Study On The Enrichment Of Antimicrobial Peptides And Purification Technology In Wheat Germ

In this paper, effects of culture condition and components in the culture solution on antimicrobial peptides accumulation during incubation of wheat germ were investigated. The processe parameters were optimized. The coefficient of nitrogen compounds was also discussed. The separation and purification of antimicrobial peptides and antibiotic activity of wheat germ accumulating peptides were studied, and the results were as follows:1,The free amino nitrogen and peptides content increased gradually as time extending, during wheat germ incubation. The content of soluble protein increased at begaining, and then decreased.The reducing sugar increased and the crude fat decreased gradually as time extending. Peptides were enriched because of protein degradation and increasement protease activity in the culture solution. The optimization incubation conditions, namely time, temperature, and solvent-solid ratio, were conducted using orthogonal test with three variables and one response functions. The results showed that culture conditions markedly affected peptides content and antibiotic activity. orthogonal test design showed that the optimal conditions for antibiotic activity in wheat germ were at an incubation time of 1h, a temperature of 45℃, a solvent-solid ratio of 25:1. The maximum of antibacterial circle diameter was 16.00 mm in the optimized culture conditions, which increased 1.23-fold as compared with unincubated wheat germ.2. Among nine variables investigated with Plackett-Burman design, Vc,L-cysteine and GSH were selected out based on their statistically significant (P<0.05) and positive effects on antibacterial circle diameter in wheat germ. Subsequently, steepest ascent method was used for further optimization of these selected variables to improve antibacterial activity. The most three effective components were Vc,L-cysteine and GSH and the optimum concentrations were obtained as follows:6.11 mmol/L,26.87 mmol/L and 6.78 mmol/L, respectively. An overall 1.60-fold increase in antibacterial circle diameter was achieved in the optimized culture solution (23.63 mg/g) as compared with control (14.80 mg/g).3,Wheat germ extract can remove most of the of the protein, sugar and other impurities with 80% ethano. Wheat germ extract was separated and purified by hyperfiltration, SephadexG-25, G-10 and HPLC. There are four components after isolated ultrafiltration and molecular weight less than 10K with antibacterial activity. The component G-25-2, G-10-2 and HPLC-2 had high antibiotic activity. The molecular of HPLC-2 was 456.84 Da by ESI-MS. Antibacterial experiment showed that such antibacterial peptide could effectively restrain the gram-positive bacteria (Bacillus subtilis, Micrococcus luteus and Staphylococcus aureus) and gram-negative bacteria (Escherichia coli and Pseudomonas aeruginosa). The MIC was 500μg/mL. The antibiotic activity of wheat germ extract could not be altered by high temperature (100℃) treatment with 1h, exhibiting highter thermal stability.