Background and Objective: There is concern about the safety of food, especially grilled foods in Africa. This study aimed to assess polycyclic aromatic hydrocarbons (PAH) and heavy metals content in grilled chicken meat and health risk for consumers in Benin. Materials and Methods: A total of 35 samples of grilled chicken meat were collected from Cotonou and Abomey-Calavi following various grilling processes and devices. PAHs were analyzed by gas chromatography coupled with mass spectrometry. Heavy metals were analyzed by atomic absorption spectrophotometer and mercury by spectrophotometer coupled with VP100 hydride generator. Estimated Daily Intake (EDI) and Hazard Quotient (HQ) were used to evaluate the risk of exposure to benzo(a)pyrene, lead and cadmium. Results: Whole grilled chicken meat (2.761-4.062 μg kg1) did not meet the maximum limit of 2 μg kg1 for benzo(a)pyrene set by European Union except whole pre-cooked grilled chicken meat (0.333±0.033 μg kg1) and whole grilled chicken meats with wood-device (1.493±0.055 μg kg1). Regardless of the device, grilled chicken meat skewers contained a tolerable level of benzo(a)pyrene (1.216-1.892 μg kg1). Lead level in whole (223.3±36 μg kg1) and skewers (153.3±18 μg kg1) chicken grilled on cabinet type charcoal-device exceeded the tolerated limit (100 μg kg1). However, levels of cadmium, mercury and arsenic were in accordance with the regulation in all analyzed samples. Only HQ of benzo(a)pyrene was above 1 (HQ>1) for whole grilled chicken meats consumed daily. Regardless of consumption frequency, HQ of lead and cadmium were less than 1 (HQ<1) for all types of grilled chicken meats. Conclusion: No single metal poses health risk to consumers of grilled chicken meat in Benin. However, there is a potential health risk associated with PAHs in the product, which should be managed.
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Chicken meat is not only an important source of protein but also the cheapest in the world. Its global consumption has grown rapidly from 5.95-10.87 kg per capita per year from 1995 to 2015. It is projected to increase to 12.79 kg per capita per year by 2025, more than double in 30 years1,2. It is often consumed in fried, grilled, boiled, roasted, or braised forms3,4. Grilling is a cooking method that submits the surface of food to dry heat, usually above, below or from the side5. With respect to meat, grilling increases protein digestibility, provides a distinctive aroma and flavor, while retaining the texture and shape of the meat when the temperature is between 200 and 260°C6-9. However, Grilling is among the main cooking methods responsible for the formation of Polycyclic Aromatic Hydrocarbons (PAHs) in foodstuffs. There is a multitude of PAHs but only 15-16 have been recognized as potentially harmful to animals and humans10,11. Of these, benzo(a)pyrene is the main indicator of PAH toxicity, with chrysene, benzo(a)anthracene, benzo(b)fluoranthene and benzo(a)pyrene added to form a group of four priority indicators (PAH 4)12. Due to their high toxicity, limits have been set in foods by different standard organizations in various countries and international institutions13-15. In Europe, the maximum tolerated levels for benzo(a)pyrene and the PAH4 group in food are 2 and 12 μg kg1, respectively, whereas in Benin they are 5 and 30 μg kg1, respectively16,17. In Benin, there is a lot of interest in PAH content in foods, especially in smoked and grilled ones. Kpoclou et al.18 reported that benzo(a)pyrene (91 μg kg1) and PAH4 (490 μg kg1) levels in smoked shrimp exceeded the limits mentioned above. Similar results were obtained by Iko Afé et al.19 who pointed out that smoked fish and smoked-dried fish also failed to meet the standards.
Apart from PAHs, toxic heavy metals are also contaminants that can also be found in chicken meat. Thus, limit values have been set in food products especially for lead (100 μg kg1), cadmium (50 μg kg1), mercury (500 μg kg1) and arsenic (100 μg kg1)16,20. Iwegbue et al.21 found that the levels of cadmium, nickel, chromium and lead in some chicken meat collected in northern Nigeria exceeded the tolerated limits. In Egypt, in the state of Qalyubia, the content of lead and cadmium in fried and grilled chicken meat exceeded the tolerated limits22. Indeed, in humans, chronic consumption of heavy metals can cause neurological, hepatic etc. and other health problems23-25.
Nowadays, grilled chicken meat is widely consumed mostly as street food in Benin. A recent study carried out in Abomey-Calavi and Cotonou found that grilling chicken meat may promote the appearance of PAH and heavy metals in the final product26. Thus, this study aimed to assess PAH and heavy metal contamination in grilled chicken meat and risk exposure to consumers.
MATERIALS AND METHODS
Sample collection: Samples was collected at Cotonou and Abomey-Calavi, the two biggest and crowded cities in South of Benin. The samples were collected using various process devices such as barrel device using charcoal, cabinet type device using charcoal, cabinet type device using gas and grilling device using wood26. Five samples of chicken meat skewer grilled with charcoal-barrel device (BGB), chicken meat skewer grilled with cabinet type charcoal-device (BGP), whole chicken meat grilled with charcoal-barrel device (PGB), whole chicken meat grilled with cabinet type charcoal-device (PGP), whole precooked and grilled chicken meat grilled with charcoal-barrel (PpGB), whole chicken meat grilled with cabinet type gas-device (PGG), indigenous chicken meat grilled with wood (PLG) were collected and analyzed.
Analysis of PAHs content in chicken meat: Chrysene, benzo(a)anthracene, benzo(b)fluoranthene and benzo(a)pyrène were analyzed in this study. They were analyzed using the following three steps:
Extraction: Extraction was carried out according to the 3540C method described by US-EPA27 with some modifications. Only 10 g of MgSO4 was added to 10 g of ground chicken meat and the mixture was homogenized and left to stand for 15 min at room temperature. To the ground material, 50 μL of 10 ng μL1 recovery standard solution were added along with 300 mL of dichloromethane and then extraction was carried out by Soxhlet at a rate of 2-5 cycles hr1 overnight. With the addition of 20 mL of hexane, the extract was evaporated to 1 mL.
Purification: The purification was carried out according to the 3630C method described by US-EPA28 with slight modifications. Prior to purification, the column was filled (prepared) with 3 g of silica soaked in enough dichloromethane. Then, 1 mL of Na2SO4 and 4 mL of dichloromethane were added. Finally, the column was conditioned with three portions of 6 mL of hexane. The purified extract was placed on the purification column and then eluted with 8 mL of hexane. A first fraction was collected. Then, a second elution was carried out with 14 mL of dichloromethane and a second fraction containing the PAHs was collected and filtered. The purified extract was concentrated under a nitrogen until a volume of less than 300 μL was obtained, then 50 μL of the 10 ng μL1 volumetric solution and 500 μL of isooctane were added. The purified extract was analyzed on a gas chromatograph coupled to a mass spectrophotometer (GC/MS).
Analysis: The analysis was carried out according to the 8270D method described by U.S. EPA29. About 1 μL of the aliquot was injected in high splitless mode into the GC (Thermo Scientific) equipped with a DB-EUPAH type chromatographic column (30 m×0.25 mm) with a stationary phase of 0.25 μm at a flow rate of 1.4 mL min1. The carrier gas was helium. The initial temperature was 80°C (for 1 min), then increased from 35°C min1 to 320°C and finally 3°C min1 to 335°C, maintained for 10 min. The mass spectrophotometer (Thermo Scientific) was configured for electron impact in ionization mode (70 eV), selective ions in acquisition mode and the transfer line was set at 300°C with a source of 340°C. The limits of detection and quantification were respectively of the order of 0.01 and 0.03 μg kg1 for each of the PAHs. Each extract was analyzed three times. The result was expressed in μg kg1 of meat.
Analysis of heavy metals in grilled chicken meat: The samples were previously dried at 70°C for approximately 4 days before being ground for heavy metal determination. The assay was performed following the method described by Joyce30 with some modifications. Afterward, 1 g of dried and ground sample, were previously added to 10 mL of H202 (9%) and then left for 24 hrs. Then the sample was digested with 4 mL of HNO3 and heated at 150°C until it became colorless. After evaporation and cooling, it was filtered through a wattman paper. Mercury was determined using cold mineralization of the sample. Pb, Cd, As, were determined in the filtrate using an atomic absorption spectrophotometer (brand SOLAAR S2, THERMO FISCHER). Mercury (Hg) was determined with the same spectrophotometer coupled with VP100 hydride generator. The heavy metal content was expressed in μg kg1 of meat.
Assessment of the risk of exposure to benzo(a)pyrene, lead and cadmium: The Estimated daily intake (EDI) for traces and the hazard quotient (HQ) were calculated as described by Adamou et al.31. Previous study found that skewers were preferred by children and consumed once a week by adolescents, whereas whole chickens were consumed twice a week by adults26. The amount ingested per intake for skewers was 70 g (in excess) and 300 g (in excess) for whole chickens. The maximum ingested amount of whole chicken was determined by dividing the maximum meat mass of boneless grilled whole chicken by 2, because a whole grilled chicken was often shared by at least two adult individuals. The average bone mass was subtracted due to the absence of the elements of interest.
The calculation was based on an average body weight of 28 kg for children (0-15 years) and 70 kg for an adult according to the US Environmental Protection Agency32. The EDIs and HQs were calculated for three consumption frequencies (once a week, twice a week and every day). HQ 1, indicated that the risk of toxicity is very low, while HQ >1 indicated a high probability of toxicity:
|HQ||= Hazard quotient|
|EDI||= Estimated daily intake of trace elements (μg kg1 b. wt. day1)|
|C||= Trace element concentration (μg kg1)|
|Q||= Quantity of grilled meat ingested per day (kg day1)|
|F||= Frequency of exposure (F = 1), it is without unit|
|P||= Body weight of the target (kg)|
|ADI||= Acceptable daily intake|
According to the report of the Institut National de l' Environnement Industriel et des Risques (INERIS) for chronic threshold effects, the ATSDR (Agency for Toxic Substances and Disease Registry) recommends the following maximum limits for the Acceptable Daily Intake33:
- Benzo(a)pyrene: 8.10-3 μg kg1 b. wt. day1 for a 70 kg adult
- Lead: 3.6 μg kg1 b. wt. day1
- Cadmium: 2.10-4 mg kg1 b. wt. day1
PAH levels in grilled chicken meat: PAHs content in grilled chicken meat are shown in Table 1.
Gas-grilled chicken meat had the highest values of chrysene, benzo(a)anthracene, benzo(b)fluoranthene and benzo(a)pyrene compared to other types of grilled chicken meats. It was followed by chicken meat grilled with the charcoal cabinet device which overtook chrysene and benzo(a)pyrene. There was no significant difference between these two grilled chicken meats with respect to benzo(a)anthracene and benzo(b)fluoranthene. Chrysene content of skewers grilled with the charcoal cabinet device was higher than that found in skewers and pre-cooked whole chicken meat grilled with the barrel device and indigenous chicken meats grilled with the wood. On the other hand, there was no significant difference in chrysene between the pre-cooked chicken and skewers grilled with the barrel device and the indigenous chickens. Likewise, there was no significant difference in the benzo(a)anthracene content of indigenous grilled chickens and grilled skewers. Pre-cooked grilled chickens contained the lowest value for benzo(a)anthracene. With regard to benzo(b)fluoranthene, the lowest values were found in indigenous grilled chicken meats, precooked chicken and skewers grilled with barrel device. They were followed by chicken meat skewers grilled with charcoal-cabinet device and the pre-cooked grilled chickens. There was a significant difference in benzo(a)pyrene content between all types of grilled chicken. In decreasing order, we have gas grilled chicken meat, grilled chicken meat with charcoal cabinet type kiln, grilled chicken with barrel kiln, grilled skewers with charcoal cabinet type kiln, indigenous chicken meat grilled with the wood, grilled skewers with barrel kiln and pre-cooked grilled chicken meats. In addition, all whole grilled chicken meat exceeded the maximum limit of 2 μg kg1 for the benzo(a)pyrene content set by the European Union17 with the exception of whole pre-cooked and grilled chicken meat with barrel device. However, the grilled chicken skewers met this limit. But, according to Beninese regulations16, all grilled chicken meats complied with the maximum limit of 5 μg kg1. Likewise, all the grilled chicken meats complied with the respective limits of 12 and 30 μg kg1 for the sum of the 4 PAHs set by the European Union and Beninese regulations.
Lead was the most abundant component in grilled chicken meats, regardless of the process considered. Lead levels in grilled skewers (153.3±18 μg kg1) and whole chicken meat (223.3±36 μg kg1) collected from operators using charcoal cabinet device were significantly highest and exceeded the tolerated limits. They were followed by indigenous grilled chicken meat (96.67±25.8 μg kg1) and grilled chicken meat skewers with the barrel device (73.33±36 μg kg1). Finally, pre-cooked and charcoal-grilled whole chicken meat (36.67±9 μg kg1) and gas-grilled chicken meat (36.67 μg kg1) contained the lowest levels. For cadmium, whole pre-cooked chicken meat grilled by charcoal-barrel device (PpGB) contained the highest level (36.67±6 μg kg1). In contrast, there was no significant difference in this level in the other grilled chicken meats. However, the average cadmium levels in all these grilled chicken meats were below the limit tolerated by the regulations in force in Benin. The mercury and arsenic levels of the grilled chicken meats analyzed were well below the maximum limits.
EDI and HQ of benzo(a)pyrene: Table 3 presents EDI and HQ of benzo(a)pyrene for a child (28 kg) and adult (70 kg).