Abstract:
:An ultrasensitive method for analysis of polysialic acid (polySia) chains, using fluorescence-assisted high-performance liquid chromatography was developed. The new method is a substantial improvement of our earlier method in which the reducing terminal Sia residues of a homologous series of oligo/polySia hydrolytically released during derivatization reaction were simultaneously labeled with a fluorogenic reagent, 1,2-diamino-4,5-methylenedioxybenzene (DMB) in situ. We first studied extensively the stability of oligo/polySia in the acid (0.02 M trifluoracetic acid) used for 1,2-diamino-4,5-methylenedioxybenzene derivatization under various conditions of reaction time and temperature, analyzing the hydrolytic products by high-performance anion exchange chromatography with pulsed electrochemical detection (HPAEC-PED). Then we optimized the reaction conditions to minimize degradation of the parent polySia while maintaining high derivatization rate. Using a DNAPac PA-100 column rather than a MonoQ column, baseline resolution of polySia peaks up to DP 90 with a detection threshold of 1.4 femtomol per resolved peak was achieved. The new method was used to analyze the degree of polymerization of a polySia-containing glycopeptide fraction derived from embryonic chicken brain, and the results were compared with those obtained by HPAEC-PED.
journal_name
Glycobiologyjournal_title
Glycobiologyauthors
Inoue S,Lin SL,Lee YC,Inoue Ydoi
10.1093/glycob/11.9.759subject
Has Abstractpub_date
2001-09-01 00:00:00pages
759-67issue
9eissn
0959-6658issn
1460-2423journal_volume
11pub_type
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