Preparation And Characteristics Of Enzymatically Hydrolyzed Sesame Protein Peptide And Its Ferrous Chelate

Iron is one of the essential trace elements in the human body.Iron deficiency can cause many diseases in the human body.Inorganic iron has poor stability,low bioavailability,and has certain toxic side effects.Inorganic iron has poor stability,low bioavailability and certain toxic and side effects,while protein peptide iron chelate can be directly absorbed through the small intestine,with high absorption rate,no digestive tract stimulation and no side effects,so it is an ideal iron supplement.Sesame meal is a by-product of sesame oil extraction and is rich in various nutrients such as protein.China's sesame meal is rich in resources,but it is usually used as feed or fertilizer,and it has not been fully utilized,resulting in great waste of resources.In this paper,sesame meal was used as raw material to study the preparation and characteristics of enzymatically hydrolyzed sesame protein peptide and its ferrous chelates,which provided a theoretical basis for the development and application of new iron supplements.The sesame meal was degreased and the sesame protein was extracted by basic acid precipitation method.Alkaline protease,neutral protease and papain were used to digest the sesame protein.The alkaline protease was determined as the best enzyme source by hydrolysis degree,ferrous ion chelation rate and DPPH·clearance rate.Using alkaline protease,single factor experiments were carried out on the influencing factors of enzymatic hydrolysis(enzymatic hydrolysis time,substrate concentration,amount of enzyme added,enzymatic hydrolysis temperature and pH value).Response surface optimization for the combined rate and DPPH·clearance rate indicators,The results showed that the optimal enzymatic conditions for the preparation of chelating protein peptides with ferrous chelate activity were:enzymatic hydrolysis time 3h,substrate concentration 4.5%,enzyme loading 1500 U/g,enzymatic hydrolysis temperature 45.5?,pH 10,under these conditions,the sequestration rate of the sesame protein peptide prepared by the sesame protein peptide was 62.02%,and the DPPH·clearance rate was 53.05%;The optimal enzymatic hydrolysis conditions for preparing sesame protein peptide with antioxidant activity were:enzymatic hydrolysis time 3 h,substrate concentration 4.2%,enzyme addition amount 1400 U/g,enzymatic hydrolysis temperature 47?,pH 10,Under these conditions,the DPPH·clearance rate of the prepared sesame protein peptide was 55.42%,and the ferrous ion chelation rate was 55.09%.The optimal enzymatic conditions after optimization of the ferrous ion chelation rate are similar to those obtained by optimizing the DPPIH·clearance rate,and the DPPH·clcarance rate reached 53.05%when the chelating ability of sesame protein peptide ferrous was the highest,which was similar to the DPPH scavenging rate under the optimal enzymatic hydrolysis condition of antioxidant active peptide,therefore,the optimal enzymatic hydrolysis conditions for sesame protein were as follows:enzymatic hydrolysis time 3 h,substrate concentration 4.5%,enzyme addition amount 1500 U/g,enzymatic temperature 45.5?,pH 10.Sesame protease hydrolysate was chelated with ferrous chloride to prepare sesame protein peptide ferrous chelate.The effects of mass ratio of Vc to ferrous salt,mass ratio of peptide to ferrous salt,pH value,reaction time,reaction temperature and volume multiple of ethanol on chelation rate and product yield of sesame-protein peptide ferrous chelate were investigated by single factor experiment.According to the results of single factor experiments,the optimal process parameters for preparing sesame protein peptide ferrous chelates were as follows:mass ratio of Vc to ferrous salt 0.2:1,mass ratio of peptide to ferrous salt 3:1,The pH was 7.41,the time was 17 min,the temperature was 32?,and the amount of ethanol added was 6 times the volume of the reaction liquid,under these condition,the theoretical values of chelation rate and product yield were 72.42%and 44.60%,respectively.In the verification experiment under the same condition,the chelation rate and product yield were 72.36%and 44.68%,respectively,which were close to the theoretical values.Sesame protein peptide with high chelating activity and antioxidant activity was obtained from sesame protease hydrolysate by ultrafiltration and ion exchange chromatography.Separation and purification of sesame protein peptide by ultrafiltration were carried out,and three components of F1(>10 kDa),F2(3 kDa-10 kDa),F3(<3 kDa)were obtained.There were significant differences in the chelating rate of ferrous ions and DPPH·clearance rate among the three components.and the chelation rate of ferrous ions in component F3 was the highest at 77.64%,followed by component F2 at 68.51%,component F3 has the highest chelating rate of ferrous ions,which is 77.64%,secondly,component F2 was 68.51%,component F1 has the lowest chelating rate of ferrous ions,which is 56.60%.component F3 had the highest DPPH·clearance rate of 62.22%.The second was component F2,which was 57.02%,component F1 had the lowest DPPH·clearance rate of 48.80%,component F3 was separated by ion exchange chromatography to obtain three components:F3-1,F3-2 and F3-3.There were significant differences in the chelating rate of ferrous ions and DPPH·clearance rate among the three components,component F3-2 has the highest chelating rate of ferrous ions,which is 84.82%,secondly,component F3-3 was 76.63%,component F3-1 had the lowest chelating rate of ferrous ions of 64.64%.component F3-1 had the highest DPPH·clearance rate of 67.98%,secondly,component F3-2 was 63.22%.component F3-3 had the lowest DPPH·clearance rate of of 58.05%.The composition of sesame protein peptide ferrous chelate was qualitatively studied by sodium sulfide method.It was determined that there was no free protein peptide in the chelate,and the protein peptide was fully chelated with ferrous ion.The structure of sesame protein peptide and its ferrous chelate was identified by infrared spectroscopy.According to the different peak values and their changing characteristics,it was indicated that sesame protein peptide ferrous chelate was a new product.The results of amino acid component analysis showed that the amino acid ratio of sesame protein peptide changed before and after chelation,and the content of acidic amino acid aspartic acid,glutamic acid and cystine,histidine,alanine,methionine and leucine increased.It is indicated that the carboxyl group of the acidic amino acid and the reactive group such as a mercapto group or an imidazolyl group of the amino acid residue play an important role in the ferrous chelate ability.The bioavailability of sesame-protein peptide-ferrous chelate was studied by simulating the gastrointestinal digestive environment in vitro.By comparing the bioavailability of ferrous gluconate and inorganic iron(FeSO4),the results showed that the bioavailability of sesamin protein iron complex was higher than that of ferrous gluconate and FeSO4:After digestion for 2 h,the release rate of Fe2+ reached more than 90%.Depending on the acidification of gastric acid,the chelate of ferrous iron of sesame protein peptide can release Fe2+ slowly and its stimulation to stomach is weaker than that of ferrous gluconate and inorganic iron.Ferrous sesame protein peptide chelate was less affected by pH than ferrous gluconate and inorganic iron,and its stability was higher.After 8 hours digestion in gastrointestinal tract,the solubility and dialysis rate of ferrous sesame protein peptide chelate were above 40%,and the solubility and permeability were higher than ferrous gluconate and FeSO4.