| Deoxynivalenol (DON) is produced in wheat, barley and corn following infestation by fungus Fusarium in the field and during storage. And DON can cause poisoning symptoms in many animals and human beings. People call this mycotoxin as "vomitoxin", because it frequently arouse vomiting of pigs. DON was the deoxygenated nivalenol derivative, which is commonly found in temperate regions worldwide as a natural contaminant of cereals. DON can damage the cellular and humoral immune function, adversely affect immune function in experimental animals.Astraglus is a kind of widely used traditional Chinese medicine with significantly improving immune function, good diuretic action and detoxification efficacy. The aim of the experiment is to investigate the toxicity of deoxynivalenol (DON) in BALB/c mice, and to study the detoxification effect of astraglus on the deoxynivalenol treated. The purpose is to study the toxicological mechanism of DON further and provide a new ideas in exploring traditional Chinese medicine to reduce or eliminate toxicity of mycotoxins.Test I Preparation and Content Detection of DON.In order to obtain a great deal of DON with high purity and concentration, Fusarium graminearum was cultured in the potato dextrose agar medium and the hypha inoculate rice medium at 28℃ with no light for 4 weeks. After dried at 65℃ and crushed into pieces, DON in the culture was edtracted with acetronile-water (84:16, V/V) and purified by a purification colum. The content of DON was detected by the methods of LC-MS/MS and UPLC. In conclusion, the rice medium could culture Fusarium graminearum with high yield. The concentrations of DON were respectively 1057 μg/g by LC-MS/MS and 1.069 mg/g by UPLC. This method of culturing Fusarium graminearum can harvest DON with high yield and low cost. The method of LC-MS/MS could be used widely to dectect the content of DON in the culture.Test II Experimental study on toxicity of DON on BALB/c miceTo study the acute, subacute and immune toxicity of deoxynivalenol (DON),60 BALB/c mice were randomly divided into 6 groups, including 5 DON treated groups with different dosages and 1 blank control group, to determined the LD50. Based on the LD50,48 BALB/c mice were randomly divided into 4 groups, including 0 (control),0.15 (low dose),0.6 (middle dose) and 2.4 (high dose) mg/kg.bw DON exposed daily via intragastric administration for 4 weeks. The items including body weight, feed consumption, viscera index, histopathological changes and hematological indexes were observed. Lienal specific immunity indexes, including CD4+, CD8+, CD3+, CD19+were detected by Flow Cytometry (FCM). The results showed that the LD50 dosage of DON purification was 3.3 mg/kg by improved Karber’s method. The average weight and food intake of mice in middle and high dose groups was significantly lower than that of control and low dose groups (p< 0.05). In addition, the ratios of kidney and liver weight to the body in high dose group were significantly increased compared with that of control and low dose groups (p< 0.05), however, the ratio of spleen index was remarkably declined (p<0.05). Obviously histopathological changes were exposed in the liver and kidney of high dosage group. Among the blood routine physiological detections, LY% of high dosage group droped significantly, while MO%, GR% and HCT rised evidently(p<0.05).The CD4+/CD8+value in high dose group was significantly decreased(p<0.05). In conclusion, the oral exposure DON at the of 2.4 mg/kg.bw have toxicity and immune inhibition in mice. Liver, kidney are the major target organs of DON toxicity.Test Ⅲ The resistance effect of astragalus on the immunosuppression of BALB/c mice caused by deoxynivalenol48 BALB/c mice were randomly divided into four groups, including blank control group, astraglus group, DON group an astraglus-DON group.1 week in advance, Mice in the astraglus group and astraglus-DON group were exposed to astraglus via drinking water at the dosage of 0.02g/mL for 5 consecutive days, then normal water for further two days. The treatment was carried for 4 consecutive weeks. Mice in DON group and astraglus-DON group were treated with DON at the dosage of 2.4 mg/kg.bw every day as intragastric administration for 28 days. Mice in blank control group and astraglus group were treated with same volume of saline. The indicators including body weight, feed consumption, viscera index, histopathological and hematological indexes were observed. Lienal specific immunity indexes CD4+, CD8+, CD3+, CD19+ were detected by Flow Cytometry. The values of cytokines in serum, including IL-1β, IL-2, IFN-γ, TNF-α, TGF-β were measured by kits, mRNA expressions of aboved cytokines genes were measured by Real-time PCR. The average weight, feed consumption and the spleenic index in astraglus-DON group were significantly higher than that of DON group (p<0.05). The kidney and liver indexes in astraglus-DON group were significantly lower than that of DON group (p < 0.05). In addition, the histological changes of the kidney and liver in astraglus-DON group were significantly weaker. Moreover, compared to DON group, the values of CD4+/CD8+, CD19+ and the concentrations of cytokines of IL-2, TNF-a, IFN-γ and TGF-β in astraglus-DON group were significantly increased(p<0.05), the mRNA expressions of cytokines genes of IL-2, TGF-β, IFN-γ in astraglus-DON group were also increased remarkably compared to the DON group. In conclusion, astraglus can resist the toxicity and reduce the damages of the liver, kidney caused by deoxynivalenol, improve the immunofunction and reduce the death of mice. |
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