Novel approach for the determination of the redox status of homocysteine and other aminothiols in plasma from healthy subjects and patients with ischemic stroke.
Novel approach for the determination of the redox status of homocysteine and other aminothiols in plasma from healthy subjects and patients with ischemic stroke.
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用于测定健康受试者和缺血性中风患者血浆中同型半胱氨酸和其他氨基硫醇氧化还原状态的新方法。
DOI：
10.1093/clinchem/47.6.1031
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发表时间：
2001
影响因子：
9.3
通讯作者：
C. Helgason
中科院分区：
文献类型：
--
作者：
R. H. Williams;J. Maggiore;R. Reynolds;C. Helgason
背景
血浆“氧化还原”状态可通过测量还原型（r）、游离型（f）、氧化型（ox）和蛋白结合型（B）高半胱氨酸（Hcy）以及相关的氨基硫醇半胱氨酸、半胱氨酰甘氨酸（CysGly）和谷胱甘肽（GSH）来评估，但样本采集较为复杂。缺血性卒中患者的氧化还原状态尚未确定，这可能会增加对其在发病机制中作用的理解。我们希望研究在容易获得的酸性柠檬酸钠中收集的样品中进行这种测量的可行性。
方法
我们使用酸性柠檬酸钠（BioPool Stabilyte，pH 4.3）与EDTA全血测定了氨基硫醇及其在稳定的无蛋白滤液中的稳定性。分析前，还对血浆样品进行超滤，以获得无蛋白滤液。总同型半胱氨酸（tHcy），fHcy和rHcy及其相关的氨基硫醇，半胱氨酸，半胱氨酰甘氨酸和谷胱甘肽的浓度同时测定酸性柠檬酸钠血液使用反相高效液相色谱荧光检测。使用总、游离和还原级分的浓度，通过差值计算结合和氧化氨基硫醇。使用这种方法，我们比较了新诊断的缺血性卒中患者（n = 20）和健康的年龄和性别匹配的受试者（n = 20）的氧化还原状态。
结果
使用Stabilyte的全血中tHCy、tCys、tCysGly和tGSH浓度可稳定8小时;每种氨基硫醇的还原级分可稳定4小时。对于酸化柠檬酸钠化血液中的所有还原硫醇，无蛋白质滤液中的回收率为90-100%。缺血性脑卒中患者血浆tHcy、fHcy、bHcy、rHcy和oxHcy水平显著升高（P <0.0005），t-、f-、r-和oxCys水平显著升高（P <0.05）。脑卒中患者t-、B-和rCysGly浓度较低（P <0.05），t-、B-和oxGSH浓度也较低（P <0.005）。
结论
在酸性柠檬酸钠（BioPool Stabilyte）中采集血液，可以测定Hcy及其相关氨基硫醇的氧化还原状态，这可能有助于了解它们与脑血管疾病病因的关系。
BACKGROUND
Plasma "redox" status can be assessed by measurements of reduced (r)-, free (f)-, oxidized (ox)-, and protein-bound (b)-homocysteine (Hcy) plus the related aminothiols cysteine, cysteinylglycine (CysGly), and glutathione (GSH), but sample collection has been complex. The redox status has not been determined in ischemic stroke patients and may provide increased understanding of its role in pathogenesis. We wished to examine the feasibility of this measurement in samples collected in readily available acidic sodium citrate.
METHODS
We measured aminothiols and their stability in stabilized protein-free filtrate using acidic sodium citrate (BioPool Stabilyte, pH 4.3) vs EDTA whole blood. Before analysis, plasma samples were also ultrafiltered to obtain a protein-free filtrate. The concentrations of total Hcy (tHcy), fHcy, and rHcy and their related aminothiols, cysteine, cysteinylglycine, and glutathione were simultaneously determined on acidic sodium-citrated blood using reversed-phase HPLC with fluorescence detection. Bound and oxidized aminothiols were calculated by difference using the concentrations of the total, free, and reduced fractions. Using this approach, we compared the redox status in newly diagnosed ischemic stroke patients (n = 20) and healthy age- and sex-matched subjects (n = 20).
RESULTS
tHcy, tCys, tCysGly, and tGSH concentrations in whole blood with Stabilyte were stable for 8 h; the reduced fraction of each aminothiol was stable for 4 h. Recovery in the protein-free filtrate was 90-100% for all reduced thiols in acidified sodium-citrated blood. Patients with ischemic stroke had higher plasma tHcy, fHcy, bHcy, rHcy, and oxHcy (P <0.0005) and higher plasma t-, f-, r-, and oxCys (P <0.05). t-, b-, and rCysGly concentrations were lower in the stroke patients (P <0.05), as were t-, b-, and oxGSH (P <0.005).
CONCLUSIONS
Collection of blood in acidic sodium citrate (BioPool Stabilyte) permits the determination of the redox status of Hcy and its related aminothiols, which may add to the understanding of their relationship to the etiology of cerebrovascular disease.
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DOI：
10.1016/s0025-7125(16)30273-5
发表时间：
1993
期刊：
The Medical clinics of North America
影响因子：
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作者：
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DOI：
10.1016/0005-2760(87)90139-1
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影响因子：
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DOI：
10.1016/s0021-9258(18)61082-8
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1987-07
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影响因子：
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MALINOW, MR;NIETO, FJ;BOND, G
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