Compound Of Probiotic Starters And Analysis Of Its Fermented Feed Effect

Recently,China has completely banned the addition of antibiotics to feed.Therefore,finding safe,green and pollution-free alternatives to antibiotics has become a top priority for animal husbandry.According to previous studies,probiotics,a safe and reliable feed additive,is one of the most suitable substitutes for antibiotics.In this study,seven probiotics were identified,with their physiological and biochemical characteristics were also studied.Three dominant strains were screened.The fermentation mixed feed was designed by an orthogonal experiment,and its effect was evaluated and analyzed.The main results were as follows:（1）The isolation,colony morphology observation,Gram staining,16S r DNA molecular identification and sequence alignment of seven probiotic strains were carried out.The results showed that among DY,KLS,KCYB,BSQ and SCQJ with unknown genetic background were Bacillus paralicheniformis,Bacillus amyloliquefaciens,Bacillus subtilis,Pediococcus pentosaceus and Enterococcus lactis,respectively.The seven probiotics were composed of a single strain,which belonged to Yeast,Bacillus and Lactic acid bacteria.The contents of Saccharomyces cerevisiae,DY,KLS,KCYB,BSQ,SCQJ and Lactobacillus plantarum（ZWR）were 0.868×10¹⁰ cfu/g,2.272×10¹⁰cfu/g,0.277 x 10¹⁰ cfu/g,4.158 x 10¹⁰ cfu/g,2.205 x 10¹⁰ cfu/g,1.686 x 10¹⁰ cfu/g and 1.167×10¹⁰ cfu/g,respectively.Their bacterial content were higher than the national standard of 10⁶ cfu/g.（2）The growth curves of Saccharomyces cerevisiae,DY,KLS,KCYB,BSQ,SCQJ and Lactobacillus plantarum（ZWR）were measured to compare the different growth characteristics,and targeted screening experiments of Bacillus and Lactobacillus strains were carried out.The results showed that,among the three kinds of Bacillus,the ability to hydrolyze protein and starch of KLS was the strongest ability,followed by KCYB,and that of DY was the weakest.Among the three Lactic acid bacteria of BSQ,SCQJ and ZWR,the capacity of acid producing of BSQ,was the strongest,the SCQJ and ZWR strains was obviously weaker than that of BSQ.The strains of BSQ,SCQJ and ZWR had the ability of DPPH（2,2-diphenyl-1-picrylhydrazyl）free-radical scavenging.The DPPH scavenging activity of BSQ at 0 h and 15 h were 58.70%and 61.33%,respectively,which were stronger than those of SCQJ and ZWR at the same culture time.The bacteriostatic effects of DY,KLS,KCYB,BSQ,SCQJ and ZWR on Escherichia coli and Staphylococcus aureus were compared.It was found that the bacteriostatic effect of BSQ was significantly better than other strains,and it also had inhibitory effects on Staphylococcus aureus and Escherichia coli.According to the growth curve and biochemical characteristics of the seven strains,Saccharomyces cerevisiae cultured for 30 h,KLS cultured for 14 h and BSQ cultured for 15h were selected for orthogonal experiment.（3）Three factors with three levels orthogonal experiments were carried out on Saccharomyces cerevisiae,BSQ and KLS to ferment the mixed feed（soybean meal:wheat bran:corn=2:2:1）.The color,smell and texture of the fermented feed were investigated,and the p H value,total energy,water content,total acidity,crude protein,acid soluble protein and volatile base nitrogen were measured and analyzed.At the same time,the bacteriostatic effect and the ability of DPPH free-radical scavenging of feed extract were determined.The results showed that the fermented feeds in the nine experimental groups all had obvious acid or wine aroma.Compared with the control group,the p H value of the experimental group decreased significantly,with the content of total acidity,acid soluble protein and volatile base nitrogen increased significantly.Compared with the control group,the p H value of the experimental group was the lowest to 4.277.The total acidity of the control group increased from 162.55%to 298.45%.The content of acid soluble protein and volatile base nitrogen in the control group were increased from 1.88%to 2.92%and 4.93%to 38.17%,respectively.The water content of the experimental group decreased from the initial 40%to 36.70%,and the water loss was lower than the control group.There was no significant difference in total energy and crude protein content between the control group and the experimental group.The results of bacteriostatic experiment showed that the experimental groups 2,3,5 and 6 with higher concentration of lactic acid bacteria and lower concentration of S.cerevisiae had certain inhibitory effects on S.aureus.In conclusion,adding compound probiotics to the fermented feed can increase the smell of feed,and has significant effects on reducing p H,increasing the content of acid-soluble protein,volatile base nitrogen and total acidity.According to the experimental results,the combination of A₁B₂C₂,0.877×10⁷ cfu/kg of Saccharomyces cerevisiae,0.740×10¹⁰ cfu/kg of Pediococcus pentosaceus and 1.217×10⁸ cfu/kg of Bacillus amyloliquefaciens,was selected as the dominant combination.The results of this study showed that the compound probiotic fermented feed can inhibit the growth and reproduction of harmful microorganisms by reducing the p H of the fermented feed and increasing the contents of total acidity,acid soluble protein and volatile base nitrogen.At the same time,probiotics can decompose the macromolecular substances in feed materials into small molecules that are easier to digest and absorb to improve the nutritional value,digestibility and utilization of feed,thus to improve the economic benefits effectively.