The Stimulatory Effects of the Antimicrobial Agents Bavistin, Cefotaxime and Kanamycin on <i>In Vitro</i>Plant Regeneration of <i>Centella asiatica</i>(L.)—An Important Antijaundice Medicinal Plant

Antimicrobial agents such as bavistin, cefotaxime and kanamycin were evaluated for their effects on the rapid shoot regeneration from nodal explants of Centella asiatica (L.). Filter sterilized bavistin (250 mg/L) was augmented alone and in combination with cytokinins such as BAP and TDZ into the media to trace the effect on regeneration. On this basis, the potential use of bavistin (150 mg/L) along with BAP (2.0 mg/L) was evaluated which showed the maximum shoot number (6.6) and shoot length (4.4 cm) respectively. Cefotaxime at the concentration of 100 μM/L was found to be effective to obtain the maximum shoot number formation (5.8) with the regeneration frequency (90%). Kanamycin at the concentration of 80 μM/L induced maximum shoot regeneration (5.12). Kanamycin at 100 μM/L or at higher concentrations reduced the shoot regeneration. The best rooting response was noticed when in vitro regenerated microshoots were transferred to the rooting medim which was supplemented with IBA (2.0 mg/L). This combination generates 90% of regeneration frequency and maximum number of roots per shoot (14.2) and high root length (4.2 cm). The rooted plants were acclimatized and transferred to field for survivalance. The addition of antibiotics was found to be more effective and safer for using since their effects on regeneration were found to be negligible.

Biosynthesis of copper nanoparticles supported on manganese dioxide nanoparticles using Centella asiatica L. leaf extract for the efficient catalytic reduction of organic dyes and nitroarenes

In this study we designed a novel,cost‐efficient and green method for the synthesis of copper nanoparticles(Cu NPs)supported on manganese dioxide(MnO2)NPs,using Centella asiatica L.leaf extract as a naturally‐sourced reducing agent,without stabilizers or surfactants.This synthetic process is environmentally‐friendly and avoids the use of toxic reducing agents.Phenolic hydroxyl groups in the leaf extract are believed to reduce Cu2+in solution to generate Cu NPs that are subsequently stabilized on the MnO2NP surfaces.The resulting Cu/MnO2nanocomposite was fully characterized using X‐ray diffraction,transmission electron microscopy,field emission scanning electron microscopy,energy‐dispersive X‐ray spectroscopy and Fourier transform infrared spectroscopy.This material was found to function as a highly active,efficient and recyclable heterogeneous catalyst for the reduction of Congo red,rhodamine B and methylene blue as well as nitro compounds such as2,4‐dinitrophenylhydrazine and4‐nitrophenol in the presence of NaBH4in aqueous media at ambient temperature.The high stability of the Cu/MnO2nanocomposite also allows the catalyst to be separated and reused several times without any significant loss of activity.?2018,Dalian Institute of Chemical Physics,Chinese Academy of Sciences.Published by Elsevier B.V.All rights reserved.

A new triterpene and a saponin from Centella asiatica

A new triterpene and a saponin, named 2α,3β23-trihydroxyurs-20-en-28-oic acid （1） and 2α,3β,23-trihydroxyurs-20-en-28-oic acid O-α-L-rhanmopyranosyl-（1→4）-O-β-D-glucopyranosyl-（1→6）-O-β-D-glucopyranosyl ester （2）, have been isolated from the aerial part of CenteUa asiatica. Their structures were elucidated by spectral methods, including 2D-NMR spectra.

Antifertility properties ofCentella asiatica ethanolic extract as a contraceptive agent: Preliminary study of sperm proteomic

Objective:To investigate the antifertility properties ofCentella asiatica L. ethanolic extract and identify the sperm proteomic changes in treated male rats.Methods: A total of 32 male Sprague-Dawley rats were divided into two groups: control group (n=16) received distilled water and treatment group (n=16) which received 300 mg/kg ofCentella asiatica L. extract for 42 d. After the treatment period, the number of implantation sites was recorded and the sperm proteomic changes were analysed by 2D gel electrophoresis. In addition, the expression of protein spots was quantified by MALDI-TOF analysis.Results:Centella asiatica L. extract resulted in low number of implantation sites in the treatment group (100.00±2.82) compared to the control group (183.00±2.14). The percentage of infertile male rats in the treatment group was higher (43.75%) compared to the control group (18.75%). Proteomic analysis showed the expression of protein spots identified in the treatment group decreased with 234 spots compared to the control group with 282 spots.Conclusions:The results revealed that there was antifertility activity in the male rats with the administration ofCentella asiatica L. ethanol extract and the identified proteins could provide understanding on the adverse effect ofCentella asiaticain male reproductive system.

Vegetative Growth and Molecular Identification of Fusarium equiseti Isolated from Wilt Disease of Centella asiatica L. in Bangladesh

Centella asiatica (L.) is commonly known as Thankuni plant and has ethnobotanical importance in Bangladesh. Present experiment was conceded to investigate the wilt disease of C. asiatica, vegetative growth and molecular characterization of pathogenic fungi. Pathogenic fungus, Fusarium equiseti was identified as a causal agent of wilt disease in C. asiatica. The effect of culture media on the mycelial growth of F. equiseti showed the highest (89.25 mm) on potato dextrose agar (PDA) medium followed by carrot agar (CA) medium and the lowest growth (40.25 mm) was measured in HA medium. The optimal temperature and pH for mycelial growth of F. equiseti were 30&deg;C and 7, respectively. The genetic variation of the selected species of fungi, the internal transcribed spacer (ITS) region was amplified using ITS4 and ITS5 primers and sequenced. The PCR product of the ITS region of F. equiseti was 535 bp. Phylogenetic tree of thirty-seven strains of Fusarium sp. based on the nucleotide sequences of the ITS region using the neighbor-joining method with 1000 bootstrapping indicated that 98% - 100% identity with MN886590.1 JUF0046 (F. equiseti). ITS sequences are generally constant, or show little variation within species, but vary between species in a genus. The ITS region is relatively short and can be easily amplified by PCR using universal single primer pairs. Genetic distance exhibited high level of similarity with identical ITS sequences. To date, no published research articles are found on the molecular identification of F. equiseti, the causal agent of fusarium wilt disease of C. asiatica in Bangladesh.

Anti-fatigue Effect of Water Extract from Centella asiatica(L.)Urban on Mice

[Objectives]To study the anti-fatigue effect of water extract from Centella asiatica(L.)Urban on mice.[Methods]After intragastric administration of low,medium and high concentration of water extract from Centella asiatica(L.)Urban for 14 d,the rotarod time and the content of serum lactic acid in mice were determined,respectively.[Results]Compared with the control group,the rotarod time of mice in the low and medium concentration groups was significantly prolonged(P<0.05),but there was no significant difference between the control group and the high concentration group;the content of serum lactic acid in the medium concentration group was significantly lower than that in the control group(P<0.01),but there was no significant difference between low concentration group and high concentration group and the control group.[Conclusions]A certain concentration of water extract from Centella asiatica(L.)Urban had a good anti-fatigue effect.

积雪草主要三萜类化合物测定方法的优化研究

【目的】建立以环糊精为流动相添加剂的高效液相色谱法测定积雪草药材中三萜类化合物积雪草苷、羟基积雪草苷和积雪草苷B的含量,为控制积雪草的质量提供依据。【方法】采用分子设计方法优选流动相添加剂-环糊精种类,效应面法分析环糊精浓度、流动相中乙腈的比例、流动相pH对分离的影响,优化色谱条件,测定积雪草中积雪草苷、羟基积雪草苷和积雪草苷B含量。【结果】优化后HPLC色谱测定方法以γ-环糊精为流动相添加剂,固定相为CAPCELLPAKC18柱(150mm×4.6mm,5μm),流动相为乙腈-3mmol/L环糊精溶液(含0.10%磷酸)=20∶80(V/V),流速1.0mL/min,柱温25℃,检测波长205nm;羟基积雪草苷和积雪草苷B分离度为4.32;积雪草苷为200~2000ng,积雪草苷B为120~1200ng,羟基积雪草苷为200~2000ng内线性关系良好,方法精密度、重现性、稳定性和回收率良好。【结论】新HPLC色谱测定方法简便、准确、可靠,且经济环保,可用于测定积雪草药材中主要三萜类化合物。

近红外光谱技术在线监测积雪草药材活性成分的大孔树脂分离纯化过程

利用近红外(NIR)光谱技术研究并建立可在线监测积雪草药材活性成分———羟基积雪草苷和积雪草苷的大孔树脂分离纯化过程的方法。收集积雪草药材提取物的50%乙醇大孔树脂洗脱液,并采集其近红外光谱信息,建立上述收集液中羟基积雪草苷、积雪草苷的高效液相色谱(HPLC)含量测定方法;然后,采用偏最小二乘法(PLS)建立上述收集液的NIR光谱与其中羟基积雪草苷及积雪草苷含量的线性关系。建模过程中,以相关系数(R2)、交叉验证均方根误差(RMSECV)为指标,确定用于建模的最优近红外波段和光谱预处理方法,即羟基积雪草苷和积雪草苷模型的最佳建模波段分别为12 000.8～7 499.8cm-1和12 000.8～9 750.3cm-1;R2分别为96.44和96.07;RMSECV分别为0.084 80和0.000 99。将上述模型用于在线监测积雪草药材提取物大孔树脂分离纯化过程中羟基积雪草苷及积雪草苷的含量,结果表明此模型预测效果良好。该方法快速、简便、准确,可用于生产过程中羟基积雪草苷及积雪草苷的在线检测和质量控制。

积雪草化学成分研究

目的研究积雪草Centella asiatica的化学成分。方法采用各种色谱技术进行分离纯化,通过理化性质和波谱分析进行结构鉴定。结果从积雪草中共分离得到14个化合物,其结构鉴定为β-谷甾醇(Ⅰ)、二十六醇辛酸酯(Ⅱ)、山柰酚(Ⅲ)、槲皮素(Ⅳ)、胡萝卜苷(Ⅴ)、香草酸(Ⅵ)、丁二酸(Ⅶ)、积雪草酸(Ⅷ)、羟基积雪草酸(Ⅸ)、ter-minolic acid(Ⅹ)、积雪草苷(Ⅺ)、积雪草二糖苷(Ⅻ)、羟基积雪草苷、积雪草苷-B。结论化合物为一新化合物;羟基积雪草酸和terminolic acid、羟基积雪草苷和积雪草苷-B为两对同分异构体。

高效液相法测定积雪草配方颗粒中积雪草苷的含量

目的:建立测定积雪草配方颗粒剂中积雪草苷的HPLC含量分析方法。方法:采用C18柱,RP-HPLC法,水-乙腈为流动相,梯度洗脱,检测波长205 nm。结果:积雪草苷在0.02-0.51 mg·mL-1范围内,峰面积与浓度呈线性关系,r=0.9999,高、中、低浓度的平均回收率分别为99.6%,101.7%,102.2%;RSD分别为2.9%,1.7%,4.9%(n=3)。结论:该方法简便,结果准确,适用于积雪草配方颗粒剂的质量控制。

酶法提取积雪草苷

采用酶法提取积雪草中的积雪草苷,对酶的种类、用量、积雪草与溶剂水的料液质量比、酶解液初始pH、酶解温度、酶解时间等因素进行了考察,结合正交实验设计筛选出酶解最优工艺参数,并将酶法提取结果与其他方法比较.研究结果表明,优化的提取条件为纤维素酶与底物积雪草质量比m(E)∶m(S)=1∶50,料液质量比为(积雪草与溶剂水的质量比)1∶25,pH 6.0,酶解温度40℃,酶解时间1.5 h.与传统提取方法相比,酶法提取速率快,工艺条件温和,得率高,是一种良好的提取方法.

积雪草提取物对抑郁模型动物行为的影响

目的观察积雪草提取物对抑郁大鼠、小鼠干预后行为学的改变。方法 60只Wistar大鼠随机分为6组,每组1 0只,分别为对照组、模型组、氟西汀组、积雪草提取物高剂量组(成分10.08mg+成分24.48 mg/kg),中剂量组(成分10.04mg+成分22.24mg/kg),低剂量组(成分10.02 mg+成分21.12 mg/kg)。采用慢性不可预知性温和应激刺激制备大鼠抑郁模型,糖水消耗量检测积雪草提取物抗抑郁效应。60只昆明小鼠,随机分为6组,分组情况同上。自主活动、强迫游泳、悬尾实验检测积雪草提取物对小鼠抑郁样行为的影响。结果积雪草提取物高、中、低剂量组能够明显增加大鼠糖水消耗量,与模型组比较(P<0.01)。积雪草提取物高、中、低剂量组能明显缩短悬尾实验和强迫游泳小鼠不动时间。结论积雪草提取物对大、小鼠抑郁样行为有一定的改善作用。

肠杆菌FM-1强化积雪草修复镉污染土壤机理

分析了肠杆菌FM-1的耐镉性及促生特性,并研究了肠杆菌FM-1对矿山型和非矿山型积雪草镉富集及根际土壤pH值的影响.结果表明,肠杆菌FM-1对镉有较强的耐受性,其分泌的吲哚乙酸含量,铁载体A/Ar值和溶磷能力分别达到(72.85±0.62)mg/L,0.21±0.01和(143.33±2.13)mg/L,表明肠杆菌FM-1具有良好的促生能力.将肠杆菌FM-1接种于采自广西柳州泗顶铅锌矿周边4种类型的土壤中,均不同程度提高了矿山型和非矿山型积雪草对镉的富集.其中,下游区土壤接种肠杆菌FM-1后,矿山型积雪草茎和叶中镉含量分别增加了87.90%～161.84%和21.85%～76.42%,非矿山型叶片中的镉含量增加了5.84%～35.20%.研究表明肠杆菌FM-1促进了积雪草对镉的富集.同时,肠杆菌FM-1的添加在一定程度上影响了积雪草根际土壤的pH值,显著降低了下游区矿山型积雪草根际土壤中的pH值.

4种中药体外抗结核分枝杆菌H_(37) RV的研究

目的研究中药夏枯草、积雪草、筋骨草和青蒿素的体外抗结核分枝杆菌H37RV的活性。方法采用改良罗氏固体培养基试管培养法进行活性检测。结果夏枯草醇提物抗结核效果明显;积雪草苷片、筋骨草及青蒿素均无效。结论夏枯草醇提物抗结核菌效果优于水提物;积雪草醇提物有微弱抑制结核杆菌活性,但其有效成分可能不是积雪草苷;青蒿素及二氢青蒿素无抑制结核分枝杆菌的活性。

制备型高效液相色谱法从积雪草提取物中分离纯化积雪草甙和羟基积雪草甙对照品

积雪草甙和羟基积雪草甙是积雪草及其相关制品质量控制的两个指标成分,本研究利用制备型高效液相色谱从积雪草提取物中同时分离纯化得到这两个成分。对制备色谱的流动相组成、流速、进样量和检测波长等制备参数进行了优化。采用的色谱柱为C18柱(50mm×200mm,5μm);流动相为甲醇-水(体积比为60∶40),流速100mL/min;二极管阵列检测器在220nm检测;进样体积为1.5mL。在20min的运行时间内,积雪草甙和羟基积雪草甙与干扰成分得到很好的分离,产品纯度达到98%以上。此方法具有快速高效、产品纯度高的特点,可以用于制备积雪草甙和羟基积雪草甙对照品。

积雪草提取纯化工艺的研究

目的研究积雪草的提取纯化工艺。方法采用正交试验进行工艺优化,薄层色谱扫描法测定羟基积雪草苷含量。结果提取工艺最佳条件为:水提取2次,第1次加10倍量水提取2h,第2次加8倍量水提取1.5h。纯化工艺最佳条件为壳聚糖用量0.4mL/g,药液浓缩比例1.2:1,絮凝温度70℃。结论本方法可节省原料,缩短工时,降低成本,利于工业化大生产。

不同产地积雪草中的微量元素比较研究

用火焰原子吸收分光光度法测定了７个不同产地的中药材积雪草中８种微量元素Ｆｅ、Ｃｕ、Ｚｎ、Ｍｎ、Ｃｏ、Ｃｄ、Ｓｒ、Ｃｒ的含量，并进行了比较，结果表明中药材中微量元素含量因地而异。为药材道地性研究、药理作用探讨及临床合理用药提供了依据。

积雪草中积雪草酸的分离、纯化及其含量测定

目的分离纯化积雪草中积雪草酸,并建立高效液相色谱法(high-performance liquid chromatography,HPLC)测定积雪草酸的含量。方法超声提取积雪草中积雪草酸,将粗提物用石油醚-丙酮体系在硅胶柱上梯度洗脱,HPLC法测定积雪草酸的含量。色谱条件:采用Waters Symmetry C18色谱柱(4.6 mm×250 mm,5μm),流动相为乙腈-10 mmol.L-1乙酸铵溶液(38∶62,V/V),检测波长为210 nm,流速为1.0 mL.min-1,柱温25℃,进样体积为20μL。结果积雪草酸在浓度10～200 mg.L-1线性关系良好(R2=0.999 5),其日内、日间RSD均小于5.4%,回收率为100.5%。积雪草中积雪草酸含量为0.99 g.kg-1。经提取、分离、纯化得到积雪草酸白色粉末,纯度为79.0%。结论本实验所用分析方法简便、准确、重复性好,可用于积雪草酸的含量测定;所用提取分离及纯化积雪草酸的方法可使中药中积雪草酸得到有效富集。

民族药届招觅的生药学鉴定

目的对多民族常用的植物药届招觅进行生药学鉴定,为其鉴别及应用提供科学依据。方法采用原植物、性状、显微鉴别的方法。结果届招觅在原植物、性状、显微等方面具有专属性的特征。结论通过原植物、性状、显微鉴别能够很好地鉴定该药材。

低糖型积雪草饮料的研制

以积雪草为主要原料,通过感官评定和正交设计试验,筛选出最佳配方并进行稳定性研究。结果表明,以积雪草浸提液与水的比例1∶4、蛋白糖浓度0.2%、柠檬酸浓度0.1%和CMC浓度0.1%为最佳配方。研制的低糖型饮料,在营养、色泽、风味、口感上均有很好的表现。