SIALIC ACIDS COMPOSITION ANALYSIS BY HPAEC

通过 HPAEC 进行唾液酸成分分析

• 批准号: 8363084
• 负责人: Parastoo Azadi
• 金额: $ 0.17万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-06-01 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8363084
• 关键词: Acetic Acids; Aliquot; Anions; Calibration; Carbohydrates; Chromatography; Equation; Funding; Gases; Glycoproteins; Grant; Ice; Injection of therapeutic agent; Methods; Mole the mammal; N-Acetylneuraminic Acid; N-glycolylneuraminic acid; National Center for Research Resources; Nitrogen; Phase; Principal Investigator; Protocols documentation; Pump; Research; Research Infrastructure; Resources; Saliva; Sampling; Sialic Acids; Sodium Acetate; Source; System; Technology; Time; United States National Institutes of Health; Vial device; base; cost; detector; programs

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Methods: An aliquot of about 150 ¿g each from NANA and saliva samples were prepared for sialic acids analysis. The aliquots were hydrolyzed with 500 ¿L of 2.0 M acetic acid at 80¿C for 3 h. All hydrolysates were dried under nitrogen gas, resuspended in H2O, sonicated for 7 min in ice and transferred to injection vials. A mix of standards for N-acetylneuraminic acid and N-glycolylneuraminic acid with a known number of moles was hydrolyzed in the same manner and at the same time as the samples. Four concentrations of standard mixture were prepared to establish a calibration equation. The number of moles of each residue in the sample was quantified by linear interpolation from the calibration equation. The sialic acids were analyzed by HPAEC using a Dionex ICS3000 system equipped with a gradient pump, an electrochemical detector, and an autosampler. The sialic acids were separated by a Dionex CarboPac PA20 (3 x 150 mm) analytical column with an amino trap. The gradient programs used the following mobile phase eluents: 100 mM NaOH, and 1M sodium acetate in 100 mM NaOH and injections were made every 40 min. All methods were based on protocols described by Hardy and Townsend (Hardy, M. R., and Townsend, R. R., "High-pH anion-exchange chromatography of glycoprotein-derived carbohydrates", 1994, Methods Enzymol. 230: 208-225).

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 方法： 从NANA和唾液样本中分别制备了约150克的等量，用于唾液酸分析。等价物用500°L和2.0M冰醋酸在80°C下水解3h，水解物在氮气中干燥，在水中悬浮，在冰中超声7min，转移到注射瓶中。 N-乙酰神经氨酸和N-羟基神经氨酸的标准混合物与已知摩尔数的N-乙酰神经氨酸和N-羟基神经氨酸的标准混合物以与样品相同的方式和同时进行水解。制备了四种浓度的标准混合物，建立了校正方程。样品中每个残留物的摩尔数由校正方程中的线性插值法定量。 唾液酸的分析采用Dionex ICS3000系统，该系统配备了梯度泵、电化学检测器和自动进样器。唾液酸采用Dionex CarboPac PA20(3×150 mm)色谱柱，氨基捕捉器。梯度程序使用以下流动相洗脱：100 mM氢氧化钠和1M醋酸钠在100 mM氢氧化钠中，每40分钟进样一次。所有方法都基于Hardy和Townsen(Hardy，M.R.和Townend，R.，“High-pH阴离子交换色层析糖蛋白衍生碳水化合物”，1994，Methods Enzymol)所描述的方案。230：208-225)。