Preliminary Study On Antiviral Effect On Newcastle Disease In Vitro And Relevent Mechanism Of Traditional Chinese Herbal Medicine-Qilankangduyin And Its Compounds

Qilankangduyin(Qilanyin,QLY) was invented by our laboratory (patent number:263651) which was made by pure traditional Chinese herbal medicine. This medicine had been approved to be a new veterinary drug by the nation, and used to be local drug. It was found that QLY had preventive and therapeutic effect on IBDV,NDV and other viral diseases in clinical application. However, as a preliminary praeparatum, QLY offen precipitates easily in storage, and should be further optimized. The effect of preventive and therapeutic on NDV and other diseases needed further confirm and enhance. Moreover, the antiviral mechanism of traditional Chinese herbal medicine needed to interpreted further.Component in Qilanyin had been extracted in this present experiment. Screening technique of antiviral in vitro and plaque formation assay were used to verificate anti-infection effect on NDV.Therefore, the antiviral mechanism was preliminary studied from immune, viral absorption and release three respects respecitively in this experiment.The research included four tests.Test1The maximal safe concentration of Qilankangduyin and its components on CEF The maximal safe concentration of Qilankangduyin and its components on chick embryo fibroblast was studied by tissue culture. The results showed that QLY, TPS, TT, RIA, RIAw and RIAa’s safe concentrations were1000,1000,500,125,31.3and62.5μg/mL respectively. The max safe concentration was lower when the corpuscular destruction rate was used as the evaluation index. Test2Effects of Qilankangduyin and Its components on NDV proliferation in CEF Cytopathic effect inhibition and MTT colorimetric assays were used to detect the antiviral ability of QLY and its components on newcastle disease virus (NDV) in chick embryo fibroblast(CEF) cells.Moreover, the drug’s antiviral ability was identificated by plaque formation assay. The results showed that QLY had the strongest ability of killing NDV directly, the highest inhibition ratio was128.85%. TPS was good at interrupting proliferation of NDV, the highest inhibition ratio was130.23%. TT showed the best ability of killing NDV directly with an inhibition ratio of146.15%. However, RIA, RIAw and RIAa showed aitivirus ability in all the three means and their aitivirus ability were confirmed by the plaque formation assay.In summary, Qilanyin and its components showed better ability in interrupting proliferation of NDV and killing NDV, while the drugs was weak in blocking.Test3Effects of Qilankangduyin and Its components on chicken peripheral-blood lymphocyte proliferation MTT colorimetric assays was used to detect the vatiation of chicken peripheral-blood lymphocyte proliferation with treatments of adding drugs alone and adding drugs with PHA. The results showed that when QLY, TPS, RIA, RIAw and RIAa’s concentration were at62.5,62.5～500,3.9～62.5,2.0and15.6μg/mL respectively with the treatment of adding drugs alone,their A570values were significantly higher than cell contral(P<0.05), the highest proliferation rate were25.26%,30.53%,34.62%,27.05%and25.96%respectively.When treated with PHA, TPS and RIAw at the concentrations of500and3.9～31.3μg/mL respectively were significantly higher than PHA contral (P<0.05), their highest proliferation rates were14.29%and13.85%. The results indicated that TPS and RIA had better effect on simulating lymphocyte alone or in coordination with PHA. They could enhance body immune function.Test4Effect of traditional Chinese herbal medicine component of RIAa on adsorption and release of NDV Effect of RIAa on adsorption and release of NDV was detected by cytopathic effect inhibition and MTT colorimetric assays. Results showed that under the treatment of adding drug and virus simultaneously, the A570values at the concentration of31.3μg/mL when absorbed60min was significantly higher than virus contro(P<0.05).However, the values of all the three concentration when absorbed120min were significantly higher than virus control(P<0.05).With the treatment of adding drug firstly, when absorbed120min, the A570values of RAIa at the concentrations of62.5and31.3μg/mL were significantly higher than virus control(P<0.05). However, the A570values of different absorbed time when the virus was added firstly showed no significant difference with the virus control (P>0.05) In the experiment of adding cell supernatant into CEF again, the A570value of RIAa+NDV was significantly higher than NDV control (P<0.05). Results indicated that absorption time and the methods of adding drugs showed significant effect on the ability of virus infecting cells.RIAa at a certain concentration could inhibit virus absorbing into cells, and protect cells from infecting virus. however, it could not remove virus which had been absorbed on cells.On the other hand, RIAa could remarkablely inhibit virus’s releasing.