Sensitive LC-MS/MS method for the temporal profiling of bile acids, fatty acids and branched-chain alpha-keto acids in maternal plasma during pregnancy and cord blood plasma at delivery

  • Clin Chim Acta. 2023 Jun 16;117449. doi: 10.1016/j.cca.2023.117449.
Daniel Zhi Wei Ng  1 ,  Sean Xian Yu Lee  1 ,  Delicia Shu Qin Ooi  2 ,  Le Duc Huy Ta  2 ,  Gaik Chin Yap  2 ,  Carina Jing Xuan Tay  2 ,  Chiung-Hui Huang  2 ,  Elizabeth Huiwen Tham  3 ,  Evelyn Xiu Ling Loo  4 ,  Lynette P C Shek  4 ,  Anne Goh  5 ,  Hugo P S Van Bever  4 ,  Oon Hoe Teoh  5 ,  Yung Seng Lee  6 ,  Fabian Yap  5 ,  Kok Hian Tan  5 ,  Yap-Seng Chong  7 ,  Shiao Yng Chan  7 ,  Johan Gunnar Eriksson  8 ,  Keith M Godfrey  9 ,  Bee Wah Lee  10 ,  Eric Chun Yong Chan  11
Affiliations
  • 1. Department of Pharmacy, National University of Singapore, 18 Science Drive 4, Singapore 117543.
  • 2. Department of Paediatrics, National University of Singapore, Yong Loo Lin School of Medicine, 1E Kent Ridge Road, NUHS Tower Block, Level 12, Singapore 119228.
  • 3. Department of Paediatrics, National University of Singapore, Yong Loo Lin School of Medicine, 1E Kent Ridge Road, NUHS Tower Block, Level 12, Singapore 119228; Division of Allergy & Immunology, Khoo Teck Puat-National University Children's Medical Institute, National University Health System, Singapore, Singapore.
  • 4. Department of Paediatrics, National University of Singapore, Yong Loo Lin School of Medicine, 1E Kent Ridge Road, NUHS Tower Block, Level 12, Singapore 119228; Singapore Institute for Clinical Sciences (SICS), Agency for Science, Technology and Research (A*STAR), Singapore, Singapore.
  • 5. Department of Paediatrics, KK Women's and Children's Hospital, Singapore, Singapore.
  • 6. Department of Paediatrics, National University of Singapore, Yong Loo Lin School of Medicine, 1E Kent Ridge Road, NUHS Tower Block, Level 12, Singapore 119228; Singapore Institute for Clinical Sciences (SICS), Agency for Science, Technology and Research (A*STAR), Singapore, Singapore; Division of Paediatric Endocrinology and Diabetes, Khoo Teck Puat-National University Children's Medical Institute, National University Health System, Singapore, Singapore.
  • 7. Singapore Institute for Clinical Sciences (SICS), Agency for Science, Technology and Research (A*STAR), Singapore, Singapore; Department of Obstetrics & Gynaecology, National University of Singapore, Singapore.
  • 8. Singapore Institute for Clinical Sciences (SICS), Agency for Science, Technology and Research (A*STAR), Singapore, Singapore; Department of Obstetrics & Gynaecology, National University of Singapore, Singapore; Folkhälsan Research Center, Helsinki, Finland and Department of General Practice and Primary Health Care, University of Helsinki, Helsinki, Finland; Department of General Practice and Primary Health Care, University of Helsinki, Helsinki, Finland.
  • 9. MRC Lifecourse Epidemiology Centre and NIHR Southampton Biomedical Research Centre, University of Southampton and University Hospital Southampton NHS Foundation Trust, Southampton, UK.
  • 10. Department of Paediatrics, National University of Singapore, Yong Loo Lin School of Medicine, 1E Kent Ridge Road, NUHS Tower Block, Level 12, Singapore 119228. Electronic address: [email protected].
  • 11. Department of Pharmacy, National University of Singapore, 18 Science Drive 4, Singapore 117543. Electronic address: [email protected].
PMID: 37331549 DOI: 10.1016/j.cca.2023.117449
Abstract

Background and aims: There are significant changes to the maternal inflammatory profile across pregnancy. Recent studies suggest that perturbations in maternal gut microbial and dietary-derived plasma metabolites over the course of pregnancy mediate inflammation through a complex interplay of immunomodulatory effects. Despite this body of evidence, there is currently no analytical method that is suitable for the simultaneous profiling of these metabolites within human plasma.

Materials and methods: We developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the high-throughput analysis of these metabolites in human plasma without derivatization. Plasma samples were processed using liquid-liquid extraction method with varying proportions of methyl tert-butyl ether, methanol, and water in a 3:10:2.5 ratio to reduce matrix effects.

Results: LC-MS/MS detection was sufficiently sensitive to quantify these gut microbial and dietary-derived metabolites at physiological concentrations and linear calibration curves with r2>0.99 were obtained. Recovery was consistent across concentration levels. Stability experiments confirmed that up to 160 samples could be analyzed within a single batch. The method was validated and applied to analyse maternal plasma during the first and third trimester and cord blood plasma of 5 mothers.

Conclusion: This study validated a straightforward and sensitive LC/MS-MS method for the simultaneous quantitation of gut microbial and dietary-derived metabolites in human plasma within 9 minutes without prior sample derivatization.

Keywords
Bile acid; Branch chain alpha keto acid; Fatty acid; Liquid chromatography tandem mass spectrometry; Liquid-liquid extraction; Short chain fatty acid.
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