The highest biomass titer and productivity of 1.42 g L-1 and 101.43 mg L-1 d-1, respectively, were recorded with an estimated CO2 fixation rate of 159.91 mg L-1 d-1 when grown at 12.5% CO2. The intracellular total protein content of 49.53 (w/w) was found to be highest in 12.5% CO2, suggesting the possible use of CT01 biomass as potential alternative aqua feed. Due to its high protein content, CT01 biomass was The total protein in the biomass and the carbon fixation rate of CT01 were also found to be higher with the light intensity of 150 µE m-2s-1. The interdependent dynamics of light intensity, growth and culture pH indicated that the process engineering strategy, based on on-demand supply of CO2 under optimized light wavelength and intensity, can result in improved biomass concentrations and productivity by maintaining optimal culture pH and eliminating CO2.
C ONTENTS
INTRODUCTION
REVIEW OF LITERATURE
- Status of feed production for aquaculture 20 2.4 Microalgae: Biology and classification 28
- Microalgal cultivation systems: growth parameters, nutrition
- Microalgal nutritional quality as fishmeal and fish oil
SAMPLING, ISOLATION AND SELECTION OF
POTENTIAL CO 2 TOLERANT MICROALGAL STRAINS
Sampling, screening and isolation of potential CO 2 tolerant
Identification of the isolated strain 82
Growth of the organism and CO2 sequestration under
Screening, isolation and identification of CO2 tolerant
4 . EVALUATION OF NOVEL MICROALGAE SPECIES PERFORMANCE AS A FEED SUPPLEMENT FOR
Assessment of feedstocks for fish growth metrices and
Biochemical composition of microalgal biomass, fish
Assessment of feedstocks for fish growth metrices, dietetics
PROCESS ENGINEERING STRATEGY FOR
CULTIVATION OF HIGH-DENSITY PROTEIN RICH CT01 MICROALGAL BIOMASS
Effect of light wavelength and intensity on growth kinetics
Statistical optimization of media for CT01 for
6.DEMONSTRATION OF AN INTEGRATED SUSTAINABLE MICROALGAL FEED TECHNOLOGY AT LARGE SCALE OF
AQUAFEED
Materials and methods 156
- Cultivation of CT01 in novel customised 50 L arlift
- Assessment of different types of feedstocks for fish growth
- Analysis of growth and substrate utilization 160 6.2.5 Biochemical composition of microalgal biomass and fish
- Development of customised energy efficient airlift
- Evaluation of fresh microalgal cell as alternative aquafeed 169
L ist of Figures
Nutrient utilization rate profile of Desmodesmus pannonicus CT01. biomass under different exposures of light wavelength 139. Nutrient utilization rate profile of Desmodesmus pannonicus CT01. biomass under different light intensity exposures.
L ist of Tables
ANOVA for quadratic regression model obtained from CCD- RSM used in media component optimization for. ANOVA for quadratic regression model obtained from CCD-RSM used in optimization of growth media components.
C HAPTER 1
Introduction
- Background and motivation
- Objectives of the study
- Approach
- Organization of the thesis
- References
1994) 'Preliminary study on the use of some oil cakes as dietary sources of protein for fingerlings of Indian giant carp (Labeo rohita)', Aquaculture p. 1992) 'Development of practical dry diets for coho salmon, Oncorhynchus kisutch, using poultry meal, feather meal, soybean meal and canola meal as major protein sources', Proc. on the diet of finned fish and fish feed technology, p. 2008) 'Triacylglycerols from microalgae as feedstocks for biofuel production:. perspectives and progress, The Plant Journal. 2018) "The promising future of microalgae: current status, challenges and optimization of a sustainable and renewable industry for biofuels, feed and other products", Microbial Cell Factories.
Review of Literature
Global CO 2 emission status
Therefore, to reduce the effects of global warming and keep the levels of carbon dioxide in the atmosphere stable, some of the technologies associated with renewable and sustainable energy sources have been taken into account (Baena-Moreno et al., 2018; . Singh and Dhar, 2019). The COVID-19 epidemic was the primary contributor to the reduction in global CO2 emissions that occurred in 2020.
Carbon sequestration technologies
EDGAR expected the year 2020 to show a decline, with global anthropogenic fossil CO2 emissions to be 5.1% lower than in 2019. This would put them slightly below the level of 36.2 Gt CO2 emissions recorded in 2013 was documented.
Status of feed for Aquaculture
The growth of the fish farming industry is expected to continue in the future. Research is being conducted to determine whether genetically modified crops with improved properties, such as canola and camelina that have a high content of omega-3 fatty acids, can serve as a viable alternative ingredient in aquafeed (Jones et al ., 2020). The alternative food components described above each have the advantages of nutritional content, presence of bioactive compounds and sustainable production.
Microalgae: biology and classification
Heterocysts are the source of nitrogen for all growing photosynthetic cells in the system. After maturation, the daughter cells are released from the mother cell by rupture of the cell wall, and the daughter cells then eat the remaining detritus left behind the mother cell (Safi et al., 2014).
Biochemistry of microalgae
Within the vegetative mother cell of Chlamydomonas sp., the multiplication process involves the creation of flagellate motile spores known as zoospores. This cooperation between PSI and PSII leads to the production of ATP and NADPH as the end products of light-dependent activities.
- Effect of temperature
- Effect of light
- Effect of nutrients
- Effect of pH
Yang et al., 2004) reported an increase in the length of the lag phase in cultures of Botryococcus braunii. Sulfur is more often digested in the form of sulfate (SO4-2), but sulfite, which is its more toxic form, is not (Oren, Padan, and Malkin, 1979).
Microalgal cultivation system
In conclusion, a series of studies determined that high pH stress suppresses the cell cycle and induces the accumulation of lipids (Vuppaladadiyam et al., 2018). Alternatively, the light can be provided in the form of artificial light, such as light-emitting diodes (LEDs), which are an energy source developed due to their low energy consumption and their light range (e.g. red LED, 624-634 nm; green LED, autotrophic cultivation considered technically and commercially scalable, typically in outdoor environments, especially to increase lipid productivity (using 2% CO2 in the air) and also to recycle industrial CO2 (Moreno-Garcia et al., 2017; Saha and Murray, 2018) .
Reactor systems used for algal culturing
Accurate calculations of the amount of incoming light and water temperature are essential to growing success. Both of these factors help to ensure that problems related to light availability in the center of the PBR are avoided.
Microalgae in aquaculture industry
The amount of polyunsaturated fatty acids (eicosapentaenoic acid, EPA); arachidonic acid, ARA; . and docosahexaenoic acid, DHA) found in microalgae is the second most important factor (Reitan et al., 1997). EPA and DHA are produced by a wide variety of autotrophic and heterotrophic microalgae species belonging to a variety of classes, but AA is often found only in trace levels (Bigogno et al., 2002).
Current challenges
2016) 'A review of biological processes and their potential for CO2 capture', Journal of Environmental Management. 2011) 'Evaluation of microalgae as sources of digestible nutrients for monogastric animals', Journal of Livestock and Forage Sciences.
C HAPTER 3
Background and motivation
In the presence of sunlight and inorganic energy sources, photoautotrophs such as microalgae or cyanobacteria and chemoautotrophs such as acetogens, crenarchaeota and betaproteobacteria use CO2 as their carbon source (Hu et al., 2019; Zeng, Alain and Shao, 2021). The growth performance of the strain, CO2 sequestration capacity and amount of removal of macronutrients such as phosphate and nitrate from the strain were investigated by growing it under a wide range of CO2 concentrations ranging from 5% to 25%, v/v.
Materials and methods
- Sampling, screening and isolation of potential CO 2 tolerant microalgal strain
- Identification of the isolated strain
- Characterization of the novel microalgal isolate under different physiochemical parameters
- Growth of the organism and CO 2 sequestration under varied CO 2 concentration in the inlet gas stream
- Analysis of growth and substrate utilization of CT01
- Statistical analysis
Morphometric analysis of the stem was performed under phase contrast microscope (Eclipse E200, Nikon, Japan) and Field-effect scanning electron microscope (FESEM, Carl Zeiss SIGMA VP, Germany). Xf and Xi represent the biomass concentration (g L-1) on the last (tf) and initial (ti) days of the culture, respectively.
Results and Discussion
- Screening, isolation and identification of CO 2 tolerant microalgal strain
- Biochemical characterization of CT01
The growth of the microalgal strains was observed to be correlated with the utilization of nitrate and phosphate throughout the screening period. The growth of the organism was compromised when the initial pH of the medium became either acidic.
Conclusions
2020) 'TRENDS IN GLOBAL CO2 AND TOTAL GREENHOUSE GAS EMISSION REPORT', Netherlands Environmental Assessment Agency PBL, (December), p. 2020) 'Recent advances in carbon capture storage and utilization technologies: a review', Environmental chemistry composition may influence heavy metal tolerance of microalgae: a comparison', Journal of Applied Phycology. 2020) "A comparative life cycle assessment of microalgae production from CO2 sequestration from flue gas in outdoor railway ponds under batch and semi-continuous regime", Journal of Cleaner Production.
C HAPTER 4
Background and Motivation
It is also predicted that there could be a 110% increase in demand for protein-enriched foods, adding more demand to conventional production (Tilman et al., 2011). Variations in protein digestibility can be attributed to changes in protein or cell wall matrices (Teuling et al., 2017).
Materials and Methods .1 Feedstock preparation
- Assessment of feedstocks for fish growth metrices and dietetics
- Statistical analysis
The FAME content of the post-extracted residual microalgal biomass was estimated using sequential two-step direct transesterification method (Kumar et al., 2014) and the FAME components were analyzed using GC-FID (Agilent Technologies, USA). Total lipid, total ash and total moisture content of the fish body was performed using the protocol mentioned above.
Results and Discussions .1 Feedstock Composition
- Assessment of feedstocks for fish growth metrices, dietetics and fish quality
Similar to the growth matrix, the diet of fish fry was found to be better when fed MixF compared to ReF. The total dry matter content of fish was found to be similar regardless of the type of raw material.
Conclusions
With the increase in the inclusion rate of Spirulina platensis in the diet of Caspian trout, the total protein and polyunsaturated fatty acid content was reported to increase linearly (Roohani et al., 2019). A pool of literature also reported no negative effect on the growth and quality characteristics of different fish species, such as red tilapia (Oreochromis sp.) or European sea bass (Dicentrarchus labrax), when their conventional meal was supplemented with different microalgae species (Rincón et al., 2012; Tulli et al., 2012b).
1956) 'Colorimetric method for the determination of sugars and related substances', Analytical Chemistry, 28(3), p. 2021a) 'Effect of chlorella-based diets on the growth of silver carp, hypophthalmichthys molitrix and Egyptian Fisheries, 25(4), pp Proceedings of the National Academy of Sciences, 108(50), pp. 2012a) 'Effect of inclusion of dried tetraselmis suecica on growth, feed utilization and fillet composition of juvenile European sea bass fed organic diets', Journal of Aquatic Food Product Technology, 21(3), pp.
C HAPTER 5
Process engineering strategy for cultivation of high-density protein rich CT01 microalgal
Background and Motivation
Although many types of microalgae have been shown to have potential as feedstock in aquaculture, the high cost of biomass production and lower biomass concentration remain major concerns to compete with conventional fishmeal (Han et al., 2019). . Therefore, species-specific optimization of medium composition, its concentration, and culture conditions can improve growth performance and thus make the process economically sustainable and feasible (Shin et al., 2018).
Material and methods
- Microorganism and inoculum preparation
- Media optimization for maximization of biomass titer and productivity
- Effect of light wavelength and intensity on growth kinetics and biochemical composition of CT01
- Growth of CT01 with pH based CO 2 feeding
- Analysis of growth, biomass composition and substrate utilization
- Statistical analysis
We studied the influence of seven different combinations of light wavelengths on the growth kinetics and biochemical composition of the strain. Cell growth was obtained by measuring the optical density of the cells as indicated in section 3.2.4.
Results and Discussions
- Statistical optimization of media for CT01 for maximisation of biomass titer and productivity
- Combinatorial effect of light wavelength and intensity towards production of high- density protein rich biomass
- Growth performance of CT01 with pH based CO 2 feeding strategy
Based on the above regression equations, the optimal concentrations of key nutrients in the Basal Bold medium for maximal biomass production and productivity of CT01 were determined to be 770 mg L-1 sodium nitrate, 174 mg L-1 potassium dihydrogen phosphate, and 0.77 units TME. The optimization of medium concentration for maximum biomass production and better productivity using RSM design provided an efficient medium with altered concentration of nutrients. This result correlated with findings from previous studies showing that the combination of the wavelengths of red and blue light performs better (Yan et al., 2013).
Conclusions
Combinations of red-blue light wavelengths were found to be optimized for CT01 in terms of biomass productivity, carbon sequestration rate and total protein content. CT01 was then subjected to different light intensities in the presence of optimized medium and combinations of red-blue light wavelengths.
2019) 'Effect of light intensity and quality on growth rate and composition of Chlorella vulgaris', Plants 2020, vol. 2015) 'Process engineering for high cell density cultivation of lipid-rich microalgal biomass of Chlorella sp. 2010) 'Effect of aeration and lighting on biomass production and protein biosynthesis in a Spirulina sp. 2015) 'Effect of different light spectra on the growth and productivity of Nannochloropsis sp. 2013) "Effect of different LED light sources and influent carbon/nitrogen ratios on synthetic sanitary wastewater treatment using Chlorella vulgaris", World Journal of Microbiology and Biotechnology.
C HAPTER 6
Demonstration of an integrated sustainable microalgal feed technology at large scale of
Background and Motivation
Most reactors are developed and scaled up using semi-empirical methods (Huang et al., 2017), making them inefficient and expensive. It is well known that light availability is a limiting element in the development of PBR cells (Richmond, 2004; Huang et al., 2011).
Materials and methods .1 Inoculum pre paration
- Cultivation of CT01 in customised 50 L arlift photobioreactor
- Assessment of different types of feedstocks for fish growth metrices and dietetics Study on live microalgal feed (LiF), dry microalgal feed (DrF) and reference feed (ReF) for
- Analysis of growth of CT01 and its substrate utilization
- Biochemical composition of microalgal biomass and fish carcass
- Statistical analysis
The growth dietetic performance and total carcass composition of the mrigal carp were assessed and compared to the dried microalgal biomass of CT01 and the reference diet. The temperature, DO and TDS of the water were measured during the feeding trials using a water test kit (Hach, USA).
Results and discussions
- Development of customised airlift photobioreactor clubbed with pH guided CO 2
- Evaluation of fresh microalgal cell as alternative aquafeed
Nutrient utilization was also found to be higher with pH-based CO2 nutrient batch. The fish survival rate was found to be similar in the LiF and DrF-fed aquariums compared to ReF-fed aquariums.
Conclusions
2008) 'Reduction of CO2 by a high-density culture of Chlorella sp. in a semi-continuous photobioreactor', Bioresource Technology. 2008) 'Comparison of two different pneumatically mixed column photobioreactors for cultivation of Artrospira platensis (Spirulina platensis)', Bioresource Technology.
C hapter 7
Conclusions
