Peterson K P, Pavlovich J G, Goldstein D, Little R, England J, Peterson C M
Sansum Medical Research Foundation, Santa Barbara, CA 93111, USA.
Clin Chem. 1998 Sep;44(9):1951-8.
Hemoglobin A1c (HbA1c) is a stable minor Hb variant formed in vivo by posttranslational modification by glucose, originally identified by using cation exchange chromatography, and containing primarily glycated N-terminal beta-chains. However, the structure(s) of the quantified species has not been elucidated, and the available methods lack a reference standard. We used electrospray ionization mass spectrometry to determine the extent of glycation of samples separated by boronate affinity and/or cation exchange chromatography. Analyses of clinical samples were consistent with the curvilinear relationship of patient glucose and HbA1c. As glycation increased, the ratio of beta-chain to alpha-chain glycation increased, and the number of glycation sites on the beta-chain increased, although these were relatively minor components. We found several glycated species that cochromatographed with HbA1c on cation exchange, including species with both glycated alpha- and beta-chains, nonglycated alpha- and glycated beta-chains, and multiply glycated beta-chains. The combined use of affinity and cation exchange chromatography with structural confirmation by electrospray ionization mass spectrometry was found to be useful in producing samples of sufficient purity for the standardization of glycohemoglobin clinical assays.
糖化血红蛋白A1c（HbA1c）是一种在体内由葡萄糖进行翻译后修饰形成的稳定的次要血红蛋白变体，最初通过阳离子交换色谱法鉴定，主要包含糖化的N端β链。然而，定量物质的结构尚未阐明，现有方法缺乏参考标准。我们使用电喷雾电离质谱法来确定通过硼酸亲和色谱法和/或阳离子交换色谱法分离的样品的糖化程度。临床样品分析与患者葡萄糖和HbA1c的曲线关系一致。随着糖化增加，β链与α链糖化的比例增加，β链上的糖化位点数量增加，尽管这些是相对次要的成分。我们发现了几种在阳离子交换色谱上与HbA1c共色谱的糖化物质，包括同时具有糖化α链和β链的物质、未糖化α链和糖化β链的物质以及多重糖化β链的物质。通过电喷雾电离质谱法进行结构确认，发现亲和色谱法和阳离子交换色谱法联合使用有助于制备纯度足够高的样品，用于糖化血红蛋白临床检测的标准化。
