TABLE OF CONTENTS

CHAPTER ONE

1.0              Introduction

1.1       Justification of the study       

1.2        Objective of the study

CHAPTER TWO

2.0       Literature Review

2.1.      Bio-degradation

2.2.      Bio-remediation and bio-degradation

2.2.1.   Natural attenuation

2.3       Bio-stimulation

2.4.      Bio-augmentation

2.5.      Crude Oil

2.5.1.   Composition Of Petroleum

2.5.1.1 Paraffins:

2.5.1.2.Olefins:

2.5.1.3.Naphthenes (Cycloparaffins)

2.5.1.4.Aromatics

2.5.2.   History Of Crude Oil In Nigeria

2.5.3.   Impact of Crude Oil On the Nigerian Economy

2.6.      Role of microorganisms in bio-degradation of pollutants.

2.7.      Some biodegradable pollutants.

2.7.1.   Hydrocarbons:

2.7.2.   Polycyclic aromatic hydrocarbons (PAHs):

2.7.3.   Polychlorinated biphenyls (PCBs):

2.7.4.   Pesticides:

2.7.5.   Dyes:

2.7.6.   Radionuclides:

2.7.7    Heavy metals:

2.8       Microfungi and mycorrhiza degradation.

2.8.1.   Yeasts degradation

2.8.2.   Filamentous fungi degradation

2.9.      Gas chromatography

2.9.1    The chromatographic process

2.9.1.1 Carrier gas

2.9.1.2.Injector

2.9.1.3 Column

2.9.1.4.Detector

2.9.2    Sample preparation techniques

2.9.3    Advantages and disadvantages of gas chromatography

2.9.3.1.Advantages of Gas Chromatography

2.9.3.2 Disadvantages of Gas Chromatography

CHAPTER THREE

3.0 Materials and Methods

3.1 Materials/Equipments used

3.2 Collection of samples

3.3 Sterilization of material

3.4 Culture media preparation

3.5 Serial dilution and enumeration of total fungi in soil sample

3.6. Sub-culturing of fungi isolates

3.7. Biochemical test for yeast

3.7.1 Gram staining

3.7.2 Sugar fermentation test

3.8 Identification of fungi

3.8.1 Cultural characteristic of fungi

3.8.2 Microscopic examination of fungi

3.9 Preservation of culture

3.10 Preparation of fungal culture

3.11 Sterilization of crude oil

3.12 Test for degradation of  crude oil

3.13.Extraction Of degraded Crude Oil

3.14 Methodology for the gas chromatography analysis

CHAPTER FOUR

4.0 Result

4.1 Microbial population of soil

4.2 Morphological and microscopic characteristic and dominant fungal isolated from soil

4.3 Rate of crude oil degradation by different fungi isolate

4.4 Gas chromatography analysis  result for the extracted crude oil samples.

CHAPTER FIVE

5.0       Discussion

CHAPTER SIX

6.0       CONCLUSION AND RECOMMENDATION

6.1       Conclusion

6.2.      Recommendation

REFERENCES

CHAPTER ONE

1.1     INTRODUCTION

Crude oils are composed of mixtures of paraffin, alicylic and aromatic hydrocarbons. Microbial communities exposed to hydrocarbons become adapted, exhibiting selective enrichment and genetic changes resulting in increased proportions of hydrocarbon-degrading bacteria and bacterial plasmids encoding hydrocarbon catabolic genes (Leahy and Colwell, 2004). Adapted microbial communities have higher proportions of hydrocarbon degraders that can respond to the presence of hydrocarbon pollutants. The measurement of biodegradation rates under favorable laboratory conditions using 14C-labelled hexadecane has led to the estimation that as much as 0.5 – 60 g oil/m3 seawater convert to carbon dioxide, depending on temperature and mineral nutrient conditions. The principal forces limiting the biodegradation of polluting petroleum in the sea are the resistant and toxic components of oil itself, low water temperatures, scarcity of mineral nutrients (especially nitrogen and phosphorous), the exhaustion of dissolved oxygen and in previously unpolluted areas, the scarcity of hydrocarbon-degrading microorganisms (Atlas, 2002). Low winter temperature can limit rates of hydrocarbon biodegradation increasing resident time of oil pollutant(Bodennec et al., 2007). Bio-degradation is nature's way of recycling wastes, or breaking down organic matter into nutrients that can be used and reused by other organisms.

In the microbiological sense, "bio-degradation" means that the decaying of all organic materials is carried out by a huge assortment of life forms comprising mainly bacteria and fungi, and other organisms. This pivotal, natural, biologically mediated process is the one that transforms hazardous toxic chemicals into non-toxic or less toxic substances. In a very broad sense, in nature, there is no waste because almost everything gets recycled. In addition, the secondary metabolites, intermediary molecules or any ‘waste products’ from one organism become the food/nutrient source(s) for others, providing nourishment and energy while they are further working-on/breaking down the so called waste organic matter. Some organic materials will break down much faster than others, but all will eventually decay. By harnessing microbial communities, the natural “forces” of biodegradation, reduction of wastes and clean up of some types of environmental contaminants can be achieved. There are several reasons for which this process is better than chemical or physical processes. For example, this process directly degrades contaminants rather than merely transforming them from one form to the other, employ metabolic degradation pathways that can terminate with benign terminal products like CO2 and water, derive energy directly form the contaminants themselves, and can be used in situ to minimize the disturbances usually associated with chemical treatment at the clean-up sites. Biological degradation of organic compounds may be considered an economical tool for remediating hazardous waste-contaminated environments. While some environments may be too severely contaminated for initial in situ treatment to be effective, most contaminated media will use some form of biological degradation in the final treatment phase.

Diverse groups of fungi have been isolated from oil contaminated environments and/or  have been shown to degrade hydrocarbons in the laboratory. Microbial degradation is the major mechanism for the elimination of spilled oil from the environment ( Atlas, 2000.). In this study, crude oil-contaminated soil samples areas in delta state were examined with the aim of isolating fungi with high crude oil degrading potentials.

1.2       Justification of the study     

Various  studies have identified some micro-organisms to be able to degrade crude oil. The degrading ability of these micro organisms have been determined using different methods such as gas chromatography and mass spectrometry (GC/MS), gas chromatography (GC), turbidometry, titrimetry e.t.c. Bio-degredation of crude oil is majorly carried out by bacteria and fungi. This study scientifically justifies the use of fungi to degrade crude oil. This project was therefore carried out to determine the degrading ability of fungi isolated from crude oil-contaminated soil samples using gas chromatography.

1.3        Objective of the study

The specific objectives of the study are to :

                               I.            Isolate and identify fungi from crude oil-contaminated soil sample.

                            II.            Screen the isolates for bio-degradative abilities.

                         III.            Assessment of the degrading abilities of the  fungi isolates by gas chromatography

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