Study On The Formation Mechanism And Functional Characteristics Of Maillard Peptides

Maillard reaction markedly contributes not only to the aroma, taste and color, but also to the strong antioxidant potential. Maillard reaction is extendedly applied in the production of salty flavor and seasoning base, and in the processing of food as antioxidants, such as meat, baking food and so on. Maillard peptides, a class of Maillard reaction products (MRPs) derived from peptides and reducing sugar, produce an enhanced effect on ?avor, including umami, continuity and mouthfulness and are the flavor cores of salty seasonings and fermented sauce products. However, the traditional method of preparation for Maillard peptides is complex and costly by twice ultrafiltration technologies, which was unfavorable to their applications in production. Therefore, the researches on the formation mechanism of Maillard peptides so as to explore more suitable precursors as well as the high conversion paths of Maillard peptides, can provide theoretical sense in large-scale production of Maillard peptides for potential application in the development of flavour enhancers.Soybean protein isolate was used as a raw material for enzymatic hydrolysis in this research. The study was focused on the peptides substrate processing of flavorings from Maillard reaction, the effect of Maillard reaction on the formation of Maillard peptides as well as the formation mechanism of Maillard peptides. In addition, the flavor properties, antioxidant activities and application of these MRPs in chicken soup were investigated. The main contents are as follows:Based on the degree of hydrolysis, peptide nitrogen rate and the sensory evaluation, an appropriate endoprotease (Alcalase) was selected. The hydrolysis condition of Alcalase and Flavourzyme enzyme was obtained as follows: a substrate concentration 6%, Alcalase concentration 2000 U/g, temperature 60℃, pH 8.0, hydrolysis time 3h; Flavourzyme concentration 80 U/g, temperature 50℃, hydrolysis time 4 h. Under this condition, the obtained soybean peptides with degree of hydrolysis of 12.43% and peptide nitrogen rate of 55.58% were proper substrate for flavorings.Five soybean peptides were prepared by ultrafiltration, namely SP1, SP2, SP3, SP4 and SP5. The molecular weight (MW) distribution revealed that each peptide contained 2.67%, 9.46%, 19.57%, 50.24% and 74.51%, respectively as peptides from 1000-5000 Da. At the present of xylose (Xyl), the formation mechanism of Maillard peptides by five peptides as a function of temperature (100~140°C) and time (0~180 min) were researched in Maillard reaction. The results showed that pH value decreased and browning intensity became strong as the heating temperatures and times increased in five systems. The changes in pH and browning intensity were the highest in SP1-Xyl system, followed by SP2-Xyl system, the lowest in SP5-Xyl system. At a low temperature, the free amino acids contents declined in all systems with the rising temperature. With a further temperature elevation, an unstable trends for the changes in the free amino acids contents were observed in SP1/SP2/SP3-Xyl systems, however, a continuously increase in other two systems. Additonally, the free amino acids contents of all systems decreased and then gradually increased with the prolonged heating time. In SP1/SP2-Xyl systems, rising temperature favored the formation of Maillard peptides and the MRPs above 5000 Da, while causing the decrease of the MRPs below 1000 Da; in SP3-Xyl system, the content of Maillard peptides was not greatly affected by heating temperature; in SP4/SP5-Xyl systems, a higher temperature was unfavorable for the effective conversion of Maillard peptides as well as to the accumulation of MRPs above 5000 Da, however, the MRPs below 1000 Da only increased in the latter system. The change trends for the different MW MRPs with prolonged time were similar with those with the temperature. The results of sensory evaluation revealed that the temperatures of 120~130℃and the times of 120~150 min were proper for preparation of flavorings with enhanced-flavor properties. As for overall acceptance, the results of the sensory evaluation in a umami solution were as follows: SP5-Xyl>SP4-Xyl>SP3-Xyl>SP2-Xyl>SP1-Xyl. Though the former two systems possessed the strongest flavor-enhanced properties, SP2/SP3-Xyl systems also exhibited acceptable effects. It was a preferable industrial choice for the production of MRPs through SP2 and SP3.By investigating the proposed structures of high MW cross-links derived from in digylcine/ glutathione (GSH) / (cys-glu-lys-his-ile-met)-Xyl systems, the formation mechanisms of Maillard peptides were proposed. The results of fluorescence analysis showed that a new peak was observed in the three systems, in addition, the maximum fluorescence intensity in 420～430 nm enhanced with prolonged heating time. According to the results of Gel Permeation Chromatography, it was observed that the percentage of high MW MRPs markedly increased in the three systems, compared with the original peptides. Especially in (cys-glu-lys-his-ile-met)-Xyl model system, the percentage of Maillard peptides (1000-5000 Da) reached about 52.90%. After the separations of MRPs derived from digylcine-Xyl model system through Sephadex G-10 and twice HPLC technologies, a high m/z value of 379 was obtained by LC-MS investigations. Likewise, a high m/z value of 616 was observed in GSH-Xyl model system. Furthermore, the levels of m/z 379 and 616 increased as a function of heating time. Taken carbon atoms originated from parent sugar were indentified. In addition, it was found that there was an intensive molecular [M+H] + ions at m/z 700 to 1000, which increased with the heating time, indicating the formation of new compounds. The high MW products from small peptides may be formed by cross-linking between peptides and reducing sugar. It may suggest that Maillard peptides can be prepared by cross-linking of sugar and small peptides with a certain range of MW distribution.One hundred and thirty-one kinds of volatile compounds were separated and indentified, among which the concentrations of pyrazine, furan, pyrrole, aldehydes, ketones, alcohols, acids, esters and ethers were higher than other volatiles. The total volatiles content of MRPs derived from SP1 was higher than that of other MRPs. From the antioxidant assay results, it could be found that the antioxidant activities of soybean peptides were considerably improved by reacting with Xyl, however, there were no distinct differences among the five MRPs. Some heterocyclic compounds of MRPs significantly correlated with reducing power and Fe2+ chelating activity by principal component analysis (PCA).Results of the application indicated that the MRPs from soybean peptides significantly enhanced the flavor characteristics in chicken soup, including umami, moulthfullness and community, but no obvious improvement of aroma was observed. An increase trend was observed in flavor-enhanced properties of MRPs derived from SP1 to SP5, which was inconsitent with the results of sensory evaluation performed on a umami solution. It may suggest that SP4 and SP5 are desirable substrates for preparing the MRPs with high quality flavorings or flavor base, however, SP2 and SP3 can be more suitable precursors for the large-scale production of Maillard peptides for potential application in the development of flavor enhancers.