Quantification of key red blood cell folates from subjects with defined MTHFR 677C>T genotypes using stable isotope dilution liquid chromatography/mass spectrometry.
Quantification of key red blood cell folates from subjects with defined MTHFR 677C>T genotypes using stable isotope dilution liquid chromatography/mass spectrometry.
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DOI：
10.1002/rcm.3624
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发表时间：
2008-08
影响因子：
2
通讯作者：
Blair, Ian A.
中科院分区：
文献类型：
--
作者：
Huang, Yuehua;Khartulyari, Stefanie;Morales, Megan E.;Stanislawska-Sachadyn, Anna;Von Feldt, Joan M.;Whitehead, Alexander S.;Blair, Ian A.
红细胞(RBC)叶酸水平是在红细胞生成时建立的，因此为个体在前四个月的平均叶酸状态提供了一个替代生物标志物。叶酸以叶基多谷氨酸的形式存在，这是一种不能从红细胞分泌的高极性分子，必须在分析之前转化为单谷氨酸形式。这是通过在抗坏血酸存在的情况下，在pH为5的全血中裂解红细胞，使用个人血浆翼酰基多谷氨酸水解酶来完成的。在pH 1.5条件下，以[13C5]-5-甲酰基四氢叶酸为催化剂，将甲酰化四氢叶酸衍生物定量转化为稳定的5，10-亚甲基四氢叶酸(5，10-MTHF)形式。然后将得到的[13C5]-5，10-MTHF用作已转化为5，10-MTHF的甲酰化形式的四氢叶酸以及原始样品中存在的任何5，10-MTHF的内标。建立了一种稳定的同位素稀释液相色谱-多反应监测/质谱法，并将其用于RBC叶酸、5-甲基四氢叶酸(5-MTHF)、四氢叶酸(THF)和5，10-MTHF的准确定量。该方法灵敏、可靠，可用于检测不同亚甲基四氢叶酸还原酶(MTHFR)677C和GT；T等位基因与红细胞叶酸表型的关系。可鉴定出四种不同的红细胞叶酸表型。根据单个红细胞叶酸的相对含量，将其分类为I类(5-MTHF和GT；95%；THF和lt；5%；5，10-MTHF和lt；5%)、II类(5-MTHF和lt；95%；THF 5%至20%；5，10-MTHF和Gt；5%)、III类(5-MTHF和GT；55%；THF和GT；20%；5，10-MTHF和GT；5%)和IV类(5-MTHF和lt；55%；THF和GT；20%；5，10-MTHF和GT；5%)。
Red blood cell (RBC) folate levels are established at the time of erythropoiesis and therefore provide a surrogate biomarker for the average folate status of an individual over the preceding four months. Folates are present as folylpolyglutamates, highly polar molecules that cannot be secreted from the RBCs, and must be converted into their monoglutamate forms prior to analysis. This was accomplished using an individual’s plasma pteroylpolyglutamate hydrolase by lysing the RBCs in whole blood at pH 5 in the presence of ascorbic acid. Quantitative conversion of formylated tetrahydrofolate derivatives into the stable 5,10-methenyltetrahydrofolate (5,10-MTHF) form was conducted at pH 1.5 in the presence of [13C5]-5-formyltetrahydrofolate. The resulting [13C5]-5,10-MTHF was then used as an internal standard for the formylated forms of tetrahydrofolate that had been converted into 5,10-MTHF as well any 5,10-MTHF that had been present in the original sample. A stable isotope dilution liquid chromatography-multiple reaction monitoring/mass spectrometry method was validated and then used for the accurate and precise quantification of RBC folic acid, 5-methyltetrahydrofolate (5-MTHF), tetrahydrofolate (THF), and 5,10-MTHF. The method was sensitive and robust and was used to assess the relationship between different methylenetetrahydrofolate reductase (MTHFR) 677C>T genotypes and RBC folate phenotypes. Four distinct RBC folate phenotypes could be identified. These were classified according to the relative amounts of individual RBC folates as type I (5-MTHF >95%; THF <5%; 5,10-MTHF <5%), type II (5-MTHF <95%; THF 5% to 20%; 5,10-MTHF <5%), type III (5-MTHF >55%; THF >20%; 5,10-MTHF >5%), and type IV (5-MTHF <55%; THF >20%; 5,10-MTHF >5%).
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影响因子：
168.9
作者：
Molloy, AM;Daly, S;Scott, JM
通讯作者：
Scott, JM
影响因子：
4.2
作者：
BAILEY, LB
通讯作者：
BAILEY, LB
影响因子：
9.3
作者：
Fazili, Zia;Pfeiffer, Christine M.;Koontz, Deborah
通讯作者：
Koontz, Deborah
影响因子：
1.8
作者：
LUCOCK, MD;HARTLEY, R;SMITHELLS, RW
通讯作者：
SMITHELLS, RW
影响因子：
2.9
作者：
Pejchal, R;Campbell, E;Ludwig, ML
通讯作者：
Ludwig, ML