Trapping And Inhibition Mechanisms Of Catechins On N?-(carboxymethyl) Lysine

Dietary advanced glycation end products(AGEs)have been paid more and more attention since their relationship with many diseases has been proved.In order to reduce the hazard of dietary AGEs,the AGEs levels in foods should be cut down.Reduction effect of exogenous additives on AGEs formation during food processing has been extensively investigated.However,additives may also have the potential elimination effect on AGEs which is ignored by food researchers.N?-(carboxymethyl)lysine(CML),as a typical AGE,was chosen as the research target in the present thesis.The involved kinetics and mechanisms of the reaction between catechins and CML were investigated,in order to replenish the mechanism of elimination and reduction CML by catechins.The main results are shown as follow:(1)Kinetics and mechanisms of the reaction between catechin quinones and CMLCatechins exert their radical scavenging ability by donating the hydrogen atom of the phenolic hydroxyl group on their B-rings,and in effect they will be oxidized to o-benzoquinones via semiquinone radicals.In addition,these o-benzoquinones can be also formed by the autoxidation of catechins.o-Benzoquinones are highly electrophilic molecules which may react with CML.Typical catechins,such as(-)-epicatechin(EC)and(-)-epigallocatechin gallate(EGCG),were chosen in this thesis.4-Methylbenzoquinone(4MBQ),as the oxidation product of a model compound(4-methycatechol,4MC)for catechin B-ring,was applied as the model of catechin quinones for the subsequently kinetic investigation.Rate constants for reaction of 4MBQ with CML at different pHs(pH 5.0,7.0 and 8.0)were determined by stopped-flow spectrophotometry under pseudo first-order conditions,with ~0 M-1s-1 at pH 5.0,9.5 ± 1.4 M-1s-1 at pH 7.0 and 164.5 ± 5.0 M-1s-1 at pH 8.0.Rate constants for reaction of o-benzoquinones with amine-(glycine,L-lysine,N?-acetyl-L-lysine and N?-acetyl-L-lysine),thiol-(GSH,L-cysteine,N?-acetyl-L-cysteine)and guanidine-(N?-acetyl-L-arginine)containing amino acids and peptides were also measured by stopped-flow spectrophotometry.The apparent second order rate constants of 4MBQ with CML were faster than those of 4MBQ with amine-or guanidine-containing amino acids and much slower than those of 4MBQ with thiols.Thus,in thiol-containing food systems the reaction of quinones with thiols is likely to be kinetically favorable.In addition,the nature of these reaction products was identified by HPLC-ESI-MS/MS.(2)The trapping ability and mechanism of catehins and catechin oxidation products on glyoxalThe effects of catechins and catechin oxidation products on glyoxal concentrations were investigated in model systems,in order to complete the reduction mechanism of catechins on CML formation.The effects of EC and EGCG on glyoxal concentrations in aqueous solutions at pH values relevant for food(pH 5.0,7.0 and 8.0)and temperatures used during food processing(60,80 and 100oC)were investigated,and we found that EC and EGCG trapped glyoxal more efficiently at higher pH conditions and temperatures.The correlation between glyoxal trapping(%)and CML inhibition(%)by catechins was examined in a model system consisting of lysine and glyoxal.As a consequence,CML inhibition was found to be positively correlated to glyoxal decrease obtained by EC and EGCG with R2 of 0.97 for both catechins.In addition,the effect of oxidation by cupric ion on EC glyoxal trapping ability was investigated,and we found that oxidation of EC did not affect its glyoxal-trapping efficiency(p>0.05),and all of the EC oxidation products still remain the ability to trap glyoxal according to the UPLC-ESI-MS/MS analysis.(3)The trapping and inhibition effect of catechins on CML in food model systemThe trapping and inhibition effects of catechins on CML were verified in food model system.The efficiency of EC and EGCG to reduce the CML formation was investigated during heating at 80 and 100 oC in a glucose-lysine model system at pH 5.0,7.0 and 8.0 to mimic different food processing conditions.0.001-5% of EC and EGCG were found to reduce CML concentration by up to 45.5 ± 1.0% and 51.0 ± 1.7% respectively in the model system.The effect of EC and EGCG on the intermediates(glyoxal and fructosamine)of CML formation was also investigated.EC and EGCG could reduce the glyoxal levels and increase the fructosamine levels in the model systems.Adducts formed via the reaction of catechin quinones with CML in the catechins additional glucose-lysine system were measure by UPLC-ESI-MS/MS.The quinones of EC and EGCG were found to react with CML under alkaline conditions(pH 8.0),while no reaction products were found under acidic conditions(pH 5.0).In addition,effects of EC and EGCG on the CML level in the thermal treated real food systems were also studied.EC and EGCG(more effective than aminoguanidine)could slow down the increasing of CML concentrations in real food systems(soy sauce,amino acid drinking,coconut milk and corn syrup)during thermal treatment.In addition,the reaction between EC quinones and CML was found in EC additional coconut milk during thermal treatment.These achievements could be supplementary materials for the exsiting inhibition mechanism of CML,and proved that catechins could reduce the hazardous compounds levels produced during the Maillard reaction.They enriched the theories in food safety research.