Molecular Identification of Alternaria alternate (Fr.) Keissl. from the Leaf Blight Disease of Centella asiatica L.

Centella asiatica (L.), frequently known as Thankuni, is an important ethnobotanical plant in Bangladesh. This study was conducted to evaluate the morphological characteristics, cultural factors and molecular identification of the causal agent of Alternaria leaf blight disease of C. asiatica. The potato dextrose agar (PDA) medium recorded the maximum mycelial growth (69 mm), followed by the yeast extract agar (YEA) medium, while the honey peptone agar (HPA) medium recorded the lowest growth (27 mm). The optimal pH and temperature for mycelial growth of Alternaria alternata were 6 and 30°C, respectively. Internal transcribed spacer (ITS) region of Alternaria alternata PCR products measured 558 bp and blast search showed 99% sequence similarity with Alternaria alternata species complex. To the best of our knowledge, Alternaria leaf blight disease caused by Alternaria alternata is the first record in Bangladesh.

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.

Production of asiaticoside from centella(Centella asiatica L.Urban)cells in bioreactor

Objective:To investigate the effects of some culture conditions on production of asialicoside from centella(Centella asiatica L.Urban)cells cultured in 5-L bioreactor.Methods:The centell cell suspension culture was conducted in 5-L bioreactor to investigate the growth and asiaticoside accumulation under various conditions.Asiaticoside content was determined by HPLC analysis.Results:The results showed that the cell growth and asiaticoside accumulation peaked after 24d of culture at an agitation speed of 150 r/min and aeration rate of 2.5 L/min.The cell biomass reached a maximum value of 302.45 g fresh weight(31.43 g dry weight)and growth index of 3.03with inoculum size of 100 g.However,asiaticoside content was the highest(60.08 mg/g dry weight)when culture was initiated with an inoculum size of 50 g.Conclusions:The present study found the suitable conditions for growth of centella cells and their asiaticoside production in bioreactor.

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°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.

车前(lantago asiatica L.)毛状根诱导技术研究

为建立发根农杆菌Ri质粒介导的车前(lantago asiatica L.)转化体系,通过发根农杆菌A4和R1601侵染车前无菌苗的外植体诱导毛状根.实验结果表明,车前外植体最佳侵染时间为8 min;两种菌液均可以诱导车前叶片、叶柄和根产生毛状根,车前叶片和叶柄毛状根的诱导率低于40%,车前根部毛状根的诱导率高于80%;经过分子鉴定,Ri质粒T-DNA的rolB基因已成功整合到毛状根中,表明车前的根是适宜毛状根诱导的最佳外植体,可为今后车前毛状根的扩大培养奠定基础.

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.

Attenuation of Collagen Induced Arthritis by Centella asiatica Methanol Fraction via Modulation of Cytokines and Oxidative Stress

Objective To investigate the anti-inflammatory, antioxidant and anti-arthritic effects of Centello asiatica methanolfraction （CAME） on collagen-induced arthritis （ClA）, an animal model of rheumatoid arthritis. Methods Arthritis was induced in female wistar rats by immunization with porcine type II collagen. The CIA rats were treated orally with CaME （50, 150, and 250 mg/kg/day） for 15 d （beginning on day 21 of the experimental period）. The clinical, histological, biochemical, and immunological parameters were assessed. Results CaME treatment （150 and 250 mg/kg） significantly attenuated the severity of CIA and reduced the synovial inflammation, cartilage erosion, and bone erosion as evident from both histological and radiographic data. The escalated plasma levels of pro-inflammatory cytokines TNF-α, IL-1β, IL-6, and IL-12 alongwith nitric oxide in CIA rats decreased significantly on CaME treatment. The serum levels of type-Ⅱ collagen antibody were significantly lower in rats of CaME （150 and 250 mg/kg） treated group than those in the arthritic group. Furthermore, by inhibiting the above mediators, CaME also contributed towards the reversal of the disturbed antioxidant levels and peroxidative damage. Conclusion Our results clearly indicate that oral administration of CaME suppresses joint inflammation, cytokine expression as well as antioxidant imbalance, thereby contributing to an amelioration of arthritis severity in CIA rats.

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.

Structural Features of a Polysaccharide from Centella asiatica

A polysaccharide, isolated from Centella asiatica, was a complicated arabinogalactan (AG), which contained a little a-(14)-linked GalpA and a-(12)-linked Rhap residues. Based on composition and methylation analyses, partial acid hydrolysis, periodate oxidation, NMR, ESI-MS experiments, it was shown to have a backbone of b-(13)-linked D-Galp residues, with heavily branched side chains. Araf residues were linked to O-3 of (13, 6)-linked Galp residues.

Antidiabetic effect of combination of fractionated-extracts of Andrographis paniculata and Centella asiatica: In vitro study

Objective: To examine the effect of combination of Andrographis paniculata herb fraction(AHF) and Centella asiatica herb fraction(CHF) on PPAR γ and GLUT4 mRNA expression in 3 T3-L1 adipocyte, and its effect on insulin-stimulated glucose uptake. Methods: 3 T3 L1 adipocyte cells were used to investigate gene expression of PPARy reverse transcription-polymerase chain reaction method. The γ and GLUT4 protein b adipocyte cells were differentiated by using insulin, dexamethasone and 3-isobutyl-1-methylxanthine from 3 T3-L1 cells. Pioglitazone, AHF, CHF and the combination of both herbs were evaluated on glucose uptake activity, PPAR γ and GLUT4 mRNA expressions in 3 T3-L1 adipocyte. Results: The results showed that combination of AHF at 30 μg/mL and CHF at 10 μg/mL could enhance insulin-stimulated glucose uptake. The combination also increased PPAR γ and GLUT4 mR NA expressions significantly in comparison to those of negative control(DMSO). These effects were equal in comparison to those of pioglitazone(0.02 μM) and its single extracts Conclusions: The combination of AHF and CHF can increase glucose uptake and insulin sensitivity through up-regulation of PPAR γ and GLUT4 mRNA expressions in 3 T3-L1 adipocyte.

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.

Comparative Efficacy of <i>Tagetes erecta</i>and <i>Centella asiatica</i>Extracts on Wound Healing in Albino Rats

The study was undertaken to evaluate the comparative efficacy of hydroalcoholic extracts of leaves of Tagetes erecta (T. erecta) and aerial parts of Centella asiatica (C. asiatica) on Excision, Incision and Dead space wound models in albino rats. Extract of T. erecta and C. asiatica (P T. erecta extract (250 and 500 mg/kg) showed significantly increased the wound breaking strength in incision wound model and wet and dry granulation tissue weights, breaking strength in a dead space wound model compare to control and C. asiatica treated group (P T. erecta extract showed potent wound healing activity then the reported C. asiatica in different wound parameters.

Comparative study of standardized Centella asiatica and combination of levodopa and benzerazid HCl on Parkinson′s model zebrafish

OBJECTIVE To investigate the motility,dopamine,serotonin and pro-BDNF level of Centella asiatica(CA)comparable to the combination of levodopa and benzerazid HCL on zebrafish model Parkinson′s.METHODS Rotenone 5μg·L-1 induced to adult zebrafish(8months)for 28 dto made zebrafish model Parkinson′s.CA concentration were used 5 and 10μg·mL-1.The combination of levodopa 100 g and 25 mg benzerazid(L-DOPA agonist)was given at 1 μg·mL-1 at the same time to rotenone,and CA as well.Motility assessment conducted for every 7dstarted on day 0until day 28.Five fish were subjected in the 2L tank(25cm×16.5cm×12cm).Three vertical lines were drawn on the tank at equal distances,dividing the tank into four zones(the length of each zone was 6.25cm).Locomotor activity was measured for 5min by counting the number of lines that adult zebrafish crossed.Fish were sacrificed by decapitating on ice water.Dopamine level measured from whole brain by ELISA,serotonin and pro-BDNF by immunoreactivity at substantia nigra.RESULTS The locomotor activity of rotenone treated fish were significantly decrease starts at 7 dcompared to control group.Interestingly on 10μg·mL-1 group there are increasing motility start at day 7 and slightly decrease until day 28,but on L-DOPA group increasing motility at day 7 followed by decreasing motility significantly(P<0.05)until day 28.Dopamine level of rotenone group decreased compared to control group,CA 10μg·mL-1 and L-DOPA group increased significantly to rotenone group which CAμg·mL-1 higher than L-DOPA group.Serotonin and pro-BDNF level had the same profile.Its significantly increased on rotenone group compared to control group.Serotonin on group with CA administration significantly decreased compared to rotenone group,while rotenone plus CA 10μg·mL-1 had no significant differet to L-DOPA group(P>0.05).Pro-BDNF on rotenone with CA 5μg·mL-1 had no significant difference,while compare to rotenone with CA 10-1 μg·mLgroup and L-DOPA group were significantly different(P<0.05).CONCLUSION Rotenone produces reliable Parkinson′s zebrafish model by decreasing motility and dopamine level.CA was more stable,increasing motility than L-DOPA administration.CA and L-DOPA increased dopamine level,but decreasing serotonin and pro-BDNF.

Decreasing a-synuclein aggregation by methanolic extract of Centella asiatica in zebrafish Parkinson's model

Objective: To observe the effects of Centella asiatica(C. asiatica) methanolic extract on a-synuclein aggregation and its expression in rotenone-exposed zebra fish.Methods: Zebra fish(Danio rerio) were exposed to 5 m g/L rotenone for 28 days and coincubated with 2.5, 5.0 and 10.0 m g/mL of C. asiatica methanolic extract. The medium was changed every 48 h for maintain the concentration of rotenone and extract. After 28 days zebra fish were sacrificed on the ice block and protein was isolated from zebra fish brain for ELISA of dopamine and Western blotting of a-synuclein. Immunohistochemistry was conducted to observe the a-synuclein expressions from histopathological preparation of zebra fish brain. The head were soaked in 10% formaline for less than 24 h and embedded onto paraffin block, then sliced for immunohistochemistry using anti a-synuclein antibody. We also measured zebra fish motility for 5 min in each week.Results: C. asiatica has important bioactive compounds such as asiaticoside that has antiin flammatory and antioxidant properties. It may inhibit cascade reaction due to oxidative stress induced by rotenone. Decreasing reactive oxygen species proposed probability of radical attack to a-synuclein protein that caused aggregation and increase of its expression.The motility of zebra fish was also maintained in C. asiatica groups due to the increasing dopamine level in rotenone-induced zebra fish. High level of reactive oxygen species inactivated enzyme for dopamine synthesis such as tyrosine hydroxylase, and oxidized dopamine itself. Oxidized dopamine increased a-synuclein aggregation. Thus, the dopamine level decreased in rotenone-induced zebra fish, but C. asiatica increased dopamine level.Conclusions: C. asiatica has a potential to be developed as an anti-Parkinson's disease treatment due to its capability for minimized the sign of Parkinson's such as a-synuclein aggregation and expression, increasing motility and dopamine as well.

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.

Standardized extract of Centella asiatica ECa 233 inhibits lipopolysaccharide-induced cytokine release in skin keratinocytes by suppressing ERK1/2 pathways

Objective:To evaluate the effect of standardized extract of Centella asiatica ECa 233 on inflammatory mediator production through cyclooxygenase-2(COX-2),extracellular signal-regulated kinase 1/2(ERK1/2)and nuclear factor-κB(NF-κB)pathway in keratinocyte Ha Ca T cells.Methods:Ha Ca T cells were treated with 0.1,1,10 and 100μg/m L ECa 233 in the presence of lipopolysaccharide(LPS).Proinflammatory cytokines and prostaglandin E2 were assessed with ELISA.Western blotting was used to determine the inhibition of COX-2,ERK1/2 and NF-κB protein expression.Results:ECa 233 suppressed LPS-induced release of interleukin-1β,tumor necrosis factor-α,and prostaglandin E2.ECa 233 also inhibited COX-2,phosphorylation of ERK1/2 and the activation of NF-κB.Moreover,the formation of reactive oxygen species(ROS)was decreased in response to LPS-inflamed keratinocytes.Conclusions:ECa 233 inhibits LPS-stimulated production of inflammatory mediators in keratinocytes via suppressing ERK1/2 and NF-κB pathways.The suppressive effect of ECa 233 may be related to an inhibition of ROS production.

The efficacy of topical spray containing Centella asiatica extract on excision wound healing in rats

Centella asiatica (Linn.) belongs to the family of Apiaceae(Umbeliferae). It has been widely used for traditional medicine, especially for wound healing, anti-inflammatory,antipsoriatic, antiulcer, antibacterial and antifungal. The biologically active ingredients of Centella asiatica are believed to be triterpenes, particularly asiatic acid, madecassic acid,asiaticoside and madecassoside (Fig. 1).

Enzymatic pretreatment and microwave extraction of asiaticoside from Centella asiatica

The extraction of asiaticoside from Centella asiatica by enzymatic pretreatment and microwave extraction (EPME) was studied in this article. The effects of several important factors such as temperature of enzymatic pretreatment, liquid to solid ratio and microwave radiation time were investigated by quadric regression orthogonal design experiment and were analyzed by response surface. An extraction model with well forecast performance was then established. The results indicate that the optimum extraction condition was as follows: liquid to solid ratio was 36mL/g, temperature of enzymatic pretreatment was 45℃, enzymatic time was 30min, and microwave ra-diation time was

A Phytosociological Study of Hydrocotyle mannfi and Centella asiatica Weed Community in Kampala （Uganda, Eastern Africa）

A phytosociological study of a weed community dominated by Hydrocotyle mannii and Centella asiatica was carried out in Kampala, the capital city of Uganda from 2000 to 2002. The Zurich-Montpellier School of Phytosociology method was used with special emphasis on floristic and synecological structure as well as on dynamic aspects. This community was found to grow on shady moist roadsides and is about 15 cm high. In total, 54 species were recorded; most of them are annual, anemochorous and widespread. The Hydrocotyle mannii and Centella asiatica community is described as a new weed association： Hydocotylo-Centelletum asiaticae Mosango ass. nova.. Hydrocotyle mannii, Centella asiatica, Dichondra repens and Sida veronicifolia have been recognized as the character species of this association. The Hydrocotylo-Centelletum is a pioneer association. It has been ascribed to the alliance Eleusinion indicae Leonard 1950 which includes plant communities growing on shady roadsides. In terms of ecological succession, it appears to be evolving to the Asystasia gangetica herbaceous fallow association which belongs to the alliance Panicion maximi. It belongs to the order Ruderali-Euphorbietalia Schmitz 1971 and the class Ruderali-Manihotetea Leonard in Taton 1949.

Efficacy Study on the Anti-Aging Emulsion Containing Carnosine and Centella Asiatica Extract

A clinical study was conducted on the anti-aging effects of an emulsion containing carnosine and centella asiatica extract to provide a reference for the evaluation and development of anti-aging cosmetics.Sixty healthy females,aged 35~60,were divided into the test sample and placebo groups.Skin elasticity and wrinkles of the participants were analyzed before they used the emulsion and after they used it for 8 weeks.Instrument measurement results showed that after 8 weeks,skin elasticity and wrinkles in the members of the test sample group improved relative to those of the placebo group.Clinical effect assessment results showed that subjects did not experience adverse skin reactions,and their skin wrinkles showed improvement.Self-assessment results showed that the score of the total satisfaction of the test sample was 4.3,and 100%subjects are satisfied with the test sample.Emulsion containing carnosine and centella asiatica extract has a certain anti-aging effect.