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.
