Analytical and Bioanalytical Chemistry

, Volume 406, Issue 22, pp 5339–5346 | Cite as

Quantification of genetically modified soya using strong anion exchange chromatography and time-of-flight mass spectrometry

  • Po-Chih Chang
  • P. Muralidhar Reddy
  • Yen-Peng HoEmail author
Research Paper


Stable-isotope dimethyl labeling was applied to the quantification of genetically modified (GM) soya. The herbicide-resistant gene-related protein 5-enolpyruvylshikimate-3-phosphate synthase (CP4 EPSPS) was labeled using a dimethyl labeling reagent, formaldehyde-H2 or -D2. The identification and quantification of CP4 EPSPS was performed using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). The CP4 EPSPS protein was separated from high abundance proteins using strong anion exchange chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Then, the tryptic peptides from the samples and reference were labeled with formaldehyde-H2 and formaldehyde-D2, respectively. The two labeled pools were mixed and analyzed using MALDI-MS. The data showed a good correlation between the peak ratio of the H- and D-labeled peptides and the GM soya percentages at 0.5, 1, 3, and 5 %, with R 2 of 0.99. The labeling reagents are readily available. The labeling experiments and the detection procedures are simple. The approach is useful for the quantification of GM soya at a level as low as 0.5 %.


Genetically modified soya Dimethyl labeling reagent Matrix-assisted laser desorption/ionization mass spectrometry Quantification Anion exchange chromatography 



The authors thank the Ministry of Science and Technology of the Republic of China for supporting this research financially.

Supplementary material

216_2014_7965_MOESM1_ESM.pdf (36 kb)
ESM 1 (PDF 36 kb)


  1. 1.
    Singh OV, Ghai S, Paul D, Jain RK (2006) Genetically modified crops: success, safety assessment, and public concern. Appl Microbiol Biotechnol 71(5):598–607CrossRefGoogle Scholar
  2. 2.
    Thomson J (2003) Genetically modified food crops for improving agricultural practice and their effects on human health. Trends Food Sci Technol 14(5–8):210–228CrossRefGoogle Scholar
  3. 3.
    Uzogara SG (2000) The impact of genetic modification of human foods in the 21st century: a review. Biotechnol Adv 18(3):179–206CrossRefGoogle Scholar
  4. 4.
    Padgette S, Kolacz K, Delannay X, Re D, LaVallee B, Tinius C, Rhodes W, Otero Y, Barry G, Eichholtz D (1995) Development, identification, and characterization of a glyphosate-tolerant soybean line. Crop Sci 35(5):1451–1460CrossRefGoogle Scholar
  5. 5.
    Haslam E, Harborne J (1993) Shikimic acid: metabolism and metabolites. Wiley, New YorkGoogle Scholar
  6. 6.
    Padgette S, Re D, Barry G, Eichholtz D, Delannay X, Fuchs R, Kishore G, Fraley R (1996) New weed control opportunities: development of soybeans with a Roundup Ready gene. Herbicide-resistant crops: agricultural, environmental, economic, regulatory, and technical aspects 53–84Google Scholar
  7. 7.
    Steinrucken H, Amrhein N (1980) The herbicide glyphosate is a potent inhibitor of 5-enolpyruvyl-shikimic acid-3-phosphate synthase. Biochem Biophys Res Commun 94(4):1207CrossRefGoogle Scholar
  8. 8.
    Elenis DS, Kalogianni DP, Glynou K, Ioannou PC, Christopoulos TK (2008) Advances in molecular techniques for the detection and quantification of genetically modified organisms. Anal Bioanal Chem 392(3):347–354CrossRefGoogle Scholar
  9. 9.
    Morisset D, Stebih D, Cankar K, Zel J, Gruden K (2008) Alternative DNA amplification methods to PCR and their application in GMO detection: a review. Eur Food Res Technol 227(5):1287–1297CrossRefGoogle Scholar
  10. 10.
    Shrestha HK, Hwu K-K, Chang M-C (2010) Advances in detection of genetically engineered crops by multiplex polymerase chain reaction methods. Trends Food Sci Technol 21(9):442–454CrossRefGoogle Scholar
  11. 11.
    Ahmed FE (2002) Detection of genetically modified organisms in foods. Trends Biotechnol 20(5):215–223CrossRefGoogle Scholar
  12. 12.
    Brett GM, Chambers SJ, Huang L, Morgan MRA (1999) Design and development of immunoassays for detection of proteins. Food Control 10(6):401–406CrossRefGoogle Scholar
  13. 13.
    Meyer R (1999) Development and application of DNA analytical methods for the detection of GMOs in food. Food Control 10(6):391–399CrossRefGoogle Scholar
  14. 14.
    Miraglia M, Berdal KG, Brera C, Corbisier P, Holst-Jensen A, Kok EJ, Marvin HJP, Schimmel H, Rentsch J, van Rie JPPF, Zagon J (2004) Detection and traceability of genetically modified organisms in the food production chain. Food Chem Toxicol 42(7):1157–1180CrossRefGoogle Scholar
  15. 15.
    Rogan GJ, Dudin YA, Lee TC, Magin KM, Astwood JD, Bhakta NS, Leach JN, Sanders PR, Fuchs RL (1999) Immunodiagnostic methods for detection of 5-enolpyruvylshikimate-3-phosphate synthase in Roundup Ready® soybeans. Food Control 10(6):407–414CrossRefGoogle Scholar
  16. 16.
    Vaïtilingom M, Pijnenburg H, Gendre F, Brignon P (1999) Real-time quantitative PCR detection of genetically modified maximizer maize and roundup ready soybean in some representative foods. J Agric Food Chem 47(12):5261–5266CrossRefGoogle Scholar
  17. 17.
    Germini A, Mezzelani A, Lesignoli F, Corradini R, Marchelli R, Bordoni R, Consolandi C, De Bellis G (2004) Detection of genetically modified soybean using Peptide Nucleic Acids (PNAs) and microarray technology. J Agric Food Chem 52(14):4535–4540CrossRefGoogle Scholar
  18. 18.
    Feriotto G, Borgatti M, Mischiati C, Bianchi N, Gambari R (2002) Biosensor technology and surface plasmon resonance for real-time detection of genetically modified roundup ready soybean gene sequences. J Agric Food Chem 50(5):955–962CrossRefGoogle Scholar
  19. 19.
    Gryson N (2010) Effect of food processing on plant DNA degradation and PCR-based GMO analysis: a review. Anal Bioanal Chem 396(6):2003–2022CrossRefGoogle Scholar
  20. 20.
    Stave JW (1999) Detection of new or modified proteins in novel foods derived from GMO—future needs. Food Control 10(6):367–374CrossRefGoogle Scholar
  21. 21.
    García-Cañas V, Simó C, León C, Ibáñez E, Cifuentes A (2011) MS-based analytical methodologies to characterize genetically modified crops. Mass Spectrom Rev 30(3):396–416CrossRefGoogle Scholar
  22. 22.
    Ocaña MF, Fraser PD, Patel RKP, Halket JM, Bramley PM (2007) Mass spectrometric detection of CP4 EPSPS in genetically modified soya and maize. Rapid Commun Mass Spectrom 21(3):319–328CrossRefGoogle Scholar
  23. 23.
    Ocaña MF, Fraser PD, Patel RKP, Halket JM, Bramley PM (2009) Evaluation of stable isotope labelling strategies for the quantitation of CP4 EPSPS in genetically modified soya. Anal Chim Acta 634(1):75–82CrossRefGoogle Scholar
  24. 24.
    Zhang Y, Lai C, Su R, Zhang M, Xiong Y, Qing H, Deng Y (2012) Quantification of Cry1Ab in genetically modified maize leaves by liquid chromatography multiple reaction monitoring tandem mass spectrometry using 18O stable isotope dilution. Analyst 137(11):2699–2705CrossRefGoogle Scholar
  25. 25.
    Hsu JL, Huang SY, Chow NH, Chen SH (2003) Stable-isotope dimethyl labeling for quantitative proteomics. Anal Chem 75(24):6843–6852CrossRefGoogle Scholar
  26. 26.
    Shevchenko A, Wilm M, Vorm O, Mann M (1996) Mass spectrometric sequencing of proteins from silver-stained polyacrylamide gels. Anal Chem 68(5):850–858CrossRefGoogle Scholar
  27. 27.
    Helmke SM, Yen C-Y, Cios KJ, Nunley K, Bristow MR, Duncan MW, Perryman MB (2004) Simultaneous quantification of human cardiac α- and β-Myosin heavy chain proteins by MALDI-TOF mass spectrometry. Anal Chem 76(6):1683–1689CrossRefGoogle Scholar
  28. 28.
    Ji C, Li L (2005) Quantitative proteome analysis using differential stable isotopic labeling and microbore LC−MALDI MS and MS/MS. J Proteome Res 4(3):734–742CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • Po-Chih Chang
    • 1
  • P. Muralidhar Reddy
    • 2
  • Yen-Peng Ho
    • 1
    Email author
  1. 1.Department of ChemistryNational Dong Hwa UniversityHualienRepublic of China
  2. 2.Department of Chemistry, Nizam CollegeOsmania UniversityHyderabadIndia

Personalised recommendations