Studies On Reduction Mechanism And Structure-activity Relationship Of Acrylamide In Foods By Bio-flavonoids

Recently, the findings, analytical methods, formation mechanism and reduction pathways of acrylamide in foods have attracted wide attention all over the world. The formation mechanism, impact factors and effective reduction methods of acrylamide during heat processing play important roles in the defense of acrylamide hazard and food safety. Studies on the reduction effect of plant-derived components on the formation of acrylamide and its precursors and intermediates through Maillard reaction in foods become a research hotspot. This dissertation systematically studied the mechanism and structure-activity relationship of bio-flavonoids in reducing acrylamide formation in foods.Two analytical methods of gas chromatography (GC) and liquid chromatography tandem mass spectrometry (LC-MS/MS) for the quantification of acrylamide were investigated. The LC-MS/MS method was further validated via the analysis in routine heat-treated foods, baby foods and complex food matrixes, trace analysis, and food analysis performance assessment scheme (FAPAS) proficiency test. The above two methods were compared with each other and applied for the analysis of acrylamide in Chinese traditional foods. Finally, the LC-MS/MS method was selected as the standard method for quantitative analysis of acrylamide.Two bio-flavonoid rich extracts of antioxidant of bamboo leaves (AOB) and extract of green tea (EGT) were first used to examine the effect on acrylamide reduction. The result showed that 74.1% and 76.1% of acrylamide were reduced in potato crisps and French fries when they were treated by 0.1% and 0.01%(w/w) of AOB solutions before frying, respectively. Similarly,59.0% of acrylamide was reduced in fried chicken wings when 0.5%(w/w) of AOB was used. Furthermore,82.9% and 72.5% of acrylamide were reduced in Chinese fried bread sticks when 0.1%(w/w) of AOB and 0.01%(w/w) of EGT were added in the flour, respectively. The optimal addition levels of AOB and EGT were finally obtained after taking the maximum permissible addition levels and sensory quality into consideration. Based on the confirmation of the reduction effect of bio-flavonoids on acrylamide in foods, the formation of acrylamide was studied in a model system of asparagine-glucose/fructose/ sucrose. The experimental design was performed by the response surface methodology (RSM) and orthogonal array methodology (OAM) while the acrylamide content was analyzed by ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). Results showed that acrylamide was optimally generated in model systems when the equimolar asparagine and glucose were heated 15 min in the low-moisture system at 180℃or 5 min in the microwave system at 180℃.The acrylamide content could be decreased either by inhibiting its formation or promoting its elimination. The effect of AOB and EGT on the formation and elimination kinetics of acrylamide was then investigated in the asparagine-glucose model system. The maximum reduction effect of acrylamide was observed in low-moisture and microwave systems when the addition levels of both extracts were 2×10-4 mg/g and 10-6 mg/mL, respectively. Meanwhile, the effect of both extracts on the kinetics of acrylamide in low-moisture and microwave systems was described via the Logistic-exponential and first-order formation/elimination kinetic models, respectively. Results indicated that addition of AOB and EGT could significantly reduce the formation kinetics of acrylamide while no effect on its elimination kinetics.The reduction effect and structure-activity relationship of totally 24 bio-flavonoid samples on the formation of acrylamide were then studied in a potato model system using the microwave heating method. Results of dose-response study showed a maximum reduction effect when the addition levels of bio-flavonoids were 10-9 mol/L. Results of antioxidant evaluation systems using DPPH, ABTS and FRAP methods revealed a significant positive correlation between the acrylamide reduction effect of bio-flavonoids and the antioxidant levels (ATEAC) in reaction systems. Meanwhile, the structure-activity relationship study on the reduction effect of bio-flavonoids was performed via the quantitative structure-activity relationship (QSAR) method, and it was demonstrated that the number and position of aromatic hydroxyl functional groups play an important role in the reduction effect of flavonoids. Furthermore, the reduction effect of flavones is more effective than isoflavones with the same number of aromatic hydroxyls. The reduction effect of flavone C-glycosides is better than flavone O-glycosides for the flavone glycosides with the same structure of aglycone. The topological structures of bio-flavonoids are also related to their reduction effects.To simultaneously quantify the contents of asparagine, glucose, fructose and acrylamide in final reaction products, an improved UPLC-MS/MS method was evaluated. Six bio-flavonoids including homoorientin (the characteristic flavone C-glycoside of AOB), (-)-epigallocatechin gallate (the characteristic flavanol ester of EGT), luteolin-7-O-glucoside, luteolin, genistein and quercetin were selected for the reduction mechanistic study according to their reduction abilities and universality. The mechanistic kinetic models were established in the potato microwave model system based on the optimal addition levels (10-9 mol/L) of 6 selected bio-flavonoids. Then, the kinetic profiles of asparagine, glucose, fructose, acrylamide and melanoidins were obtained. The reduction positions of bio-flavonoids were finally analyzed after estimating the change of kinetic parameters. Results indicated that bio-flavonoids can significantly reduce the formation of intermediates such as Schiff base in the reaction between asparagine and fructose, the transform action from glucose to fructose via isomerization, and the formation of acrylamide from intermediates. However, they cannot reduce the reaction between asparagine and glucose and no effect on the elimination of acrylamide. Furthermore, bio-flavonoids have reduction effect on the formation of melanoidins, which is one of the final products of the Maillard reaction. However, such effect seems not significant.