| Rapeseed meal(RSM)is by-product of rapeseed after oil extraction.It contains approximately 32～40%crude protein,consists of well-balanced amino acid with higher essential amino acid.Its production was second only to that of soybean meal.Except protein,rapeseed meal composes of anti-nutritional factors,such as glucosinolates(Gls),fiber,and phytic acid.They can induce negative effects on the animal digestion and absorption of nutrients.In this study,we optimized the conditions that affecting the enzymatic hydrolysis of fiber-degrading enzymes and the fermentation of rapeseed meal by lactobacilli,the enzymatic hydrolysis and fermentation were combined to reduce the anti-nutritional factors in rapeseed meal and improve its nutritional value;the effects of enzymatic hydrolysis and lactobacilli fermentation on in vitro pig digestibility and microbial fermentation of rapeseed meal were also investigated by simulating the intestinal environment of pigs which suggested great practical significance to the development of unconventional protein-source feed,and to alleviate the shortage of protein feed resources and to reduce the cost.1.The study of optimization of lactobacilli ferment rapeseed mealIn this experiment,three strains of Lactobacillus reuteri L45,Lactobacillus plantarum L47 and Lactobacillus johnsonii L63 were used as fermentation broth to study the feasibility of lactobacilli ferment rapeseed meal.The conditions of lactobacilli fermentation of rapeseed meal were optimized by the combination of single factor experiment and orthogonal experiment.The results showed that lactobacilli could ferment rapeseed meal to produce lactic acid,reduce the pH of fermentation medium,and reach the stable phase at 24 h,indicating that lactobacilli could take rapeseed meal as the only nutrient source for growth and fermentation.By optimizing the fermentation conditions,we obtained the optimal fermentation conditions(0%of wheat bran addition,1:1.1 of feed-water ratio,7%of inoculum size,48 h of fermentation time).Compared with the unfermented group,the crude protein(CP)of rapeseed meal increased by 6.4%(39.61 vs 42.15)(P<0.05)and the trichloroacetic acid soluble protein(TCA-SP)increased by 78.5%(8.40 vs 14.99)(P<0.05).There was no significant effect on crude fiber(CP),neutral detergent fiber(NDF)and acid detergent fiber(ADF)(P>0.05).The contents of glucosinolates and phytic acid were significantly reduced(P<0.05).The results showed that lactobacilli fermentation of rapeseed meal could reduce its anti-nutritional factors and improve its nutritional value.2.The study of optimization of fiber-degrading enzymatic hydrolysis rapeseed mealBecause of the high fiber content in rapeseed meal,it is difficult to be digested and absorbed by monogastric animals.Through experiment 1,we found that lactobacilli had limited ability to degrade rapeseed meal fiber.In order to reduce the fiber of rapeseed meal and release fermentable sugar,the enzymatic hydrolysis of rapeseed meal was carried out by using cellulose and pectinase.Firstly,the optimum amount of cellulase and pectinase was determined and the compound enzyme solution was prepared.The effects of enzymatic hydrolysis pH(3.2～6.4),temperature(40～60℃),enzymatic hydrolysis time(6～30 h)and substrate concentration(50～250 g/L)on the enzymatic hydrolysis of rapeseed meal reducing sugar conversion were studied by single factor experiment and response surface experiment.The interaction among the factors is also studied.The results showed that the effect of cellulase and pectinase complex enzyme on reducing sugar conversion was greater than that of single enzyme enzymatic hydrolysis(P<0.05).The pH,temperature and enzymatic hydrolysis time are the main factors affecting the reducing sugar conversion.In the process of enzymatic hydrolysis,pH to temperature,temperature to time had interaction on reducing sugar conversion(P<0.05).Under the optimized enzymatic hydrolysis conditions(pH 4.0,temperature 50℃,enzymatic hydrolysis time 28 h,substrate concentration 200 g/L),the reducing sugar conversion rate reached 21.111%,which was close to the predicted value of 21.180%.Besides,fiber-degrading enzymes significantly decrease the crude fiber content of RSM(P<0.05).These results indicating that the established model could predict the reducing sugar conversion rate of rapeseed meal enzymatic hydrolysis by fiber-degrading enzymes.3.The effect of fiber-degrading enzymes and lactobacilli cooperate fermentation to the nutritional values of rapeseed mealEnzymatic hydrolysis or fermentation can improve the nutritional value of rapeseed meal,but there are few studies on the application of fiber-degrading enzyme cooperate with lactobacilli fermentation in rapeseed meal.Thus,the effects of two methods respectively or together on improving the nutritional value of rapeseed meal were studied.This experiment was carried out in 4 groups(control group,enzymatic hydrolysis for 28 h,fermentation for 48 h and enzymatic hydrolysis for 28 h+fermentation for 48 h)to explore the effect of enzymatic hydrolysis of rapeseed meal on the fermentation efficiency of lactobacilli,and to study the effect of enzymatic hydrolysis and fermentation on the nutritional values of rapeseed meal.The effects of enzymatic hydrolysis and fermentation on in vitro digestibility and in vitro fermentation efficiency of rapeseed meal nutrients were evaluated by simulating gastrointestinal digestion and fermentation in pigs with in vitro two-step enzymatic hydrolysis(pepsin and trypsin)and in vitro gas production method.The results showed that enzymatic hydrolyzed rapeseed meal could promote the number of viable bacteria and lactic acid content of lactobacilli after 48 h fermentation compared with not enzymatic rapeseed meal,(P<0.05).The combination of enzymatic hydrolysis and fermentation significantly increased the crude protein(CP)content of rapeseed meal compared with the unfermented group(P<0.05).Compared with not enzymatic rapeseed meal,enzymatic hydrolysis significantly reduced the contents of crude fiber(CF),neutral detergent fiber(NDF)and acid detergent fiber(ADF)(P<0.05).In addition,enzymatic hydrolysis also significantly improved the in vitro dry matter digestibility(IVDMD)and in vitro crude protein digestibility(IVCPD)of rapeseed meal(P<0.05).In vitro gas production,total short-chain fatty acids(TSCFAs),acetic acid and propionic acid contents of rapeseed meal were decreased by enzymatic hydrolysis or fermentation(P<0.05).Moreover,the degree of SCFAs decline caused by enzymatic hydrolysis was significantly higher than that caused by lactobacilli fermentation(P<0.05).There was no synergistic effect between the two processes on in vitro digestibility and in vitro fermentation efficiency(P>0.05).The results showed that the fermentable reducing sugar produced by enzymatic hydrolysis of rapeseed meal by fiber degradation could promote the further fermentation of lactobacilli,and the digestibility of nutrients in rapeseed meal could be improved in vitro. |
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