Study On The Formation Pathways As Well As Reduction And Control Of Heterocyclic Amines Based On The Common Aldehyde Intermediates

Heterocyclic amines(HAs)are a large class of carcinogenic and mutagenic compounds produced during the cooking of animal-derived protein-rich food materials.Recent studies about HAs reduction are mainly based on the existing HAs formation pathways,such as scavenging phenylacetaldehyde to block 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine(Ph IP)formation and scavenging free radicals to inhibit quino(xa)line HAs.However,these are not the only pathways responsible for HAs formation,thus,considering them as targets may not achieve effective inhibition against HAs formation,resulting in a large amount of HAs remaining in the food system.Therefore,it is necessary to explore unknown HAs formation pathways to reduce them effectively.Our study has confirmed that small-molecule aldehydes such as phenylacetaldehyde,benzaldehyde,glyoxal,and methylglyoxal are important intermediates for the generation of corresponding HAs.Referring to the inhibitory strategy of adding exogenous ingredients to form adducts with phenylacetaldehyde,if these common aldehyde intermediates can be simultaneously scavenged,the inhibitory efficiency of HAs is expected to be enhanced.In this study,aldehyde compounds in chemical model system for pyridine and quinoline HAs formation were screened by gas chromatography coupled with mass spectrometry(GC-MS)to determine the aldehyde intermediates.Then,considering the quantitative relationship between aldehyde intermediates-creatinine-HAs combined with the common molecular structure characteristics of HAs,the pathways for HAs generation by condensation of corresponding aldehyde intermediates and creatinine were proposed.The combination of non-precursor amino acids was employed to scavenge these common intermediates simultaneously,and thus achieving the synergistic inhibition against HAs formation.It was found that both benzaldehyde and phenylacetaldehyde were the critical intermediates responsible for Ph IP generation,and there was a mutual conversion between the two intermediates.The pathway for the reaction of benzaldehyde and phenylacetaldehyde with creatinine to form Ph IP was also proposed.Then,the benzaldehyde/creatinine and phenylacetaldehyde/creatinine model systems were established to investigate the effects of six non-precursor amino acids on Ph IP generation,and the concentration of HAs precursor and intermediate in the system was also monitored.The results showed that histidine could decrease Ph IP formation in benzaldehyde/creatinine model system dose-dependently,and the inhibitory efficiency reached 99.51% at the 20 mg/m L addition level.Histidine also showed a good scavenging ability against benzaldehyde,with a scavenge rate of 69.13% at the highest addition level(20 mg/m L).Proline achieved a good Ph IP inhibitory efficiency of 97.03% at the highest dosage(20 mg/m L)in phenylacetaldehyde/creatinine model system and also reduced the phenylacetaldehyde content to 40.90±1.09 μg/m L compared to the control group(51.62±1.32μg/m L).Methylglyoxal and glyoxal were demonstrated to be the critical intermediates responsible for quino(xa)line HAs generation.It was found that methylglyoxal could convert to acrolein at a high temperature,and then acrolein could condense with creatinine to form 2-amino-3,7,8-trimethyl-imidazo[4,5-f]quinoxaline(7,8-Di Me IQx)and 2-amino-3,4,8-trimethyl-imidazo[4,5-f]quinoxaline(4,8-Di Me IQx).Methylglyoxal could directly react with creatinine to form 2-amino-1,5,6-trimethylimidazo[4,5-b]pyridine(1,5,6-TMIP).Glyoxal could react with creatinine to form 2-amino-1,6-dimethylimidazo[4,5-b]pyridine(DMIP),2-amino-3-methyl-3H-imidazo[4,5-f]quinoline(IQ),and 2-amino-3-methyl-imidazo[4,5-f]quinoxaline(IQx).Cysteine can inhibit the formation of 4,8-Di Me IQx and 7,8-Di Me IQx dose-dependently in methylglyoxal/creatinine system,with inhibition rates of 95.42% and 95.66% at 20 mg/m L addition level,respectively.Principal component analysis also showed that the methylglyoxal content decreased significantly after adding cysteine.The obtained results also showed that proline could efficiently scavenge glyoxal in glyoxal/creatinine system,and the scavenging rate reached 68.33% at the highest addition level.Moreover,principal component analysis showed that proline showed a good inhibitory effect on the corresponding HAs generation.The results showed that histidine and proline had a stable inhibitory effect on Ph IP generation in glucose/creatinine/phenylalanine model system,with the inhibition rates reaching100.00% and 87.37% at the highest addition level.Histidine also dose-dependently decreased the concentration of benzaldehyde in glucose/creatinine/phenylalanine system,with a maximum scavenging rate of 46.30%.Proline also had a stable phenylacetaldehyde scavenging rate,with a maximum scavenging rate of 53.05% at 20 mg/m L addition level.Principal component analysis revealed that the HAs content of cysteine and proline group significantly decreased compared with the control group in glucose/creatinine/glycine model system,and methylglyoxal and glyoxal showed the same trend.The obtained results indicated that cysteine and proline can effectively reduce the quinoline-type HAs generation by scavenging methylglyoxal and glyoxal.The inhibitory potential of histidine-proline and cysteine-proline combinations at different mass ratios was further explored in glucose/creatinine/phenylalanine and glucose/creatinine/glycine model systems.The histidine-proline combination at all mass ratios could simultaneously scavenge benzaldehyde and phenylacetaldehyde to achieve synergistic inhibition of Ph IP generation.The cysteine-proline combination could also efficiently reduce quinoline-type HAs by simultaneously scavenging the aldehyde intermediates in the system,but the inhibitory effect was greatly influenced by the mass ratio.The benzaldehyde-histidine,phenylacetaldehyde-proline,methylglyoxal-cysteine,and glyoxalproline adducts were respectively identified in the corresponding model systems.Furthermore,referring to the results calculated by Discovery Studio,the toxicity of these adducts was obviously lower than HAs.The amino acid combinations were found to have synergistic inhibitory effects on the HAs generation in roast beef patties.However,the inhibitory effect in three amino acids combination was not significantly enhanced compared to the two amino acids combinations.The contents of these HAs intermediates and precursors was positively correlated with the HAs generation,and it was verified that scavenging these aldehyde intermediates could inhibit HAs generation.Furthermore,amino acids and their combinations had no significant effect on the major volatile compounds such as pyrazines but could promote fat oxidation resulting in the content of hexanal increased.The obtained results show that the combination of amino acids has a great potential in efficiently reducing the HAs in food system,and may not significantly influence its flavor.