IVACE - FEDER FIVEX

Objective

The general objective of this project is to study and evaluate the methodologies for the production, isolation and characterisation of the functionality of EVs obtained from different sources for their application as functional ingredients in different industrial sectors.

The project aims to address everything from the identification of new sources of EVs, as well as the production and development of scalable isolation methodologies, through to the design of functional evaluation tests, characterisation and conservation strategies. To achieve the proposed general objective, the following specific objectives are presented:

Scientific-technical review of the main sources and functionalities associated with them, isolation methodologies currently used to obtain EVs,and conservation strategies: Identification and selection of sources and functionalities of EVs, strategies for their purification and conservation. In addition to the literature review, collaboration with companies providing information on these aspects will be key.
Legal framework of EVs as a functional ingredient in the agricultural, food, cosmetics and pharmaceutical sectors: Understanding the legal framework for the commercialisation of EVs as a new ingredient for incorporation into products in the identified sectors.
Standardisation of the scalable EV isolation process, regardless of their origin: Design of a purification system that enables the separation of EVs scalable at industrial level and applicable to EVs from different sources.
Obtaining EVs from different origins to validate the standardised workflow Morphological and functional characterisation of the EVs obtained, considering possible by-products derived from the agri-food industry as a source of EVs.
Development of techniques for the functional evaluation of EVs: Optimised design of more complex evaluation methods for the bioactive potential of the EVs produced and isolated within the framework of the project, selecting those functions demanded by companies in the sectors involved. This will be complemented by proteomic characterisation of the EVs with the greatest potential for industrial application.
Definition of a long-term EV conservation protocol: Application of current conservation strategies, as well as others applied to similar biological structures, to define a conservation protocol that enables their long-term storage without compromising their viability.

Activities

Scientific-technical review of the state of the art of the application of EVs in different sectors according to their origin. Legal framework
Isolation of EVs. Design of a standardised workflow
Production of EVs from different sources of interest.
In vitro evaluation of the functional activity of EVs
Conservation of EVs
Communication and dissemination
Transfer and promotion of results
Management and coordination

Results obtained

During the execution of FIVEX, the bibliographic and legal review was completed, providing a global view of the use of extracellular vesicles (EVs) in different sectors (food, cosmetics, pharma and agriculture) and selecting sources, isolation and conservation techniques applied in the different technical activities within the project.

A standardised workflow was designed for the isolation of EVs, adaptable according to the source, and validated through the obtaining and initial characterisation of EVs (size, number of particles, morphology, biocompatibility and immunomodulatory effect). In the functional evaluation of EVs, ad hoc in vitro protocols were developed for oral and topical applications of EVs, evaluating resistance to gastric digestion and transport through epithelial and dermal barriers, as well as functionalities associated with dermal health in EVs obtained from three selected probiotics.

Finally, progress was made in EV conservation strategies, optimising parameters for spray drying and comparing with reference methods (PBS -80 °C, lyophilisation with trehalose), although pending improvements were identified to ensure industrial processability.