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Combined Ultrasonic and Enzyme treatment of Lignocellulosic Feedstock as Substrate for Sugar Based Biotechnological Applications

Periodic Reporting for period 3 - US4GREENCHEM (Combined Ultrasonic and Enzyme treatment of Lignocellulosic Feedstock as Substrate for Sugar Based Biotechnological Applications)

Reporting period: 2018-07-01 to 2019-06-30

Rising concern over fossil fuels supply and the impacts of GHG have led to high interest in new feedstocks for manufacturing industries. Using biomass as sustainable renewable resource is a way to replace carbon from fossil sources in the manufacture of chemicals, materials and fuels.
Due to the limited arable surface of Europe, the major potential for European biorefineries lies with lignocellulosic substrates derived from agriculture and forestry residues and from energy crops (SRC).

.Develop ultrasound (US) pretreatment that effectively disrupts the lignocellulosic matrix
.Degrade lignocellulose with SC-CO2 technologies to maximize release of sugars
.Optimize yield and reduce the cost of enzymatic hydrolysis
.Test for fermentability of the sugar fraction
.Upscaling for pilot scale
.Life Cycle Assessment for comparison to existing technologies.

Conclusion of the Action: aims above mentioned above were achieved. During lab scale phase, wheat straw showed much more efficient results than poplar wood, which led eventually to the trials of pilot scale with wheat straw and wheat bran, a more similar lignocellulose biomass in characteristics. Life Cycle and cost benefit analysis showed that for all five environmental categories ( raw materials and depletion potential, energy consumption for treatment and losses, land use and risk potential , toxicity of by-products, and emissions) the conversion and resources efficiency the US pre-treatment had advantages in comparison with the standard industrial steam explosion pre-treatment. US4Greenchem pre-treatment has more advantages in its Eco-Efficiency. The values generated in the CBA were also lower.
WP1: Mechanical Disruption
Task 1.1: pre-requisites for cavitation
biomass (hardwood chips and straw) chemical characterization, different milling techniquesperformed. surface characterization showed difference in the pore structure.
Task 1.2: optimization of cavitation parameters
The influence of particle size on the sonication pre-treatment was evaluated.
Task 1.3: impact of cavitation on biomass structure
Ultrasound treated samples surface characterization performed. Further pretreatment with ultrasonication and hydrodynamic cavitation in 10 % sodium hydroxide(UNITO).
WP2: Matrix Disintegration
Task 2.1: CO2 Hemicellulose Hydrolysis
subcritical and supercritical carbon dioxide hydrolysis at a high pressure
Task 2.2: High frequency sonochemical disruption
high-frequency transducer was developed for biomass treatment Whand samples were sonicated at 500 kHz.
Task 2.3: Integration of the proposed technologies
ultrasound probe (40 kHz) installed into existing Sc-CO2 extraction unit CO2 (ScCO2) tests were carried out. A specialized transducer design was needed
Task 2.4: Characterization of the lignin-derived fractions
analysis of solid and liquid samples and characterized lignin-derived fractions
WP3: Biocatalysis Design
Task 3.1: Enzymatic Digestibility studies
Samples tested with enzymatic hydrolysis The best soluble sugar yields reached with ultrasonication in sodium hydroxide.
Task 3.2: New enzymes for improved hydrolysis
Three novel fungal beta-glucosidases used for EH.
Task 3.3: Increase of enzyme yields
For increase, homemade enzyme cocktails from Trichoderma reesei 101 strain were used.
WP4: Sugar Recovery
Task4.1: Optimized Hydrolysis
Efforts focused on search for greener pre-treatment conditions.
Task 4.2: Sugar Recovery
NF allows obtaining two aqueous streams: one enriched in xylose and another in glucose.
Task 4.3: sugar suitability for fermentation
enzymatic hydrolysis liquor is fermentable without further purification.
WP5: Value-added products
Task 5.1: energetic valorization solids
pellets analysed
Task 5.2: lignin derived antioxidants
lignins precipitated revealed an antioxidant activity significantly poorer than that in commercial lignins.
Task 5.3:Polymer building blocks
lignin enriched solids were characterized for application as a component of polymeric composites.
Task 5.4:adhesives and resins
valuable phenolic blocks and good yields of bio-oil, up to 42% of the initial lignocellulosic biomass, were obtained .

WP6:Upscaling and Pilot testing
Task 6.1: Upscaling the pretreatment step
An upscaled modular of two 3 kW ultrasonic reactors Vertical reactor.
Task 6.2: Upscaling the hydrolysis step
Enzyme cocktail developed and produced from shake flasks to fed batch fermentation

Task 6.3 : Upscaling of the extraction and valorisation technologies
two sugar streams enriched in xylose and glucose obtained by countercurrent multistage NF cascade
Task 6.4: Biorefinery Integration
integrated system built in partner’s facilities, VTT and after again at TTZ.

Task 6.5: Pilot scale Testing
Pilot scale testing was performed in March 2019 on wheat straw biomass and pre-treatment process evaluation with wheat bran in May-June 2019.

WP7: Process Evaluation
Task 7.1: Cost-benefit Analysis adaptation
Eco- efficiency approach for lignocellulose decomposition processes developed.

Task 7.2: LCA by LDEE approach
Using UMBERTO NXT LCA, the comparative material flows performed and applied “ecoinvent v3” database

WP8:Dissemination Activities
Task 8.1: Scientific Dissemination performed (publication, conferencea, posters)

Task 8.2: Business Dissemination
Bio Market Insights banner produced. Project video was produced, published press release

WP9: Project Management
General coordination, Administrative project coordination, and IPR & innovation management
VTT has submitted 2 patents application on novel beta-glucosidases in June 2018

Results exploitation and dissemination:
business-oriented activities were performed also in the last 6 months of the project during pilot scale tests. A key focus within the marketing strategy was the demonstration of the process itself and push market penetration forward. Contacts with potential clients (biorefineries, bioethanol plants, bio-plastics producer, pharmaceutical industry, etc.) were facilitated during the project by an introduction of the technology in trade fairs and other promotional events by presenting in scientific conferences and scientific articles and via the Bio Based Industries Network. The project also produced a video. US4greenchem marketing value of technologies developed must be made in demo scale, where large industrial-scale process brings costs down and efficiency of process up.
US4GREENCHEM optimized 3 major steps of a state-of-the-art biorefinery for conversion pathway of lignocellulosic substrates into biofuel and biomaterials, reducing processing costs: physical pre-treatment with ultrasound and supercritical CO2 technologies, a multi-target hydrolysis and valorization of process effluents.
Socio-economic impacts include knowledge creation, increased academia-industry cooperation, creation of new markets, reutilization of local residues and support the regional development, valorization of agricultural residues, reducing dependence on imports of fossil oil, new skilled jobs in rural regions and in product development and engineering.