Assessment of Heavy Metal Concentrations in Pawpaw (Carica papaya Linn.) around Automobile Workshops in Port Harcourt Metropolis, Rivers State, Nigeria

Background. Consumption of plants such as Carica papaya grown around automobile workshops is common in big cities in Nigeria. However, little is known about the heavy metals contamination of these consumables due to the influence of automobile emissions during maintenance activities. Objectives. This study aimed to assess heavy metal concentrations in C. papaya and supporting soils around automobile workshops in Port Harcourt Metropolis, Rivers State, Nigeria. Methods. Seven automobile workshops were used for the present study. First, 20 m × 20 m quadrats were laid out for soil and C. papaya tissue sampling. One composite soil sample was collected from the topsoil (0–15 cm depth) around each of the automobile workshops. Three C. papaya stands at least 30 cm apart around each workshop were used for the study and from these stands, tissues (root, stem, leaf, fruit) of C. papaya were collected. Standard laboratory techniques were used to determine the pH, electrical conductivity (EC) and heavy metals (lead (Pb), mercury (Hg), cadmium (Cd), copper (Cu), and zinc (Zn)) in the soil samples and C. papaya tissues. Pairwise t-test was used to determine significant differences (p<0.05) in the heavy metal concentrations in soil and C. papaya tissues between the sample and control sites, while correlation statistics were used to determine the relationship of heavy metal concentrations between soil and C. papaya tissues. Results. C. papaya tissues and supporting soil had significantly higher levels of pH, EC and heavy metals in the sampled plots than the control plot. The heavy metal concentrations in C. papaya and soil occurred in the decreasing order of Pb>Cu>Hg>Zn>Cd. The fruit of C. papaya had the highest mean concentrations of Pb (51.4±14.1 mg/kg) and Zn (26.4±1.9 mg/kg), while the leaf had the highest mean concentration of Hg (32.0±2.3 mg/kg). The pH, Cu and Zn in the supporting soil were significantly correlated with the levels in the C. papaya tissues. Conclusion. Bio-accumulation of heavy metals by C. papaya is evident around automobile workshops, and Pb, Hg, Cd concentrations were found to be above the permissible limits for human consumption according to World Health Organization (WHO) standards. Consumption of food materials grown around automobile workshops could pose health risks.


Introduction
Automobile workshops are sources of auto wastes in cities and pose environmental hazards in developing countries. Auto wastes include used oil and fluids, dirty shop rags, waste from solvents used for cleaning parts, metallic particles from machinery wear, used batteries, asbestos from brake pads, oxidation products, and organic and inorganic chemicals used in oil additives and metals. 1, 2 The release of automobile wastes is responsible for the accumulation of heavy metals in soils and vegetation around the automobile workshops. 3 Automobile workshops accumulate heavy metals concentrations in soil, creating environmental and health hazards. 4 The soils in the tropics are capable of supporting complete food production and modifying carbon cycling, but soil pollution from automobile waste has devastated the soil. 5 Automobiles in Nigeria mainly use petrol and the smoke releases and other factors. 7 The effects of contaminants from automobile workshops have reached an alarming rate in Nigeria as heavy metal contamination has affected food at the trophic level as well as organisms. 8 Soil pollution is a main cause of soil resource degradation. 9 Heavy metals are absorbed by growing crops and transferred to other parts of the plant. 10 The destroyed soil leads to low yields in crop production and low quality crops. 11,12 Vegetables, tubers, flowers and trees such as pawpaw (Carica papaya) and other medicinal plants are often cultivated around these automobile workshops.
There have been a number of studies on heavy metals in soils and plants, including an assessment of heavy metal contents in roadside dust of Dhaka Metropolitan City, Bangladesh, a study of the effects of automobile fumes on the concentration of Pb in bread in Port Harcourt City, Nigeria, a study of the concentration of Pb in sweet orange in Port Harcourt, and an examination of the influence of automobile workshops along major roads of Port Harcourt City on soil properties. 3,7,13, 14 Other studies include an examination of the levels of total hydro-carbon in plants along the roads Port Harcourt City, the effects of heavy metals on topsoil at the vicinity of automobile mechanic villages in Owerri, Nigeria, and the level of pollution of the soil caused by automobile workshop waste. [15][16][17][18][19][20] None of these studies have investigated the influence of automobile workshop wastes on C. papaya, despite the fact that it is a common plant growing naturally around mechanic workshops in Nigeria. C. papaya is a fruit commonly consumed all around the world, especially in the developing world. This is due in part to its reputation for having medicinal properties. 21 This study examined the impact of automobile wastes on the tissues (root, leaf, bark and fruit) of C. papaya in Port Harcourt Metropolis, Nigeria.

Study Site
The study was carried out around mechanic workshops in selected communities in Obio/Akpor, Ikwerre and Emohua Local Government Areas of Rivers State, Nigeria. The communities included Choba, Alakahia, Rumuosi, Rumualogu, Igwuruta and Emohua. An automobile mechanic workshop was selected in each of the communities based on the abundance of pawpaw in that environment and whether the automobile workshop had been in existence for at least 10 years. All study locations had the same soil type, climate and ecological zone (rainforest). The study area was located between latitudes 4°51' 30''N and 4° 57' 30''N and longitudes 6°50' 00''E and 7°00' 00''E ( Figure 1), in the sub-equatorial region with a tropical climate. The mean annual temperature and relative humidity are 30 o C and 85%, respectively. The rainfall is seasonal, variable and heavy, and mean annual rainfall is about 2,300 mm. 22 The prevailing wind is basically south-westerly and north-easterly, with a wind speed between 5 and 17 m/s. 23 The topography of the study area ranges between 16 m and 40 m above sea level. 22 The vegetation type resembles that of a tropical rainforest. The vegetation is consistently nourished with high rainfall and high temperatures, providing favorable conditions for the growth of varieties of tall trees such as Swietenia macrophylla, Triplochiton scleroxylon, Terminalia superba, Elaeis guineensis and Raphia hookeri. 24 The soils of the area can be categorized as freshwater brown and sandy loams.

Research Design and Site Description
The study included experimental plots (automobile workshops) and a purposively selected control plot (natural forest). The control plot is about 1000 m away from any automobile shops or dumpsites of any sort. Seven automobile workshops were identified at least 200 m apart and with respect to their locations, these included S1 (Wilbros); S2 (UNIPORT); S3 (Choba); S4 (Alakahia); S5 (Rumuosi); S6 (Igwuruta); S7 (Rumualogu) and the control (Emohua). Study Research various illnesses. The soil around the automobile workshops is black in color as a result of petrol, diesel, engine oil and spray paints. In addition, there are piles of automobile waste products including containers for spray paints, thinner, petrol, diesel, and residue of carbide (CaC 2 ) around the automobile workshops.

Collection of Carica papaya Leaf, Stem, Root and Fruit and Soil Samples
A test plot measuring 20 m x 20 m was laid out around the pile of refuse in each automobile workshop and a control plot also measuring 20 m x 20 m was sited on the floor in the natural forest in Emuohua. In each quadrat, three mature C. papaya plants measuring 30 m were identified. Soil samples were collected from the topsoil (0-15 cm) around the identified C. papaya plants. Three soil samples were collected around each C. papaya stand with a soil auger and mixed in a bowl. A composite sample was taken from the C. papaya stand and put into a labeled polythene bag. Altogether, 21 soil samples were collected from both the test and control plots and taken to the laboratory for further analysis. C. papaya leaf, stem, root and fruit were collected from the same C. papaya stand identified for soil sample collection. From each C. papaya stand, a total of three leaf, stem (base, middle, top), root and fruit samples were collected. Papaya leaves, stems, roots and fruits collected from each stand were homogenized to make a composite sample of each of the three sites and labeled accordingly. 25 The C. papaya tissue sample collection was performed using a stainless steel knife. All samples were well washed with distilled water to remove adsorbed soil particles and then air-dried.

Laboratory Analysis of Carica papaya Tissues
Fruit samples were scraped to separate the seed from the fruit and thereafter only the fruit was crushed. Subsequently, the leaves, stems and roots were crushed. All tissues were oven-dried at 70 o C for 24 hours. After cooling at ambient temperature, the dried tissues were milled into fine powder and sieved using a 2 mm mesh criteria for the sampled workshops were existence for a minimum of 10 years and an abundance of pawpaw around the workshop. C. papaya was selected for the study because of its abundance around all of the automobile workshops. In addition, C. papaya produces edible fruit, and the leaf, stem and roots are used to treat The blank reagent and standard reference soil materials were included in each sample batch to verify the accuracy and precision of the digestion procedure and for subsequent analyses. All samples were quantified in quadruplicate. The elemental analyses were performed in the agronomy laboratory of the University of Ibadan, Ibadan, Nigeria.

Method of Data Analysis
Descriptive and inferential statistics were applied in the present study.

Field Procedures
All field procedures were in accordance with general and standard Quality Assurance and Quality Control requirements of the USEPA. 33 Contamination of samples was avoided by using clean and sterile sampling containers. Separate samples were used for analyzing parameters requiring different treatment or preservation before analysis. A composite sampling technique was adopted for soil. 34 Control samples were collected at appropriate points remote from focus areas (about 1000 m away from the nearest automobile workshop). Samples were appropriately preserved and labeled. Proper chain of custody was applied.

Laboratory Procedures
Analyses were carried out within the holding time of respective parameters. Only functional and calibrated equipment was used for sample and thereafter kept in pre-cleaned screw-capped polyethylene containers for further analysis. The sample digests of the fruit, leaves, stems and roots were analyzed in three replicates for pH, electrical conductivity (EC), Pb, cadmium (Cd), Cu, mercury (Hg) and Zn. The heavy metals (Pb, Cd, Cu, Hg and Zn) in the C. papaya tissues were determined by atomic absorption spectrophotometry (Buck Scientific Model 210/211 VGP, US) using the Association of Official Agricultural Chemists (AOAC) standard. 26 Then pH was determined using a water solution, while EC was determined using a conductivity meter. 27 The elemental analyses were done in the agronomy laboratory of the University of Ibadan, Ibadan, Nigeria.

Laboratory Analysis for Soil Samples
Soil pH was measured potentiometrically in 0.01 M calcium chloride solution using 1:2 soil solutions. Electrical conductivity was determined using a 1:2 ratio of soil sample to distilled water measured with a digital conductivity meter. 25 Extracts used for determining heavy metals (trace elements) were obtained by leaching soil samples using 0.1 N ethylenediaminetetraacetic acid (EDTA) and 5 g of each sample was weighed into a clean, dry silica dish, covered and ignited in a furnace for 6 hours at 500°C until a grey white ash was obtained. 28 The cover of the dish was opened to allow for escape of gases. To cool ash samples, 5 ml of 10% hydrochloric acid was added to enhance dissolution and 5 ml of 10% nitric acid was subsequently added and set in a water bath to dissolve completely. The solution was later relocated into a clean dry 50 ml standard volumetric flask and made to mark with distilled water. 29 The concentrations of extractable trace metals including Cu, Pb, Cd, Hg and Zn were determined using atomic Research analysis. Only trained and experienced staff were involved in the analytical work.

Effects of automobile workshops on heavy metals uptake in Carica papaya tissues
Findings revealed that pH ranged between 5.0 and 6.1 in the root of C. papaya with mean pH of 5.67 around automobile workshops, while in the control site, pH ranged between 7.8 and 8.3 with a mean pH of 8.09.
Similarly, the mean pH in the leaf, fruit and stem were lower than that of the control plot, suggesting that the pH level in the C. papaya tissues around the automobile workshops was acidic and alkaline in the control plot. Differences between the control and test plots are shown in Table 1 workshops occurred in the decreasing order of Pb>Cu>Hg>Zn>Cd (Figure 2). Comparing the level of concentrations of heavy metals with the WHO permissible levels, all investigated heavy metals were higher than the permissible level, indicating that the plants were polluted ( Table 1).

Effects of Automobile Workshops on pH, EC and Heavy Metals in Soil
The pH level at the test site was significantly more acidic than at the control plot (t=6.  Table 2). Compared with control samples, the concentrations of Hg, Cd, Cu and Zn were significantly higher in the test samples ( Table 2). The concentrations of Hg, Cd and Cu around the automobile workshops were higher than the permissible limit of the DPR, while the concentrations of Pb, Cd, Cu and Zn from the test and control plots were lower than the permissible limits of the USEPA (Table 2). Similar to the tissue of C. papaya, the mean concentrations of heavy metal contents in soils around the mechanic workshops occurred in the decreasing order of Pb>Cu>Hg>Zn>Cd.

Contamination Factor and Pollution Loading Index of Heavy Metals in Soil
Contamination factor and pollution load index were used to assess heavy metal contaminations in soil located

Table 2-pH, EC and Heavy Metals Concentrations in Soil
Eludoyin. Ogbe

Figure 2 -Trend of heavy metals concentrations in Carica papaya tissues
around the automobile workshops ( Table 3). The contamination factors for the five metals in different locations were high, ranging from 0.8 to 32.5 in Zn. In all the locations, Zn had the highest contamination factor, except in S4, while Hg was the lowest, except in S4, S5 and S6. The contamination factors of all the heavy metals indicate severe to very severe contamination ( Table 4). The PLIs of all the study locations were greater than unity and the majority indicated moderate pollution levels, except locations S5 and S6 (severe pollution levels) and S7 (very severe pollution) ( Table 3).

Translocation Factor
Translocation factor, the ratio of the concentration of metal in the aerial portion of the plant to the total concentration in the soil sample, is presented in  (4.47). The TFs in the study area were generally very high, as they were greater than 1.

Relationships between pH, EC and Heavy Metals between Carica papaya Tissues and Soil
The correlation analysis revealed in Table 6 shows that pH in soil was positively correlated with pH in the root (r=0.785, p<0.05); leaf (r=0.761, Eludoyin

Discussion
The pH in the tissues of C. papaya and soil around the automobile workshops was acidic in nature. However, previous studies have reported that the soil around the automobile workshops was relatively alkaline. The concentration of Pb was observed to be the highest in all of the tissues of C. papaya around the automobile workshops and significantly higher than the control site and the permissible levels of 0.3 mg/kg as specified by the WHO. The bioaccumulation of Pb was higher in the underground organs (rhizomes and roots) of Hieracium pilosella. 57 The higher Pb concentration in C. papaya tissues and soils around automobile workshops is in agreement with findings of previous studies.  66 Pb is known to be toxic even at low concentrations, especially in young children. 67 It has been reported that automobile workshops may be used as playgrounds or be located near residential areas where children play about freely and it is possible for children to ingest Pb through hand to mouth behavior, especially during the developmental years from ages 0-2. 47 Thus, ingestion is considered to be the most significant pathway for Pb, which could lead to sensory disturbances, kidney disorders and brain damage. 67 The concentration of Cu was significantly higher in the soil and C. papaya tissues around the automobile workshops compared to the control plot and the permissible level of 36 mg/kg in soil as specified by the DPR.
The high concentration of Cu may be dangerous as it damages the liver and kidney when it gets into the human body through the food chain. The presence of Cu may be attributed to automobile workshops, traffic densities and the use of auto paints. 62,68 Findings also showed that heavy metals in soil were positively correlated with heavy metals in the tissues of C. papaya, except for Cd in root. This suggests that the absorption of heavy metals in the tissues of C. papaya was greatly controlled by the content of heavy metals in the soil solution and bioavailability of metals in the supporting soil, and as result, C. papaya can be potentially used in the bio-monitoring of environmental contamination of heavy metals. 25, 57 However, pH in soil significantly influenced Pb levels in all the tissues, Cu in the root, fruit and stem; and Zn in all the C. papaya tissues investigated. Zinc in soil was negatively and significantly correlated with Pb in root, leaf, fruit and stem.

Research
The high negative correlation between Zn in soil and Pb in tissues gives a strong indication that they have different sources of contamination in the environment. 69,70

Conclusion
Bio-absorption of heavy metals (Pb, Cu, Hg, Cd, Zn) by the tissues of C. papaya is evident around mechanic workshops in Port Harcourt Metropolis, and their uptake is directly linked to the supporting soil. The accumulation of heavy metals in the root, leaf, fruit and stem of C. papaya and soils around mechanic workshops was significantly higher than in the control plot. The heavy metal concentrations in the tissues of C. papaya and soil around mechanic workshops occurred in the decreasing order of Pb>Cu>Hg>Zn>Cd. Mercury had the highest plant/soil metal concentration ratio and translocation factor despite its lower bioavailability compared to Pb and Cu. Positive and significant correlations existed between the heavy metals in the tissues of C. papaya, except for Cd in root and heavy metal contents in soil. We strongly recommend that the consumption of the leaf, stem, root and fruit of C. papaya grown around automobile workshops be discouraged. Indiscriminate disposal of automobile waste should be properly examined by the government so that this practice can be reduced or completely eradicated. Finally, since C. papaya has the tendency and potential to absorb heavy metals, it can be grown in surplus around automobile workshops for phyto-extraction of heavy metals and further studies on the level of accumulation of heavy metals in the human body, especially among auto-mechanic technicians and residents living around the automobile workshops are needed.