Study On The Extraction Of Silkworm Pupa Protein With Ionic Liquid And Its Phosphorylation Process

China is the world’s largest silk producer.The output of silkworm cocoons and raw silk accounts for about 80% of the world’s total.The annual silk trade volume exceeds 42 billion.dollars.Silkworm pupae are one of the main by-products of the sericulture industry.The annual output of silkworm pupae in my country exceeds 500,000 tons.It is reported that the protein content of silkworm pupa is about 60%,composed of 18 kinds of amino acids,of which 8 essential amino acids account for 42% of the total amino acid content.It is a high-quality edible protein with high nutritional value.In order to establish a green and efficient method for extracting protein from silkworm pupa,this thesis uses reeled silkworm pupa as raw material to study the ionic liquid method to extract silkworm pupa protein(SPP).The orthogonal experiment optimizes the silkworm pupa protein extraction process and compares the protein molecules.Changes in structure and physical and chemical properties;at the same time,in order to improve the processing characteristics of silkworm pupa protein,sodium tripolyphosphate(STP)was used to phosphorylate the silkworm pupa protein,and the response surface methodology was used to optimize the modification process to study the phosphorylation of silkworm pupa protein.Molecular structure and physical and chemical properties.The specific research results are as follows:Extraction of protein from silkworm pupa by ionic liquid methodTaking silkworm pupa as raw material,comparing different ionic liquids,such as choline acetate,choline dihydrogen phosphate and choline hydroxide,to extract silkworm pupa protein,and orthogonal experiment to optimize the extraction process.The results show that choline hydroxide is a suitable ionic liquid for extracting silkworm pupa protein.The factors affecting the extraction effect are: choline hydroxide content,solid-liquid ratio,temperature,reaction time,and the optimal extraction conditions are: choline hydroxide concentration 3 %,solid-liquid ratio(g/ml)1:30,dissolution temperature 40℃,dissolution time 1h,under the above conditions,the protein recovery rate of silkworm pupa is 64.52%,and the protein content is 90.45%.2.The molecular structure and physical and chemical properties of silkworm pupa proteinThe molecular structure of silkworm pupa protein was analyzed by scanning electron microscopy,infrared spectroscopy,circular dichroism and other methods,and the cytotoxicity of silkworm pupa protein was verified by MTT method.The results show that the microstructure of silkworm pupa protein is loose and porous,with four protein infrared characteristic peaks of amide Ⅰ,Ⅱ,Ⅲ and A.In the secondary structure,α-helix accounts for 7%,β-sheet accounts for 72.3%,and β-turn accounts for 1%.%,random curling accounted for 19.7%;the solubility of silkworm pupa protein extracted by ionic liquid method was increased by 82.87% compared with alkali-soluble acid precipitation method,emulsification increased by 15.46%,emulsification stability increased by 30.15%,foamability increased by 15.8%,Bubble stability increased by 31.5%,disulfide bond content decreased by 16.4%,and sulfhydryl group content increased by 4.09%;the silkworm pupa protein extracted by ionic liquid method had no effect on the growth of human umbilical vein endothelial cells(HUVEC).3.Preparation of phosphorylated silkworm pupa proteinTaking silkworm pupa protein as the raw material,using sodium tripolyphosphate to phosphorylate the silkworm pupa protein,taking the solubility of silkworm pupa protein as the inspection index,and optimizing the modification process by response surface methodology.The results showed that the solubility of silkworm pupa protein was17.42g/L under the conditions of STP/SPP 44.04%,temperature 50.01℃,p H 8.89,and time2.10 h.The factors affecting silkworm pupa protein are: STP/SPP > p H > reaction temperature > reaction time.4.The molecular structure and physical and chemical properties of phosphorylated silkworm pupa proteinPhosphorylation of silkworm pupa protein with STP increased its solubility by 204%,and its foamability,foaming stability,emulsification and emulsification stability were increased by 108.24%,17.63%,48.99% and 9.94%,respectively.The secondary structure of phosphorylated silkworm pupa protein is mainly composed of β-sheets and random curls.Compared with the unmodified silkworm pupa protein,they are increased by 26.9% and12.2%,respectively;while the unmodified silkworm pupa protein is composed of α-helix and β-turn.Mainly,accounting for 35.7% and 25.3% respectively.The MTT method detected the cytotoxicity of phosphorylated silkworm pupa protein,which showed that phosphorylated silkworm pupa protein had no inhibitory effect on the growth of human umbilical vein endothelial cells(HUVEC).