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Globalno zagrijavanje kritičan je problem koji utječe na naš planet i uzrokuje značajne promjene u temperaturi i klimi na Zemlji. Budući da su biljke poikilotermni organizmi, najznačajniji utjecaj globalnog zatopljenja na život biljaka jesu sposobnost kratkotrajne aklimatizacije, kad su temperaturni skokovi značajni i dugotrajna adaptacija na novonastale klimatske uvjete uzrokovane povećanjem temperature. Toplinski stres kritičan je okolišni čimbenik koji može imati ozbiljan utjecaj na rast i razvoj biljaka. Kada su biljke izložene visokim temperaturama tijekom duljeg razdoblja, to može dovesti do širokog spektra fizioloških i biokemijskih promjena, što u konačnici može dovesti do smanjenog rasta i reprodukcije. Jedno od potencijalnih rješenja koja su predložena za ublažavanje utjecaja globalnog zagrijavanja je uporaba algi, posebno vrste mikroalgi Nannochloropsis sp., koje su poznate po svojoj visokoj fotosintetskoj učinkovitosti i stopama rasta. Međutim, globalno zagrijavanje također ima značajne implikacije za alge, uključujući Nannochloropsis sp. čiji molekularni mehanizmi odgovora na stres uključuju aktivaciju kaskade proteina koja u konačnici rezultira sintezom proteina toplinskog udara (Heat shock protein-Hsp). Hsps su grupni naziv za nekoliko različitih evolucijski konzerviranih obitelji proteina od kojih su neki molekularni šaperoni. Hsp70 predstavlja obitelj šaperona čija se koncentracija povećava u stanicama prilikom toplinskog stresa te isti pridonose pravilnom smatanju i stabilizaciji proteina. Bioinformatičkom analizom genoma alge Nannochloropsis sp. utvrdilo se postojanje deset sekvenci proteina iz Hsp70 obitelji i određena su njihova svojstva na temelju fizikalno-kemijskih svojstava, genomskih lokacija, predviđena je stanična lokalizacija pojedinih proteina te su određeni filogenetski odnosi među njima. Global warming is a critical problem that affects our planet and causes significant changes in temperature and climate on Earth. Since plants are poikilothermic organisms, the most significant impacts of global warming on plant life are the ability to acclimate to short-term temperature changes, and long-term adaptation to new climate conditions caused by increased temperature. Heat stress is a critical environmental factor that can have a serious impact on plant growth and development. When plants are exposed to high temperatures for an extended period, it can lead to a wide range of physiological and biochemical changes, ultimately resulting in reduced growth and reproduction. One potential solution proposed to mitigate the impact of global warming is the use of algae, especially the microalga species Nannochloropsis sp., known for their high photosynthetic efficiency and growth rates. However, global warming also has significant implications for algae, including Nannochloropsis sp., whose molecular response mechanisms to stress conditions involve the activation of a protein cascade that ultimately results in the synthesis of heat shock protein (Hsp). Hsps are a group name for several evolutionarily conserved families of proteins, some of which are molecular chaperones. Hsp70 represents a family of chaperones whose concentration increases in cells during heat stress, contributing to proper protein folding and stabilization. Bioinformatic genome analysis of Nannochloropsis sp. algae revealed the existence of ten protein sequences from the Hsp70 family, and their properties were determined based on their physicochemical properties, genomic locations, predicted cellular localization of individual proteins, and their phylogenetic relationships.

Mikroalge rodova Nannochloropsis i Isochrysis predstavljaju sirovinu koja se može koristiti za proizvodnju energije. Cilj ovog rada bio je opisati uzgoj mikroalgi rodova Nannochloropsis i Isochrysis u različitim tehnološkim sustavima te laboratorijskim metodama istražiti njihove kemijske i fizikalne karakteristike za preliminarnu procjenu energetskog potencijala za proizvodnju bioplina i biodizela. Analizom uzoraka mikroalgi Nannochloropsis oculata, utvrđene su srednje vrijednosti sadržaja vode 95,28 %, sadržaja pepela 76,5 %, sadržaja masti 0,3271 %. Salinitet otopine uzoraka mikroalge N. oculata iznosio je 37,114 ppt. Utvrđena je kemijska potrošnja kisika (KPK), od ostataka mikroalgi nakon proizvodnje biodizela, koja je iznosila 1244 mg/L. Analizom uzoraka mikroalgi Isochrysis galbana, utvrđene su srednje vrijednosti sadržaja vode 95,42 %, sadržaja pepela 77,91 %, sadržaja masti 0,3395 %. Salinitet otopine uzoraka mikroalge I. galbana je iznosio 36,516 ppt. Utvrđena je kemijska potrošnja kisika (KPK), od ostataka mikroalgi nakon proizvodnje biodizela, koja je iznosila 999 mg/L. Može se zaključiti da postoji potencijal u proizvodnji biogoriva iz mikroalgi, te da je moguća integrirana kružna proizvodnja. Međutim, potrebno je optimizirati sustav proizvodnje kako bi prinosi bili što veći, a troškovi proizvodnje što manji. Nannochloropsis and Isochrysis microalgae represent a raw material that can be used for energy production. The aim of this research was to describe the cultivation of the Nannochloropsis and Isochrysis microalgae in different technological systems and to determine their chemical and physical characteristics using laboratory methods for a preliminary assessment of the energy potential for the production of biogas and biodiesel. Analysing the Nannochloropsis samples, the average values of water content was measured at 95.28 %, ash content at 76.5 %, fat content at 0.3271 % were determined. The solution salinity of the Nannochloropsis samples was measured at 37.114 ppt. The chemical oxygen consumption (COD) of microalgae residues after biodiesel production was determined, which was 1244 mg/L. Through the analysis of the Isochrysis galbana samples, the average values of water content was measured at 95.42 %, ash content at 77.91 %, fat content at 0.3395 % were determined. The solution salinity of the I. galbana samples was measured at 36,516 ppt. The chemical oxygen consumption (COD) of microalgae residues after biodiesel production was determined, which was 999 mg/L. It can be concluded that there is potential in the production of biofuel from microalgae, and that integrated circular production is possible. However, it is necessary to optimize the production system so that yields are as high as possible and production costs are as low as possible.

Leachate from landfill contains concentrated nutrients that may enter the terrestrial and aquatic environment, including nearby coastal areas. The nutrient contaminants eventually bring harm to marine organisms, including microalgae. This study was performed to investigate the growth of two green microalgal species, i.e. Chlorella sp. and Nannochloropsis sp. in diluted landfill leachate. Besides, the ability of nutrient removal by these microalgal was also explored from the changes of chemical oxygen demand (COD) and nutrients content. The initial and final concentrations of COD, NH3-N, and PO43- in the diluted leachate (5%, 10% and 15%) were measured and the growth patterns of these species were determined by counting the cell numbers for 12 days. Comparison of these microalgae showed that the growth rate of Nannochloropsis was significantly higher compared to Chlorella in all leachate concentrations. Leachate at 5% enhanced the growth of both microalgae, while leachates at 10% and 15% decreased their growth as early as at the beginning of the test. It is apparent that the less concentrated leachate discharged into seawater would not pose any toxicity to the environment and would not bear adverse effect to microalgae yet could promote their growth. This study also revealed that the microalgae could remediate leachate pollution by its ability of nutrient removal; thus, leading to the potential application in wastewater bioremediation, including industrial waste and palm oil mill effluent.

Microalgae have been immobilized in alginate matrixes for the ease of harvesting of the microalgae. However, not all microalgae thrive when immobilized. Immobilized cells are sensitive to the micro-environment in alginate matrixes. Hence, this study aims to improve the growth of immobilized cells and also ammoniacal-nitrogen (NH4 +-N) removal by adding growth medium in alginate beads during the immobilization process. Different strength of the Guillard’s f/2 medium (0%, 30%, 50%, and 100%) was enriched in alginate beads inoculated with Nannochloropsis sp. The immobilized cells were then cultivated in seawater with 1000 µM NH4 +-N as the sole nitrogen source. The growth and NH4 +-N uptake by immobilized cells enriched with different strength of f/2 medium were compared. There were no significant differences on specific growth rate and specific uptake rate of immobilized cells after nutrient enrichment in alginate beads. However, maximum cells attained and maximum NH4 +-N removed increased when strength of f/2 medium enriched in alginate beads was increased. Significantly higher maximum cells attained and maximum NH4 +-N removal were found in immobilized cells enriched with half strength of f/2 medium than those without nutrient enrichment. The present study demonstrated that nutrient enriched in alginate beads can be used by immobilized cells, and subsequently improves the performance of immobilized cells in marine water treatment and production of microalgal biomass simultaneously.

MicroalgaeNannochloropsissp. is a widely recognized renewable biodiesel feedstock. The ability of this microalgae to absorb CO2constitutes an added value toward reducing global warming. However, the process of optimizing its growth still involves many challenges. Photoinhibition, which takes places during microalgae cultivation when using continuous lighting, constitutes an unresolved problem. Therefore, the optimum light/dark cycle method is considered necessary. The experiments were conducted using a designed, tubular airlift photobioreactor and blue, energy-saving, light-emitting diode (LED) lights for the purpose of internal illumination. We observed that a 45:15 min (light:dark) cycle increased the production ofNannochloropsissp. biomass significantly, with a cell density, wet weight, and lipid content of 17 × 106cell/ml, 7.11 g, and 10.1% dry weight, respectively. By using the blue LED lights, our designed, airlift photobioreactor increased cell growth by 70% compared to the growth ofNannochloropsissp. in nature and produced 61 times higher lipid content compared toNannochloropsissp. that is exposed to natural light.

Nannochloropsis sp. is a green alga that is widely used in the aquaculture industry as a feed in Malaysia, but genetic engineering studies of this alga are still underexplored even though there is a growing interest in microalgae genetic engineering for various industrial purposes. This study aims to investigate the efficiency of three transformation methods normally done on microalgae, namely polyethylene glycol (PEG), electroporation, and glass beads on Malaysian indigenous Nannochloropsis sp. using two commercially available plasmids, pUC19 and pGEM-T easy vector as well as an amplicon of ampicillin resistance (AMPR) gene. In this study, out of three transformation methods tested, positive transformants of Nannochloropsis sp. were successfully obtained via electroporation method. Further verification via polymerase chain reaction (PCR) and sequencing confirmed that the electroporation method was found to be the sole successful method in producing transgenic lines of our locally isolated Nannochloropsis sp. Results from this study proved the efficiency of electroporation for delivery of transgene to this green alga which has been reported to be tedious. The described method also provides the gateway for developing Nannochloropsis sp. as a delivery system to aquatic organism due to its importance in the industry.

In this study, microalgal phenolic extracts (MPE) of Nannochloropsis sp. and Spirulina sp. were tested in in vitro experiments and, in comparison with synthetic fungicides, in field experiments, for their ability to control Fusarium graminearum development and limit deoxynivalenol (DON) contamination. In in vitro experiments, the Nannochloropsis and Spirulina extracts inhibited fungal biomass by 34 and 25%, respectively, compared with the untreated control. This effect was confirmed by a reduction in ergosterol production (-80% for Nannochloropsis and -75% for Spirulina) and in DON content (-97% for Nannochloropsis and -62% for Spirulina). In field experiments, application of the fungicide prothioconazole and prothioconazole + tebuconazole resulted in control of Fusarium head blight (FHB) and foliar disease, leading to a significant increase in grain yield (+13%) and a reduction in DON content (-46%) compared to the untreated control. The application of MPE at wheat flowering reduced the severity of FHB compared with the control (-35% for Spirulina and -39% for Nannochloropsis). However, the MPE did not significantly control foliar diseases (Septoria tritici blotch) and therefore did not enhance the grain yield. Moreover, no effect in reducing the DON content in comparison to the control was observed in the field. In view of that, the use of MPE in wheat fields as real alternatives to conventional fungicides requires the discovery of solutions to empower their persistence and efficacy.

Microalgae contain a significant amount of carbohydrates that can be converted further to produce valuable compounds. To extract carbohydrates from microalgae, sub-critical water extraction (SWE) is a viable and novel method. Compared to other existing chemical and biological extraction methods, SWE is more economical, effective, and efficient in terms of process conditions. This process uses high temperature and pressure of water at just below supercritical conditions to keep water at a liquid state. This study aims to investigate the factors that affect the extraction yield of carbohydrates from microalgae Nannochloropsis sp. using the SWE process. The extraction was carried out at a temperature of 160 – 320 oC, a duration of 5 – 25 min, and microalgal biomass loading of 5 - 25% w/v. The kinetics study was conducted in an extraction time of 5 – 25 min at 200 oC and 5% (w/v) biomass loading. A simplified kinetic model based on a consecutive reaction was used to describe carbohydrate production and decomposition via SWE. It was found that the maximum yield of total carbohydrate was 18.04 g/100 g which was obtained at 200 oC, 10 min, and 5% (w/v) microalgal biomass loading. The carbohydrate concentration was decreased as the temperature, extraction time and microalgal biomass loading increased. This result indicated that the SWE is a promising extraction method for carbohydrate recovery from microalgae, and the factors investigated here had a significant effect on the extraction process. The proposed kinetic model was also able to capture the experimental data well within the range of operating conditions studied in this work.

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Biomass of algae is a very potent adsorbent for absorbing aqueous waste containing heavy metals and organic dyes. This study purposes to confirm the ability of adsorbents from green algae Nannochloropsis sp. modified with silica (ASN) and followed by coating magnetite particles (ASN-MPs) to absorb simultaneously the mixture of Methylene Blue (ME) and Cu(II) cations in aqueous solution. Simultaneous sorption of ME and Cu(II) cations to ASN and ASN-MPs was carried out by the batch method with the interaction pH condition 7, contact time 90 min, and initial concentrations of ME and Cu(II) cations (0.1–1.0 mM). Based on adsorption data, Cu(II) cations have a greater adsorption rate and capacity (qm) compared to ME at the same contact time and initial concentration. The adsorption capacity (qm) values of the bi-component ME and Cu(II) cation mixture in ASN and ASN-MPs were 1.39 × 10− 1 and 5.32 × 10− 1 mmol g− 1, respectively, with the binary Langmuir adsorption isotherm constant for Cu(II) cations greater than ME. Modified adsorbent from algae Nannochloropsis sp. with silica matrix and magnetite particle coating is an adsorbent that has a high effectiveness in the collective sorption of ME and Cu(II) cations. Therefore, these adsorbents can be used for the adsorption of cation mixtures of heavy metals and organic dyes that are cationic in solution.

The increased concentration of Nannochloropsis sp. microalgae during the harvesting process is of great importance economically. Separation of Nannochloropsis sp. using Polyvinylidene Fluoride (PVDF) polymeric membrane was studied with the addition of 2% LiCl to improve up-concentration performance. The Hermia model is used to predict the type of fouling on the membrane. Cleaning with NaOCl 1000 ppm solution was carried out to overcome fouling on the membrane. PVDF/LiCl membranes were prepared using the immersion-precipitation phase inversion method. The results showed that the addition of LiCl caused an increase in membrane hydrophilicity as indicated by a decrease in the value of the contact angle from 94.8° to 70-84°. Membrane morphology changed from finger-like to sponge-like. The concentration of Nannochloropsis sp. increased from 563.92 to 5097.5 ppm. The membrane performance after cleaning is still very good because it produces recovery is more than 89.924%, and the microalgae rejection is 100%. It was concluded that the PVDF membrane after the addition of LiCl could improve the up-concentration performance of Nannochloropsis sp. and NaOCl solution was very effective in overcoming fouling on the membrane.

Powdered biomass of Nannochloropsis sp. and Tetraselmis sp. were used as innovative ingredients in wheat tortillas at flour substitution levels of 0.5–3.0%. Incorporation of microalgae into the tortilla formulations led to increased protein and fat content. The content of phenolic and carotenoids was also higher in microalgae-enriched tortillas, especially for those enriched in Nannochloropsis sp. at a flour substitution level of 3.0%. Not only the phenolic content but also the antioxidant capacity of the tortillas was higher after microalgae incorporation. Bioaccessible polyphenols were also higher in microalgae-containing tortillas as well as the antioxidant capacity of the enzymatic digestive extracts. No major differences in physical parameters (besides colour) were observed, and the overall acceptance of the microalgae-enriched tortillas assessed after a sensorial analysis was comparable to that of the wheat-only controls. Moreover, the purchase intention of the products as well as the acceptability index suggested that the tortillas would have a good acceptance. Several reports highlighted the nutritional value of microalgae. The production of microalgae and their utilisation as novel ingredients in innovative foods will increase the sustainability of the food industry and promote health. info:eu-repo/semantics/acceptedVersion

Lipidi izolirani iz biomase mikroalgi Nannochloropsis sp. predstavljeni su kao perspektivna sirovina za proizvodnju biodizela. U ovom radu proveden je fototrofni šaržni uzgoj s pritokom supstrata u otvorenom sustavu na izvornoj f/2 podlozi uz dodatak natrijevog bikarbonata te fototrofni šaržni uzgoj s pritokom supstrata u barbotirajućoj koloni na izvornoj f/2 podlozi bez dodatnog izvora ugljika. Tijekom uzgoja praćene su promjene udjela proteina, ugljikohidrata i lipida u izdvojenoj biomasi kao i sastav pigmenata te profil masnih kiselina u ukupnim lipidima. Utvrđeno je da u periodu smanjene koncentracije fosfora, metabolizam mikroalge se usmjerava prema akumulaciji lipida i ugljikohidrata kao rezervnih makromolekula uz smanjenje udjela proteina. Posljednjeg dana šaržnog uzgoja u otvorenom bazenu postignuta je koncentracija biomase od 0,515 g Lˉ¹, a maksimalna produktivnost biomase od 0,8796 mg Lˉ¹ hˉ¹ postignuta je osamnaestog dana uzgoja. Šaržnim uzgojem u barbotirajućoj koloni koncentracija biomase na kraju uzgoja iznosila je 0,458 g Lˉ¹, a produktivnost sinteze biomase tada je bila 0,8297 mg Lˉ¹ hˉ¹. Kemijska flokulacija sa željezovim (III) kloridom potvrđena je kao alternativna metoda za izdvajanje stanica mikroalgi. Naposljetku, u radu je predstavljen elaborat tehničko-tehnološkog rješenja postrojenja za proizvodnju biodizela. Lipids isolated from biomass of microalgae Nannochloropsis sp. are presented as a promising feedstock for biodiesel production. Cultivation methods used in this paper were phototrophic fed-batch cultivation with substrate inflow in an open pond system on the original f/2 medium with sodium bicarbonate and phototrophic fed-batch cultivation with substrate inflow on the original f/2 medium without supplementation of carbon source in a bubble column. Changes in the content of proteins, carbohydrates and lipids were monitored in the separated algae biomass during cultivation. The composition of pigments and the profile of fatty acids in total lipids were as well monitored simultaneously. During the period of reduced phosphorus concentration it was confirmed that metabolic responses of microalgae lead to the accumulation of lipids and carbohydrates as reserve macromolecules and to a decrease in protein content. On the last day of fed-batch cultivation in open pond, biomass concentration of 0,515 g Lˉ¹ was achieved. The maximum productivity of 0,8796 mg Lˉ¹hˉ¹ was achieved eighteenth day of cultivation. Additionally, on the last day of fed-batch cultivation in bubble column, biomass concentration was 0,458 g Lˉ¹ and productivity of biomass synthesis was 0,8297 mg Lˉ¹ hˉ¹. Chemical flocculation with iron (III) chloride has been confirmed as an alternative method for microalgae cell harvesting. Lastly, technical-technological solution for a biodiesel production facility was presented in this paper.

This work proposes a way to maximize the potential of a Nannochloropsis sp. biorefinery process, through membrane technology, producing an extract enriched in soluble proteins, free from the insoluble protein fraction, with a low lipid content and eliminating the colored chlorophyll-a. This procedure, following the principles of a circular economy approach, allows for the valorization of a stream from the biorefining of Nannochloropsis sp. that, otherwise, would be considered a residue without commercial value. The process proposed minimizes fouling phenomena at the membrane surface, making it possible to achieve high permeate fluxes, thus reducing the need for membrane cleaning and, therefore, contributing to an extended membrane lifetime. Supernatant obtained after centrifugation of a suspension of ruptured Nannochloropsis sp. cells was processed by ultrafiltration using a membrane with a cut-off of 100 kDa MWCO. Two different operating approaches were evaluated—controlled transmembrane pressure and controlled permeate flux—under concentration and diafiltration modes. Ultrafiltration operated in a diafiltration mode, under controlled permeate flux conditions, led to the highest soluble protein recovery (78%) with the highest constant permeate flux (12 L·m−2·h−1) and low membrane fouling.

The microalgae Aurantiochytrium sp. (AUR), Isochrysis sp. (ISO), and Nannochloropsis sp (NAN) were studied as possible alternative feeds to well established commercial compound feeds for both rotifers (Brachionus plicatilis) and Artemia franciscana. Fatty acid (FA) composition —relative (in % of total FAs) and absolute (in mg/g dw)—was determined in order to assess their potential for providing essential FAs. The FA profiles showed relevant differences between the four feeds (compound feed and the three microalgal species), but less stark than in the feeds themselves. Whereas Isochrysis sp. was relatively rich in DHA and poor in EPA, 18.6 ± 1.7% vs 0.6 ± 0.0%, respectively, Nannochloropsis sp. had the opposite pattern, 0.2 ± 0.3% vs 28.3 ± 0.7%. Aurantiochytrium sp. was rich in DHA (19.1 ± 0.2% corresponding to 89.8 ± 0.2 mg/g dw), but posed difficulties as a feed for both rotifers and artemia, given its low lipid incorporation and, in particular, poor DHA deposition. Rotifers fed the compound feed had the best combination of n3 PUFA levels (22.1 ± 0.1 mg/g dw), DHA contents (13.6 ± 0.4 mg/g dw), and DHA/EPA ratios (~3), being rotifers fed AUR and ISO feeds second best. Hence, these microalgae may deserve to be further explored as potential sources of specific FAs in rotifers and artemia., Food science; Food technology; Biotechnology; Aurantiochytrium sp.; Isochrysis sp.; Nannochloropsis sp.; Lipid composition rotifers; Artemia franciscana.

Nannochloropsis sp. is a green alga that is widely used in the aquaculture industry as a feed in Malaysia, but genetic engineering studies of this alga are still underexplored even though there is a growing interest in microalgae genetic engineering for various industrial purposes. This study aims to investigate the efficiency of three transformation methods normally done on microalgae, namely polyethylene glycol (PEG), electroporation, and glass beads on Malaysian indigenous Nannochloropsis sp. using two commercially available plasmids, pUC19 and pGEM-T easy vector as well as an amplicon of ampicillin resistance (AMPR) gene. In this study, out of three transformation methods tested, positive transformants of Nannochloropsis sp. were successfully obtained via electroporation method. Further verification via polymerase chain reaction (PCR) and sequencing confirmed that the electroporation method was found to be the sole successful method in producing transgenic lines of our locally isolated Nannochloropsis sp. Results from this study proved the efficiency of electroporation for delivery of transgene to this green alga which has been reported to be tedious. The described method also provides the gateway for developing Nannochloropsis sp. as a delivery system to aquatic organism due to its importance in the industry., Graphical Abstract

Nannochloropsis sp. has received increased attention by researchers in recent years due to its complexity and abundance of lipid structures. The lipids of this microalgae species have been identified to contain large quantities of neutral lipids which are capable of producing raw materials for nutraceuticals, food additives and biofuels. The production of biodiesel has received the greatest attention as there is an increase in global demand for both more fuel and more environmentally sustainable methods to produce such resources. The greatest challenges facing industries to mass produce viable products from microalgae involve the degradation of the cell wall and extracting the fatty acid of interest due to high costs. Various studies have shown that the extraction lipids from the microalgae can greatly influence the overall fatty acid composition. Different extraction methods can re- sult in recovering higher quantities of either saturated fatty acids, monounsaturated fatty acids or polyunsatu- rated fatty acids. Biodiesel production requires higher quantities of saturated fatty acids and monosaturated fatty acids as increased quantities of polyunsaturated fatty acids result in oxidation which decreases the perfor- mance of the biodiesel. Whereas, polyunsaturated fatty acids are required in order to produce pharmaceuticals and food additives such as omega 3. This review will focus on how different in-situ extraction methods for lipid and fatty acid recovery, influence the fatty acid composition of various Nannochloropsis species (oculate, gaditana, salina and oceanica). The mechanical methods (microwave, ultrasonic and supercritical‑carbon dioxide) of extraction for Nannochloropsis sp. will be critically evaluated. The use of enzymes will also be addressed, for their ability to extract fatty acids in a more environmentally friendly manner. This paper will report on the viable byproducts which can be produced using different extraction methods.

Pharmaceuticals and their metabolites are released into the environment by domestic, hospital, and pharmaceutical industry wastewaters. Conventional wastewater treatment technology does not guarantee effluents of high quality, and apparently clean water may be loaded with pollutants. In this study, we assess the performance and efficiency of free and immobilised cells of microalgae Nannochloropsis sp. in removing four pharmaceuticals, chosen for their occurrence or persistence in the environment. These are paracetamol, ibuprofen, olanzapine and simvastatin. The results showed that free microalgae cells remain alive for a longer time than the immobilised ones, suggesting the inhibition of cell proliferation by the polymeric matrix polyvinyl alcohol. Both cells, free and immobilised, respond differently to each pharmaceutical. The removal of paracetamol and ibuprofen by Nannochloropsis sp., after 24 h of culture, was significantly higher in immobilised cells. Free cells removed a significantly higher concentration of olanzapine than immobilised ones, suggesting a higher affinity to this molecule than to paracetamol and ibuprofen. The results demonstrate the effectiveness of Nannochloropsis sp. free cells for removing olanzapine and Nannochloropsis sp. immobilised cells for removing paracetamol and ibuprofen.

The current study evaluated the effects of fish oil replacement with eicosapentaenoic acid-enriched single-cell microalgae in the diets of larval and postlarval kuruma shrimp. Experimental diets containing different level of Nannochloropsis sp. powder (10, 40, 70 g/kg) and lipids (2, 8, 10, 14, 35, 58 g/kg) were evaluated. The substitution of fish oil with algal powder significantly improved shrimp growth compared to that in the control group, with the highest final body weight recorded at 70 g/kg in larvae and 40 g/kg in postlarvae. Larvae fed algal lipids with a dry weight percentage in the diet of greater than 10 g/kg showed significantly lower performance than those in the control group. The neutral lipids in the tissue of the postlarvae that had been fed algal lipids had an improved fatty acid profile, as the content of highly unsaturated fatty acids increased compared to that in the other groups. The content of docosahexaenoic acid increased in the tissue, indicating the possible occurrence of the metabolism and accumulation of Nannochloropsis sp. essential fatty acids. Algal powder could replace FO up to 140 g/kg in shrimp diets without compromising growth and FA profiles.

Microalgae are photosynthetic microorganisms widely used at the industrial level because of their high content of compounds with bioactive properties such as antimicrobial peptides. These compounds have a wide range of action mechanisms, do not produce the resistance of conventional antibiotics and could be effective against bacteria, fungi, viruses, parasites, and even tumor cells. In this work, by mechanical cell lysis treatment, a protein extract of dry biomass of Nannochloropsis sp was obtained, subsequently, it was hydrolyzed with an enzyme complex of proteases, papain, and pancreatin, for 6 hours, at 44 ° C and pH 7. Finally, the antimicrobial activity of the fraction of hydrolyzed extract with a size less than 3 kDa was evaluated against Staphylococcus aureus, Escherichia coli, and Candida albicans, obtaining a percentage of growth inhibition of 93.1%, 96.6%, and 14.4% respectively, which represents a peptic extract with bacteriostatic and fungistatic properties. Resumen Las microalgas son microorganismos fotosintéticos ampliamente utilizados a nivel industrial gracias a su alto contenido de compuestos con propiedades bioactivas como lo son los péptidos antimicrobianos. Estos compuestos son eficaces contra bacterias, hongos, virus, parásitos e incluso células tumorales, poseen una amplia variedad de mecanismos de acción y no generan la resistencia de los antibióticos convencionales. En este trabajo, a través de lisis celular mecánica, se obtuvo un extracto de proteínas a partir de biomasa seca de Nannochloropsis sp., posteriormente fue hidrolizado usando un complejo enzimático de proteasas, papaína y pancreatina, durante 6 horas, a 44°C y pH 7. Finalmente, se evaluó la actividad antimicrobiana de la fracción de extracto hidrolizado que poseía un tamaño menor a 3 kDa contra Staphylococcus aureus, Escherichia coli y Candida albicans, obteniendo un porcentaje de inhibición de crecimiento de los microorganismos de 93,1%, 96,6%, y 14,4% respectivamente, lo cual representa un extracto peptídico con propiedades bacteriostáticas y fungistáticas.

Harvesting, a challenging step of microalgae processing appears to be promising using flocculation method by the addition of tannin based natural biopolymer, AFlok-BP1. In this study, marine microalgae Nannochloropsis sp. was flocculated using AFlok-BP1 to evaluate its effectiveness in terms of flocculation efficiency, pH and dosages used, zeta potential, floc morphology characterization with comparison to the uses of aluminium sulphate and alkaline-induced pH. Oleaginous marine microalgae Nannochloropsis sp. was found to work effectively using AFlok-BP1 at an acidic pH 5 with more than 90% efficiency obtained at 160 mg/L. The finding was further supported with great instability of microalgae surface charge of -0.33 ± 0.27 mV, hence maximum destabilization of microalgae was believed to occur. The use of alum also effective to harvest Nannochloropsis sp. at neutral pH 7 with comparable efficiency as AFlok-BP1, but high dosage needed for more than 300 mg/L was at major concerns. Similarly, improved efficiency was observed for alkaline-induced pH for almost 100% efficiency. This would provide a better alternative for marine microalgae flocculation however, high concentration of minerals in the harvested biomass due to pH increase in need of further evaluation. To summarize, AFlok-BP1 would be a viable option to harvest marine microalgae Nannochloropsis sp. for the production of valuable compounds as it is an environmental friendly flocculant and provide no potential toxicity to the harvested biomass.

Fungicides and, for the first time, microalgal phenolic extracts (MPE) from Spirulina sp. and Nannochloropsis sp. were applied on maize culture media under field conditions to evaluate their ability to minimize Fusarium species development and fumonisin production. An in vitro assay against F. verticillioides was carried out using maize grains as the culture medium. An open-field experiment was carried out in Northwest Italy under natural infection conditions. The compared treatments were factorial combinations of two insecticide treatments (an untreated control and pyrethroid, used against European Corn Borer), four antifungal treatments (an untreated control, MPE from Spirulina sp., MPE from Nannochloropsis sp., and a synthetic fungicide), and two timings of the application of the antifungal compounds (at maize flowering and at the milk stage). The MPE compounds were capable of inhibiting fumonisin production in vitro more efficiently than tebuconazole. Insecticide application reduced the infection by Fusarium species and subsequent fumonisin contamination. However, fumonisins in maize fields were not significantly controlled by either fungicide or MPE application.

Recently, microalgal biomass has attached much attention due to the wide diversity of compounds synthesized from different metabolic pathways. This work attempts to study metabolites recovery from Nannochloropsis sp. biomass concentrated by centrifugation and flocculation. Carbohydrates were obtained using acid and alkaline hydrolysis required for cell disruption. Protein extraction was performed after alkaline pretreatment and lipids were recovery by acid hydrolysis- Soxhlet and alkaline hydrolysis- Soxhlet extraction routes. It was found that carbohydrates were recovered by acid hydrolysis in 41 % and 35.39 % for centrifuged and flocculated biomass, respectively, values higher than thus reported using alkaline hydrolysis. For protein extraction, centrifuged biomass exhibited higher recovery yield (55.48%) than flocculated biomass (38.40%). The lipid extraction route that achieved highest yield (43.45%) was acid hydrolysis with HCl followed by Soxhlet extraction with hexane. In addition, statistical analysis by T test suggested that flocculants affect negatively biomass culture, hence, efficiency of metabolites extraction.

Data of this article describes growth, survival rate, water quality and hemato-biochemical indices of Nile Tilapia (Oreochromis niloticus) fry. To collect the data, the Nile Tilapia fry was reared in 30 L glass aquarium (18 fish/ tank) for 56-days under controlled environmental condition. Feed was prepared with 25 and 50% replacement of commercial fish meal with Nannochloropsis sp. and Tetraselmis sp. microalgae, while no replacement was made for control feed. Initial and final body weight of fish was recorded to find the data of growth rate; survival rate was calculated from the initial and final live individuals recorded during the experiment; physico-chemical parameters were analyzed to collect water quality data; hemato-biochemical indices were collected using hematology analyzer and photometry. The data on growth, survival rate and hemato-biochemical indices were statistically significant (p < 0.05). Therefore, these data might contribute to the selection of marine microalgae to improve the water quality during fish farming which could enhance the growth and survivability of fish. In addition, the data of hemato-biochemical indices represent that feeding selected marine microalgae might result in the production of healthy and disease-free fish.

The effects of Ca2+ in converter slag and electric arc furnace slag leaching solutions on CO2 content, the effects of CO2 content in solutions on the accumulation of chlorophyll a of Nannochloropsis sp., and the mechanisms were studied. Results showed that addition of 50 mg/L converter slag leaching solution and 400 mg/L electric arc furnace slag leaching solution (lower than 50 vol% concentration) promoted the accumulation of chlorophyll a of Nannochloropsis sp., while an increased concentration of 70 vol% inhibited its accumulation. The highest concentration of chlorophyll a was obtained on the addition of 10 vol%. With 10 vol% leaching solutions added, chlorophyll a concentration for converter slag reached 4.2 mg/L, 1.45 times as much as that of pure F/2 medium, and chlorophyll a concentration for electric arc slag reached 3.2 mg/L, 1.10 times as much as that of pure F/2 medium. Under the same addition proportion, the promoting or inhibiting effect from converter slag on the accumulation of chlorophyll a of Nannochloropsis sp. was more obvious than that for electric arc furnace slag. The reason for the promotion of chlorophyll a accumulation of Nannochloropsis sp. was that an increase of Ca2+ in the solutions led to an increase of CO2, thereby promoting the photosynthetic rate of microalgae. The study provided new ideas for the green application of solid waste and industrial production of microalgae.

The efficiency of harvesting the algal biomass plays a dominant role in the production of biodiesel from microalgae. On a commercial scale the choice and efficiency of the harvesting techniques may be a deciding factor for economic viability of this renewable energy source. The highly energy intensive centrifugation and filtration process are followed by flocculation for harvesting microalgae. Therefore the choice, cost and efficiency of the flocculating techniques can effectively lower the cost of harvesting and in turn minimize the cost of biodiesel production. This study aims to investigate the electroflocculation technique for harvesting microalgae under optimizing the operating parameters viz. voltage and current. Chaetoceros calcitrans and Nannochloropsis sp. were cultured and electrofloculated with a harvesting efficiency of 92 and 95% respectively at the optimum voltage and current.

En la producción de alimento vivo, representado principalmente por microalgas, se buscan nuevas técnicas de medios de cultivo sustituyendo las fórmulas tradicionales en aras de reducir costos. Teniendo en cuenta esto, se usó un medio no convencional (fertilizantes industriales) para producir las microalgas Isochrysis galbana y Nannochloropsis sp. en cultivos estáticos, y se alcanzaron densidades de 7,5 × 106 cel mL–1 de Nannochloropsis sp. en 1000 L y 0,265 × 106 cel mL–1 de I. galbana en 250 L. Hubo un crecimiento exponencial, que se alcanzó entre los tres y los ocho días, en que se duplicaron las densidades iniciales. Se sugiere cosechar en lapsos cortos (hasta cuatro días), debido a la proliferación de contaminantes y la disminución de la calidad de las células con el tiempo. Se concluye que el medio diseñado para el enriquecimiento del cultivo es una alternativa viable para la producción de las microalgas que permite satisfacer la demanda alimenticia en el levante de larvas de peces marinos.

Harvesting microalgae from medium is a major challenge due to their small size and low concentrations. In an attempt to find a cost-effective and eco-friendly harvesting technique, mung bean (Vigna radiata) protein extract (MBPE) was used for flocculation of Nannochloropsis sp. The effects of parameters such as pH, flocculant dose, algae concentration, and mixing time were used to study the flocculation efficiency (FE) of MBPE. Optimum parameters of MBPE dosage of 20 mL L−1 and a mixing rate of 300 rpm for 6 min achieved a FE of >92% after 2 h of settling time. MBPE-aggregated microlga flocs were characterized by microscopy. Zeta potential values decreased with increasing flocculant dose, and the values obtained were −6.93 ± 0.60, −5.36 ± 0.64, and −4.44 ± 0.22 for doses of 10, 20, and 30 mL L−1, respectively. In conclusion, MBPE flocculants used in this study are safe, nontoxic, and pollution free, so they could be used for an effective, convenient, and rapid harvesting of microalgae in an eco-friendly approach. These methods are sustainable and could be applied in industrial scale for aquaculture nutrition.

In this article, microalgae Nannochloropsis sp. was used for fatty acid (FA) extraction, using a supercritical fluid-carbon dioxide (SF-CO2) extraction method. This study investigated the influence of different pre-treatment conditions by varying the grinding speed (200&ndash, 600 rpm), pre-treatment time (2.5&ndash, 10 min), and mixing ratio of diatomaceous earth (DE) and Nannochloropsis sp. biomass (0.5&ndash, 2.0 DE/biomass) on FAs extraction. In addition, the effect of different operating conditions, such as pressure (100&ndash, 550 bar), temperature (50&ndash, 75 &deg, C), and CO2 flow rate (7.24 and 14.48 g/min) on eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) recovery, was analyzed. Experimental data evidenced that, keeping constant the extraction conditions, the pre-treatment step enhanced the FAs extraction yield up to 3.4 fold, thereby the maximum extracted amount of FAs (61.19 mg/g) was attained with the pre-treatment with a ratio of DE/biomass of 1 at 600 rpm for 5 min. Moreover, by increasing both SF-CO2 pressure and temperature, the selectivity towards EPA was enhanced, while intermediate pressure and lower pressure promoted DHA recovery. The highest amount of extracted EPA, i.e., 5.69 mg/g, corresponding to 15.59%, was obtained at 75 &deg, C and 550 bar with a CO2 flow rate of 14.48 g/min, while the maximum amount of extracted DHA, i.e., ~0.12 mg/g, equal to 79.63%, was registered at 50 &deg, C and 400 bar with a CO2 flow rate of 14.48 g/min. Moreover, the increased CO2 flow rate from 7.24 to 14.48 g/min enhanced both EPA and DHA recovery.

Las microalgas son microorganismos fotosintéticos que requieren nutrientes relativamente simples, que permiten la generación de biomasa útil en la producción de lípidos y proteínas para la alimentación animal, aceites para biocombustibles, la producción de pigmentos, como sistemas de biorremediación de aguas residuales, entre otras aplicaciones. Sin embargo, los costos de producción son relativamente altos en comparación con otras fuentes de proteínas de origen vegetal o animal comunes en la producción de alimentos concentrados para la acuicultura. Hay varios factores que influyen en los costos de producción de la biomasa de microalgas, entre los que se encuentran el medio de crecimiento y la fuente de luz, el último para cultivos que utilizan lámparas eléctricas en entornos cerrados o de laboratorio. En este trabajo, la producción de biomasa de la microalga Nannochloropsis sp, se evaluó utilizando un fertilizante N-P-K en fotobiorreactores exteriores utilizando la luz solar como fuente de energía. Para el cultivo al aire libre, se utilizaron 8 fotobiorreactores de columna de burbujeo hechos de fibra de vidrio, ubicados en la Sede de Estudios Ecológicos y Agroindustriales del SEEA de la Universidad de Antioquia en el municipio de Carepa Antioquia, Colombia. Se utilizó la microalga Nannochloropsis sp, se evaluaron las mezclas de aire y CO2 con tres niveles (0,022; 3 y 6% de CO2 en la corriente de aire) y se evaluó un fertilizante comercial tipo NPK como medio de cultivo (13-40-13), con tres niveles en concentraciones de 6,17; 12,34 y 18,51 mg Nitrógeno / L y como control, el medio de referencia Guillard F / 2 (12,34 mg nitrógeno / L). Para los cultivos en el laboratorio, a una concentración de fertilizante de 18,51 mg Nitrógeno/L, se logró la mayor producción de biomasa y porcentaje de proteína (biomasa de 289 mg/L y 21% de proteína), valores que fueron más altos que los obtenidos con el medio de referencia. Sin embargo, para cultivos al aire libre, los mejores resultados se obtuvieron con el medio de referencia. Los experimentos se llevaron a cabo en condiciones ambientales de luz y temperatura del municipio de Carepa-Antioquia, logrando una producción de biomasa microalgal rica en proteínas, siendo mayor en el medio de referencia al comparar el fertilizante agrícola, 300 mg/L de biomasa con 22 % de proteína con el medio de referencia y 200mg/L con 23% de proteína con el fertilizante. Además, se concluye que es posible cultivar la biomasa de Nannochloropsis sp bajo las condiciones ambientales del municipio de Carepa y los costos de usar fertilizantes como medio de cultivo son 90 veces más bajos en comparación con el medio de referencia. Microalgae are photosynthetic microorganisms that require relatively simple nutrients, which allow the generation of biomass useful in the production of lipids and proteins for animal feed, oils for biofuels, the production of pigments, among other applications as bioremediation systems for wastewater. However, production costs are relatively high compared to other protein sources of plant or animal origin common in the production of concentrated feeds for aquaculture. There are several factors that influence the production costs of microalgae biomass, among which are the growing medium and the light source, the latter for crops that use electric lamps in closed or laboratory environments. In this work, the biomass production of the microalga Nannochloropsis sp, was evaluated using an N-P-K fertilizer in outdoor Photobioreactors using sunlight as an energy source. For outdoor culture, 8 bubble column photobioreactors made of glass fiber were used, located in the Headquarters of Ecological and Agroindustrial Studies of the SEEA of the University of Antioquia in the municipality of Carepa Antioquia, Colombia. The microalga Nannochloropsis sp was used, the air and CO2 mixtures were evaluated with three levels (0.022, 3 and 6% of CO2 in the air stream) and a commercial NPK type fertilizer was evaluated as a culture medium (13-40-13), with three levels in concentrations of 6.17; 12.34 and 18.51 mg Nitrogen / L and as control, the Guillard reference medium F/2 (12.34 mg nitrogen / L). For the cultures in the laboratory, at a fertilizer concentration of 18.51 mg Nitrogen / L, the highest biomass production and the percentage of protein (biomass of 289 mg / L and 21% protein) were achieved, values that were higher than those obtained With the reference medium. However, for outdoor cultures, the best results were obtained with the reference medium. The experiments were carried out under environmental conditions of light and temperature of the municipality of Carepa-Antioquia, achieving a microalgae production of protein-rich biomass, being higher in the reference medium when comparing the agricultural fertilizer, 300 mg / l of biomass with 22% protein with the reference medium and 200mg / L with 23% protein with the fertilizer. In addition, it is concluded that it is possible to grow the biomass of Nannochloropsis sp under the environmental conditions of the municipality of Carepa and the costs of using fertilizers as a culture medium are 90 times lower in comparison with the reference medium. Maestría

A feeding trial was conducted to assess the effects of dietary Nannochloropsis sp. as powder or lipid on survival, growth performance and fatty acids content in Kuruma shrimp juvenile. Seven isonitrogenous, isolipidic and isocaloric diets were formulated to contain in first group graded levels of Nannochloropsis powder: 0% (CON, control diet), 4% (A), 7% (B) and 10% (C) dry matter then in second group algae oil proportionally extracted (AE, BE and CE, respectively). The feeding trial was conducted for 50 days. The results showed that the substitution of fish oil (FO) with algae powder diets A, B and C significantly (P

Las microalgas son microorganismos fotosintéticos, ampliamente utilizados a nivel industrial por su alto contenido de proteínas, lípidos, carbohidratos, vitaminas, pigmentos entre otros. Específicamente la proteína microalgal es una fuente prometedora de péptidos con propiedades bioactivas, entre ellas, antimicrobianas. Los péptidos antimicrobianos poseen un amplio rango de acción contra bacterias, hongos, virus, parásitos y células tumorales y no producen la resistencia de los antibióticos convencionales. En este trabajo, a partir de biomasa de la microalga Nannochloropsis sp. se obtuvo un extracto de proteínas mediante el desarrollo de un protocolo de disrupción celular mecánica por homogenización a pH neutro, el cual alcanzó un porcentaje de lisis del 60%. El extracto y la biomasa intacta fueron sometidos a hidrolisis enzimática con dos proteasas de manera simultánea, papaína y pancreatina, alcanzando un grado de hidrólisis superior al 80% y al 20% respectivamente, después de 6 horas de reacción, demostrando la resistencia que ejerce la pared celular frente la acción enzimática. Se evaluó el efecto antimicrobiano de diferentes concentraciones del extracto hidrolizado con un tamaño menor a 3KDa contra Staphylococcus aureus, Escherichia coli y Candida albicans, obteniendo porcentajes de inhibición de 93,14%, 96,65% y 14,46% respectivamente, convirtiendo a Nannochloropsis sp, en una microalga con un gran potencial para la obtención de péptidos bioactivos con actividad antimicrobiana. Palabras clave: péptidos antimicrobianos, proteína microalgal, hidrolizado enzimático, Nannochloropsis sp. Abstract: Microalgae are photosynthetic organisms, widely used at industrial level for their high content of proteins, lipids, carbohydrates, vitamins, pigments, and others. Specifically, the microalgal protein is a promising source of peptides with bioactive properties, including antimicrobial. Antimicrobial peptides have a wide range of action against bacteria, fungi, viruses, parasites and tumor cells and do not produce the resistance of conventional antibiotics. In this work, from Nannochloropsis sp biomass, a protein extract was obtained through the development of a mechanical cell disruption protocol by homogenization, which reached a 60% lysis percentage. The extract and the intact biomass were subjected to enzymatic hydrolysis with two proteases simultaneously, papain and pancreatin, reaching a degree of hydrolysis higher than 80% and 20% respectively, after 6 hours of reaction, demonstrating the resistance exerted by the cell wall against enzymatic action. The antimicrobial effect of different concentrations of the fraction less than 3KDa of the hydrolyzed extract against Staphylococcus aureus, Escherichia coli and Candida albicans was evaluated, obtaining average inhibition percentages of 93,14, 96,65% and 14,46% respectively, converting Nannochloropsis sp, into a microalgae with a great potential for obtaining bioactive peptides with antimicrobial activity. Keywords: antimicrobial peptides, microalgal protein, enzymatic hydrolyzate, Nannochloropsis sp. Maestría

Microalgae have a wide range of applications in industries such as feed and aquaculture, functional food, chemicals, pharmaceuticals, cosmetics and biofuels. Despite their high potential, widespread large-scale cultivation of microalgae especially for low-value applications struggles to get off the ground. To increase the economic viability of bulk cultivation, production costs need to be reduced. Recycling growth medium not only allows salvaging the unused nutrients remaining in the water after harvesting, but also permits to reduce water demand and to avoid or reduce waste water treatment costs. To be able to implement a recycling method in a fully automated production system of microalgae, several challenges such as cleaning of spent medium, replenishment of nutrients and comprehending positive or negative effects on microalgal growth needed to be addressed. The present study explored a method to rejuvenate centrifuge supernatant (centrate) in a robust and inexpensive way, mapped nutrient consumptions and formulated dosing strategies for an automated replenishment, and gained understanding in the effect of medium recirculation on the cultivation of Nannochloropsis sp. and Tisochrysis lutea, separately and across species borders. In Chapter 2, a polishing treatment was sought for centrate originating from centrifugation of high density cultures of Nannochloropsis sp. to feed into a microfiltration setup generating contaminant-free growth medium. High pH-induced coagulation/flocculation (with NaOH or Ca(OH)2) and ‘rapid’ sand filtration (start exhaust flow rate of 9 m³·m-2·h-1) were investigated separately, and linked at optimal settings to reduce membrane fouling and increase microfiltration performance. Stand-alone sand filtration provided a meagre improvement in centrate filterability; ‘modified fouling index’ decreased 21 ± 21% after 55 min. High pH induced coagulation/flocculation with NaOH and Ca(OH)2 at pH 11.5 reached flocculation efficiencies of 90 ± 1% and 92 ± 4% respectively. When combining both techniques, the obtained flocculated centrates, however, resulted in rapid clogging of the sand bed. Consequently, only the combination of NaOH coagulation/flocculation at pH 10.5 and subsequent sand filtration was further investigated. This combination removed 78 ± 18% of particles and improved ‘modified fouling indices’ with 75 ± 19%. After 60 minutes filtration runs, the sand bed was backwashed with (acidified) water during 5 minutes to regenerate the sand bed. The results indicated that there is room for improvement by optimizing washing regimes of the sand filter. In any case, the suggested strategy will greatly decrease membrane fouling thus allowing prolonged recirculation of growth medium. The impact of medium recycling on a continuous monoculture of Nannochloropsis sp. cultivated at high density was assessed in Chapter 3. It was observed at two different scales (1.3 L laboratory-scale reactors and a 24 m³ outdoor pilot setup) that it is feasible to cultivate Nannochloropsis sp. with reused, microfiltered and replenished medium. Average yields on light of 0.8 ± 0.2 g·mol-1 PAR and 0.26 ± 0.08 g·mol-1 PAR were obtained at indoor laboratory and outdoor pilot scale setups, respectively. Additionally, nutrient consumption rates were documented in order to guide the replenishment of reused media. It turned out that, due to a disproportionate consumption of nutrients, the replenishment strategy had to be adjusted to limit accumulation or depletion of nutrients in the medium upon repeated medium recycling. The N/P feed concentration was therefore altered from 37/1.3 to 24.7/1.75 (mM/mM). For each recycle round, an important decrease in nutrient (56%), salt (66%) and water (80%) consumption was obtained. Unfiltered centrate too was applied successfully as reused growth medium. However, if one wishes to prevent unwanted contamination in the production of multiple monocultures on a large scale facility, the use of this unfiltered media source is, while cost-efficient, not recommended. When multiple monocultures are cultivated in a large-scale installation, mixing of filtered harvest broths sourced from these cultures will occur. While reusing this medium, allelopathic effects may appear and induce favourable growth responses for some genera but not others. In Chapter 4, the potential of reusing medium from two algae species (Nannochloropsis sp. and Tisochrysis lutea) for growing monocultures of the opposite algal strain was evaluated at three successive cultivation setups with increasing volume; 400 mL in turbidostat mode, 2.6 L and 220 L in semi-continuous mode. Whereas small scale tests were principally used to detect major differences in growth characteristics when using recycled media, pilot scale experiments provided important information on nutrient consumption and generated productivity data that can be extrapolated to large scale cultivation systems. No contamination or negative effect on productivity occurred during cultivation of either species with reused, microfiltered and replenished medium derived from the other species. Microfiltration and balanced nutrient supplementation of the recycled medium contributed to this result. Average yields on light of 1.6 ± 0.5 and 0.6 ± 0.2 g·mol-1 PAR were obtained for Nannochloropsis sp. and T. lutea respectively. By reusing medium, a significant reduction in water footprint (77%) and salt expenditure (71%) was obtained. At pilot scale, a total nutrient cost reduction of 68% was achieved. In conclusion, the results of this thesis led to the implementation of a medium recycling method in a fully automated production system of microalgae, which in turn contributes to the economic viability of a large-scale cultivation of microalgae. Overall, compared to no medium recirculation, a reduction of 67.5% in medium cost and 34.0% in total variable production cost, or € 7.3 kg-1algae, is accomplished when 80% of the medium is reused on a 0.38 ha, 108 m³ microalgae cultivation plant.

Microalgae as a source of biodiesel precursor are promising by pointing out several advantages of its cultivation, such as able to be cultivated in non-arable land, high growth rate with high biomass yield, high lipid content, and renewable. Mass cultivation of microalgae requires large amount of nutrients which makes it economically impractical. To overcome this problem, an alternative medium which contains cheaper nutrients sources should be addressed. In this research, the growth and biomass productivity of Navicula sp. and Nannochloropsis sp. were compared in the commonly used F/2 medium and modified medium containing commercial NPK fertilizer. The results indicated that Navicula sp. and Nannochloropsis sp. can thrive in modified F/2-NPK medium under continuous illumination, while NPK-only medium didn’t show any significant increase in growth and biomass accumulation for both strains compared to initial cell inoculation. Cell optical density at 750 nm and biomass dry weight of 80% F/2 and 50% F/2 medium were comparable to that in F/2 control medium for both strains, indicating that Navicula sp. and Nannochloropsis sp. have a similar requirement for nutrients types. Furthermore, higher specific growth rate of Nannochloropsis sp. than Navicula sp. seen in both modified medium showed its favorable condition for growth.

One of the factors influencing the growth of Nannochloropsis sp. is the composition of culture media. The addition of growth regulators in the form of auxins and cytokines in culture media can increase the growth of microalgae. This study aims to examine the effect of IAA (auxin) and BAP (cytokinin) with various concentrations on biomass, chlorophyll content and carbohydrate content of Nannochloropsis sp. culture. Nannochloropsis sp. culture was treated with IAA and BAP in concentration variations each consisting of 0, 0.1, 1, 10 ppm with 3 replications. Data were analyzed with two way ANOVA at 95% confidence level and Tukey follow-up test. The results showed that the combination treatment of IAA and BAP did not affect the chlorophyll-a content of Nannochloropsis sp. culture, but it affected the biomass with the highest P16 (I10B10) of 3.65 g/L and carbohydrate content with the highest content in P4 (I10B0) of 0.30 mg/L. The highest chlorophyll-a content was found in P15(I1B10) of 5.574 mg/L, increased by 6 % compared to controls. Whereas the lowest chlorophyll-a content was found in P12(I10B1) of 1.563 mg/L, decreased by 70 % compared to the control.

Coral trout, Plectropomus leopardus (Lacepède, 1802)is a visual feeder. Turbidity caused by phytoplankton or clay particle in the water will affect the visual foraging of coral trout larvae. Addition of Nannochloropsis sp. has been included in standard operational procedure for marine fish larval rearing as green water. However, the density of Nannochlorposis sp. in coral trout larval rearing system has not been evaluated. This study aimed to evaluate the optimal of Nannochloropsis sp. required for rearing of coral trout larvae. Nannochloropsis sp. was given to two days old larvae (D-2), with the densities of 2 x 105, 4 x 105, and 6 x 105cell/mL. After 50 days rearing period (D-50), evaluation on the average size and total harvest were recorded. The results showed that the density of 2 x 105 cell/mL Nannochloropsis sp. was the best in survival rate (2.35 ± 1.05%) than other densities, but they were not significantly different (P>0.05) than those of 4 x 105 cell/mL (1.67 ± 0.70%) and 6 x 105 cell/mL (1.26 ± 1.05%). The lower densities, 2 x 105 and 4 x 105cell/mL, were dominated by more than 50% of > 2.7 cm sized juvenile. Histological analysis of fish eyes supported that the two lower densities produced dominant cone shape as the receptor cells in the retina observed. From an economical aspect, addition of 2 x 105 cells/mL resulted on the higher profit, hence optimum density of Nannochloropsis sp. added in coral trout larval rearing on a mass scale was 2 x 105 cells/mL.

In this paper we describe our approach to solving t he problem of the simultaneous trans -esterification of lipids (that require a basic catalyst) and esterifi cation of FFAs (that require an acid catalyst), usi ng mixed oxides ad hoc prepared. New mixed oxides based on calcium, cerium and aluminium have been used as catalysts in the reaction of trans -esterification of oils of different composition an d quality, namely: i) extra virgin olive oil as a test case (low acidity) and ii) non-edible oil (high acidity), such as tha t extracted from aquatic biomass ( Nannochloropsis sp microalgae). Among the several catalysts used, 12C aO 7Al 2O3 7CeO 2 has shown unprecedented properties in terms of act ivity and resistance in the simultaneous trans esterification/esterification.