Page 48 - 2020年2月第31卷第3期

胡桃醌对大鼠血清中与脑组织损伤相关生物标志物的影响 Δ

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董文婷 ，霍金海，孙国东，王伟明（黑龙江省中医药科学院中药所分析室，哈尔滨 150036）
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中图分类号 R965 文献标志码 A 文章编号 1001-0408（2020）03-0298-05
DOI 10.6039/j.issn.1001-0408.2020.03.09

摘 要 目的：考察胡桃醌对大鼠脑组织的影响及其与脑组织损伤相关生物标志物的关系。方法：将 40只大鼠分为空白组和胡
桃醌高、中、低剂量组（34.832、17.416、8.708 mg/kg），每组 10只，每天灌胃给药 1次，连续给药 4周。末次给药后，考察各组大鼠的
一般行为学、脑指数及脑组织病理形态。采用超高效液相色谱-多反应检测质谱（UPLC-MRM-MS）法检测各组大鼠血清中左旋多
巴（L-Dopa）、L-酪氨酸（L-Tyr）、L-色氨酸（L-Trp）的含量。色谱条件为色谱柱Waters Acquity UPLC BEH C18，流动相为乙酸铵水溶
液-乙腈，梯度洗脱，流速为 0.3 mL/min，进样量为 5 μL，柱温为 30 ℃；质谱条件为电喷雾离子源，正离子模式下检测，毛细管电压
为3 500 V，去溶剂气流量为 650 L/h，去溶剂化温度为 350 ℃，离子源温度为 110 ℃。结果：与空白组比较，胡桃醌各剂量组大鼠
出现倦怠、四肢无力的行为；脑组织可见大、小脑皮质内血管充血，锥体细胞层部分神经元固缩，核深染，形态不规则，边界不清晰
等病理性改变；胡桃醌高剂量组脑指数显著增加（P＜0.05）。建立的UPLC-MRM-MS 法专属性良好，L-Dopa、L-Tyr、L-Trp的检测
质量浓度线性范围分别为 31.25～32 000、31.25～32 000、15.625～16 000 ng/mL（r＝0.999 1～0.999 9），检测限分别为 6.250、
5.625、3.125 ng/mL，定量限分别为 15.625、18.75、10.00 ng/mL，精密度、准确度、稳定性（24 h）试验 RSD 均小于 2.6％（n＝6），基
质效应为 95.1％～100.1％（RSD均未超过3.25％，n＝3）。动物实验结果显示，与空白组比较，胡桃醌中、高剂量组L-Dopa含量显
著升高（P＜0.01）；胡桃醌低、中、高剂量组 L-Tyr含量显著升高，L-Trp含量显著降低（P＜0.05或 P＜0.01）。结论：胡桃醌对大鼠的
行为学、脑指数、脑组织形态均有明显影响，可能通过升高血清中 L-Dopa 、L-Tyr 的含量，降低 L-Trp 的含量影响大鼠的脑组织
功能。
关键词 胡桃醌；脑组织；生物标志物；超高效液相色谱-多反应检测质谱法；含量测定

Effects of Juglone on Serum Biomarkers Related to Brain Tissue Injury in Rats
DONG Wenting，HUO Jinhai，SUN Guodong，WANG Weiming（Analysis Room，Institute of Traditional Chinese
Medicine，Heilongjiang Academy of Chinese Medical Sciences，Harbin 150036，China）

ABSTRACT OBJECTIVE：To investigate the effects of juglone on brain tissue of rats and its relationship with the biomarkers
related to brain tissue injury. METHODS：Totally 40 rats were divided into blank group，juglone high-dose，medium-dose and
low-dose groups（34.832，17.416，8.708 mg/kg），with 10 rats in each group. They were given medicine intragastrically once a day，
for consecutive 4 weeks.After last administration，the general behavior，brain index and brain tissue morphology were investigated.
UPLC-MRM-MS method was used to determine the serum contents of L-dopa（L-Dopa），L-tyrosine（L-Tyr）and L-tryptophan（L-Trp）
in rats. The chromatographic condition included Waters Acquity UPLC BEH C18 column，mobile phase consisted of ammonium
acetate aqueous solution-acetonitrile，gradient elution，at the flow rate of 0.3 mL/min，sample size of 5 μL，column temperature of
30 ℃；MS condition include electrospray ion source，in positive ion mode，capillary voltage of 3 500 V，desolvent gas flow of 650
L/h，desolvent temperature of 350 ℃，ion source temperature of 110 ℃. RESULTS：Compared with blank group，the rats in each
dose group showed the behavior of tiredness and weakness of limbs. The brain tissue morphology showed pathological changes，
which contained blood vessel congestion in the cerebral and cerebellar cortex，partial cell nucleus pyknosis in the pyramidal cell
layer，deep staining of nuclei，irregular shape and unclear boundary and other pathological changes；the brain index of juglone
high-dose group increased significantly（P＜0.05）. The established UPLC-MRM-MS method showed good specificity and the range
of L-Dopa，L-Tyr and L-Trp were 31.25-32 000，31.25-32 000，15.625-16 000 ng/mL（r＝0.999 1-0.999 9），respectively. The limits
of detection were 6.250，5.625，3.125 ng/mL，respectively. The limits of quantitation were 15.625，18.75，10.00 ng/mL，respectively.
RSDs of precision，accuracy and stability（24 h）tests were all
Δ 基金项目：国家自然科学基金青年科学基金资助项目（No. lower than 2.6％（n＝6）. Matrix effects were 95.1％ -100.1％
81503348）；黑龙江省自然科学基金项目（No.H2017044）；哈尔滨市应
（RSD are not more than 3.25％，n＝3）. Compared with blank
用技术研究与开发项目（No.2017RAQXJ224）
group，the contents of L-Dopa were increased significantly in
＊助理研究员，硕士。研究方向：中药药效与物质基础。电话：
juglone medium-dose and high-dose groups（P＜0.01）. The
0451-55653086。E-mail：1776427432@qq.com
# 通信作者：研究员，博士生导师。研究方向：中药新药研发及剂 contents of L-Tyr were increased significantly in juglone
型改革。电话：0451-55653086。E-mail：zyyjy@163.com lowdose，medium-dose and high-dose groups，while the

·298 · China Pharmacy 2020 Vol. 31 No. 3 中国药房 2020年第31卷第3期