附子中双酯型、单酯型、非酯型生物碱对佐剂性关节炎作用的实验研究

[目的]明确附子中双酯型生物碱、单酯型生物碱及非酯型生物碱对佐剂性关节炎大鼠血清一氧化氮（NO）、肿瘤坏死因子（TNF-α）水平及足趾容积变化的影响,初步明确其治疗佐剂性关节炎的药效物质。[方法]雄性wistar大鼠,左后足趾皮内一次性注射弗氏完全佐剂（FCA）0.1mL/只造模,2W后,取造模成功的大鼠,随机分为模型组、单酯型生物碱组、双酯型生物碱组及非酯型生物碱组,各给药组灌胃给予相应生物碱药液,给药2W后,检测血清中NO及TNF-α含量,造模及给药期间同时记录体重及足趾容积变化情况。[结果]与正常组比较,模型组与各给药组大鼠体重增长均受到抑制（P〈0.05）;与模型组比较,各给药组血清中NO、TNF-α含量亦显著性降低（P〈0.01）,单酯型组及非酯型组在降低NO作用上较双酯型明显（P〈0.05）;且各给药组大鼠的足趾容积显著降低（P〈0.01）,其中单酯型组效果更为明显。[结论]附子中单酯型、双酯型及非酯型生物碱对大鼠佐剂性关节炎有良好的治疗作用,其中单酯型生物碱的治疗作用更为明显。     

拟青霉类生物碱的抗排斥作用及其机制研究

采用DBA／2小鼠耳廓皮下同种异体乳鼠心脏组织移植方法，研究蝙蝠蛾拟青霉人工发酵菌丝培养物(PHC)中类生物碱部位的抗排斥作用。结果发现：3.0mol／L拟青霉类生物碱(PAK)能显著延长心脏移植物存活期，降低术后免疫应答排斥反应初期小鼠体内腹腔巨噬细胞(PMΦ)吞噬作用及产生IL-1活性，与空白对照组相比差异有极显著意义。本实验未发现PAK的淋巴细胞毒性作用。初步证实PAK是PHC发挥抗排斥作用的活性有效部位之一。     

葛根芩连方药中黄连总生物碱的含量测定

目的 建立葛根芩连汤及其制剂中黄连总生物碱的含量测定方法。方法 氧化铝柱纯化,紫外分光光度法350 nm处测定含量。结果 葛根芩连汤水提物中总生物碱含量为32.4 mg/g,加样回收率为98.1% (RSD =2.9%)。葛根芩连微丸中总生物碱含量为19.6 mg/g。结论 该法测定黄连总生物碱方法简单、结果准确,可为葛根芩连微丸质量标准、葛根芩连方及其他含黄连的方剂中生物碱的含量测定提供参考。     

正交试验设计优选九节龙皂苷I聚乳酸微球的制备工艺

目的 筛选溶剂蒸发法制备九节龙皂苷I聚乳酸微球（ADS-I-PLA-MS）最佳工艺。方法 采用HPLC-ELSD测定方法,以包封率和载药量为评价指标,W/O/W溶剂蒸发法制备微球;通过单因素和正交试验设计,考察内水相九节龙皂苷I（ADS-I）甲醇溶液的质量浓度、ADS-I甲醇溶液与聚乳酸（PLA）二氯甲烷溶液体积比、PLA二氯甲烷溶液质量浓度和聚乙烯醇（PVA）体积等因素对ADS-I-PLA-MS包封率及载药量的影响。结果 溶剂蒸发法制备ADS-I-PLA-MS的最佳工艺条件为ADS-I甲醇溶液质量浓度为8 mg/mL、ADS-I甲醇溶液与PLA二氯甲烷溶液体积比为1∶13、PLA二氯甲烷溶液质量浓度为90 mg/mL、PVA体积为500 mL。结论 优选出的ADS-I-PLA-MS制备工艺合理可行。     

巴豆生物碱对Lewis小鼠肺腺癌细胞生长和凋亡的影响及机制研究

目的 通过研究巴豆生物碱（CA）对Lewis小鼠肺腺癌细胞生长及对凋亡相关蛋白（Survivin、Caspase-3、Bax）、YAP蛋白的调控作用,探讨CA对肺腺癌细胞生长和凋亡的影响及其具体的作用机制。方法 培养Lewis鼠源性肺腺癌细胞（LLC）,复制小鼠皮下成瘤模型,分为5组,每组10只,分别为对照组、低剂量CA组、中剂量CA组、高剂量CA组及顺铂组。皮下成瘤1周后给予药物干预,第22天对5组小鼠取瘤称重,计算抑瘤率并通过光学显微镜观察各组病理形态学的变化,判断CA对瘤体生长的抑制作用;Western blotting检测Survivin、Caspase-3、Bax及YAP蛋白相对表达量;qRT-PCR法检测Survivin、Caspase-3、Bax及YAP mRNA相对表达量。结果 中剂量CA组和高剂量CA组小鼠的一般生存状态较对照组、低剂量CA组及顺铂组好。低剂量CA组、中剂量CA组、高剂量CA组及顺铂组肿瘤细胞出现不同程度细胞凋亡现象,对照组肿瘤细胞多为坏死细胞。低剂量CA组、中剂量CA组、高剂量CA组及顺铂组的抑瘤率分别是13.91％、14.83％、27.84％和68.45％,4组抑瘤率比较,差异有统计学意义（P <0.05）,各组抑瘤率依次升高,顺铂组肿瘤生长最慢。不同剂量CA组中Survivin mRNA相对表达量低于对照组（P <0.05）,CA剂量越高,Survivin mRNA相对表达量越低;而不同剂量CA组中Caspase-3和Bax mRNA相对表达量高于对照组（P <0.05）,CA剂量越高,Caspase-3和Bax mRNA的相对表达量越高;各组YAP mRNA的相对表达量比较,差异无统计学意义（P >0.05）。结论 CA对Lewis肺腺癌小鼠瘤体生长有抑制作用,并引起细胞凋亡。CA通过促进Bax和Caspase-3表达及抑制Survivin表达来调控Lewis小鼠肺腺癌细胞凋亡;其调控机制可能是通过下调Survivin的表达,从而使得Survivin对Caspase-3的抑制作用减弱,而Bax上调可进一步激活Caspase-3,从而诱导Caspase级联效应促进细胞凋亡,为肺腺癌治疗提供了新思路。     

Four new C19-diterpenoid alkaloids from Aconitum hemsleyanum var. circinatum

Four new C19-diterpenoid alkaloids, hemaconitines A–D (1–4), were isolated from the roots of Aconitum hemsleyanum var. circinatum. Their structures were elucidated as 19R-hydroxyl-secoyunnaconitine (1), (3R)-hydroxyl-liwaconitine (2), 14-anisoyl-leucanthumsine E (3), and 19R -acetonyl-8-O-methyltalatisamine (4) by extensive spectroscopic analysis (IR, UV, HR-ESI-MS, 1D, and 2D NMR).     

Drug target interaction of tubulin-binding drugs in cancer therapy

Microtubules and their component protein, tubulin, constitute a popular target for the treatment of cancer. Many drugs that are presently used in clinics or in clinical trials and drugs that show promise as anticancer drugs bind to tubulin and microtubules. There are three conventional binding sites on β-tubulin where many of these drugs bind. The binding properties, conformational changes upon binding, association constants and thermodynamic parameters for the drug–tubulin interaction on these three sites are discussed. The antiproliferative activities of these drugs and the possible correlation with the binding properties are also described.     

Simultaneous Determination of Six Quaternary Ammonium Alkaloids in Coptidis Rhizoma by UPLC

Objective To establish a new, rapid, and reliable reversed-phase ultra performance liquid chromatography (RP-UPLC) method for the simultaneous determination of six quaternary ammonium alkaloids (QAAs) in Coptidis Rhizoma. Methods The effect of different experimental parameters on the analysis of QAAs by RP-UPLC was evaluated. Results Optimal resolution was achieved with an Acquity UPLC BEH C18 column using a gradient elution profile and a mobile phase consisting of water spiked with 10 mmol/L ammonium bicarbonate (A, pH adjusted to 10.0 by ammonia water) and acetonitrile (B), at a flow rate of 0.30 mL/min and wavelength of 345 nm. The column temperature was set at 30 ℃. The proposed method was found to be reproducible, precise, and rapid according to the method validation. Conclusion The proposed method, which is compatible with MS analysis and the preparation of QAA, provides some helpful insights into the quality control of Coptidis Rhizoma.     

Real-time toxicity prediction of Aconitum stewing system using extractive electrospray ionization mass spectrometry

Due to numerous obstacles such as complex matrices, real-time monitoring of complex reaction systems (e.g., medicinal herb stewing system) has always been a challenge though great values for safe and rational use of drugs. Herein, facilitated by the potential ability on the tolerance of complex matrices of extractive electrospray ionization mass spectrometry, a device was established to realize continuous sampling and real-time quantitative analysis of herb stewing system for the first time. A complete analytical strategy, including data acquisition, data mining, and data evaluation was proposed and implemented with overcoming the usual difficulties in real-time mass spectrometry quantification. The complex Fuzi (the lateral root of Aconitum)–meat stewing systems were real-timely monitored in 150 min by qualitative and quantitative analysis of the nine key alkaloids accurately. The results showed that the strategy worked perfectly and the toxicity of the systems were evaluated and predicated accordingly. Stewing with trotters effectively accelerated the detoxification of Fuzi soup and reduced the overall toxicity to 68%, which was recommended to be used practically for treating rheumatic arthritis and enhancing immunity. The established strategy was versatile, simple, and accurate, which would have a wide application prospect in real-time analysis and evaluation of various complex reaction systems.     

Pinellia Total Alkaloids Modulate the GABAergic System in Hippocampal Formation on Pilocarpine-Induced Epileptic Rats

Objective: To investigate the neuromodulatory effect of pinellia total alkaloids(PTA) on the gamma-aminobutyric acidergic(GABAergic) system in epileptic rats, and preliminarily evaluate the anti-epileptic effect of PTA. Methods: Ninety-one male Sprague-Dawley rats were randomized to a control group(n=17) or an epileptic group(n=74) using computer-generated random numbers. Status epilepticus(SE) was induced with pilocarpine in the epileptic group. Epileptic rats that survived SE were randomly divided into 4 groups, namely an epilepsy group(n=13), a topiramate(TPM, 60 mg/kg) group(n=12), a high-dose PTA(800 mg/kg) group(n=12), and a low-dose PTA(400 mg/kg) group(n=10). Treatments were given intragastrically once daily for 14 days. The control group and epilepsy group received normal saline. Spontaneous recurrent seizures(SRSs) were monitored 8-h daily for 7 days after treatment. Then, the hippocampal formation tissues were collected. GABA level was measured using enzyme-linked immunosorbent assay. Protein and mRNA expression levels of glutamate decarboxylase 65(GAD65), GABA transporter-1(GAT-1), GABA transaminase(GABA-T), and GABAA receptor(GABAAR) α4, α5, γ2 and δ subunits were measured using Western-blotting analysis and quantitative polymerase chain reaction. Results: PTA lowered the incidence and frequency of SRS(both doses vs. the TPM group, P0.05). Compared with the epilepsy group, PTA increased the levels of GABA(both doses P0.01) and GAD65(mRNA, 800 mg/kg, P0.01), and suppressed the levels of GAT-1(mRNA, 800 mg/kg, P0.01; 400 mg/kg, P0.05), GABA-T(mRNA, both doses P0.01), and GABAAR δ subunit(protein, 800 mg/kg, P0.05) and γ2 subunit(protein, both doses P0.01). PTA upregulated the low-expressed mRNA levels of GABAAR α5 subunit(400 mg/kg, P0.01), δ subunit(800 mg/kg, P0.05), and γ2 subunit(400 mg/kg, P0.05). Conclusions: PTA regulated the GABAergic system through modulating GABA levels and the expression levels of GAD65, GAT-1, GABA-T, and GABAAR α4, α5, γ2 and δ subunits. PTA may exert antiepileptic effects on the pilocarpine-induced epilepsy model.