In vivo metabolic investigation of moxifloxacin using liquid chromatography/electrospray ionization tandem mass spectrometry in combination with online hydrogen/deuterium exchange experiments

Abstract

RATIONALE Tuberculosis is a leading cause of death from an infectious disease and moxifloxacin is an effective drug as compared to other fluoroquinolones. To date only two metabolites of the drug are known. Therefore, the present study on characterization of hitherto unknown in vivo metabolites of moxifloxacin using liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI‐MS/MS) is undertaken. METHODS In vivo metabolites of moxifloxacin have been identified and characterized by using LC/ESI‐MS/MS in combination with an online hydrogen/deuterium (H/D) exchange technique. To identify in vivo metabolites, blood, urine and faeces samples were collected after oral administration of moxifloxacin to Sprague–Dawley rats. The samples were prepared using an optimized sample preparation approach involving protein precipitation, liquid‐liquid extraction followed by solid‐phase extraction and LC/MS/MS analysis. RESULTS A total of nine phase I and ten phase II metabolites of moxifloxacin have been identified in urine samples including N‐sulphated, glucuronide and hydroxylated metabolites which are also observed in plasma samples. In faeces samples, only the N‐sulphated metabolite is observed. The structures of metabolites have been elucidated based on fragmentation patterns, accurate mass measurements and online H/D exchange LC/MS/MS experiments. Online H/D exchange experiments are used to support the identification and structural characterization of drug metabolites. CONCLUSIONS A total of 19 in vivo metabolites of moxifloxacin have been characterized using LC/ESI‐MS/MS in combination with accurate mass measurements and online H/D exchange experiments. The main phase I metabolites of moxifloxacin are hydroxylated, decarbonylated, desmethylated and desmethylhydroxylated metabolites which undergo subsequent phase II glucuronidation pathways. Copyright © 2012 John Wiley & Sons, Ltd.

Fragmentation of Moxifloxacin and Its Analogs by Electrospray Ionization Time-of-Flight Mass Spectrometry

“…However, the relative abundances of the product ions resulting from the same fragmentation patterns varied greatly. Raju et al (2012) reported several characteristic ions formed by cleavage of the benzene ring, whereas negligible amounts of these products were observed in this analysis. Many abundant radical ions resulting from loss of CH 3 or CH 3 O were detected in this work, although Raju et al (2012) observed small numbers of these ions.…”

“…Raju et al (2012) reported several characteristic ions formed by cleavage of the benzene ring, whereas negligible amounts of these products were observed in this analysis. Many abundant radical ions resulting from loss of CH 3 or CH 3 O were detected in this work, although Raju et al (2012) observed small numbers of these ions. Although the structures of moxifloxacin and analog 1 are similar, the relative abundances of products from the same fragmentation patterns varied greatly ( Figure 2 and Table 2).…”

“…Interestingly, many of the abundant product ions at m=z 298, 278, 260, and 232 were radicals (Cai, Mo, and Rannulu 2010;Xu et al 2012;Wu et al 2012). The fragmentation pathways of protonated moxifloxacin were different from those reported by Raju et al (2012), although the analyses were performed with instruments from different manufacturers. The losses of neutral species, such as CO, H 2 O, CO 2 , and HF, and the cleavage of piperidine or pyrrolidine occurred in both studies.…”

“…Raju et al (2012) recently determined the fragmentation pathways of moxifloxacin by electrospray ionization (ESI)-quadrupole ion trap mass spectrometry and ESI-quadrupole-time-of-flight (QTOF). Unexpectedly, in studying this compound, a different tandem mass spectrum spectrum using ESI-QTOF was determined.…”

Editorial: Drug Metabolism and Toxicology

“…These metabolism studies are generally carried out by employing in vitro and in vivo systems [8] followed by using modern analytical techniques. Liquid chromatography-mass spectrometry (LC-MS) is the most popular and versatile analytical technique which has been widely used for the identification of trace levels of drugs and their metabolites in various biological matrices due to its high sensitivity [9][10][11][12]. Several strategies, such as MS/MS, MS n , on-line hydrogen/ deuterium exchange experiments, accurate mass measurements, softwares dedicated to the metabolite prediction (in silico tools), chemical derivatization and radio-labeling of parent drugs were used for structural characterization of metabolites [9][10][11][12][13][14][15].…”