Preliminary Preparation Of Branched-chain Amino Acids From Flaxseed Cake

Flaxseed cake is the by-product of flaxseed after oil extraction.The content of protein in flaxseed cake can be as high as 30% in flaxseed cake.At present,it is often used as animal fed or waste disposal,its protein resources are not only not fully utilized,but also add a certain burden to the environment.Flaxseed cake contains a complete range of amino acids,including 8 essential amino acids required by the human body,of which branched chain amino acids(BCAAs)account for about 15% of the total amino acids.As an important energy substance in the body,BCAAs has a variety of physiological functions.It is widely used in the fields of food and medicine and has great market value.At present,BCAAs is produced by microbial fermentation,and there are no reports on BCAAs extracted from plant materials.In view of the rich BCAAs in flaxseed cake,this paper selected flaxseed cake as the raw material to preliminarily explore the methods of extracting proteins from it,and further hydrolyzing proteins to prepare branched chain amino acids,so as to lay a scientific foundation for making full use of flaxseed protein resources,especially for the development of branched chain amino acid-related products.The main research results are as follows:(1)The contents of basic nutrients in flaxseed cake were determined.The results showed that the content of BCAAs in flaxseed cake accounted for 17.35% of the total amino acids,indicating that flaxseed cake was a good source of BCAAs.(2)The crude protein from flaxseed cake were extracted by alkali solution and acid precipitation method.On the basis of single factor experiment,orthogonal experiment was designed to optimize the process of flaxseed protein extraction.The best process parameter combination was: p H 9.5,and solid-liquid ratio(v/w),extracting temperature of 55 ?,extracting time 2.0 h.The results of validation test showed that the extraction rate of crude protein was 53.96% under the optimal conditions.(3)The hydrolysis of flaxseed crude protein was carried out by ultrasound-assisted enzymatic method,and the effects of ultrasonic frequency,enzyme addition,p H,hydrolysis temperature and hydrolysis time on the hydrolysis effect of flaxseed crude protein were investigated.On the basis of single factor experiment,Box-Behnken response surface analysis tests was designed to optimize flaxseed protein hydrolysis process.The optimal hydrolysis process parameters combination was: ultrasonic frequency of 6463.29 Hz,the adding quantity 3.47%,p H 9.40,hydrolysis temperature of 47.60 ?.In actual operation process parameter is adjusted to:ultrasonic frequency 6500 Hz,the adding amount of 3.5%,p H 9.4,hydrolysis temperature 45 ?,the degree of hydrolysis reached 31.72%,and the predicted value of relative error is 0.72%,shows that the response surface model is accurate and effective.(4)The method of separation and identification of branched chain amino acids in linseed crude protein hydrolysate by thin layer chromatography was preliminarily explored,and the optimal ratio of layer extender was determined to be n-butanol-butyl acetic acid-water:5:2:3,which could effectively realizes the separation and identification of branched chain amino acids.(5)Amino acids in crude protein hydrolysate were purified by using Membrane separation technology,collecting the liquid sample which can through the ultrafiltration,nanofiltration membrane filtered of different intercept molecular weight(4000 Da,2000 Da,1000 Da,500 Da,200 Da).The content of amino acids were determined by using High performance liquid chromatography(HPLC).The results showed that,the effect of 500 Da membrane on concentration of BCAAs was best:the content of total free amino acid and BCAAs were 4.644 mg/m L and was 1.349mg/m L,respectively;and the extraction rate of BCAAs was 29.85% in the permeate liquid.