Preparation Of Casein Phosphopeptides And Studies On Its Calcium－holding Ability

Casein phosphopeptides(CPP)is using casein as the raw material,a single enzyme or enzyme hydrolysis,then purified and obtained the natural physiological activity peptide containing phosphoserine clusters.It is in the bottom of animal small intestine,which pH neutral to slightly alkaline,can combine with Ca²⁺,Fe²⁺ and Zn²⁺ and other metal ions, preventing metal ions and phosphate precipitation,so that the small intestine soluble calcium,iron and zinc concentration,and promote the absorption and utilization of these essential metal ions.This article was prepared CPP from casein,and studied the CPP's preparation process and its ability to hold calcium.Using L₉(3⁴)orthogonal experiment,to optimize the process of CPP preparation using alkaline protease 2709 to hydrolysis casein.The optimum hydrolysis conditions were:substrate concentration 5%,temperature 50℃,enzyme dosage 9000 U/g,pH 9.0,hydrolysis time 3 h.Under these conditions,the degree of hydrolysis of casein reached 22.85%.Using calcium-ethanol precipitation to isolate crude CPP from casein peptide, analyzed the factors that affect the yield and nitrogen/phosphorus molar ratio(N/P)of CPP,it was found that,with the increase of degree of hydrolysis(DH),CPP's N/P decreased.The yield of CPP increased with the increase of pH,pH at 5.5 N/P to reach the maximum.With the casein peptide solution concentration increased,CPP's N/P also will be elevated,the yield of CPP was reach the highest when casein peptide solution at a concentration of 10%.The yield and N/P of CPP were raised with the ethanol concentration and precipitate time increased.Using 001×7 cation exchange resin and 717 anion exchange resin purified crude CPP to obtain refined CPP,analyze the factors that affect nitrogen recovery and desalination rate in the process of desalination, it was found that the peptide concentration of 5%,flow rate of 6m/h,temperature 25℃as the best desalination conditions,in these conditions,the nitrogen recovery rate of 77.19%,desalination rate of 78.93%.Using high performance liquid chromatography(HPLC)to determine the molecular weight distribution and amino acid composition of refined CPP,it was found that the vast majority of CPP molecular weight distribute in the small molecular weight range,molecular weight of CPP below 3000 Da accounted for 98.22%,which,molecular weight distribute between the 1000⁵00 Da and between the 500¹80 Da each accounted for 23.29% and 60.66%.Using vitro simulation experiment to study the calcium-holding ability of CPP,analysis the factors that affect the calcium-holding capacity of CPP.The results showed that,CPP's ability to hold calcium increased with the degree of hydrolysis was enhanced;but also increased with the addition of the CPP increased and the purity of CPP improved,refined CPP can delay the formation of calcium phosphate precipitation 25³0 min.Short-term high temperatures heating in food processing will not make a major impact to CPP's ability of hold calcium;glucose can accelerate the calcium phosphate precipitation,and add the starch,sodium benzoate and citric acid have a role to postpone the formation of calcium phosphate precipitation,which citric acid is especially effective.