Sensitive LC MS quantitative analysis of carbohydrates by Cs+ attachment

  • Eduard Rogatsky
  • Harsha Jayatillake
  • Gayotri Goswami
  • Vlad Tomuta
  • Daniel Stein


The development of a sensitive assay for the quantitative analysis of carbohydrates from human plasma using LC/MS/MS is described in this paper. After sample preparation, carbohydrates were cationized by Cs+ after their separation by normal phase liquid chromatography on an amino based column. Cesium is capable of forming a quasi-molecular ion [M + Cs]+ with neutral carbohydrate molecules in the positive ion mode of electrospray ionization mass spectrometry. The mass spectrometer was operated in multiple reaction monitoring mode, and transitions [M + 133]→133 were monitored (M, carbohydrate molecular weight). The new method is robust, highly sensitive, rapid, and does not require postcolumn addition or derivatization. It is useful in clinical research for measurement of carbohydrate molecules by isotope dilution assay.


  1. 1.
    Tserng, K.-Y.; Kalhan, S. C. Estimation of glucose carbon recycling and glucose turnover with [U-13C] glucose. Am. J. Physiol 1983, 245, E476-E482.Google Scholar
  2. 2.
    Wolfe, R. R. Radioactive and stable isotope tracers in biomedicine: Principles and practice of kinetic analysis; Wiley-Liss: New York, 1992, pp. 424–426.Google Scholar
  3. 3.
    Beylot, M.; Martin, C.; Beaufrere, B.; Riou, J. P.; Mornex, R. Determination of steady state and nonsteady state glycerol kinetics in humans using deuterium-labeled tracer. J. Lipid Res 1987, 28, 414–422.Google Scholar
  4. 4.
    Esteban, N. V.; Liberato, D. J.; Sidbury, J. B.; Yergey, A. L. Stable isotope dilution thermospray liquid chromatography/mass spectrometry method for determination of sugars and sugar alcohols in humans. Anal. Chem 1987, 59, 1674–1677.CrossRefGoogle Scholar
  5. 5.
    Reid, S.; Shackleton, C.; Wu, K.; Kaempfer, S.; Hellerstein, M. K. Liquid chromatography/mass spectrometry of plasma glucose and secreted glucuronate for metabolic studies in humans. Biomed. Environ. Mass Spectrom 1990, 19, 535–540.CrossRefGoogle Scholar
  6. 6.
    Takatsu, A.; Nishi, S. Stable isotope dilution method for the determination of serum glucose using discharge-assisted thermospray liquid chromatography/mass spectrometry. Biol. Mass Spectrom 1991, 20, 415–418.CrossRefGoogle Scholar
  7. 7.
    Magni, F.; Arnoldi, L.; Monti, L.; Piatti, P.; Pozza, G.; Galli, K. M. Determination of plasma glycerol isotopic enrichment by gas chromatography-mass spectrometry: An alternative glycerol derivative. Anal. Biochem 1993, 211, 327–328.CrossRefGoogle Scholar
  8. 8.
    Gilker, C. D.; Pesola, G. R.; Matthews, D. E. A mass spectrometric method for measuring glycerol levels and enrichments in plasma using 13C and 2H stable isotopic tracers. Anal. Biochem 1992, 205, 172–178.CrossRefGoogle Scholar
  9. 9.
    Ackermans, M. T.; Ruiter, A. F.; Endert, E. Determination of glycerol concentrations and glycerol isotopic enrichments in human plasma by gas chromatography/mass spectrometry. Anal. Biochem 1998, 258, 80–86.CrossRefGoogle Scholar
  10. 10.
    Flakoll, P. J.; Zheng, M.; Vaughan, S.; Borel, M. J. Determination of stable isotopic enrichment and concentration of glycerol in plasma via gas chromatography-mass spectrometry for the estimation of lipolysis in vivo. J. Chromatogr. B 2000, 744, 47–54.CrossRefGoogle Scholar
  11. 11.
    McIntosh, T. S.; Davis, H. M.; Matthews, D. E. A liquid chromatography-mass spectrometry method to measure stable isotopic tracer enrichments of glycerol and glucose in human serum. Anal. Biochem 2002, 300, 163–169.CrossRefGoogle Scholar
  12. 12.
    Schoemaker, R. C.; Burggraaf, J.; Cohen, A. F. Assessment of hepatic blood flow using continuous infusion of high clearance drugs. Br. J. Clin. Pharmacol 2000, 45, 463–469.CrossRefGoogle Scholar
  13. 13.
    Matthews, D. E.; Bier, D. M. Stable isotope methods for nutritional investigation. Annu. Rev. Nutr 1983, 3, 309–339.CrossRefGoogle Scholar
  14. 14.
    Kalhan, S. C. Stable isotope tracers for studies of glucose metabolism. J. Lab. Clin. Med 1990, 116, 615–622.Google Scholar
  15. 15.
    Tserng, K. Y.; Kalhan, S. C. Estimation of glucose carbon recycling and glucose turnover with [U-13C] glucose. Am. J. Physiol 1983, 245, E76-E82.Google Scholar
  16. 16.
    Tetsuo, M.; Zhang, C.; Matsumoto, H.; Matsumoto, I. Gas chromatographic-mass spectrometric analysis of urinary sugar and sugar alcohols during pregnancy. J. Chromatogr. B 1999, 731, 111–120.CrossRefGoogle Scholar
  17. 17.
    Kato, Y.; Numajiri, Y. Chloride attachment negative-ion mass spectra of sugars by combined liquid chromatography and atmospheric pressure chemical ionization mass spectrometry. J. Chromatogr 1991, 562, 81–97.CrossRefGoogle Scholar
  18. 18.
    Tannenbaum, H. P.; Roberts, J. D.; Dougherty, R. C. Negative chemical ionization mass spectrometry—chloride attachment spectra. Anal. Chem 1975, 47, 49–54.CrossRefGoogle Scholar
  19. 19.
    Kumaguai, H. Application of liquid chromatography/mass spectrometry to the analysis of sugars and sugar-alcohol; Agilent Technologies: Application Note no. 5988-4236EN, 10/2001.Google Scholar
  20. 20.
    Staneke, P. O.; Groothuis, G.; Ingemann, S.; Nibbering, N. M. M. Formation, stability, and structure of radical anions of chloroform, tetrachloromethane, and fluorotrichloromethane in the gas phase. Int. J. Mass Spectrom. Ion Processes 1995, 142, 83–93.CrossRefGoogle Scholar
  21. 21.
    Kusakavitch, M. Investigation of discoloration of chloroform solution. Poster 370 at the Eastern Analytical Symposium; Somerset, NJ, November, 2004.Google Scholar
  22. 22.
    Liang, H.; Foltz, R.; Bennett, P. Sensitive and selective LC/MS/MS method for determination of endogenous polyols in human nerve tissues. Poster ThPK 220 at the 52nd Conference of American Society of Mass Spectrometry; Nashville, TN, June, 2004.Google Scholar
  23. 23.
    Zhu, J.; Cole, R. Ranking of gas-phase acidities and chloride affinities of monosaccharides and linkage specificity in collision-induced decompositions of negative ion electrospray-generated chloride adducts of oligosaccharides. J. Am. Soc. Mass Spectrom 2001, 112, 1193–1204.CrossRefGoogle Scholar
  24. 24.
    Antonopoulos, A.; Bonnet, P.; Botek, E.; Debrun, J. L.; Hakim, B.; Herbreteau, B.; Morin-Allory, L. Study of the attachment of Na+ on glucose and on some of its methylated derivatives. Rapid Commun. Mass Spectrom 2003, 17, 122–125.CrossRefGoogle Scholar
  25. 25.
    Botek, E.; Debrun, J. L.; Hakim, B.; Morin-Alory, L. Attachment of alkali cations on β-D-glucopyranose: Matrix-assisted laser desorption/ionization time of flight studies and ab initio calculations. Rapid Commun. Mass Spectrom 2001, 15, 273–276.CrossRefGoogle Scholar
  26. 26.
    Ngoka, L. C.; Gal, J.-F.; Lebrilla, C. B. Effects of cations and charge types on the meta-stable decay rates of oligosaccharides. Anal. Chem 1994, 66, 692–698.CrossRefGoogle Scholar
  27. 27.
    Kohler, M.; Leary, J. LC/MS/MS of carbohydrates with postcolumn addition of metal chlorides using triaxial electrospray probe. Anal. Chem 1995, 67, 3501–3508.CrossRefGoogle Scholar
  28. 28.
    Rogatsky, E.; Stein, D. T. A novel, highly robust method of carbohydrate prepurification by 2D LC prior to LC/MS or GC/MS. J. Chromatogr. A 2005, 1073, 11–16.CrossRefGoogle Scholar
  29. 29.
    Young, C. S.; Dolan, J. W. Success with evaporative light scattering detection, Part II: Tips and techniques. LC/GC North Am 2004, 22, 244–250.Google Scholar
  30. 30.
    Schoor, A.; Erdmann, N.; Effmert, U.; Mikkat, S. Determination of the cyanobacterial osmolyte glucosylglycerol by high-performance liquid chromatography. J. Chromatogr. A 1995, 704, 89–97.CrossRefGoogle Scholar
  31. 31.
    Tseng, K.; Lindsay, L. L.; Penn, S.; Hedrick, J. L.; Lebrilla, C. B. Characterization of neutral oligosaccharide-alditols from Xenopus laevis egg jelly coats by matrix-assisted laser desorption Fourier transform mass spectrometry. Anal. Biochem 1997, 250, 18–28.CrossRefGoogle Scholar
  32. 32.
    Hager, J. W. Recent trends in mass spectrometer development. Anal. Bioanal. Chem 2004, 378, 845–850.CrossRefGoogle Scholar
  33. 33.
    Bruggink, C.; Maurer, R.; Herrmann, H.; Cavalli, S.; Hoefler, F. Analysis of carbohydrates by anion exchange chromatography and mass spectrometry. J. Chromatogr. A 2005, 1085, 104–109.CrossRefGoogle Scholar
  34. 34.
    Guignard, C.; Jouve, L.; Bogeat-Triboulot, M. B.; Dreyer, E.; Hausman, J.-F.; Hoffmann, L. Analysis of carbohydrates in plants by high-performance anion-exchange chromatography coupled with electrospray mass spectrometry. J. Chromatogr. A 2005, 1085, 137–142.CrossRefGoogle Scholar

Copyright information

© American Society for Mass Spectrometry 2005

Authors and Affiliations

  • Eduard Rogatsky
    • 1
  • Harsha Jayatillake
    • 1
  • Gayotri Goswami
    • 1
  • Vlad Tomuta
    • 1
  • Daniel Stein
    • 1
  1. 1.General Clinical Research CenterAlbert Einstein College of MedicineBronxUSA

Personalised recommendations