Preparation Of Cyperus Serratus Polypeptide And Its Immunoregulatory Function

Cyperus esculentus is an annual herbaceous oil crop of Cyperaceae and a multipurpose economic crop with high commercial value.Now it is widely planted and promoted in China and mainly used for extracting oil.Its byproduct,the Cyperus meals,which is rich in plant protein,is a high-quality raw material for protein deep processing.Recent studies have shown that the protein absorbed by human body is mostly absorbed in the form of active peptide through the action of digestive enzymes.More and more studies have shown that bioactive peptides have a variety of physiological functions,including antioxidant,antihypertensive,anti-inflammatory,antibacterial and other properties,which have been widely concerned by the scientific community.In this experiment,Cyperus meals was used as raw material to extract the protein isolate of Cyperus esculentus through alkali precipitation and acid extraction,the alkaline protease was selected to optimize the enzymatic hydrolysis process of Cyperus esculentus,and the preparation process of Cyperus esculentus active peptide with High-DH was obtained.At the same time,in this experiment we also researched the improvement of active peptides from Cyperus esculentus on immunosuppressive model mice and the effect of active peptides on the secretion of anti-inflammatory cytokines from spleen lymphocytes of mice so as to provide theoretical and experimental basis for the industrial utilization of protein resources and the development of immune active peptide products.The main results are as follows:(1)The protein isolate of Cyperus esculentus was prepared from the oil extracted Cyperus meal(oil content 10.36%).The protein isolate(protein content 68.49%)obtained through alkali precipitation and acid extraction was used as raw material to select alkaline protease as the most suitable protease from four different proteases.The single factor experiment was carried out by selecting four factors of enzymolysis:time,temperature,p H value and enzyme dosage.After determining the enzyme hydrolysis time which had little influence on the experiment was 2 hours,the hydrolysis degree was taken as the optimization index to optimize the enzymatic hydrolysis process of Cyperus esculentus by response surface methodology.The optimum technological parameters were obtained as follows: p H 10.10,enzyme dosage 9000 U / g,enzymolysis temperature 46.2 ?.The degree of hydrolysis of the hydrolysate was 24.87%.(2)According to the optimal enzymatic hydrolysis conditions and different hydrolysis time,four kinds of Cyperus oleifera hydrolysates with different degrees of hydrolysis were prepared.The proliferation of splenic lymphocytes in mice was tested with such enzymatic hydrolysates of Cyperus oleifera at the concentration of 100 mg / L.The results showed that 18.64%,22.79%,23.45% and 24.82% of the hydrolysates could significantly enhance the proliferative activity of splenic lymphocytes,and 24.82% of the hydrolysates had the most significant proliferative activity(P < 0.05 or P <0.01)?(3)Study on immunoregulation of Cyperus oleifera polypeptide in vitro: The Cyperus polypeptides with molecular weight less than 3 k Da were obtained from the hydrolysate of Cyperus esculentus with a degree of hydrolysis of 24.82% by ultrafiltration.Using such Cyperus polypeptide as raw material,the proliferation and cytokine secretion of splenic lymphocytes in mice were determined by different concentrations of Cyperus polypeptide.The results showed that except 200 mg / L,Compared with Con A stimulation group,the proliferation of splenic lymphocytes could significantly or extremely significantly be enhanced in the other four concentrations of peptide groups(P < 0.05 or P < 0.01).(4)Compared with Con A model group,50 mg / L,100 mg / L and 150 mg / L polypeptide groups could significantly or extremely significantly enhance the secretion of IL-2 and IFN-? by spleen lymphocytes(P < 0.05 or P < 0.01).These results indicate that the < 3kda fraction of Cyperus oleifera polypeptide can regulate the cellular immune function and control the inflammatory reaction to a certain extent.(5)Study on immunoregulation of Cyperus polypeptide in vivo: Cyperus polypeptide with molecular weight less than 3 k Da was divided into three dose groups and given to the mice immunized with cyclophosphamide to prove that Cyperus polypeptide could improve the immunosuppression model induced by cyclophosphamide in mice.The results showed that Cyperus polypeptide could improve the immune organ index and body weight growth rate of mice,and increase the contents of IL-2,TNF-? and IFN-? in serum of mice.According to the results of the examination of mouse spleen sections under the electron microscope,it was found that Cyperus polypeptide alleviated the pathological changes of mouse spleen.