KSP: Bio-farming for bioactive compounds

The revised application for the project «Biofarming for bioactive products» with the short name «bioACTive» was approved by the Research Council on 23 April 2021 and the project had a start-up meeting on 12 May. A post doc employee started work on 30 August at Nofima in Ås. The aim of the project is to increase the value of the Norwegian agricultural sector by producing bioactive plants (phytogens) for feed supplements and bioactive packaging through 1) increasing the shelf life of salmon feed 2) reducing winter sores in farmed fish 3) improving intestinal flora and health in chicken, and 4) to design degradable and antimicrobial packaging materials. NIBIO Apelsvoll and Særheim, which is responsible for work package 1, have focused on selecting, propagating and producing plant material of the species and lines selected in the project. Plants have been propagated and produced in greenhouses, tunnels and outdoors, a total of approx. 90 plant samples. From Artemesia annua both stems and leaves have been harvested, and from Portulaca oleracea both leaves and seeds. The seeds are analyzed according to the results of the leaf analyzes. Several varieties of Origanum vulgare and A. annua are grown under different lighting conditions to investigate whether the content of bioactive substances can be increased with different light effects and A. annua and Humulus lupulus are further grown in tunnels and outdoors to provide guidance to producers about optimization of growing conditions. We have also gained further knowledge about the growth requirements of the selected species and the differences between the various lines. Results from this first phase have been presented at the OneHealth seminar in Oslo on 3 November 2021. The aim of work package 2 is to make plant extracts and, based on bioactivity-guided fractionation (in vitro tests), to isolate compounds that show antioxidant, antimicrobial and anti-inflammatory effects. Extracts will also be characterized by targeted and non-targeted metabolomics. So far 117 plant samples are received from 10 herb species. Each sample (5?15 g dry weight) has been extracted under sonication with dichloromethane (DCM), ethanol and water, successively. In total, 3x117 excerpts have been made. The total extraction yield was determined to be 24 ± 11%. The different solvents contributed to the total yield in the following range: water> ethanol> DCM. The radical removal capacity of each extract was determined. Rosemary revealed the highest capacity among the DCM extracts while oregano gave the strongest effect within both the EtOH and water extracts. Among the herbs, it was found that oregano contains the most phenols. It was found to be a positive correlation between radical scavenging capacity and the content of total phenols with respect to both the ethanol extracts and the water extracts. In work package 3, one of the aims was to evaluate the antimicrobial potential of nine different plant species: Achillea millefolium, Artemisia annua ?Anamed?/?Apollon?, Rosmarinus officinalis, Humulus lupulus, Portulaca oleracea, Mentha piperita, Rhodiola rosea, Origanum vulgare, Leuzea carthamoides. Total of 100 samples from different parts of the plant (flower, leaf, seed, root) have been used to make water, ethanolic (EtOH) and dichloromethane (DCM) extracts. The postdoc within the project has centered work regarding the antimicrobial activity of these 300 extracts. The agar disk diffusion method has been used to determine the antimicrobial activities of the different plant extracts against two indicator organisms: the Gram-positive Staphylococcus aureus, and the Gram-negative bacteria Escherichia coli. The three Atlantic salmon pathogens Moritella viscosa, Tenacibaculum finnmarkense and Aliivibrio wodanis have also been used applying two different types of growth media mimicking effects when bacteria are growing in seawater or simulating nutrient rich host conditions. Water extracts exhibited low activity. The DCM extracts showed the highest antibacterial activity against all the tested bacterial strains. However, none extracts showed antimicrobial activity against all strains. The DCM Rosemary extracts showed activity against all salmon pathogens. Preliminary results indicate that plant extracts can be of great value as natural antimicrobials in aquaculture. The main aim of WP5 is to create an active food packaging system by creating a coating via spraying, electrospinning, etc. of a biopolymer casein containing the selected (the best antioxidant and antimicrobial properties) herb extract on a biobased substrate. To create a coating on the surface of the substrate, surface treatment is required via cold atmospheric plasma. The parameters for the plasma treatment have already been optimized. Some, initial trails of electrospinning for creating nanofibers as coating has also been conducted but requires further optimization for an effective coating.

The objective for the “Bio-farming for bioactive compounds” projects is to document that it is possible to reduce fish diseases of farmed salmon, improve poultry health and gut microbiota and to design biodegradable packaging materials with antimicrobial properties using bioactive compounds from herbs. We have selected 9 Norwegian herbs expected to give high production of bioactive compounds. Growth conditions will be described related to climatic stress and upscaling of production for advanced closed green-house systems, open field, and growth tunnels. Bioactive compounds in herbs will then be extracted, fractionated, and isolated and screened for antioxidant, antimicrobial, anti-inflammatory characteristics. Analytical samples will be characterized by means of targeted and non-targeted metabolomics and in invitro testing. Effects of additives in fish feed will be examined in invitro test systems, determining antimicrobial activity of extracts, fractions, and isolates of herbs on the growth of Atlantic salmon pathogens. The benefits will be measured as impacts on growth, health, and immunity in fish, and physiological impacts on specific tissues (head kidney, skin, gut, blood). Benefitial effects from extracts as feed additives in poultry feed will be examined by in vitro and in vivo testing. The effects will be documented on anti-oxidant and anti-inflammatoric potential, performance, counts of intestinal pathogens, and microbiota profile. The third application is to design and optimize sustainable methods for incorporation of antimicrobial herb extract into food packaging materials, giving longer shelf life. Different cutting-edge technologies (High pressure processing, ultra-sound, and electrospinning) will be tested for an enhanced incorporation. The project is interdisciplinary, and collaboration and competence dissemination are important goals in the work.