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Jeremiah Makarau ILIYA

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Download the complete chemistry project topic and material (chapter 1-5) titled BIOREMEDIATION OF OIL-CONTAMINATED SOIL USING EMULSIFIER (LIQUID SOAP), NPK FERTILIZER AND MICROBIAL (Bacillus sp) TREATMENT here on PROJECTS.ng. See below for the abstract, table of contents, list of figures, list of tables, list of appendices, list of abbreviations and chapter one. Click the DOWNLOAD NOW button to get the complete project work instantly.

 

PROJECT TOPIC AND MATERIAL ON BIOREMEDIATION OF OIL-CONTAMINATED SOIL USING EMULSIFIER (LIQUID SOAP), NPK FERTILIZER AND MICROBIAL (Bacillus sp) TREATMENT

The Project File Details

  • Name: BIOREMEDIATION OF OIL-CONTAMINATED SOIL USING EMULSIFIER (LIQUID SOAP), NPK FERTILIZER AND MICROBIAL (Bacillus sp) TREATMENT
  • Type: PDF and MS Word (DOC)
  • Size: [1,519kb]
  • Length: [141] Pages

 

ABSTRACT

Two different set of soil samples were collected near Kaduna Refining and Petrochemical Company Limited (KRPC), a Subsidiary of Nigerian National Petroleum Corporation (NNPC) Kaduna. The first set of samples labelled (D) was obtained from areas where diesel from the refinery spilled into the environment and the second set of samples labelled (P) was collected from areas where petrol leaked and spilled into the environment. The pH of the soil was found to be 5.9 and 6.2 for D and P samples respectively. The cation exchange capacity (CEC) was higher in sample P than in sample D (32.0 and 30.0 mmol/kg of soil respectively). P has high concentrations of cations ( Ca2+, Mg2+, K+ and Na+ with concentration values 3.6, 1.17, 0.50 and 0.22 mol/kg respectively) because of its high CEC while sample D with a lower CEC has a lower concentration of cation (Ca2+, Mg2+, K+ and Na+ with concentration values 1.20, 0.27, 0.25 and 0.17 mol/kg respectively). The oil (contaminant) was extracted in dichloromethane and a GC-MS analysis was run to determine the nature (composition) of the oil and the concentration of the contaminant was determined using gravimetric method. Results revealed that the total concentration of the contaminant (oil) before treatment was 398 g/kg and 194 g/kg for sample D and P respectively. The GC-MS results obtained showed that in both samples (D and P) linear and branched alkanes(n-Tetratetracontane, 3,6-Dimethyldecane, n-Pentadecane, 2-Bromodecane, n-Heptadecane etc. and n-Tetradecane, 2-Bromododecane, n-Octadecane, 14-Methyl-8-hexadecenal etc. respectively) were the main contaminants. Bioremediation was initiated and examined by applying separately fertilizer (F), bacteria inoculation (B), emulsifier (E) on the contaminated soil and by combining bacteria inoculation and fertilizer (B and F), fertilizer and emulsifier (F and E), bacteria inoculation and emulsifier (B and E) also a combination of bacteria inoculation, emulsifier and fertilizer (B,E and F) on both sample D and P . Bioremediation was determined by weight lost method throughout the 28days of treatment and the trend of
degradation is thus: (B, E and F) > B > ( B and E ) > (B and F) > (E and F) > E > F. Results obtained showed that treatment with B,E and F combined together yielded the highest percentage of oil degradation (97%, and 95% for samples P and D respectively), followed by B (96% and 81% for samples D and P respectively). This finding suggests that Bacillus sp is a viable microbial strain for bioremediation of oil-contaminated soil when biostimulated by adding fertilizer combined with emulsification (B, E and F). The percentage of oil degraded in sample P and D are almost the same (97 and 95% respectively) and there is significant difference (P < 0.05) in the trend of degradation for D and P this may be because, even though the samples contained oil (contaminant) with almost the same composition but differ in the number of carbon chain and other physical factors like density, viscosity etc.

TABLE OF CONTENTS

Title page – – – – – – – – – – i Declaration – – – – – – – – – – ii Certification – – – – – – – – – – iii Dedication – – – – – – – – – – iv Acknowledgement – – – – – – – – – v Abstract – – – – – – – – – – vi Table of contents – – – – – – – – – – viii List of Tables – – – – – – – – – – xiii List of Figures – – – – – – – – – – xiv List of Appendices – – – – – – – – – xvi Abbreviations – – – – – – – – – – xvii 1.0 CHAPTER ONE: Introduction – – – – – – 1 1.1 Background Study – – – – – – – – 1 1.2 Soil Contaminated with Oil – – – – – – – 3 1.3 Microbial Degradation of Petroleum Hydrocarbon – – – – 4 1.4 Justification for the Research. – – – – – – – 5 1.5 Aim and Objectives – – – – – – – – 6 2.0 CHAPTER TWO: Literature Review – – – – – 7 2.1 Oil Clean-up Strategies – – – – – – 7 2.1.1 Physico-chemical Methods – – – – – – – 8 2.1.2 Biological Methods – – – – – – – 10 2.2.0 Principle of Bioremediation – – – – – – 13
2.2.1 Nutrients – – – – – – – – 14 2.2.2 Optimizing Bioremediation Performance through Nutrients Addition – 16
2.2.3 Electron Acceptor / Oxygen – – – – – – – 17 2.2.4 Detergent – – – – – – – – – 20 2.2.5 Biosurfactants (Emulsifier) – – – – – – – 20 2.3.0 Environmental Requirements – – – – – – – 23 2.4.0 Factors Influencing Bioremediation of Hydrocarbons – – – 25 2.4.1 Temperature – – – – – – – – – 25 2.4.2 pH – – – – – – – – – – 26 2.4.3 Salinity – – – – – – – – – 26 2.4.4 Oxygen – – – – – – – – – 26 2.4.5 Nutrients – – – – – – – – – 26 2.4.6 Chemical Composition of Petroleum – – – – – – 27 2.4.7 Physical State of Hydrocarbons – – – – – – 27 2.4.8 Hydrocarbon Concentration – – – – – – – 28 2.5.0 Advantages of bioremediation – – – – – – 28 2.6.0 Strategies and Techniques used for Monitoring Biodegradation – – 29 2.6.1 Strategies – – – – – – – – – 29 2.6.2 In situ Bioremediation of Soil – – – – – – 29 2.6.3 Intrinsic Bioremediation – – – – – – – 29 2.6.4 Engineered Bioremediation – – – – – – – 30 2.6.5 Biostimulation – – – – – – – – – 30 2.6.6 Injection of Hydrogen Peroxide – – – – – – 31 2.6.7 Bioaugmentation – – – – – – – – 31 2.6.8 In Situ Bioremediation of Groundwater – – – – – 32 2.6.9 System Design – – – – – – – – 32
2.6.10 Application – – – – – – – – – 33 2.6.11 Ex situ Bioremediation – – – – – – – 33 2.6.11.1 Ex situ Remediation Techniques – – – – – – 33 2.6.11.2 Advantages of Ex situ Remediation – – – – – – 35 2.6.12 Land Farming – – – – – – – – – 36 2.6.13 Composting – – – – – – – – – 36 2.6.14 Biopiles – – – – – – – – – 36 2. 6.15 Bioreactors – – – – – – – – – 37 2.6.16 Phytoremediation – – – – – – – – 37 2.7.0 Petroleum Hydrocarbon Degrading Bacteria – – – – – 38 2.8.0 Anaerobic Degradation – – – – – – – 39 2.9.0 Techniques for Studying Oil Degradation – – – – – 42 2.9.1 Respirometry – – – – – – – – – 42 2.9.2 Gas Chromatography (GC) – – – – – – – 43 2.9.3 Luminescence Techniques – – – – – – – 43 2.9.4 Fluorescence Analysis – – – – – – – – 43 2.9.5 Determination of Total Petroleum Hydrocarbons (TPH) by Infrared Spectrometry (IR) and Gas Chromatography (TPH-IR and TPH-GC) – 44 3.0 CHAPTER THREE: Materials and Methods – – – – 45 3.1 Basic Materials used – – – – – – – – 45 3.2 Sample Location – – – – – – – – 46
3.3 Sample Collection. – – – – – – – – 46 3.4 Sample Preparation – – – – – – – – 48
3.5 Culture Medium Preparation – – – – – – – 48 3.6 Sample Culturing – – – – – – – – 48 3.7 Gram Staining – – – – – – – – – 48 3.8.0 Preparation of Reagents – – – – – – – 49 3.8.1 Preparation of Methyl Orange Indicator – – – – – 49 3.8.2 Preparation of Calcimeter Bernard Filling Solution – – – – 49 3.8.3 Preparation of Solution of 4M HCl – – – – – – 49 3.8.4 Preparation of 0.16M Potassium Dichromate (K2Cr2O7) – – – 50 3.8.5 Preparation of 1.0 M Ferrous Sulphate (FeSO4 • 7H2O) – – – 50 3.8.6 Preparation of Ortho-Phenanthroline Ferrous Complex Indicator- – – 50 3.8.7 Preparation of 0.01M CaCl2 .2H2O – – – – – – 50 3.9.0 Moisture Content Determination – – – – – – 50 3.10 pH Determination – – – – – – – – 51 3.11 Determination of Total Nitrogen in Soil – – – – – 51 3.12 Determination of Cation Exchange Capacity (CEC) – – – – 51 3.13 Chloride Determination – – – – – – – 52 3.14 Determination of Carbonate and Bicarbonate – – – – 53 3.15 Organic Carbon Determination – – – – – – 53 3.16 Determination of Exchangeable Bases (Mg, Ca, K and Na) – – – 54 3.17 Soil Particle Size Analysis – – – – – – – 54 3.18 Phosphorus determination- – – – – – – – 55 3.19 The Experimental Sets – – – – – – – 55 3.20. Extraction of oil for GC-MS Analysis – – – – – 56 3.21. Biodegradation of Crude Oil – – – – – – – 56
3.22 The Emulsifier (Liquid Soap) – – – – – – – 57 3.23 Statistical Tool – – – – – – – – 57 4.0 CHAPTER FOUR: Results – – – – – – – 59 4.1 Physicochemical Properties – – – – – – – 59 4.2 The Percentage of Oil (Contaminant) Degraded With Time – – – 59 4.3 Gram staining – – – – – – – – – 79 5.0 CHAPTER FIVE: Discussion of Results – – – – – 82 5.1 Physicochemical Properties – – – – – – – 82 5.2 Gram stain – – – – – – – – – 83 5.3 Degradation of oil (contaminant) by Natural processes – – – 84 5.4 Effect of inoculation with Bacillus sp (B) only (bioaugmentation) – – 84 5.5 Effect of Adding Fertilizer (F) Only (Biostimulation) – – – 85 5.6 Effect of Emulsification (E) of Sample on Degradation of its oil – – 86 5.7 Effects of combining inoculation and emulsification (B and E) on soil – 86 5.8 Effect of Emulsification and Adding Fertilizer (E and F) – – – 87 5.9 Effect of inoculation with bacteria and adding Fertilizer (B and F) – – 87 5.10 Effect of Combining Emulsifier, Fertilizer and Bacillus sp (B, E and F) – 88 5.11 Effect of Oil Concentration and Composition of The Sample – – 89 6.0 CHAPTER SIX: Summary, Conclusions and Recommendations – 92 6.1 Summary – – – – – – – – – 92 6.2 Conclusion – – – – – – – – – 92 6.3 Recommendations – – – – – – – – 93
REFERENCE – – – – – – – – – 94 APPENDICES – – – – – – – – –

CHAPTER ONE

Background Study
One of the major environmental problems in the world today is hydrocarbon contamination resulting from the activities related to the petrochemical industry. In Nigeria, oil pollution problems have been prevalent since the commencement of oil exploration and development of the petroleum industry (Okoh et al., 2001). Accidental release of petroleum products are of particular concern in the environment. Hydrocarbon components have been known to belong to the family of carcinogens and neurotoxic organic pollutants. Currently accepted disposal methods of incineration or burial insecure landfills can become prohibitively expensive when amounts of contaminants are large. The deleterious effect of pollutants on the environment has led to increased awareness and vigilance against contamination of the Niger Delta environment. In relatively recent times in Nigeria, there has been remarkable increase in population, urbanization and industrial activities, (Eze and Okpokwasili, 2010). The release of crude oil into the environment by oil spills is receiving worldwide attention (Millioli et al., 2009). Bioremediation which has been defined as biological response to environmental abuse has continued to receive research attention across the globe (Hammer, 1993). Bioremediation has been described as the use of living microorganisms to degrade environmental pollution. In order words, it is a technology for removing pollutants from the environment thus restoring the original natural environment (Sasikuma and Papmazath, 2003). The long term aim of bioremediation designs is to present cost effective designs which reduces the pollutant to a level referred to as low as reasonable and practicably possible (ALARP). In order to achieve this cost effectiveness, researchers all over the world have begun to pay research attention to the use of organic waste as the source of limiting nutrients for effective bioremediation (Ibiene et al., 2011).
Mechanical and chemical methods generally used to remove hydrocarbons from contaminated sites have limited effectiveness and can be expensive (Das and Chandran, 2011). Bioremediation is the promising technology for the treatment of these contaminated sites since it is cost-effective and will lead to complete mineralization. Bioremediation functions basically on biodegradation, which may refer to complete mineralization of organic contaminants into carbon dioxide, water, inorganic compounds, and cell protein or transformation of complex organic contaminants to other simpler organic compounds by biological agents like microorganisms (Das and Preethy, 2010). Many indigenous microorganisms in water and soil are capable of degrading hydrocarbon contaminants.
Petroleum-based products are the major source of energy for industry and daily life. Leaks and accidental spills occur regularly during the exploration, production, refining, transport, and storage of petroleum and petroleum products. The amount of natural crude oil seepage was estimated to be 600000 metric tons per year with a range of uncertainty of 200,000 metric tons per year (Kvenvolden and Cooper, 2003). Release of hydrocarbons into the environment whether accidentally or due to human activities is one of the main causes of water and soil pollution. Soil contamination with hydrocarbons causes extensive damage of body system since accumulation of pollutants in animals and plant tissue may cause death or mutations (Sheetal, 2012). These oil spills can even cause damage to the sea and shoreline organisms (Rodríguez-Martínez , 2006). The other sources of contamination include service stations, garages, scrap yard, waste treatment plants, saw mills, etc. Many microorganisms have the ability to utilize hydrocarbons as sole sources of carbon as energy for metabolic activities and these micro organisms are ubiquitous and widely distributed in nature (Jyothi et al., 2012). The microbial utilization of hydrocarbons depends on the chemical nature of the compounds within the petroleum mixture and on environmental determinants (Adeline et al., 2009). Hydrocarbons enter into the environment through waste disposal, accidental spills, as pesticides and via losses during transport, storage, and use. Hydrocarbon (petroleum) degrading bacteria’s ability to degrade and/or detoxify organic contaminants has been established as an efficient, economical, versatile and environmentally sound treatment (Atlas, 1981) The extensive use of petroleum products leads to the contamination of almost all compartments of the environment, and biodegradation of the hydrocarbons by natural populations of microorganisms has been reported to be the main process acting in the cleaning of hydrocarbon-polluted environment (Chaillan et al., 2004). Due to extensive increase in environmental pollution, numerous biodegradative bacteria have been isolated in the past, and their physiology, biochemistry, and genetics have been intensively studied. Biodegradation, which is the destruction of organic compounds by microorganisms, is carried out largely by diverse bacterial populations, mostly by Pseudomonas species (Boboye et al., 2010). The fuel is a complex mixture of normal, branched and cyclic alkanes, and aromatic compounds obtained from the middle-distillate fraction during petroleum separation. Bioremediation processes rely on the ability of microorganisms present naturally which are highly efficient due to their simplicity and cost-effectiveness when compared to other technologies (Jyoth et al., 2012). Hydrocarbon utilizing microorganisms are ubiquitously distributed in the marine environment following oil spills (Dua et al., 2002).
1.2. Soil contaminated with oil Soil contaminated with Petroleum is hazardous to human health, pollutes ground water and decreases the agricultural productivity of soils (Wang et al., 2008). The health concern stems primarily from direct contact with the contaminated soil, vapors from the contaminants, and from secondary contamination of water supplies within the soil. The technology commonly used for the soil remediation includes mechanical, burying, evaporation, dispersion and washing (Antai and Mgomo, 1989; Das and Mukherjee, 2011; Johnsen et al., 2005; Colwell et al., 1973; Leahy and Colwell, 1990). However, these technologies are expensive and can lead to incomplete decomposition of contaminants. The use of bioremediation process to detoxify or remove pollutants is an evolving method for the removal and degradation of many environmental pollutants including the products of petroleum industry. Bioremediation technology is believed to be non-invasive and relatively cost-effective (Millioli et al., 2009). Biodegradation by natural populations of microorganisms represents one of the primary mechanisms by which petroleum and other hydrocarbon pollutants can be removed from the environment (Rosenberg and Ron, 1996) and is cheaper than other remediation technologies (Vidali, 2001). The success of oil spill bioremediation depends on the ability to establish and maintain conditions that favor enhanced oil biodegradation rates in the contaminated environment. One important requirement for the degradation of oil is the presence of microorganisms with the appropriate metabolic capabilities. If these microorganisms are present, then optimal rates of growth and hydrocarbon biodegradation can be sustained by ensuring that adequate concentrations of nutrients and oxygen are present. 1.3 Microbial Degradation of Petroleum Hydrocarbon
Biodegradation of petroleum hydrocarbons is a complex process that depends on the nature and on the amount of the hydrocarbons present. Petroleum hydrocarbons can be divided into four classes: the saturates, the aromatics, the asphaltenes (phenols, fatty acids, ketones, esters, and porphyrins), and the resins (pyridines, quinolines, carbazoles, sulfoxides, and amides). Different factors influencing hydrocarbon degradation have been reported by Cooney and Summers, 1976; Chris, 2000). One of the important factors that limit biodegradation of oil pollutants in the environment is their limited availability to microorganisms (Providenti et al., 1995). Hydrocarbons differ in their susceptibility to microbial attack and can be generally ranked as follows: linear alkanes > branched alkanes > small aromatics > cyclic alkanes (Rosenberg and Ron, 1996). Some compounds, such as the high molecular weight polycyclic aromatic hydrocarbons (PAHs), may not be degraded at all. Microbial degradation is the major and ultimate natural mechanism by which one can clean up the petroleum hydrocarbon pollutants from the environment. Hydrocarbons biodegraded by bacteria, yeast, and fungi have been reported to have efficiency that ranged from 6% (Jones et al., 1970) to 82% (Pinholt et al., 1979) for soil fungi, 0.13% (Jones et al., 1970) to 50% (Pinholt et al., 1979) for soil bacteria, and 0.003% (Hollaway et al., 1980) to 100% (Mulkins and Stewart, 1974) for marine bacteria. Mixed populations with overall broad enzymatic capacities are required to degrade complex mixtures of hydrocarbons such as crude oil in soil( Bartha and Bossert, 1984), fresh water (Cooney, 1984), and marine environments (Atlas, 1985; Floodgate, 1984). 1.4 Justification for the Research.
In relatively recent times in Nigeria, there has been remarkable increase in population, urbanization and industrial activities (Eze and Okpowasil, 2010). The Kaduna Refinery produces 1.5milion litres of fuel daily and has recently produced 14.989 million litres of petrol, evacuated by 454 trucks , 6 million litres of kerosene have also been produced and evacuated by 184 trucks, another 12 million litres of diesel have been produced and evacuated by 411 trucks, while 13 million litres of low pour fuel oil have been produced within one month. This implies that large volume of oil contaminants (particularly around the oil related companies) are released into the environment. Pollution caused by petroleum and its derivatives is the most prevalent in the refinery environment. The low solubility and adsorption of high molecular weight hydrocarbons limit their availability to microorganisms. Oils are hydrophobic in nature, their availability to bacteria are limited which leads to the
slow degradation because petroleum hydrocarbon compounds bind to soil components, and they are difficult to be removed or degraded. When a major oil spill occurs the supply of carbon is dramatically increased and the availability of nitrogen and phosphorus generally becomes the limiting factor for oil degradation (Atlas, 1984; Leahy and Colwell, 1990), hence there is need to enhance bioremediation. Bioremediation offers several advantages over conventional techniques such as land filling or incineration. Bioremediation can be done on site, is often less expensive and site disruption is minimal, it eliminates waste permanently, eliminates long-term liability, and has greater public acceptance, with regulatory encouragement, and it can be coupled with other physical or chemical treatment methods. The release of crude oil into the environment by oil spill is receiving worldwide attention (Millioli, et al., 2009). 1.5 Aim and objectives The overall aim of the research is to bioremediate oil contaminated soil using NPK fertilizer, Bacillus sp and emulsification by liquid soap. The specific objectives are to determine the effect of: 1. physicochemical parameters on oil degradation 2. NPK fertilizer as nutrient source in bioremediation 3. microbial seeding on oil degradation. 4. emulsifier (liquid soap) on oil degradation. 5. combination of i-iii above on oil degradation.

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