A combined landfarming-phytoremediation method to enhance remediation of mixed persistent contaminants

Contamination of soil and water with petroleum hydrocarbons and metals can pose a signi�cant threat to the environment and human health. This study aimed to investigate the establishment and growth of high fescue and agropyron in two petroleum-contaminated soils (soil S1 and soil S2) with previous landfarming treatments, and to assess the phytoremediation potential for heavy metal removal from these polluted soils. The results showed that the presence of petroleum hydrocarbons signi�cantly (P < 0.05) reduced plant growth, but plant development was facilitated in soils with prior landfarming treatments. Urease activity in the rhizosphere of agropyron for soil S1 was about 47% higher than the unplanted control soil. The rhizosphere of agropyron and tall fescue eliminated more than 40% and 20% of total hydrocarbon amounts in soil S1, respectively, compared to the unplanted soil. Moreover, the plants grown in the landfarming treatment exhibited higher concentrations of metals (Fe, Zn, Mn, Cu, and Ni) than the control. Based on the �ndings, the combination of landfarming and phytoremediation techniques can provide an optimal solution for removing mixed pollutants, including petroleum hydrocarbons and metals, from the environment.


Introduction
Petroleum hydrocarbons, which are widely distributed in the environment, are a group of carcinogenic contaminants that raise serious concerns due to their potential for bioaccumulation in food chains (Truskewycz et al., 2019).Contaminated soils with petroleum hydrocarbons are prevalent worldwide, requiring the development of effective and low-cost in-situ technologies for remediation (Akpokodje et al., 2022).
Phytoremediation offers a promising alternative to expensive and disruptive engineering-based methods of remediation.It involves utilizing plants to effectively eliminate, break down, con ne, or neutralize environmental pollutants, thereby making them harmless (Majumder et al., 2017).Plant-associated micro ora and organic/inorganic substances exuded by plants in their rhizosphere facilitate the breakdown or transformation of pollutants into non-toxic forms (Siddiqua et al., 2022).
Several studies have demonstrated the effectiveness of speci c plant species in stimulating the degradation of total petroleum hydrocarbons (TPHs) in soil and con rming the relationship between microbial activity in the rhizosphere and the breakdown of petroleum compounds (Akpokodje & Uguru, 2019; Majumder et al., 2017).For instance, Nedunuri et al. (2000) reported that some plants release enzymes and exudates that can intensify the microbial activity in the rhizosphere that have a variety of petroleum-degradation characteristics (Nedunuri et al., 2000).
To be effective, the rate of TPH removal and degradation must be accelerated above mechanical or microbial processes.Heavy fractions of petroleum hydrocarbons are less phytotoxic than complete hydrocarbons, and soils polluted with petroleum hydrocarbon contaminants elevate the levels of heavy metals in the environment, posing extreme challenges to human and animal health (Correa Garcia et al., 2018).
In addition, soils polluted with petroleum hydrocarbon contaminants cause to elevate the levels of heavy metals in the environment, which hold extreme challenges to human and animal health (Koshlaf & Ball, 2017).Phytoremediation technology has been demonstrated to be a long-lasting solution for the remediation of such contaminated soils (Koshlaf & Ball, 2017).This research investigates the establishment and growth of plants in petroleum-polluted soils and evaluates the effects of vegetation on microbial activity indicators and TPH removal from soil.Additionally, the study assesses the potential of a combined landfarming-phytoremediation for the remediation of heavy metals from petroleum-polluted soil.

Soil sampling and analysis
Bulk soil samples were gathered from two different locations: an area designated for oily wastes disposal (referred to as soil S1), and petroleum-contaminated farmlands (referred to as soil S2), situated near the Tehran Oil Re nery in Iran (coordinates 35° 30' N, 51° 26' E).Additionally, an uncontaminated soil sample was collected from a nearby unaffected area for comparison.To ensure an even distribution of petroleum pollutants, the collected soil samples were air-dried and then passed through a 4-mm sieve.They were subsequently subjected to a land-farming technique, where the soil was mixed with a garden hoe at 3-day intervals over a period of 21 days.Following the treatment process, 1-kg subsamples of the treated soils were sieved through a 2-mm sieve for further chemical and physical analyses.Several parameters were evaluated for the soil samples.Soil pH and electrical conductivity (EC) were measured in a 1:5 ratio of solid-to-liquid aqueous extract.The available-phosphorus content was extracted from the soil using a 0.5 M NaHCO3 solution.Total nitrogen (N) content was determined using the micro-Kjeldahl method.The physical and chemical properties of the soil samples, as measured, are presented in Table 1.To assess the levels of DTPA-extractable metals in the soil samples, including Zn, Fe, Mn, Cd, Ni, and Cu, the Lindsay and Norvell (1978) method was employed (Besalatpour et al., 2008).The results of these metal analyses are provided in Table 2.

PAH and TPH analysis
The study used a Soxhlet apparatus to extract petroleum hydrocarbon contents from soil samples using a 1:1 (v/v) mixture of dichloromethane and n-hexane (150 ml) for 24 hours (Ngwenya & Mahlambi, 2023).The extracted samples were then analyzed for selected polycyclic aromatic hydrocarbons (PAHs) using gas chromatography (GC) with a Delsi DI 200 chromatograph equipped with a direct injection port and ame ionization detector (FID).The GC settings for both sets were 340 ºC, with helium used as the carrier gas under 0.08 MPa.The column was a CP Sil 5 CB (Chrompack) capillary column (50 m by 0.32 mm, with a lm thickness of 0.25 µm), and the temperature program was 100 to 320 ºC, at 3 ºC min−1.
The initial concentrations of PAHs and total petroleum hydrocarbons (TPHs) in the soil samples are presented in Table 3.

Landfarming experiments
Before conducting phytoremediation trials, the study evaluated the effects of landfarming operations on the remediation of petroleum-contaminated soils to determine their impact on the phytoremediation process.During the landfarming operations, the contaminated soils were manually mixed using a garden hoe every 3 days.This process facilitated the distribution and interaction of the pollutants within the soil.Irrigation was carried out at approximately 0.7 eld capacity, ensuring that the soil moisture level was optimal for microbial activity and pollutant degradation.Before each irrigation event, the soils were turned over to expose a fresh layer to sunlight and air, which enhanced the remediation process.In addition to the landfarming plots, control plots were set up for comparison.The control plots underwent the same treatment procedures as the landfarming plots, including manual mixing, but without irrigation or aeration.This allowed for a clear distinction between the effects of landfarming and the natural attenuation processes occurring in the control plots.The experiment spanned a duration of 4 months, allowing su cient time for the remediation process to take place and for the effectiveness of the landfarming method to be evaluated.

Phytoremediation trials
After the completion of the landfarming trials, soil samples were immediately utilized to create contamination treatments for phytoremediation experiments.Two types of contamination treatments were prepared: one involved soil samples from the landfarming treatment, which had undergone landfarming operations, and the other served as a control without landfarming operations.To set up the experiments, approximately 3 kg of soil samples were placed in plastic pots with a diameter of 150 mm and a height of 250 mm.To prevent soil from escaping through the drainage holes, a lter paper disc was positioned at the bottom of each pot.Seeds of tall fescue (Festuca arundinacea L.) and agropyron (Agropyron smithi L.) were then planted in the pots, with a planting depth of 1 to 2 cm.These plant species were chosen for their potential to aid in the phytoremediation process.Unplanted soil was also included in each contamination treatment.The pots were watered from the top to keep soil moisture near 70% eld capacity during the 18-week experimental period.No additional nutrients were added to the greenhouse to replicate a natural environment, and the minimum and maximum temperatures in the greenhouse during the experiment were 22 ºC and 37 ºC, respectively.Upon concluding the experiment, all plants were harvested, and their shoot and root components were separated.Both the shoots and roots were carefully washed using tap water followed by distilled water to remove any external debris.After washing, the plant materials were dried and weighed to determine the biomass yields, providing insight into the growth and productivity of the plants in each treatment.In addition to plant analysis, soil samples were collected from the rhizosphere, which refers to the soil surrounding the plant roots, speci cally targeting the area in uenced by root activities.Furthermore, soil samples were also taken from a depth of 5 to 8 cm in the unplanted soil of each treatment.This allowed for the assessment of the impact of the plants on the soil and the potential effects of the different treatments on soil composition and characteristics.

Microbiological analysis
To assess the soil microbial activity in the phytoremediation experiments, we measured the basal soil respiration and urease activity.For the basal soil respiration, we placed 50 g of soil samples into 250-ml glass containers, closed them with rubber stoppers, and collected the evolved CO2 by trapping it in NaOH solution.We titrated the alkali excess with HCl to determine the amount of CO2 produced (Kassem Alef, 1st Edition -June 16, 1995).For the urease activity, we added 2.5 ml of urea solution to 5 g of soil samples in 100-ml Erlenmeyer asks, and then incubated them for 2 h at 37°C.A control without urea was also included for each sample.After incubation, we added 50 ml of KCl solution, shook the mixture for 30 min, and ltered the suspension.We then analyzed the ltrates for the ammonium content, which re ects the urease activity.

Metal analysis in plant tissues
The root and shoot samples were ashed following the protocol described by Munter et al. (1984).Brie y, plant material weighing 0.5 to 1 g was placed in a ceramic crucible and dry-ashed in a mu e furnace for 18 hours at 550°C.After cooling, the ash was dissolved in 5 mL of 20% HCl and diluted to 25 mL with distilled water.The concentration of Cu, Zn, Ni, Cd, Mn, and Fe in the samples was determined using an atomic absorption spectrophotometer (Munter et al., 1984).

Statistical analysis
Statistical analysis was conducted using a completely randomized design and a factorial trial in the SAS statistical computer program (Parsad, 2010).Analysis of variance was performed to assess the signi cance of differences between the means.

In uence of TPHs on plant growth
The presence of petroleum hydrocarbons in the soil had a signi cant impact on the growth of the plants being studied.The in uence of total petroleum hydrocarbons (TPHs) on the root dry matter yield (RDMY) of the plants is depicted in Fig. 1.In soil S1, the RDMY of agropyron decreased by approximately 35% and 50% in the landfarming treatment and control group (without landfarming operations), respectively, compared to the uncontaminated soil.However, in soil S2, the RDMY of agropyron in the landfarming treatment was 40% higher than that of the control group.The RDMY of tall fescue in soil S1 declined by 75% and 85% in the landfarming treatment and control group, respectively, compared to the uncontaminated soil.Additionally, tall fescue seedlings exhibited high sensitivity to petroleum pollutants in soil S2, resulting in poor growth and eventual death within 8 to 9 weeks (Fig. 1).
The presence of petroleum hydrocarbons in the soil signi cantly affected the growth of the plants under investigation.Figure 1 illustrates how TPHs impacted the root dry matter yield (RDMY) of these plants.In soil S1, the RDMY of agropyron was approximately 35% and 50% lower in the landfarming treatment and control group (without landfarming operations), respectively, compared to the uncontaminated soil.Conversely, in soil S2, the RDMY of agropyron in the landfarming treatment was 40% higher than that of the control group.The RDMY of tall fescue in soil S1 decreased by 75% and 85% in the landfarming treatment and control group, respectively, compared to the uncontaminated soil.Additionally, the presence of petroleum pollutants in soil S2 had a detrimental effect on tall fescue seedlings, leading to poor growth and eventual death within 8 to 9 weeks (Fig. 1).
Furthermore, the shoot dry matter yield (SDMY) of the plants indicated a slow start with low aboveground biomass production in both soils S1 and S2, particularly in the control treatment compared to the uncontaminated soil (Fig. 1).In soil S1, tall fescue produced an average accumulated shoot dry biomass of 1.4 g and 0.9 g in the landfarming treatment and control, respectively, representing a decrease of approximately 66% and 77% compared to the unpolluted soil.The aboveground biomass of agropyron in the landfarming treatment and control for soil S1 was signi cantly reduced by about 55% and 61%, respectively, compared to the unpolluted soil.Furthermore, the shoot dry weight of agropyron in both the landfarming and control treatments for soil S2 was signi cantly lower than that of the uncontaminated soil.It seems that the phytotoxic effects of petroleum hydrocarbons were lower in the soil in which the landfarming operations were conducted before the phytoremediation trial.This facilitated the establishment and development of the plants.Landfarming operations remove the light fractions of petroleum hydrocarbons that induce the most signi cant phytotoxic effects on seed germination and plant seedlings' growth (Besalatpour et al., 2010).The reduction in dry matter yield of the studied plants was most likely due to the inherent toxicity of remaining hydrocarbons in the soils.The hydrophobic properties of the TPH contaminants might have also caused restrictions in water and nutrient availability to the plants and perturbed their root development, which can be another crucial factor that affected plant growth and biomass production (Koshlaf & Ball, 2017).
Multiple studies have provided evidence of the detrimental effects of petroleum hydrocarbons on soil, speci cally in relation to plant germination and growth.For example, Kulakow et al. conducted a study where all legumes died within 60 days when cultivated in sediments contaminated with hydrocarbons over a long period (Kulakow et al., 2000).Merkl et al. observed a signi cant decline in shoot and root dry matter production in various grass species when grown in petroleum-contaminated soil (Merkl et al., 2004).Rui Liu et al. also discovered phytotoxic effects of hydrocarbons (Liu et al., 2012).Chaineau et al.
concluded that the inhibition of plant growth increased as the concentration of total petroleum hydrocarbons (TPH) increased, although it was not a linear relationship.They reported over an 80% reduction in biomass for wheat and beans at a hydrocarbon concentration of 0.3%, and a decrease of less than 30% for maize at 1.2% concentration (Chaîneau et al., 1997).

Soil microbial respiration
The measurement of soil microbial respiration provides insights into the microbial activity in the soil, as well as the quality and quantity of mineralizable substrates (Weaver et al., 1994).The results of the study are presented in Fig. 2, which illustrates the CO2 evolution values generated by soil microbial respiration in the rhizosphere of the plants under investigation and in unplanted soil for each treatment in soils S1 and S2.In soil S1, the microbial respiration in both vegetated treatments was signi cantly higher (P < 0.05) than in the unplanted soil.Additionally, the CO2 evolution values in the rhizosphere of agropyron were higher than those of tall fescue in both soils S1 and S2.Moreover, in the landfarming treatment for soil S1, microbial respiration in the rhizosphere of agropyron increased by approximately 42%, while in the control group, it increased by about 26%, compared to the unplanted soil.Similarly, in the landfarming treatment and control for tall fescue, the CO2 evolution due to microbial activity in the rhizosphere was 28% and 25% higher than in the unplanted soil, respectively.These ndings suggest that the superior establishment and subsequent growth of agropyron, particularly in terms of higher root biomass production (Fig. 1), resulted in increased microbial activity in its rhizosphere compared to tall fescue.Although microbial respiration in the vegetated landfarming treatment was relatively higher than in the control treatment for both soils S1 and S2, the differences were not signi cant for tall fescue in soil S2.
The results of this study provide strong evidence that vegetated petroleum-contaminated soil exhibits greater microbial activity than unvegetated contaminated soil.It has been suggested that plants release exudates such as amino acids, organic acids, carbohydrates, growth factors, and smaller soluble proteins in their rhizosphere that can have an impact on the microbial community and activity, resulting in increased soil microbial respiration in the rhizosphere compared to the bulk soil (Egamberdieva et al., 2011).Lie et al. found that the microbial population and respiration in the rhizosphere of plants, particularly grasses, was higher than in unplanted soils, which is consistent with our ndings (Li et al., 2002).In their study, Angela Sessitsch et al. (2013) observed that the CO2 evolution in the rhizosphere of perennial ryegrass exhibited an increase following a lag phase of 6 days upon the addition of 500 mg/g of diesel hydrocarbons to the soil (Sessitsch et al., 2013).Additionally, Gunther et al. demonstrated that in a polluted soil with PAHs and aliphatic hydrocarbons, soil microbial respiration rates in the rhizosphere of ryegrass were higher than in the bulk soil (Günther et al., 1996).

Urease activity
The measurement of soil enzymes such as urease, produced by microorganisms, animals, and plants, can be utilized to evaluate microbial activity in polluted soils, especially in the nitrogen cycle.The ndings of the current study demonstrate that the urease activity in the rhizosphere of plants was signi cantly higher than in the unplanted soil for both soils S1 and S2 (Fig. 2).Speci cally, in the landfarming treatment for soil S1, the urease activity in the rhizosphere of both agropyron and tall fescue was approximately 47% higher than in the unplanted soil.However, there was no signi cant difference (P < 0.05) in the urease activity between agropyron and tall fescue (Fig. 2).Furthermore, in soil S2, the urease activity in the rhizosphere of agropyron increased by about 28% and 20% in the landfarming treatment and control, respectively, compared to the unplanted soil.Overall, the vegetated landfarming treatment exhibited higher urease activity compared to the control treatment, particularly in soil S1.However, the differences in urease activity for tall fescue in soil S2 were minimal.The establishment and subsequent growth of plants were more successful in the landfarming treatment than in the control treatment (Fig. 1), and the urease activity in the rhizosphere of plants was greater in the landfarming treatment than in the control treatment.Plants release some exudates, such as amino acids, organic acids, carbohydrates, growth factors, and soluble proteins in their rhizosphere, which can increase microbial and enzymatic activity in the soil.The higher soil microbial respiration in the rhizosphere of studied plants than in the unplanted soil (Fig. 2

TPH remediation
The results presented in Fig. 3 demonstrate that vegetation signi cantly enhanced the removal of petroleum hydrocarbon contaminants from the soil.The greater dissipation of TPHs in the landfarming treatment as compared to the control treatment can be attributed to better establishment and growth of the tested plants, as well as greater microbial activity in the treatment (as observed in Figs. 1 and 2).In the rhizosphere of agropyron and tall fescue, more than 40% and 20% of the total amount of hydrocarbons in soil S1 were removed, respectively, as compared to the unplanted soil.TPH degradation in the rhizosphere of agropyron in the landfarming treatment for soil S2 was approximately 7% higher than the control treatment (Fig. 3).The brous root systems of grasses like agropyron and tall fescue, along with their extensive surface area for microbial colonization and root exudates, seem to create more favorable conditions for the degradation of petroleum hydrocarbons than unplanted soil (Baghaie & Daliri, 2020).Xu et al. suggested that the enhanced degradation of total petroleum hydrocarbons (TPH) in the rhizosphere of the plants under investigation could be attributed to various changes taking place in the soil as a result of root presence (Xu et al., 2006).These changes include modi cations in chemical characteristics, alterations in microbial composition, and increased microbial activity.
In previous studies, Rui Liu et al. discovered that the dissipation of diesel fuel in the rhizosphere of legumes was greater than in unplanted soil (Liu et al., 2012).Similarly, Sara Correa-García et al. found that the TPH degradation by bermuda grass (Cynodon dactylon L.) and tall fescue during the rst 6 months was not signi cantly different from the unplanted control, but after one year, the TPH degradation was signi cantly higher in both vegetated treatments, with Bermuda grass achieving a mean TPH reduction of 68% and tall fescue achieving a mean TPH reduction of 62% (Correa Garcia et al., 2018).In addition, Merkl et al. evaluated legumes and grasses to determine their ability to stimulate microbial degradation in soils contaminated with 5% (w/w) of heavy crude oil, and discovered that the TPH concentration in planted soil was lower than in bulk soil (Merkl et al., 2004).Brachiaria brizantha L. was found to be the most effective plant for phytoremediation of petroleum-contaminated soils in the tropics.Xu et al. reported a reduction of 92% and 88% in the concentration of phenanthrene and pyrene, respectively, in soils planted with maze after 60 days (Xu et al., 2006).Furthermore, Wiltse et al. found that various strains of alfalfa (Medico sativa L.) were able to reduce crude oil contamination in the rhizosphere by 33-56% compared to the control (Wiltse C.C., 1998).

Iron (Fe)
Figure 4 illustrates the metal concentration in the plant tissues under different treatments for soil S1.The results of the metal contents in the tissues of agropyron for soil S2 are also presented in Fig. 5.As previously mentioned, the presence of petroleum contaminants in soil S2 made the tall fescue seedlings sensitive and unable to produce dry matter yield at the end of the trial period (Fig. 1).The concentration of Fe in the underground tissues of both agropyron and tall fescue was considerably higher than in the aboveground tissues.Moreover, the Fe accumulation in the root and shoot tissues of agropyron was greater in the landfarming treatment for both soils S1 and S2 than in the control treatment.The better establishment and growth of agropyron in the landfarming treatment, compared to the control (Fig. 1), resulted in a higher Fe uptake by the plant.On the other hand, no signi cant difference (P < 0.05) was observed between the landfarming treatment and control in the Fe concentration in tall fescue tissues.However, the Fe accumulation in its underground tissues was signi cantly higher than that in its aboveground tissues.The amount of Fe taken up by agropyron in both the landfarming and control treatments was considerably higher than in the unpolluted soil.The Fe concentration in the agropyron underground tissues in the landfarming treatment and control for soil S1 was 64% and 54% higher than in the unpolluted soil, respectively.Additionally, the Fe accumulation in the agropyron root tissues in the landfarming treatment and control for soil S2 was 63% and 51% higher than in the unpolluted soil, respectively.

Zinc (Zn)
The zinc uptake by Agropyron plants growing in soil S1 was higher compared to soil S2 (see Figs. 4 and   5).The concentration of zinc in the underground tissues of Agropyron in the landfarming treatment for soil S1 showed a signi cant difference (P < 0.005) compared to the control treatment.However, there was no signi cant difference between the landfarming treatment and control in the concentration of zinc in the root and shoot tissues of Agropyron for soil S2.The accumulation of zinc in the underground tissues of Agropyron in the landfarming treatment and control for soil S1 was 51% and 34% higher than the unpolluted soil, respectively.Additionally, the uptake of zinc by tall fescue was signi cantly enhanced in the landfarming treatment for soil S1 compared to the control treatment, and the concentration of zinc in its root tissues in the landfarming treatment was 32% higher than in the control treatment.The accumulated concentration of zinc in the aboveground tissues of tall fescue in the landfarming treatment for soil S1 was also 10% higher than in the control treatment.

Manganese (Mn)
In terms of manganese (Mn), no signi cant differences (P < 0.05) were observed between the landfarming treatment and control in the accumulation of Mn in the root tissues of tall fescue, while the concentration of Mn in its root tissues in the landfarming treatment and control was 40% and 24% higher than the unpolluted soil, respectively (see Fig. 4).Furthermore, the Mn contents in the underground tissues of Agropyron in the landfarming treatment and control for soil S1 were 21% and 15% higher than the unpolluted soil, respectively.A signi cant difference in the accumulation of Mn in the aboveground tissues of Agropyron was observed between the landfarming treatment and control for soil S1.Similar to Fe and Zn, the accumulation of Mn in the roots was greater than in the shoots, indicating low metal translocation in the plants.The total concentration of Mn in the aboveground tissues of Agropyron in the landfarming treatment was also about 35% higher than in the control treatment for soil S2.However, there was no signi cant difference in the accumulation of Mn in its underground tissues (see Fig. 5).

Copper (Cu)
The root tissues of the studied plants, especially agropyron in the landfarming and control treatments, had considerably higher Cu contents compared to the unpolluted soil (Figs. 4 and 5).The average copper concentration in the underground tissues of agropyron in the landfarming treatment and control for soil S1 was 140 and 89 mg kg−1 dry weight, respectively, while it was only 21 mg kg−1 dry weight in the unpolluted soil.There was no signi cant difference (P < 0.05) found between the landfarming treatment and control for soil S1 in the Cu accumulation in the tall fescue root tissues.However, the Cu concentrations in the tall fescue shoot tissues in the landfarming and control treatments were 59 and 82% higher than the unpolluted soil, respectively.Moreover, the differences in the Cu concentration in the agropyron aboveground tissues were not signi cant.

Nickel (Ni)
As with other measured metal concentrations in plant tissues, the Ni accumulation in the root tissues was greater than in the shoot tissues (Figs. 4 and 5).There was also a signi cant difference (P < 0.05) in the Ni concentration taken up by the plants in the landfarming treatment and control.However, accumulated Ni in the aboveground of agropyron in the landfarming treatment was not signi cantly higher than in the control treatment for both soils S1 and S2.In addition, the Ni concentration in the agropyron root tissues in the landfarming treatment for soils S1 and S2 was 78 and 62% higher than the unpolluted soil, respectively.The Ni accumulation in the root and shoot tissues of tall fescue in the landfarming treatment and control for soil S1 was also greater than in the unpolluted soil.

Cadmium (Cd)
The concentration of cadmium in the soils was found to be high (Table 2), however, the accumulation of cadmium in the plant tissues in both the landfarming treatment and control was lower than the sensitivity of the atomic absorption spectrophotometer.It seems that the plants investigated in this study were not capable of accumulating a relatively high amount of cadmium in their tissues, indicating that harvesting agropyron and tall fescue would not be an effective method for Cd-removal from contaminated soils.
Although from a toxicological perspective, this is a desirable property, as cadmium would not pass into the food chain via herbivores, thereby avoiding potential risks to the environment.
The ndings of this study demonstrate that the plants analyzed from the contaminated soils exhibited higher levels of metal accumulation in their tissues compared to the uncontaminated soil (Table 2).In general, the average concentration of metals in the belowground tissues of the studied plants was signi cantly higher than in the aboveground tissues, suggesting that the substrate metals were readily available to the plants but had limited mobility within the plant once absorbed.These results is consistent with earlier studies (Bai et al., 2018).Phytostabilization can be used to minimize the passage of pollutants in the soils.Plants have the ability to immobilize heavy metals through various mechanisms, including absorption and accumulation by their roots, adsorption onto root surfaces, or precipitation within the rhizosphere.These processes help to reduce the mobility of metals and prevent their release into groundwater (Seneviratne et al., 2017).

Conclusion
The present study demonstrated that the prior implementation of landfarming operations facilitated plant establishment and growth, particularly in soil contaminated with oily waste.Microbial respiration and urease activity in the rhizosphere of the examined plants were signi cantly higher than in the unplanted soil.Additionally, the presence of vegetation greatly enhanced the removal of hydrocarbon pollutants from the soil, although total petroleum hydrocarbon (TPH) dissipation was higher in the landfarming treatment compared to the control.The concentrations of Fe, Zn, Mn, Cu, and Ni in the plant tissues from the landfarming treatment were higher than in the control.Notably, the average concentration of metals in the belowground tissues of the plants was signi cantly higher than in the aboveground tissues.
From these ndings, it can be concluded that the combination of landfarming and phytoremediation techniques may overcome the limitations of using each method individually.This suggests that employing a landfarming-phytoremediation approach can be an optimal solution for effectively remedying mixed persistent pollutants such as petroleum hydrocarbons and metals from the environment.Furthermore, it is worth noting that the plants used in this study were able to grow in conditions without the addition of nutrients, which could be advantageous in the revegetation of petroleum-contaminated soil as it reduces the need for costly fertilizer amendments.

Declarations
Con ict of Interest Statement: The authors (Ali Asghar Besalatpour and Mohamad Reza Fadaei Tehrani) con rm that they have no known competing nancial interests or personal relationships that could have in uenced the work reported in this paper.The authors have no relevant nancial or non-nancial interests to disclose.
Funding Statement: This research was not supported by any speci c grant from public, commercial, or not-for-pro t sectors.The authors (Ali Asghar Besalatpour and Mohamad Reza Fadaei Tehrani) declare that no funds, grants, or other support were received during the preparation of this manuscript.
Data availability: The data used in this study are available from the author upon request (mfadaei@nri.ac.ir).
Authors' contribution statements: Both authors (Ali Asghar Besalatpour and Mohamad Reza Fadaei Tehrani) contributed to the study conception and design of the research as well as eld and laboratory analysis.Both authors were also involved in writing the manuscript and have read, commented and approved the nal manuscript.
Ethical responsibilities of Authors: Both authors Asghar Besalatpour and Mohamad Reza Fadaei Tehrani) have carefully reviewed, comprehended, and adhered to the "Ethical Responsibilities of Authors" statement outlined in the Instructions for Authors.They are aware that, with few exceptions, no modi cations can be made to the authorship once the paper has been submitted."The percentage reduction of TPH in the rhizosphere of the examined plants and unplanted soil for each treatment in soils S1 and S2.Signi cant differences at P<0.05 are denoted by different letters The metal concentrations in the plant tissues for different treatments for soil S 1 .Signi cant differences at P<0.05 are denoted by different letters.
) con rms this expectation.Lie et al. and Besalatpour et al. have reported similar ndings regarding microbial activity and urease activity in the plant rhizosphere and the landfarming treatment, respectively (Besalatpour et al., 2011; Li et al., 2002).Margesin et al. stated that urease activity, as well as other biological activity indicators, can be used to monitor the degradation of diesel oil in contaminated soil (Margesin et al., 2000).

Figure 1 Root
Figure 1

Table 1
Physical and chemical properties of the soils collected from an oily wastes land ll (soil S 1 ) and farm lands contaminated by petroleum pollutants (soil S 2 ).

Table 3
Concentration of measured PAHs and TPHs in the soils sampled from oily wastes land ll (soil S 1 ) and petroleum-contaminated farm lands (soil S 2 ) at the beginning of experiment.