A COMPREHENSIVE PROCEDURE FOR PREPARATION OF PARTIALLY METHYLATED ALDITOL ACETATES FROM GLYCOPROTEIN CARBOHYDRATES
A COMPREHENSIVE PROCEDURE FOR PREPARATION OF PARTIALLY METHYLATED ALDITOL ACETATES FROM GLYCOPROTEIN CARBOHYDRATES
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DOI：
10.1016/0003-2697(92)90048-c
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发表时间：
1992-05-15
影响因子：
2.9
通讯作者：
TAYLOR, PB
中科院分区：
文献类型：
--
作者：
ANUMULA, KR;TAYLOR, PB
改进了从糖蛋白衍生的碳水化合物制备部分甲基化阿迪醇乙酸酯(PMAA)的各种步骤，以快速、高产率地获得这些衍生物。碳水化合物在二甲基亚砜(DMSO)中全甲基化，使用氢氧化钠和甲基碘(CH3I)的精细悬浮液。以市售50%氢氧化钠水溶液为原料，采用超声破碎、二甲基亚砜洗涤的方法，方便地制备了氢氧化钠的微细悬浮液。利用NaOH和CH3I的精细悬浮液，卵清蛋白和胎球蛋白糖肽的甲基化反应在5分钟内完成，并且甲基化反应没有产生任何非糖制品。未经任何纯化的甲基化碳水化合物在挥发性有机酸的混合物中被水解，这使得样品中的酸可以通过蒸发快速去除。以4-N，N‘-二甲氨基吡啶为催化剂，将部分甲基化糖醇与乙酸酐在室温下进行乙酰化反应2~4h，反应不产生非糖反应伪影。确定了甲基化、水解和乙酰化反应的时间进程，以获得制备PMAAs的最佳反应条件。该程序有助于快速鉴定和定量PMAAs，因为减少了反应伪影，并且色谱图的质量仅取决于原料和用于甲基化分析的试剂的纯度。这些用于快速甲基化分析的简单方法在使用碳水化合物分析仪对少量分离的低聚糖进行表征中得到了证明。
Various steps involved in the preparation of partially methylated alditol acetates (PMAAs) from glycoprotein-derived carbohydrates were improved to obtain the derivatives in a rapid manner with excellent yields. Carbohydrates were permethylated in dimethyl sulfoxide (DMSO), using a fine suspension of sodium hydroxide and methyl iodide (CH3I). The fine suspension of NaOH was prepared conveniently from commercially available 50% aqueous NaOH in DMSO by sonication and washing the precipitate with DMSO. Methylation of ovalbumin and fetuin glycopeptides using the fine suspension of NaOH and CH3I was complete within 5 min, and the methylation reaction did not generate any nonsugar artifacts. Methylated carbohydrates without any purification were hydrolyzed in a mixture of volatile organic acids, which permitted rapid removal of the acids from samples by evaporation. Acetylation of partially methylated alditols with acetic anhydride for 2–4 h at ambient temperature using 4-N,N′-dimethylaminopyridine as a catalyst and the reaction was free from generating nonsugar reaction artifacts. The reaction time course for methylation, hydrolysis, and acetylation was determined to obtain optimum reaction conditions for preparation of the PMAAs. The procedure facilitated rapid identification and quantitation of PMAAs due to diminished reaction artifacts and the quality of the chromatogram depended only on the purity of starting material and the reagents used for the methylation analysis. Utility of these simple methods for rapid methylation analysis was demonstrated in the characterization of oligosaccharides isolated in small amounts using a carbohydrate analyzer.