Downloaded from orbit.dtu.dk on: Oct 09, 2021 Micro -algae biomass as an alternative resource for fishmeal and fish oil in the production of fish feed Safafar, Hamed Publication date: 2017 Document Version Publisher's PDF, also known as Version of record Link back to DTU Orbit Citation (APA): Safafar, H. (2017). Micro -algae biomass as an alternative resource for fishmeal and fish oil in the production of fish feed. National Food Institute, Technical University of Denmark. General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights. Users may download and print one copy of any publication from the public portal for the purpose of private study or research. 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Micro -algae biomass as an alternative resource for fishmeal and fish oil in the production of fish feed PhD Thesis Hamed Safafar 2017 Micro -algae biomass as an alternative resource for fishmeal and fish oil in the production of fish feed PhD Thesis by Hamed Safafar National Food Institute Technical University of Denmark November 2016 Micro -algae biomass as an alternative resource for fishmeal and fish oil in the production of fish feed Copyright: National Food Institute, Technical University of Denmark Photo: Hamed Safafar ISBN: 978-87-93565-02-9 This PhD Thesis is available at www.food.dtu.dk National Food Institute Technical University of Denmark Mørkhøj Bygade 19 DK-2860 Søborg Tel: +45 35 88 70 00 Fax: +45 35 88 70 01 PREFACE This PhD project entitled as “Micro -algae biomass as an alternative resource for fishmeal and fish oil in the production of fish feed” was carried out at the National Food Institute, Technical University of Denmark, under the supervision of Professor Charlotte Jacobsen as main supervisor (DTU-Food) and Dr. Per Møller as co-supervisor (Department of development, Kalundborg commune). The project was started in November 2013 and ended in November 2016. This project was a part of a big project “Development of filtering technologies for microalgae and sustainable, high-quality feed for fry, (FIMAFY)” funded by Grønt Udviklings og Demonstrations Program (GUDP). The project partners include DTU-Aqua, BioMar A/S, LiqTech International A/S, IFAU and Ecolipids. The PhD project aimed at developing new technologies for downstream processing of microalgae as a fish feed ingredient. The PhD project screened and selected the proper microalgae species, characterised the biochemical composition of algal biomass during the cultivation, processing and the storage and finally developed new technologies for downstream processing of microalgal biomass. During the project, several algae-specific analytical methods were developed. Moreover, a microalgae cultivation laboratory including pilot scale downstream processing facilities (harvest, up- concentration, pasteurisation and drying) was established at DTU Food. I am deeply proud to see that young researchers are employing these methods and facilities for their research. November 15, 2016 Kongens Lyngby, Denmark Hamed Safafar i ACKNOWLEDGEMENT After an intensive period of three years, today is the day: writing this note of thanks is the finishing touch on my thesis. First and foremost I wish to express my deepest gratitude to my supervisor Professor Charlotte Jacobsen for her continuous support, patience and motivation during the project. Your guidance helped me in all the time of research and writing of this thesis. I am heartily thankful to you. My sincere thanks also goes to Per who was a supportive co-supervisor. I learned a lot from both of you. My thanks to all of colleagues and friends who are behind this work: To Professor Odilio Alves-Filho for his scientific supports during my stay in NTNU, Norway. To Dr. Susan Holdt for her scientific supports during the project. To our amazing laboratory technicians: Inge, Lis, Trang, Rene and Heidi who are incredibly helpful. In particular, I am grateful to Inge: without your precious helps it would not be possible to conduct this research. To Klaus, who provided the best of his technical expertise that greatly assisted me in every part of this research. To Birgitte for sharing the office these years and being such a good friend. Nina, Alireza, Pedro ,Urd, Ann-Doritt, Ditte and Betul, I had great times with you and will always remember you. To Jørgen, Ivar, Karin and Haris, members of FIMAFY steering group, Micahel from Kalundborg microalgae facility, Jon and Davide from DTU-ENV for their collaborations. My deepest thanks to all the amazing and talented students I have the chance to work with: Jenia, Patrick, Anita, Anja and Simon, you did an incredibly great job. My beloved family Maryam and Arian deserve a particular note of thanks. Maryam, your supports and patience during these years helped me to finish this project as it is now. Final thanks to my parents, for all of the sacrifices that you’ve made on my behalf. Your prayer for me was what sustained me thus far. Hamed Safafar ii Summary In recent years, intense efforts have been made to find new, alternate and sustainable aquatic feed ingredients, primarily in anticipation of an increasing world population and predicted insufficient fishmeal supply which is a critical component of aquaculture feed. Now it is becoming increasingly evident that the continued exploitation of industrial fish as a resource fish feed will ultimately become both environmentally and economically unsustainable. Microalgae are at the base of the entire aquatic food chain and play a major role in the diet of aquatic animals such as fish. Microalgae’s main application for aquaculture are related to nutrition, being used as a sole fresh feed or an additive, e.g. source of pigment. Algae produce almost all nutritious compounds which are required for fish. The diverse biochemical composition of microalgae represents them as a promising candidate for the formulation of fish feed. The nutritional composition of microalgae depends on the species, environmental conditions and growth medium composition. Microalgae for use in aquaculture should be non-toxic and possess the essential nutritive constituents, in a reasonable price. Photosynthetic production of algae either in outdoor or indoor photobioreactor systems is costly since cultures must be maintained at low densities. Consequently, large volumes of media must be processed to recover small quantities of algae, and since most algal cells are minuscule, unspecific expensive harvesting processes must be employed. Strategies such as cultivation of microalgae on low price growth media, selection of microalgae capable of growing on such media and produce biomass with desired chemical composition and development of specific harvest and downstream processing represent basic solutions to improve the applicability of microalgae biomass as a fish feed ingredient. Moreover, storage of the algae biomass at optimum conditions minimise the deterioration of valuable compounds. This project has employed the strategies mentioned above to provide a clear concept for the cultivation, processing and the storage of microalgae biomass intended to be used as a fish feed ingredient. A pre-gasified industrial process water with high concentration of ammonia and free from toxic compounds, representing effluent from a local biogas plant was used as a low price growth medium. Therefore, the biomass production benefits from low cultivation price and also from valorization of the nutrients. Screening of various microalgae species has been extensively done to find proper microalgae capable of growing on industrial process water and producing a biomass containing high levels of protein, long-chain polyunsaturated fatty acids (LC PUFA), and bioactive compounds such as natural antioxidants. Effects of growth media composition/concentration and cultivation time on the nutritional composition of the biomass, variations in proteins, lipid, fatty acid composition, amino acids, tocopherols, and pigments were evaluated. Among all studied species including Nannochloropsis salina, nannochloropsis limnetica, Chlorella sorokiniana, Chlorella vulgaris, iii Chlorella pyrenoidosa, Desmodesmus sp. and Arthrospira platensis, the microalgae Chlorella pyrenoidosa grew well on the industrial process water, efficiently valorized the compounds in the growth medium (ammonia and phosphorous) and produced reasonable amounts of the biomass (6.1 g/L). The resulting biomass included very high levels of protein (65.2±1.30% DW) as well as promising amino acid and carotenoid compositions. Chlorella pyrenoidosa was selected as a source of proteins and amino acids while lacking LC PUFA’s. The microalgae Nannochloropsis salina which was grown on a mixture of standard growth medium and industrial process water produced a biomass containing high eicosapentaenoic acid (C20:5 n-3, as 44.2% ± 2.30% of total fatty acids), representing a rich source of LC PUFA. Data from laboratory scale experiments were translated to large scale and both of these species have been successfully cultivated