| ACE(Angiotensin Converting Enzyme) inhibitory peptides, which are a class of bioactive peptides that have ACE inhibitory activity and have a significant hypotensive effects, can be prduced by lactic acid bacteria. Therefore,to expand the functionality of fermentation milk and goat milk, and enhance the added-value of their products, the main subject of the research is to prepare Direct Vat Set(DVS) freeze-dried powder of Lactobacillus bulgaricus(Lactobacillus bulgaricus LB6, LB6) which was pre-screened with high yield of ACE inhibitory peptides. The main results are as follows.1. The results of Plackett-Burman and Factorial experimental design showed that lactose, trehalose and leucine influenced proliferation and antifreeze ability of LB6 signifacantly. The Box-Behnken response surface experiment indicated that, the optimal proliferation-antifreeze medium for LB6was: 2.1 % lactose, 1.0 % soy peptone, 0.9 % trehalose, 0.4 % yeast extract, and0.0012 % leucine. The validated viable counts and survival rate reached to(7.55±0.27) ×108 CFU/m L and(68.2±0.39) %, respectively. It is close to the predicted value(p<0.05). Storage stability of freeze-dried LB6,which cultured by the optimal proliferation-antifreeze medium before freeze-drying, was tested under the conditions of refrigerator freezing(-18℃).The result indicated that the prediction shelf life of the freeze-dried LB6 was about 1339 days before the viable count down to 1×106 CFU/m L.2. The results of lyoprotectants(oligosaccharides, amino acids, polymer substances, sugar alcohols, proteins, and salts, etc.) by Plackett-Burman and Factorial experiments showed that lactose, xanthan gum and polyvinylpyrrolidone(PVP) were the significant factors. Box-Behnken response surface experiments showed that the optimized lyoprotectants of LB6 cells were:16.4 % lactose, 1.1 % xanthan gum, 2.3 % PVP, 10 % oligofructose, 10 %xylooligosaccharides, 10 % galactooligosaccharides, 0.5 % Fe SO4. The verification test showed that survival rate was(81.02±1.46) %, viable counts of freeze-dried LB6 were(8.83±0.0067) ×1010 CFU/g, similar to the predicted values(p<0.05). Based on the above lyoprotectants, the optimal dose of skim milk was 24 %, and the validated survival rate and viable counts of freze-dried LB6 were(84.26±0.68) % and(1.14±0.043) ×1011 CFU/g, respectively.3. Accelerated storage testing(test at 45℃, 50℃ and 55℃) of freeze-dried LB6 powder and LB6 goat mlik powder suggested that the speculated inactivation rate constant of freeze-dried LB6 powder under 4℃ and 25℃were:k4=2.41×10-4 and k25=6.88×10-3, respectively. Thus, the predicted shelf lives of LB6 under above coditions were approximately 929 days and 32.5 days,respectively, before viable count decreasing to 1×106 CFU/m L. Moreover, the speculated inactivation rate constant of LB6 goat mlik powder at 4℃and25℃were k4 = 5.64×10-4 and k25 = 1.49×10-2, respectively. And predicted shelf life was about: 358 days and 13.5 days, respectively.4. To prepare a DVS fermenting agent, the freeze-dried LB6 powder was mixed with Streptococcus thermophilus at ratio of 1 ∶ 1. The results of fermentation experiments showed that DVS fermentation agent is better than subculture fermentation agent on proliferation and the productivily of ACE peptides. The final total viable count of LB6 was more than 1×106 CFU/m L,ACE inhibition rate dropped to 77.71 %, and p H maintained above 4.0,maximum acidity was 116.8 o T at the 28 days storage, it’s suggested that the above yogurt had good storage stability. Then, the LB6 powder was pre-mixed in goat milk powder to preparing fermentable goat milk powder and yogurt. The test of fermentation performance of fermentable goat milk powder showd that the curd time is about 5 h, the final p H, acidity, ACE inhibition rate and total viable count of LB6 were 4.34, 104 o T, 89.87 % and 2.5×109 CFU/m L,respectively, therefore, it showed a good fermentation ability and the fermentable goat milk powder is feasible. |
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