Capillary electrophoresis analysis of glucooligosaccharide regioisomers.
Complex gluco-oligosaccharide mixtures of two regioisomer series were successfully separated by CE. The gluco-oligosaccharide series were synthesized, employing a dextransucrase from Leuconostoc mesenteroides NRRL B-512F, by successive glucopyranosyl transfers from sucrose to the acceptor glucose or maltose. The glucosyl transfer to both acceptors, occurring through the formation of alpha1-->6 linkages, differed for the two series only in the glucosidic bond to the reducing end namely alpha1-->6 or alpha1-->4 bond for glucose or maltose acceptor, respectively. Thus, the combination of the two series results in mixed pairs of gluco-oligosaccharide regioisomers with different degrees of polymerization (DP). These regioisomer series were first derivatized by reductive amination with 9-aminopyrene-1,4,6-trisulfonate (APTS). Under acidic conditions using triethyl ammonium acetate as electrolyte, the APTS-gluco-oligosaccharides of each series were separated enabling unambiguous size determination by coupling CE to electrospray-mass spectrometry. However, neither these acidic conditions nor alkaline buffer systems could be adapted for the separation of the gluco-oligosaccharide regioisomers arising from the two combined series. By contrast, increased resolution was observed in an alkaline borate buffer, using differential complexation of the regioisomers with the borate anions. Such conditions were also successfully applied to the separation of glucodisaccharide regioisomers composed of alpha1-->2, alpha1-->3, alpha1-->4, and alpha1-->6 linkages commonly synthesized by glucansucrase enzymes.
Département de Génie Biochimique et Alimentaire, Centre de Bioingénierie, Gilbert Durand, Toulouse, France.
Gilles
G
Joucla
Thérèse
T
Brando
Magali
M
Remaud-Simeon
Pierre
P
Monsan
Germain
G
Puzo
D000586:Amination; D002240:Carbohydrate Sequence; D019075:Electrophoresis, Capillary / Q000379:methods*; D005947:Glucose / Q000737:chemistry*; D005964:Glucosyltransferases / Q000737:chemistry / Q000378:metabolism*; D007536:Isomerism; D007934:Leuconostoc / Q000201:enzymology; D008320:Maltose / Q000737:chemistry; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000138:chemical synthesis / Q000737:chemistry / Q000302:isolation & purification*; D011721:Pyrenes / Q000737:chemistry; D013395:Sucrose / Q000737:chemistry; D013451:Sulfonic Acids / Q000737:chemistry
C492698:9-aminopyrene-1,4,6-trisulfonic acid; D009844:Oligosaccharides; D011721:Pyrenes; D013451:Sulfonic Acids; D013395:Sucrose; D008320:Maltose; D005964:Glucosyltransferases; C020025:dextransucrase; D005947:Glucose