Study On The Preparation Of Protein From Rapeseed Meal And Its Character And Enzymatic Digestion

In this thesis,ultrasonic-assisted extraction of proteins from the double-1ow defatted rapeseed meal was studied.The basic extraction conditions obtained through single effect factor and the best extraction conditions were determined by Box-Benhnken center united experimental design. Combined with second stage acid-sedimentary method and ultra-filtrated method to separate the rapeseed protein , rapeseed precipitated protein RP5.8 , RP3.6 ,rapeseed ultra-filtrated protein RPs and soybean prorein SP were obtained. There were four different molecular weight fractions of RPsⅠ,RPsⅡ,RPsⅢand RPsⅣby separated and purified from the ultra-filtrated protein RPs. Physicochemical and functional properties of rapeseed protein were analysed comprehensively and preliminary discussion was carried on enzymatic hydrolysis of rapeseed protein.1,The optimum ultrasonic-assisted extraction conditions were determined by response surface methodology as follows : ultrasonic power of 450W, ultrasonic time of 80min, ratio of material VS buffer solution at 1:25(g/ml),pH=11.6,extraction temperature 35℃,extracted times of 3, under which the content of protein in the extracts and its extraction rate was improved 8.31% and 20.43%,respectively , which compared with traditional alkali extraction and acid precipitation method. The results showed that ultrasonic-assisted extraction was significantly better than traditional alkali extraction and acid precipitation method on extraction rate, protein extraction yield,crude protein content and extraction time.2,Rapeseed ultra-filtrated protein RPs was separated and purified by gel filtration chromatography and four different molecular weight fractions were obtained,which the ratio of content was 8:3:1:1. The fractions of RPsⅠand RPsⅡhad high contents , which was 61.54% and 23.08% of the total protein respectively. The physicochemical properties of rapeseed ultra-filtrated protein RPs and its fractions showed that RPs contains highly conservedβ-sheet structure. The content and length ofα-helix were different from soybean protein .The secondary structure of two fractions was similar to each other but there was great difference from RPs inβ-turns. Both thermal stability and hydrophobicity of RPs were superior to soybean protein.3,The high crude content of rapeseed ultra-filtrated protein RPs had good solubility which removed small molecule toxic substance .The functional properties of RPs was better than soybean protein. The crude content of rapeseed precipitated protein RP5.8 , RP3.6 were lower than RPs and SP, but with better oil adsorption ability ,emulsifying capacity,foaming and foaming stability than soybean protein. The amino acid analysis of three types rapeseed protein showed rational amino acid composition, the essential amino acid content, E/T,E/N in which were higher than soybean protein . From theoretical point of view rapeseed protein was high quality protein. The best choice for isoelectric point precipitation method to separate rapeseed protein should using two stage or multi stage precipitation method , favorable at the pH value around 6 and 8 from the result of 2-D electrophoresis and further around pH=5.2 and pH=10 in next precipitates. It could obtain more proteins and improve the extraction rate and crude content.4,The solubility of rapeseed protein was better at the condition ion strengthμ=0.08thanμ=0.5. It was increased along with pH increasing and it was strongest at pH=12.5. The strongest enzyme activity of alkaline protease hydrolysating rapeseed protein was 1335U/g and the degree of hydrolysis (DH)was 20.49% at condition of t=45℃,[s]= 35.41%,enzyme added0.05%,dissolved in I=0.08 ,pH=10.5 buffer. At this optimal condition, the DH of ultra-filtrated protein RPs and 2S rapeseed protein were 24.14% and 14.6%. The result of gel chromatography analysis showed the component with molecular weight≤140Da account 67.8% and the content of polypeptide account 32.2% in solutions of 2S rapeseed protein hydrolysis. It was demonstrated that alkaline protease hydrolysating rapeseed protein at this optimal condition was completely, mainly generated polypeptide, oligopeptide and amino acid, providing theoretical basis for preparing rapeseed bioactive peptide.