Effective spectrophotometric methods for resolving the superimposed spectra of Diclofenac Potassium and Methocarbamol.

The UV spectra of Diclofenac Potassium DIC and Methocarbamol MET are superimposed making their analysis using direct or derivative spectrophotometric methods quite complicated. This study presents four effective spectrophotometric methods that enable simultaneous determination of both drugs without interference. The first method is based on application of simultaneous equation method on their zero order spectra where DIC has shown absorbance maxima at 276 nm and MET displays two absorbances maxima at 273 nm and 222 nm in distilled water. The second method relies on dual wavelength method, the two wavelengths (232 and 285 nm) were chosen for determination of DIC where the absorbance differences at these wavelengths are proportional to DIC concentration while the absorbance differences of MET are equal to zero. For the determination of MET, the two wavelengths (212 and 228 nm) were selected. The third method of first-derivative ratio has been applied where the derivative ratio absorbances of DIC and MET were measured at 286.1 and 282.4 nm, respectively. The fourth method utilizing ratio difference spectrophotometric (RD) method was eventually performed on the binary mixture. The amplitude difference between the two wavelengths (291and 305 nm) was calculated for DIC estimation while the amplitude difference between the two wavelengths (227and 273 nm) for MET determination. All methods show linearity range from 2.0-25 µg. mL-1 and 6.0-40 µg. mL-1 for DIC and MET respectively. The developed methods have been statistically compared with a reported method based on first derivative method and the results of statistical comparison confirm the accuracy and precision of the proposed methods therefore they can be effectively applied for determination of MET and DIC in pharmaceutical dosage form.

Development and Validation of a Stability-Indicating High-Performance Liquid Chromatographic Method for the Quantification of Methocarbamol and Its Impurities in Pharmaceutical Dosage Forms.

A novel, delicate, stability-indicating, gradient, reversed-phase high-performance liquid chromatographic method has been established for the quantitative estimation of methocarbamol (MTC) and its impurities present in a pharmaceutical oral suspension. XBridge C18, 5 µm, 250 mm × 4.6 mm column was used to accomplish chromatographic separation with a buffered mobile phase consisting of a mixture of 0.01 M of sodium dihydrogen phosphate (pH 7.0 buffer) and methanol in the ratio of 95:05 (v/v), respectively, were used as solvent A and a mixture of methanol and Milli-Q water in the ratio 90:10 (v/v), respectively, was used as solvent B. Analysis was carried out at 0.8 mL/min flow rate and the detection wavelength at 225 nm. The compartment temperature of the column is put at 25°C. The resolution of MTC and its four impurities has been attained >2.0 for all pairs of compounds. Significant degradation of MTC was photolytic, thermal and oxidative stress conditions. Validation of the developed method was performed as stated by the International Conference on Harmonization guidelines with regard to all validation parameters like specificity, accuracy, linearity, precision, limit of detection, limit of quantitation and robustness.

A New Application of PARAFAC Model to UPLC Dataset for the Quantitative Resolution of a Tri-Component Drug Mixture.

In the presented work, a three-way analysis of ultra-performance liquid chromatography-photodiode array (UPLC-PDA) dataset was performed by parallel factor analysis (PARAFAC) for quantitatively resolving a ternary mixture containing paracetamol and methocarbamol with indapamide selected as an internal standard in their co-eluted chromatographic conditions. Paracetamol and methocarbamol were quantified in the working range between 3-24 and 5-50 µg/mL by applying PARAFAC decomposition to UPLC-PDA data array obtained under unresolved chromatographic peak conditions. To compare the experimental results provided by co-eluted UPLC-PARAFAC method, an ordinary UPLC method was developed ensuring proper separation of the peaks. The performance of both PARAFAC and ordinary UPLC methods were assessed by quantifying independent test samples, intra- and inter-day samples and spiked samples of pharmaceutical preparations. Then, both methods were applied for quantitative estimation of the related drugs in a commercial pharmaceutical preparation. In this study, PARAFAC method was proved to be a very powerful alternative for the quality control of pharmaceutical preparations containing paracetamol and methocarbamol even in their co-eluted chromatograms with high precision and accuracy in a short chromatographic runtime of 1.2 min.

Simultaneous determination of methocarbamol and aspirin in presence of their pharmacopeial-related substances in combined tablets using novel HPLC-DAD method.

Objective and Significance: Methocarbamol (MET) and aspirin (ASP) are widely used as a muscle relaxant combination. The USP reports guaifenesin (GUA) and salicylic acid (SAL) as related substances and hydrolytic products of MET and ASP, respectively. This work aimed at developing and validating a simple and sensitive RP-HPLC method for the determination of both drugs as well as their related substances (at their pharmacopeial limits) in their bulk powders, laboratory prepared mixtures, and MET-ASP combined tablets. Methods and Results: Chromatographic separation was achieved in less than 9 min with the required resolution, peak symmetry, and accuracy on C18 column using isocratic elution system of diluted acetic acid (pH 3.2): acetonitrile at the ratio of 79: 21, v/v, at a flow rate of 1 mL/min. Detection was achieved with photodiode array at 233 nm for MET, GUA, and SAL and at 273 nm for ASP. The developed method has been validated as per ICH guidelines and the calibration plots were linear over the concentration ranges of 2-150, 0.4-30, 25-450, and 0.2-27 µg/mL for MET, GUA, ASP, and SAL, respectively. Conclusion: The optimized method proved to be specific, robust and precise for the quality control of the studied drugs in pharmaceutical preparations to ascertain that their related substances are not exceeding the permitted pharmacopeial limits.

Smart manipulation of ratio spectra for resolving a pharmaceutical mixture of Methocarbamol and Paracetamol.

Two smart, specific, accurate and precise spectrophotometric methods manipulating ratio spectra are developed for simultaneous determination of Methocarbamol (METH) and Paracetamol (PAR) in their combined pharmaceutical formulation without preliminary separation. Method A, is an extended ratio subtraction one (EXRSM) coupled with ratio subtraction method (RSM), which depends on subtraction of the plateau values from the ratio spectrum. Method B is a ratio difference spectrophotometric one (RDM) which measures the difference in amplitudes of ratio spectra between 278 and 286 nm for METH and 247 and 260 nm for PAR. The calibration curves are linear over the concentration range of 10-100 µg mL(-1) and 2-20 µg mL(-1) for METH and PAR, respectively. The specificity of the developed methods was investigated by analyzing different laboratory prepared mixtures of the two drugs. Both methods were applied successfully for the determination of the selected drugs in their combined dosage form. Furthermore, validation was performed according to ICH guidelines; accuracy, precision and repeatability are found to be within the acceptable limits. Statistical studies showed that both methods can be competitively applied in quality control laboratories.

Simultaneous determination of methocarbamol and aspirin by RP-HPLC using fluorescence detection with time programming: its application to pharmaceutical dosage form.

A new simple, rapid and sensitive reversed-phase liquid chromatographic method was developed and validated for the simultaneous determination of methocarbamol (MET) and aspirin (ASP) in their combined dosage form. The separation of these compounds was achieved within 6.0 min on a CLC Shim-pack C8 column (250 × 4.6 mm, 5 µm particle size) using isocratic mobile phase consisting of acetonitrile and 0.02 M dihydrogenphosphate buffer (30:70, v/v) at pH = 5.0. The analysis was performed at a flow rate of 1.0 mL/min with fluorescence detection at 277/313 nm for MET and 298/410 nm for ASP using real-time programming. The selectivity, linearity of calibration, accuracy, inter- and intra-day precision and recovery were examined as parts of the method validation. The concentration-response relationship was linear over concentration ranges of 0.02-0.20 and 0.02-0.40 µg/mL for MET and ASP, respectively, with a limit of detection of 6 and 32 ng/mL for MET and ASP, respectively. The proposed method was successfully applied for the analysis of both MET and ASP in prepared tablets with average recoveries of 99.88 ± 0.65% for MET and 100.44 ± 0.78% for ASP. The results were favourably compared to those obtained by a reference method.

Simultaneous determination of methocarbamol and ibuprofen or diclofenac potassium using mean centering of the ratio spectra method.

Accurate and sensitive methods were developed for simultaneous determination of methocarbamol and ibuprofen or diclofenac potassium in their binary mixtures, and in the presence of a methocarbamol related substance (guaifenesin) using mean centering of the ratio spectra method. The results obtained were statistically compared with the reported HPLC methods; no significant difference between the proposed methods and the reported methods was found regarding either accuracy or precision.

Simultaneous spectrophotometric determination of diclofenac potassium and methocarbamol in binary mixture using chemometric techniques and artificial neural networks.

In this study, the simultaneous determination of diclofenac potassium (DP) and methocarbamol (MT) by chemometric approaches and artificial neural networks using UV spectrophotometry has been reported as a simple alternative to using separate models for each component. Three chemometric techniques-classical least-squares (CLS), principal component regression (PCR), and partial least-squares (PLS)-along with radial basis function-artificial neural network (RBF-ANN) were prepared by using the synthetic mixtures containing the two drugs in methanol. A set of synthetic mixtures of DP and MT was evaluated and the results obtained by the application of these methods were discussed and compared. In CLS, PCR, and PLS, the absorbance data matrix corresponding to the concentration data matrix was obtained by the measurements of absorbances in the range 260-310 nm in the intervals with Δλ = 0.2 nm in their zero-order spectra. Then, calibration or regression was obtained by using the absorbance data matrix and concentration data matrix for the prediction of the unknown concentrations of DP and MT in their mixtures. In RBF-ANN, the input layer consisting of 251 neurons, 9 neurons in the hidden layer, and 2 output neurons were found appropriate for the simultaneous determination of DP and MT. The accuracy and the precision of the four methods have been determined and they have been validated by analyzing synthetic mixtures containing the two drugs. The proposed methods were successfully applied to a pharmaceutical formulation containing the examined drugs.

Spectrofluorometric determination of methocarbamol in pharmaceutical preparations and human plasma.

A simple, sensitive and rapid spectrofluorometric method for determination of methocarbamol in pharmaceutical formulations and spiked human plasma has been developed. The proposed method is based on the measurement of the native fluorescence of methocarbamol in methanol at 313 nm after excitation at 277 nm. The relative fluorescence intensity-concentration plot was rectilinear over the range of 0.05-2.0 µg/mL, with good correlation (r=0.9999), limit of detection of 0.007 µg/ mL and a lower limit of quantification of 0.022 µg/ mL. The described method was successfully applied for the determination of methocarbamol in its tablets without interference from co-formulated drugs, such as aspirin, diclofenac, paracetamol and ibuprofen, The results obtained were in good agreement with those obtained using the official method (USP 30).The high sensitivity of the method allowed the determination of the studied drug in spiked human plasma with average percentage recovery of 99.42 ± 3.84.

Simultaneous determination of diclofenac potassium and methocarbamol in ternary mixture with guaifenesin by reversed phase liquid chromatography.

New, simple, rapid and precise reversed phase liquid chromatographic (RP-LC) method has been developed for the simultaneous determination of diclofenac potassium (DP) and methocarbamol (MT) in ternary mixture with guaifenesin (GF), degradation product of methocarbamol. Chromatographic separation was achieved on a Symmetry Waters C18 column (150 mm x 4. 6mm, 5 microm). Gradient elution based on phosphate buffer pH (8)-acetonitrile at a flow rate of 1 mL min(-1) was applied. The UV detector was operated at 282 nm for DP and 274 nm for MT and GF. Linearity, accuracy and precision were found to be acceptable over the concentration ranges of 0.05-16, 0.5-160 and 0.5-160 microg mL(-1) for DP, MT and GF, respectively. The optimized method proved to be specific, robust and accurate for the quality control of the cited drugs in pharmaceutical preparation.

Electrochemical determination of methocarbamol on a montmorillonite-Ca modified carbon paste electrode in formulation and human blood.

Utilizing the fascinating strong adsorptive ability, high chemical and mechanical stability properties of montmorillonite-calcium (MMT-Ca) clay, a MMT-Ca modified carbon paste electrode was developed for the sensitive determination of methocarbamol. Methocarbamol has been oxidized in buffered solutions at the developed MMT-Ca-modified CPE in a single 2-electron irreversible anodic peak. A simple and sensitive square-wave adsorptive anodic stripping voltammetric method was optimized for determination of methocarbamol in bulk form, pharmaceutical formulation and in spiked human serum using the developed modified CPE. This was carried out without the necessity for samples pretreatment and/or time-consuming liquid-liquid or solid-phase extraction steps prior to the analysis. The developed electrode exhibited an excellent sensitivity and selectivity towards methocarbamol even in the presence of 10(2)-10(4)-fold of its active ingredient "ibuprofen," common excipients, common metal ions or co-administrated drugs. Limits of detection of 3 x 10 (-9) and 1.2 x 10(-8) M and limits of quantitation of 1 x 10(-8), and 4 x 10(-8) M methocarbamol were achieved in the bulk form and in spiked human serum, respectively. Moreover, the developed method was successfully applied for determination of methocarbamol in human plasma of subjects following administration of an oral dose of Ibuflex tablets.

A stability-indicating high-performance liquid chromatographic method for the determination of methocarbamol in veterinary preparations.

An isocratic HPLC method was developed and validated for the quantitation of methocarbamol in the presence of its degradation products. Quantitation was achieved using a reversed-phase C18 column at ambient temperature with mobile phase consisting of methanol-water-tetrahydrofuran (25 + 65 + 10, v/v). The flow rate was 0.9 mL/min. The detection was by UV light at 274 nm. The proposed method was validated for selectivity, precision, linearity, and accuracy. The assay method was found to be linear from 159.0 to 793.2 microg/mL (3.2 to 15.9 microg injected). All validation parameters were within the acceptable range. The developed method was successfully applied to estimate the amount of methocarbamol in a veterinary injection.

Simultaneous determination of paracetamol and methocarbamol in tablets by ratio spectra derivative spectrophotometry and LC.

The application of the ratio spectra derivative spectrophotometry and high-performance liquid chromatography (HPLC) to the simultaneous determination of paracetamol (PAR) and methocarbamol (MET) in combined pharmaceutical tablets is presented. The spectrophotometric procedure is based on the use of the first derivative of the ratio spectra obtained by dividing the absorbtion spectrum of the binary mixtures by a standard spectrum of one of the compounds. The first derivative amplitudes were measured at 243.0 and 230.3 nm for the assay of PAR and MET, respectively. Calibration graphs were established for 2-30 microg ml for PAR and 2-36 microg/ml for MET in binary mixture. The detection limits for PAR and MET were found 0.097 and 0.079 microg/ml, respectively; while the quantification limits were 0.573 microg/ml for PAR and 1.717 microg/ml for MET. For the HPLC procedure, a reversed-phase column with a mobile phase of methanol-water (60:40, v/v), was used to separate both compounds with a detection of 274.0 nm. Linearity was obtained in the concentration range of 2 300 and 1.5-375 microg/ml for PAR and MET, respectively. The detection and quantification limits were found to be 0.42 and 1.4 microg/ml for PAR and 0.36 and 1.2 microg/ml for MET, respectively. The relative standard deviations were found to be less than 0.52%, indicating reasonable repeatibility of both methods. The proposed methods were successfully applied to the determination of these drugs in commercial tablets.

A new HPLC technique for the separation of methocarbamol enantiomers.

We have developed a stereoselective high-performance liquid chromatography technique for analytical separation of methocarbamol enantiomers. Precolumn derivatization was performed at room temperature using (-)-menthylchloroformate as a chiral reagent in the presence of pyridine as catalyst. The resulting diastereomers were separated on two Resolve C18 columns connected in series. The mobile phase was phosphate buffer (pH 7.5)-acetonitrile (50: 50, v/v) at a flow rate of 1 mL min(-1). UV detection was set at 274 nm. The optimum amount of reagent and the maximum peak intensity of the diastereomers were determined. The resolution of the diastereomers was satisfactory (alpha = 1.04) under the conditions used.

A fatal interaction of methocarbamol and ethanol in an accidental poisoning.

A case is presented of a fatal drug interaction caused by ingestion of methocarbamol (Robaxin) and ethanol. Methocarbamol is a carbamate derivative used as a muscle relaxant with sedative effects. Therapeutic concentrations of methocarbamol are reported to be 24 to 41 micrograms/mL. Biological fluids were screened for ethanol using the Abbott TDx system and quantitated by gas-liquid chromatography (GLC). Determination of methocarbamol concentrations in biological tissue homogenates and fluids were obtained by colorimetric analysis of diazotized methocarbamol. Blood ethanol concentration was 135 mg/dL (0.135% w/v) and urine ethanol was 249 mg/dL (0.249% w/v). Methocarbamol concentrations were: blood, 257 micrograms/mL; bile, 927 micrograms/L; urine, 255 micrograms/L; gastric, 3.7 g; liver, 459 micrograms/g; and kidney, 83 micrograms/g. The combination of ethanol and carbamates is contraindicated since acute alcohol intoxication combined with carbamate usage can lead to combined central nervous system depression as a result of the interactive sedative-hypnotic properties of the compounds.

Liquid chromatographic determination of methocarbamol in injection and tablet dosage forms: collaborative study.

A reverse phase liquid chromatographic method was developed for determining methocarbamol in injection and tablet dosage forms. The injections require dilution only; the tablets require a filtration step before introduction into the chromatograph. Response for methocarbamol was linear over the range 0-18 micrograms, using an ultraviolet detector at 274 nm. Recoveries by the author ranged from 96.1 to 101.9% for authentic injection formulations and 98.0 to 101.0% for authentic tablet formulations. A collaborative study of the method by 6 laboratories resulted in standard deviations of 1.70 and 2.22 for injection and tablet dosage forms, respectively. The method has been adopted official first action.

Liquid chromatography in pharmaceutical analysis XI: determination of muscle relaxant--analgesic mixture using reversed-phase and ion-pair techniques.

High pressure liquid chromatography using reversed-phase and/or ion-pair techniques was used to optimize resolution of aspirin-containing muscle relaxant mixtures as well as other therapeutic agents commonly found in muscle relaxant-analgesic mixtures. The compounds were chromatographed on an octadecylsilane column using methanol--water solvent systems, some of which contained tetrabutylammonium cation as counterion. Mixtures of methocarbamol--aspirin and chlorzoxazone--acetaminophen were selected to demonstrate separation and quantification. The methocarbamol--aspirin mixture was chromatographed with methanol--water (40:60, pH 6.8) containing 0.01 M tetrabutylammonium cation at a flow rate of 2.0 ml/min. The chlorzoxazone--acetaminophen mixture was chromatographed with methanol--water (50:50) at a 2.0 ml/min flow rate. The separation and quantitation of each mixture were achieved in approximately 8 min with accuracy in the 2--3% range.