Study On Active Constituents From The Roots Of Polygala Tenuifolia Willd Against Dactylogyrus Infecting Fish

With increasing intensification of the aquaculture industry in China,the incidence of diseases caused by viruses,bacteria,fungi,parasites and other undiagnosed and emerging pathogens is also expected to increase proportionately.Monogenean parasites such as Dactylogyrus intermedius which are common pathogenic parasites living on the gills of fish,present the largest group of metazoan fish parasites and are of major importance in the fish pathology.The parasites can cause a serious damage to the host such as lowered growth performance,loss of appetite,and high mortalities,resulting in serious economic losses.Various parasiticides such as praziquantel,mebendazole,and toltrazuril were well known species compounds used to deal with the parasitism and deleterious consequences in aquaculture.However,problems have emerged with the frequent use of these chemical-based agents,notably the development of resistance in parasites,as well as environmental contamination and toxicity problems.Because of the features of botanical drugs such as degradable and eco-friendly,currently,investigators have diverted their approach towards the development of plant-derived anthelmintics.Based on this,in this study a wellknown traditional Chinese medicine(TCM)"Yuan-Zhi"(the root of Polygala tenuifolia Willd.)was put to control D.Intermedius of Carassius auratus and the active component were obtained by using bio-guided isolation method.The structures of the isolated compounds were elucidated by spectroscopic analyses.The results obtained in this work were as follows:1.Crude extracts were obtained from five solvents(petroleum ether,chloroform,ethyl acetate,acetone,methanol)with increasing solvent polarity.The results of the anthelmintic efficacy of the five extracts against D.Intermedius showed that methanol extract exhibited the highest anthelmintic efficacy of 100%at the lowest concentration of 500.0 mg/L.Furthermore,2.Methanol crude extract of P.tenuifolia was successively fractioned with n-butanol and water,and the different fractions were investigated indicating that the n-butanol fraction is responsible for anthelmintic activity with 100%effective concentrations of 200 mg/L.Bioactivity-guided fractionation and isolation of the compounds responsible for anthelmintic activity was carried out with the n-butanol fraction yielding a active compounds.According to in vivo anthelmintic assays,it was found to be 100%effective at the concentrations of 10 mg/L.By MS,1H NMR,13C NMR spectroscopic analysis and comparing the spectral data with the reported values,the compound was identified as 3,4,5-trimethoxycinnamic acid.3.The acute toxicity of methanol extract and the active compound from P.tenuifolia was assayed using the healthy goldfish.The results showed that the LC50 of methanol extract of P.tenuifolia were 775.5 mg/L after 48 h;LC50 of 3,4,5-trimethoxycinnamic acid was 26.3 mg/L after 48 h.