Pfeiffer C M, Gregory J F
Food Science and Human Nutrition Department, University of Florida, Gainesville 32611, USA.
Clin Chem. 1996 Nov;42(11):1847-54.
Erythrocyte (RBC) folates occur mainly as 5-methyltetrahydrofolate polyglutamates. Determination of RBC folate concentration requires an initial deconjugation of these polyglutamates. In this study, existing HPLC methods were adapted to investigate the rate and extent of this deconjugation process. The action of endogenous plasma pteroyl-polyglutamate hydrolase activity was strongly affected by the conditions of sample preparation, with pH of the incubation mixture more critical to effective deconjugation than incubation time. Dilution of whole blood with 10 g/L ascorbic acid yielded fast hydrolysis of long-chain polyglutamates, and total conversion to 5-methyltetrahydrofolate monoglutamate occurred after 90 min of incubation at 37 degrees C. In contrast, dilution of whole blood with 10 g/L sodium ascorbate, with up to 90 min of incubation at 37 degrees C, yielded a mixture of polyglutamates of 5-methyltetrahydrofolate (glun = 1-8). As documented by direct HPLC analysis and in concurrent assays with Lactobacillus casei, acidification provided by ascorbic acid can have dramatic effects on the measurement of RBC folates.
红细胞（RBC）叶酸主要以5-甲基四氢叶酸多聚谷氨酸盐的形式存在。测定红细胞叶酸浓度需要首先将这些多聚谷氨酸盐解聚。在本研究中，对现有的高效液相色谱（HPLC）方法进行了调整，以研究这种解聚过程的速率和程度。样品制备条件对内源性血浆蝶酰多聚谷氨酸水解酶活性的作用影响很大，孵育混合物的pH值对有效解聚比孵育时间更为关键。用10 g/L抗坏血酸稀释全血可使长链多聚谷氨酸快速水解，在37℃孵育90分钟后可完全转化为5-甲基四氢叶酸单谷氨酸。相比之下，用10 g/L抗坏血酸钠稀释全血，在37℃孵育长达90分钟，会产生5-甲基四氢叶酸多聚谷氨酸（谷氨酰基=1-8）的混合物。如直接HPLC分析和与干酪乳杆菌的同步测定所记录，抗坏血酸提供的酸化对红细胞叶酸的测量可能有显著影响。
