Preparation And In Vitro Immunological Study Of Sacha Inchi Polypeptide

Sacha inchi is a kind of oil plant with rich fat and protein which is introduced from the Guardo in recent years,and has been approved as a national new resource of food.The seed of Sacha inchi is mainly used to extract oil by low temperature pressing and has the fruit cake as the residue.This fruit cake which contains 60% of high nutritional protein is mainly used for feed processing,resulting in a waste of resources.If the fruit cake protein is use to prepare polypeptide for relative product development.On the one hand it will contribute to the development of deep processing and high value-added products,one the other hand it is helpful for acceleration the development of C hinese polypeptide market.In this paper,enzymolysis for preparation of Sacha inchi polypeptide was studied.The immunomodulatory effects of Sacha inchi polypeptides was preliminary evaluated through the induction of lymphocyte proliferation and cytokine release.Conclusions were drawn as follows:(1)Six kinds of protease,alkaline protease,papain,compound protease,pepsin,neutral protease and trypsin,were used to prepared Sacha inchi polypeptides,in which alkaline protease was the most suitable one.(2)Among three kinds of pretreatment methods,sodium sulfite adding,surfactant adding and ultrasonic treatment,ultrasonic treatment could most obviously improve the protein hydrolysis.0.15-0.45% of sodium sulfite adding could improve the protein hydrolysis,and 0.25% of sodium sulfite adding could obtain 10% improvement of protein hydrolysis.Among surfactants of CTAB,SDS,tween-80 and betaine,only CTAB could significantly improve(12%)the protein hydrolysis of Sacha inchi.The protein hydrolysis also could be significantly improve by ultrasonic treatment with 20-80 Hz for 15 min.40 Hz ultrasound assisted enzymatic treatment improved more than 50% of protein hydrolysis compared to single enzymolysis.(3)The optimum process conditions of Sacha inchi polypeptide preparation by ultrasound assisted enzymatic treatment were as follows: the protein was treated by ultrasound at 40 Hz for 15 min,then enzymolysis for 6 h at 50℃ with alkaline protease of 7500 U/g.Under these conditions,the hydrolysis degree of Sacha inchi protein was 17.92%,and the molecular weight of 94.64% peptide was below 1000 Da.(4)In vitro lymphocyte immunity test showed that polypeptide of Sacha inchi contained certain immunomodulatory effects.5-640 g/mL concentration of Sacha inchi polypeptide significantly improved the proliferation of spleen lymphocytes and obvio us dose effect was showed in medium low dose.Sacha inchi polypeptide plus lipopolysaccharide(LPS)possessed a significantly(p<0.05)synergistic effect for stimulating proliferation of lymphocyte in high dose(20-640 μg/mL).Sacha inchi polypeptide plus concanavalin A(ConA)also had a significant(p<0.05)promoting effect of lymphocyte proliferation in middle dose(20-160 μg/mL).Sacha inchi polypeptide at certain concentration also had a positive role in promoting lymphocyte secretion of interleukin IL-2 and tumor necrosis factor alpha TNF-α.