Reverse-Phase High Performance Liquid Chromatographic Determination of Clidinium Bromide and Chlordiazepoxide in Tablet Formulations

Abstract

Clidinium bromide is a quaternary ammonium anticholinergic agent with peripheral effects. It is commercially available in combination with chlordiazepoxide. There is no official compendial method available for the analysis of both compounds concurrently. Conventional methods for determining both compounds involve extensive extraction and thus are time-consuming, and lack precision. The analytical scheme described in this paper provides a fast and reliable reverse-phase HPLC for chlordiazepoxide and clidinium bromide in pharmaceutical combinations. The mobile phase was 0.04 M ammonium acetate in 70% acetonitrile solution with 1% dimethylformamide (pH 6). The column was microPak MCH-10 (300.0 mm × 4.0 mm), UV detection at 254 nm and benzophenone was used as internal standard. The method was confirmed for linearity, recovery, specificity, accuracy, and applicability.     

Determination of Propantheline Bromide in Tablet Formulations by Reverse-Phase HPLC

Abstract

The reported reverse-phase HPLC method for the determination of propantheline bromide, xanthanoic acid, xanthone, and 9-hydroxy-propantheline bromide in tablets is fast, sensitive, specific, accurate and reporoducible. Methyl xanthanoate is used as internal standard. The total elution time is 6 min. The method is stability-indicating since it can determine the degradation products. The column utilized was suplecosil LC-8 (5 micron), 250 mm × 4.6 mm i.d. The mobile phase was 0.03 M solution of ammonium acetate in acetonitrile: water: THF (60:38:2); the pH was adjusted to 4.5 with acetic acid, the detection was at 254 nm. A wavelength of 248 nm was used to quantitate xanthanoic acid and the flow rate was 1.5 mL/min. The proposed HPLC method was verified for linearity, accuracy, precision, and applicability.     

Thin layer Chromatographic (TLC) Determination of Phenytoin in Pharmaceutical Formulations and Identification of Its Hydroxylated Urinary Metabolites

Abstract

A simple quantitative TLC method is described for the determination of phenytoin (PHT) in pharmaceutical formulations (capsules and injectables) using a scanning densitometer. The average percent recoveries for PHT capsules and injectables were found to be 98.9 ± 0.46 and 100.52 0.25 respectively. The method proposed is rapid, stability-indicating and can be adopted for routine analysis in quality control laboratories. The method was also developed for the identification of PHT and its hydroxylated metabolites namely p-hydroxy phenytoin (p-HPPH) conjugated and unconjugated and the phenytoin dihydrodiol (DHD) in the urine of epileptic adult patients.     

A rapid and highly sensitive stability-indicating high-performance thin-layer chromatography technique for the determination of tedizolid phosphate with a classical univariate calibration

No high-performance thin-layer chromatography (HPTLC) techniques have been established for the determination of tedizolid phosphate (TDZP) in pharmaceutical products or physiological fluids. Therefore, a rapid and highly sensitive stability-indicating HPTLC technique has been developed for the determination of TDZP in commercial formulations with a classical univariate calibration. The HPTLC‒densitometry analysis of TDZP was carried out via chloroform‒methanol (90:10, V/V) mobile phase. The determination of TDZP was performed at the wavelength of 300 nm. The proposed HPTLC technique was linear in the range of 10‒2000 ng band^‒1. In addition, the method was found to be highly accurate (% recovery = 98.53‒101.74%), precise (%CV = 0.67‒0.91%), robust (%CV = 0.83‒0.86%), highly sensitive (LOD = 3.41 ng band^‒1, LOQ = 10.23 ng band^‒1) for the determination of TDZP. The proposed technique was also able to detect TDZP in the presence of its degradation products under various stress conditions and it can be considered as a stability-indicating method. The proposed HPTLC technique was applied for the analysis of TDZP in its commercial formulations. The TDZP contents of commercial tablets and injection were determined as 98.41% and 101.23%, respectively. These results suggested that the proposed HPTLC technique can be applied for the routine analysis of TDZP in its commercial products and newly established formulations.

     

Determination of Metronidazole Benzoate in Liquid Preparations by Reversed Phase HPLC

Abstract

Despite the availability of several commercial suspension formulations of metronidazole benzoate, there is no official method for its analysis. The conventional analytical method requires extensive extraction following each extraction with evaporation then non-aqueous titration with perchloric acid. The method is time-consuming and is not stability-indicating. This proposed HPLC methodology has the advantages of being fast since it requires no chemical manipulation. The assay is stability-indicating being capable of separating the hydrolysis products (metronidazole and benzoic acid) and the common impurity (2-methyl-5-nitroimidazole) from metronidazole benzoate. The method is also specific as it is free of interferences from excipients. The proposed HPLC method was verified for linearity, accuracy and precision and was applied successfully to several commercial suspensions.     

Determination of Atenolol and Its Related Compounds by Ion Pair High Performance Liquid Chromatography

Abstract

A rapid, simple, accurate, and stability-indicating ion pair high performance liquid chromatography (IP HPLC) procedure is presented for the determination of atenolol in pharmaceutical tablets. The related compounds of atenolol were separated, making the determination specific for atenolol. An aliquot of the sample is dissolved in methanol containing N-butyryl-4-aminophenol as an internal standard and chromatographed on a supelcosil LC-8-DB (5μ) (25Omm × 4.6mm i.d.) column using 1.0 mM ammonium acetate and 2.0 mM octanesulfonic acid sodium salt in acetonitrile: water (25:75) solution adjusted to pH 3.5 with glacial acetic acid as the mobile phase. The detection was carried at 225 nm. The relative standard deviations are less than 1.0% for the two commercial products analyzed. The method was tested for linearity, recovery, and specificity.     

Rapid separation and determination of process-related substances of paracetamol using reversed-phase HPLC with photo diode array as a detector.

A simple and rapid gradient reversed-phase high-performance liquid chromatographic method for simultaneous separation and determination of paracetamol and its related compounds in bulk drugs and pharmaceutical formulations has been developed. As many as nine process impurities and one degradation product of paracetamol have been separated on a Symmetry C18 column (4.6 x 250 mm i.d., particle size 5 microm) with gradient elution using 0.01 M potassium dihydrogen phosphate buffer (pH 3.0) and acetonitrile as mobile phase and photo diode array detection at 215 nm. The chromatographic behavior of all the compounds was examined under variable compositions of different solvents, temperatures, buffer concentrations and pH values. The correlation coefficients for calibration curves for paracetamol as well as impurities were in the range of 0.9951 - 0.9994. The proposed RP-LC method was successfully applied to the analysis of commercial formulations; the recoveries of paracetamol were in the range of 99-101%. The method could be of use not only for rapid and routine evaluation of the quality of paracetamol in bulk drug manufacturing units but also for detection of its impurities in pharmaceutical formulations.     

Stability-Indicating LC Method for the Determination of Olmesartan in Bulk Drug and in Pharmaceutical Dosage Form

A novel stability-indicating LC assay method was developed and validated for quantitative determination of olmesartan in bulk drugs and in pharmaceutical dosage form in the presence of degradation products generated from forced degradation studies. An isocratic, reversed phase LC method was developed to separate the drug from the degradation products, using an Ace5-C18 (250 mm × 4.6 mm, 5 μm) column, and 50 mM ammonium acetate (pH-5.5 by acetic acid) and acetonitrile (70:30 v/v) as a mobile phase. The detection was carried out at the wavelength of 235 nm. The olmesartan was subjected to stress conditions of hydrolysis (acid, base), oxidation, photolysis and thermal degradation. Degradation was observed for olmesartan in acid, base and in 30% H₂O₂ conditions. The drug was found to be stable in the other stress conditions attempted. The degradation products were well resolved from the main peak. The percentage recovery of olmesartan ranged from (99.89 to 100.95%) in pharmaceutical dosage form. The developed method was validated with respect to linearity, accuracy (recovery), precision, specificity and robustness. The forced degradation studies prove the stability-indicating power of the method.

     

Validated Stability-Indicating HPLC–DAD Determination of the Antihypertensive Binary Mixture of Carvedilol and Hydrochlorothiazide in Tablet Dosage Forms

A simple and selective HPLC with diode array detection stability-indicating method was developed for the simultaneous determination of the antihypertensive drugs carvedilol (CRV) and hydrochlorothiazide (HCT) in their combined formulations. Effective chromatographic separation was achieved using Zorbax SB-C8 column (4.6 × 250 mm, 5 μm) with gradient elution of the mobile phase composed of 0.025 M phosphoric acid and acetonitrile at a flow rate of 1 mL min^?1. The multiple wavelength detector was set at 242 nm for measurement of CRV and 271 nm for HCT. Quantification was based on measuring the peak areas. The cited drugs were resolved with retention times 4.9 and 6.7 min for HCT and CRV, respectively. Analytical performance of the proposed HPLC procedure was thoroughly validated with respect to system suitability, linearity, ranges, precision, accuracy, specificity, robustness, detection and quantification limits. The linearity ranges were 5–300 and 5–200 μg mL^?1 for CRV and HCT, respectively, with correlation coefficients >0.9996. Both drugs were subjected to stress conditions of acidic and alkaline hydrolysis, oxidation, photolysis and thermal degradation. The proposed method proved to be stability-indicating by resolution of the drugs from their forced degradation products. Moreover, specificity of the method was verified by resolution of both drugs from more than 20 pharmaceutical compounds of various medicinal categories. The validated HPLC method was applied to the analysis of the cited antihypertensive drugs in their combined tablet dosage forms. The proposed method made use of DAD as a tool for peak identity and purity confirmation.     

A Novel Ultra-High Performance Liquid Chromatography Method for the Determination of Erdosteine,Related Impurities and Degradation Products in New Effervescent Tablets

A novel and automated, stability-indicating, reversed phase ultra performance liquid chromatography (UPLC) method was developed and validated for the quantitative determination of erdosteine, its known impurities and two novel degradation products in a new pharmaceutical dosage form (effervescent tablets). The chromatographic separations were performed on a Waters Acquity UPLC HSS T3, 1.8 µm (2.1 mm?×?150 mm, I.D.) stainless steel column. The mobile phase consisted of 0.1% TFA in water and methanol under gradient elution conditions, at a flow rate of 0.29 mL/min, for the assay and impurities analysis. UV detection was set at a wavelength of 238 nm. Erdosteine raw material, placebo and effervescent tablets were subjected to forced degradation. The new degradation products (labeled OX1 and OX2) were found after oxidative treatment and characterized by ultra performance liquid chromatography mass spectrometry. The validation parameters such as linearity, limit of detection (LOD) and quantification (LOQ), accuracy, precision, specificity and robustness were highly satisfactory for all analyzed compounds. LOD (0.020 and 0.011–0.385 µg/mL for erdosteine and impurities, respectively) and LOQ values show the high sensibility of the method. Specificity of the method was confirmed by testing the matrix components. The validated method demonstrated to be suitable for routine quality control purposes and for routine stability studies of erdosteine in effervescent formulations.     

Stability-Indicating LC Method for Assay of Cholecalciferol

This paper discusses the development of a stability-indicating reversed-phase LC method for analysis of cholecalciferol as the bulk drug and in formulations. The mobile phase was acetonitrile–methanol–water 50:50:2 (v/v). The calibration plot for the drug was linear in the range 0.4–10 μg mL⁻¹. The method was accurate and precise with limits of detection and quantitation of 64 and 215 ng, respectively. Mean recovery was 100.71%. The method was used for analysis of cholecalciferol in pharmaceutical formulations in the presence of its degradation products and commonly used excipients.

     

Development and validation of a stability-indicating LC method for the assay of adapalene in bulk drug and pharmaceutical formulations

A simple and sensitive reverse phase HPLC method for the determination of adapalene (ADP) in bulk drug samples and pharmaceutical formulations has been developed and validated. The separation of ADP was achieved on an Inertsil ODS-3V (5 ??m, 15 cm ± 4.6 mm i.d.) column using UV detector at 230 nm. The mobile phase consisted of ammonium acetate (25 mM, pH 3.0), methanol and tetrahydrofuran (18: 42: 40 v/v). The linear range of detection was 2?C200 ??g/mL (R = 0.9991). Intra- and inter-day assay relative standard deviation values were less than 0.5%. The method has been successfully applied to the determination of ADP in pharmaceutical preparations. The excipients commonly present in formulations did not interfere with the assay of ADP. Analytical parameters were calculated and complete statistical evaluation was performed.     

Validation of an HPLC Analytical Method for Determination of Pancuronium Bromide in Pharmaceutical Injections

Abstract

Pancuronium bromide is used with general anesthesia in surgery for muscle relaxation and as an aid to intubation. A high performance liquid chromatographic method was fully validated for the quantitative determination of pancuronium bromide in pharmaceutical injectable solutions. The analytical method was performed on an amino column (Luna® 150 mm × 4.6 mm, 5 µm). The mobile phase was composed of acetonitrile:water containing 50 mmol L^?1 of 1-octane sulfonic acid sodium salt (20:80 v/v) with a flow rate of 1.0 mL min^?1 and ultraviolet (UV) detection at 210 nm. The proposed analytical method was compared with that described in the British Pharmacopoeia.     

Validated Stability-Indicating HPLC–DAD Determination of the Antihypertensive Binary Mixture of Carvedilol and Hydrochlorothiazide in Tablet Dosage Forms

A simple and selective HPLC with diode array detection stability-indicating method was developed for the simultaneous determination of the antihypertensive drugs carvedilol (CRV) and hydrochlorothiazide (HCT) in their combined formulations. Effective chromatographic separation was achieved using Zorbax SB-C8 column (4.6 × 250 mm, 5 μm) with gradient elution of the mobile phase composed of 0.025 M phosphoric acid and acetonitrile at a flow rate of 1 mL min⁻¹. The multiple wavelength detector was set at 242 nm for measurement of CRV and 271 nm for HCT. Quantification was based on measuring the peak areas. The cited drugs were resolved with retention times 4.9 and 6.7 min for HCT and CRV, respectively. Analytical performance of the proposed HPLC procedure was thoroughly validated with respect to system suitability, linearity, ranges, precision, accuracy, specificity, robustness, detection and quantification limits. The linearity ranges were 5–300 and 5–200 μg mL⁻¹ for CRV and HCT, respectively, with correlation coefficients >0.9996. Both drugs were subjected to stress conditions of acidic and alkaline hydrolysis, oxidation, photolysis and thermal degradation. The proposed method proved to be stability-indicating by resolution of the drugs from their forced degradation products. Moreover, specificity of the method was verified by resolution of both drugs from more than 20 pharmaceutical compounds of various medicinal categories. The validated HPLC method was applied to the analysis of the cited antihypertensive drugs in their combined tablet dosage forms. The proposed method made use of DAD as a tool for peak identity and purity confirmation.

     

RP‐HPLC Method for the Determination of Tenofovir in Pharmaceutical Formulations and Spiked Human Plasma

Abstract

A simple reverse‐phase high‐performance liquid chromatographic method for the determination of tenofovir disoproxil fumarate (TDF) in pharmaceutical formulations and human plasma samples has been developed and validated. Piroxicam (PRX) was used as an internal standard. The assay of the drug was performed on a CLC C₁₈ (5 μ, 25 cm×4.6 mm i.d.) with UV detection at 259 nm. The mobile phase consisted of acetonitrile–water mixture in the ratio of 75∶25, and a flow rate of 1 ml/min was maintained. The standard curve was linear over the range of 0.2–10 µg/ml (r ²=0.9966). Analytic parameters have been evaluated. Within‐day and between‐day precision as expressed by relative standard deviation was found to be less than 2%. The method has been applied successfully for the determination of TDF in spiked human plasma samples and pharmaceutical formulations.     

Spectrophotometric Determination of Bio-Active Compounds in Commercial Samples with Nitrous Acid and Cresyl Fast Violet Acetate

Abstract

A simple and sensitive method for the spectrophotometric determination of bio-active compounds (drugs and non-nutritive sweeteners) containing reactive functional groups, viz., aromatic primary amine (drugs: Dapsone, Sulphamethoxazole), aromatic secondary amine (drug: Pindolol), aliphatic secondary amine (non-nutritive sweetener: Cyclamate), acid hydrazide (drug: Isoniazid) and thiol (drug: Captopril), is proposed. The method involves the addition of excess of sodium nitrite to the compound in the presence of 0.25 M hydrochloric acid solution and the unreacted nitrous acid is determined by the measurement of corresponding decrease in the absorbance of cresyl fast violet acetate (Δ_max : 555 nm), the most suitable one out of several dyes tested. This method was applied for the determination of bio-active compounds in commercial samples. (drugs: pharmaceutical formulations; non-nutritive sweetener (cyclamate): foodstuffs). The newly proposed method enabled the determination of the bio-active compounds in microgram quantities (0.1 - 0.5 μg/ml). Standard deviation values evaluated through replicate determinations were found to be < 0.5 mg per dose (RSD : 0.5 - 1.2%). The common excipients do not effect the determination of the drugs in pharmaceutical formulations. Many of the usually occurring additives in food stuffs are tolerated to a very high level in the determination of cyclamate in beverages, syrup, ice candy and ice cream. The validity of the method was tested against the reference method. Recoveries to the tune of 99.2 - 101.1% were observed by adopting this method.     

Development and validation of a single RP-HPLC assay method for analysis of bulk raw material batches of four parabens that are widely used as preservatives in pharmaceutical and cosmetic products

A stability-indicating, robust, fast, and user friendly reversed-phase high-performance liquid chromatographic (RP-HPLC) assay method has been developed and validated for the analysis of commercial raw material batches of methylparaben, ethylparaben, propylparaben, and butylparaben. These four parabens are widely used as preservatives in pharmaceutical and cosmetic products. Accurate assay value of each of the parabens in their respective commercial lots is critical to determine the correct weight of the paraben that is needed to obtain the target concentration of the paraben in a specific lot of pharmaceutical or cosmetic products. Currently, there are no single HPLC assay methods (validated as per ICH requirements) available in the literature that can be used to analyze the commercial lots of each of the four parabens. The analytical method reported herein analyzes all four parabens in less than 10 min. The method presented in this report was successfully validated as per ICH guidelines. Therefore, this method can be implemented in QC laboratories to analyze and assay the commercial bulk lots of the four parabens.     

Determination of glimepiride in pharmaceutical formulations using HPLC and first-derivative spectrophotometric methods

Two validated analytical methods have been developed to determine glimepiride in pharmaceutical formulations using HPLC and 1st order derivative spectrophotometric techniques. Employing reverse phase HPLC method, the drug was analyzed by pumping a mixture of acetonitrile and 2% formic acid solution, pH 3.5 (80: 20 v/v) through a C₁₈ column (250 × 4.6 mm, 5 μm) and detecting the eluents at 228 nm. The linearity range was found to be 20–140 μg/mL with mean recovery of 100.52 ± 0.33%. The second method was based on the formation of a complex of the drug with 2,3,5-triphenyl-2H-tetrazolium chloride in basic media. 1st order derivative spectrum made it possible to detect the complex at 413.5 nm. The linearity range was found to be 40–160 μg/mL, with mean recovery of 100.33 ± 0.47%. Both the proposed methods can reliably be used for routine analysis of glimepiride in raw material as well as in pharmaceutical formulations. The article is published in the original.     

Development and Validation of LC Method for Simultaneous Determination of Two Binary Mixtures Containing Indapamide

A new sensitive, simple, rapid, and precise RP LC method with hydrochlorothiazide as internal standard has been developed for resolving two binary mixtures, perindopril with indapamide and captopril with indapamide, in pharmaceutical formulations. The drugs were separated at room temperature on a 250 mm × 4.6 mm, 5-μm particle, cyanopropyl column with 10 mm KH₂PO₄, pH 6.0-methanol 55:45 (v/v) as mobile phase at a flow rate of 1 mL min^?1. Detection was at 210 nm. Factors affecting the separation process were studied and optimized. The linearity, accuracy, and precision of the method were good, and the method was successfully applied to the determination of the two binary combinations in synthetic mixtures and commercial pharmaceutical products.     

A porous graphitized carbon column HPLC method for the quantification of paracetamol, pseudoephedrine, and chlorpheniramine in a pharmaceutical formulation

A simple, rapid, and stability-indicating HPLC method has been developed, fully validated, and applied to the quantification of paracetamol, pseudoephedrine hydrochloride, and chlorpheniramine maleate in a pharmaceutical formulation, using hydrochlorothiazide as an internal standard. Chromatographic separation was achieved isocratically on an RP porous graphitized carbon analytical column (125 x 2.1 mm id, particle size 5 microm) using 5.0 mM ammonium acetate-acetonitrile (35 + 65, v/v) mobile phase at a flow rate of 0.50 mL/min. UV spectrophotometric detection at 220 nm was used. The method had linear calibration curves over the range of 30-70 microg/mL for paracetamol, 1.8-4.2 microg/mL for pseudoephedrine hydrochloride, and 120-280 ng/mL for chlorpheniramine maleate. The intraday and interday RSD values were less than 3.2% for all compounds, while the relative error was less than 2.9%. Accelerated stability studies performed under various stress conditions proved the selectivity of the method. The developed method was applied successfully to QC and content uniformity tests of commercial tablets.