Investigation of the mechanisms of Genkwa Flos hepatotoxicity by a cell metabolomics strategy combined with serum pharmacology in HL-7702 liver cells

1. To investigate Genkwa Flos hepatotoxicity, a cell metabolomics strategy combined with serum pharmacology was performed on human HL-7702 liver cells in this study.

2. Firstly, cell viability and biochemical indicators were determined and the cell morphology was observed to confirm the cell injury and develop a cell hepatotoxicity model. Then, with the help of cell metabolomics based on UPLC-MS, the Genkwa Flos group samples were completely separated from the blank group samples in the score plots and seven upregulated as well as two down-regulated putative biomarkers in the loading plot were identified and confirmed. Besides, two signal molecules and four enzymes involved in biosynthesis pathway of lysophosphatidylcholine and the sphingosine kinase/sphingosine-1-phosphate pathway were determined to investigate the relationship between Genkwa Flos hepatotoxicity and these two classic pathways. Finally, the metabolic pathways related to specific biomarkers and two classic metabolic pathways were analyzed to explain the possible mechanism of Genkwa Flos hepatotoxicity.

3. Based on the results, lipid peroxidation and oxidative stress, phospholipase A₂/lysophosphatidylcholine pathway, the disturbance of sphingosine-1-phosphate metabolic profile centered on sphingosine kinase/sphingosine-1-phosphate pathway and fatty acid metabolism might be critical participators in the progression of liver injury induced by Genkwa Flos.     

红心大白菊挥发油成分的GC-MS分析

目的 :分析我国中药材GAP基地红心大白菊挥发油的化学成分 ,为其质量评价提供科学依据。方法 :采用水蒸气蒸馏法提取红心大白菊挥发油 ,用GC毛细管柱进行分析 ,归一化法测定其相对含量 ,并用GC-MS法鉴定化学成分。结果 :检出 136个色谱峰 ,鉴定出 47个化合物 ,占挥发油总量的 62.24%。结论 :红心大白菊挥发油中 ,雪松醇、喇叭醇和 1-(1,5-二甲基-4-己烯基 )-4-甲基苯是其主要成分 ,以倍半萜类化合物喇叭醇含量最高 ,占总挥发油量的 13.9%。     

铁皮石斛花对高糖高脂饮酒致高血压大鼠的降压作用及机制研究

观察铁皮石斛花对高糖高脂饮酒致高血压大鼠的降压作用及可能机制。采用高糖高脂饲料喂养复合梯度饮酒造模4周制备高血压模型大鼠,模型成功后根据血压分为模型对照组、缬沙坦（5.7 mg·kg^-1）阳性对照组、铁皮石斛花2个剂量组（3,1 g·kg^-1）,连续给予相应药物6周。定期测量大鼠血压;末次给药后,检测血浆中内皮素-1（ET-1）、血栓素B2（TXB2）、前列环素（PGI2）、一氧化氮（NO）水平;检测胸主动脉内皮型一氧化氮合酶（eNOS）表达及病变情况;采用离体血管环张力实验检测胸主动脉血管内皮依赖性舒张情况。结果显示,铁皮石斛花能显著降低高血压大鼠收缩压和平均动脉压,并抑制胸主动脉中膜增厚及内皮细胞脱落;减少血浆中ET-1和TXB2含量,升高PGI2和NO含量;长期给药后胸主动脉血管内皮依赖性舒张明显增加,且能被eNOS抑制剂（L-NAME）所阻断,并能增加eNOS表达。因此,铁皮石斛花对高糖高脂饮酒致高血压大鼠具有明显的抗高血压作用,其机制可能是通过保护血管内皮细胞,平衡血管内皮舒缩因子,从而改善血管舒张功能,最终起到抗高血压作用。     

草棉花花瓣提取物对大鼠肝损伤的保护作用

目的：探讨草棉花花瓣采用不同工艺提取的黄酮类化合物即提取物Ⅰ(FGF-Ⅰ)和提取物Ⅱ(FGF-Ⅱ)对D-氨基半乳糖(D-GalN)引起的实验性肝损伤的保护作用并比较2种提取物的作用强度，对药物筛选提供依据。方法：Wistar大鼠随机分为正常组、模型组、阳性对照组(联苯双酯组，150 mg·kg^-1)、FGF-Ⅰ大、小剂量组(50，25 mg·kg^-1)和FGF-Ⅱ大、小剂量组(50，25 mg·kg^-1)7个组，腹腔注射350 mg·kg^-1的D-GalN造成急性肝损伤动物模型；观察2种提取物对血清谷丙转氨酶(ALT)、谷草转氨酶(AST)，肝组织匀浆超氧化物歧化酶(SOD)活性、丙二醛(MDA)含量、谷胱甘肽过氧化物酶(GSH-PX)活性等的影响。结果：对D-GalN引起的肝损伤动物模型FGF-Ⅰ和FGF-Ⅱ均能降低血清ALT，AST活性(P＜0.01)；FGF-Ⅰ大、小剂量组能增加肝组织SOD活性(P＜0.01)，大剂量可以降低MDA含量(P＜0.05)，但是对GSH-PX活性没有影响；FGF-Ⅱ大、小剂量组可以增加肝脏SOD，GSH-PX活性(P＜0.05)并可以降低MDA含量(P＜0.05)。结论：FGF-Ⅰ和FGF-Ⅱ均有一定的保肝作用， FGF-Ⅱ的作用优于FGF-Ⅰ。     

气相色谱-质谱联用法测定金银花中192种农药多残留

 目的 建立气相色谱-质谱联用法（GC-MS）测定金银花中192种农药多残留的分析方法,对样品进行普查,了解金银花中农药残留情况。 方法 用丙酮进行提取,采用凝胶渗透色谱（GPC）与固相萃取(SPE)结合净化的分析方法,采用气相色谱质谱联用法进行测定。结果 回收率总体处于70% ～110%之间,相对标准偏差≤15%,方法检出限总体处于0.01 mg·kg^-1以下。结论 此方法灵敏度高,准确,可靠,可作为金银花中的农药多残留测定法。样品普查结果表明,金银花中农药残留污染情况较为严重。
     

复方丁香洗剂中丁香酚和苦参碱的含量测定

目的建立紫外分光光度法及酸性染料比色法测定复方丁香洗剂中丁香酚和苦参碱的含量.方法丁香酚、苦参碱均可与氢氧化钠成盐而溶于碱,易于分离,前者于(296±2)nm波长处测定吸收度,后者与溴麝香草酚蓝络合后于(415±2)nm波长处测定吸收度.结果丁香酚平均回收率为98.30%,RSD为0.76%,在10.26～23.95 mg·L-1范围有良好的线性关系(r=0.999 8);苦参碱平均回收率为98.63%,RSD为0.83%,在3～10.5 mg·L-1范围线性关系良好(r=0.999 6).结论本方法准确,灵敏度好,可用于复方丁香洗剂或中药制剂中的丁香酚、苦参碱含量测定.     

舒肤软膏的薄层色谱鉴别

目的:建立舒肤软膏的质量控制标准.方法:采用薄层色谱法对舒肤软膏中的苦参、百部、金银花、青蒿油、薄荷脑进行定性鉴别.结果:在薄层层析色谱中均能检出苦参、百部、金银花、青蒿油、薄荷脑.结论:方法简便、专属性强,可用于该制剂的质量控制.     

Effect of a Korean Traditional Formulation, Hwaotang, on Superoxide Generation in Human Neutrophils, Platelet Aggregation in Human Blood, and Nitric Oxide, Prostaglandin E2 Production and Paw Oedema Induced by Carrageenan in Mice

Hwaotang, a traditional Korean medicinal formulation, is a dried decoctum of a mixture of 7 herbal medicines, consisting of Angelica gigantis Radix, Rehmanniae radix, Paeoniae radix, Ciniamomi cortex, Cnidii rhizoma, Persicae semen and Carthami flos. We have investigated that Hwaotang water extract (HOT) has various effects on stimulus-induced superoxide generation in human neutrophils. The effects of HOT on superoxide generation in human neutrophils were investigated. HOT significantly inhibited N-formyl-methionyl-leucyl-phenylalanine (fMLP)-induced superoxide generation in a concentration-dependent manner, but not that induced by arachidonic acid (AA). On the other hand, HOT enhanced superoxide generation induced by phorbol 12-myristate 13-acetate (PMA) in a concentration-dependent manner. The superoxide generation induced by PMA with HOT was suppressed by staurosporine, an inhibitor of protein kinase C, but was not suppressed by genistein, an inhibitor of protein tyrosine kinase. Tyrosyl phosphorylation of a 58 kDa protein, which was increased by fMLP, was inhibited by HOT. HOT also inhibited the generation of a 47 kDa protein and platelet aggregation in human blood. The results suggest that protein tyrosine kinase participates in fMLP-mediated superoxide generation by HOT-treated human neutrophils. HOT inhibited neutrophil functions, including degranulation, superoxide generation, and leukotriene B4 production, without any effect on 5-lipoxygenase activity. HOT reduced nitric oxide (NO) and prostaglandin E2 production in mouse peritoneal macrophages stimulated with lipopolysaccharide, whereas no influence on the activity of iNOS, COX-2 or COX-1 was observed. HOT significantly reduced mouse paw oedema induced by carrageenan. Western blot analysis showed that HOT reduced the expression of iNOS and COX-2. The results indicate that HOT exerts anti-inflammatory effects related to the inhibition of neutrophil functions and of NO and prostaglandin E2 production, which could be due to a decreased expression of iNOS and COX-2.     

RP-HPLC法测定金银花及金芪降糖片中的绿原酸

目的 建立同时测定金银花及金芪降糖片中绿原酸的反相高效液相色谱法。方法 色谱柱为Diamondsil C18（250 mm×4.6 mm，5 μm），以乙腈0.4%磷酸溶液（13∶87）为流动相，检测波长为327 nm，体积流量为1.0 mL/min。结果 在上述条件下测定金银花中绿原酸为2.49%，金芪降糖片中绿原酸为8.776 mg/片。结论 本方法操作简单、结果准确、回收率高、重现性好，可用于金银花及金芪降糖片的质量控制。     

沙生蜡菊花降脂活性部位的化学成分（III）

In Europe, the therapeutic application of Helichrysum arenarium (L.) Moench has been used as a folk medicine due to its choleretic, diuretic, and detoxify function for long time. The sap of its flos could be used to treat the disease of digestive system. During the course of our studies on activity screening, the methanolic extract of the flowers of Helichrysum arenarium (L.) Moench (Asteraceae, Everlasting in English) was found to have the lipid-lowering activity. For the purpose of finding the chemical constituents with the lipid-lowering activity, further chemical studied was carried out on its MeOH extracts. By various methods of isolation (silica gel column, ODS gel column, HPLC), six compounds were isolated, and with the methods of physico-chemical property and spectrographic analysis, six compounds were identified as lcariside F2 (1), icariside D1 (2), phenethanol-β-vicianoside (3), benzyl alcohol-β-vicianoside (4), benzoic acid-β-D-gentiobioside (5), phenethanol-β-D-gentiobio- side (6). Compounds 1-6 were obtained from Helichrysum Mill. for the first time.