Metabolic Engineering and Process Intensification for Muconic Acid Production Using Saccharomyces cerevisiae.

The efficient production of biobased organic acids is crucial to move to a more sustainable and eco-friendly economy, where muconic acid is gaining interest as a versatile platform chemical to produce industrial building blocks, including adipic acid and terephthalic acid. In this study, a Saccharomyces cerevisiae platform strain able to convert glucose and xylose into cis,cis-muconic acid was further engineered to eliminate C2 dependency, improve muconic acid tolerance, enhance production and growth performance, and substantially reduce the side production of the intermediate protocatechuic acid. This was achieved by reintroducing the PDC5 gene and overexpression of QDR3 genes. The improved strain was integrated in low-pH fed-batch fermentations at bioreactor scale with integrated in situ product recovery. By adding a biocompatible organic phase consisting of CYTOP 503 and canola oil to the process, a continuous extraction of muconic acid was achieved, resulting in significant alleviation of product inhibition. Through this, the muconic acid titer and peak productivity were improved by 300% and 185%, respectively, reaching 9.3 g/L and 0.100 g/L/h in the in situ product recovery process as compared to 3.1 g/L and 0.054 g/L/h in the control process without ISPR.

Industrial side streams as sustainable substrates for microbial production of poly(3-hydroxybutyrate) (PHB).

Poly(3-hydroxybutyrate) (PHB) is a microbially produced biopolymer that is emerging as a propitious alternative to petroleum-based plastics owing to its biodegradable and biocompatible properties. However, to date, the relatively high costs related to the PHB production process are hampering its widespread commercialization. Since feedstock costs add up to half of the total production costs, ample research has been focusing on the use of inexpensive industrial side streams as carbon sources. While various industrial side streams such as second-generation carbohydrates, lignocellulose, lipids, and glycerol have been extensively investigated in liquid fermentation processes, also gaseous sources, including carbon dioxide, carbon monoxide, and methane, are gaining attention as substrates for gas fermentation. In addition, recent studies have investigated two-stage processes to convert waste gases into PHB via organic acids or alcohols. In this review, a variety of different industrial side streams are discussed as more sustainable and economical carbon sources for microbial PHB production. In particular, a comprehensive overview of recent developments and remaining challenges in fermentation strategies using these feedstocks is provided, considering technical, environmental, and economic aspects to shed light on their industrial feasibility. As such, this review aims to contribute to the global shift towards a zero-waste bio-economy and more sustainable materials.

Impact of Thermomechanical Fiber Pre-Treatment Using Twin-Screw Extrusion on the Production and Properties of Renewable Binderless Coriander Fiberboards.

The aim of this study consisted of manufacturing renewable binderless fiberboards from coriander straw and a deoiled coriander press cake, thus at the same time ensuring the valorization of crop residues and process by-products. The press cake acted as a natural binder inside the boards owing to the thermoplastic behavior of its protein fraction during thermopressing. The influence of different fiber-refining methods was evaluated and it was shown that a twin-screw extrusion treatment effectively improved fiber morphology and resulted in fiberboards with enhanced performance as compared to a conventional grinding process. The best fiberboard was produced with extrusion-refined straw using a 0.4 liquid/solid (L/S) ratio and with 40% press cake addition. The water sensitivity of the boards was effectively reduced by 63% through the addition of an extrusion raw material premixing operation and thermal treatment of the panels at 200 °C, resulting in materials with good performance showing a flexural strength of 29 MPa and a thickness swelling of 24%. Produced without the use of any chemical adhesives, these fiberboards could thus present viable, sustainable alternatives for current commercial wood-based materials such as oriented strand board, particleboard and medium-density fiberboard, with high cost-effectiveness.

Characterization of French Coriander Oil as Source of Petroselinic Acid.

Coriander vegetable oil was extracted from fruits of French origin in a 23% yield. The oil was of good quality, with a low amount of free fatty acids (1.8%) and a concurrently high amount of triacylglycerols (98%). It is a rich source of petroselinic acid (C18:1n-12), an important renewable building block, making up 73% of all fatty acids, with also significant amounts of linoleic acid (14%), oleic acid (6%), and palmitic acid (3%). The oil was characterized by a high unsaponifiable fraction, comprising a substantial amount of phytosterols (6.70 g/kg). The main sterol markers were ß-sitosterol (35% of total sterols), stigmasterol (24%), and Δ7-stigmastenol (18%). Squalene was detected at an amount of 0.2 g/kg. A considerable amount of tocols were identified (500 mg/kg) and consisted mainly of tocotrienols, with γ-tocotrienol as the major compound. The phospholipid content was low at 0.3%, of which the main phospholipid classes were phosphatidic acid (33%), phosphatidylcholine (25%), phosphatidylinositol (17%), and phosphatidylethanolamine (17%). About 50% of all phospholipids were non-hydratable. The ß-carotene content was low at 10 mg/kg, while a significant amount of chlorophyll was detected at about 11 mg/kg. An iron content of 1.4 mg/kg was determined through element analysis of the vegetable oil. The influence of fruit origin on the vegetable oil composition was shown to be very important, particularly in terms of the phospholipids, sterols, and tocols composition.

Pressure fermentation to boost CO2-based poly(3-hydroxybutyrate) production using Cupriavidus necator.

CO2-based poly(3-hydroxybutyrate) (PHB) can be produced by the versatile bacterium Cupriavidus necator through chemolithoautotrophic fermentation, using a gas mixture consisting of CO2, H2, and O2. Despite offering a propitious route for carbon-neutral bioplastic manufacturing, its adoption is currently hampered by the wide explosive range of the required gas mixture, as well as the limited gas-to-liquid mass transfer rates. To address these challenges, pressure fermentation was applied as a robust and effective strategy, while ensuring safe operation by adhering to the limiting O2 concentration, utilizing state-of-the-art bioreactors. Consequently, exponential growth could be prolonged, boosting CO2-based PHB production from 10.8 g/L at 1.5 bar up to 29.6 g/L at 3 bar. The production gain closely aligns with the theoretical calculations, except for when the pressure was increased up to 4 bar. Overall, the demonstrated increase in PHB production underscores the potential of pressure fermentation to enhance aerobic gas fermentation.

Does poly-3-hydroxybutyrate biodegradation affect the quality of soil organic matter?

The search for eco-friendly substitutes for traditional plastics has led to the production of biodegradable bioplastics. However, concerns have been raised about the impact of bioplastic biodegradation on soil health. Despite these concerns, the potential negative consequences of bioplastics during various stages of biodegradation remain underexplored. Therefore, this study aims to investigate the impact of micro-bioplastics made of poly-3-hydroxybutyrate (P3HB) on the properties of three different soils. In our ten-month experiment, we investigated the impact of poly-3-hydroxybutyrate (P3HB) on Chernozem, Cambisol, and Phaeozem soils. Our study focused on changes in soil organic matter (SOM), microbial activity, and the level of soil carbon and nitrogen. The observed changes indicated an excessive level of biodegradation of SOM after the soils were enriched with micro-particles of P3HB, with concentrations ranging from 0.1% to 3%. The thermogravimetric analysis confirmed the presence of residual P3HB (particularly in the 3% treatment) and underscored the heightened biodegradation of SOM, especially in the more stable SOM fractions. This was notably evident in Phaeozem soils, where even the stable SOM pool was affected. Elemental analysis revealed changes in soil organic carbon content following P3HB degradation, although nitrogen levels remained constant. Enzymatic activity was found to vary with soil type and responded differently across P3HB concentration levels. Our findings confirmed that P3HB acts as a bioavailable carbon source. Its biodegradation stimulates the production of enzymes, which in turn affects various soil elements, indicating complex interactions within the soil ecosystem.

Challenges and progress towards industrial recombinant protein production in yeasts: A review.

Recombinant proteins (RP) are widely used as biopharmaceuticals, industrial enzymes, or sustainable food source. Yeasts, with their ability to produce complex proteins through a broad variety of cheap carbon sources, have emerged as promising eukaryotic production hosts. As such, the prevalence of yeasts as favourable production organisms in commercial RP production is expected to increase. Yet, with the selection of a robust production host on the one hand, successful scale-up is dependent on a thorough understanding of the challenging environment and limitations of large-scale bioreactors on the other hand. In the present work, several prominent yeast species, including Saccharomyces cerevisiae, Pichia pastoris, Yarrowia lipolytica, Kluyveromyces lactis and Kluyveromyces marxianus are reviewed for their current state and performance in commercial RP production. Thereafter, the impact of principal process control parameters, including dissolved oxygen, pH, substrate concentration, and temperature, on large-scale RP production are discussed. Finally, technical challenges of process scale-up are identified. To that end, process intensification strategies to enhance industrial feasibility are summarized, specifically highlighting fermentation strategies to ensure sufficient cooling capacity, overcome oxygen limitation, and increase protein quality and productivity. As such, this review aims to contribute to the pursuit of sustainable yeast-based RP production.

Environmental life cycle assessment of nutraceuticals: A case study on methylcobalamin in different packaging types.

The growing production of pharmaceuticals and nutraceuticals, e.g., methylcobalamin supplements, improves the health of people. This study assesses the environmental footprint of chewable methylcobalamin supplements in four packaging types: blister packs or bottles made of HDPE, PET, or glass. A cradle-to-grave life cycle assessment is conducted to evaluate the supply to Belgian consumers of the recommended daily dose of methylcobalamin supplementation (1.2 mg) in case of deficiency. The impact of methylcobalamin manufacturing in major producing countries (China as baseline and France) is analyzed based on detailed synthesis modeling of data points coming from patents. The overall carbon footprint (CF) is dominated by the transport of consumers to the pharmacy and methylcobalamin powder manufacturing in China (while its mass share per supplement is only 1 %). The impact is the lowest for supplements in HDPE bottles (6.3 g CO2 eq) and 1 %, 8 %, and 35 % higher for those in PET bottles, glass bottles, and blister packs, respectively. Tablets in blister packs have for other investigated impact categories (fossil resource footprint (FRF); acidification; eutrophication: freshwater, marine, and terrestrial; freshwater ecotoxicity; land use; and water use) the highest footprint and those in HDPE and PET bottles for most the lowest. The CF of methylcobalamin powder manufacturing in France is 22 % lower than in China (2.7 g CO2 eq), while the FRF is similar in both locations (26-27 kJ). The FRF and the difference in the CF are chiefly due to energy use and solvent production emissions. Similar trends as the CF are found for other investigated impact categories. Valuable conclusions are drawn for environmental studies on pharmaceuticals and nutraceuticals: (i) including accurate data on consumer transport, (ii) using more environmentally-friendly active ingredients, (iii) choosing appropriate packaging types considering multiple aspects: convenience, environmental footprint, etc., and (iv) providing a holistic picture through assessing various impact categories.

Advanced PHB fermentation strategies with CO2-derived organic acids.

Over the past decade, formic acid and acetic acid have gained increasing attention as alternative feedstocks for poly-3-hydroxybutyrate (PHB) production as these potentially CO2-derived molecules are naturally assimilated by Cupriavidus necator. Both organic acids were individually evaluated in fed-batch fermentations at bioreactor scale. Acetic acid was revealed as the most promising carbon source yielding 42.3 g L-1 PHB, whereas no significant amount of PHB was produced from formic acid. Hence, acetic acid was further used as the substrate during process intensification. Key performance characteristics, including process stability, PHB titer, and productivity were optimized by introducing NH4-acetate as the nitrogen source, extending the growth phase, and implementing a repeated fed-batch procedure, respectively. These advanced fermentation strategies resulted in the establishment of a stable fermentation process reaching 58.5 g L-1 PHB, while doubling the productivity to 0.93 g L-1 h-1 PHB.

VOC and carbonyl compound emissions of a fiberboard resulting from a coriander biorefinery: comparison with two commercial wood-based building materials.

Indoor air quality is a major public health issue. It is related to the choice of construction materials and associated with VOC emissions. Two wood-based commercial panels were tested: a medium-density fiberboard (MDF) and a chipboard (CH), and they were compared to a material produced from a coriander biorefinery (COR). Indicators chosen to compare the materials were physical properties (density, bending properties, surface hardness, thickness swelling, and water absorption) and VOC emissions. Emissions were evaluated in an environmental chamber at 23 °C, 31 °C, and 36 °C, and during 28 days. Carbonyl emissions on day 1 at 23 °C were 74, 146, and 35 µg m-2 h-1, respectively, for MDF, CH, and COR. Terpenic emissions were 12, 185, and 37 µg m-2 h-1, respectively. Higher temperature resulted in higher emissions which decreased over time, except for formaldehyde. VOC emissions depended largely on material and temperature. Formaldehyde emission was 300 to 600 times lower for coriander boards (< 0.2 µg m-2 h-1), making them significantly more environmentally friendly materials in comparison with MDF and chipboard. These results highlight the interest of coriander by-products as raw materials for producing fiberboards with low impact on indoor air quality.

Petroselinic acid purification and its use for the fermentation of new sophorolipids.

Petroselinic acid, a positional isomer of oleic acid, was isolated from the vegetable oil of Coriandrum sativum fruits. This uncommon fatty acid was subsequently used as substrate for sophorolipid fermentation with a Starmerella bombicola lactone esterase overexpression (oe sble) strain. A petroselinic acid based diacetylated sophorolipid lactone was obtained in high purity without incorporation of de novo synthesized fatty acids such as oleic acid. A total production of 40 g/L was obtained. The petroselinic acid based sophorolipid lactone was subsequently hydrolyzed towards the petroselinic acid based sophorolipid acid. For both compounds, their critical micelle concentration (CMC) and corresponding surface tension were compared to their oleic acid based counterparts. Both petroselinic acid based sophorolipids displayed a much lower CMC value than their oleic acid based counterparts, although their minimal surface tension was the same. Besides, the sophorolipid fermentation product was chemically modified towards a novel C12 sophorolipid aldehyde. This derivative constitutes an interesting building block for further modification towards new-to-nature sophorolipids with high potential for self-assembly applications.