Study On Safety Evaluation And Pharmacological Effect In Vitro Of Toxic Mussel Water Decoction

Toxic mussel powder collected in Chinese Veterinary Pharmacopoeia (2010edition) are composed of Scutellaria baicalens60g, Cortex phellodendir20g,Folium isatidis10g, Rhubarb10g. It has the effects of heat-clearing and detoxifying.And it is used to treat the plague of mussel in the clinic. Toxic mussel water decoctionfollows the prescription of Toxic mussel powder, after crushed and sieved withtraditional method to get the water decoction. Our previous experiments showed thatToxic mussel powder has a therapeutic effect on chicken E. coli, however, its moderntoxicological effect and pharmacological effect in vitro are not clear. In this study, weset out to investigate the toxicological effect and pharmacological effect in vitro ofToxic mussel water decoction, Our aim is to provide a basis for further research andclinical application of Toxic mussel water decoction.Acute toxicity test According to the maximum concentration (1g/mL) andthe maximum volume (40mL/kg weight) with intragastric administration for only onetime, and its maximum concentration with intragastric administration for several timesa day, we studied the acute toxicity of Toxic mussel water decoction. The resultsshowed that no changes in mental condition, diet, motion of mice. With intragastricadministration of Toxic mussel water decoction at a dose of the maximumconcentration and the maximum volume by one time, there was no mice died. At adose of40.0g/kg body weight three times within24h, the maximum cumulativedosages up to120g/kg body weight, the mice still did not appear obvious toxic effect,which indicates that Toxic mussel water decoction is actually nontoxic.Sub-chronic toxicity test With intragastric administration of high doses (10g/kg), moderate doses (6g/kg), low doses (2g/kg) of Toxic mussel water decoction towistar rats of3groups for4weeks, and at a dose of10mL/kg body weight one time tomice, we studied the sub-chronic toxicity of Toxic mussel water decoction. The resultsshowed that Toxic mussel water decoction has no impact on health condition, body weight, food intake, water intake, hematology, biochemical parameters of blood,organ coefficient of the wistar rats. These results indicated that Toxic mussel waterdecoction has good safety for clinical application.Antibacterial test in vitro To study the antimicrobial effect of Toxicmussel water decoction, we used MH broth trace dilution method to test the minimum inhibitory concentration of Toxic mussel water decoction to O78standard strains and other three E. coli strains isolated from chicken; We used fullyautomatic identification of bacteria susceptibility instrument to test the sensitivity and the tolerance levels of four strains of E. coli to antimicrobial agents. Weused dig holes method and plate method to study the inhibitory effect of Toxic mussel water decoction to four strains of E. coli. The results showed that the minimum inhibitory concentration of Toxic mussel water decoction are0.5,0.3,0.3,1g/mL, respectively. It indicated that Toxic mussel water decoction hasan inhibitory effect on the tested strains; The results showed that three E. coli strains isolated from chicken were resistance strains, each strain was resistantto four kinds of antibiotics at least. The standard strain was only resistant totetracycline, and sensitive to the rest commonly antibiotics. By using dig holesmethod, we found that the Toxic mussel water decoction has no inhibitory effect on the three drug-resistance bacteria. By using the plate point test, we found that Toxic mussel water decoction has inhibitory effect on strain2(ciprofloxacin resistant strain), and the inhibitory rate was only9.09%.The anti-inflammatory effects test The safety concentration range ofShanhuang water decoction was measured by MTT test. RAW264.7cells werestimulated with LPS, and the same time treated with different concentrations ofShanhuang water decoction. After incubation of12h, the supernatant was collected,then we use ELISA assay method to determined the inflammatory cytokines (TNF-α、IL-6、IL-10) of supernatant to evaluate the anti-inflammatory effect of Shanhuangwater decoction. The results showed that0.5-2mg/mL concentration of Shanhuangwater decoction has no toxicity effect on RAW-264.7cells. According to the result ofELISA assay, we found that Shanhuang water decoction (1-2mg/mL) significantlyinhibited the secretion of TNF-α (p <0.01), remarkably inhibited the secretion of IL-6(p <0.01), and significantly increased the secretion of IL-10(p <0.01). The resultsshowed that Toxic mussel water decoction can not only significantly reduced the secretion of TNF-α and IL-6, but also can significantly increase the secretion of IL-10.It indicated that Toxic mussel water decoction exhibited an anti-inflammatory effect.In summary, Toxic mussel water decoction has no adverse effects on the centralnervous system, respiratory system and cardiovascular system of mice. We can notmeasured the LD50of Toxic mussel water decoction. It has no subchronic toxicity andhas good security on mice. Toxic mussel water decoction not only exhibited ananti-inflammatory effect, but also has inhibitory effect on avian E.coli in vitro. Theseprovide a basis for clinical application of Toxic mussel water decoction.