calcium (ATV Ca) purity and level of impurities in drug substances.

Development and Validation of UV Spectrophotometry and RP-HPLC Method for simultaneous determination of Rosuvastin and Clopidogrel in Tablet Dosage Form

Development and validation of RP-HPLC method for simultaneous determination of diclofenac sodium and tizanidine hydrochloride in bulk and tablet formulation

Development and validation of RP-HPLC method for simultaneous determination of ondansetron hydrochloride and granisetron hydrochloride in their admixtures with pantoprazole sodium

A simple and precise stability indicating method for the simultaneous estimation of dapagliflozin and saxagliptin in combined tablet dosage form was developed and validated using RP-HPLC.