Dr. Fransisco Gabriel Acien Fernandez, Associate Professor, University of Almeria

1. This project has received funding from the European Union’s Horizon 2020 Research and Innovation program
under the Grant Agreement No. 727874
Prof. F. Gabriel Acien
Dpt. Chemical Engineering, University of Almeria, SPAIN
Sustainable integrated Algae Biorefinery for the
production of bioactive compounds for Agriculture
and Aquaculture (SABANA)

2. Description
Sustainable Algae Biorefinery for Agriculture aNd Aquaculture (SABANA)
Call H2020-BG-2016-2017
Blue Growth - Demonstrating an ocean of opportunities
Organism European Commission
Topic/Type/Budget BG-01-2016, Innovation action, 10,5 M€
Duration (months) 48
Fixed keywords Bioproducts (products that are manufactured using biological materials
and methods)
Free Keywords Biorefinery, Microalgae, Marine water, Large Scale, Biopesticides,
Biostimulants, Aquafeed, Wastewater

3. WATER
Feed additives, aquafeed
biostimulants, biopesticides
Nitrogen
Phosphorus
Objective
• Large scale production: Develop robust and scalable microalgae production and processing
processes, in continuous mode all the year around. Economic analysis limiting technologies.
• Sustainable production: To integrate the treatment of wastes to increase the sustainability of
the entire process. Life Cycle Analysis determines what is possible or not.
• Markets/commercialization: Only products now requested by the markets and that legally
accepted are considered. Business plan is the driver of the project.

4. Block diagram of the project

5. Suitable wastewater
Effluent COD/BOD,
mg/l
N,
mg/l
P,
mg/l
Total Suspended
Solids, mg/l
Turbidity,
NTU
Biomass
production, kg/m3
Digestate 9000/7000 8000 400 10000 30000 100.0
Manure 16000/12000 9000 500 3000 9000 112.5
Agro-industrial
(Breweries)
4000/3800 30 10 1000 3000 0.4
Centrate 300/200 500 12 1000 3000 6.3
Sewage 700/500 65 11 300 900 0.8
Microalgae
culture medium
50 10 0 0
• The composition of sewage and centrate are similar to standard microalgae culture
media, but digestate and manure contain too much contaminants
• Microalgae can be produced using whatever of these wastewater as only nutrients
source, recovering up 90% of nutrients inlet

6. After 24 months of activities…

7. UNIVERSITY OF ALMERIA
DEMO FACILITY
DEMO1 Facility
Recent achievements

8. R&D Facility
Recent achievements
Raceway reactors
Thin-layer cascade
Tubular reactors
Large scale reactors
Auxiliary facilities
• Air, flue gas
• Culture medium
• Harvesting
• Spry-dryer
• Biomass processing

9. DEMO1 Industrial facility (1.5 ha)
Recent achievements

10. Fully automatized production facility
Control and data acquisition systems already completed, but continuously improved…
Recent achievements

11. Fully automatized production facility
Control and data acquisition systems already completed, but continuously improved…
Recent achievements

12. Fully automatized production facility
Control and data acquisition systems already completed, but continuously improved…
Recent achievements

13. Fully automatized production facility
Control and data acquisition systems already completed, but continuously improved…
Recent achievements

14. Fully automatized production facility
Control and data acquisition systems already completed, but continuously improved…
Recent achievements

15. Fully automatized production facility
Control and data acquisition systems already completed, but continuously improved…
Recent achievements

16. Microalgae production cost
-
0,5
1,0
1,5
2,0
2,5
3,0
RW+fertilizers RW-sewage RW-centrate RW-manure
Productioncost,€/kg
Raceway
0,0
0,5
1,0
1,5
2,0
2,5
3,0
TL+fertilizers TL-sewage TL-centrate TL-manure
Productioncost,€kg
Thin-layer
• Production cost below 2 €/kg is possible only when using nutrients from
wastewaters
• Production cost is lower when using Thin-Layer reactors due to the higher
productivity on these systems
Recent achievements

17. Optimal harvesting strategy
Recent achievements
• Pre-concentration step is mandatory
• A final dewatering step is required to
achieve final concentration of 100 g/L
• Robust large scale processing is not
easy…
Cost
aprox(€/kg)
Energy
(kWh/m3)
0.30 1.00
0.05 0.13
0.07 0.15
0.03 0.11
Culture
1 g/L
Sludge
10 g/L
Paste
100 g/L
Option 1 Centrifugation
Option 2 Sedimentation Centrifugation
Option 3 Dissolved air flotation Centrifugation
Option 4
Non-pressure
membrane
Centrifugation

18. WET
BIOMASS
CELL
DISRUPTION
ENZYMATIC
HYDROLISIS
LOW VALUE
BIOFERTILIZER
LIQUID
EXTRACTION
CELL
DISRUPTION
HIGH VALUE
BIOSTIMULANTS
HIGH VALUE
BIOPESTICIDES
WET
BIOMASS
CELL
DISRUPTION
STABILIZATION
PROCESS
LOW VALUE
AQUAFEED
LIQUID
EXTRACTION
CELL
DISRUPTION
BIOACTIVE
CONCENTRATES
AQUACULTURE PRODUCTS
Processing strategies
AGRICULTURE PRODUCTS
Recent achievements

19.  Effectiveness of cell disruption method strongly depends on strain/biomass
Scenedesmus are more robust against cell disruption than Spirulina
 Energy consumption of ultrasound treatment is excessive
 Among conventional methods: HPH exhibits highest energy-efficiency
 Energy demand for PEF treatment and HPH is in the same range
Energy content of the biomass (20 MJ/kgdw)
Cell disruption
HPH US
HPH US
PEF treatment energy range
dw
dw
Recent achievements

20. Optimal harvesting strategy
Recent achievements
• PRODUCTION scale:
• Design, Engineering and Planning of GEA Separator SDA 40 + Homogenizer Ariete
NS 3030H – Skid Unit

21. Microalgae strains: selection
Agriculture uses…
Collection Strains Biostimulants Biopesticides Aquaculture Selected
SZE 21 freshwater green microalgae
24 freshwater cyanobacteria
10
5
5
5
5
0
3
2
BEA 10 seawater green microalgae
10 seawater cyanobacteria
5
3
2
3
8
2
3
2
Biostimulant effect on watercress seed germination
Control
Antagonistic effect against phytopatogens
Control

22. 0 10 20 30 40 50 60 70
SABANA-1
SABANA-2
SABANA-3
SABANA-4
SABANA-5
SABANA-6
SABANA-7
SABANA-8
SABANA-9
SABANA-10
SABANA-11
SABANA-12
Xanthomonas campestris
Clavibacter michiganensis
Rhizoctonia solani
Phytophthora capsici
Fusarium oxysporum
Pythium ultimun
Biostimulant effect Biopesticide effect
Microalgae strains: in vitro trials
Agriculture uses…
Strains

23. Biostimulation In vivo: Application of algal extracts (Cucumber, Melon, others..)
T10 days
26 ºC
First treatment (25
mL / plant)
T20 days
26 ºC
Second treatment (25
mL/plant)
T40 days
26 ºC Stem length
Root length
Leaf number
Fresh-Weight
Dry-Weight
0
2
4
6
8
10
12
14
16
18
20
MACC-612-05 MACC-612-01 MACC-683-05 MACC-683-01 MACC-430-05 MACC-430-01 Control
Tallo (cm) Raíz (cm) Longitud (cm) Ratio R/T
SAB-M683-01 SAB-M430-01 ControlSAB-M430-05SAB-M683-05SAB-M612-05 SAB-M612-01
Stem (cm) Root (cm) Length (cm) Ratio root/stem
Microalgae strains: in vivo trials
Agriculture uses…

24. Microalgae strains: field trials
Agriculture uses…
Regular High Premium
Biostimulants €/L 5 10 20
Biopesticides €/L 10 15 25
Major requirements:
• No large volumes requested, medium size facilities
• Enhanced biomass containing target compounds
• Demonstrate the bioactivity in real field conditions
• Safety and sustainability of produced biomass

25. Aquaculture uses…
Cytotoxicity
microalgae
Scenedesmus (T.10 0% I) mg ml
-1
0 2 4 6 8 10 12
%Survival(SAF-1)
70
80
90
100
110
120
130
140
150
Scenedesmus (1-100% I) mg ml
-1
0 2 4 6 8 10 12
%Survival(SAF-1)
40
60
80
100
120
140
160
180
200
Scenedesmus (4-25% I) mg ml
-1
0 2 4 6 8 10 12
%Survival(SAF-1)
60
80
100
120
140
160
180
percentage of cell viability compared with a control without microalgae.

26. COM CT TISO NAN SCE p
Initial body weight (g) 11.4±0.26 11.4±0.20 11.4±0.43 11.4±0.12 11.4±0.39 0.9984
Final body weight (g) 29.8±2.77a 47.9±3.62b 47.7±4.95b 45.2±0.28b 45.7±4.87b 0.0006
Daily gain (DG, mg day-1) 0.22±0.03a 0.43±0.04b 0.43±0.05b 0.40±0.01b 0.40±0.05b 0.0003
Specific growth rate, SGR (%) 1.12±0.09a 1.69±0.08b 1.68±0.08b 1.63±0.01b 1.63±0.08b <0.0001
Feed efficiency ratio (FER) 0.52±0.05a 0.81±0.06b 0.78±0.04b 0.77±0.01b 0.80±0.05b 0.0001
Feed conversion ratio (FCR) 1.92±0.18b 1.24±0.08a 1.28±0.06a 1.30±0.02a 1.26±0.08a <0.0001
Protein efficiency ratio (PER) 0.95±0.09a 1.47±0.10b 1.42±0.07b 1.40±0.02b 1.45±0.09b 0.0001
Survival (%) 96.8±2.75 88.9±5.50 90.5±0.01 92.1±5.50 95.2±0.01 0.1129
Values are mean ± SD of triplicate determination. Values in the same row with different lowercase letter indicate significant difference (p < 0.05)
1.- Senegalese sole juveniles
2.- Seabream juveniles
3.- Seabream larvae
Microvilli length (μm) Microvilli diameter (μm) Number of microvilli
μm2
Total absorption surface per
microvilli (μm2)
CT 1.38 ± 0.17 a 0.10 ± 0.01 61.93 ± 12.12 a 28.80 ± 3.25 a
TISO 1.57 ± 0.16 b 0.10 ± 0.01 76.60 ± 10.17 b 39.14 ± 3.44 b
NAN 1.99 ± 0.25 c 0.10 ± 0.01 70.57 ± 9.80 ab 45.93 ± 3.68 c
SCE 1.35 ± 0.25 a 0.10 ± 0.01 66.78 ± 15.66 a 26.30 ± 10.41 a
p <0.0001 0.0616 <0.0001 <0.0001
Aquaculture uses…
In vivo feeding trials

27. Ultraestructural study
Transmision electron microscopy (TEM) images
CTCOM ISO NAN SCE
None of the dietary treatments cause damage in the brush border integrity of intestinal mucosa
ISO and NAN-fed fish showed significant
increase in microvilii length and microvilli
absorption surface compared to fish fed
microalgae-free diets
Aquaculture uses…
In vivo feeding trials
Microalgae biomass must not be considered to replace nutrients
for aquafeed, but as feed additive to enhance the “health” of
fishes and “safety” of production

28. Dissemination
Training-Center: International Courses
International Summer Course 2017 Almeria (Spain)
International Summer Course 2018 Almeria (Spain)
8th Symposium on Microalgae and Seaweed Products on Plant/Soil Systems 2017 Mosonmagyarovar (Hungary)
International training course GAP’17 held in Trebon (Check Republic)
EUALGAE Training Course 2017 Almeria (Spain)
Symposium Biotechnology of microalgae 2017 Guadalajara (Mexico)
Training course Biotechnology of microalgae 2017 La Paz (México)
Workshop Microalgae production for aquaculture uses 2017 Coquimbo (Chile)

29. Dissemination
Training-Center: students and researchersName DNI/Passport Entity Start date
Duration
months
Comments
Daissy Lorena Repreto 1053838288
Universidad Nacional de Colombia
sede Manizales
September 2017 2 Company contract
Mirka Rojas 115920712 Instituto Tecnológico de Costa Rica December 2016 10
Colombian National
Grant
Luis Alfredo Barboza Fallas 115950399 Instituto Tecnológico de Costa Rica December 2016 5 Company contract
Maria José Salar García 48453090W
Universidad Politécnica de
Cartagena, España
December 2016 4
Spanish National
Grant
Victor Manuel Ortíz Martínez 77712047S
Universidad Politécnica de
Cartagena, España
December 2016 4
Spanish National
Grant
Ana Isabel Guzmán Carrasco 77153485P Univ. Almeria (Spain) January 2017 6 Company contract
Claudia Sepúlveda Vega 13419422-7
Universidad de Antofagasta en
Chile
January 2017 18
Chilean National
Grant
Isaac Lisondro 1854741
Universidad Autónoma de Chiriquí,
Panamá
January 2017 4
Panama National
Grant
Jose Raul Roman Fernandez 76630378J Univ. Almeria (Spain) January 2017 6
Spanish National
Grant
Ainoa Moilla España 41527775X Univ. Almeria (Spain) February 2017 6 Company contract
Beatriz Roncero Ramos 44373274H Univ. Almeria (Spain) February 2017 12
Spanish National
Grant
Beatrice Peter 322344221
University on Insubria in Varese
Italy
March 2017 9 Erasmus+ grant
Blas Manuel Franco Ortellado Y3090374-Q Universidad de Valladolid March 2017 6
Spanish National
Grant
Jazmin Bazaes Donoso 15021083-6 Universidad de Antofagasta, Chile March 2017 3 Company contract
Maria Cristina Purice X6445513Q Univ. Almeria (Spain) March 2017 4
Spanish National
Grant
Mario Alberto Lopez Alonso 75729818H Univ. Almeria (Spain) March 2017 4
Spanish National
Grant
Victor Pizarro Quezada P10754103 Universidad de Antofagasta, Chile March 2017 3 Company contract
Begoña Sanchez Astrain 16640657-L Univ. Almeria (Spain) April 2017 3 Master practices
Cristian Inostroza Gonzalez 16062968-1
Insituto de Transferencia
Tecnologica y Emprendimiento,
Chile
April 2017 3 Company contract
Darío Ruiz Barbudo 17473057-A Univ. Almeria (Spain) April 2017 3 Master practices
Gabriel Martins da Rosa P10754103
Universidad Federal do Rio Grande,
Brasil
April 2017 8
Brazilian National
Grant
Luiza Moraes P17412536
Universidad Federal do Rio Grande,
Brasil
April 2017 8
Brazilian National
Grant
Marlon Gonzalo Carvajal Proaño 1721161055 Univ. Almeria (Spain) April 2017 3 Master practices
Ana Sánchez Zurano 75536955N Univ. Almeria (Spain) June 2017 24
Spanish National
Grant
Carla Cristina Mestre Verissimo Dias 13554073 9ZX2
Laboratorio Nacional de Energia e
Geologia
June 2017 3
Portuguese National
Grant
Alessandro Solimeno Y2440644J Univ. Barcelona July 2017 6 Company contract
Sahar Akaberi Y09K0YT0H Karlsruhe Institute of Technology July 2017 2 Company contract
Victor Alonso 26819754Y Universidad de Malaga August 2017 6 months Master practices
EL BAHRAOUI Naoufel 328909730 SETEC (France) September 2017 2 months Company contract
Davide Veronesi AY1919319 Universidad de Milan, Italia November 2017 6 months Company contract
Klaus Leber 643220342 Karlsruhe Institut Technologu November 2017 1 month Company contract
Alejandro Bueso Sánchez 76663908D Univ. Almeria (Spain) January 2018 6 months Company contract
Joyce Gloria Villachica Llamosas 7078441 Smallvill S.A.C. January 2018 3 months Company contract
Leslye Karla Villachica Llamosas 7078206 Smallvill S.A.C. January 2018 3 months Company contract
Mariana Jimenez Veuthey AAB993712
Universidad Nacional de Entre Ríos,
Argentina
January 2018 1 month
Argentina National
Grant
Pablo Martín Valencia 44732608T Univ. Almeria (Spain) March 2018 4 months Master Practices

30. Dissemination
Training Center: Massive Online Open Course
https://www.youtube.com/watch?v
=-eg_TyaMUNY

31. Dissemination
Data Center

32. Acknowledgements…
EU Commission…
Partners…
Our team and collaborators…

33. SABANASustainable Algae Biorefinery for Agriculture aNd Aquaculture
www.eu-sabana.eu | info@sabana.eu @sabana.eu
sabana.eu
sabana.eu