Study On The Preparation, Preliminary Purification And Antioxidant Activity Of Bee Pupa Polypeptide

Biologically active peptides,also known as functional peptides,has different biological activities,such as anti-oxidation,immune enhancement,blood pressure lowering and blood sugar lowering and so on,according to their structural properties and amino acid composition and sequence.The bee pupa is the larvae and pupae of wild bees such as wasps,wasps,black bees,and bumblebees,and contains biologically active substances such as proteins,vitamins,minerals,and flavonoids,of which proteins are the main active substances and have antioxidant and blood pressure-lowering properties.Enhance immunity and anti-tumor biological activities.However,the research and development of the bee pupa polypeptides are relatively lagging behind.There are few studies on the preparation,isolation,purification and biological activity of the bee pupa polypeptides.Based on this,systematically study the nutrition composition and active ingredients of the bee sturgeon;use the alkaline protease hydrolysis method.Polypeptides were prepared and single factor and response surface methodology were used to optimize the proteolytic conditions.The anion exchanger DEAE-52was used to separate and purify the bee pupa polypeptides.The amino acid composition and the purified components were studied,and the antioxidant activity of the purified components was discussed.It provides the basis for the research and application of functional foods of bee pupa polypeptides.The main research content is as follows:（1）Analysis of main components and nutritional components of bee pupae.The national standard method was used to study the moisture,ash,protein,fat,total sugar and flavonoid content of bee pupa powder.The study found that the bee meal was mainly composed of moisture,ash,protein,fat,total sugar and flavonoids,and its content was 15.15%,3.42%,48.13%,19.81%,10.13%,and 44.70 mg/100 g,respectively.The bee pupa powder contained 18 kinds of amino acids,and the total amount of amino acids was 42.46%.E/T%was 37.09%and E/N%was 70.65%,basically meeting the ideal protein standard.The main fats of bees were oleic acid,myristic acid,palmitic acid and stearic acid.The highest content was unsaturated fatty acid-oleic acid,accounting for 26.62%of the total fat.It can be seen that there are abundant nutrients in bee pupa powder and it is worth further research and development.（2）Enzymatic hydrolysis preparation process of bee pupa polypeptide.The bee pupa was used as the raw material,and the bee pupa protein was extracted by soxhlet extraction method.The best protease was selected and its technology was studied.Studies have found that alkaline protease is the best protease for the preparation of apical polypeptides.Single factor experiments found that the optimal extraction conditions were 0.32 g/L enzyme concentration,2 g/L substrate concentration,pH 8.5,and 40°C temperature.Response surface methodology found that the optimal extraction process was 0.31 g/L enzyme,pH 8.71 and 44°C.Under this condition,the degree of hydrolysis of the peptone peptide was 14.46%,which was close to the predicted value.（3）Preliminary purification and antioxidation of bee pupa polypeptides.The anion exchanger DEAE-52 was used to preliminarily purify the enzymatically purified bee pupa polypeptide and its antioxidative activity was studied.In vitro antioxidation experiments showed that purified fraction IV had the best ability to eliminate ABTS²⁺·;Trinece-SDS-PAGE gel electrophoresis analysis showed that purified fraction IV of the bee pupa polypeptides were mainly composed of polypeptides less than 3.3 kDa;amino acid studies found that the components IV contains 18 kinds of amino acids,and its amino acid content is significantly increased compared with the raw material;antioxidant test results show that the purified component IV of bee pupa polypeptides can effectively eliminate DPPH·、ABTS⁺·、·OH,and chelate Fe²⁺.Its EC50 is At 0.74,0.67,3.02,and 0.75 mg/mL,theβ-carotene/linoleic acid oxidation system was significantly inhibited,and the reducing ability was significantly stronger than that of the positive control carnosine.It can be seen that the peptone hydrolyzed by alkaline protease has strong antioxidant activity.