Antiviral And Antioxidant Activity Of Aqueous Extract Of Rhizome Of Phyllostachys Nigra(Lodd.ex Lindl.) Munro

Phyllostachys nigra(Lodd.ex Lindl.)Munro is a plant of Phyllostachys of Gramineae.The plant is mainly distributed in the provinces of Zhejiang,Jiangsu,Anhui,Hubei,Fujian and Shanxi.According to the records in "Cao Mu Simple Prescription" and "Herbs in Chongqing",the rhizome of the plant was used for dehumidification and detoxification.Previous studies suggested that Phyllostachys nigra(Lodd.ex Lindl.)Munro contained water-soluble-polysaccharidesin,flavonoids,lactones,phenolic acids and so on.Phyllostachys nigra(Lodd.ex Lindl.)Munro has functions in apoptosis inhibition and vasodilation properties.However,there were few researches that have been done on the antiviral and antioxidant effect of the material.Therefore,this study investigated the antiviral and antioxidant effect of aqueous extract of rhizome of Phyllostachys nigra(Lodd.ex Lindl.)Munro.Experiment 1 Preparation of aqueous extract of rhizome of Phyllostachys nigra(Lodd.ex Lindl.)Munro and determination of safety Aqueous extract of rhizome of Phyllostachys nigra(Lodd.ex Lindl.)Munro(PNR)was extracted by water decoction.In order to evaluate the safety of PNR,the acute toxicity and sub-chronic toxicity test were conducted.The results of acute toxicity test showed that the mice had some untoward reactions half an hour after administration,such as depressed spirit and anorexia,but they became normal quickly.There was no death in all groups within 14 days,and the maximum dosage was 5000 mg/kg.During sub-chronic toxicity test,the body weight of mice in high dose group was decreased,but there were no significant differences between each group in later period.The total white cell count of medium dose group was significantly more than control group and the Lymphocyte rate of medium dose group was significantly lower than control group.There were no significant differences in ALT,AST,TG and UN among all groups.There were no significant differences in histopathology among all groups.These results indicated that PNR had no toxicity.Therefore,the clinical application of aqueous extract of rhizome of Phyllostachys nigra(Lodd.ex Lindl.)Munro was safe.Experiment 2 Effects on inhibition of VSV of PNR in vitro Cytotoxicities of PNR on Vero cell lines were evaluated by MTT method.The percent protection of PNR were examined by MTT method in three ways and VSV inhibitory rates on Vero cells were measured by plaque reduction assay.Ability of PNR to inhibit replication of VSV-G gene was determined by real-time PCR.Morpholine hydrochloride(ABOB)was used as positive control.The result showed that safe concentration of PNR on Vero cells was 500μg/mL.PNR had good percent protection in three ways and PNR had the best percent protection when given after VSV inoculation.In pre-treatment assay,simultaneous treatment assay and post treatment assay,the IC50 values of PNR were 511.8 μg/mL,38.6 μg/mL and 28.5μg/mL,respectively,and the SIs were 3.3,12.0 and 58.9.The result of plaque reduction assay matched the test of percent protection.PNR inhibited replication of VSV-G gene,so that the viral proliferation were inhibited.Experiment 3 Effects of PNR on inhibition of VSV in vivo The ICR mice were randomly divided into six groups(n=10):Normal control group(Normal-C),VSV-infected control group(VSV-C),ABOB treated group(ABOB),low,moderate and high dose of PNR treated group.The mice were intranasally infected with 1.4×105 TCID50 of VSV in a total volume of 0.01 mL to both nostrils except Normal-C.And the Normal-C group was treated with physiological saline at the same time.Two hours later,the treatments of PNR were implemented.In the ABOB group,mice were infected and received the treatment of ABOB at 0.09 g/kg bw/d.In other treatment groups,mice were treated with PNR at low(0.1 g/kg bw/d),moderate(0.2 g/kg bw/d)and high(0.4 g/kg bw/d)does,respectively.Mice in Normal-C and VSV-C group were treated with the normal saline.Physiological condition and body weight were recorded daily,respectively.The treatments were administered daily via gavage for 6 d.The lung,spleen,and brain were excised,fixed and examined by H&E staining.And western blot was analyzed.The results showed that PNR increased the body weight of mice infected by VSV.The pathological damages of lung,brain and spleen tissues in infected mice were reduced in a dose-dependent manner.And PNR increased the secretions of antiviral protein IFN-α,IFN-γ and Mx2 in a dose-dependent manner.Experiment 4 The research of antioxidant activity of PNR in vitro The antioxidant activity of PNR was determined with experiment of the scavenging power of DPPH,hydroxyl radical,superoxide anion and the total reducing power.Vc was used as the positive control.As the results shown,in the superoxide anion scavenging assay,the IC50 values of PNR and Vc were 31.96±0.27 μg/mL and 39.32±1.19 μg/mL,respectively.In the DPPH radical scavenging,hydroxyl radical scavenging and total reducing power assay,the IC50 values of PNR were 83.34±3.20 μg/mL,237.82±0.95 μg/mL and 169.25±3.98 μg/mL,respectively,while the IC50 values of Vc was 19.58±0.18 μg/mL,153.46±0.22 μg/mL and 103.4±5.21μg/mL,respectively.