发育期大鼠海马氨基酸类神经递质的研究

The amino acid neurotransmitters of the hippocampus in rats

目的用高效液相色谱法检测大鼠海马组织中氨基酸类神经递质,研究它们在发育期大鼠海马中含量的变化.方法采用安捷伦反相高效液相色谱柱(ZORBAX,XDB,C18,4.6mm×250mm,5μm,Agilent),异硫氰酸苯酯为柱前衍生剂,在柱温38℃下采用二元梯度洗脱,于254 nm波长处用VARIAN-Prostar 325紫外检测器和LC-WORKSTATION-V6.2色谱数据处理系统检测和处理对照品和大鼠海马样品的氨基酸色谱数据,并计算乳鼠、幼鼠和成鼠海马氨基酸含量.结果氨基酸含量在Asp0.036～0.586 mg/ml、Glu0.027～0.4336 mg/ml、Gly0.053～0.8438 mg/ml、Tau0.039～0.6312 mg/ml和GABA0.0268～0.4288 mg/ml时,其峰面积与氨基酸含量的线性相关系数均大于0.9995;五种氨基酸回收率在96.0%～106%之间.三组样品间五种氨基酸含量的差别均有极显著性意义(P<0.01).结论该法检测大鼠海马组织中氨基酸类神经递质灵敏,特异性好,且分析时间短.氨基酸类神经递质含量随大鼠海马的发育而增高.

Objective To study the changes of trace amino acid neurotransmilters Glu, Asp, Gly, Tau and GABA in hippocampus during the developing stage.Methods By means of the Angelon Column of reversing phase high performance liquid chromatography ( 0.46 cm×15 cm, 5～6 μm), VARIAN Prostar 325 UV detector and LC WORKSTATION V6.2 Chromatography date manager, the trace amino acid neurotransmitters were determined respectively in the hippocampus of mammal, juvenile and matured 30 rats, which involved pre column derivatization with phenylisothiocyanate, reversed phased gradient elution, using an eluent of acetonitrice acetate, current velocity being 1.0 mL/min and detecting wave length 254nm.Results While the amino acid neurotransmitters being Asp 0.036 ～ 0.586 mg/ml, Glu 0.027 ～ 0.4336 mg/ml, Gly 0.053 ～ 0.8438 mg/ml Tau 0.039 ～ 0.6312 mg/ml and GABA 0.0268 ～ 0.4288 mg/ml respectively, their concentrations had a high linear relationship with its peak area ( r > 0.99 ) and the precisions(%) among 96.0 ～106. The differences of amino acid neurotransmitters among the three samples were significant ( P < 0.01 ).Conclusions This method has sufficient sensitivity, specificity and less separation time for determination of trace amino acid neurotransmitters in the hippocampus of rats. With the developing and maturing of rat hippocampus, the concentrations of the amino acid neurotransmitters are increasing.