Characterizing Polycyclic Aromatic Hydrocarbons in Smoked Chicken from Ilorin and Implications for Human Health ^†

Abstract

Smoked chicken products from the Nigerian Stored Products Research Institute and three locations in Ilorin, Nigeria, were analyzed for polycyclic aromatic hydrocarbons (PAHs) using gas chromatography. Total PAH levels ranged from 490.893 to 509.064 μg/kg across samples, with benzo(α)pyrene levels (0.947–1.072 μg/kg) within the safe limit of 2 μg/kg. However, PAH4 levels (477.771–491.757 μg/kg) exceeded the European Union’s safe limit of 30 μg/kg. Mean Estimated Daily Intake, Carcinogenic Risk, and Toxicity Equivalent Factor were 0.299 µg/kg/day, 45.341, and 1.443, respectively. Regular monitoring and enforcement of quality control standards are crucial for consumer safety while promoting best practices across the industry.

1. Introduction

Smoking meat remains essential for food preservation in regions with limited refrigeration, particularly in developing countries [1]. However, this process introduces polycyclic aromatic hydrocarbons (PAHs)—compounds formed during incomplete combustion that pose significant health risks [2].

Polycyclic aromatic hydrocarbons comprise over 200 compounds characterized by fused aromatic ring structures. The US Environmental Protection Agency has classified 16 PAHs as priority pollutants based on toxicity profiles [3]. These PAHs are further categorized based on their structural characteristics and health impacts. Light PAHs (2–3 rings) typically exhibit acute toxicity but lower carcinogenicity, while heavy PAHs (4–6 rings) demonstrate greater carcinogenic and mutagenic potential. Benzo(α)pyrene (BaP), a 5-ring structure, serves as a marker compound and is classified as a Group 1 human carcinogen by the International Agency for Research on Cancer [4]. The PAH4 group consists of four high-concern compounds—benzo[a]pyrene, benzo[a]anthracene, chrysene, and benzo[b]fluoranthene—and their combined presence is assessed using Toxicity Equivalent Factors to determine cumulative carcinogenic risk [4].

In Nigeria, smoked chicken is popular among street vendors, yet data on PAH levels, estimated daily intake (EDI), and associated health risks remain scarce [5]. This study analyzes BaP, total PAHs, and PAH4 in smoked chicken samples from Ilorin, Kwara State, with results compared against safety limits established by the Joint FAO/WHO Expert Committee on Food Additives [6]. The findings will help evaluate potential health implications from consuming PAH-exposed smoked chicken in this region.

2. Methodology

2.1. Study Area and Sampling Points

The study was conducted in Ilorin, Kwara State, Nigeria (8.5373° N, 4.5444° E), in June 2023. The map of Ilorin metropolis and the sample collection areas are shown in Figure 1 and Figure 2, respectively. The areas include NSPRI (Sample A), Maraba (Sample B), Al Hikmah University Area (Sample C), and Tanke Junction (Sample D). The maps were created using Google Earth Pro, with all geographic coordinates verified against satellite imagery.

2.1.1. NSPRI Processed Chicken Sample

Two broiler chickens were slaughtered, cleaned, and seasoned (with a 1:2:2 mixture of salt, ginger, and seasoning powder, respectively). The seasoned samples were smoked using kiln technology at the Nigerian Stored Products Research Institute (NSPRI). Hot, moist air at temperatures ranging from 90 to 110 °C was applied for six hours, followed by an additional six hours of drying. NSPRI smoked chicken (Sample A) serves as the control (Figure 3).

2.1.2. Street-Sampled Chicken

Smoked chicken samples were collected from three locations in Ilorin: Maraba (Sample B), Figure 4; Al Hikmah University Area (Sample C), Figure 5; and Tanke Junction (Sample D), Figure 6. These samples were separately homogenized and kept encased in foil at 4 °C for further use.

2.2. Sample Preparation, Extraction, and Detection of PAHs

2.2.1. Extraction and Cleanup

Following Smith et al. [7], 2 g of homogenized smoked chicken samples were mixed with analytical-grade (Merck, Boston, MA, USA) anhydrous sodium sulphate (Na₂SO₄) to remove moisture. Analytical-grade (Merck, USA) dichloromethane (CH₂Cl₂) was used for extraction. The extract was cleaned using a Na₂SO₄-packed column, concentrated via rotary evaporation, and re-dissolved in 1 mL of CH₂Cl₂.

2.2.2. Gas Chromatography Analysis

A Hewlett-Packard 5890 Series II gas chromatograph, equipped with a flame ionization detector (FID), was utilized to analyze the polycyclic aromatic hydrocarbons (PAHs) in the samples. PAH compounds were identified by comparing their retention times to those of a standard PAH mixture. This mixture included a range of PAHs: Naphthalene, Acenaphthylene, Acenaphthene, Fluorene, Phenanthrene, Anthracene, Fluoranthene, Pyrene, Benzo(α)anthracene, Chrysene, Benzo(β)fluoranthene, Benzo(κ)fluoranthene, Benzo(α)pyrene, Indeno(1,2,3-cd)pyrene, Dibenzo(a,h)anthracene, and Benzo(g,h,i)perylene. Then, the concentrations of the PAHs were quantified using external calibration curves. This method ensured the accurate identification and quantification of the PAHs present in the samples.

2.2.3. PAH Classification

In our study, PAHs were categorized as carcinogenic or non-carcinogenic based on International Agency for Research on Cancer (IARC) and US Environmental Protection Agency (EPA) evaluations. The non-carcinogenic group includes Fluorene, Naphthalene, Pyrene, Acenaphthylene, Anthracene, and Phenanthrene, which are not classified as significant human carcinogens by IARC. The carcinogenic group consists of Indeno(1,2,3-cd)pyrene, Benzo(g,h,i)perylene, Benzo(a)pyrene, Chrysene, Benzo(b)fluoranthene, Benzo(a)anthracene, Dibenzo(a,h)anthracene, and Benzo(k)fluoranthene. These compounds are classified by IARC as either Group 1 (known human carcinogens) or Group 2B (possible human carcinogens). This classification enables more accurate health risk assessment and supports targeted risk management strategies for smoked chicken products.

2.2.4. Method Validation

Analytical validation of the method was conducted through calibration curves, limits of detection (LOD), limits of quantification (LOQ), and recovery studies to ensure reliability and accuracy. Calibration curves were established using three standard solutions with concentrations ranging from 0.2 to 10 mg/L, providing the basis for quantifying the PAH congeners. The LOD and LOQ for each PAH class were determined to evaluate the method’s sensitivity and ensure accurate detection and quantification at low concentration levels. Recovery studies were performed by spiking the samples with a known concentration of PAHs (0.05 mg/L) and analyzing them in triplicate.

2.2.5. Health Risk Assessment

The health risks from PAH exposure in smoked chicken were evaluated using standardized methods from multiple studies [8,9,10,11]. The assessment incorporates four key parameters:

The Estimated Daily Intake (EDI), which measures how much PAH-contaminated food a person consumes daily, accounting for their body weight, is calculated using the following expression [8]:

$$EDI=C\times {\displaystyle \frac{IR}{BW}}$$

(1)

where C = PAH concentration, IR = ingestion rate, and BW = body weight (70 kg).

The Toxic Equivalent Concentration (TEQ) evaluates overall toxicity by considering both the amount of each PAH present and its relative toxicity compared to a reference compound and is evaluated using the following formula [9]:

$$TEQ=\sum \mathrm{C}\mathrm{i}\times \mathrm{T}\mathrm{E}\mathrm{F}\mathrm{i}$$

(2)

where Ci = PAH concentration, and TEFi = toxicity factor.

The sum of four key carcinogenic PAHs, specifically looking at the total amount of four major cancer-causing PAHs (BaA, Chr, BbF, and BaP) present in food, is calculated using the following equation [10]:

$$\mathrm{P}\mathrm{A}\mathrm{H}4=\mathrm{B}\left(\mathsf{\alpha }\right)\mathrm{A}+\mathrm{C}\mathrm{h}\mathrm{r}+\mathrm{B}\left(\mathrm{b}\right)\mathrm{F}\mathrm{L}+\mathrm{B}\left(\mathsf{\alpha }\right)\mathrm{P}$$

(3)

The Cumulative Cancer Risk (CR), which estimates lifetime cancer risk by combining daily exposure levels with established cancer slope factors, is evaluated by the following equation [11]:

$$CR=\mathrm{S}\mathrm{l}\mathrm{o}\mathrm{p}\mathrm{e} \mathrm{f}\mathrm{a}\mathrm{c}\mathrm{t}\mathrm{o}\mathrm{r}\times \mathrm{E}\mathrm{D}\mathrm{I}\times \mathrm{E}\mathrm{x}\mathrm{p}\mathrm{o}\mathrm{s}\mathrm{u}\mathrm{r}\mathrm{e} \mathrm{d}\mathrm{u}\mathrm{r}\mathrm{a}\mathrm{t}\mathrm{i}\mathrm{o}\mathrm{n}$$

(4)

The final assessment uses carcinogenic toxicity equivalents (TEQs) to estimate the overall cancer risk by summing the potencies of individual PAHs [12].

3. Statistical Analysis

Data on LOD, LOQ and contamination indices were analyzed for significance using ANOVA (SPSS v2.0, IBM, Chicago, IL, USA).

4. Results of Validation Method, Total Polyaromatic Hydrocarbons, Benzo (α) Pyrene, and the Four Polyaromatic Hydrocarbons

4.1. Validation and Quality Control of PAH Analysis

Table 1. Limits of detection (LOD) and limits of quantification (LOQ) for the 16 PAHs (means and standard deviation of triplicates).

S/N	PAH	LOD (mg/L)	LOQ (mg/L)
1.	Naphthalene	0.0000023 ± 0.0000001	0.0000015 ± 0.0000001
2.	Acenaphthylene	0.0000034 ± 0.0000002	0.0000025 ± 0.0000001
3.	Acenaphthene	0.0000030 ± 0.0000001	0.0000021 ± 0.0000001
4.	Fluorene	0.0000026 ± 0.0000001	0.0000017 ± 0.0000001
5.	Phenanthrene	0.0000041 ± 0.0000002	0.0000032 ± 0.0000002
6.	Anthracene	0.0000084 ± 0.0000004	0.0000052 ± 0.0000003
7.	Fluoranthene	0.0000091 ± 0.0000005	0.0000063 ± 0.0000003
8.	Pyrene	0.0000075 ± 0.0000004	0.0000049 ± 0.0000002
9.	Benzo(α)anthracene	0.0000069 ± 0.0000003	0.0000044 ± 0.0000002
10.	Chrysene	0.0000042 ± 0.0000002	0.0000036 ± 0.0000002
11.	Benzo(β)fluoranthene	0.0000066 ± 0.0000003	0.0000039 ± 0.0000002
12.	Benzo(κ)fluoranthene	0.0000033 ± 0.0000002	0.0000022 ± 0.0000001
13.	Benzo(α)pyrene	0.0000052 ± 0.0000003	0.0000039 ± 0.0000002
14.	Indeno(1,2,3)pery-lene	0.0000036 ± 0.0000002	0.0000028 ± 0.0000001
15.	Dibenzo(α,h)anthracene	0.0000014 ± 0.0000001	0.0000011 ± 0.0000001
16.	Benzo(g,h,i)perylene	0.0000014 ± 0.0000001	0.0000012 ± 0.0000001

shows the limits of detection (LOD) and limits of quantification (LOQ) for the 16 PAH classes.

Table 2. ANOVA results for comparing LOD and LOQ (mean concentrations) of 16 PAHs, showing F-statistic, degrees of freedom, and p-value.

Source of Variation	Degrees of Freedom (df)	Sum of Squares (SS)	Mean Square (MS)	F-Statistic	p-Value
Between Groups	15	1.23 × 10−10	8.20 × 10−12	82.0	<0.0001
Within Groups	32	3.20 × 10−12	1.00 × 10−13	-	-
Total	47	1.26 × 10−10	-	-	-

shows that the ANOVA analysis revealed a p-value well below the 0.05 significance level, indicating significant differences in mean PAH concentrations. This finding validates our method’s ability to distinguish between various PAHs.

The recovery rates for PAH analyses, ranging from 95.3% to 104.8%, demonstrate high accuracy and reliability of the method. These values, close to the ideal recovery of 100%, indicate that the method consistently performs well across different PAHs, ensuring precise and dependable results. Chromatograms from the recovery studies conducted on the four sample sets are shown in Figure 7, Figure 8, Figure 9 and Figure 10.

4.2. Total Polyaromatic Hydrocarbons, Benzo (α) pyrene, and the Four Polyaromatic Hydrocarbons

Figure 11 illustrates the total concentrations of polycyclic aromatic hydrocarbons (PAHs), Benzo(α)pyrene (BaP) and PAH4 (BaA, Chr, BbF, and BaP) in smoked chicken samples, with the data summarized as follows.

Total PAHs: The total PAH content varied significantly across samples, with values ranging from 490.88 µg/kg to 509.06 µg/kg (p < 0.05). Sample A had the highest concentration, while Sample D had the lowest.

Benzo(α)pyrene (BaP): Levels of BaP, a key carcinogenic congener, ranged from 0.94 µg/kg to 1.07 µg/kg. Sample A exhibited the highest BaP concentration, and Sample B had the lowest.

PAH4: The combined concentration of the four major carcinogenic PAHs (BaA, Chr, BbF, and BaP) was highest in Sample A (6.97 µg/kg) and lowest in Sample B (5.34 µg/kg). PAH4 levels generally exceeded those of BaP, except in Samples B and D, where no significant difference was observed (p > 0.05).

4.3. Assessment of Carcinogenic Risk Indices

Figure 12 presents the estimated carcinogenic risk indices of PAHs in smoked chicken samples, which exhibited significant variation (p < 0.05). The indices assessed include the Estimated Daily Intake (EDI), Toxicity Equivalent Factor (TEQ), and Carcinogenic Risk (CR).

Toxicity Equivalent Factor (TEQ): The TEQ values, which measure the overall toxicity of PAHs relative to benzo(α)pyrene, ranged from 1.33 to 1.57 across the samples. Sample C had the highest TEQ, indicating a greater relative toxicity compared to the other samples.

Carcinogenic Risk (CR): The CR levels, which estimate the lifetime cancer risk, varied between 39.01 and 50.91. Sample B had the lowest CR, suggesting a reduced cancer risk, while Sample A had the highest value. However, Samples C and D did not show a significant difference in CR (p > 0.05), indicating similar cancer risk profiles for these samples.

Estimated Daily Intake (EDI): The EDI, which estimates the daily consumption of PAHs, was generally low in all samples, including the control. Sample D had the highest EDI at 0.36, while Sample B had the lowest at 0.23.

5. Discussion and Health Risk Implications

Total PAH concentrations in smoked chicken from Ilorin ranged between 490.88 and 509.06 µg/kg, aligning with levels found in other smoked meats [13], although differences in smoking methods make direct comparisons challenging [14,15]. Benzo(α)pyrene (BaP) levels (0.94 to 1.07 µg/kg) were found below the EU limit, suggesting safety, but local practices may affect PAH formation [16]. Existing research primarily focuses on other meats, highlighting the need for more studies on PAH contamination in street-vended smoked chicken [17].

6. Conclusions

PAHs were found in all smoked chicken samples, with BaP levels below the EU limit but total PAH and PAH4 concentrations exceeding safe levels, posing health risks. TEQ and CR values indicated increased carcinogenicity risk, highlighting the need for improved processing, vendor training, and quality control.

7. Limitations and Future Directions

This study’s limitations include a focus on Ilorin and a lack of detailed consumer data on smoked chicken consumption. Future research should expand the sampling, source additional data on consumption in children and adults, explore PAH formation, and calculate the EDI and TEQ for each individual PAH compound and for each specific subgroup of PAHs, as well as develop standardized protocols and local safety guidelines to improve public health and smoking practices.