Fu Ling, Wang Pan, Sun Yiqun, Wang Yangyang, Zhao Jing, Ye Yuting, Zhang Yanbin, Bi Yuefeng
Department of Pharmacognosy, Zhengzhou University School of Pharmaceutical Science, Ke Xue Da Dao 100, Zhengzhou, 450001, P. R. China.
Pharmacogn Mag. 2015 Oct-Dec;11(44):740-4. doi: 10.4103/0973-1296.165574.
Chrysanthemi indici Flos, a traditional herbal medicine is used to clearing heat-toxicity, removing the liver fire, and improving eyesight. In our preliminary work, an active extract of CTC in C. An indici Flos with anti-hepatitis B virus and liver protective activity was found by HepG2.2.1.5 test and experiment of protein synthesis in mice's injured liver. In this work, we aimed to study the active faction CTC further by qualitative and quantitative analysis method.
High performance liquid chromatography time of flight electrospray ionization mass spectrometry (HPLC TOF ESI-MS) analysis method of the CTC was established. Cumambrin A and angeloylcumambrin B in CTC were analyzed by high performance liquid chromatography-ultraviolet-evaporative light scattering detector (HPLC-UV-ELSD) analysis methods. A binary gradient elution program was conducted for chromatographic separation with acetonitrile (A) and ultrapure water (B) as follows: 0-10 min, 42-46% A; 10-20 min, 46-55% A; 20-25 min, 55-60% A; and 25-35 min, 60-75% A. The column temperature and UV wavelength were set at 30°C and 205 nm.
Ten constituents including (3R, 5R, 6S, 7S, 10R)-7-(2-hydroxy-2-propyl)-10-methyl-4-methyleneperhy, dronaphthal-ene-3, 5, 6-triol acetone solvate; (+)-edusmance-4, (14)-ene-11, 13-diol; linarin; luteolin; apigenin; tricin; 5, 3',4'- trimethyl-6,7-dimethoxy flavones; cumambrin A; acacetin; and angeloylcumambrin B in CTC were identified by HPLC TOF ESI-MS. The contents of sesquiterpenes in CTC were decreased by storing years.
The results showed that both UV and ELSD methods were feasible, accurate, and the determination results were in good consistency. The contents of two sesquiterpenes decreased with storing years. Two sesquiterpenes could be used as quality control for C. indici flos CTC.
野菊花是一种传统草药，用于清热解毒、清肝泻火和明目。在我们的前期工作中，通过HepG2.2.1.5试验和小鼠损伤肝脏蛋白质合成实验，发现了野菊花中具有抗乙型肝炎病毒和肝脏保护活性的活性提取物野菊花总成分（CTC）。在本研究中，我们旨在通过定性和定量分析方法进一步研究活性组分CTC。
建立了CTC的高效液相色谱-飞行时间-电喷雾电离质谱（HPLC-TOF-ESI-MS）分析方法。采用高效液相色谱-紫外-蒸发光散射检测器（HPLC-UV-ELSD）分析方法对CTC中的野菊花内酯A和当归酰野菊花内酯B进行分析。以乙腈（A）和超纯水（B）为流动相进行二元梯度洗脱，洗脱程序如下：0～10 min，42%～46%A；10～20 min，46%～55%A；20～25 min，55%～60%A；25～35 min，60%～75%A。柱温设定为30℃，紫外波长设定为205 nm。
通过HPLC-TOF-ESI-MS鉴定出CTC中的10种成分，包括（3R，5R，6S，7S，10R）-7-（2-羟基-2-丙基）-10-甲基-4-亚甲基十氢萘-3,5,6-三醇丙酮溶剂化物、（+）-桉叶-4（14）-烯-11,13-二醇、木犀草苷、木犀草素、芹菜素、小麦黄素、5,3',4'-三甲基-6,7-二甲氧基黄酮、野菊花内酯A、刺槐素和当归酰野菊花内酯B。CTC中倍半萜类成分的含量随储存年限的增加而降低。
结果表明，UV和ELSD方法均可行、准确，测定结果一致性良好。两种倍半萜类成分的含量随储存年限的增加而降低。两种倍半萜类成分可作为野菊花CTC的质量控制指标。
