摘要
研究了丹酰氯柱前衍生反相高效液相色谱法测定人脑脊液中γ-氨基丁酸(Gaba)和谷氨酸(Glu)的方法。检测波长为UV254nm;流动相A:甲醇;流动相B:四氢呋喃-甲醇-0.05mol/L醋酸钠(pH6.2)(575420,V/V);流速为1mL/min;梯度洗脱。讨论了有关测试条件。方法的线性范围分别为5~1000μmol/L(Glu)和1~600μmol/L(Gaba);最低检出限(μmol/L)分别为0.002(Glu)和0.001(Gaba);Glu和Gaba的回收率分别为96.9%~98.8%和97.5%~98.8%;3种浓度的Glu、Gaba标准液的峰高测定(n=5)表明,其变异系数分别为2.2%~3.4%和3.8%~5.6%。方法简便、快速、可靠。
Е ? Aminobutyric acid (Gaba) and glutamic acid (Glu) are believed to be the major neurotransmitter. Levels in cerebrospinal fluid (CSF) may reflect their metabolism in various neurotic and psychiatric diseases. Measurements of Gaba and Glu in body fluids will help to elucidate their metabolic role and diagnostic value. In the present work, the concentrations of glutamic acid and γ aminobutyric acid in human cerebrospinal fluid were determined by reversed phase high performance liquid chromatography using pre column derivatized with dansyl chloride and UV detection at 254nm. The mobile phase was A:CH 3OH and B: THF/CH 3OH/0.05mol/L NaAc(pH 6.2)(5/75/420,V/V) with gradient elution. The flow rate was lmL/min CSF samples were deproteinizated with methanol. After centrifugation at 15 000 r/min for 10 min, the supernatant was introduced into a screw capped vial and evaporated to near dryness at 80℃. Derivatization was carried out by the addition of 250μL of bicarbonate solution (pH 9.8) and 250 μL of dansyl chloride solution (4g/L) followed by heating at 40℃ water bath for 30 min. Extraction was performed with 300μL of ethyl acetate and the organic layer separated was dried at room temperature under nitrogen. The dry residue was dissolved and injected into the HPLC system. The linear range of the method was 5 1 000μmol/L (r= 0.998 8 ) for Glu and 1 600μmol/L (r=0.998 4) for Gaba. The lower limit of detection were 0.002 μmol/L and 0.001 μmol/L respectively. The recoveries were 96.9% 98.8% and 97.5% 98.8% and the coefficients of variation of peak height measurements were 2.2% 3.4% and 3.8% 5.6% respectivery. The method enables a simple, rapid and reproducible quantification of Glu and Gaba neurotransmitter.
出处
《色谱》
CAS
CSCD
北大核心
1997年第3期237-239,共3页
Chinese Journal of Chromatography
关键词
高效液相色谱
氨基丁酸
谷氨酸
脑脊液
high performance liquid chromatography, γ aminobutyric acid,glutamic acid, cerebrospinal fluid