Isotopomer analysis of lipid biosynthesis by high resolution mass spectrometry and NMR.
Isotopomer analysis of lipid biosynthesis by high resolution mass spectrometry and NMR.
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DOI：
10.1016/j.aca.2009.08.032
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发表时间：
2009-10-05
影响因子：
6.2
通讯作者：
Higashi RM
中科院分区：
文献类型：
--
作者：
Lane AN;Fan TW;Xie Z;Moseley HN;Higashi RM
我们将2D-核磁共振和输注FT-ICR-MS与计算机辅助分配相结合，直接在粗细胞提取物中分析甘油磷脂(GPL)的13C同位素，导致在几分钟内产生非常高的>；3000等压分子的信息吞吐量。要将同分异构体与其他GPL结构区分开来，需要优于1ppm的质量精确度和在测量m/z时的100,000分辨率。从生长在[U-13C]-葡萄糖上的LCC2乳腺癌细胞中提取的GPLs的同源分析提供了关于GPLs的生物合成和转化的丰富信息。从FT-ICR-MS得到的同位素强度比在自然丰度背景下的准确度为≈1%或更高，并取决于信鼻子比。详细分析了来自[U-13C]-葡萄糖的13C掺入特定磷脂酰胆碱的时间过程，以提供生长中的LCC2细胞中甘油、乙酰辅酶A和总GPL池大小的定量测量。对从高分辨率2D核磁共振获得的甘油和脂肪酰链中13C的位置浓缩进行了独立和互补的分析，以验证模型的关键方面。这项技术能够简单快速地制备样品，具有快速分析，一般适用于几乎任何头基的普通GPL，以及其他类别代谢物的混合物。
We have coupled 2D-NMR and infusion FT-ICR-MS with computer-assisted assignment to profile 13C-isotopologues of glycerophospholipids (GPL) directly in crude cell extracts, resulting in very high information throughput of >3000 isobaric molecules in a few minutes. A mass accuracy of better than 1 ppm combined with a resolution of 100,000 at the measured m/z was required to distinguish isotopomers from other GPL structures. Isotopologue analysis of GPLs extracted from LCC2 breast cancer cells grown on [U-13C]-glucose provided a rich trove of information about the biosynthesis and turnover of the GPLs. The isotopologue intensity ratios from the FT-ICR-MS were accurate to ≈ 1% or better based on natural abundance background, and depended on the signal-to-nose ratio. The time course of incorporation of 13C from [U-13C]-glucose into a particular phosphatidylcholine was analyzed in detail, to provide a quantitative measure of the sizes of glycerol, acetyl CoA and total GPL pools in growing LCC2 cells. Independent and complementary analysis of the positional 13C enrichment in the glycerol and fatty acyl chains obtained from high resolution 2D NMR was used to verify key aspects of the model. This technology enables simple and rapid sample preparation, has rapid analysis, and is generally applicable to unfractionated GPLs of almost any head group, and to mixtures of other classes of metabolites.
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影响因子：
2.9
作者：
Neese, RA;Siler, SQ;Hellerstein, MK
通讯作者：
Hellerstein, MK
DOI：
10.1016/s1096-4959(02)00006-4
发表时间：
2002-03-01
影响因子：
2.2
作者：
Carrasco, MP;Jiménez-López, JM;Marco, C
通讯作者：
Marco, C
影响因子：
11.2
作者：
Glunde, K;Jie, C;Bhujwalla, ZM
通讯作者：
Bhujwalla, ZM
影响因子：
3.6
作者：
Fan, Teresa W. M.;Bandura, Laura L.;Lane, Andrew N.
通讯作者：
Lane, Andrew N.
影响因子：
6.5
作者：
Milne, SB;Ivanova, PT;Brown, HA
通讯作者：
Brown, HA