Semicontinuous system for the production of recombinant mCherry protein in Chlamydomonas reinhardtii.

Biotechnology advances have allowed bacteria, yeasts, plants, mammalian and insect cells to function as heterologous protein expression systems. Recently, microalgae have gained attention as an innovative platform for recombinant protein production, due to low culture media cost, compared to traditional systems, as well as the fact that microalgae such as Chlamydomonas reinhardtii are considered safe (GRAS) by the Food and Drug Administration (FDA). Previous studies showed that recombinant protein production in traditional platforms by semicontinuous process increased biomass and bio product productivity, when compared to batch process. As there is a lack of studies on semicontinuous process for recombinant protein production in microalgae, the production of recombinant mCherry fluorescent protein was evaluated by semicontinuous cultivation of Chlamydomonas reinhardtii in bubble column photobioreactor. This semicontinuous cultivation process was evaluated in the following conditions: 20%, 40%, and 60% culture portion withdrawal. The highest culture withdrawal percentage (60%) provided the best results, as an up to 161% increase in mCherry productivity (454.5 RFU h-1 - Relative Fluorescence Unit h-1 ), in comparison to batch cultivation (174.0 RFU h-1 ) of the same strain. All cultivations were carried out for 13 days, at pH 7, temperature 25°C and, by semicontinuous process, two culture withdrawals were taken during the cultivations. Throughout the production cycles, it was possible to obtain biomass concentration up to 1.36 g L-1 .

Biomass and pigment production for Arthrospira platensis via semi-continuous cultivation in photobioreactors: Temperature effects.

This study describes the response of Arthrospira platensis to a variety of temperature conditions as reflected in variations of photosynthetic parameters, pigmentation, and biomass productivity in indoor photobioreactor (PBR) cultivations. These experiments are designed to better understand the impact of temperature, seasonal variations, and acclimation effects on outdoor biomass production. The irradiance level and temperature range (20-39°C) are chosen to enable modeling of semi-continuous operation of large-scale outdoor PBR deployments. Overall, the cultivations are quite stable with some pigment-related instabilities after prolonged high-temperature exposure. Changes in productivity with temperature, as reflected in measured photosynthetic parameters, are immediate and mainly attributable to the temperature dependence of the photosaturation parameter, a secondary factor being variation in pigment content on a longer time scale corresponding to turnover of the culture population. Though pigment changes are not accompanied by significant changes in productivity, prolonged exposure at 35°C and above yields a clear degradation in performance. Productivities in a semi-continuous operation are quantitatively reproduced with a productivity model incorporating photosynthetic parameters measured herein. This study confirms the importance of temperature for biomass and pigment production in Arthrospira cultivations and provides a basis for risk assessments related to temperature mitigation for large-scale outdoor cultivations.

Algae as green energy reserve: Technological outlook on biofuel production.

Depletion of fossil fuel sources and their emissions have triggered a vigorous research in finding alternative and renewable energy sources. In this regard, algae are being exploited as a third generation feedstock for the production of biofuels such as bioethanol, biodiesel, biogas, and biohydrogen. However, algal based biofuel does not reach successful peak due to the higher cost issues in cultivation, harvesting and extraction steps. Therefore, this review presents an extensive detail of deriving biofuels from algal biomass starting from various algae cultivation systems like raceway pond and photobioreactors and its bottlenecks. Evolution of biofuel feedstocks from edible oils to algae have been addressed in the initial section of the manuscript to provide insights on the different generation of biofuel. Different configuration of photobioreactor systems used to reduce contamination risk and improve biomass productivity were extensively discussed. Photobioreactor performance greatly relies on the conditions under which it is operated. Hence, the importance of such conditions alike temperature, light intensity, inoculum size, CO2, nutrient concentration, and mixing in bioreactor performance have been described. As the lipid is the main component in biodiesel production, several pretreatment methods such as physical, chemical and biological for disrupting cell membrane to extract lipid were comprehensively reviewed and presented. This review article had put forth the recent advancement in the pretreatment methods like hydrothermal processing of algal biomasses using acid or alkali. Eventually, challenges and future dimensions in algal cultivation and pretreatment process were discussed in detail for making an economically viable algal biofuel.

Statistical optimization of light intensity and CO2 concentration for lipid production derived from attached cultivation of green microalga Ettlia sp.

Attached cultivation systems have been receiving extensive attention as a breakthrough in microalgae cultivation technology. However, there is a lack of studies that emphasize precise optimization of important parameters in attached cultivation of microalgae. In this study, the effects of two major environmental parameters in photoautotrophic cultivation, light intensity and CO2 concentration, on the biomass and lipid surface productivity of Ettlia sp. YC001 were optimized by employing Response Surface Methodology (RSM) and validated experimentally. The optimum initial conditions for attached cultivation were use of seed from the late exponential phase (LE) and an inoculum surface density of 2.5 g/m2. By optimization, maximum biomass surface productivity of 28.0 ± 1.5 g/m2/day was achieved at 730 µE/m2/s with 8% CO2. The maximum lipid surface productivity was 4.2 ± 0.3 g/m2/day at 500 µE/m2/s with 7% CO2. Change of the fatty acid composition with respect to changes in environment parameters led to improvement of biodiesel quality at higher light intensity and higher CO2 concentration. Attached cultivation of Ettlia sp. YC001 has successfully produced biomass and lipids at a high production rate with relatively low light energy demand and high CO2 utilization.

Increasing the Photoautotrophic Growth Rate of Synechocystis sp. PCC 6803 by Identifying the Limitations of Its Cultivation.

Many conditions have to be optimized in order to be able to grow the cyanobacterium Synechocystis sp. PCC 6803 (Synechocystis) for an extended period of time under physiologically well-defined and constant conditions. It is still poorly understood what limits growth of this organism in batch and continuous cultures in BG-11, the standard medium used to grow Synechocystis. Through a series of batch experiments in flasks and continuous mode experiments in advanced photobioreactors, it is shown that the limiting nutrient during batch cultivation is sulfate, the depletion of which leads to ROS formation and rapid bleaching of pigments after entry into stationary phase. In continuous mode, however, the limiting nutrient is iron. Optimizing these growth conditions resulted in a so far highest growth rate of 0.16 h-1 (4.3 h doubling time), which is significantly higher than the textbook value of 0.09 h-1 (8 h doubling time). An improved medium, BG-11 for prolonged cultivation (BG-11-PC) is introduced, that allows for controlled, extended cultivation of Synechocystis, under well-defined physiological conditions. The data present here have implications for mass-culturing of cyanobacteria.