小鼠脑组织中单胺类神经递质含量测定方法的建立与比较

目的 比较高效液相色谱(HPLC)-紫外光(UV)、HPIC-荧光(FLD)和HPLC-质谱(MS)3种检测方法测定小鼠不同脑组织中单胺类神经递质含量的优劣性,并应用最优方法测定血管性抑郁小鼠不同脑组织中单胺类神经递质的含量.方法 分别采用HPLC仪器搭配UV、FLD和MS检测器建立小鼠不同脑组织中多巴胺(DA)、去甲肾上腺素(NE)和5-羟色胺(5-HT)的含量测定方法,并进行系统的方法学验证,采用HPLC-FLD法对血管性抑郁小鼠不同脑组织中单胺类神经递质DA、NE和5-HT含量进行测定.结果 HPLC-UV法的定量限分别为DA 103.5 ng/mL、NE 107.5 ng/mL、5-HT 93.6 ng/mL;HPLC-FLD法的定量限分别为DA 10.35 ng/mL、NE 10.75 ng/mL、5-HT9.36 ng/mL;HPLC-MS法的定量限分别为DA 10.35 ng/mL、NE 32.25 ng/mL、5-HT 9.36 ng/mL.HPLC-FLD和HPLC-MS法对DA和5-HT的测定优于HPLC-UV法,HPLC-FLD法对于NE的测定优于HPLC-MS法,而HPLC-MS法存在较强的基质效应.HPLC-FLD法检测结果示血管性抑郁小鼠海马组织中DA含量最低,大脑皮质中5-HT含量最低,而不同脑组织中NE含量没有明显差别.结论 与HPLC-UV法和HPLC-MS法相比,HPLC-FLD法更适用于小鼠脑组织中单胺类神经递质(DA、NE、5-HT)的含量测定.DA和5-HT可以用作血管性抑郁症疾病的诊断标志物.

Establishment and comparison of content determination methods for monoamine neurotransmitters in brain tissue of mice

Objective To compare the advantages and disadvantages between high performance liquid chromatography (HPLC)-ultraviolet (UV, HPLC-UV), HPLC-fluorescence detector (FLD, HPLC-FLD) and HPLC-mass spectrometry (MS, HPLC-MS) for determination of the monoamine neurotransmitters (MNs) in brain tissues of mice, and to determine the content of MNs in brain tissues of vascular depression mice using the optimal method. Methods We used the UV, FLD and MS detectors to establish different methods for determination of dopamine (DA), norepinephrine (NE) and 5-hydroxytryptamine (5-HT) in brain tissues. Then the methods was verified with methodology. The HPLC-FLD was applied for the content determination of DA, NE and 5-HT in different brain tissues of vascular depression mice. Results It was observed that the limits of quantification for HPLC-UV, HPLC-FLD and HPLC-MS were DA 103.5 ng/mL, NE 107.5 ng/mL and 5-HT 93.6 ng/mL; 10.35 ng/mL, 10.75 ng/mL and 9.36 ng/mL; and 10.35 ng/mL, 32.25 ng/mL and 9.36 ng/mL, respectively. Both HPLC-FLD and HPLC-MS for DA and 5-HT were superior to HPLC-UV. HPLC-FLD for NE was superior to HPLC-MS. Compared with HPLC-UV and HPLC-FLD, HPLC-MS showed remarkable matrix effects. HPLC-FLD determination results showed that content of DA was the lowest in hippocampus, 5-HT was the lowest in cerebral cortex, and NE did not differ in different brain tissues of vascular depression mice. Conclusion Compared with HPLC-UV and HPLC-MS, HPLC-FLD can more effectively determine the contents of MNs (DA, NE and 5-HT). DA and 5-HT can be used as the diagnostic markers of the vascular depression diseases.