Enhancement of biogas production rate from bioplastics by alkaline pretreatment

Producción CientíficaThe effect of alkali-based pretreatment on the methanization of bioplastics was investigated. The tested bioplastics included PHB [poly(3-hydroxybutyrate)], PHBH [poly(3-hydroxybutyrate-co-3-hydroxyhexanoate)], PHBV [poly(3-hydroxybutyrate-co-3-hydroxyvalerate], PLA (polylactic acid), and a PLA/PCL [poly(caprolactone)] 80/20 blend. Prior to methanization tests, the powdered polymers (500–1000 μm) at a concentration of 50 g/L were subjected to alkaline pretreatment using NaOH 1 M for PLA and PLA/PCL, and NaOH 2 M for PHB-based materials. Following 7 days of pretreatment, the amount of solubilized carbon for PLA and its blend accounted for 92–98% of the total initial carbon, while lower carbon recoveries were recorded for most PHB-based materials (80–93%), as revealed by dissolved total organic carbon analysis. The pretreated bioplastics were then tested for biogas production by means of mesophilic biochemical methane potential tests. Compared to unpretreated PHBs, methanization rates of pretreated PHBs were accelerated by a factor of 2.7 to 9.1 with comparable (430 NmL CH4/g material feed) or slightly lower (15% in the case of PHBH) methane yields, despite featuring a 1.4–2.3 times longer lag phases. Both materials, PLA and the PLA/PCL blend, were only extensively digested when pretreated, yielding about 360–380 NmL CH4 per gram of material fed. Unpretreated PLA-based materials showed nearly zero methanization under the timeframe and experimental conditions tested. Overall, the results suggested that alkaline pretreatment can help to enhance the methanization kinetics of bioplastics.Ministerio de Ciencia e Innovación - AEI and European Union NextGenerationEU/PRTR (RYC2021‐034559‐I)Junta de Castilla y León - EU-FEDER (CLU 2017-09, CL-EI-2021-07 y UIC 315

Biodegradation of bioplastics under aerobic and anaerobic aqueous conditions: Kinetics, carbon fate and particle size effect

Producción CientíficaThe biodegradation of PHB, PHBV, PBS, PBAT, PCL, PLA, and a PLA-PCL blend was systematically compared under aerobic and anaerobic aqueous conditions assessing biodegradation kinetics, extent, carbon fate and particle size influence (in the range of 100–1000 µm). Under standard test conditions, PHB and PBHV were biodegraded anaerobically (83.9±1.3% and 81.2±1.7%, respectively) in 77 days or aerobically (83.0±1.6% and 87.4±7.5%) in 117 days, while PCL was only biodegraded (77.6±2.4%) aerobically in 177 days. Apparent biomass growth accounted for up to 30.5% of the total initial carbon depending on the bioplastic and environment. Maximum aerobic and anaerobic biodegradation rates were improved up to 331 and 405%, respectively, at the lowest particle size tested (100–250 µm). This study highlights the usefulness of a more detailed analysis of biodegradation kinetics and carbon fate to improve both the development and testing of biodegradable materials/products in the context of a circular bioeconomy.Junta de Castilla y León - Fondo Europeo de Desarrollo Regional (projects CLU 2017-09 and UIC 315

Co-cultures of Propionibacterium freudenreichii and Bacillus amyloliquefaciens cooperatively upgrade sunflower seed milk to high levels of vitamin B12 and multiple co-benefits

Background Sunflower seeds (Helianthus annuus) display an attractive source for the rapidly increasing market of plant-based human nutrition. Of particular interest are press cakes of the seeds, cheap residuals from sunflower oil manufacturing that offer attractive sustainability and economic benefits. Admittedly, sunflower seed milk, derived therefrom, suffers from limited nutritional value, undesired flavor, and the presence of indigestible sugars. Of specific relevance is the absence of vitamin B12. This vitamin is required for development and function of the central nervous system, healthy red blood cell formation, and DNA synthesis, and displays the most important micronutrient for vegans to be aware of. Here we evaluated the power of microbes to enrich sunflower seed milk nutritionally as well as in flavor. Results Propionibacterium freudenreichii NCC 1177 showed highest vitamin B12 production in sunflower seed milk out of a range of food-grade propionibacteria. Its growth and B12 production capacity, however, were limited by a lack of accessible carbon sources and stimulants of B12 biosynthesis in the plant milk. This was overcome by co-cultivation with Bacillus amyloliquefaciens NCC 156, which supplied lactate, amino acids, and vitamin B7 for growth of NCC 1177 plus vitamins B2 and B3, potentially supporting vitamin B12 production by the Propionibacterium. After several rounds of optimization, co-fermentation of ultra-high-temperature pre-treated sunflower seed milk by the two microbes, enabled the production of 17 µg (100 g)−1 vitamin B12 within four days without any further supplementation. The fermented milk further revealed significantly enriched levels of L-lysine, the most limiting essential amino acid, vitamin B3, vitamin B6, improved protein quality and flavor, and largely eliminated indigestible sugars. Conclusion The fermented sunflower seed milk, obtained by using two food-grade microbes without further supplementation, displays an attractive, clean-label product with a high level of vitamin B12 and multiple co-benefits. The secret of the successfully upgraded plant milk lies in the multifunctional cooperation of the two microbes, which were combined, based on their genetic potential and metabolic signatures found in mono-culture fermentations. This design by knowledge approach appears valuable for future development of plant-based milk products

Production of volatile fatty acids (VFAs) from five commercial bioplastics via acidogenic fermentation

Producción CientíficaThe feasibility of producing volatile fatty acids (VFAs) from five commercial bioplastics via acidogenic fermentation by a non-pretreated anaerobic sludge was investigated. Mesophilic, anaerobic, acidogenic batch assays at 1, 10 and 20 g/L feed concentrations revealed the feasibility of producing VFAs from polyhydroxyalkanoates (PHA), i.e., PHB and PHBV, but not from PBS, PCL and PLA under the test conditions and time. However, only high PHA substrate concentrations (10–20 g/L) resulted in organic overloading and decreasing the pH of the culture broth down to 4–5, which in turn induced the accumulation of VFAs via kinetic imbalance between acidogenesis and methanogenesis. Gaseous carbon (C-CO2 and C-CH4) accounted for 8–35% of the total initial carbon, while C-VFAs represented 10–18%, mainly as acetate and butyrate. This study represents the first systematically assessed proof-of-concept to produce VFAs from PHA, which is key for the design of bioplastic-to-bioplastic recycling (bio)technologies.Junta de Castilla y Leon - Fondo Europeo de Desarrollo Regional (grants CLU 2017-09, CL-EI-2021-07 and UIC 315

Screening Enzymes That Can Depolymerize Commercial Biodegradable Polymers: Heterologous Expression of <i>Fusarium solani</i> Cutinase in <i>Escherichia coli</i>

In recent years, a number of microbial enzymes capable of degrading plastics have been identified. Biocatalytic depolymerization mediated by enzymes has emerged as a potentially more efficient and environmentally friendly alternative to the currently employed methods for plastic treatment and recycling. However, the functional and systematic study of depolymerase enzymes with respect to the degradation of a series of plastic polymers in a single work has not been widely addressed at present. In this study, the ability of a set of enzymes (esterase, arylesterase and cutinase) to degrade commercial biodegradable polymers (PBS, PBAT, PHB, PHBH, PHBV, PCL, PLA and PLA/PCL) and the effect of pre-treatment methods on their degradation rate was assessed. The degradation products were identified and quantified by HPLC and LC-HRMS analysis. Out of the three enzymes, Fusarium solani cutinase (FsCut) showed the highest activity on grinded PBAT, PBS and PCL after 7 days of incubation. FsCut was engineered and heterologous expressed in Escherichia coli, which conferred the bacterium the capability of degrading solid discs of PBAT and to grow in PBS as the sole carbon source of the medium