Development and validation of a HPLC method for quantification of rivastigmine in rat urine and identification of a novel metabolite in urine by LC-MS/MS

Karthik Arumugam, Mallikarjuna Rao Chamallamudi, Ravindranath Reddy Gilibili, Ramesh Mullangi, Subramanian Ganesan, Sidhartha S. Kar, Ranjithkumar Averineni, Gopal Shavi, Nayanabhirama Udupa

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A sensitive, specific and accurate HPLC method for the quantification of rivastigmine (RSM) in rat urine was developed and validated. The method involves the simple liquid-liquid extraction of RSM and pyridostigmine as an internal standard (IS) from rat urine with tertiary methyl butyl ether. The chromatographic separation of RSM and IS was achieved with 20mm ammonium acetate buffer (pH 6.5) and acetonitrile (65:35, v/v) delivered at flow-rate of 1mL/min on a Kromasil KR-100. The method was in linear range from 50 to 5000ng/mL. The validation was done as per FDA guidelines and the results met the acceptance criteria. The method was successfully applied for the quantification of RSM in rat urine. Besides method validation, we have identified two metabolites of RSM in urine. Both the metabolites were characterized by HPLC-PDA and LC-MS/MS and it was found that one metabolite is novel.

Original languageEnglish
Pages (from-to)353-361
Number of pages9
JournalBiomedical Chromatography
Issue number3
Publication statusPublished - 03-2011


All Science Journal Classification (ASJC) codes

  • Analytical Chemistry
  • Biochemistry
  • Molecular Biology
  • Pharmacology
  • Drug Discovery
  • Clinical Biochemistry

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