| Diet is an important factor in the global variation of human cancer. Heterocyclic aromatic amines (HAAs) are mutagenic/carcinogenic compounds formed at low levels via the Maillard reaction and a free radical mechanism during cooking of animal tissue at high temperature. The formation and yield of HAAs are dependent on physical parameters, such as cooking temperature and time, cooking technique and equipment, heat and mass transport, and on chemical parameters, especially the precursors to HAAs. To date, researchers all over the world are busy studying various aspects of HAAs such as their formation, occurrence in different food products, bioavailability, bio-transformation, carcinogenicity, levels of exposure to consumers, in efforts to determine their significance in human cancer development. However, the processing methods and conditions of Chinese traditional food is different from Western, and to date little is known about HAAs content in the traditional meat products of China. The objectives of the present study were to investigate the formation of HAAs in Chinese traditional food, and to evaluate their inhibition through the addition of natural antioxidants to meat before cooking. The contents and results are as follows.1. Detection of heterocyclic aromatic amines in traditional meat products by HPLCHAAs content was investigated in five traditional meat products (including roasted duck, charbroiled mutton, dried pork floss, marinated beef, roasted chicken). The clean-up procedure included tandem solid-phase extraction (SPE) using Extreltut-type columns filled with diatomaceous earth, propylsulphonic acid and chemically bounded phase-C18. Separation was achieved using TSK-gel ODS-80TM column and identification and quantitative analysis of HAAs fraction was carried out using a HPLC system with DAD and FLD. Results showed that the linear ranges were 0.01 ng·μL-1-1.0 ng·μL-1 and the limit of detection were0.02 ng·μL-1-0.3 ng·μL-1 for 15 HAAs. Recoveries were 61%-94%, the relative standard deviation were 2.4%-8.0%. Six HAAs were detected in the traditional meat products, the various and content of HAAs differentiated among the meat products. 9H-pyrido[4,3-b]indole (Norharman) and 1-methyl-9H-pyrido[4,3-b]indole (Harman) were detected in all of the meat products, and the content of it were up to 42.75 ng·g-1, 33.9 ng·g-1 respectively. 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) was only detected in roasted duck and charbroiled mutton. 2-amino-1-methyl-6-phenylimidazo [4,5-f]pyridine (PhIP) was detected in roasted duck, charbroiled mutton and dried pork floss. However, 2-amino-9H-pyrido[2,3-b]indole (AaC) and 2-amino-3-methyl-9H-pyrido[2,3-b] indole (MeAaC) were only detected in dried pork floss. The content of it were less 2 ng·g-1.2. Formation of heterocyclic aromatic amines during processing of duck meatThe effects of various processing methods, boiling, microwave cooking, charcoalgrilling, roasting, deep-frying and pan frying on the formation of heterocyclic aromatic amines (HAAs) in duck breast were studied. Results showed that both the varieties and contents of HAAs and the cooking loss of duck breast increasing along with increasing cooking temperature and time. Pan frying of duck breasts contained the highest amount of total HAAs, followed by charcoal grilling, deep-frying, roasting, microwave cooking and boiling. 9H-pyrido[4,3-b]indole (Norharman) and 1-methyl-9H-pyrido[4,3-b]indole (Harman) were detected in all of the cooked duck meat, and the content of it were from 0.1 ng·g-1 up to 33 ng·g-1. 2-amino-1-methyl-6-phenylirnidazo[4,5-f]pyridine (PhIP) was formed easily in cooked duck meat by pan frying and charcoal grilling, the content of it was from 0.9 ng·g-1 up to 17.8 ng·g-1. 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) was identified in duck meat by charcoal grilling and pan frying, the content of it was from 0.39 ng·g-1 up to 4.19 ng·g-1. 2-amino-3, 8-dimethyl-imidazo[4,5-f]quinoxaline (MeIQx) was detected in amounts below 4.49 ng·g-1 in duck meat by charcoal grilling, roasting, deep-frying and pan frying. The other HAAs were detected in amounts below 10 ng·g-1. Colour development increases with cooking temperature, but no correlation with HAAs content was observed.3. Effect of precursor composition in raw meat on the formation on heterocyclic aromatic amines in pan fried meatsThe meat pieces of following animals species, beef, pork, mutton, chicken, duck and goose were pan fried at 200℃for 10 min to study the occurrence of HAAs in fried meat of various animal species and the effect of precursor composition in raw meat on the formation of HAAs in pan fried meat. The composition of precursor (free amino acids, creatine and glucose) was examined in raw meat and the HAAs were detected in pan fried meat by HPLC with DAD and FLD detection. Results showed that great differences were found between the precursor content, both between the animal species and within any species. The molar ratio of total creatine/glucose was from 0.89 up to 9.84. Ten HAAs were detected in pan fried meat. 9H-pyrido[4,3-b]indole(Norharman) and 1-methyl-9H-pyrido [4,3-b]indole (Harman) were detected at high levels in cooked meat, the concentrations of the two compounds ranged from 10.90 ng·g-1 up to 24.16 ng·g-1 and from 4.64 ng·g-1 to 14.04 ng·g-1 respectively. 2-amino-3, 8-dimethyl-imidazo[4,5-f]quinoxaline(MeIQx) was detected in amounts up to 3.55 ng·g-1 in beef. 2-amino-1-methyl-6-phenylimidazo-[4, 5-f]pyridine (PhIP) was formed easily in poultry, the content of PhIP in chicken was up to 15.26 ng·g-1. A significant linear correlation only existed between PhIP and the molar ratio of total creatine/glucose(P<0.05), the increased glucose contents reduced the concentrations of PhIP.4. Kinetics of heterocyclic aromatic amines formation in meat model systemMutton was used as a model system to examine the kinetics of the formation ofherterocyclic aromatic amines and elucidate the effects of time, temperature, and precursors on the formation of HAAs in cooked ground mutton. Freshly ground mutton was heated at four different temperatures (160, 180, 200, 220℃) for four different time intervals (5, 10, 15, 20 min). HAAs were extracted and purified using solid-phase extraction and quantified using reverse-phase HPLC equipped with DAD and scanning fluorescence detectors. The results showed that 10 HAAs (IQ, MeIQx, 4, 8-DiMeIQx, PhIP, Norharman, Harman, Trp-P-2, Trp-P-1, AaC and MeAaC) were identified. The formation of HAAs increased with increasing time and temperature. The highest amount of HAAs formation of 4.51 ng·g-1, 10.99 ng·g-1, 5.75 ng·g-1, 7.44 ng·g-1, 25.71 ng·g-1, 22.04 ng·g-1, 2.17 ng·g-1,1.80 ng·g-1, 6.55 ng·g-1,6.32 ng·g-1, respectively were found after heating for 20 min at 220℃. Glucose and creatine decreased with increasing time and temperature, while creatinine increased with increasing time and temperature. The formation of HAAs and the degradation of precursors were inversely correlated. The formation of HAAs followed a bimolecular mechanism with one reactant concentration in large excess.5. Inhibition effect of natural extract on the formation of heterocyclic aromatic amines in cooked muttonThe effects of vitamin E, grape seed extract and quercetin on HAAs formation in fried ground mutton patties were studied. Antioxidants were added directly to the surface of the patties before frying. Patties were fried at 200℃for 10 min. HAAs were isolated by solid phase extraction and quantitated by HPLC with DAD and FLD detection. Results showed that 10 HAAs were detected in fried ground mutton patties, the total amount of HAAs was up to 51.68 ng·g-1. 9H-pyrido[4,3-b]indole (Norharman), 1-methyl-9H-pyrido [4,3-b]indole (Harman) and 2-amino-1-methyl-6- phenylimidazo-[4,5-f]pyridine (PhIP) were detected at high levels in fried ground mutton patties, the concentrations of the two compounds were 19.66 ng·g-1,12.23 ng·g-1 and 10.50 ng·g-1 respectively. Compared to the control, the HAAs were not significantly decreased by 0.05% vitamin E, 0.1% quercetin and 0.5% quercetin, but it showed the decreased trend. IQ was effectively decreased by 0.5% grape seed extract in the treatment compared to the control. IQ concentrations in cooed patties were reduced by 10.32%. Grape seed extract at 0.5% levels significantly reduced the formation of IQ, MeIQx, 4,8-DiMeIQx and PhIP by 36.65%, 67.73%, 35.63%and 30.00% respectively (p<0.05). Antioxidants were not effectively inhibited the formation of apolar HAAs in cooked mutton. |
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