Quantitation of itopride in human serum by high-performance liquid chromatography with fluorescence detection and its application to a bioequivalence study

• Published in: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Vol. 818; no. 2; pp. 213 - 220
• Main Authors: Singh, Sonu Sundd, Jain, Manish, Sharma, Kuldeep, Shah, Bhavin, Vyas, Meghna, Thakkar, Purav, Shah, Ruchy, Singh, Shriprakash, Lohray, Brajbhushan
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier Science 25.04.2005
• Subjects: Absorption; Adult; Analysis; Analytical, structural and metabolic biochemistry; Benzamides - blood; Benzamides - isolation & purification; Benzamides - pharmacokinetics; Benzyl Compounds - blood; Benzyl Compounds - isolation & purification; Benzyl Compounds - pharmacokinetics; Bioequivalence; Biological and medical sciences; Chromatography, High Pressure Liquid - methods; Drug Stability; Freezing; Fundamental and applied biological sciences. Psychology; General pharmacology; Human serum; Humans; Itopride; Male; Medical sciences; Pharmacology. Drug treatments; Reproducibility of Results; Sensitivity and Specificity; Spectrometry, Fluorescence; Therapeutic Equivalency; Validation; Human; Validation; Biological fluid; Human serum; Fluorescence detector; Bioequivalence; Itopride; HPLC chromatography; Benzamide derivatives; Serum; Pharmacokinetics; Quantitative analysis
• ISSN: 1570-0232
• DOI: 10.1016/j.jchromb.2004.12.035

A new method was developed for determination of itopride in human serum by reversed phase high-performance liquid chromatography (HPLC) with fluorescence detection (excitation at 291 nm and emission at 342 nm). The method employed one-step extraction of itopride from serum matrix with a mixture of tert-butyl methyl ether and dichloromethane (70:30, v/v) using etoricoxib as an internal standard. Chromatographic separation was obtained within 12.0 min using a reverse phase YMC-Pack AM ODS column (250 mm x 4.6 mm, 5 microm) and an isocratic mobile phase constituting of a mixture of 0.05% tri-fluoro acetic acid in water and acetonitrile (75:25, v/v) flowing at a flow rate of 1.0 ml/min. The method was linear in the range of 14.0 ng/ml to 1000.0 ng/ml. The lower limit of quantitation (LLOQ) was 14.0 ng/ml. Average recovery of itopride and the internal standard from the biological matrix was more than 66.04 and 64.57%, respectively. The inter-day accuracy of the drug containing serum samples was more than 97.81% with a precision of 2.31-3.68%. The intra-day accuracy was 96.91% or more with a precision of 5.17-9.50%. Serum samples containing itopride were stable for 180.0 days at -70+/-5 degrees C and for 24.0 h at ambient temperature (25+/-5 degrees C). The method was successfully applied to the bioequivalence study of itopride in healthy, male human subjects.