ECOTOXICITY OF TITANIUM DIOXIDE NANOPARTICLE TO CHLORELLA VULGARIS BEYERINCK (TREBOUXIOPHYCEAE, CHLOROPHYTA) UNDER LIMITED NITROGEN CONDITIONS IN VITRO


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TABLE OF CONTENTS

CONTENTS
ABSTRACT
TABLE OF CONTENTS
APPENDICES
LIST OF ABBREVIATIONS

CHAPTER ONE
1.0 INTRODUCTION
1.1 Background to the Study
1.2 Statement of the Research Problem
1.3 Justification
1.4 Aim of the Study
1.5 Objectives
1.6 Hypotheses

CHAPTER TWO
2.0 LITERATURE REVIEW
2.1 Taxonomy and Morphology of Chlorella vulgaris
2.2 Environmental Factors Affecting Algal Growth
2.2.1 Light
2.2.2 Temperature
2.2.3 pH
2.2.4 Test medium
2.2.5 Nutrient
2.2.5.1 Nitrogen
2.3 Ecotoxicology: Chemicals and the Environment
2.4 Nanoparticles in the Environment
2.4.1 Natural nanopartices
2.4.2 Anthropogenic nanopartices
2.5 Ecotoxicology of Metal oxide Nanoparticles
2.5.1 Titanium dioxide nanoparticles
2.6 Enzyme Biomarkers
2.6.1 Antioxidants and oxidative stress in algae

CHAPTER THREE
3.0 MATERIALS AND METHODS
3.1 Algal Culture Species
3.2 Culture Media
3.2.1 Preparation of culture media
3.3 Titanium dioxide Nanoparticles Treatment
3.3.1 Preparation of nanoparticles dispersion and treatment
3.4 Nutrient Source
3.5 Growth and Biomass Determination
3.5.1 Dry weight measurement
3.5.2 Cell counts (cells mL-1)
3.6 Chlorophyll Determination
3.7 Biochemical Composition Determination
3.7.1 Total carbohydrates
3.7.2 Protein extraction
3.7.3 Total proteins
3.7.3.1 Bradford reagent preparation
3.7.3.2 Protein Assay
3.7.4 Antioxidants enzyme determination
3.7.4.1 Assay of glutathione-s-transferase
3.7.4.2 Assay of peroxidase
3.7.4.3 Assay of lipid peroxidation
3.8 Data Analyses

CHAPTER FOUR
4.0 RESULTS
4.1 Growth and Biomass Production of Chlorella vulgaris
4.1.1 Dry weight
4.1.2 Cell counts
4.1.3 Chlorophyll a
4.1.4Total chlorophyll
4.1.5 Specific growth rate
4.2 Biochemical Composition of Chlorella vulgaris
4.2.1 Carbohydrate content
4.2.2 Protein content
4.3. Antioxidant Enzyme Activities
4.3.1 Glutathione-s-transferase
4.3.2 Lipid peroxidation
4.3.3 Peroxidase activity
4.4       Correlations of Growth, Biomass Production, Biochemical Composition, and Antioxidant Enzymes Activity with Nitrogen/Titanium dioxide Nanoparticles Treatments

CHAPTER FIVE
5.0 DISCUSSION

CHAPTER SIX
6.0 SUMMARY, CONCLUSIONS AND RECOMMENDATIONS
6.1 Summary
6.2 Conclusions
6.3 Recommendations
REFERENCES
APPENDICES



ABSTRACT


The broad application of titanium dioxide nanoparticles in many consumer products has resulted in the release of substantial amounts into aquatic system, which serve as the terminal sink for nanomaterials. These titanium dioxide nanoparticles may induce some unexpected toxic effects to aquatic organisms such as microalgae. This study was carried out to evaluate the toxicity of limited nitrogen and titanium dioxide nanoparticles to the microalgae Chlorella vulgaris. The nanoparticles were prepared in Organisation for Economic Cooperation and Development (OECD) algal test medium and also the nitrogen concentration in the algal test medium was adjusted to match limited environmental nitrogen level. The growth, biomass production, biochemical composition (Carbohydrate and protein content), and antioxidant response (Glutathione-s-transferase, peroxidase, and lipid peroxidation) of the algae were monitored over a 96h period. The results showed that limited nitrogen (2.8 × 10 -6 M) decrease growth, biomass (dry weight, cell counts, chlorophyll content), carbohydrate content, and increase protein content, antioxidant enzyme activity and lipid peroxidation (malondialdehyde content) in the alga. Titanium dioxide nanoparicle treatments (0.2mg/L, 8mg/L, 16mg/L and 32mg/L) decrease growth, biomass, carbohydrate content and increase glutathione-s-transferase activity. The combination of limited nitrogen with titanium dioxide nanoparticle decrease growth, dry weight, chlorophyll content, and carbohydrate content of the alga. This result suggests that limited nitrogen and titanium dioxide nanoparticle treatments affects growth, biomass production, biochemical composition, induces oxidative stress and also induces the oxidation of lipids while titanium dioxide nanoparticle combined with limited nitrogen affect growth, biomass, and carbohydrate content of Chlorella vulgaris under white fluorescent light.




CHAPTER ONE


1.0                                                                       INTRODUCTION


1.1              Background to the Study


Microalgae are a very important component of the aquatic ecosystem; they are a group of fast growing unicellular or simple multicellular microorganisms that have the ability to fix CO2 while capturing solar energy with efficiency 10 to 50 times greater than that of terrestrial plants (Wang et al., 2008). They also have higher biomass production compared to energy crops (Wang et al., 2008). They inhabit the pelagic as well as benthic environments of the hydrosphere in a variety of forms. This variation is more pronounced in green algae, which taxonomically belongs to different phyla, such as Prochlorophyta, Volvocophyta, Euglenophyta, Chlorophyta, and Charophyta (Graham and Wilcox, 2000).

Chlorella sp belongs to the Division Chlorophyta, class Chlorophyceae, order Chlorococcales, and family Chlorococcaceae.

Titanium dioxide, also known as titanium (IV) oxide or Titania is the naturally occurring oxide of titanium, the ninth most abundant element in the world. It is five times less abundant than iron but 100 times more abundant than copper (IARC, 2010). Generally titanium dioxide is sourced from Ilmenite ore, rutile and anatase, which are mined from deposits located throughout the world. Ilmenite ore is the widest spread of titanium dioxide bearing ore in the world. Rutile (TiO2) and Ilmenite (FeTiO3) are commonly found as accessory minerals in plutonic and metamorphic rocks but occur also as detrital.....


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