Skip to main content

Statistical Analysis of Metabolomics Data

  • Protocol
  • First Online:
Metabolomics Tools for Natural Product Discovery

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1055))

Abstract

Statistical matters form an integral part of a metabolomics experiment. In this chapter we describe several important aspects in the analysis of metabolomics data such as the removal of unwanted variation and the identification of differentially abundant metabolites, along with a number of other essential statistical considerations.

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

Access this chapter

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. Fiehn O (2002) Metabolomics—the link between genotypes and phenotypes. Plant Mol Biol 48:155–171

    Article  PubMed  CAS  Google Scholar 

  2. Roessner U, Bowne J (2009) What is metabolomics all about? Biotechniques 46(5):363–365

    Article  PubMed  CAS  Google Scholar 

  3. Roessner U, Beckles DM (2009) Metabolite measurements. Springer, New York

    Google Scholar 

  4. De Livera AM, Dias DA, De Souza D, Rupasinghe T, Pyke J, Tull D, Roessner U, McConville M, Speed TP (2012) Normalising and integrating metabolomics data. Anal Chem 84(24):10768–10776. DOI:10.1021/ac302748b

    Google Scholar 

  5. Glass DJ (2007) Experimental design for biologists. Cold Spring Harbor Laboratory, New York

    Google Scholar 

  6. Montgomery DC (2008) Design and analysis of experiments. Wiley, Hoboken

    Google Scholar 

  7. O’Callaghan S, Desouza DP, Isaac A, Wang Q, Hodkinson L, Olshansky M, Erwin T, Appelbe B, Tull DL, Roessner U, Bacic A, McConville MJ, Likic VA (2012) PyMS: a Python toolkit for processing of gas chromatography–mass spectrometry (GC–MS) data. Application and comparative study of selected tools. BMC Bioinformatics 13(1):115

    Google Scholar 

  8. Schleif F-M (2007) Preprocessing of nuclear magnetic resonance spectrometry data. Technical report, August 2007

    Google Scholar 

  9. Katajamaa M, Orešič M (2007) Data processing for mass spectrometry-based metabolomics. J Chromatogr A 1158:318–328

    Article  PubMed  CAS  Google Scholar 

  10. Xia J, Psychogios N, Young N, Wishart DS (2009) MetaboAnalyst: a web server for metabolomic data analysis and interpretation. Nucleic Acids Res 37:W652–W660

    Article  PubMed  CAS  Google Scholar 

  11. Hrydziuszko O, Viant MR (2012) Missing values in mass spectrometry based metabolomics: an undervalued step in the data processing pipeline. Metabolomics 8(1):161–174

    Article  CAS  Google Scholar 

  12. Katajamaa M, Oresic M (2005) Processing methods for differential analysis of LC/MS profile data. BMC Bioinformatics 6:179

    Article  PubMed  Google Scholar 

  13. Steuer R, Morgenthal K, Weckwerth W, Selbig J (2007) A gentle guide to the analysis of metabolomic data. Methods Mol Biol (Clifton, NJ) 358:105–126

    Google Scholar 

  14. Smilde AK, van der Werf MJ, Bijlsma S, van der Werff-van der Vat BJC, Jellema RH (2005) Fusion of mass spectrometry-based metabolomics data. Anal Chem 77(20):6729–6736

    Google Scholar 

  15. van den Berg RA, Hoefsloot HCJ, Westerhuis JA, Smilde AK, van der Werf MJ (2006) Centering, scaling, and transformations: improving the biological information content of metabolomics data. BMC Genomics 7:142

    Article  PubMed  Google Scholar 

  16. Temmerman L, De Livera AM, Bowne J, Sheedy RJ, Callahan DL, Nahid A, De Souza DP, Schoofs L, Tull DL, McConville JM, Roessner U, Wentworth JM (2012) Cross-platform urine metabolomics of experimental hyperglycemia in type 2 diabetes. Diab Metab vol S6:002. DOI:10.4172/2155-6156.S6-002

    Google Scholar 

  17. Roessner U, Nahid A, Chapman B, Hunter A, Bellgard M (2011) Metabolomics—the combination of analytical biochemistry, biology, and informatics, vol 1, 2nd edn. Elsevier B.V., New York

    Google Scholar 

  18. Troyanskaya O, Cantor M, Sherlock G, Brown P, Hastie T, Tibshirani R, Botstein D, Altman RB (2001) Missing value estimation methods for DNA microarrays. Bioinformatics (Oxford, England) 17(6):520–525

    Article  CAS  Google Scholar 

  19. Oba S, Sato M, Takemasa I, Monden M, Matsubara K, Ishii S (2003) A Bayesian missing value estimation method for gene expression profile data. Bioinformatics 19(16):2088–2096

    Article  PubMed  CAS  Google Scholar 

  20. van Buuren S, Groothuis-Oudshoorn K (2011) Mice: multivariate imputation by chained equations in R. J Static Softw 45(3):1–67

    Google Scholar 

  21. Goodacre R, Broadhurst D, Smilde AK, Kristal BS, Baker JD, Beger R, Bessant C, Connor S, Capuani G, Craig A, Ebbels T, Kell DB, Manetti C, Newton J, Paternostro G, Somorjai R, Sjöström M, Trygg J, Wulfert F (2007) Proposed minimum reporting standards for data analysis in metabolomics. Metabolomics 3(3):231–241

    Article  CAS  Google Scholar 

  22. Schlesselman J (1971) Power families: a note on the Box and Cox transformation. J R Stat Soc Ser B (Methodol) 307–311

    Google Scholar 

  23. Callahan DL, Roessner U, Dumontet V, De Livera AM, Doronila A, Baker AJM, Kolev SD (2012) Elemental and metabolite profiling of nickel hyperaccumulators from New Caledonia. Phytochemistry 81:80–89

    Article  PubMed  CAS  Google Scholar 

  24. Gullberg J, Jonsson P, Nordström A, Sjöström M, Moritz T (2004) Design of experiments: an efficient strategy to identify factors influencing extraction and derivatization of Arabidopsis thaliana samples in metabolomic studies with gas chromatography/mass spectrometry. Anal Biochem 331(2):283–295

    Article  PubMed  CAS  Google Scholar 

  25. Bijlsma S, Bobeldijk I, Verheij ER, Ramaker R, Kochhar S, Macdonald I, Van Ommen B, Smilde AK (2006) Large-scale human metabolomics studies: a strategy for data (pre-) processing and validation. Anal Chem 78(2):567–574

    Article  PubMed  CAS  Google Scholar 

  26. Redestig H, Fukushima A, Stenlund H, Moritz T, Arita M, Saito K, Kusano M (2009) Compensation for systematic cross-contribution improves normalization of mass spectrometry based metabolomics data. Anal Chem 81(19):7974–7980

    Article  PubMed  CAS  Google Scholar 

  27. Sysi-Aho M, Katajamaa M, Laxman Y, Oresic M (2007) Normalization method for metabolomics data using optimal selection of multiple internal standards. BMC Bioinformatics 8:93

    Article  PubMed  Google Scholar 

  28. Crawford LR, Morrison JD (1968) Computer methods in analytical mass spectrometry. Identification of an unknown compound in a catalog. Anal Chem 40(4):1464–1469

    CAS  Google Scholar 

  29. Wang W, Zhou H, Lin H, Roy S, Shaler TA, Hill LR, Norton S, Kumar P, Anderle M, Becker CH (2003) Quantification of proteins and metabolites by mass spectrometry without isotopic labeling or spiked standards. Anal Chem 75(18):481848–26

    Article  Google Scholar 

  30. Scholz M, Gatzek S, Sterling A, Fiehn O, Selbig J (2004) Metabolite fingerprinting: detecting biological features by independent component analysis. Bioinformatics (Oxford, England) 20(15):2447–2454

    Article  CAS  Google Scholar 

  31. Cairns DA, Thompson D, Perkins DN, Stanley AJ, Selby PJ, Banks RE (2008) Proteomic profiling using mass spectrometry—does normalising by total ion current potentially mask some biological differences? Proteomics 8(1):21–27

    Article  PubMed  CAS  Google Scholar 

  32. Gika HG, Macpherson E, Theodoridis GA, Wilson ID (2008) Evaluation of the repeatability of ultra-performance liquid chromatography-TOF-MS for global metabolic profiling of human urine samples. J Chromatogr B Anal Technol Biomed Life Sci 871(2):299–305

    Article  CAS  Google Scholar 

  33. Zelena E, Dunn WB, Broadhurst D, Francis-McIntyre S, Carroll KM, Begley P, O’Hagan S, Knowles JD, Halsall A, Wilson ID, Kell DB (2009) Development of a robust and repeatable UPLC-MS method for the long-term metabolomic study of human serum. Anal Chem 81(4):1357–1364

    Article  PubMed  CAS  Google Scholar 

  34. Lai L, Michopoulos F, Gika H, Theodoridis G, Wilkinson RW, Odedra R, Wingate J, Bonner R, Tate S, Wilson ID (2010) Methodological considerations in the development of HPLC-MS methods for the analysis of rodent plasma for metabolomic studies. Mol Biosyst 6(1):108–120

    Article  PubMed  CAS  Google Scholar 

  35. Dunn WB, Broadhurst D, Begley P, Zelena E, Francis-McIntyre S, Anderson N, Brown M, Knowles JD, Halsall A, Haselden JN, Nicholls AW, Wilson ID, Kell DB, Goodacre R (2011) Procedures for large-scale metabolic profiling of serum and plasma using gas chromatography and liquid chromatography coupled to mass spectrometry. Nat Protoc 6(7):1060–1083

    Article  PubMed  CAS  Google Scholar 

  36. Kamleh MA, Ebbels TMD, Spagou K, Masson P, Want EJ (2012) Optimizing the use of quality control samples for signal drift correction in large-scale urine metabolic profiling studies. Anal Chem 84(6):2670–2677

    Article  PubMed  CAS  Google Scholar 

  37. Gagnon-Bartsch JA, Speed TP (2011) Using control genes to correct for unwanted variation in microarray data. Biostatistics 13(3):539–552

    Article  PubMed  Google Scholar 

  38. Leek JT, Storey JD (2007) Capturing heterogeneity in gene expression studies by surrogate variable analysis. PLoS Genet 3(9):1724–1735

    Article  PubMed  CAS  Google Scholar 

  39. Leek JT, Storey JD (2008) A general framework for multiple testing dependence. Proc Natl Acad Sci USA 105(48):18718–18723

    Article  PubMed  CAS  Google Scholar 

  40. Tusher VG, Tibshirani R, Chu G (2001) Significance analysis of microarrays applied to the ionizing radiation response. Proc Natl Acad Sci USA 98(9):5116

    Article  PubMed  CAS  Google Scholar 

  41. Efron B (2007) Correlation and large-scale simultaneous significance testing. J Am Stat Assoc 102(477):93–103

    Article  CAS  Google Scholar 

  42. Lonnstedt I, Speed TP (2002) Replicated microarray data. Stat Sin 12:31–46

    Google Scholar 

  43. Smyth GK (2004) Linear models and empirical Bayes methods for assessing differential expression in microarray experiments. Stat Appl Genet Mol Biol 3(1):1544–6115

    Google Scholar 

  44. Holm S (1979) A simple sequentially rejective multiple test procedure. Scand J Stat 6(2):65–70

    Google Scholar 

  45. Benjamini Y, Hochberg Y (1995) Controlling the false discovery rate: a practical and powerful approach to multiple testing. J R Stat Soc Ser B 57:289–300

    Google Scholar 

  46. Westfall PH, Young SS (1993) Resampling-based multiple testing: examples and methods for p-value adjustment. Wiley-Interscience, New York

    Google Scholar 

  47. Efron B, Tibshirani R, Storey JD, Tusher V (2001) Empirical Bayes analysis of a microarray experiment. J Am Stat Assoc 96(456):1151–1160

    Article  Google Scholar 

  48. Storey JD, Tibshirani R (2001) Estimating false discovery rates under dependence, with applications to DNA microarrays. Technical report

    Google Scholar 

  49. Friedman J, Hastie T, Tibshirani R (2001) The elements of statistical learning, 2nd edn. Springer, New York

    Google Scholar 

  50. Frank IE, Friedman JH (1993) A statistical view of some chemometrics regression tools. Technometrics 35(2):109–135

    Article  Google Scholar 

  51. Wold S, Sjostrom M (2001) PLS-regression: a basic tool of chemometrics. Chemom Intell Lab Syst 58(2):109–130

    Article  CAS  Google Scholar 

  52. Vapnik V (1999) The nature of statistical learning theory. Springer, Berlin

    Google Scholar 

  53. Breiman L, Friedman JH, Olshen RA, Stone CJ (1984) Classification and regression trees. Wadsworth International Group, Belmont

    Google Scholar 

  54. Breiman L (2001) Random forests. Mach Learn 45(1):5–32

    Google Scholar 

  55. Cox TF, Cox MAA (2001) Multidimensional scaling. Chapman and Hall, Boca Raton

    Google Scholar 

  56. MacQueen J (1967) Some methods for classification and analysis of multivariate observations. In: Proceedings of the fifth Berkeley symposium on mathematical statistics and probability. University of California Press, Berkeley, pp 281–297

    Google Scholar 

  57. Kohonen T (1982) Self-organized formation of topologically correct feature maps. Biol Cybern 43(1):59–69

    Article  Google Scholar 

  58. De Livera AM, Bowne J (2013) metabolomics: A collection of functions for analysing metabolomics data. R package version 0.1.1

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2013 Springer Science+Business Media, LLC

About this protocol

Cite this protocol

De Livera, A.M., Olshansky, M., Speed, T.P. (2013). Statistical Analysis of Metabolomics Data. In: Roessner, U., Dias, D. (eds) Metabolomics Tools for Natural Product Discovery. Methods in Molecular Biology, vol 1055. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-577-4_20

Download citation

  • DOI: https://doi.org/10.1007/978-1-62703-577-4_20

  • Published:

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-576-7

  • Online ISBN: 978-1-62703-577-4

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics