1. Introduction
According to the Food and Agriculture Organization (FAO) of the United Nations, the primary sources of meat are domesticated animal species, such as cattle, pigs, birds, sheep, and goats, respectively [
1]. The content of major elements (Ca, Fe, Mg, K, Na, and P) and trace elements (Zn, Mn, Cu, As, Cd, Hg, Pb, and Se) in meats [
2], lipids, carbohydrates, proteins, color, and texture depend on several aspects, such as the region in which the animal is raised, genetics, age, food, type of cut, and tissue [
3,
4].
The Commission of the European Communities stipulated the maximum allowed limit of As, Cd, and Pb in poultry, cattle, pork, and sheep muscle and offal [
5]. Different methods are used to determine the ingested amount of food contaminants, water, and other liquids. International organizations such as the Joint FAO/WHO Expert Committee on Food Additives (JECFA) and the European Food Safety Authority (EFSA) recommends the use of health-based guidance values (HBGVs) as provisional tolerable weekly intake (PTWI) and benchmark dose lower confidence limit (BMDL) for metals and metalloids considered contaminants that may accumulate in the body [
6,
7,
8]. Other approaches to estimate the possible exposure to food additives from a diet comprise the estimated daily intake (
EDI) [
9] and the hazard quotient (HQ) method. The
EDI is the amount of an additive ingested by the average consumer of the food [
10,
11], and the
HQ is the ratio of the potential exposure to a substance at a level at which no adverse effects are expected, respectively [
12]. Moreover, the minimal risk level (MRL) is an estimate of the daily human exposure to a hazardous substance that contains elements such as Al, Cd, Co, Cr, Cu, Mo, V, and Zn that is likely to be without appreciable risk of adverse [
13]. On the other hand, for some elements, such as Cu, Fe, Mg, Mn, Mo, Ni, V, and Zn, there is a maximum level of daily nutrient intake (UL) that aims to avoid the risks of toxicity [
14].
The way meats of different types are prepared [
15], and especially storage conditions and handling, are potential sources of contamination and can significantly alter the levels of metals present in food [
16], by placing food in contact with the contaminants. During the thermal processing of food, reactions that change color and taste occur, including the ones that result in toxic and mutagenic substances, such as furan, acrylamide, and acrolein [
17]. Besides, fuels such as coal, which is used in barbecue grills, release heavy metals [
18], becoming a possible food pollutant.
According to studies, metals are an expressive route of exposure to toxic compounds [
19,
20,
21]. Total and bioaccessible concentrations of trace elements (Al, As, Cd, Cu, Fe, Hg, Mn, Ni, Pb, and Zn) were measured in charcoals from 15 barbecue products available from UK retailers [
19]. According to an estimate of daily intake for the bioaccessible trace elements in the coal samples available in the United Kingdom, As and Al are the elements that caused the most significant concern of impact on human health [
19]. The Canadian government considers charcoal as a source of hazardous emissions for health; thus, its sale and advertising are under regulation [
20].
In many countries, like Brazil, Argentina, Paraguay, and Bolivia, the use of wood on a barbecue grill for roasting meat is a cultural issue. In Brazil, along with the various types of wood sold as barbecue fuels, some people use the remains of construction wood, without knowing that a number of them have undergone chemical treatment, such as
Eucalyptus and other wood that is treated with Chromed Copper Arsenate (CCA) [
22]. Consequently, wood smoke contains several toxic, harmful air pollutants, including As and Cr [
23,
24,
25]. In view of the above, studies considering the
EDI and comparisons with reference values for each element (PTWI, UL, BMDL, and MRL) can provide an overview of human-health-risk assessment [
6,
7,
8,
9,
10,
11,
12,
13,
14].
Some studies reported the concentration of toxic and essential elements in several commercial types of meats, using different thermal preparation methods (roasting, boiling, and microwave cooking) [
26], determination of metal in raw meats by inductively coupled plasma optical spectrometry (ICP-OES) [
27] and inductively coupled plasma mass spectrometry (ICP-MS) [
28], the role of detergent washing on levels of metals in meat [
29], and validation of mineralization procedures for the determination of contents in raw meats samples by ICP-MS [
30]. However, there is limited data on the major and trace elements composition of roast meats consumed by the public in Brazil and other countries, especially regarding the metal and metalloids content after barbecuing-smoke exposure. To date, there are no studies that have quantified the accumulation of macro- and microelements in different types of meat when roasted on barbecue grills, using different kinds of wood and charcoal as fuels. The aim of this study was (i) to determine the levels of Cd, Co, Cr, Mn, Mg, Cu, Al, Mo, Ni, Pb, V, Fe, Zn, and As in beef topside, pork loin, chicken breast, and lamb shank roasted on barbecues, using different types of fuels, such as woods and charcoal; (ii) obtain the
EDI and
HQ due to exposure to metals consumed from roasted meats, using wood and coals; and (iii) compare the
EDI values in roasted meats with the values established by the PTWI, UL, BMDL, and MRL.
2. Materials and Methods
2.1. Sample Collection
Three samples of each type of raw meat were purchased from six different butcheries of Campo Grande, MS, in the Midwest region of Brazil, while considering the different origins in terms of the production system (meat labels) and anatomical location: beef (topside), pork (loin), lamb (shank), and chicken (breast without skin).
The following woods were acquired through direct purchase in commercial establishments in Campo Grande, Brazil: Eucalyptus citriodora wood, Guazuma ulmifolia wood, Anadenanthera falcata wood, and treated Eucalyptus (CCA-treated Eucalyptus wood). Two charcoals were purchased: Eucalyptus citriodora coal and Guazuma ulmifolia coal. The purchase of wood and coals were in 2019.
2.2. Preparation of Raw Meat Samples
Triplicate portions of proximate to 50 g of each meat sample were obtained by using stainless steel scalpels, individually, to avoid contamination. All samples were ground in a domestic processor with stainless steel blades (Thermomix TM5 equipment—Vorwerk L.L.C., Wuppertal, Germany) and homogenized to secure faithful batches of each type of meat. The samples were individually placed in universal sterilized plastic collectors previously identified and stored in a freezer at −20 °C until the time of analysis.
2.3. Preparation of Raw Meat Samples for Barbecue Grills
Slices proximate to 90 g in triplicate of three different samples of raw meat with 1.70 cm thickness were made, using stainless steel scalpels. Samples were divided into three groups: raw samples, samples roasted on a barbecue with wood or charcoal as fuels, and samples of hamburger roasted on an electric barbecue. The whole process of possible roasting combinations of the various meat samples on the masonry barbecue grill, using different types of wood and coals, was tested in triplicate and according to
Table 1.
A masonry barbecue that was 60 cm long, 37 wide, and 31 cm deep was used to roast the meat. The wood or charcoal was evenly distributed inside the masonry barbecue. Meat samples were placed on a stainless steel grid, at the height of 40 cm from the wood or charcoal. Each sample of meat on the masonry barbecue grill remained for 30 min in the presence of firewood or charcoal, until roasting to the point. After the meat was roasted, samples were sliced, using a scalpel with stainless steel blades. Then, the samples were weighed and homogenized (Thermomix TM5 equipment—Vorwerk L.L.C., Wuppertal, Germany), and then stored in a universal plastic collector and frozen at −20 °C until analysis.
The following procedures were performed for hamburger samples roasted on an electric barbecue: Before the start of the cooking experiment, the electric barbecue grill was washed with detergent, rinsed thoroughly, and dried. Burgers of proximate to 50 g of four different samples of raw meat previously processed were made. The meat was heated by using an electric barbecue grill (Electric Barbecue Mister Grill Plus equipment—Cotherm LTDA, São Paulo, Brazil), at its maximum setting. After fifteen minutes of grilling, the meat was flipped, using a disposable plastic spoon, and grilling continued for fifteen more minutes, for a total of 30 min of grilling. The electric barbecue grill was switched off, and it was found that the meat was cooked in the center and roasted to the point. After the burgers were roasted, samples were sliced, using a scalpel with a stainless steel blade. Then, they were weighed, homogenized (Thermomix TM5 equipment—Vorwerk L.L.C., Wuppertal, Germany), and stored in a universal plastic collector and taken for analysis. Experiments were performed in triplicate.
2.4. Preparation of Raw and Roasted Meat by Microwave Digestion
A 300 mg mass of each sample was cut with stainless steel scalpels and then weighed (raw meat and roasted meat) in the digestion vessels. We added 2 mL of HNO
3 (65% Merck, Darmstadt, Germany), 1.5 mL of H
2O
2 (30% Merck, Darmstadt, Germany), and 2 mL of ultrapure deionized water (18 MΩcm, Milli-Q Millipore, Bedford, MA, USA) to each sample. The samples were digested in a microwave digestion system (Speedwave four
®, Berghof, Germany). This application used the temperature program shown in
Table 2. The samples were then transferred to clean a polyethylene tube and diluted with ultrapure water to 10 mL. All digestions were performed in triplicate for fresh and roasted meat samples and analytical blanks.
2.5. Elemental Measurement by Using ICP-OES
The determination of major and trace elements in samples of different types of raw and roasted meat was performed by using an inductively coupled plasma optical emission spectrometer (ICP-OES) with an axial view (iCAP 6300 Series, Thermo Scientific, Waltham, MA, USA). The instrumental and operating parameters for ICP-OES are shown in
Table 3.
An addition/recovery test for the elements under study was carried out in a meat sample by spiking (0.5 and 1.0 mg/L of each analyte).
Table 4 shows that the method had a recovery interval of 90–111% for the spike of 0.5 mg/L and 93–112% for the spike of 1.0 mg/L. Thus, the recovery test shows that there were no systematic errors or losses of elements during the digestion process.
2.6. Calibration Procedure
The calibration curves for all the analytes were built on nine different concentrations over the range of 0.005–2 mg/L. A multielement solution containing 100 mg/L Al, Cu, Fe, Mg, Mn, Mo, Ni, Co, V, and Zn (SpecSol-Quinlab, São Paulo, Brazil) and a monoelementar solution containing 100 mg/L As, Cd, Cr, and Pb (SpecSol-Quinlab, São Paulo, Brazil) of each element were used to build calibration curves.
The calculation of the limits of detection (LOD) and limits of quantification (LOQ) was according to the analytical standards established by the IUPAC [
31].
Table 5 shows the parameters of the calibration curve, as well as LOD and LOQ values and the obtained correlation coefficient (
R2). The values of LOD were in the range 0.0002–0.0045 (mg/L) and the LOQ were 0.007–0.0151 (mg/L).
2.7. Human Health Risk Assessment
The method used in this study was for non-carcinogenic and adapted from the method described by Onsanit et al. [
9]. The human health risk was assessed by considering the
EDI for a chemical contaminant in meats, as well as the intake amount of the meats. It is possible to calculate the
EDI (µg/kg bw/day) through consumption of food, using the following equation:
where,
Cmeat,
dcmeat, and
bw (bodyweight) represent heavy metal content in raw and roasted meats (µg/g), daily meat consumption per capita (g/day), and the adult’s body weight. The average weight of a Brazilian adult is 70 kg, and the average daily consumption of beef, pork, poultry, and lamb in Brazil is 63.2, 8.5, 36.5, and 0.8 g/person/day, respectively [
32]. Other than that, the risk to human health by intake of heavy-metal-contaminated meats was characterized by using a hazard quotient (
HQ). The
HQ is a ratio of
EDI and chronic oral reference dose (
RfD), which is determined by the following equation:
The
RfD values for the risk calculation were established by the Joint Food and Agriculture Organization/World Health Organization Expert Committee on Food Additives [
6] and the United States Environmental Protection Agency (USEPA) [
33]. The
RfD values for the elements are as follows: Al = 0.4 µg/kg bw/day, As = 0.3 µg/kg bw/day, Cr = 3 µg/kg bw/day, Cu = not available, Fe = 700 µg/kg bw/day, Mn = 140 µg/kg bw/day, Mo = 5 µg/kg bw/day, V = 9 µg/kg bw/day, Mg = 11,000 µg/kg bw/day, Ni = 20 µg/kg bw/day, Cd = 1 µg/kg bw/day, Pb = 4 µg/kg bw/day, Co = 30 µg/kg bw/day, and Zn = 300 µg/kg bw/day [
33]. In Equation (2), toxic risk is considered to occur if
HQ > 1, while
HQ < 1 represents negligible hazard (adverse non-carcinogenic effects).
2.8. Statistical Analysis
One-way ANOVA statistically analyzed differences between the groups, with post hoc Tukey’s test multiple comparison in GraphPad Prism 8.0 software. The significance level was set at p < 0.05.
4. Conclusions
The present study showed that there are statistical differences in the contents of major (Mg) and trace elements (Al, As, Cr, Cu, Fe, Mn, Mo, V, and Zn) in raw meats, meats roasted with an electric grill with electric grill, and roasted meats on barbecues grill, using different fuels.
The concentrations of Al, Cr, Cu, and Fe in raw meats were below the values obtained by other countries. However, high levels of As in the raw beef topside, raw pork loin, raw chicken breast, and Mg, V, and Zn in all raw meats were quantified in this study. The content of Cu, Cr, and As were higher in meats roasted using wood. On the other hand, there was an increase in the values of V, Al, and Fe when the meat was roasted using charcoal.
The concentration of Ni and Cd was high when the meats were roasted using an electric grill. In fact, the material from which electric grills are made can contaminate meat. Mo levels were quantified in the raw and roasted leg of lamb shank with the electric grill, and in the roasted beef and leg of lamb shank, using different fuels. On the other hand, the variations in the concentrations of Mg, Mn, and Mo in the roasted meat are due to the presence of particles and gases from wood and coal. The levels of all measured elements in raw meats and in electric grilled meats are usually lower than those in the roasted meats with coal and wood, except for Ni and Cd.
There are no values established by MRL for Fe, Mg, Ni, and Mn. In addition, there is no value of PTDI suggested by JECFA for Cr, Mo, Mg, and V. The EDI of Al, Cu, and Zn are below the limit of JECFA and MRL reference levels. However, the EDI values of Cr, Mo, and V found in this study did not exceed the MRL standard. The EDI values of Fe and Mn are below the PTDI value. The EDI values of Mg in all samples of meat were below the permissible limits by UL. However, EDI values of As in some roasted meats are above of EFSA and JECFA limits.
Samples of roast chicken breast with wood fire and coal presented HQ values of Al and As for adults above 1. Moreover, HQ values of Cr, Cu, Fe, Mn, Mg, Mo, V, and Zn for adults in all roasted meats were less than 1. According to the Agency for Toxic Substances and Disease Registry, the effects of exposure to any hazardous substance depend on the dose and principally the duration.
Although HQ is less than 1, the health hazards associated with the use of woods and charcoal are of significant concern, and smoke associated with this meat-roasting process represents a significant source of contaminants and can add to the ingestion source. Levels of Cd and Ni were detected in the meats prepared in the electric grill, in opposition to the meats roasted with different types of woods and coals, which raises a concern on the day-to-day use of this equipment present in several households.
Finally, the metals and metalloids emitted by the combustion of coal or wood combustion in restaurants or domestic kitchens, wood stoves, and barbecue grills should be investigated, considering the risk to children, the elderly, and pregnant women, especially. High exposures to a long-term contaminant may cause damage to health.