期刊
JOURNAL OF CHROMATOGRAPHY A
卷 1216, 期 9, 页码 1520-1523出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.chroma.2009.01.015
关键词
Neutral oligosaccharides; Anthranilic acid; Derivatization; HPLC; Lacto-N-fucopentaose III; 2 '-Fucosyllactose
An improved analytical scheme for human milk neutral oligosaccharides determination was developed, in which, the oligosaccharides were pooled in two fractions (pools 1 and 2) after gel filtration, and then were quantitatively derivatized with a single fluorescent reagent, 2-anthranilic acid. Separation was by reversed-phase HPLC on an ODS-100Z column with a mobile phase of 50 mM ammonium acetate pH 4.0 and 150 mM citrate buffer pH 4.5 and monitored by a fluorescence detector at 360 nm excitation and 425 nm emission wavelengths. The method improved on the separation of neutral tetra- and hexasaccharide isomers, namely, lacto-N-tetraose (LNT) and lacto-N-neotetraose (LNnT) as well as of lacto-N-difucohexaose I (LNDFH I) and lacto-N-difucohexaose II (LNDFH II). The separation of trisacccharide isomers, 3-fucosyllactose (3-FL) and 2'-fucosyllactose (2'-FL) was also successful. Limits of detection and quantification were in the range of 1-10 ng/l and 2-30 ng/l, respectively. The methods' accuracy was good with its precision at <20% RSD and <1% RSD, respectively, for oligosaccharide concentration and retention time. The recoveries were in the range of 80-100%. This method was successfully applied to the separation and determination of representative neutral oligosaccharide contents in Samoa women milk. (C) 2009 Elsevier B.V. All rights reserved.
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