The World Health Organization’s International Agency for Research on Cancer (IARC) established that outdoor air pollution is carcinogenic to humans, as the association between increased incidence of cancer and particulate matter have been established . The presence of daily PM2.5 at 32–158 µg/m3 and PM10 at 33–467 µg/m3 beyond WHO acceptable limits, confirms the likelihood of Port Harcourt residents being exposed to hazardous compounds [60, 61]. High amounts of fine particulate matter in indoor and outdoor environs noted in the study were similar to those found in highly petroleum-based industrial areas [23–27, 62–64]. The observed increase in particulate matter is most likely responsible for the relative upsurge in trends of morbidity and mortality recorded over the years in Port Harcourt, confirming reports from previous studies [19, 20, 62], as toxicity and mortality, cancers, upsurge in respiratory, cardiovascular, and renal dysfunctions resulting from exposure to particulate matter bonded with PAHs have been reported in a plethora of epidemiological studies [47, 65, 44]. Although the ration of PM10 and PM2.5 have not been studied for atmospheric particulate matter of Port Harcourt, studies have implicated infants and children being at risk of fine aerodynamic particulates [22, 66]. The study findings also aggress with earlier reports of weight loss and skin toxicity as observed on the outdoor experimental rats, due to exposure to particulate matter and effects of PAHs [43, 67]. Over 500 PAHs and its related compounds have been found in the air  and the carcinogenic PAHs such as benz[a]anthracene, benzo[a]pyrene, benzo[b]fluoranthene, benzo[k]fluoranthene, chrysene, dibenzo[a, h]anthracene, benzo[ghi]perylene and indeno[1,2,3-cd]pyrene [10, 28]. Most of the PAHs found in the study were mostly of the carcinogenic class. There is strong evidence that injuries to the lung tissues and DNA damage could result from sub-chronic exposure to low amounts of particles, even below the WHO’s 24hours threshold .
Five established carcinogenic PAHs such as benz[a]anthracene, benzo[b]fluoranthene, benzo[k]fluoranthene, chrysene, benzo[ghi]perylene and indeno[1,2,3-cd]pyrene [5, 23] including Phenanthrene were not found in samples of the baseline animal group as against those of indoor and outdoor animal groups (see Table 1). Also, of the mean concentrations noted, the indoor exposed animal group constituted a mere 5% (see Figs. 3 and 4) implicating a relatively clean air in the University of Nigeria Nsukka, where the experimental animals were imported from for the study assessment. This suggests that people in non-hydrocarbon and petroleum-free environments are at lower risk of cancers and general tissue toxicity compared to residents inhaling air polluted with PAHs bonded particulate matter due to petroleum and hydrocarbon related industrial activities [61–67]. Also, the continuous inhalation of PAHs bounded particulate increases the concentrations of these compounds in the tissues, enabling toxicity through oxidative stress, inflammation, and DNA methylation [5, 48].
Nsukka is located in Enugu, south-east Nigeria. It is an agricultural town, with the University of Nigeria (UNN) situated in it and having minimal commercial and industrial activities that are likely to generate pollutants such as PAHs . As seen in Fig. 2 above, the experimental animals were imported from this non-petroleum and hydrocarbon pollution-free environment (UNN) which should ideally record no concentrations of PAHs. Although in lesser amounts, the study found some concentrations of the individual PAHs, including B[a]P in blood samples of the baseline animal group. This is likely to result from pyrogenic sources, including domestic combustion and most probably not from hydrocarbon or petrogenic events [36, 69, 70]. This calls for a further forensic investigation of the presence of abundant alkyl homologs in such an environment.
The indoor air concentration constitutes about 13% of PAHs levels investigated. Though absent in the baseline animal group, the presence of fluoranthene in the ambient of the indoor animal group indicates toxicity (see Table 1), as fluoranthene is said to be a complementary indicator to B[a]P . Studies have demonstrated the burning of biomass, fossil fuel, and solid fuel as important sources of airborne particulates and its component pollutants, including PAHs, which are emitted into indoor environs through unvented or flueless combustion [70, 71]. Reports have shown that low molecule (less than for rings) PAHs predominantly concentrates indoor air , thus the high concentrations of large-molecule PAHs (including dibenz[a,h]anthracene, benzo[b]fluoranthene, benzo[k]fluoranthene, and indeno[1,2,3-cd]pyrene ) know for high carcinogenicity and toxicity noted in the indoor animal groups as against those of baseline, suggests intrusion of outdoor air [5, 73]. Cooking activities and indoor combustion sources such as fossil fuels and biofuel  and indoor environments impacted by ETS  have been reported. Suggesting a higher concentration of some PAHs in indoor exposures likely to occur during the cooking period [17, 74]. In as much as outdoor air influences the concentrations of PAHs in indoors, the type of cooking fuel used impacts greatly on indoor PAHs levels in descending order of dung cake > dung cake/wood mixture > wood > coal > kerosene > LPG [5, 17, 74]. Also cooking temperature has been found to influences the production of most indoor PAHs as increasing temperatures affects evaporation of PAHs into the air and pyrolysis from partially cracked organic compounds .
Higher indoor concentrations of 12 PAHs (naphthalene, acenaphthene, fluorene, phenanthrene, anthracene, fluoranthene, pyrene, benz[a]anthracene, chrysene, benzo[k]fluoranthene, benzo[e]pyrene and benzo[a] pyrene) have been recorded across homes with an unvented fireplace, lacking airtight stoves and windows, homes and public places ( such as restaurants, discotheques, and clubs ) with ETS, while lower levels were noted across offices, hospitals, schools, libraries and coffee shops [17, 75–80] and children were noted to be adversely affected . Conducting the study during the dry season could also have influenced the increased PAHs concentration in the ambient. Though seasonal variations were not investigated, earlier reports have established seasonal differences in PAHs concentrations, with higher levels in rainy (winter) than in dry or summer seasons . Thus, outdoor air, indoor combustion sources, cooking activities, cooking temperatures, indoor ETS, ventilation characteristics of the living house, climatic factors, and seasonal variations all affect indoor PAHs concentrations. This implicates women; especially in Africa have a higher chance of being affected by indoor air pollutants such as PAHs  as their activities are observed to be predominantly indoors.
As seen in Fig. 2 above, 82% of total mean concentrations of PAHs noted, were from the outdoor experimental animal group. This indicates high toxicity of ambient air inhaled by residents and implicates finding of the previous study that used lichen as a bioindicator for air pollution . Combustion in energy and transformation industries, nonindustrial combustion plants, combustion in the manufacturing industry, production processes, traffic road transport, ETS, other mobile sources, waste incineration, agriculture and forestry, natural sources were all found to contributes significantly to outdoor PAHs concentrations in industrialized nations with the variety of sources and emission factor across the countries [82, 83]. High toxicity was predicted for pregnant women in these studies . While in developing countries, pyrogenic and petrogenic combustions for industrial, commercial, and domestic uses involving biomass, fossil fuel and solid fuel, including petroleum and hydrocarbon refining sources contribute to outdoor particulates matter containing PAHs [70, 71]. Although no empirical emission inventory has been conducted for Port Harcourt, places with petrochemical economic and industrial ecology have been reported to have ambient air largely polluted by petroleum and hydrocarbon [23–26, 62, 70, 84, 85] most probably responsible for the increase in trends of morbidity and mortality [20, 23] noted in the metropolis.
With outdoor animal group recording higher concentrations of total PAHs (see Figs. 3 and 4), especially the larger-molecule including dibenz[a,h]anthracene, benzo[b]fluoranthene, benzo[k]fluoranthene, and indeno[1,2,3-cd]pyrene and the recorded experimental dead Wistar rats notably from the outdoor group, continues inhalation of ambient air of Port Harcourt suggests higher chances of tissue toxicity. Especially inhabitants engaged in outdoor activities such as road sweepers, street hawkers, traffic wardens, building, and road constructors. This could be responsible for the clinician’s hematological findings of rare toxins in blood samples of some young patients at tertiary hospitals in Port Harcourt . Hence the medical advice to residents to avoid places where crude oil is cooked, as management of neoplasms and myelofibrosis conditions resulting from crude oil pollution remains expensive, especially for low-income people .
From the study result of outdoor PAHs concentration, it is most probable that the indoor air conditions were influenced by outdoor air [5, 73] This might be further worsened by ETS at open spaces adjacent to night clubs. Unlike risk to indoor air PAHs concentration, men in Africa are highly at risk of PAHs concentration in outdoor air environs  as they carry out more of outdoor occupations. Ambient air in densely populated and metropolitan locations have been demonstrated to have higher PAHs concentrations than less dense, rural, forest, and agricultural areas because of the numerous sources of fossil fuel combustions . Carcinogenic PAHs such as dibenzo[a, h]anthracene, benzo[ghi]perylene, indeno[1,2,3-cd]pyrene and pyrene had the predominant concentrations of PAHs in the study. This agrees with previous studies in which toxicity resulting from ambient exposure in petroleum industrial places was demonstrated [23, 62, 84].
While levels of the PAHs especially Benz(a)anthracene, Benzo(b)fluoranthene, and Fluoranthene remained significantly higher in the outdoor, they were often undetected in the baseline animals. This suggests that, unlike residents in an environment free from petroleum-based ambient air pollution, inhabitants of Port Harcourt metropolis are inhaling into her body system, air poisoned with PAHs due to petroleum activities. Figures 3 and 4 also implicatively show that residents of Port Harcourt are more predisposed to morbidity and mortality associated with PAHs [19, 38, 43, 87, 88], unlike those in hydrocarbon pollution-free environment . This could explain hematologists’ sudden notice of myelofibrosis in some young residents of Port Harcourt involved in artisanal petroleum refining . The clinician observation negates empirical evidence of myelofibrosis found to be common in the older person of 60 + years [23, 89, and 90]. Studies by the United Nations Environmental Programme have shown that substandard oil explorations and artisanal petroleum refining significantly added to the effect of hydrocarbon pollutions which affect both the environment and health . Adversely, exposure to PAHs poses dangers to residents of Port Harcourt, as continuous exposures to outdoor and indoor air polluted with PAHs resulting from petroleum industrial activities likely to increase cases of cancers [59, 62, 92] and deformities, especially in pregnant women, infants and children and obese persons [20, 62]. Adverse birth outcomes such as preterm birth, early pregnancy fetal death, low birth weight likely to be common, as well as intrauterine growth retardation with the capacity to distort the academic performance of children in the future when compared to cohort children from the non-polluted environment [19, 91–93].
Unlike studies were both indoor and outdoor PAHs concentrations were observed to be similar , the PAH levels found in this study were significantly higher in the outdoor animals (82%) when compared to the indoor animals (13%) as shown in Fig. 3 and the mean Pyrene levels in all animals were significantly different. With significantly higher values observed for outdoor animals (Figs. 3 and 4). These findings implicate exposure to the variation of doses of PAHs . Suggesting that when compared to indoor workers, outdoor workers such as street traders, beggars, market sellers, road construction workers, and children in open schools within Port Harcourt metropolis are more at risk of adverse health effects associated with PAHs induced by petroleum-based particulate matter pollution [16, 18]. The notable negation of study results from similar studies [96–98] in which higher levels of indoor PAHs were observed, is likely to result from the difference in geographic location, seasonal variations, and source of particulate matter inducing the PAHs [72, 75]. Deaths recorded from the outdoor experimental exposed rats are likely to result from tissue toxicity beyond the bearable limit and immune suppression prompted by oxidative stress and alteration of DNA methylation and expression of specific genes [44, 65, 94, 95, 99–102], as caution was taken prevent physical stress and trauma on the rats. The study findings explain and implicate previous reports on increased trends of selected mortality and morbidity associated with particulate matter pollution in Port Harcourt between 2016 and 2018 as well as an upsurge in health burden [20, 23]. Similarly, the result also proves that the risk of toxicity, carcinogenicity, and mutagenicity due to high concentrations of PAHs are likely higher in inhabitants living in hydrocarbon industrial polluted environments than those in non-hydrocarbon industrial areas [5, 44, 54, 62, 84, 102–104].
The high levels of carcinogenic PAHs such as dibenzo[a, h]anthracene, benzo[ghi]perylene, indeno[1,2,3-cd]pyrene and pyrene detected in the study, indicate the likelihood of tissue toxicity of exposed inhabitants [17, 23]. Toxicity resulting in high incidences of deaths and cancers of several types and at different tissue locations have been documented in places with exposure to carcinogenic PAHs in petroleum industrial ecology [17, 62, 105–109]. The death of seven outdoor experimental rats before the end of 90days study period, loss of weight, and change in skin color of exposed animals noted in the study remain a concern and the likelihood of such effects on human inhabitants is most probable [17, 23, 62, 107, 109]. And interactions of PAHs with tissues have been found to results in cancers and mortality, . Animal model studies have demonstrates lymphoreticular system tumours associated with exposures to dibenzo[a, h]anthracene, benzo[ghi]perylene, indeno[1,2,3-cd]pyrene [135, 136]. Implicatively, this suggests, that exposed inhabitants were most probably susceptible to the toxicity of PAHs [137–141], due to its suppression of immunosurveillance and immunocompetence of body .