Mechanism And Safety Evaluation Of The Insect-resistant Mung Bean To Callosobruchus Chinensis(L.)

Mungbean[Vigna radiata(L.)],a kind of edible bean,is planted fairly largely in China.Its quality and yield are seriously affected by Callosobruchus chinensis(L.).The traditional physical and chemical controlling methods not only increase the cost,but also affect the health of human beings and the survival environment.The use of the insect-resistant mungbean is the most economical and effective way to prevent this damage.Up to now,there has been no report about mechanism and safety evaluation of the C.chinensis(L.)of insect-resistant mungbean.Therefore,using different mungbean varieties as experimental material,this study filtered variety of insect-resistant mungbean and explored the change of enzymes activities serving for digestion,protection,and detoxification after treatment of C.Chinensis(L.)with insect-resistant mungbean.Using two-dimensional electrophoresis(2-DE)and mass spectrometry identification technology,it analyzed different protein expression of the function in the insect-resistant and insect-sense mungbean and conducted the subchronic toxicity study of insect-resistant mungbean on mice,aimed at clarifying the biochemical mechanism and safety of insect-resistant mung bean on the C.chinensis(L.),hoping to provide theoretical basis for the exploration and utilization of insect-resistant mungbean ingredients.The results are showed as the following:Experiment 1:The results showed that among the 13 mungbean varieties 6 are high-resistant,including B18,B20,B23,B27,A22 and Jinlv no.7.Seed's Dam Rate(SDR)of these mungbean varieties is below 10%,while SDR of the other 7 varieties are above 90%,which is high suspect mungbean.There is no significant difference in rate of egg hatch among all mungbean varieties.Compared to the control group,the larval weight,the weight of the male and female adults and the adult eclosion rate of C.chinensis(L.)treated with insect-resistant mungbean was 0.69,0.14,0.69 and 0.71 times sespectively.There is no significant effect on the hatch,adult eclosion rate and SDR of C.chinensis(L.)for the same variety,whether it's peeled or unpeeled.Experiments with artificial mungbean showed that the rate of adult eclosion gradually decreased with the increase in the protein ratio in insect-resistant mungbean,but did not alter with the increase of the proportion of starch.Experiment 2:In this experiment,the activities of enzymes serving for digestion,protection,and detoxification of larva treated with insect-resistant mungbean were analyzed.Results showed that the activity of total protease,a-amylase,and a-glucosidase in intestine of larva were significantly lower than the control group after treatment with insect-resistant mung bean for 8 to 16 days,with a concomitant increase in activity of superoxide dismutase(SOD),peroxidase(POD),carboxylesterase(CarE),and GSH-S-transferase(GST)for treatment for 8 to 12 days.However,no significant difference was observed in the activities of CarE and GST for treatment for 14 to 16 days.These results indicated that insect-resistant mungbean could disturb the normal development of C.chinensis(L.)larva by suppressing the digestive enzyme activity and adversely affecting the activities of protective and detoxifying enzymes.Experiment 3:By comparing the two-dimensional electrophoresis(2-DE)of resistant and susceptible mung beans,this study found there were altogether 15 points that showed differential protein expressions of resistant mungbean were higher than susceptible mung beans.Mass spectrometry showed that 6 among the 15 protein spots were known functional proteins and the functional proteins identified were alpha and beta subtype 8 s globulin as nutrient storage protein,Ribulose-1,5-bisphosphate carboxylase/oxygenase,(RuBis CO)subunits binding protein,precursor peptide chains for amylase inhibitor and trypsin inhibitor.The two inhibitor may inhibit protease and amylase of the insect.The function of two protein spots were unknown yet.The other seven protein point were not verified in UniProt database.Experiment 4:In this experiment,we aimed to evaluate the safety of insect-resistant mungbean Jinlv no.7 on mice with different proportions.Results showed that the insect-resistant mungbean is safe as animal feed during the period.Because the mice not only were healthy from the appearance,but their blood biochemistry and the organ development were not significantly different from that of the control group.In addition,results of histopathology examination showed no obvious abnormalities in the major organs of mice.The sequencing results of liver and kidney for cancer and apoptosis related gene showed no mutation occurred.The normal growth and development of the C.chinensis(L.)in mungbean Jinlv no.7 can is influenced by inhibiting the activities of intestinal digestion enzymes of larvas,but it is safe to mice,which would provide theoretical basis for the insect-resistant mechanism of insect-resistant mungbean as well as references for edible safety and commercial prospects to insect-resistant mungbean Jinlv no.7.