Sub-chronic exposure to PhIP induces oxidative damage and DNA damage, and disrupts the amino acid metabolism in the colons of Wistar rats.

Heterocyclic amines (HCAs) are a group of mutagenic compounds produced during thermal processing of protein-rich foods. One of the most abundant HCAs, 2-amino-1-methyl-6-phenylimidazo[4,5-b] pyridine (PhIP) has potential carcinogenic and mutagenic effects on human organs, especially the colon. This study aimed to explore the toxic effects of PhIP on amino acid metabolism in the colon of Wistar rats using RNA-seq and LC-MS/MS. Exposure to PhIP for 4 weeks induced oxidative damage and DNA damage in the colons, and disrupted the expression of related genes involved in tryptophan metabolism, beta(ß)-alanine metabolism, valine, leucine, and isoleucine degradation, and glutathione metabolic pathways. Moreover, the levels of fecal metabolites related to amino acid metabolism were affected by PhIP. Cumulatively, these results indicate that PhIP can induce colonic oxidative injury and disorders related to amino acid metabolism, thereby providing a new theoretical basis for the study of PhIP toxicity.

Formation of a creatinine thermal degradation product and its role and participation in the radical pathway of forming the pyridine ring of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP).

Creatinine, commonly found in muscle tissue, has been demonstrated as an essential precursor of 2-amino-1-methyl-6-phenylimidazo [4, 5-b] pyridine (PhIP) in thermally possessed foods. In this current study, formamide and N-methylformamide were identified as the main thermal degradation products of creatinine. The raised production of PhIP and the decreased level of aldol condensation product occurred simultaneously with the increased addition of formamide in the model system of creatinine and phenylacetaldehyde. It was demonstrated that formamide reacted with the aldol condensation product to close the pyridine ring to form PhIP. The radical pathway of PhIP formation was then proposed and verified by density functional theory (DFT) computation and an ion-trap time-of-flight mass spectrometer (IT-TOF-MS). This is the first report of the role of the creatinine thermal degradation product in the pathway of PhIP formation, which could facilitate potential strategies against the inhibition of PhIP formation in thermally processed foods.

Co-Extraction and Co-Purification Coupled with HPLC-DAD for Simultaneous Detection of Acrylamide and 5-hydroxymethyl-2-furfural in Thermally Processed Foods.

Acrylamide and 5-hydroxymethyl-2-furfural (5-HMF) are two of the most abundant compounds generated during thermal processing. A simple method for the simultaneous quantitation of acrylamide and 5-HMF was developed and successfully applied in thermally processed foods. Acrylamide and 5-HMF were co-extracted with methanol and then purified and enriched by an Oasis HLB solid-phase extraction cartridge, simultaneously analyzed by high-performance liquid chromatography and detected with a diode array detector, respectively, at their optimal wavelength. The linear concentration range was found to be 25-5000 µg/L with high linear correlation coefficients (R > 0.999). The limit of detection and the limit of quantitation for acrylamide and 5-HMF were 6.90 µg/L and 4.66 µg/L, and 20.90 µg/L and 14.12 µg/L, respectively. The recovery of acrylamide and 5-HMF in biscuits, bread, Chinese doughnuts, breakfast cereals, and milk-based baby foods was achieved at 87.72-96.70% and 85.68-96.17% with RSD at 0.78-3.35% and 0.55-2.81%, respectively. The established method presents simplicity, accuracy and good repeatability, and can be used for the rapid simultaneous quantitation of acrylamide and 5-HMF in thermally processed foods.

Dual Effects of Creatinine on the Formation of 2-Amino-1-Methyl-6-Phenylimidazo [4,5-b]pyridine (PhIP).

Creatinine was found to not only act as a precursor of 2-amino-1-methyl-6-phenylimidazo [4,5-b]pyridine (PhIP) formation but also inhibit PhIP formation in a creatinine/phenylalanine model system. The dual mechanistic effects of creatinine on PhIP formation were then investigated in a model system. Adducts of creatinine-PhIP were detected by quadrupole-time-of-flight mass spectrometry and were found to be a likely explanation for the substantial decrease in the yield of PhIP when excess creatinine was supplied. Structures of probable adducts were predicted in molecular docking studies, which showed that hydrogen bonds were formed between creatinine and PhIP in 1:1 and 2:1 ratios. Furthermore, the active sites during creatinine-PhIP adduct formation (the primary amino groups [N2 -] and sp2 nitrogen atoms [N3 ] of creatinine and PhIP) match the active sites of PhIP metabolism and adducts of PhIP/lipid-derived reactive carbonyls. This verifies that creatinine inhibits PhIP production via the formation of adducts with hydrogen bonds at the N2 and N3 sites. PRACTICAL APPLICATION: This study enhances the understanding of how creatinine affects PhIP formation, reveals a new PhIP inhibition mechanism, and will be useful for developing technology to control PhIP formation during food processing.

Inhibition effects of flavonoids on 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline and 2-amino-3,7,8-trimethylimidazo[4,5-f]quinoxaline formation and alkoxy radical scavenging capabilities of flavonoids in a model system.

BACKGROUND: Heterocyclic aromatic amines (HAAs) have been considered as carcinogenic and mutagenic chemicals generated during thermal processing of protein-rich foods that can be inhibited by some flavonoids. Free radical scavenging is a major characteristic of flavonoids. RESULTS: The half-maximal inhibitory concentration (IC50 ) values of nine flavonoids were determined by evaluating their capacity to inhibit 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and 2-amino-3,7,8-trimethylimidazo[4,5-f]quinoxaline (7,8-DiMeIQx) formation in a model system. The results of the 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) test validated that MeIQx and 7,8-DiMeIQx formed via a free radical pathway. Electron spin resonance (ESR) spectroscopic analysis with spin trapping (α-(4-pyridyl N-oxide)-N-tert-butylnitrone (POBN) spin adduct, aN = 15.2 G and aH = 2.7 G) revealed that an alkoxy radical was the generated intermediate. The scavenging capacities of the nine flavonoids on alkoxy radicals were then evaluated based on the ESR spectra of the POBN spin adducts. CONCLUSION: The weak correlation between the alkoxy radical scavenging capacities and IC50 of the flavonoids suggested that their inhibitory activity against MeIQx and 7,8-DiMeIQx formation operates by a more complex mechanism than simply scavenging alkoxy radicals. © 2017 Society of Chemical Industry.

Inhibition of 2-Amino-1-methyl-6-phenylimidazo [4,5-b]pyridine (PhIP) Formation by Alkoxy Radical Scavenging of Flavonoids and Their Quantitative Structure-Activity Relationship in a Model System.

The inhibitory effect of 10 flavonoids on the formation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in a creatinine-phenylalanine model system was investigated through electronic spin resonance and a quantitative structure-activity relationship. Alkoxy radicals were observed during the heating process, providing evidence for a radical pathway in the formation of PhIP. The alkoxy radical scavenging capability of the flavonoids was proportional to their inhibition of PhIP formation (IC50 ). We deduced that flavonoid inhibition of PhIP generation occurs via scavenging of alkoxy radicals during the heating process. Multiple linear regression and partial least squares models were used to elucidate the relationship between PhIP inhibition activity and structure characteristics of the flavonoids. The lipo-hydro partition coefficient and molecular fractional polar surface area of the flavonoids were found to be predictive of the inhibition effect.

Effects of antioxidants of bamboo leaves and flavonoids on 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) formation in chemical model systems.

The inhibitory effects of antioxidants of bamboo leaves (AOB) and flavonoids against 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) formation were investigated in creatinine and phenylalanine model systems. AOB and the tested flavonoids (orientin, homoorientin, vitexin, isovitex, apigenin, luteolin, isorhamnetin, fisetin, and hesperetin) had significant dose-dependent inhibition effects on PhIP formation with different IC50 values. The superoxide anion (O2(•-)) scavenging activities of these nine flavonoids were evaluated using the pyrogallol autoxidation system. The EC50 values of compounds that showed antioxidant activity were found to correlate well (R(2) = 0.8003) with the corresponding IC50 values representing their inhibition of PhIP formation. It was assumed that the inhibitory effects of flavonoids on PhIP formation were probably achieved by scavenging free radicals generated in the reaction system. These findings provide valuable information for the development of effective strategies to minimize heterocyclic amine content in thermally processed food.