基于硫同位素稀释质谱法的β淀粉样多肽绝对定量研究

Absolute Quantification of Beta Amyloid Peptide Based on Sulfur Isotope Dilution Mass Spectrometry

采用液相色谱-同位素稀释质谱(HPLC-ID-ICPMS)在线联用技术,通过对β淀粉样多肽中硫元素的测定,实现了β淀粉样多肽的准确定量分析。首先通过优化4种液相色谱条件,将β淀粉样多肽纯品中的Aβ42与杂质进行分离,然后将液相洗脱液与34S稀释剂在线混合,针对34S稀释剂对34S/32S信号强度和灵敏度的影响,对34S稀释剂的流速进行了优化,并根据同位素稀释法的公式以及Aβ42中硫元素含量,得到β淀粉样多肽中Aβ42的含量为(0.763±0.004) g/g。本方法在5~60μg/g范围内,线性相关系数为0.999,检出限为140 pg/g,定量限为467 pg/g,目标物的回收率均在98%~105%之间,相对标准偏差均小于6%。本样品采用氨基酸水解法的测量结果为(0.768±0.009) g/g,两种方法的测量结果一致性良好。

The accurate quantification of β-amyloid peptides was achieved by the accurate measurement of sulfur elements in β-amyloid peptides based on high performance liquid chromatography isotope dilution inductively coupled plasmon mass spectrometry(HPLC-ID-ICPMS). Aβ42 in β-amyloid peptide was separated from impurities by optimizing liquid chromatography conditions, and then the eluent was mixed online with enriched 34S diluent. In view of the influence of enriched 34S diluent on 32S/34S signal strength and sensitivity, the flow rate of 34S diluent was optimized. According to the formula of isotope dilution method and the number of S elements in Aβ42, the content of Aβ42 in β amyloid peptide was(0.763±0.0044) g/g. The linear correlation coefficient was 0.999 within the range of 5-60 μg/g, the detection limit was 140 pg/g, and the quantitative limit was 467 pg/g. The recovery of the Aβ42 was between 98% and 105%, and the relative standard deviation was less than 6%. The measurement result of this sample by amino acid hydrolysis method was(0.768±0.0091) g/g, which was in good consistency with the result by this method.