HPLC测定愈伤平胶囊中芍药苷的含量

目的:建立愈伤平胶囊中芍药苷的含量测定方法。方法:采用高效液相色谱法测定,Dikma DiamonsilC18柱,以乙腈-0.1%磷酸(17∶83)为流动相,流速1.0mL/min,柱温:30℃,检测波长:230nm。结果芍药苷的线性范围为0.2112～2.1120μg;平均回收率为99.36%,RSD为1.96%(n=9);结论:所用方法测定样品灵敏度高,重复性好,可用于愈伤平胶囊的质量控制。     

栀子叶片愈伤组织诱导优化研究

目的:对栀子(Gardenia jasminoides Ellis)叶片愈伤组织诱导进行优化研究。方法:通过以叶片为外植体,MS和WPM为基本培养基,添加不同激素组合(6-BA、NAA和IBA)进行愈伤组织诱导研究。结果:栀子叶片在WPM+6-BA 0.5 mg/L+NAA 0.5 mg/L以及6-BA 1.0 mg/L+NAA 0.1(或0.5)mg/L中均有100%的诱导率;在MS培养基上,MS+6-BA 1.0(或1.5)mg/L+NAA 0.5 mg/L的诱导率最好,分别为70%和74%。栀子叶片愈伤组织在再分化培养基上均未诱导出丛生芽。结论:WPM培养基比MS培养基更易诱导栀子叶片愈伤组织,且栀子愈伤组织的再分化较困难。     

归芍浓缩丸中芍药内酯苷和芍药苷的含量测定

目的建立归芍浓缩丸中芍药内酯苷和芍药苷的含量测定方法。方法采用反相高效液相色谱(RP-HPLC)法测定归芍浓缩丸中芍药内酯苷和芍药苷的含量。色谱条件:用C18分析柱,以乙腈-0.05%冰醋酸溶液(17∶83)为流动相,检测波长为230 nm,进样量为20μl。结果高效液相色谱法测定芍药内酯苷在0.67~2.02μg范围内线性关系良好(r=0.999 8),平均回收率为100.5%,RSD=0.29%(n=7);芍药苷在0.41~1.63μg范围内线性关系良好(r=0.999 9),平均回收率为101.2%,RSD0.37%(n=7)。结论该方法简便快速、精密度高、准确度好,可作为该制剂的质量控制方法。     

八角莲愈伤组织中黄酮苷类化学成分研究

综合运用硅胶柱色谱、Sephadex LH-20凝胶柱色谱、制备薄层色谱和制备型HPLC等多种色谱技术对八角莲愈伤组织的化学成分进行分离纯化,并采用UV,IR,MS和NMR等波谱技术鉴定化合物的结构。从八角莲愈伤组织的95%乙醇提取物中共分离得到7个黄酮苷类化合物,分别鉴定为山柰酚-3-O-[6″-(3''-甲氧基)-丙二酰基]-β-D-吡喃葡萄糖苷(1),山柰酚-3-O-(6″-O-乙酰基)-β-D-吡喃葡萄糖苷(2),山柰素-3-O-β-D-吡喃葡萄糖苷(3),山柰酚-3-O-β-D-吡喃葡萄糖苷(4),异槲皮苷(5),槲皮素-4'-O-β-D-吡喃葡萄糖苷(6)和山柰酚-3-O-(6″-O-丙二酸单酰基)-β-D-吡喃葡萄糖苷(7)。以上化合物均为首次从八角莲愈伤组织中分离得到,其中化合物1,2,3,6和7为首次从八角莲植物材料中分离得到,化合物1是一个新化合物。     

芍药苷和芍药内酯苷的抗抑郁作用与NO/cGMP信号转导通路的相关性

目的:比较赤芍、白芍及其有效成分芍药苷、芍药内酯苷对小鼠的抗抑郁作用及其与NO/c GMP信号转导通路的相关性。方法:选取小鼠悬尾实验模型,给予赤芍水提物、白芍水提物,记录小鼠悬尾不动时间。采用相同实验方法给予小鼠芍药苷和芍药内酯苷,并检测小鼠大脑皮质及海马组织中NO/c GMP通路相关指标。结果:与模型组比较,2 g/kg赤芍和2 g/kg、1 g/kg白芍组小鼠悬尾不动时间减少,差异有统计学意义(P0.05)。与2 g/kg赤芍同剂量组比较,白芍的小鼠悬尾不动时间减少,差异有统计学意义(P0.05)。与模型组比较,20 mg/kg芍药苷组、20 mg/kg芍药内酯苷组的小鼠悬尾不动时间及NO、c GMP含量明显减少,差异有统计学意义(P0.05);且与10 mg/kg芍药苷同剂量组比较,芍药内酯苷组的小鼠悬尾不动时间及NO含量明显减少,差异有统计学意义(P0.05)。与模型组比较,20 mg/kg芍药苷组及20mg/kg芍药内酯苷组的小鼠n NOS、GluR1mRNA表达明显减少,差异有统计学意义(P0.05)。结论:白芍具有抗抑郁作用,芍药苷和芍药内酯苷是白芍抗抑郁作用的物质基础;与白芍比较,赤芍的抗抑郁作用较弱,与其主要含有抗抑郁作用较弱的芍药苷,而不含抗抑郁作用较强的芍药内酯苷有关。     

培养条件对罗布麻愈伤组织中金丝桃苷和异槲皮苷积累的影响

目的 探究促进罗布麻愈伤组织积累金丝桃苷、异槲皮苷的培养条件.方法 以罗布麻无菌苗子叶为材料,在诱导培养基上诱导愈伤组织并继代,探索不同激素配比、培养基碳源、氮源和盐浓度对罗布麻愈伤组织生长率和金丝桃苷、异槲皮苷积累的影响.结果 (1)孵育在MS+0.5 mg/L KT+0.5 mg/L NAA培养基上的愈伤组织中,金丝桃苷、异槲皮苷含有量最高,分别为6.12 mg/g、7.23 mg/g;(2)培养基碳源为2.0 mg/L蔗糖+2.0 mg/L葡萄糖时,有利于愈伤组织中金丝桃苷、异槲皮苷的积累,两种成分分别为7.45 mg/g、8.69 mg/g,是植物叶片的1.24倍和1.22倍;(3)当培养基氮源为2 475 mg/L NH4NO3,0 mg/L KNO3时,金丝桃苷、异槲皮苷含有量均最高,分别为8.67 mg/g、9.86 mg/g,是植物叶片的1.44倍和1.38倍;(4)培养基中添加150 mmol/L NaCl时,愈伤组织中金丝桃苷、异槲皮苷含有量最高,分别为10.07 mg/g、11.15 mg/g,是植物叶片的1.67倍和1.56倍.结论 通过改变培养条件,可以促进罗布麻愈伤组织中金丝桃苷和异槲皮苷的积累,为进一步以罗布麻愈伤组织为材,工业化生产金丝桃苷和异槲皮苷这两种重要的药效成分奠定基础.     

黄芩愈伤组织中多酚氧化酶活性与黄芩苷合成的关系研究

目的 研究黄芩愈伤组织中多酚氧化酶活性变化规律及其与黄芩苷次生合成之间的关系.方法 采用紫外分光光度法测定多酚氧化酶活性,采用高效液相色谱法测定黄芩苷的量.研究不同添加物(抗坏血酸、氯化钠、苯甲酸、聚乙烯吡咯烷酮、硫酸铜)对多酚氧化酶和黄芩苷量的影响.结果 在黄芩愈伤组织生长周期的前20 d,黄芩苷基本没有合成,多酚氧化酶的活性低水平表达;20～35 d,黄芩苷大量合成,多酚氧化酶的活性有所降低,但降低趋势较小;35～50 d,多酚氧化酶活性高水平表达,黄芩苷次生合成受到抑制.结论 多酚氧化酶的高水平表达不利于黄芩苷的次生合成,抗坏血酸、氯化钠、苯甲酸等均可抑制多酚氧化酶的活性,促进黄芩苷次生合成,其中以添加0.02%的抗坏血酸效果最为明显,可使黄芩苷的量提高17.6%(82.3 mg/g),而聚乙烯吡咯烷酮、硫酸铜对多酚氧化酶的活性有着明显的促进作用,抑制了黄芩苷的积累.     

肉苁蓉愈伤组织培养及所含有效成分量的研究

目的为扩大和提高肉苁蓉Cistanchedeserticola资源的利用效率。方法以肉苁蓉肉质茎的不同组织部位作为外植体,采用正交实验方法,筛选诱导愈伤组织产生的不同培养基和培养条件,并继代培养。应用HPLC方法对愈伤组织培养物中松果菊苷和洋丁香苷(毛蕊花糖苷)的量进行测定。结果肉苁蓉肉质茎的维管组织部分是诱导愈伤组织的最适外植体,鳞片叶次之,髓组织部分诱导效果较差。在暗培养、25~27℃条件下以B5为基本培养基附加6-BA(0.5~2mg/L)与IAA(0.5~1.5mg/L)诱导愈伤组织效果最佳;在半光照(光培养10h/d,暗培养14h/d)条件下,愈伤组织生长正常,最佳继代时间为25~30d。愈伤组织培养物中松果菊苷和洋丁香苷(毛蕊花糖苷)达4.37%。结论筛选出了适宜的肉苁蓉愈伤组织培养方法,且培养物中主要药用有效成分松果菊苷和洋丁香苷(毛蕊花糖苷)的量达到并超过了《中国药典》要求(0.3%)的标准。     

LC-MS/MS同时检测大鼠血浆中芍药苷和芍药内酯苷的浓度及其药代动力学研究

目的:研究白芍中芍药苷和芍药内酯苷在大鼠体内的药代动力学。方法:应用高效液相色谱-串联质谱联用分析方法,测定大鼠灌胃给予白芍提取物后芍药苷和芍药内酯苷的时间-血药浓度,根据药-时曲线计算药代动力学参数。结果:芍药苷和芍药内酯苷线性范围分别为1.0~1000 ng·m L-1,2.0~2000 ng·m L-1,回收率均大于80%,日间RSD≤10%,日内RSD≤11%,RE在-4.9%~2.4%之间。芍药苷和芍药内酯苷药代动力学参数如下:Tmax为(0.89±0.27)、(0.97±0.37)h;t1/2为(1.97±0.63)、(2.25±0.59)h;Cmax为(301.3±96.8)、(122.9±42.2)μg·L-1。AUC0-t为(1495.1±521.2)、(688.5±222.9)μg·h·L-1;AUC0-∞为(1541.1±548.0)、(720.1±241.0)μg·h·L-1。结论:该法简便、灵敏、特异,适用于血浆中芍药苷和芍药内酯浓度的测定。白芍中芍药苷和芍药内酯苷在大鼠体内药代动力学行为均符合二室模型。     

灵发素对三七愈伤组织发生和增殖的影响

为探讨三七Panax notogingseng愈伤组织快速高效的诱导和增殖,以三七茎段为外植体,进行了5组培养基的对比研究:(1)MS 2,4-D 2 mg/L(CK);(2)MS 2,4-D 2 mg/L N6-BA 2 mg/L;(3)MS 2,4-D 2 mg/L KT 2 mg/L;(4)MS 2,4-D 2 mg/L ZT 2 mg/L;(5)MS 2,4-D 2mg/L LFS 2 mg/L.结果表明:1.在CK的基础上添加灵发素(LFS),可以促进茎段愈伤组织早发生1～2周,诱导率达81%,比CK高出30%以上,而N6BA、KT和ZT作用不明显.2.LFS能使愈伤组织的鲜重在40 d内增加360.2%,而KT、ZT和N6-BA仅增加13.4%～21.8%.以平均接种1 g愈伤组织计,40 d内含LFS的第(5)组收获干物质81.5 mg,另4组只收21.6～25.9 mg.3.LFS可以使愈伤组织继代保存3年以上未表现老化,一直保持增殖能力.     

HPLC法测定白芍总苷胶囊中芍药内酯苷、芍药苷和苯甲酰芍药苷

目的建立HPLC对白芍总苷胶囊中芍药内酯苷、芍药苷和苯甲酰芍药苷同时定量的分析方法。方法采用HPLC法。色谱柱Zorbax SB-C18色谱柱(250mm×4.6mm,5μm);流动相A为乙腈,B为0.015%磷酸溶液,梯度洗脱程序为0min(8%A),5min(12%A),20min(20%A),25min(20%A);35min(45%A),40min(45%A);检测波长230nm;柱温30℃。结果芍药内酯苷、芍药苷和苯甲酰芍药苷质量浓度与峰面积均呈良好的线性关系。平均回收率芍药内酯苷为96.99%,RSD为0.56%;芍药苷为103.6%,RSD为0.77%;苯甲酰芍药苷为104.2%,RSD为1.35%(n=5)。结论该方法分离度好,简便易行,具有良好的重现性,可用于白芍总苷胶囊中芍药内酯苷、芍药苷和苯甲酰芍药苷的同时测定。     

Paeonia lactiflora Pall inhibits bladder cancer growth involving phosphorylation of Chk2 in vitro and in vivo

Ethnopharmacological relevance

Extracts of Paeonia lactiflora Pall (RPA), a traditional Chinese medicines has been shown to treat cancers.

Aim of the study

The purpose of this study is to evaluate the anticancer effect of RPA in urinary bladder carcinoma in vitro and in vivo.

Materials and methods

The cell viability was analyzed with DAPI. Flow cytometry and Western blot were used to study the apoptosis and cell cycle related mechanism. A rat model of bladder cancer was induced by N-butyl-N-(4-hydroxybutyl) nitrosamine (OH-BBN). Tumors were analyzed with immunohistochemical analysis.

Results

Our data suggested that RPA inhibits growth of bladder cancer via induction of apoptosis and cell cycle arrest. Treatment of TSGH-8301 cells with RPA resulted in G2-M phase arrest that was associated with a marked decline in protein levels of cdc2, cyclin B1, cell division cycle 25B (Cdc25B) and Cdc25 C. We also reported that RPA-mediated growth inhibition of TSGH-8301 cells was correlated with activation of checkpoint kinase 2 (Chk2). Herein, we further evaluated urinary bladder cancer using a model of bladder cancer induced by OH-BBN. Analysis of tumors from RPA-treated rats showed significant decrease in the expression of Bcl2, cyclin D1, and PCNA, and increase in the expression of p-Chk2 (Thr-68), Bax, and Cip1/p21.

Conclusion

Our data provide the experimental evidence that RPA could modulate apoptosis in models of bladder cancer.     

星点设计-效应面法优化牛蒡子中牛蒡子苷和牛蒡子苷元提取工艺

目的：优化牛蒡子中双指标成分最佳提取工艺。方法：采用星点设计-效应面法,在3个主要因素作用下,以牛蒡子苷及牛蒡子苷元提取率为评估指标,经过多元线性回归和二项式拟合,利用岭嵴分析法优化牛蒡子苷及其苷元提取工艺。结果：确定牛蒡子苷及牛蒡子苷元最佳提取工艺为：70%乙醇,24倍量,超声提取15 min。结论：上述工艺能够大量提取牛蒡子苷及牛蒡子苷元,为制备牛蒡子苷及其苷元提供试验依据,也为充分开发利用牛蒡资源提供参考。     

中心复合设计-效应面法优化黄芩苷磷脂复合物鼻用原位凝胶的制备工艺

目的优化黄芩苷磷脂复合物鼻用原位凝胶的制备工艺。方法采用中心复合设计-效应面优化法,以胶凝温度为评价指标,对黄芩苷磷脂复合物鼻用原位凝胶的制备工艺关键影响因素Poloxamer407(P407)、Poloxamer188(P188)、聚乙二醇6000(PEG 6000)进行研究,利用多元数学统计矩模型,对制备工艺参数和胶凝温度的相关性进行研究,建立工艺参数与胶凝温度的相关方程,确立最佳工艺参数。结果先将黄芩苷磷脂复合物1.0 g溶于0.1%三乙醇胺溶液中,再加入18 gP407、6 g P188、1 g PEG 6000、葡萄糖5 g、苯扎氯胺0.02 g,加去离子水至100 g,于冰浴(4℃)磁力搅拌下使其分散均匀,置4℃冰箱中保存24 h以上,直至聚合物完全溶解得到澄明溶液。结论采用中心复合设计-效应面法优化的黄芩苷磷脂复合物鼻用原位凝胶的制备工艺稳定可行。     

“醒脑安神调补肝肾”针刺法治疗绝经期综合征选穴应用

"醒脑安神调补肝肾"针刺法主穴选择:小醒脑(内关、印堂、三阴交)、四神聪、神门、关元、肾俞、肝俞、太溪、足三里、太冲。辨证配穴:肝肾阴虚加阴谷、照海;脾肾阳虚加命门、腰阳关、脾俞;辨病配穴:失眠配申脉、照海;精神不振、烦躁不安配大陵;潮热盗汗配合谷、复溜;头晕耳鸣配百会、风池、听会;大便溏薄配天枢、阴陵泉;浮肿配水分等。失眠、潮热汗出、情绪改变是绝经期阶段最显著的症状。"醒脑安神调补肝肾"针刺法重在醒脑安神,调补肝肾,调节机体阴阳平衡。     

A comparative study of the antimicrobial and phytochemical properties between outdoor grown and micropropagated Tulbaghia violacea Harv. plants

Aim of the study

The study aimed to compare the antimicrobial and phytochemical properties of in vitro cultured and outdoor grown Tulbaghia violacea plants in the quest to validate the use of micropropagated plants as alternatives to outdoor grown plants in traditional medicine. Tulbaghia violacea is used extensively in South African traditional medicine for HIV/AIDS patients and in the treatment of gastrointestinal ailments, asthma, fever and tuberculosis.

Materials and methods

Extracts of micropropagated and outdoor grown Tulbaghia violacea plants were evaluated for their antibacterial and antifungal activities against Bacillus subtilis, Escherichia coli, Klebsiella pneumoniae, Staphylococcus aureus and a fungus Candida albicans using microdilution methods. Saponins and phenolic compounds including condensed tannins, gallotannins and flavonoids were quantitatively determined using spectrophotometric methods. A qualitative test for saponins was also carried out.

Results

The petroleum ether (PE) extracts of micropropagated plants and dichloromethane (DCM) extracts of outdoor grown plants showed good antibacterial activity, each against two bacterial test strains. PE extracts of micropropagated plants showed the best antibacterial activity with a minimum inhibitory concentration (MIC) of 0.39 mg/ml against Bacillus subtilis. Good MIC (<1 mg/ml) and minimum fungicidal concentration (MFC) values of 0.78 mg/ml were only obtained in DCM extracts of outdoor grown plants. MIC and MFC values for water and ethanol extracts of both micropropagated and outdoor grown plants were similar and in the range 3.125-12.5 mg/ml. Total phenolics, gallotannins, flavonoids and saponins were significantly higher in micropropagated plants than in outdoor grown ones. In all cases, the amounts of phytochemical compounds in micropropagated plants were more than twice that of outdoor grown plants except for condensed tannins.

Conclusion

The results form a good basis for the use of Tulbaghia violacea micropropagated plants as a complement to the outdoor grown plants in traditional medicine.     

Pharmacological potential and conservation prospect of the genus Eucomis (Hyacinthaceae) endemic to southern Africa

Ethnopharmacological relevance

The genus Eucomis (Hyacinthaceae) consists of 10 species that are extensively used in African traditional medicine. This review is an appraisal of current information on the distribution and morphology, traditional uses, pharmacology, toxicology and approaches devised to enhance the conservation of the genus.

Methods

A systematic and comprehensive literature search using electronic searches such as Scopus, Google Scholar, Web of Science and ethnobotanical books was conducted.

Results

Evidence from traditional medicine usage shows wide utilization of this genus for ailments such as respiratory, venereal diseases, rheumatism as well as kidney and bladder infections. Pharmacological screening reported antimicrobial, antiplasmodial, antitumor, cytotoxic, phytotoxic and anti-inflammatory properties.

Conclusions

The potential of the genus Eucomis especially in terms of pharmacology cannot be overemphasized. Apart from the anti-inflammatory properties, the antifungal activity of Eucomis remains a valuable reservoir with potential application in the agriculture sector as a source of an affordable biocontrol agent. Based on the speculated toxic constituents in the genus Eucomis, it will be valuable to conduct detailed toxicological studies. Extensive utilization of members of the genus Eucomis is causing severe strain on wild populations. Although conventional propagation has been relatively effective in the alleviation of the declining status, micropropagation of members may be vital to guarantee the conservation of wild populations.     

益气活血利水不同配伍对慢性心衰大鼠AT1、ERK2影响的实验研究

目的:通过研究血管紧张素1型受体(AT1)及心肌细胞外信号调节激酶(ERK2)信号通路在慢性心衰中的作用,从分子生物学角度探讨中药对慢性心衰心室重构的影响,并进一步阐明其作用机理。方法:以冠脉结扎法配合力竭式游泳、减食等方法造成大鼠慢性心衰动物模型,对造模成功大鼠分为模型组、西药组、益气中药组、活血中药组、益气活血中药组和益气活血利水中药组,没有进行左冠脉结扎手术的假手术大鼠为正常组。利用实时荧光定量PCR技术及SABC免疫组化法检测慢性心衰大鼠AT1、ERK2的变化情况。结果:慢性心衰动物模型组大鼠心肌AT1、ERK2表达明显升高,与假手术组比较,差异有统计学意义(P0.01);经过治疗,各用药组大鼠心肌组织AT1、ERK2表达明显降低,其中益气活血组、益气活血利水组中药与西药组比较,差异无统计学意义(P0.05),且益气活血利水组疗效明显优于益气活血组(P0.01)。结论:证明益气、活血、利水中药复方可通过抑制心肌组织中AT1、ERK2的表达,抑制或逆转心室重构的过程,从而达到治疗慢性心衰的目的。     

Phenological and geographical influence in the concentration of selected bioactive 4-phenylcoumarins and chlorogenic acid in Hintonia latiflora leaves

Ethnopharmacological relevance

Hintonia latiflora is a Mexican medicinal plant with well-documented ethnomedical record comprising more than 400 years; in modern Mexico is used for treating several maladies such as diabetes and gastric ulcers. Although the pharmacological actions of the stem-bark and leaves have been demonstrated, the phenological and geographical effect on the concentration of active principles remains unexplored.

Aim of the study

The main goals of this study were to analyze the amount of selected 4-phenylcoumarins and chlorogenic acid in the leaves in order to assess the best harvesting period, and consequently their pharmacological efficacy. In addition, the preclinical antidiabetic efficacy of the infusion of the leaves was corroborated using standard pharmacological tests.

Materials and methods

The aqueous extracts from the leaves of Hintonia latiflora were prepared by infusion. For phenological and geographical comparison, leaves of Hintonia latiflora were collected in two different regions in Chihuahua and Michoacán. The material was analyzed by UPLC applying an analytical method that developed and validated for this purpose following the ICH guidelines. Investigation of the antidiabetic action was accomplished using an acute hypoglycemic test and oral glucose and sucrose tolerance tests.

Results

The validated analytical method was successfully applied for quantifying chlorogenic acid (1) and 4-phenylcoumarins (2–5) in the leaves of 12 different batches (1–12) during one-year period, and seven different batches for each geographical region; the concentration of the metabolites at the phenological cycle was significantly different, their concentration increased during the pre-senescence phase whereas in the leaf renovation stage the highest concentration of 2–5 was reached. The overall analysis of the active compounds concentration between the two populations investigated seems to be less important than the phenological variations. The aqueous extract of the leaves of Hintonia latiflora exerted its antidiabetic effect by different mechanisms showing comparable effect to the organic extract.

Conclusions

The findings of the present investigation reveal that the best harvest season for the leaves of Hintonia latiflora is between the leaves renovation and senescence stages avoiding the flowering period. In addition, no significant differences were found among the two different geographical populations analyzed. The infusions of the leaves, rich in 4-phenylcoumarins and chlorogenic acid, showed comparable antidiabetic action than the organic extract.