Isolation and quantification of alginate in choline chloride-based deep eutectic solvents.

Extraction of seaweed compounds using Deep Eutectic Solvents (DES) has shown high interest. Quantification, however, is challenging due to interactions with DES components. In this research work, three chemical separation techniques were investigated to isolate and quantify alginate from a set of choline chloride-based DES. While choline chloride served as the hydrogen bond acceptor (HBA); Urea, Ethylene Glycol, Propylene Glycol, Glycerol, Sorbitol, Xylitol and Glucose were used as hydrogen bond donors (HBD). DES containing sodium alginate were subjected to precipitation with sulfuric acid 0.2 M (pH 1.6), ethanol-water mixture (80 % v/v) and calcium chloride (1 % w/v CaCl2·2H2O). Alginate in precipitates was quantified and used to evaluate the performance of each separation technique. The highest recovery yields (51.2 ± 1.3 %) were obtained using the ethanol-water mixture followed by calcium chloride (45.7 ± 1.2 %), except for polyols (e.g. sorbitol). The lowest recovery yields were obtained with acid, with a particularly low recovery yield when urea was used as HBD (9.6 ± 1.3 %). Estimations of ManA/GulA ratios showed lower values for precipitates from DES compared to the ones obtained from water. This research shows ethanolic precipitation as a suitable method for alginate separation from the studied set of choline chloride-based DES.

Pseudomonas putida as a platform for medium-chain length α,ω-diol production: Opportunities and challenges.

Medium-chain-length α,ω-diols (mcl-diols) play an important role in polymer production, traditionally depending on energy-intensive chemical processes. Microbial cell factories offer an alternative, but conventional strains like Escherichia coli and Saccharomyces cerevisiae face challenges in mcl-diol production due to the toxicity of intermediates such as alcohols and acids. Metabolic engineering and synthetic biology enable the engineering of non-model strains for such purposes with P. putida emerging as a promising microbial platform. This study reviews the advancement in diol production using P. putida and proposes a four-module approach for the sustainable production of diols. Despite progress, challenges persist, and this study discusses current obstacles and future opportunities for leveraging P. putida as a microbial cell factory for mcl-diol production. Furthermore, this study highlights the potential of using P. putida as an efficient chassis for diol synthesis.

The genome of the Arctic snow alga Limnomonas spitsbergensis (Chlamydomonadales).

Snow algae are a diverse group of extremophilic microeukaryotes found on melting polar and alpine snowfields. They play an important role in the microbial ecology of the cryosphere, and their propagation on snow and ice surfaces may in part accelerate climate-induced melting of these systems. High-quality snow algae genomes are needed for studies on their unique physiology, adaptive mechanisms, and genome evolution under multiple forms of stress, including cold temperatures and intense sunlight. Here, we assembled and annotated the genome of Limnomonas spitsbergensis, a cryophilic biciliate green alga originally isolated from melting snow on Svalbard, in the Arctic. The L. spitsbergensis genome assembly is based primarily on the use of PacBio long reads and secondly Illumina short reads, with an assembly size of 260.248 Mb in 124 contigs. A combination of 3 alternative annotation strategies was used including protein homology, RNA-seq evidence, and PacBio full-length transcript isoforms. The best merged set of annotations identified 18,277 protein-coding genes, which were 95.2% complete based on Benchmarking Universal Single-Copy Orthologs analysis. We also provide the annotated mitogenome, which is a relatively large 77.942 kb circular mapping sequence containing extensive repeats. The L. spitsbergensis genome will provide a new resource for research on snow algae adaptation, behavior, and natural selection in unique, low-temperature terrestrial environments that are under threat from climate change.

Outdoor cultivation of Picochlorum sp. in a novel V-shaped photobioreactor on the Caribbean island Bonaire.

Microalgae are a promising renewable feedstock that can be produced on non-arable land using seawater. Their biomass contains proteins, lipids, carbohydrates, and pigments, and can be used for various biobased products, such as food, feed, biochemicals, and biofuels. For such applications, the production costs need to be reduced, for example, by improving biomass productivity in photobioreactors. In this study, Picochlorum sp. (BPE23) was cultivated in a prototype of a novel outdoor V-shaped photobioreactor on Bonaire (12°N, 68°W). The novel photobioreactor design was previously proposed for the capture and dilution of sunlight at low-latitude locations. During several months, the biomass productivity of the local thermotolerant microalgae was determined at different dilution rates in continuous dilution and batch dilution experiments, without any form of temperature control. Reactor temperatures increased to 35°C-45°C at midday. In the continuous dilution experiments, high average biomass productivities of 28-31 g m-2 d-1 and photosynthetic efficiencies of 3.5%-4.3% were achieved. In the batch dilution experiments, biomass productivities were lower (17-23 g m-2 d-1), as microalgal cells likely experienced sudden light and temperature stress after daily reactor dilution. Nonetheless, dense cultures were characterized by high maximum photosynthetic rates, illustrating the potential of Picochlorum sp. for fast growth under outdoor conditions.

Pressure-driven membrane processes for the recovery and recycling of deep eutectic solvents: A seaweed biorefinery case study.

Deep eutectic solvents (DES) are green alternatives for conventional solvents. They have gained attention for their potential to extract valuable compounds from biomass, such as seaweed. In this framework, a case study was developed to assess the feasibility of pressure-driven membrane processes as an efficient tool for the recovery of deep eutectic solvents and targeted biomolecules. For this purpose, a mixture composed of the DES choline chloride - ethylene glycol (ChCl-EG) 1:2, water and alginate was made to mimic a DES extraction from seaweed. An integrated separation process design was proposed where ultrafiltration-diafiltration-nanofiltration (UF-DF-NF) was coupled. UF and DF were found to be effective for the separation of alginate with an 85 % yield. DES was likewise recovered by 93 %, proving the membrane filtrations' technical feasibility. The NF performance to separate the DES from the water, for its recycling, laid by a 45 %-50 % retention and a final concentrated DES solution of 18 %(v/v).

Triple Space-Time Yield in Discontinuous Antibody Biomanufacturing by Combination of Synergetic Process Intensification Strategies.

Monoclonal antibodies are the workhorse of the pharmaceutical industry due to their potential to treat a variety of different diseases while providing high specificity and efficiency. As a consequence, a variety of production processes have been established within the biomanufacturing industry. However, the rapidly increasing demand for therapeutic molecules amid the recent COVID-19 pandemic demonstrated that there still is a clear need to establish novel, highly productive, and flexible production processes. Within this work, we designed a novel discontinuous process by combining two intensification strategies, thus increasing inoculation density and media exchange via a fluidized bed centrifuge, to fulfill the need for a flexible and highly productive production process for therapeutic molecules. To establish this new process, firstly, a small-scale experiment was conducted to verify synergies between both intensification strategies, followed by a process transfer towards the proof-of-concept scale. The combination of these two-process intensification measures revealed overall synergies resulting in decreased process duration (-37%) and strongly enhanced product formation (+116%) in comparison to the not-intensified standard operation. This led to an impressive threefold increase in space-time yield, while only negligible differences in product quality could be observed. Overall, this novel process not only increases the ways to react to emergency situations thanks to its flexibility and possible short development times, but also represents a possible alternative to the current established processes due to high increases in productivity, in comparison to standard fed-batch operations.