Flavor Formation Mechanism,and Safety Evaluation Of Flaxseed Derived Meaty Maillard Reaction Products

In this dissertation,flaxseed meal was used as the raw material to study the effects of enzymatic hydrolysis process and Maillard reaction process on flavor formation system for Maillard reaction products(MRPs).Based on the toxicological tests,the effects of MRPs on intestinal microorganisms and their metabolites were also studied,and measures were taken to suppress the possible toxic components in MRPs,the main contents of our study are as follows:Enzymatic hydrolysis of flaxseed protein and sensory characterization of MRPs.More than 50% activity of immobilized enzymes were retained after repeated use.Subsequently,after five weeks,the activities of the immobilized enzymes were also observed to be higher after storage at 4 °C.The optimum conditions for production of flaxseed protein hydrolysates using sequential enzymatic hydrolysis were as follow: 8%(w/v)of substrate concentration,3,000 U/g of Alcalase at 60 °C and p H 8.0 for 2 h and120 U/g of Flavourzyme at 50 °C and p H 6.5 for 2 h.Peptides with the molecular weight(MW)higher than 1,000 Da could improve the mouthfulness and stability in umami soup;whereas,peptides with MW of 128?1,000 Da mainly contributed to the generation of meat-like flavor compounds with a significant effect on umami taste and bitterness.Therefore,flaxseed protein hydrolysates prepared under the best technology can be used as a high-quality raw material for extended use of flaxseed derived MRPs.Analysis of key precursor peptides based on i BAQ mass spectrometry and flavor components of flaxseed derived MRPs.The peptides and amino acids of flaxseed protein hydrolysates were separately collected by G-15 gel chromatography.Taste dilution analysis(TDA)showed that peptides-MRPs had high umami,mouthfulness,and continuity enhancement.Further,HPLC-MS/MS revealed that flaxseed protein hydrolysates consumed almost 41 peptides after Maillard reaction.Particularly,DLSFIP and ELPGSP accounted for 42.22% and 20.41% of total consumption,respectively.Aroma extract dilution analysis(AEDA)indicated that formation of sulfur-containing flavors was dependent on cysteine,while peptides were more reactive than amino acids for nitrogen-containing heterocycles.On the other hand,11 flavor compounds with flavor dilution(FD)? 64 were identified for flaxseed derived MRPs.Flaxseed derived MRPs had higher peptide reactivity than free amino acids,while the formation of meaty flavor mainly comes from the Maillard reaction involving cysteine.Color and flavor of flaxseed derived MRPs: effect of cysteine,initial p H,andthermal treatment.Higher cysteine or initial p H suppressed the cross-linking of128?1,000 Da peptides and accelerated the degradation of peptides with a higher MW(>1,000 Da).The higher temperature and longer reaction time resulted into a marked increase in flavor formation in the Maillard reaction model.Volatile compounds,MW distribution,and amino acids of MRPs had a substantial influence on MRPs sensory characteristics.The generation of umami,mouthfulness,and continuity was observed at relatively lower temperatures and longer heating time.The sulfur-and nitrogen-containing compounds from the low MW peptides(<1,000 Da)may lead to meaty aroma in MRPs.It was found through the intermediates2-threityl-thiazolidine-4-carboxylic acids(TTCA)and Cysteine-Amadori(Cys-Amadori)in the early Maillard reaction that preparation process of 1.0 g of flaxseed protein hydrolysates,0.3 g of xylose,0.15 g of cysteine(initial p H is 7.5)heat-treated at 120 °C for 120 min are conducive to the accumulation of Cys-Amadori.The flaxseed derived MRPs prepared under the optimal process can be used as meat flavorings.The process parameters are beneficial to accumulate more Cys-Amadori in the early stage reaction,thereby releasing more meat-flavored volatile components in the continuous thermal reaction.Aromatic effects of immobilized enzymatic oxidation of chicken fat on flaxseed derived MRPs.More than 50% activity of immobilized LOX was retained after repeated use for five times or five weeks.The oxidized chicken fats were prepared by thermal,free LOX,and immobilized LOX treatments.After the addition of chicken fats,Maillard reaction resulted in more aliphatic aldehydes and alcohols which manifested the enhanced reaction,but the content of sulfur compounds decreased significantly,and the structure of sulfur compounds could easily form alkyl side chains.However,there was no significant difference in sensory and flavors between oxidized chicken after treatments,which may be related to oxidized degree.The mechanism was proposed or aromatic effects of oxidized chicken fat on flaxseed derived MRPs were also evident.The characteristic flavor of chicken originates from the aliphatic aldehydes and alcohols produced by Maillard reaction after adding chicken fat.The sulfur element in the meat flavor component is derived from H2 S produced by Strecker degradation of cysteine.During the Maillard reaction,dicarbonyl compound capable of catalyzing was produced by Strecker degradation.The MRPs involved in chicken fat were endowed with the characteristic flavor of chicken.Acute,genetic,and sub-chronic toxicities of flaxseed derived MRPs.Acutetoxicity results showed that the 50% lethal dose(LD50)of MRPs in rats was > 15.0 g/kg body weight(BW);whereas,the 50% effective dose(ED50)of MRPs was 12.3 g/kg BW.Ames test demonstrated that the back-mutation colonies for MRPs were negative,which displayed no mutagenicity.There were no significant differences in micronucleus rate and sperm deformity rate among different dose groups.The sub-chronic toxicity confirmed that less than 0.75 g MRPs/kg BW intake did not affect weight,food intake,mortality,gross pathology,histology,hematology,and serum biochemistry.Flaxseed derived MRPs showed food safety at an intake of 0.75 g MRPs / kg BW.Bacteria and fermentation metabolites were altered in the intestine of SD rats fed flaxseed derived MRPs during sub-chronic toxicity.MRPs promoted the digestion of pepsin,but it was not conducive to the subsequent in vitro digestion of trypsin.MRPs ingestion could increase the propionate in intestine,but it was not enough to change the short-chain fatty acids(SCFAs)and branched-chain fatty acids(BCFAs).However,MRPs ingestion led to an increase in relative abundance of gut lactobacilli.In the high-dose groups,the abundance of key genes responsible for partial amino acid and monosaccharide metabolism increased,while it was lowered in lipid metabolism compared with the middle dose groups.Therefore,the rat body could absorb MRPs in lower amount than that of protein and carbohydrates.Daily intake of2.25 g/kg BW reduced it and weight and food intake in turn produced similar effects that of dietary fiber.Tryptophan inhibited the potential carcinogenicity of the products in the Maillard reaction.One of the compounds,3,3'-Diindolylmethane(DIM)exhibited a higher activity against cancer cells,which was further isolated,screened,and its formation process was determined by an isotope labeling test.DIM could induce cell cycle arrest in the G2/M phase and promote apoptosis in a dose-dependent manner.DIM also effectively promoted the death receptor and mitochondrial pathways,PI3K/AKT signaling pathway,and endoplasmic reticulum pathway which played an important role during this process.In summary,the thermal reaction involving tryptophan is expected to be an effective way to improve the potential in vitro mutagenicity of the product in the Maillard reaction,which can provide a future reference for wide range of thermally processed foods.