Effects Of Ultrasonic Pretreatment Combined With Maillard Reaction On Antioxidant Activity Of Casein And Its Enzymatic Hydrolysates

Casein is an important component of 80% of milk,and it is also the parent protein of many bioactive peptides,which need to be released by protease to release it for better biological activity.Studies have shown that the antioxidant activity of casein hydrolysate is more significant than that of its parent casein,but the antioxidant activity of natural casein hydrolysate is much lower than that of synthetic antioxidants.Therefore,if we want to meet the human needs for food antioxidant capacity more effectively,the core issue is how to use the modification technology to further enhance the antioxidant activity of casein and its hydrolysate.Ultrasonic technology is a highly effective modification method in physical modification.It can change the functional structure of protein and change the expression of biological activity by changing the conformation of protein molecules.Maillard reaction is also widely used to enhance protein resistance.At the same time,ultrasound can promote the Maillard reaction by changing the structure of the protein to increase its involvement in the Maillard reaction.At present,research on protein modification is concentrated,but there are few studies on the modification of proteolytics.Therefore,this s tudy combines two modification methods to study the effects of ultrasonic pretreatment combined with Maillard reaction on the antioxidant activity of casein and its hydrolysate,thus providing a theoretical basis and new methods for the development of natu ral antioxidant bases for food processing.In this study,the casein and casein hydrolysate were firstly pretreated with ultrasound,and then the two were combined with glucose for Maillard reaction.The non-maillard reaction group and the non-ultrasound treatment group were used as the control group to study the effects of ultrasonic pretreatment combined with Maillard reaction on the reaction process,antioxidant activity and structure of casein and its hydrolysate,By analyzing the structural changes of casein and its hydrolysate and exploring the relationship between it and the change of antioxidant activity,the research results are as follows:?1?Casein was hydrolyzed by papain,alkaline protease,trypsin,pepsin and neutral protease to study the effects of different proteases on the degree of hydrolysis of casein and the content of enzymatic hydrolysate and DPPH free radical scavenging ability.The results showed that the hydrolysis of casein by neutral protease had higher hydrolysis degree,the same r esults with the enzymatic hydrolysate content and DPPH free radical scavenging ability,which were?40.56±0.72?%,?22.37±0.64?mg/m L and?28.43±1.37?%,respectively.Therefore,the neutral protease is selected as the most suitable hydrolase for enzymatic hydrolysis of casein to produce antioxidant activity.?2?To study the effect of ultrasonic pretreatment combined with Maillard reaction on the reaction process of casein and its hydrolysate.The results showed that the degree of grafting,intermediate content and the degree of browning of casein hydrolysate-glucose Maillard reaction products?CH-Glc MRPs?and casein-glucose Maillard reaction products?CS-Glc MRPs?after ultrasonic pretreatment is significant higher than the untreated group?p<0.05?and the increase of CH-Glc MRPs was more significant than that of CS-Glc MRPs?p<0.05?.With the increase of ultrasonic power and time,the grafting degree of CS-Glc MRPs and CH-Glc MRPs reached the maximum at 450 W,20 min and 300 W,20 min,which were seperately26.98% and 29.57%.The A294 nm of the absorbance of the intermediate products reached the maximum at 450 W,30 min and 300 W,20 min and the maximum was separately1.26 and 0.968.The browning degree of CS-Glc MRPs reached the maximum at 450 W,20 min and the maximum was 0.481.However,the effect of ultrasonic pretreatment on the browning degree of CH-Glc MRPs was not significant?p>0.05?.?3?To study the effect of ultrasonic pretreatment combined with Maillard reaction on the antioxidant activity of casein and its hydrolysate.The results showed that ultrasonic pretreatment combined with Maillard reaction significantly increased the reducing power,DPPH free radical scavenging ability,Fe²⁺ chelation ability,Cu²⁺ chelation ability and lipid oxidation inhibition ability of casein and its enzymatic hydrolysate.?p<0.05?.With the increase of ultrasonic power and ultrasonic time,the maximum reducing power of CS-Glc MRPs and CH-Glc MRPs increased by 16.46 times and 12.26 times,respectively,compared with the origi nal casein and enzymatic hydrolysate;DPPH free radical scavenging capacity increased by 3.43 times and 2.74 times,respectively;Fe²⁺ chelation ability increased by 4.96 times and 3.70 times,respectively;Cu²⁺ chelation ability increased by 4.40 times and 3.54 times,respectively;lipid oxidation inhibition ability increased by 2.98 times and 2.97 times,respectively.At the same time,compared with the commonly used antioxidants,the reducing power and Fe²⁺ chelation ability and Cu²⁺ chelation ability of CS-Glc MRPs and CH-Glc MRPs were higher than 0.01% of butyl hydroxyanisole?BHA?and VC?p<0.05?.?4?To study the effects of ultrasonic pretreatment combined with Maillard reaction on the structure of casein and its hydrolysate.The results showed that u ltrasonic pretreatment combined with Maillard reaction enhanced the UV absorption of casein and its enzymatics,indicating that the hydrophobic chromophores inside the molecule were gradually exposed.The results of endogenous fluorescence spectroscopy showed that ?max red shift occurred in casein and its hydrolysate after ultrasonic pretreatment combined with Maillard reaction.It indicates that the two structures has become loose.The results of infrared spectroscopy showed that the absorption peaks of casein and its enzymatic hydrolysates are changed,indicating that the C=O stretching vibration and the N-H bending vibration occurred in the amide I and amide II bands,indicating that the ultrasonic pretreatment and the introduction of glucose molecules ch ange the secondary structure of casein and its hydrolysate.From the results of circular dichroism,it is known that the ordered structure of casein and its hydrolysate is reduced,the disordered structure is increased,and the secondary structure becomes disordered.The protein molecules tend to stretch.In summary,ultrasonic pretreatment combined with Maillard reaction can promote the reaction process of casein and its hydrolysate.The oxidation resistance of both is enhanced,and at the same time,the structure of the two is more loose and tends to be disordered,which is due to changes in its structure leading to changes in its antioxidant activity.