Characterizing the electrospray-ionization mass spectral fragmentation pattern of enzymatically derived hyaluronic acid oligomers.
Oligosaccharides derived from hyaluronic acid by action of bovine testicular hyaluronidase (BTH) (hyaluronate 4-glycanohydrolase, E.C. 3.2.1.35) were characterized by mass spectrometry (MS) with electrospray-ionization mass spectrometry (ESIMS) and compared with results obtained by matrix-assisted laser desorption/ionization. Both oligomers with an odd number and even number of sugar units with molecular masses up to 8 kDa were observed in the ESI spectra. However, the generation of odd-numbered oligomers is not consistent with the regiospecificity of the enzyme and with the MALDI results, which indicated even-numbered oligomers exclusively. In addition, a third method of characterization, high-performance anion-exchange chromatography (HPAEC), showed only even-numbered oligomers. Relative intensities of the odd-numbered oligomers demonstrated in ESIMS a cone-voltage dependence suggesting the odd-numbered oligomers resulted from collisional activation. In order to achieve results by ESI that mirror results from other techniques, the cone voltage must be kept low and precisely controlled. This study displays the usefulness and possible vulnerabilities of ESIMS when utilized for carbohydrate analysis without corroborating data from other methods of analysis.
Prebyl
Benjamin S
BS
Department of Chemistry and the Center of Excellence for Structural Biology, The University of Tennessee, 667 Buchler Hall, Knoxville, TN 37996-1600, USA.
Kaczmarek
Conrad
C
Tuinman
Albert A
AA
Baker
David C
DC
D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D006820:Hyaluronic Acid / Q000737:chemistry* / Q000378:metabolism; D006821:Hyaluronoglucosaminidase / Q000378:metabolism; D008297:Male; D009844:Oligosaccharides / Q000032:analysis*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D009844:Oligosaccharides; D006820:Hyaluronic Acid; D006821:Hyaluronoglucosaminidase