COMPOSITION ANALYSIS BY GC-MS

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 样品转移到螺帽管中，加入2g肌醇作为内标，冷冻干燥。样品在80℃下用1M盐酸在甲醇中甲醇裂解过夜，生成甲基糖苷。在水解后，样品在氮气气流下完全干燥。甲基糖苷在甲醇中与吡啶和乙酸酐在室温下重新N-乙酰化至少30min(用于检测氨基糖)。然后，样品用Tri-Sill试剂(Thermo Science)在80℃下进行全O-三甲基硅烷化(TMS)20分钟。这些程序如前所述在Merkle和Poppe(1994)的方法Enzyol中进行。230：1-15；York等人。(1985)方法Enzyol。118：3-40。TMS甲基糖苷的分析在Hewlett Packard Series II 5890气相色谱仪上进行，配备Supelco EC-1熔融石英毛细管柱(30m×0.25 mm内径)，并连接到Hewlett Packard 5970MSD。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The samples were transferred into screw-cap tubes, added with 2 ¿g inositol as internal standard, and lyophilized. The samples were subjected to methanolysis with 1 M HCl in methanol at 80¿C overnight to form methyl glycosides. After hydrolysis, the samples were dried completely under a stream of nitrogen gas. The methyl glycosides were re-N-acetylated with pyridine and acetic anhydride in methanol (for detection of amino sugars) at room temperature for at least 30 min. The samples then were per-O-trimethylsilylated (TMS) with a Tri-Sil reagent (Thermo Scientific) at 80¿C for 20 min. These procedures were carried out as described previously in Merkle and Poppe (1994) Methods Enzymol. 230: 1-15; York, et al. (1985) Methods Enzymol. 118:3-40. Analysis of the TMS methyl glycosides was performed on a Hewlett Packard Series II 5890 gas chromatograph equipped with a Supelco EC-1 fused silica capillary column (30m ¿ 0.25 mm ID) and interfaced to a Hewlett Packard 5970 MSD.