A Study On Digestive Enzymes Activity Of Bacillus Subtilis Fermented Feed From Cottonseed Meal

Object: The objective of this paper was to study the enzyme activity (Phytase, protease, amylase andcellulose) and nutrient content of Bacillus subtilis fermented cottonseed meal. Orthogonal design4factorsand4levels were used to optimize content of metal ions, digestive enzyme activities as indicators todetermine the amount of four metal ions (K+, Na+, Ca2+, Mg2+) in fermentation substrate. The fermentedcottonseed meals were dried by different ways and periods, which were used to explore the influence ofenzyme activity. The cotton pulp fermented product was evaluated by comprehensive evaluation ofnutrition value, which could lay the foundation for the production of high quality bio-fermented feed. Themain objective of this research was to develop B. subtilis fermented feed from cotton meal, so as to exploitthe rich cotton meal resource of Xinjiang and apply the biological characteristics of B. subtilis culture, todevelop fermented feed of B. subtilis from cotton pulp source, and it will provide a theoretical basis for theprocessing of fermented feed.Methods: The dissertation consist five experiments. Experiment1: Bacillus subtilis Z3(researchsubject), Lactobacillus acidophilus S1and Candida tropicalis ZD-3(CK) were used to ferment cottonseedmeal in the same conditions. Study the effect of different strains on enzymatic activity and nutrient contentof the fermented cottonseed meal. Experiment2: the4factors and4levels orthogonal design was used,from MgCl2(0.1%,0.2%,0.3%,0.4%),CaCl2(0.25%,0.5%,0.75%,1%), KCl(0.1%,0.2%,0.3%,0.4%), NaCl(0.2%,0.4%,0.6%,0.8%) four factors four levels, in accordance with the orthogonal designtable L44,16experiment deals in total, to design experiment and to measure enzyme activity from differenttreatments. Then, through analyzing the enzyme activity of different combinations of fermented product, byorthogonal analysis of variance and extreme difference analysis, obtaining optimal pigment addition ofMgCl2, CaCl2, KCl, NaCl. Experiment3: based on the experiment2results above, the optimal fermentationprocess parameters were composed, and then the high-quality B. subtilis culture could be produced. Afterfermentation, the fermented products were dried by different ways. one of them was dried by normal drying,the temperature was set as40℃,45℃,50℃,55℃,60℃,65℃,70℃, each temperature was continued24h,28h and72h, the second method was vacuum freeze-drying. After that, explore the influence of enzymeactivity in different temperature and time. Experiment4: the dried fermented product was packed byvacuum packaging and general packaging. After10,20,30,40,50,60days, the enzyme activity was tested.The method of experiment5was that, after fermentation, nutrients content (NDF, small peptides and phytic)was determined, so as to assess the nutritional value of fermented products.Results: The results showed that as follows, Experiment1:(1) In aspect of enzymatic activity，Z3andS1fermented cottonseed meal contained phytase and phytase activity of group Z3was higher than S1, therewas no phytase in group ZD-3. Protease and amylase were from three strains, which were the hightestprotease of group Z3and the hightest amylase of group ZD-3. Cellulase was found just in group Z3.(2) Inaspect of nutrition facts, compared with control group,the composition of CP、EE、Ca and P in cottonseedmeal fermented by those three strains had increased in some degrees of compared with control group, Ashand NDF were dicreased. The composition of CP、EE、Ca、P were the hightest and NDF was the lowest ingroup Z3. The Ash was the lowest in group ZD-3, which was decreased significantly (P <0.01). Inconclusion, cottonseed meal which was fermented by Bacillus subtilis Z3had different kinds and level ofenzymes, and it can improve the cottonseed meal quallity best of all.Experiment2: The optimized amount of inorganic salts in the fermentation substrate was MgCl2:0.2%, CaCl2:0.5%, KCl:0.1%, NaCl:0.2%, under this condition, phytase activity was73.82U/g,protease activity was57.4U/g, amylase activity was50.46U/g and cellulase activity was5.09U/g.Experiment3: Under the normal drying condition, phytase, amylase, cellulase would keep a highactivity at40℃,24h; but the optimized contidion of protease was45℃,24h. Under vacuum freeze-dryingcondition, all of the enzyme activities were better than normal drying. Experiment4: Vacuum packing has minimal impact on activity, When storage60days, phytase,protease, amylase and cellulase retained96.02%,97.02%,97.3%,98.1%.Experiment5: The study found that, the effective small peptide content in the fermented feed fromcotton meal increased from2.5%to3.84%, significant difference (P <0.01); NDF decreased by25.17%and phytic acid decreased by12.75%, significant difference (P <0.01). Indicates cottonseed meal byBacillus subtilis after fermentation can significantly increase the level of small peptides, reducing NDF,phytic acid, to improve the utilization of fermented cottonseed meal has some practical significance.