| Bacillus licheniformis D-1was separated by Institute of Feed and Science, Anhui Agricultural University. It could be used as a strain of feed microbial. In order to make the industrial production of Bacillus licheniformis come true, culture condition for alkaline protease production by Bacillus licheniformis D-1, the medium components and the fermentation conditions of fermented soybean meal were preliminarily investigated, Comparison of performance, egg quality and serum biochemical indices of laying hen for the experiment with Hens fed on fermented soybean meal Instead of soybean meal.Experiment1: Optimization of Culture Conditions of Alkaline Protease Production by Bacillus licheniformis D-1. In order to optimize culture condition for alkaline protease production by Bacillus licheniformis D-1, factors such as the culture temperature, pH, stirring speed and air flow rate during fermentation were optimized by the L49(3) orthogonal test in10L bioreactor. The optimized culture conditions were determined as a culture temperature of37℃, a pH of7.5, an air flow rate4L/min, and a stirring speed of300r·min-1. Under these optimization conditions, the confirmative experiment shows that the activity of alkaline protease produced by Bacillus licheniformis D-1reached91.26U/mL.Experiment2: Optimization of medium by Bacillus licheniformis D-1. To optimize the medium components for Bacillus licheniformis D-1, the effect of ingredient of medium on growth of strain was as the main value index. Firstly, the optimum carbon source, nitrogen source and inorganic were defined by single-factor test, and then the optimal medium of Bacillus licheniformis D-1was optimized by the L（933） orthogonal experiment. The optimal medium components was identified as follows: carbon source, nitrogen source, inorganic salts, inoculum size, culture temperature, initial pH, cultivation time were2.5%wheat bran,2%soybean power,0.06%KH2PO4,5%,37℃,7.5and24h respectively, the quantity of medium100mL in a250mL flask, The bacterium concentration reached3.06×1012cfu/mL.Experiment3: Optimization of Fermentation Conditions of Bacillus licheniformis D-1on Fermented Soybean Meal. Ordinary raw soybean was used as material. Fermentation conditions of Bacillus licheniformis D-1was optimized using L516（4） orthogonal experiment design and testing in500mL flask with aerobic solid-state fermentation. This experiment was designed to investigate the effects of five effect factors of fermentation (inoculum size, material to liquid ratio, fermentation time, poured material volume and fermentation temperature) on the contents of acid soluble protein, trypsin inhibitors (TI), alkaline protease, neutral protease and lipase activity. The results showed that the optimization fermentation conditions were as acid soluble protein: inoculum size7%, material to liquid ratio1:1.0, fermentation time58h, poured material volume60g and fermentation temperature39℃. With the fermentation, the content of acid soluble protein was23.46%the optimization fermentation conditions of trypsin inhibitors (TI): inoculum size3%, material to liquid ratio1:0.9, fermentation time48h, poured material volume55g and fermentation temperature33℃. With the fermentation, the content of was4.28mg/g the optimization fermentation conditions of alkaline protease: inoculum size7%, material to liquid ratio1:0.7, fermentation time43h, poured material volume55g and fermentation temperature33℃. With the fermentation, the activity of alkaline protease was up to46.58U/g the optimization fermentation conditions of neutral protease: inoculum size9%, material to liquid ratio1:0.7, fermentation time58h, poured material volume55g and fermentation temperature35℃. With the fermentation, the activity of neutral protease was up to31.58U/g the optimization fermentation conditions of lipase activity: inoculum size9%, material to liquid ratio1:0.7, fermentation time48h, poured material volume55g and fermentation temperature33℃. With the fermentation, lipase activity was up to51.39mg/g. Bacillus licheniformis D-1on fermented soybean meal not only improve the contents of acid soluble protein and digestive enzyme, but also reduce anti-nutrient factor clearly and lead the raw soybean meal becoming into a more nutritive protein feed.Experiment4: Effect of different Fermentation of Soybean Meal in Dietary on Performance, Egg Quality and Serum Biochemical Indices of Laying Hen. This study was designed to investigate the effects of different fermented soybean meal in dietary on performance, egg quality and serum biochemical indices of laying hen. Three hundred and twenty-four29-week-age laying hens were randomly allocated into3groups,3replicate groups of36hens each. One group received the basal diet as control group, trial one received fermented soybean meal of high acid soluble protein replaced by the proportion of 2.5%soybean meal in the basal diet, trial two received fermented soybean meal of trypsin inhibitors replaced by the proportion of2.5%soybean meal in the basal diet. The experiment lasted for42days. The results showed that, compared to the control group, the laying rates of trial one and trial two were significantly increased by3.08%and3.59%（P<0.01）, egg broken and weaken ratio was significantly decreased by0.14%and0.25%（P<0.05）, The feed/egg ratio of trial two was significantly decreased by4.00%（P<0.05）, but there were no significantly differences in egg weight and egg quality（P＞0.05） IgA, IgM, IgG, AKP and TP in each test group were all increased in different degrees, while BUN、C A and P were all decreased in different degrees compared to the control group, net benefits of trial one and trial two was increased by6.33yuan (per100grams)and10.31yuan(per100grams), respectively. |
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