Skip to main content
SpringerLink
Log in

SWATH: A Data-Independent Tandem Mass Spectrometry Method to Quantify 13C Enrichment in Cellular Metabolites and Fragments

  • Protocol
  • First Online:
Metabolic Flux Analysis in Eukaryotic Cells

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2088))

Abstract

Recently, the sequential windowed acquisition of all theoretical fragment ion mass spectra (SWATH) method coupled with liquid chromatography has been demonstrated for the quantification of isotopic 13C enrichment in a large number of cellular metabolites and fragments. SWATH, a data–independent acquisition (DIA) method, alleviates the need for data deconvolution and shows greater accuracy in the quantification of low abundance isotopologs of fragments thereby resulting in a lower systematic error. Here we provide a detailed protocol for the design of Q1 mass isolation windows and the post–acquisition data analysis with emphasis on the untargeted nature of SWATH.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Zamboni N, Sauer U (2009) Novel biological insights through metabolomics and13C-flux analysis. Curr Opin Microbiol 12:553–558. https://doi.org/10.1016/j.mib.2009.083

    Article  CAS  PubMed  Google Scholar 

  2. Sauer U (2006) Metabolic networks in motion: 13C-based flux analysis. Mol Syst Biol 2:62. https://doi.org/10.1038/msb4100109

    Article  PubMed  PubMed Central  Google Scholar 

  3. Rühl M, Rupp B, Nöh K et al (2012) Collisional fragmentation of central carbon metabolites in LC-MS/MS increases precision of 13C metabolic flux analysis. Biotechnol Bioeng 109:763–771. https://doi.org/10.1002/bit.24344

    Article  CAS  PubMed  Google Scholar 

  4. McCloskey D, Young JD, Xu S et al (2016) MID Max: LC-MS/MS method for measuring the precursor and product mass isotopomer distributions of metabolic intermediates and cofactors for metabolic flux analysis applications. Anal Chem 88:1362–1370. https://doi.org/10.1021/acs.analchem.5b03887

    Article  CAS  PubMed  Google Scholar 

  5. Jaiswal D, Prasannan CB, Hendry JI, Wangikar PP (2018) SWATH tandem mass spectrometry workflow for quantification of mass isotopologue distribution of intracellular metabolites and fragments labeled with isotopic 13C Carbon. Anal Chem 90:6486–6493. https://doi.org/10.1021/acs.analchem.7b05329

    Article  CAS  PubMed  Google Scholar 

  6. Li Z, Li Y, Chen W et al (2017) Integrating MS1 and MS2 scans in high-resolution parallel reaction monitoring assays for targeted metabolite quantification and dynamic 13 C-labeling metabolism analysis. Anal Chem 89:877–885. https://doi.org/10.1021/acs.analchem.6b03947

    Article  CAS  PubMed  Google Scholar 

  7. Zhu X, Chen Y, Subramanian R (2014) Comparison of information-dependent acquisition, SWATH, and MS all techniques in metabolite identification study employing ultrahigh-performance liquid chromatography-quadrupole time-of-flight mass spectrometry. Anal Chem 86:1202–1209. https://doi.org/10.1021/ac403385y

    Article  CAS  PubMed  Google Scholar 

  8. Tsugawa H, Cajka T, Kind T et al (2015) MS-DIAL: data-independent MS/MS deconvolution for comprehensive metabolome analysis. Nat Methods 12:523–526. https://doi.org/10.1038/nmeth.3393

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Li H, Cai Y, Guo Y et al (2016) MetDIA: Targeted metabolite extraction of multiplexed MS/MS spectra generated by data-independent acquisition. Anal Chem 88:8757–8764. https://doi.org/10.1021/acs.analchem.6b02122

    Article  CAS  PubMed  Google Scholar 

  10. Hendry JI, Prasannan C, Ma F et al (2017) Rerouting of carbon flux in a glycogen mutant of cyanobacteria assessed via isotopically non-stationary 13 C metabolic flux analysis. Biotechnol Bioeng 114:2298–2308. https://doi.org/10.1002/bit.26350

    Article  CAS  PubMed  Google Scholar 

  11. Prasannan CB, Jaiswal D, Davis R, Wangikar PP (2018) An improved method for extraction of polar and charged metabolites from cyanobacteria. PLoS 2018:1–16

    Google Scholar 

  12. Mccloskey D, Gangoiti JA (2015) A pH and solvent optimized reverse-phase ion-paring-LC–MS/MS method that leverages multiple scan-types for targeted absolute quantification of intracellular metabolites. Metabolomics 11:1338–1350. https://doi.org/10.1007/s11306-015-0790-y

    Article  CAS  Google Scholar 

  13. Luo B, Groenke K, Takors R et al (2007) Simultaneous determination of multiple intracellular metabolites in glycolysis, pentose phosphate pathway and tricarboxylic acid cycle by liquid chromatography-mass spectrometry. J Chromatogr A 1147:153–164. https://doi.org/10.1016/j.chroma.2007.02.034

    Article  CAS  PubMed  Google Scholar 

  14. Lu W, Clasquin MF, Melamud E et al (2010) Metabolomic analysis via reversed-phase ion-pairing liquid chromatography coupled to a stand alone orbitrap mass spectrometer. Anal Chem 82:3212–3221

    Article  CAS  Google Scholar 

  15. Arnhard K, Gottschall A, Pitterl F, Oberacher H (2015) Applying “Sequential Windowed Acquisition of All Theoretical Fragment Ion Mass Spectra” (SWATH) for systematic toxicological analysis with liquid chromatography-high-resolution tandem mass spectrometry. Anal Bioanal Chem 407:405–414. https://doi.org/10.1007/s00216-014-8262-1

    Article  CAS  PubMed  Google Scholar 

  16. Röst HL, Rosenberger G, Navarro P et al (2014) OpenSWATH enables automated, targeted analysis of data-independent acquisition MS data. Nat Biotechnol 32:219–223. https://doi.org/10.1038/nbt.2841

    Article  CAS  PubMed  Google Scholar 

  17. Schubert OT, Gillet LC, Collins BC et al (2015) Building high-quality assay libraries for targeted analysis of SWATH MS data. Nat Protoc 10:426–441. https://doi.org/10.1038/nprot.2015.015

    Article  CAS  PubMed  Google Scholar 

  18. Oswald S, Gröer C, Drozdzik M, Siegmund W (2013) Mass spectrometry-based targeted proteomics as a tool to elucidate the expression and function of intestinal drug transporters. AAPS J 15:1128–1140. https://doi.org/10.1208/s12248-013-9521-3

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Jaiswal D, Sengupta A, Sohoni S et al (2018) Genome features and biochemical characteristics of a robust, fast growing and naturally transformable cyanobacterium Synechococcus elongatus PCC 11801 isolated from India. Sci Rep 8:16632. https://doi.org/10.1038/s41598-018-34872-z

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Tautenhahn R, Patti GJ, Rinehart D, Siuzdak G (2012) XCMS online: a web-based platform to process untargeted metabolomic data. Anal Chem 84:5035–5039. https://doi.org/10.1021/ac300698c.XCMS

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Kanehisa M, Goto S (2000) KEGG: kyoto encyclopedia of genes and genomes. Nucleic Acids Res 28:27–30. https://doi.org/10.1088/1751-8113/44/8/085201

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Millard P, Letisse F, Sokol S, Portais JC (2012) IsoCor: correcting MS data in isotope labeling experiments. Bioinformatics 28:1294–1296. https://doi.org/10.1093/bioinformatics/bts127

    Article  CAS  Google Scholar 

  23. Tautenhahn R, Cho K, Uritboonthai W et al (2012) An accelerated workflow for untargeted metabolomics using the METLIN database. Nat Biotechnol 30:826–828. https://doi.org/10.1038/nbt.2348

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Bateman KP, Castro-Perez J, Wrona M et al (2007) MSE with mass defect filtering for in vitro and in vivo metabolite identification. Rapid Commun Mass Spectrom 21:1485–1496. https://doi.org/10.1002/rcm.2996

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This work was supported by a grant from Department of Biotechnology (DBT), Government of India, awarded to PPW toward DBT-Pan IIT Center for Bioenergy (Grant no. BT/EB/PAN IIT/2012).

Author information

Authors and Affiliations

  1. Department of Chemical Engineering, Indian Institute of Technology Bombay, Mumbai, India

    Damini Jaiswal & Pramod P. Wangikar

  2. DBT-PAN IIT Centre for Bioenergy, Indian Institute of Technology Bombay, Mumbai, India

    Pramod P. Wangikar

  3. Wadhwani Research Centre for Bioengineering, Indian Institute of Technology Bombay, Mumbai, India

    Pramod P. Wangikar

Authors
  1. Damini Jaiswal
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Pramod P. Wangikar
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Pramod P. Wangikar .

Editor information

Editors and Affiliations

  1. Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA

    Deepak Nagrath

Rights and permissions

Reprints and permissions

Copyright information

© 2020 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Jaiswal, D., Wangikar, P.P. (2020). SWATH: A Data-Independent Tandem Mass Spectrometry Method to Quantify 13C Enrichment in Cellular Metabolites and Fragments. In: Nagrath, D. (eds) Metabolic Flux Analysis in Eukaryotic Cells. Methods in Molecular Biology, vol 2088. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0159-4_9

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-0159-4_9

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0158-7

  • Online ISBN: 978-1-0716-0159-4

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation