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11-02-2023 | Clostridium | Original Article

Proteomic identification and quantification of Clostridium perfringens enterotoxin using a stable isotope-labelled peptide via liquid chromatography–tandem mass spectrometry

Authors: Hiroshi Koike, Maki Kanda, Souichi Yoshikawa, Hiroshi Hayashi, Yoko Matsushima, Yumi Ohba, Momoka Hayashi, Chieko Nagano, Kenji Otsuka, Junichi Kamiie, Takeo Sasamoto

Published in: Forensic Toxicology | Issue 2/2023

Abstract

Purpose

Detection of Clostridium perfringens enterotoxin (CPE) in human stool is critical evidence of food poisoning. However, processing patient-derived samples is difficult and very few methods exist to confirm the presence of CPE. In this study, a technique was developed using proteomic analysis to identify and quantify CPE in artificial gut fluid as an alternative.

Methods

The standard CPE was spiked into artificial gut fluids, and effective methods were developed by employing both a stable isotope-labelled internal standard peptide and liquid chromatography–tandem mass spectrometry (LC–MS/MS).

Results

Proteotypic peptide EILDLAAATER formed by tryptic digestion was selected for quantitation of CPE. The peptide was identified using product ion spectra. Although the nontoxic peptides originating from CPE showed very low detectability in extraction and tryptic digestion, they could be detected with sufficient sensitivity using the method we developed. Based on a spiked recovery test at two concentrations (50 and 200 µg/kg), the recovery values were 85 and 78%, respectively. The relative standard deviations of repeatability and within-laboratory reproducibility were less than 8 and 11%, respectively. These standard deviations satisfied the criteria of the Japanese validation guidelines for residues (MHLW 2010, Director Notice, Syoku-An No. 1224–1). The limit of quantification (LOQ) was estimated to be 50 µg/kg. The combination of the product ion spectra and relative ion ratio supported CPE identification at the LOQ level.

Conclusions

To the best of our knowledge, this is the first report of proteomic analysis of CPE using LC–MS/MS. The method would greatly help in assessing CPE reliably.

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Title: Proteomic identification and quantification of Clostridium perfringens enterotoxin using a stable isotope-labelled peptide via liquid chromatography–tandem mass spectrometry
Authors: Hiroshi Koike
Maki Kanda
Souichi Yoshikawa
Hiroshi Hayashi
Yoko Matsushima
Yumi Ohba
Momoka Hayashi
Chieko Nagano
Kenji Otsuka
Junichi Kamiie
Takeo Sasamoto
Publication date: 11-02-2023
Publisher: Springer Nature Singapore
Keyword: Clostridium
Published in: Forensic Toxicology / Issue 2/2023
Print ISSN: 1860-8965
Electronic ISSN: 1860-8973
DOI: https://doi.org/10.1007/s11419-023-00660-2

Keynote Webinar | Spotlight on Diet and Diabetes

Springer Medicine

Proteomic identification and quantification of Clostridium perfringens enterotoxin using a stable isotope-labelled peptide via liquid chromatography–tandem mass spectrometry

Abstract

Purpose

Methods

Results

Conclusions

Keynote Webinar | Spotlight on Diet and Diabetes

Springer Medicine

Abstract

Purpose

Methods

Results

Conclusions

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