Introduction
Diesel is one of the important petroleum products widely used in Iran and other parts of the world. Its release into the environment can cause an imbalance of nutrient levels, especially carbon/nitrogen (C/N) ratios in soils, which is unfavourable for microorganisms' growth. This condition negatively impacts the bioremediation of hydrocarbon pollutants (Molina-Barahona et al., 2004; Sarkar et al., 2005; Khosravinodeh et al., 2013). However, in such conditions in contaminated soil systems, by providing nutrients through fertilization and optimizing the growth conditions of these organisms, it will be possible to adjust these imbalanced ratios (Lee and Ward, 1985; Kim et al., 2005). Using nutrient modifiers is one of the main methods to reduce petroleum pollutants in contaminated soils, accelerating these compounds' biodegradation by providing critical nutrients for microorganisms.
Total nitrogen is the second most influencing factor affecting the distribution and activity of bacteria in soil which mainly affects biodegradation rates of hydrocarbon pollutants (Wang et al., 2018). Many studies have investigated the effect of nitrogen on the biological remediation of contaminated soils (Brook et al., 2001; Gallego et al., 2001; Walecka-Hutchison and Walworth, 2006), and nutritional supplementation of contaminated soils using inorganic N sources is proposed as an effective method in bioremediation process (Toffoletto et al., 2005).
Kinetics analysis of biodegradation reactions has a crucial role in determining the fate of petroleum hydrocarbons in polluted soils (Nabgan et al., 2016; Sharma and Shane, 2016; Yuan et al., 2017; Guo et al., 2018; Safdari et al., 2018; Doustaky et al., 2022) which is based on providing practical trends of hydrocarbon elimination from the contaminated environments. Kinetic modelling is usually performed using first- and second-order models to evaluate the biodegradation rates of total petroleum hydrocarbons (TPH) over time (Yeung et al., 1997; Namkoong et al., 2002; Sarkar et al., 2005; Adesodun and Mbagwu, 2008; Chemlal et al., 2013; Nwankwegu et al., 2016). Based on a computational approach, this modelling analysis can provide estimations of biodegradation constant (K), half-life values (t1/2) and correlation coefficients (R2) for pollutant removal trends (Yuan et al., 2017; Safdari et al., 2018). These valuable indices could be used practically in predicting residual levels of hydrocarbons and the time required for their removal from contaminated soil systems.
The present study aimed to evaluate the effects of different inorganic nitrogenous modifiers on the bio-elimination of hydrocarbon pollutants in diesel-contaminated soil. Three inorganic nitrogen sources, including NPK, Urea, and NH4SO4, were used as soil modifiers at different initial diesel contamination levels of 0, 5 and 10% during three months. Also, kinetic modelling was applied as an essential part of the bioremediation process assessment due to the addition of nitrogen modifiers.

Methodology
The uncontaminated soil was sampled in farmlands (depth = 0 – 20 cm) of Gorgan City, Golestan Province, north of Iran. Three initial gasoil concentrations (0, 5 and 10%) were considered as pollution levels, and initial pollution was applied through complete mixing. The resulting soil mixtures were incubated for one month before N modifier applications (Nwankwegu et al., 2018; Onwosi et al., 2018). The soil moisture was held at 60% FC during this period. Inorganic nitrogen modifiers were applied to the contaminated soils as 80 gr N modifiers per 3 kg soil in each treatment. Total petroleum hydrocarbon (TPH) content, degradation ratio (%D) and microbial respiration (CO2-C) values were weekly measured for three months. TPH measurements were done using the standard methods of the United States Environmental Protection Agency (EPA 4113.1) (Hutchinson et al., 2001). Degradation ratio (%D) was calculated using the following equation:

Where TPHi and TPHr were initial and residual levels of TPH in soil samples, respectively. The Anderson (1982) method was used to determine the microbial respiration (CO2-C). Kinetic modelling analysis was done using the first- and second-order models (Agarry et al., 2013). Also, degradation constant (K), half-life (t1/2) and correlation coefficient (R2) values for TPH data were calculated using these kinetic models. Two-way analysis of variance (ANOVA) was used to evaluate the effects of different nitrogen modifiers and the initial contamination level on TPH and CO2-C. The normality of experimental data was checked using the Shapiro-Wilk test, and if necessary, data transformation was applied to normalize the data. The Comparison of means for experimental treatments was conducted using the Tukey-HSD test at a significance level of 0.05. Statistical analysis was performed using R (version 4.1.2).
Conclusion
The results showed that using inorganic nitrogen modifiers led to significant (p < 0.05) reductions in TPH concentrations in the contaminated soils compared with the control condition. A similar trend was observed for decreasing TPH concentrations in all N modifiers with all initial gasoil concentrations (0, 5 and 10%). Among nitrogen modifiers, the highest degradation ratio belonged to Urea, followed by NPK and NH4SO4. The lowest elimination of TPHs in all initial concentration treatments was found in the control (i.e. no modifier application) condition, and the highest TPH decrease belonged to the Urea application condition. Using Urea fertilizer led to the highest biodegradation ranging from 79.31% (10% initial concentration) to 85.00% (0% initial concentration). The levels of natural degradation of TPHs in the control conditions varied between 41.35 to 48.08% for 10% and 0% initial gasoil concentrations, respectively.
Microbial respiration (CO2-C) levels were significantly (p < 0.05) elevated during the first six weeks of the experimental period due to the effects of nitrogen sources. Using Urea and NPK nitrogen sources had the highest microbial respiration levels (NPK: 2.93 – 3.66 mg C/kg soil; Urea: 2.83 – 3.66 mg C/kg soil) while the lowest levels were obtained with the control condition (1.61 – 2.66 mg C/kg soil). The microbial respiration levels were higher at 5 and 10% initial gasoil concentrations compared to the 0% level.
The first-order kinetic model had better fits and higher predictive performance than the second-order model for the obtained data. Considering all modifier treatments, the biodegradation constant (K) scores decreased with increasing initial pollution concentrations. The highest K values were calculated for Urea application with 0% pollution concentration (first-order: 0.1321 day-1; second-order: 0.4374 day-1), and the lowest K values were obtained for the control condition at 10% pollution level (first-order: 0.0364 day-1; second-order: 0.0258 day-1). Conversely, the lowest half-life (t1/2) scores were obtained using Urea at 0% gasoil pollution (first-order: 5.25 days; second-order: 3.84 days), and the highest scores were calculated with no application of N modifiers (control condition) at the highest initial gasoil concentration (10%) (first-order: 19.4 days; second-order: 22.86 days).
The findings of the present study indicated that using different inorganic nitrogen sources (NPK, Urea and NH4SO4) as soil modifiers in the bioremediation of diesel contamination in soils improved biodegradation efficiency by stimulating microbial activity and metabolism in the contaminated soils. Among nitrogen sources, the highest practical improvement of pollutant removal was obtained with Urea. Also, kinetic modelling showed better performance of the first-order model in predicting the fate of petroleum hydrocarbons than the second-order model. It could be said that using inorganic nitrogen sources could practically result in reductions in diesel contamination levels and their persistence in the soil environment.