High-performance Liquid Chromatography (HPLC) And DNA Fingerprint Technology To Detect Pests In Wheat Flour

In the milling industry, insect infection seriously influence qualities of the cereal products, including lower of the processing quality?nutritional quality?sensory quality etc. However, there still lacks an efficient?rapid and economic test method for insect detection in the wheat flour in present. In order to develop a new rapid detection method, the determination of uric acid content of wheat flour by high performance liquid chromatography to determine the degree of infection of insect pests in wheat flour were carried out in this paper. Detection of wheat powder by DNA fingerprinting technology of Tribolium confusum(Duval) and Oryzaephilus surinamensis(Linnaeus) was also carried out in this paper. The main results in this paper were as follows:Detection limitation of uric acid standard solution by the HPLC method was 1?g/m L, that of uric acid content in wheat flour was 1.48 ?g uric acid/g wheat flour. Results from HPLC method showed that uric acid content in wheat flour infested with adults of T. castaneum and O. surinamensis was higher than that infested with larvae ones of corresponding species. There were a good correlation between uric acid content in wheat flour and insect infection density and infection time. The use of design expert software and the method of response surface analysis content of uric acid(Z), pest infection time(X), with population density(Y) relationship. After data processing, the regression equation with adults of Tribolium castaneum is:Z=10.35+4.42 X+6.11 Y+1.32 XY-0.06 X2+1.06Y2; the regression equation with larvae of T. castaneum is: Z=4.35+2.78 X+2.34 Y-0.48 X Y+0.45 X2+0.55 Y2. The prediction accuracy of insect infection density through detecting uric acid content in wheat flour reached 83.0%~99.6%. Experimental data of wheat flour sampling in flour mill also showed there was good correlation between infection numbers of insects and uric acid content in wheat flour. These suggest that method of determination of uric acid content in wheat flour by high performance liquid chromatography to predict the degree of insect infection in wheat flour is feasible.Based on the specificed elongation factors gene of insects in the flour, the degenerate primers was designed, and elongation factors DNA fragment in T. confusum and O. surinamensis were cloned. Different content of T. confusum and O. surinamensis eggs, larvae and adult according 20%, 10%, 5%, 3%, 1%, 0.5%, 0.1%, 0.05% respectively to 50 mg wheat flour by mass were detected by DNA fingerprinting technology. The results showed that the lowest detection limitation for T. confusum adult fragments is 1% of 50 mg wheat flour weight; the lowest detection limitation for T.confusum eggs and larvae is 0.05% of 50 mg wheat flour weight. The lowest detection limitation for O. surinamensis adult fragments is 1% of 50 mg wheat flour weight. The lowest detection limitation for O. surinamensis eggs and larvae is 0.05% of 50 mg wheat flour weight. Good correlation between insect content(X) and brightness of DNA agarose gel electrophoresis banding(Y) was Y=858.36X+74.540, coefficient of correlation is 0.9619. The prediction accuracy of insect infection density through detecting DNA electrophoresis banding brightness in wheat flour reached 33.33%~79.17%. Results showed that the DNA fingerprinting technology can be used to predict the infection degree of insect pests in wheat flour, but the detection accuracy needs to be improved.