Mohammed Abdelkawy

Development and validation of stability indicating HPLC and HPTLC methods for determination of sulpiride and mebeverine hydrochloride in combination

Validated sensitive and highly selective stability indicating methods are adopted for simultaneous quantitative determination of sulpiride and mebeverine hydrochloride in presence of their reported impurities and hydrolytic degradates whether in pure forms or in pharmaceutical formulation. The first method is High Performance Liquid Chromatography, where the mixture of sulpiride and mebeverine hydrochloride together with the reported interferents plus metopimazine as internal standard are separated on a reversed phase cyano column (5 microm ps, 250 mm x 4.6 id) using acetonitrile: water (70:30 v/v) adjusted to pH = 7 as a mobile phase. The drugs were detected at 221 nm over a concentration range of 5-40 microg ml(-1) and 5-60 microg ml(-1) with mean percentage recoveries 99.75% (S.D. 0.910) and 99.99% (S.D. 0.450) for sulpiride and mebeverine hydrochloride respectively. The second method is High Performance Thin Layer Chromatography, where sulpiride and mebeverine hydrochloride are separated on silica gel HPTLC F(254) plates using absolute ethanol:methylene chloride:triethyl amine (7:3:0.2 by volume) as mobile phase and scanning of the separated bands at 221 nm over a concentration range of 0.4-1.4 and 0.2-1.6 microg band(-1) with mean percentage recoveries 101.01% (S.D. 1.991) and 100.40% (S.D. 1.868) for sulpiride and mebeverine hydrochloride respectively.

Quantitative determination of oxybutynin hydrochloride by spectrophotometry, chemometry and HPTLC in presence of its degradation product and additives in different pharmaceutical dosage forms

Simple, accurate, sensitive and validated UV spectrophotometric, chemometric and HPTLC-densitometric methods were developed for determination of oxybutynin hydrochloride (OX) in presence of its degradation product and additives in its pharmaceutical formulations. Method A is the first derivative of ratio spectra (DD(1)) which allows the determination of OX in presence of its degradate in pure form and tablets by measuring the peaks amplitude at 216 nm. Method B and C are principal component regression (PCR) and partial least-squares (PLS) for determination of OX in presence of its degradate in pure form, tablets and syrup. While, the developed high performance thin layer chromatography HPTLC-densitometric method was based on the separation of OX from its degradation product, methylparaben and propylparaben followed by densitometric measurement at 220 nm which allows the determination of OX in pure form, tablets and syrup. The separation was achieved using HPTLC silica gel F(254) plates and chloroform:methanol:ammonia solution:triethylamine (100:3:0.5:0.2, v/v/v/v) as the developing system. The accuracy, precision and linearity ranges of the developed methods were determined. The results obtained were statistically compared with each other and to that of a reported HPLC method, and there was no significant difference between the proposed methods and the reported method regarding both accuracy and precision.

Development and validation of simultaneous spectrophotometric and TLC-spectrodensitometric methods for determination of beclomethasone dipropionate and salbutamol in combined dosage form

Spectrophotometric and TLC-spectrodensitometric methods were developed and validated for the simultaneous determination of beclomethasone dipropionate (BEC) and salbutamol (SAL). The spectrophotometric methods include dual wavelength, ratio difference, constant center coupled with a novel method namely, spectrum subtraction and mean centering with mean percentage recoveries and RSD 99.72±1.07 and 99.70±1.12, 100.25±1.12 and 99.89±1.12, 99.66±1.85 and 99.19±1.32, 100.74±1.26 and 101.06±0.90 for BEC and SAL respectively. The TLC-spectrodensitometric method was based on separation of both drugs on TLC aluminum plates of silica gel 60 F254, using benzene: methanol: triethylamine (10:1.5:0.5 v/v/v) as a mobile phase, followed by densitometric measurements of their bands at 230 nm. The mean percentage recoveries and RSD were 99.07±1.25 and 101.35±1.50 for BEC and SAL respectively. The proposed methods were validated according to ICH guidelines and were applied for the simultaneous analysis of the cited drugs in synthetic mixtures and pharmaceutical preparation. The methods were found to be rapid, specific, precise and accurate and can be successfully applied for the routine analysis of BEC and SAL in their pharmaceutical formulation with no need for prior separation. The results obtained were statistically compared to each other and to that of the reported HPLC method. The statistical comparison showed that there is no significant difference regarding both accuracy and precision.

Spectrophotometric and spectrodensitometric determination of triamterene and xipamide in pure form and in pharmaceutical formulation.

Sensitive and validated UV-spectrophotometric, chemometric and TLC-densitometric methods were developed for determination of triamterene (TRM) and xipamide (XIP) in their binary mixture, formulated for use as a diuretic, without previous separation. Method A is the isoabsorptive point spectrophotometry, in which TRM concentration alone can be determined at its λ(max) while XIP concentration can be determined by measuring total concentration of TRM and XIP at their isoabsorptive point followed by subtraction. Method B is the ratio subtraction spectrophotometry, where XIP can be determined by dividing the spectrum of the mixture by the spectrum of TRM (as a divisor) followed by subtracting the constant absorbance value of the plateau region, then finally multiplying the produced spectrum by the spectrum of the divisor, while TRM concentration can be determined at its λ(max). Method C is a chemometric-assisted spectrophotometry where classical least squares, principal component regression, and partial least squares were applied. Method D is a TLC-densitometry; this method depends on quantitative densitometric separation of thin layer chromatogram of TRM and XIP using silica gel plates at 254 nm. The proposed methods were successfully applied for the analysis of TRM and XIP in their pharmaceutical formulation and the results were statistically compared with the established HPLC method.

Simultaneous determination of methocarbamol and its related substance (Guaifenesin) in two ternary mixtures with ibuprofen and diclofenac potassium by HPTLC spectrodensitometric method

Accurate, sensitive, and precise high-performance thin-layer chromatographic (HPTLC) method was developed and validated for the determination of Methocarbamol (ME) and its related substance (guaifenesin (GU)) in two ternary mixtures with ibuprofen (IB) and diclofenac potassium. The method depends on separation and quantification of the studied drugs on TLC silica gel 60 F254 plates using ethyl acetate-acetone-triethylamine-formic acid (62:35:6:0.3, by volume) as the developing system followed by densitometric measurement of the bands at 222 nm for the first mixture containing methocarbmol, IB, and GU and at 278 nm for the second mixture containing methocarbmol, diclofenac potassium, and GU. The proposed methods were successfully applied for the determination of ME, IB, and diclofenac potassium in the presence of ME-related substance (GU) either in bulk powder or in their pharmaceutical formulations.

Application of Successive and Progressive Spectrophotometric Resolution for the Analysis of Partially or Completely Overlapping Ternary Mixtures

Abstract In this work application of a recently developed progressive resolution technique and the well-established successive resolution techniques either separately or in combination for the analysis of ternary mixtures without prior separation steps. The resolution steps verified according to the degree of spectra overlapping either partially or completely. All the proposed spectrophotometric techniques consist of several consecutive steps utilizing ratio and/or derivative spectra. The proposed methods can be used for the determination of ternary mixture of pseudoephedrine sulphate (PSE), loratadine (LOR) and paracetamol (PAR). Pseudoephedrine (PSE) could be determined using successive one and applying zero crossing derivative ratio (DD1) at 218.6 nm and the derivative of the resolved spectrum of PSE and PAR at 217 nm and 235 nm via amplitude factor. Loratadine (LOR) could be analyzed either by constant center or dual wavelength (DWL) as well as progressively using their derivative spectrum by applying amplitude factor method at 261 nm and 305 nm without any contribution of PSE. Paracetamol (PAR) could be measured at its maxima at 248 nm either directly after resolution of LOR or via constant multiplication method. Moreover it could be analyzed using the first derivative of the mixture at 305 nm without any interference of others. Regression analysis showed good correlation in the concentration ranges 4-40 μg/mL, 3.5-35 μg/mL and 1.5-17 μg/ml for PSE, LOR and PAR, respectively. The specificity of the developed methods was investigated by analyzing laboratory prepared mixtures and was successfully applied for the analysis of pharmaceutical formulations containing the cited drugs with no interference from additives. The proposed methods were validated according to the ICH guidelines. The obtained results were statistically compared with those of the official or reported methods of the cited drugs; using student t-test, F-test, and one way ANOVA, showing no significant difference with respect to accuracy and precision.

Two Validated Liquid Chromatographic Methods for the Simultaneous Determination of Flumethasone Pivalate, Its Related Substance (Flumethasone), and Clioquinol

Two liquid chromatographic methods were developed and validated. Simple and sensitive thin-layer chromatography (TLC)—densitometric and high-performance liquid chromatographic (HPLC) methods were used for the simultaneous determination of flumethasone pivalate (FP), flumethasone pivalate related substance and impurity, flumethasone (FL), and clioquinol (CL). The proposed TLC — densitometric method has been developed using silica gel plates 60 F254 as a stationary phase with benzene—hexane—acetone—formic acid (5:4:2:0.13, by volume) as a developing system followed by densitometric measurements at 235 nm. The studied components were quantified in the range of 0.3–4, 0.3–3, and 1.5–5 μg band−1, respectively. For HPLC method, chromatographic separation was achieved within 11 min with the required peak symmetry, accuracy, and precision on ODS column using acetonitrile—water (70:30, v/v) as the mobile phase at a flow rate of 1 mL min−1 with ultraviolet (UV) detection at 235 nm. The calibration plots were linear over the concentration range of 5–50, 2–35, and 10–70 mg mL−1, respectively. The proposed methods were validated as per International Conference on Harmonization (ICH) guidelines; accuracy, precision, and repeatability were found to be within the acceptable limits.

LC-MS as a Stability-Indicating Method for Analysis of Hyoscine N-Butyl Bromide under Stress Degradation Conditions with Identification of Degradation Products

Hyoscine N-Butyl Bromide (HBB) was subjected to different ICH prescribed stress conditions. It showed extensive decomposition under base hydrolytic conditions, while it was less liable to stress acid hydrolytic conditions. It showed also moderate degradation in response to oxidation stress of hydrogen peroxide. The drug showed no changes under photolysis conditions. In total, a number of major degradation products were detected by HPLC and identified by LC-MS. For establishment of stability-indicating assay, the reaction solutions in which different degradation products were formed were prepared, and the separation was optimized by varying the HPLC conditions. An acceptable chromatograms was achieved using a C18 column using (water: methanol 50: 50 v/v, pH adjusted to 3.9 with triflouroacetic acid) as a mobile phase with flow rate of 1.0 ml min−1 and UV detection wavelength at 210 nm. The percent of degradation was calculated in each run by measuring the intensity of the peak area of the intact drug at 6.2 min. Complete degradation only occur in case of 5 N NaOH indicates that the drug is very sensitive to alkaline hydrolysis. The LC-MS study was carried out to identify the major degradation products using a sunfire (waters) C-18 column and a mobile phase comprising of acetonitrile: 0.1M ammonium acetate (80:20, v/v) with flow rate of 1.0 ml min−1. MS measurements were acquired in positive ion full scan modes from 50 to 400 amu. The m/z values of the main peaks were investigated with the expected chemical structure of degradates.