Microalgal cultivation characteristics using commercially available air-cushion packaging material as a photobioreactor.

Air-cushion (AC) packaging has become widely used worldwide. ACs are air-filled, dual plastic packaging solutions commonly found surrounding and protecting items of value within shipping enclosures during transit. Herein, we report on a laboratory assessment employing ACs as a microalgal photobioreactor (PBR). Such a PBR inherently addresses many of the operational issues typically encountered with open raceway ponds and closed photobioreactors, such as evaporative water loss, external contamination, and predation. Using half-filled ACs, the performance of microalgal species Chlorella vulgaris, Nannochloropsis oculata, and Cyclotella cryptica (diatom) was examined and the ash-free dry cell weight and overall biomass productivity determined to be 2.39 g/L and 298.55 mg/L/day for N. oculata, 0.85 g/L and 141.36 mg/L/day for C. vulgaris, and 0.67 g/L and 96.08 mg/L/day for C. cryptica. Furthermore, maximum lipid productivity of 25.54 mg/L/day AFDCW and carbohydrate productivity of 53.69 mg/L/day AFDCW were achieved by C. cryptica, while maximum protein productivity of 247.42 mg/L/day AFDCW was attained by N. oculata. Data from this work will be useful in determining the applicability and life-cycle profile of repurposed and reused ACs as potential microalgal photobioreactors depending upon the end product of interest, scale utilized, and production costs.

A two-prong mutagenesis and adaptive evolution strategy to enhance the temperature tolerance and productivity of Nannochloropsis oculata.

Incorporation of microalgae in biorefineries intended to help society reach carbon neutrality is hindered by algal growth inhibition at high temperatures, necessitating the use of costly and carbon-intensive cooling systems. In the present study, a two-prong strategy of random mutagenesis and adaptive laboratory evolution to generate robust thermotolerant strains of Nannochloropsis oculata, was used. The best mutants demonstrated increased productivity at 35 °C, which was 10 °C higher than the optimal temperature of the wild type. In a 2-L photobioreactor at 35 °C, biomass and lipid productivity were 1.43-fold and 2.24-fold higher, respectively, than wild type at 25 °C. Higher pigment and carbohydrate content contributed to the mutants' rapid growth and enhanced photosynthetic efficiency. Metabolomics and lipidomics showed rewiring of the central carbon metabolism and membrane lipid synthesis in thermotolerant strains to ensure cellular homeostasis without compromising productivity. Tagatose and phosphatidylethanolamine upregulation were identified as future genetic targets for further enhancing lipid production.

Dissecting Enhanced Carbohydrate and Pigment Productivity in Mutants of Nannochloropsis oculata Using Metabolomics and Lipidomics.

Random mutagenesis is an effective strategy for enhancing cellular traits. In this study, we used the mutagen ethyl methanesulfonate to create fast-growing Nannochloropsis oculata mutants. When cultivated in a photobioreactor with a diel cycle, two mutants exhibited 2.2-fold higher carbohydrate productivity and 3.5-4.0-fold higher pigment productivity than the wild type, while one of them also showed 2.5-fold higher lipid productivity. A comprehensive physiological, metabolomic, and lipidomic study showed that the mutants had high levels of glucose-, galactose-, and xylose-based carbohydrates. Their high growth rate was attributed to increased chlorophyll a content, improved nitrogen assimilation, storage, and recycling, and low monogalactosyldiacyl glycerol/digalactosyldiacyl glycerol ratio, which was responsible for higher biomass productivity. The investigation revealed upregulation of lipid precursors, shedding light on high lipid accumulation. The derived algae strains are capable of increasing the biosynthesis of value-added storage molecules without impairing growth, rendering them promising candidates for commercial development in future biorefineries.

Physiological insights into enhanced lipid accumulation and temperature tolerance by Tetraselmis suecica ultraviolet mutants.

High outdoor temperatures significantly inhibit the growth and lipid production of the industrially promising marine microalga Tetraselmis suecica, which is viewed as a potential feedstock for high-value bioproducts and biofuels. To overcome this limitation, T. suecica was subjected to ultraviolet irradiation to generate mutants capable of being productive at higher temperatures. The top two high-lipid mutants UV-25 and UV-31 isolated at 25 °C and 31 °C, respectively, were compared to the wild type (WT) to delineate physiological alterations and shed light on the mutants' increased biomass and lipid productivity. At 25 °C, UV-25 and UV-31 exhibited lipid productivity of 36.12 and 31.33 mg/L day, which were 1.4- and 1.2-fold higher than WT, respectively. This increase in lipid biosynthesis correlated well with increased carotenoid content in UV-25 (2.2-fold) and UV-31 (3.6-fold), indicating an improved capacity to quench reactive oxygen species. At 31 °C, the growth and lipid accumulation of UV-31 remained high, signifying adaptation to higher temperatures. This is attributed to a well-coordinated modulation of the mutant's cellular metabolism through an increase in galactose and phosphatidylglycerol levels, as well as in protein, all of which contributed to its performance at elevated temperatures. The study successfully established a UV mutagenesis strategy for producing superior- performing microalgae strains with industrially desired traits, paving the way for future outdoor cultivation deployment.

Deciphering metabolic alterations in algae cultivated in spent media as means for enhancing algal biorefinery sustainability.

The recycling of unfiltered spent media during cultivation of Chlorella vulgaris was studied using metabolomics in an effort to enhance water and nutrient sustainability and reduce operating costs in algal biorefineries. Cultivation in spent media resulted in reduced biomass and lipid productivity by 14% and 19%, respectively, compared to fresh media. The decrease was related to a detected lower nutrient uptake. Nevertheless, carbohydrate content (28% of dry cell weight) and α-linolenic acid content (27 % of fatty acids) were higher in spent media cultures than in fresh media. Metabolomics analysis of intracellular metabolites revealed downregulation of nitrogen assimilation, tricarboxylic acid cycle, structural lipids, and energy metabolism, but upregulation of stress mitigation and carbohydrate synthesis. No growth was supported by spent media during a second cultivation cycle and was likely due to the identified extracellular accumulation of humic acid and free fatty acids that acted as growth auto-inhibitors.

Biochemical conversion of sweet sorghum bagasse to succinic acid.

Succinic acid, an important intermediate in the manufacture of plastics and other commodity and specialty chemicals, is currently made primarily from petroleum. We attempted to biosynthesize succinic acid through microbial fermentation of cellulosic sugars derived from the bagasse of sweet sorghum, a renewable feedstock that can grow in a wide range of climates around the world. We investigated pretreating sweet sorghum bagasse (SSB) with concentrated phosphoric acid at mild conditions (40-85°C) at various residence times and biomass concentrations. We then subjected the pretreated SSB to enzymatic hydrolysis with a commercial cellulase to release glucose. The highest glucose yield was obtained when SSB was pretreated at 50°C for 43 min at 130 g/L biomass concentration on dry basis. Fermentation was carried out with Actinobacillus succinogenes 130Z, which readily converted 29.2 g/L of cellulosic glucose to 17.8 g/L of succinic acid in a 3.5-L bioreactor sparged with CO2 at a rate of 0.5 vvm, thus reducing the carbon footprint of the process. Overall, we demonstrated, for the first time, the use of SSB for production of succinic acid using practices that lower energy use, future equipment cost, waste generation, and carbon footprint.