Preparation And Properties Of Rice Protein Peptide-Calcium Chelate

Calcium is an essential mineral nutrient for the body,and calcium intake or absorption in the diet can cause many diseases.Metal chelate peptides have potential applications in improving mineral element deficiency.At present,more and more metal chelate peptides that promote and enhance the bioavailability of mineral elements have been discovered and identified.Food source chelating peptides have been used for the development of functional foods and health care products related to calcium fortification because of their nutrition and safety.In this experiment,rice protein was prepared from broken rice,and rice protein peptide with rice protein chelate ability was prepared by alkaline protease.The chelate was separated and purified by size exclusion chromatography and reversed-phase high performance liquid chromatography.After sequencing,the calcium bioavailability of the chelate was verified by cell experiments.The rice protein peptide-calcium chelate obtained in this study provides a new idea for improving calcium utilization and nutrition and health food industry.The specific research results are as follows:(1)Ultrasound-assisted extraction of rice protein,preparation of rice protein peptide with optimal protease,and acquisition of peptides with higher calcium binding capacity by ultrafiltration.The results showed that the optimum conditions for ultrasonic assisted extraction of rice protein were: ratio of material to liquid 1:12,ultrasonic time 40 min,extraction temperature 60 ?,p H 9.5.Under this condition,the extraction rate of rice protein was 67.01%.Alkaline protease,flavor protease,neutral protease,papain,and rice protein were selected,and the best protease was screened by metal chelation and hydrolysis.The results showed that the alkaline chelation and hydrolysis degree of alkaline protease were the highest,67% and 16.89%,respectively.The optimal process of rice proteolysis is: temperature 63.7 ?,substrate mass fraction 4.4%,enzyme addition 6.8%,p H 8.5,under this condition,the hydrolysis degree of rice protein peptide is the highest,25.2%.The calcium chelating ability of the fractions of rice protein hydrolysate(RPH)obtained by enzymatic hydrolysis was different after ultrafiltration,and the P2(3-5 k Da)fraction had the highest calcium chelation ability.At 65.66 mg/g,the following chelation experiments were performed using P2(3-5 k Da)rice protein peptide(RPH2).(2)The rice protein peptide-calcium chelate was prepared by using the peptide P2(3-5k Da)with the highest calcium binding ability,and the structure of rice protein peptide and its chelate was analyzed by ultraviolet and infrared spectroscopy.The results showed that the optimal preparation process of rice protein peptide-calcium chelate(RPH2-Ca)was as follows:peptide to calcium mass ratio 5.76:1,p H 8.18,temperature 42.38 ?,reaction time 91.31 min,Under these conditions,the calcium chelation ability of RPH2 is 123 mg/g.Ultraviolet spectroscopy(UV-vis)and infrared spectroscopy(FTIR)spectra showed that the amide bond in Ca2+and rice protein peptides acted to form a ground state complex.The oxygen atom of the carboxyl group and the nitrogen atom of the amino group may participate in the chelation.Reacts and produces new substances.(3)Separation and purification of RPH2-Ca by size exclusion chromatography(SEC)and reversed-phase high performance liquid chromatography(RP-HPLC),and calcium was pervaporated by LC-MS/MS.The fraction with the highest peptide ratio is identified,The results showed that the fraction with the highest calcium/peptide ratio F22(RPH2-Ca22)was obtained by two-step purification,and a peptide chain with a molecular weight of 1013.429 Da was identified by mass spectrometry,Its amino acid sequence is Asn-Arg-Gly-Asp-Glu-Phe-Gly-Ala-Phe(abbreviated as NRGDEFGAF),and its calcium chelation ability is 136.2±0.75 mg/g.(4)Caco-2 cell absorption transport model was used to determine the calcium bioavailability of rice protein peptide-calcium chelate and Ca Cl2,The experimental results show that the calcium bioavailability of Ca Cl2 is 44.1%,while the calcium bioavailability of RPH2-Ca22 and NRGDEFGAF-Ca is 72% and 73.4%,respectively,Both were significantly higher than Ca Cl2(P<0.05),indicating that the rice protein peptide-calcium chelate can effectively promote calcium absorption in the intestine.Therefore,The rice protein peptide-calcium chelate obtained in this study provides a new idea for improving calcium utilization and nutrition and health food industry.