Solid-state fermentation of the microalgae Scenedesmus sp. for improved conservation and protein digestibility

Abstract: As part of a research grant aimed at producing chicken feed from the novel microalgae Scenedesmus sp., called the ReMAPP project, this thesis investigates opportunities to increase the conservation potential and protein digestibility utilizing solid-state lactic acid fermentation and commercial enzyme additives. The lactic acid fermentation mimics the traditional agricultural ensiling process, where crops are anaerobically fermented at room temperature for upwards of a year. The ensiling process conserves the material by using lactic acid bacteria to convert water soluble carbohydrates in the biomass to mainly lactic acid, lowering the pH and preventing growth of pathogens. This approach to microalgae conservation has not been attempted previously. Enzyme additives are hypothesized to degrade cell wall polymers to monosaccharides and are used to increase the substrate for lactic acid production, further decreasing the pH and thus improving the silage quality of the microalgae. At the same time, the protein digestibility of the specific microalgae species is hypothesized to be limited by bioavailability as the incalcitrant cell wall of Scenedesmus could prevent access to intracellular proteins, rendering them indigestible. The same enzyme additives used to improve fermentation could also affect the protein digestibility, since degradation of cell wall polymers could release intracellular proteins. To investigate the fermentation properties and protein digestibility, lab scale fermentations are carried out where Lactoplantibacillus plantarum and a variety of enzyme additives are mixed with Scenedesmus sp. biomass and left in vacuum-packed bags to ferment for 21 days. The results show bacterial inoculation by itself to be effective in decreasing pH to desirable levels but fermentation with β-glucanases, which seem to degrade polyglucans to glucose which are consumed by the lactic acid bacteria, proved the most effective. Only fermentations with β-glucanases achieved pH < 4.0, a benchmark for product safety. The effect on protein digestibility did however seem to be largely negligible. Lactic acid fermentation by itself did increase the protein digestibility, measured in PDCAAS, from 0.79 to 0.83 which is a relevant increase but still below the gold standard set by soybean. Using additional enzymes had only a marginal effect on increasing protein digestibility, and as such the hypothesis of degrading the cell wall to increase protein digestibility cannot be confirmed by this study. Endoproteases increased protein digestibility the most, the do likely not increase protein digestibility by degrading the cell wall but rather by hydrolyzing peptides. The most successful non-protease in terms of protein digestibility was a pectinase which was unexpected since no galacturonic acid, the main monomer of pectin, could be detected in the sample. To determine the cause of increase in protein digestibility and the effect pectinases have on the biomass, in-depth characterization on the biochemical composition of Scenedesmus sp. would be required. In conclusion, solid-state fermentation of Scenedesmus sp. biomass using lactic acid bacteria and enzyme additive was successful and could potentially be a useful conversation method for green microalgae in general. While some enzymes tested did yield an increase in protein digestibility where a pectinase and an endoprotease proved most effective, the hypothesis of releasing intracellular proteins to increase digestibility in combination with fermentation could not be confirmed.