Published on International Journal of Engineering & Industry
Publication Date: January, 2020
Chris U. Aghalibe, Jude C. Igwe, Anthony I. Obike & Kelechi E. Onwuka
Department of Pure and Industrial Chemistry, Abia State University
In this study, the effects of cow dung, poultry manure and NPK amendment/nutrient as supplements to biostimulate autochthonous microflora for polycyclic aromatic hydrocarbon (PAH) biodegradation were investigated in microcosms containing soil spiked with crude oil (10 % w/w).The rates of biodegradation of PAH were studied for 8 weeks remediation period under laboratory conditions. The results showed that there was a positive relationship between the biodegradation rate and presence of the soil amendment agents in microcosms simulated crude oil contaminated soil. The biodegradation data fitted well to second-order kinetic model. From the biodegradation rate constant obtained from the application of second order kinetics, it can be concluded that the order of PAH removal is NPK > PM > CD. This is because rate constant values (ki) are 0.0315 (NPK) > 0.0196 (PM) > 0.015 (CD). The system proposed here takes advantage of the bulking properties of the amendment agents as well as the autochthonous microorganism metabolic activity to efficiently degrade petroleum hydrocarbons. This system is inexpensive, efficient, and environmentally friendly and may thus offer a viable choice for petroleum hydrocarbons-contaminated soil remediation.
Keywords: Biodegradation; Biostimulation; Crude oil; PAH; Biodegradation kinetics.
Crude oil is the parent material for petroleum-based products used as major sources of energy for industry and daily life. Leakages and accidental spills occur regularly during the exploration, production, refining, transport, and storage of petroleum and petroleum products. The contamination of soil by crude oil and petroleum products has become a serious problem that represents a global concern for its potential consequences on ecosystem and human health . These spills cause soil pollution which is a major issue in Nigeria, especially in the Niger Delta states leading to huge environmental degradation .
The technology commonly used for soil remediation includes evaporation, dispersion, and washing. However, these technologies are expensive and can lead to incomplete decomposition of contaminants . For this reason an increasing attention has been directed toward the research of new strategies and environmental-friendly technologies to be applied for the remediation of soil contaminated by petroleum hydrocarbons. Among these, bioremediation technology which involves the use of microorganisms to detoxify or remove pollutants through the mechanisms of biodegradation has been found to be an environmentally-friendly, non -invasive and relatively cost-effective option . Crude oil bioremediation in soil can be promoted by stimulation of the indigenous microbial population, by introducing nutrients and oxygen into the soil (biostimulation) .
Following oil pollution, nutrients are rapidly assimilated by soil microorganisms thus depleting the nutrient reserves . Therefore, apart from the environmental problems caused by oil pollution, the agronomic and economic aspects are significant . The objective of using soil amendments is to augment the native fertility status of such soil and to enhance the rate of oil degradation, thus minimizing the contamination of scarce groundwater sources and to improve crop production . The addition of inorganic or organic nitrogen-rich nutrients (biostimulation) is an effective approach to enhance the bioremediation process. Positive effects of nitrogen amendment using nitrogenous fertilizer on microbial activity and/or petroleum hydrocarbon degradation have been widely demonstrated [9, 10, 11].
The search for cheaper and environmentally friendly options of enhancing petroleum hydrocarbon degradation through biostimulation has been the focus of research in recent times [11, 12, 13, 14]. One of such option is the use of organic wastes derived from plant and animals. Few workers have investigated the potential use of plant organic wastes such as rice husk and coconut shell , plantain peels and cocoa pod husk , moringa oleifera and soya beans  and animal organic wastes like cow dung, pig dung, and goat dung [10, 16] as biostimulating agents in the clean-up of soil contaminated with petroleum hydrocarbons and were found to show positive influence on petroleum hydrocarbon biodegradation in a polluted environment
Cow dung, poultry droppings and NPK fertilizer are readily available manures for soil. Therefore the main aim of this study is to use these manures to degrade polycyclic aromatic hydrocarbon in a crude oil impacted soil, compare the degradation efficiency of the amendment agents and suggest appropriate kinetic models to show the order of degradation.
2. Materials and methods
2.1 Sample collection
The soil sample used for the study was collected from the top surface soil (0 – 15cm) behind science block, Abia State University, Uturu. Abia State, Nigeria. The soil sample was air dried for two weeks, homogenized, passed through a 2-mm (pore size) sieve, stored in a polyethylene bag and kept in the laboratory prior to use. The cow dung was obtained from the cow market in Lokpa, Abia state, Nigeria, the poultry droppings from a local poultry farm in Okigwe, Imo State Nigeria and NPK (15:15:15) fertilizer obtained from Eke market in Okigwe, Imo State, Nigeria. All the different amendment agents were air dried for two weeks, ground and sieved to obtain uniform sized particles. The crude oil used for the study is Bonny light crude obtained from the core analysis laboratory of the Nigeria National Petroleum Corporation (NNPC), Moscow Rd., Port Harcourt, Nigeria. Each amendment agent was stored in a polyethylene bag and kept in the laboratory prior to use.
2.2 Characterization of soil sample and amendment agents
The soil sample and amendment agents were characterized for Total organic carbon (TOC), total nitrogen (N), Total phosphorus (P), Moisture Content (MC) and pH according to standard methods. The pH was determined by glass electrode pH meter in (1:2.5) soil: water ratio; Total organic carbon was determined by the modified wet combustion method  and Total nitrogen was determined by the semi-micro-Kjeldhal method . Available phosphorus was determined by Brays No.1 method .
2.3 Solid-Phase Experimental Design and Soil Treatment
150g each, of the soil sample was measured out, placed in plastic containers, labelled A- P, respectively. Sample A was used as the control while samples B-P was divided into five (5) groups to represent the different time intervals of 1, 2, 4, 6 and 8 weeks respectively. The soil in each plastic container was spiked with 10% (w/w) bonny light crude oil and thoroughly mixed together to achieve complete artificial contamination. 10% spiking was adopted in order to achieve severe contamination because above 3% concentration, oil has been reported to be increasingly deleterious to soil biota and crop growth . One week after contamination, the different remediation treatments was applied. 30g each, of the amendment agents which is equivalent to 2000kg/hectare, was measured out and added, one to each container in a group. The same was repeated for all five (5) groups representing each time interval. Each container was made up to 50% volume by distilled water for proper percolation. The contents of each container was tilled daily to ensure homogenization and adequate aeration. The contaminated soil in plastic container A, was without amendment agents and thus served as control.
2.4 Sample analysis
10g of the soil sample was weighed into extraction bottle and 20mL of extraction mixture (DCM: Hexane: acetone) in ratio 2:2:1 was added. The mixture was sonicated for 1hr and the organic aqueous layer was decanted. Extracted organic phase was dried using anhydrous sodium sulphate and concentrated using vacuum rotary evaporator gas to about 1.0mL. Extract was fractionated by using column packing. The column was packed by placing 1g of glass wool into the column and gently packed. 1ml of silica gel was placed on it and 1ml of sodium sulphate was added on top of the silica gel. The column was pre-conditioned by running 10ml of n-hexane through the column. 1ml of the concentrated extract was placed on the column and eluted with 10ml n-hexane, 10ml of DCM was allowed to drain through the column and 1.0 ml of the aromatic extract was injected into already PAH calibrated standard GC and result was expressed in mg/kg.
3. Results and Discussion
The results of the soil and amendment agents characterization is as shown in table 1
Table 1: Characterization of soil and amendment agents
The PAH removal efficiency of the amendment agents (cow dung, poultry manure and NPK) is as shown in figure 1.
Figure 1: Removal efficiency of PAH
Figure 1 presents the profile for the removal of PAH such that values of 86.9 % (CD), 90 % (PM) and 93 % (NPK) removal efficiency were recorded after eight weeks. The performance of the materials (CD, NPK and PM) are very good, but the performance of NPK was best.
The analysis of variance (ANOVA) for the biodegradation of PAH with rence to time and materials is presented in table 2.
Table 2: ANOVA results for biodegradation of PAH
Table 2 presents analysis of variance (ANOVA) results for the biodegradation of PAH. Table 1 presents values of significance for material and time as 0.000 and 0.009 respectively. These values are less than 0.05 suggesting that the use of different materials (CD, PM and NPK) and the time had significant effects on the biodegradation of PAH. In addition, values of mean square (MS) are 6.367 and 918.332 respectively for material and time. Higher MS values for time indicates that time had more effects than materials.
Biodegradation kinetic data for the biodegradation of PAH was fitted to zeroth, first and second order kinetic equations. These kinetic equations are:
Zeroth order: (1)
First order: (2)
Second order: (3)
where [A]0 and [A]t are the amounts of PAH present at the beginning of the experiment and at various time intervals, ki is the ith order rate constant and t stands for the various time intervals. Kinetics of biodegradation of PAH gave the following results:
Zeroth order: (4)
First order: (5)
Second order: (6)
Of the R2 values obtained, those of the second order kinetics was highest for biodegradation by CD, NPK and PM. This suggests that the biodegradation of PAH followed a second order kinetics. On this basis, the following reaction equation is suggested:
Chemical reaction is usually associated with second order kinetics [21, 22, 23, 24, 25, 26] therefore, we propose that the biodegradation of PAH involved a chemical reaction mechanism. Figure 2 presents the profiles of PAH biodegradation from experimental and kinetic equations.
Figure 2: Profiles of PAH biodegradation by a) Cow Dung; b) NPK; c) PM
We therefore propose rate equations for the biodegradation of PAH by CD, NPK and PM thus:
The present studies confirm that the use of cow dung, poultry manure and NPK improved the rate of PAH biodegradation in contaminated soil. The biodegradation rate constant obtained from the application of second order kinetics described the rate of PAH biodegradation with the biostimulants, it can be concluded that the order of PAH removal is NPK > PM > CD. This is because rate constant values (ki) are 0.0315 (NPK) > 0.0196 (PM) > 0.015 (CD).
The bioremediation technique proposed here for soils contaminated with crude oil and other petroleum hydrocarbons could be suitable in field, because of its low costs and its low environmental risk associated with volatile hydrocarbon losses. However, the success and efficiency of these biostimulation techniques may vary considerably from one site to another; hence, there is no universal soil treatment regimen for the bioremediation of all petroleum hydrocarbon-contaminated soils. The effectiveness of any soil treatment applied for such purpose has to be evaluated on a case-specific basis.
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