| Protein was hydrolyzed to generate some peptides and a small number of amino acid. The solubility, the thermo stability and the nutrition of protein were obviously improved after hydrolysis, so that it contributed to be absorbed and used. If the condition was right, some bio-peptidase would appear. Since the hydrophobic amino acid residues which were wrapped in protein before hydrolysis were exposed, its hydrolysate generally appeared bitter and the application was limited. Aminopeptidases could be debittering exopeptidases that selectively release N-terminal hydrophobic amino acid residues from bitter polypeptides, so it had the debitterizing function. In this paper, the fermentation condition in the shake-flask and in small fementor, the preliminary extraction process abled to enlarge and the application of anti-oxidation activity of SPI hydrolysate were investigated.Optimizations of the fermentation condition in the shake-flask was fixed as follows: corn starch 30, soybean dregs 10, K2HPO4 3,MgSO4?7H2O 0.6,CoCl2 0.2mmol per liter, initiative pH 7.5,inoculum size 4%, volume 50mL/250mL,rotate speed 200r/min, temperature 38℃, fermentation time 26h.The aminopeptidase activity was about 3000U/mL. After optimization the enzyme activity in unit volume was greatly improved, meanwhile the raw materials were decreased in the unit volume and the volume in the shake-flask was increased.The fermentation conditions in the 7L fermentor were determined as follows: 4.5L in the automatic type 7L-fermentor(Bioflo 110, the United States NBS company), temperature 38℃, inoculum for 4% (V/V), stirring speed 300 ~ 800rpm, ventilation ratio of 1.5:1, dissolved oxygen and stirring speed phase coupling, the minimum dissolved oxygen is set to 60%. By controlling the DO, pretreating raw materials and applying feed batch, the activity was further improved and the greatest activity could reach 3900 U/mL.The fermentation broth after centrifugation, the recovery rate of enzyme was about 70% by volume fraction of 45% ethanol precipitation, while it was up to 90%using mass fraction of 40% (NH4)2SO4 salting-out precipitation. On the point of view that the production and extraction was enlarged in the future, saving the dosage of ethanol and (NH4)2SO4, the preliminary extraction process was established: the fermentation broth was clarified by using 15%(NH4)2SO4, followed with 30kDa-ultrafiltration membrane treatment, so that the enzyme was concentrated by 2.5 fold, with 6.1-fold purification and 87.7% recovery, respectively, 90% of the enzyme in the retention fluid was precipitated by 25%(NH4)2SO4. The aminopeptidase obtained kept 80% activity after 85 days and remained stable. The influence of some metal ions was discussed. Co2+ had obvious activation and Mg2+,K+ and Na+ had less impact on activity, otherwise, Zn2+,Fe2+,Mn2+and Ca2+ inhibited aminopeptidase activity.In the research of the soy protein hydrolyzate antioxidant activity with aminopeptidase, it is found that the size of the order of degree of hydrolysis as follows: the degree of hydrolysis dual enzyme complex hydrolysis was the largest followed by Alcalase hydrolysis,the least was aminopeptidase hydrolysis, meanwhile antioxidant capacity with two-enzyme hydrolyzate was significantly better than the other two of single-enzymatic hydrolysis.
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