Freshwater microalgae Nannochloropsis limnetica for the production of ß-galactosidase from whey powder.

This study investigated the first-ever reported use of freshwater Nannochloropsis for the bioremediation of dairy processing side streams and co-generation of valuable products, such as ß-galactosidase enzyme. In this study, N. limnetica was found to grow rapidly on both autoclaved and non-autoclaved whey-powder media (referred to dairy processing by-product or DPBP) without the need of salinity adjustment or nutrient additions, achieving a biomass concentration of 1.05-1.36 g L-1 after 8 days. The species secreted extracellular ß-galactosidase (up to 40.84 ± 0.23 U L-1) in order to hydrolyse lactose in DPBP media into monosaccharides prior to absorption into biomass, demonstrating a mixotrophic pathway for lactose assimilation. The species was highly effective as a bioremediation agent, being able to remove > 80% of total nitrogen and phosphate in the DPBP medium within two days across all cultures. Population analysis using flow cytometry and multi-channel/multi-staining methods revealed that the culture grown on non-autoclaved medium contained a high initial bacterial load, comprising both contaminating bacteria in the medium and phycosphere bacteria associated with the microalgae. In both autoclaved and non-autoclaved DPBP media, Nannochloropsis cells were able to establish a stable microalgae-bacteria interaction, suppressing bacterial takeover and emerging as dominant population (53-80% of total cells) in the cultures. The extent of microalgal dominance, however, was less prominent in the non-autoclaved media. High initial bacterial loads in these cultures had mixed effects on microalgal performance, promoting ß-galactosidase synthesis on the one hand while competing for nutrients and retarding microalgal growth on the other. These results alluded to the need of effective pre-treatment step to manage bacterial population in microalgal cultures on DPBP. Overall, N. limnetica cultures displayed competitive ß-galactosidase productivity and propensity for efficient nutrient removal on DPBP medium, demonstrating their promising nature for use in the valorisation of dairy side streams.

Nitrogen and phosphate removal from dairy processing side-streams by monocultures or consortium of microalgae.

Acid-casein production generates waste streams that are rich in nitrogen (in the form of protein and nitrate) and phosphate. This makes this type of waste very difficult to treat using conventional techniques resulting in a high amount of operating cost and costly investment. In this research, the application of single culture or consortium of microalgae for uptake of nitrogen and phosphate in the wastewater of an acid-casein factory was investigated. The waste was a 1:1 mixture of nanofiltered whey permeate and dairy processing wastewater. Monocultures of Chlorella vulgaris, Tetradesmus obloquus, Nonnochlropsis ocenica and a consortium of the three microalgae were analyzed. The results showed that the consortium exhibited more efficient nitrogen and phosphate removal compared to the individual species. The consortium was able to rapidly hydrolyse exogenous protein present in the waste medium, removing 88% of protein and breaking down complex protein molecules into simpler compounds (such as nitrate) for assimilation into the biomass. In the first fourteen days of cultivation, the rate of nitrate assimilation by the consortium biomass was lower than that of nitrate formation from protein degradation, leading to a net increase in nitrate concentration in the medium. As protein source was depleted and biomass concentration increased, however, the rate of nitrate assimilation began to exceed that of nitrate formation allowing for net removal of nitrate. The microalgae consortium was shown to successfully bioremediate all nitrates by day 21. It was indicated that Chlorella and Nannochloropsis species were responsible for nitrogen removal in monocultures. Phosphate, on the other hand, was efficiently removed by Tetradesmus. The results indicated that a consortium cultivation of three species of microalgae led to effective elimination of both nitrogen and phosphate. Combined flow-cytometry and microscopy analyses revealed that Chlorella overtook Tetradesmus and Nannochloropsis to emerge as the dominant population in the consortium by the end of the cultivation cycle. It can be concluded that the application of microalgae consortium for simultaneous recovery of nitrogen and phosphate is a promising approach for treating acid-casein wastewater.

The effect of cell disruption on the extraction of oil and protein from concentrated microalgae slurries.

Novel cell-disruption combinations (autolytic incubation and hypotonic osmotic shock combined with HPH or pH12) were used to investigate the fundamental mass transfer of lipids and proteins from Nannochloropsis slurries (140 mg biomass/g slurry). Since neutral lipids exist as cytosolic globules, their mass transfer was directly dependent on disintegration of cell walls. Complete recovery was obtained with complete physical disruption. HPH combinations exerted more physical disruption and led to higher yields than pH12. In contrast, proteins exist as both cytosolic water-soluble fractions and cell-wall/membrane structural fractions and have a complex extraction behaviour. Mass transfer of cytosolic proteins was dependent on cell-wall disintegration, while that of structural proteins was governed by cell-wall disintegration and severance of protein linkage from the wall/membrane. HPH combinations exerted only physical disruption and were limited to releasing soluble proteins. pH12 combinations hydrolysed chemical linkages in addition to exerting physical disruption, releasing both soluble and structural proteins.

Thermally coupled dark-anoxia incubation: A platform technology to induce auto-fermentation and thus cell-wall thinning in both nitrogen-replete and nitrogen-deplete Nannochloropsis slurries.

Nitrogen-deprived Nannochloropsis cells invested their fixed carbon into the accumulation of triacylglycerol and cell wall cellulose (thickness of N-replete cell walls = 27.8 ±â€¯5.8, N-deplete cell walls = 51.0 ±â€¯10.2 nm). In this study, the effect of nitrogen depletion on the ability of the cells to weaken their own cell walls via autolysis was investigated. Autolytic cell wall thinning was achieved in both N-replete and N-deplete biomass by incubating highly concentrated slurries in darkness at 38 °C. The incubation forced cells to anaerobically ferment their intracellular cellulose and resulted in 30-40% reduction in cell wall thickness for both biomass types. This wall depletion weakened the cells and increased the extent of cell rupture by mechanical force (from 42 to 78% for N-replete biomass, from 36 to 62% for N-deplete biomass). Importantly, autolysis did not adversely impact the amino acid content of protein-rich N-replete biomass or the fatty acid content of lipid-rich N-deplete biomass.

Conversion and recovery of saponifiable lipids from microalgae using a nonpolar solvent via lipase-assisted extraction.

A single-step method for transesterifying and recovering lipids in concentrated slurries (ca 20% w/w solids) of ruptured microalgae is presented. A soluble Rhizomucor miehei lipase (RML) was used to directly transesterify the lipids in the marine microalgae Nannochloropsis salina. This allowed both triglycerides (TAG) and polar saponifiable lipids to be recovered as fatty acid methyl esters (FAME) using a nonpolar solvent (hexane). Up to 90 wt% of the total saponifiable lipids (SL) were converted to FAME within 24 h, approximately 75% of which was recovered in the hexane by centrifugation. Two pathways for the conversion and recovery of polar lipids were identified. The water in the slurry buffered against potential lipase inhibition by methanol, but necessitated a high methanol dose for maximal FAME conversion. Nonetheless the method enables the recovery of polar lipids as FAME while avoiding the need for both drying of the biomass and a downstream transesterification step.

Mechanical cell disruption for lipid extraction from microalgal biomass.

Cell disruption is an integral part of the downstream operation required to produce biodiesel from microalgae. This study investigated the use of ultrasonication and high-pressure homogenization (HPH) as cell disruption methods for two microalgal species, Tetraselmis suecica (TS) and Chlorococcum sp. (C sp.). The kinetics of cell disruption followed a first-order model (0.65

Extraction of oil from microalgae for biodiesel production: A review.

The rapid increase of CO(2) concentration in the atmosphere combined with depleted supplies of fossil fuels has led to an increased commercial interest in renewable fuels. Due to their high biomass productivity, rapid lipid accumulation, and ability to survive in saline water, microalgae have been identified as promising feedstocks for industrial-scale production of carbon-neutral biodiesel. This study examines the principles involved in lipid extraction from microalgal cells, a crucial downstream processing step in the production of microalgal biodiesel. We analyze the different technological options currently available for laboratory-scale microalgal lipid extraction, with a primary focus on the prospect of organic solvent and supercritical fluid extraction. The study also provides an assessment of recent breakthroughs in this rapidly developing field and reports on the suitability of microalgal lipid compositions for biodiesel conversion.

Oil extraction from microalgae for biodiesel production.

This study examines the performance of supercritical carbon dioxide (SCCO(2)) extraction and hexane extraction of lipids from marine Chlorococcum sp. for lab-scale biodiesel production. Even though the strain of Chlorococcum sp. used in this study had a low maximum lipid yield (7.1 wt% to dry biomass), the extracted lipid displayed a suitable fatty acid profile for biodiesel [C18:1 (∼63 wt%), C16:0 (∼19 wt%), C18:2 (∼4 wt%), C16:1 (∼4 wt%), and C18:0 (∼3 wt%)]. For SCCO(2) extraction, decreasing temperature and increasing pressure resulted in increased lipid yields. The mass transfer coefficient (k) for lipid extraction under supercritical conditions was found to increase with fluid dielectric constant as well as fluid density. For hexane extraction, continuous operation with a Soxhlet apparatus and inclusion of isopropanol as a co-solvent enhanced lipid yields. Hexane extraction from either dried microalgal powder or wet microalgal paste obtained comparable lipid yields.