CHAPTER 1
Introduction
1.1 Background of Research
Energy is the main component for human being to sustain daily activities and improving quality of life. The definition of energy is the ability to do work (Demirbas, 2008). The presence of energy could not be seen but only the effects of it are experienced. There are a few major form of heat which includes heat, light, motion and sound. Energy are then divided into two categories of kinetic and potential energy. From the vast amount of energy available, energy usage had definitely benefits the entire civilization since the beginning of life. In conjunction with the advancement of technology and human civilization, the amount of energy reserve must be sufficient to meet the demand in future.
The energy sources are derived from three main groups which are fossil, renewable and fissile energy. Fossil fuels are non-renewable energy as it takes million years for fossil to develop from decomposition of dead living material buried in the earth. Currently, fossil fuels accounts for more than 80% for the most consumed energy worldwide and predicted to keep rising with the increase of population number thus higher demand (IEA, 2013) with 36%, 27%, 23% and 14% share of energy consumption are from oil, gas, coal, renewable and alternative energy, respectively (Abas et al., 2015). The high dependency on fossil fuels had produce an uncertainty for the future supply of fossil fuels due to its nature as non-renewable energy. Many researchers discussed this concerned issue and had come out with a prediction that the fossil fuels reserve are only available until the year 2042 (Shafiee and Topal, 2009).
The usage of fossil fuels had also produce another costs on the other side, the emission of greenhouse gases (GHG) which causes climate change, ecological damage and concern on human’s health (Petinrin and Shaaban, 2015). A serious amounts of air pollutant including sulphur dioxide (SO2), nitrogen oxides (NOx), carbon monoxide (CO) carbon dioxide (CO2) are being released since the beginning of mining fossil fuels until it is fully burned at the end of consumer chain (Eipstein et al., 2011). While there is no possibility to completely terminate the toxic gases from being released , initiative to reduce the amount of pollutants are by transitioning into a more environmental and public health friendly, which is implementing renewable energy. According to Abas et al. (2015) the transitioning are most ideal at the renewable energy utilization rate of 20% by 2020, 50% by 2050 and 100% by 2100.
Renewable energy are derived from easily regenerate and available sources which includes biomass, hydropower, wind, geothermal and solar. From the variety of sources available, biofuel was suggested to be one of the most high potential to reinstate the diminishing fossil fuels (Liew et al., 2014). According to Demirbas (2009), biofuel is defined as solid, liquid and gaseous fuels that are derived from renewable sources. Biofuel productions are achieved through oil synthesis from the biomass feedstocks. Biomass, as one of the renewable energy sources includes all living matters in this planet which contains organic matter that could be converted into energy sources. Biofuel is non-toxic and carbon neutral which would produce less carbon emission compared to fossil fuels (Cucek et al., 2012). There are various type of biofuel available which comprises of biodiesel, bioalcohol, biocrude/synthetic oils, biochar, biogas and bio-hydrogen (Liew et al., 2014). Malaysia had also launched National Biofuel Policy in March 2006 in strategy to encourage more energy usage from renewable energy (NBP, 2006).
Biodiesel is an alternative to conventional petroleum-derived diesel fuel (petrodiesel) and referred as the mono-alkyl esters of vegetable oils or animal fats (Knothe et al., 2010). The term biodiesel is describing vegetable oils that has been transesterified and can be used as diesel fuel. The advantages of biodiesel are listed as follows; derived from renewable energy, miscibility with petrodiesel at all blend levels, positive energy balance, reduce most of frequently emitted toxic gases except for nitrogen oxides (NOx), biodegradable, less sulfur and aromatic content, high flash point and inherent lubricity (Knothe, 2013). On top of that, biodiesel has greater properties in terms of sulphur content, flash point, aromatic content, and biodegradability compared to conventional diesel (Martini and Schell, 1997). The trend of biodiesel demands thus increase in production had also significantly increase within the last decade showing a good sign of biodiesel acceptance across the globe (IEA, 2011).
The third-generation biofuel, microalgae is a promising candidate for biodiesel production because of their high photosynthetic efficiency compared to the conventional crops which are classified as first and second generation (Ahmad et al., 2011). The first-generation biofuel is derived from edible crops which are corn, sugarcane, soy or wheat while the second generation is the inedible plants or their parts which are grass, stems or leaves. The advantages of microalgae as feedstock especially for biodiesel production are they can accumulate large quantities of triacylglycerols (TAGs), high growth rates, fix CO2 from atmosphere, able to adapt to wide condition including extreme environment and utilize nutrients from wastewater (Hu et al., 2008). Moreover, the alternative of choosing microalgae as the main biomass source could be further contributed by the concerns of food availability if the first and second-generation biofuel being implemented at a large-scale production.
Microalgae is able to accumulate significant amount of TAGs which are studied to be the best substrate for biodiesel. This is due to the production of biodiesel in transesterification process where the TAGs react with methanol to produce fatty acid methyl esters (FAME) and glycerol (Chisti, 2007). The marine species of microalgae Nannochloropsis sp. has been identified and discovered it’s potential to produce a promising quantity of oils to being further developed into biodiesel. It is due to its ease of growth and high oil content (28.7%) of dry weight, mainly unsaturated fatty acids and a significant percentage of palmitic acid (Gouveia and Oliveira, 2009). The species Nannochloropsis oculata (N.oculata) are also discovered to have a high percentage of monounsaturated fatty acids which in turns would act as a balance between oxidative and low-temperature properties (Ma et al., 2014).
At the present time, mass cultivation of microalgae is being carried out in open ponds or tanks with artificial light. In conjunction with this practice, there are several problems associated with it for instances, susceptibility to contamination, and low cell concentration and productivity (Imamoglu et al., 2015). There has been plenty of studies have shown that the quality and quantity of microalgae biomass production and lipid content are affected by the growth conditions (temperature, light intensities, pH, duration of cultivation, CO2 concentration, salinity) and nutrient composition (nitrogen, phosphate, carbon, iron concentration) (Malakootian et al., 2015). In order to maximize its full potential into biodiesel production, the cultivation of Nannochloropsis sp. needed to be optimized with precise parameters which is suitable for the maximal growth of microalgae in a closed system which offer better process control and higher biomass and lipid productivity, instead of an open system.
To achieve high biomass and lipid production at the same time are often a challenging task in microalgae cultivation as they are concomitant with each other where only one the elements would produce a significant amount to satisfy the needs of biodiesel oil production. Therefore, there is in need to imply a favorable cultivation method to produce high biomass and lipid production concurrently. Thus, this proposed research is to study the relationship CO2 concentration and how it would affect the biomass and lipid production via the batch cultivation while experiencing nitrogen and salt stress. Besides, an optimized lipid extraction with the highest lipid yield from N.oculata are determined from three different lipid extraction methods which are solvent extraction method, sonicator-assisted or microwave-assisted.
1.2 Problem Statements
The natural sources of fossil fuels is depleting promptly because of high demands and its nature as a finite element (Kumar et al., 2015). The vast reduction of crude oil resource require a need of alternatives from other renewable energy which are more sustainable and does not cause detrimental effect to the environment. As a result, biodiesel is emerging as the renewable energy where it could reduce the dependence on fossil fuels and can be readily introduced into the existing transportation without major modification (Mathimani et al., 2015). Microalgae as the candidate for biodiesel production has come to limelight as it possess wide advantage compared to other oleaginous crops (Mata et al., 2010). It has been classified as the third generation biofuel after the usage of first and second generation biofuel, which are crops and non-edible plant parts, respectively, did not come out with a satisfied outcome. However, the commercialization of microalgae as biodiesel is still on extensive research in finding the most optimized and suitable methods for its biodiesel production.
One of the main challenge in the mass production cultivation of microalgae is to obtain a substantial amount of biomass and lipid productivity to meet the biodiesel production demand. Both high biomass and lipid production are the indicators for a desirable qualities that are being search for in a microalgae strain. The biomass production could be greatly influenced by the nutrients and other biotic factors supplied during cultivation while to obtain high lipid accumulation. However, contrast steps are needed such as stress applied on temperature, salinity, light intensity or nitrogen or phosphorus starvation (Pal et al., 2011). When stress are applied, the lipids are accumulated as a results of the cellular carbon flux from protein synthesis to lipid synthesis as its mechanism to adapt to a deprived culture conditions (Sheehan et al., 1998). There are two stress conditions that are proposed in this research which are nitrogen and salt stress.
On the other hand, the application of stress during microalgae cultivation leads to decrease in biomass productivity as the microalgae could not replicates its cells resulted from deprivation thus halted the cell growth (Guschina and Harwood, 2006). As both of biomass and lipid productivity are needed in producing a cost efficient and effective biodiesel, the proposed study is to determine whether there is a direct effect and relationship when the stress are combine together in batch microalgae cultivation. To overcome challenges in improving economical biodiesel production, a novel strategy of combination of stress factors, co-culture with other microorganisms, addition of phytohormones and chemical additives (Singh et al., 2016). Fields et al., (2014) suggested that combinations of potential stressors would impact lipid accumulation in other ways rather than just implementing one sole stress factor.
The relationship of growth and lipid production and the magnitude of nitrogen deficiency needed to stimulate lipid accumulation are still in need of depth research and understanding (Adams et al., 2013). From the statement, the stress approach alone are not enough to find a better cultivation method of microalgae to be implemented commercially. The CO2 concentration have been studied as one of the prevalent factors that are contributing in the effectiveness in microalgae productivity (Shene et al.,2016), but there are still no research to study the effect of CO2 enriched medium to enhance the rate of photosynthesis and reduce photorespiration while applying combination of growth stress condition on microalgae N.oculata.
After a successful upper stream production, the lipid extraction by using an optimized cell disruption technique is crucial as it would determine the lipid recovery of microalgae. The lipid recovery factor is a good criteria of a strain for biodiesel production especially the FAME production (Mathimani et al., 2015). The lipid extraction process is also one of the major economic challenge towards the commercial biodiesel production from microalgae (Li et al., 2014) thus the highest lipid yield extraction method for its specific strain need to be identified. Various type of lipid extraction from microalgae had been attempted such as physical method by using screw press or chemical method by using organic solvents, water (by three-phase partitioning extraction methods), supercritical methods or solvent extraction. Amongst the methods available, solvent extraction had been proven to be the most effective compared to the others (Grima et al., 2013).
However, the presence of cell wall and plasma membrane surrounding the algae cell are one of the obstacle while extracting lipid using solvent. The cell wall acts as an outer layer barrier protecting microalgae from any mechanical stress while plasma membrane is the boundary between the cell wall and cytoplasm of an algal cell. Therefore, studies of biomass pre-treatment are necessary to maximized the amount of lipid that could be extracted. The breakage of cell wall and plasma membrane prior to lipid extraction to enable more solvent penetration into the cell and release more cell contents thus increasing lipid yield. Cell-disruption techniques includes, sonication, autoclaving microwaves, homogenization via tissue grinder, osmotic shock, blender or in high-pressure flow device and freezing and grinding using pestle and mortar (Grima et al., 2013).
1.3 Research Objectives
The overall goal of this research work is to produce an efficient cultivation strategy in order to produce a good quality of biodiesel at the end of experiments. The strategies includes stress application during microalgae cultivation, CO2 application during stress cultivation and determining the most effective lipid extraction method.
The main objectives of this research are:
” To investigate the effect of various type of stress which are nitrogen and salt stress on biomass and lipid production from Nannochloropsis oculata via batch cultivation strategy.
” To determine the relationship between CO2 concentration and its effect on biomass and lipid production from Nannochloropsis oculata when combination of nitrogen and salt stress are applied.
” To determine the optimum lipid extraction method between Bligh-Dyer, sonication-assisted or microwave-assisted for highest lipid yield from Nannochloropsis oculata.
1.4 Scope of Research
This research is comprising a study of microalgae cultivation to enhance its biomass and lipid productivity for a maximized biodiesel production in batch cultivation growth strategy by applying stress condition to the culture which are nitrogen and salt stress. The optimized nitrogen (NaNO3) and salt (NaCl) concentration are determined via separate batch cultivation. The biomass and lipid production are being analysed and the lipid properties which are the FAMEs are being studied as well to determine the most optimized concentration of nitrogen and salt. Later, the selected concentration are being combined and added various CO2 concentration to enhance the productivity of microalgae while experiencing stress condition. The factor of on how CO2 concentration on how it could affect the biomass and lipid production of N.oculata when cultivated together with the applied stress condition are also being studied. This study will also focus on various lipid extraction method which are solvent extraction, sonicator-assisted or microwave-assisted to obtain the most optimized and high yield lipid extraction from N.oculata. The extracted lipids from microalgae are later converted into biodiesel through transesterification process. After undergoes transesterification, the FAMEs are analysed using a GC-FID to determine the biodiesel properties and its suitability as a commercial biodiesel.
1.5 Thesis Outline
This dissertation consists of five chapters and the particulars are as follows:
i. Chapter 1 is the introduction and brief overview about the study which comprises background of research, problem statements, research objectives, scope of research, and the outline of the thesis.
ii. Chapter 2 is the literature review of the study. It consists of in depth understanding of the nature of study, critical evaluation of the related field, significant previous findings by other researcher and active discussions on the specific scope of study. Additionally, it also provides a fundamental base as an overall idea of this research.
iii. Chapter 3 is the methodology section where the steps of actions taken in this research are being discussed in detail. The method of investigation includes experimental procedure and sample analysis together with the instrumentations and apparatus that has being used.
iv. Chapter 4 is the results and discussions part. This section includes the representation of data which comprises of figures, graphs and tables followed with the interpretation of the data and comprehensive discussion on the data analysis. This section emphasizes on the significant findings which has met the research objectives.
v. Chapter 5 is the final component of this dissertation. It generally concludes the whole summary of study and its profound effect. There are also future recommendations that could be done to improve on the related field.
This completed thesis is beneficial for a reference for the implementation in-depth cultivation strategy of commercialization Nannochloropsis oculata microalgae to be develop into biodiesel.