Periodic Reporting for period 3 - InnCoCells (Innovative high-value cosmetic products from plants and plant cells)
Période du rapport: 2024-05-01 au 2025-09-30
InnCoCells aimed to develop innovative and sustainable plant-based production processes for the exploitation of scientifically validated cosmetic ingredients based on underutilised plant resources. We aimed to optimise these resources for production using cell and hairy root cultures, aeroponics and greenhouse/field cultivation. We also applied systematic approaches such as metabolic engineering to optimise growth conditions and the yields of valuable bioactive compounds. The optimised processes were demonstrated by pilot-scale production and downstream processing. We aimed to bring at least 10 cosmetic ingredients to the pre-commercial stage. InnCoCells included a cascade biorefinery concept in which by-products and biowaste from the agri-food industry were utilised to extract further bioactive molecules. The production processes were characterised by techno-economic analysis and life cycle assessment to ensure economic feasibility and a small environmental footprint. The ingredients and extracts were evaluated using a unique panel of innovative enzymatic, metabolomic and cell-based in vitro and ex vivo assays to ensure their safety and validate the efficacy based on robust scientific data without animal testing. We implemented a unique stakeholder engagement strategy, including a Stakeholder Group to guide our research based on the needs of industry, academia, farmers, policymakers and consumers. This industry-driven and interdisciplinary project will ultimately increase the strength of the European bioeconomy by supporting the development of innovative biobased cosmetics.
We selected a portfolio of almost 100 plant species early in the project, some already available to the partners as cell suspension cultures, hairy roots or whole plants growing on aeroponic surfaces or in greenhouse/field plots, and others via our bioprospecting program. Protocols for in vitro cultivation, the use of elicitors and genetic engineering tools, developed for model species, were adapted for the InnCoCells species portfolio to optimise biomass production and metabolite yields.
Cell suspension and hairy root cultures were first tested at laboratory scale (up to 20 L) in darkness or with illumination in multiple bioreactor systems. Cryopreservation protocols were also developed. Growth and the accumulation of bioactive compounds were optimised, allowing five cell lines to be scaled up to 200–300 L with yields > 10 g DW/L and one to 1000 L. Having established a robust seed train, one hairy root line was also scaled up to 1000 L. Two of four species were scaled up to 100 m2 aeroponic surfaces and three species were grown in large-scale field plots.
For the extraction of ingredients from plant biomass, including a range of agri-food by-products, we studied different drying and milling methods, pre-treatment with enzymes and pulsed electric fields, and a range of different hydrophilic, lipophilic and supercritical CO2 solvents to improve extraction efficiency and stability. These methods were optimised at the pilot and industrial scales to match upstream production. A spiral filter press was suitable for the extraction of pilot-scale cell cultures but process parameters and yields were species-dependent. Standard operating procedures for sample extraction, stability testing and bioactivity assays were compiled. We used targeted and untargeted metabolomics to determine the chemical composition of bioactive extracts. More than 100 extracts were tested for safety (cytotoxicity and phototoxicity) and their antioxidant, antimicrobial, anti-inflammatory and anti-aging properties in vitro. We retested the top 26 extracts in vitro for anti-aging and anti-inflammatory activities, and then confirmed their efficacy in seven different ex vivo human skin models to gain insight into the mode of action. The ex vivo assays were selected to match consumer demands for safe ingredients with efficacy against acne, inflammation, ageing, sun damage and dehydration.
We compiled the safety, toxicology, chemical composition and bioactivity data for 11 of our most promising ingredients. Bioactivity data from scientific and technical dossiers were used to substantiate product efficacy and safety claims, and to develop a market positioning and commercial documentation for more than 10 ingredients. Techno-economic evaluation and life cycle assessment were carried out to confirm economic viability and sustainability for three process/product combinations. Thus far, eight of our ingredients have already reached commercial development.
We set out the project’s strategy for dissemination, communication and knowledge/IP management and implemented a broad range of activities (project website, social media, project brochures, promotional videos, podcast series, press releases, newsletters and articles in industry periodicals, and collaboration with the three other projects funded under the same H2020 topic). We have published > 10 scientific articles so far (~20 more submitted or in preparation) and have presented talks or posters at > 50 conferences. We have raised awareness of the project at > 10 high-profile cosmetic industry exhibitions, hosted nine webinars, and organised a training workshop on plant cell cultures. Our Stakeholder Group has attended four project meetings and helped with the selection of promising cosmetic ingredients as well as amplifying our dissemination and communication program and our commercialisation activities. Four patents were submitted during the project, with more to follow.
Cell suspension and hairy root cultures were first tested at laboratory scale (up to 20 L) in darkness or with illumination in multiple bioreactor systems. Cryopreservation protocols were also developed. Growth and the accumulation of bioactive compounds were optimised, allowing five cell lines to be scaled up to 200–300 L with yields > 10 g DW/L and one to 1000 L. Having established a robust seed train, one hairy root line was also scaled up to 1000 L. Two of four species were scaled up to 100 m2 aeroponic surfaces and three species were grown in large-scale field plots.
For the extraction of ingredients from plant biomass, including a range of agri-food by-products, we studied different drying and milling methods, pre-treatment with enzymes and pulsed electric fields, and a range of different hydrophilic, lipophilic and supercritical CO2 solvents to improve extraction efficiency and stability. These methods were optimised at the pilot and industrial scales to match upstream production. A spiral filter press was suitable for the extraction of pilot-scale cell cultures but process parameters and yields were species-dependent. Standard operating procedures for sample extraction, stability testing and bioactivity assays were compiled. We used targeted and untargeted metabolomics to determine the chemical composition of bioactive extracts. More than 100 extracts were tested for safety (cytotoxicity and phototoxicity) and their antioxidant, antimicrobial, anti-inflammatory and anti-aging properties in vitro. We retested the top 26 extracts in vitro for anti-aging and anti-inflammatory activities, and then confirmed their efficacy in seven different ex vivo human skin models to gain insight into the mode of action. The ex vivo assays were selected to match consumer demands for safe ingredients with efficacy against acne, inflammation, ageing, sun damage and dehydration.
We compiled the safety, toxicology, chemical composition and bioactivity data for 11 of our most promising ingredients. Bioactivity data from scientific and technical dossiers were used to substantiate product efficacy and safety claims, and to develop a market positioning and commercial documentation for more than 10 ingredients. Techno-economic evaluation and life cycle assessment were carried out to confirm economic viability and sustainability for three process/product combinations. Thus far, eight of our ingredients have already reached commercial development.
We set out the project’s strategy for dissemination, communication and knowledge/IP management and implemented a broad range of activities (project website, social media, project brochures, promotional videos, podcast series, press releases, newsletters and articles in industry periodicals, and collaboration with the three other projects funded under the same H2020 topic). We have published > 10 scientific articles so far (~20 more submitted or in preparation) and have presented talks or posters at > 50 conferences. We have raised awareness of the project at > 10 high-profile cosmetic industry exhibitions, hosted nine webinars, and organised a training workshop on plant cell cultures. Our Stakeholder Group has attended four project meetings and helped with the selection of promising cosmetic ingredients as well as amplifying our dissemination and communication program and our commercialisation activities. Four patents were submitted during the project, with more to follow.
We aimed to move beyond the state of the art by addressing four major bottlenecks: (1) target compounds tend to accumulate in plants at low and variable concentrations; (2) compounds can only be isolated from extremely rare or endangered plants, which are poorly characterised and unsuitable for large-scale production; (3) bioactive compounds cannot be produced economically on a scale that meets demand; and (4) the discovery of bioactive compounds in plants is not based on systematic scientific analysis. These bottlenecks were overcome by implementing systematic approaches to establish and scale up cell lines/hairy roots/plants and corresponding extraction and bioactivity screening processes. The expected results were: (1) sustainable exploitation of at least 10 relevant metabolic pathways in plant species; (2) a multi-step evaluation pipeline yielding at least 50 scientifically verified active ingredients; (3) optimised production processes and technologies for at least 20 ingredients; (4) evaluation of at least 10 agri-food by-products/waste fractions to generate added value; and (5) pilot-scale production of at least 10 active ingredients, along with economic and sustainability evaluation. All these aims were exceeded during the project. The major direct scientific and technological impacts will include novel, natural, sustainable and eco-friendly cultivation and processing methods leading to products with clear scientifically proven bioactive properties as well as the full exploitation of waste streams in a cascade biorefinery approach. The longer-term societal and socioeconomic impacts will include more public–private cooperation in the European biotechnology industry, greater public knowledge of biodiversity, ecosystems and the potential for sustainable exploitation, as well as new market opportunities in the European cosmetics industry.