柱前衍生RP-HPLC荧光检测法同时测定大鼠血浆和脑脊液中6种运动疲劳相关氨基酸  

Determination of Six Kinds of Amino Acid Related Exercise Fatigue in Blood Plasma and Cerebrospinal Fluid in Rat by RP-HPLC with Fluorescence Detection after OPA Precolumn Derivatization

作　　者：任淑萌[1] 蒋晔[1] 郝福[1] 李艳荣[2] 

机构地区：[1]河北医科大学药学院,河北石家庄050017 [2]承德医学院中药研究所,河北承德067000

出　　处：《体育科学》2008年第3期67-72,共6页China Sport Science

摘　　要：目的:建立邻苯二甲醛(OPA)柱前衍生反相高效液相色谱荧光检测法,同时测定大鼠血浆和脑脊液中5-羟色氨酸(5-HTP),γ-氨基丁酸(GABA),色氨酸(Trp),缬氨酸(Val),异亮氨酸(Ile)和亮氨酸(Leu)6种与运动疲劳相关氨基酸。方法:样品经甲醇沉淀后,以OPA进行柱前衍生。色谱柱为Phenomenex C_(18)(250×4.6 mm,5μm),以A甲醇-B0.05 mol/L NaAc·3H_2O(pH6.0,内含0.05%THF)为流动相进行梯度洗脱,柱温30℃,λ_(ex)为340 nmn,λ_(em)为450 nm。结果:在上述色谱条件下,5-HTP、GABA、Trp、Val、Ile、Leu和内源性物质在22.5min内分离度良好;6种氨基酸分别在2.87～368μmlol/L、6.31～807μmol/L、2.04～262μmol/L、3.52～451μmol/L、2.58～331μmol/L、2.54～326μmol/L的范围内呈良好的线性关系;运动过度组中上述6种氨基酸含量均比对照组高。结论:本法简便灵敏,结果准确可靠,可以快速地测定大鼠血浆和脑脊液中6种运动疲劳相关氨基酸的含量,为准确地反映中枢神经系统的活动状态和变化提供合理的依据。An RPLC-FL method was established for the determination of 5-Hydroxy tryptohan （5-HTP）, γ-aminobutyric acid （GABA）, tryptophan （Trp）, valine （Val）, isoleucine （Ile） and leucine （Leu）,related to exercise fatigue in blood plasma and cerebrospinal fluid in rat. Samples preparation involved a protein precipitation with methanol and derivatization with OPA. The separation was reformed on a phenomenex C18 （250 × 4.6ram, 5μm）, using a gradient elution mode with mobile phase of 0. 05mol/L NaAc. 3H20 （contained 0. 05% THF and adjusted pH to 6.0 with acetic acid） methanol at 30℃. The λ ex and λ em were 340 nm and 450 nm, respectively. These 6 kinds of were separated within 22.5min. The good linearity was achieved in the range of 2.87-368 μmol/L,6.31-807 μmol/L, 2. 04-262 μmol/L, 3.52-451 μmol/L, 2.58-331 μmol/L,2.54-326 μmol/L respectively. The contents of six kinds of amino acid in hyperkinesias group were higher than that of control group. This method is simple, flexible, accurate and suitable for determining these amino acids and provides a rational basis for reflecting the state and changes in central nervous system.

关 键 词：运动疲劳 氨基酸 邻苯二甲醛 柱前衍生 血浆 脑脊液 动物实验 鼠 

分 类 号：G804.7[文化科学—运动人体科学]