Fermentation of biodiesel-derived crude glycerol to 1,3-propanediol with bio-wastes as support matrices: Polynomial prediction model.

Biodiesel production from used cooking oil is sustainable alternative, for bio-energy production. The process generates residual crude glycerol (RCG) as the major energy-rich waste which can be used to produce various bio-based chemicals like 1,3-propanediol (1,3-PDO) through biotechnological interventions. This RCG contains several impurities like methanol, soap, organic materials, salts non-transesterified fatty acids and metals in varied concentrations. These impurities significantly affect yield and productivity of the bio-process due to their marked microbial toxicity. In this work, previously isolated Clostridium butyricum L4 was immobilized on various abundantly available cheap bio-wastes (like rice straw, activated carbon and corn cob) to explore advantages offered and improve tolerance to various feed impurities. Amongst these, shredded rice straw was found most suitable candidate for immobilization and results in maximum improvement in 1,3-PDO production (18.4%) with highest porosity (89.28%), lowest bulk density (194.48Kg/m3), and highest cellular biofilm density (CFU/g-8.4 ×1010) amongst the three matrices. For practical purposes, recyclability was evaluated and it was concluded that even after reusing for five successive cycles the production retained to ∼82.4%. Subsequently, polynomial model was developed using 30 runs central composite factorial design experiments having coefficient of regression (R²) as 0.9520, in order to predict yields under different immobilization conditions for 1,3-PDO production. Plackett-Burman was employed (Accuracy= 99.17%) to screen significant toxic impurities. Based on statistical analysis six impurities were found to be significantly influential on PDO production in adverse manner. With negative coefficient of estimate (COE) varying in decreasing order: Linoleic acid >Oleic acid >Stearic acid >NaCl>K2SO4 >KCl. The study illustrates practical application for repurposing waste glycerol generated from biodiesel plants, thus developing improved agnostic process along with yield production models.

Fermentative reforming of crude glycerol to 1,3-propanediol using Clostridium butyricum strain L4.

Repurposed used cooking oil is a sustainable alternative to other feedstocks for biodiesel production offering enviro-economic benefits. Residual crude glycerol (RCG) from such biodiesel production plants is difficult to utilize due to presence of numerous toxic impurities with various inhibitory effects on biological fermentative reforming process. However, it is a new industrial feedstock for bio-based production of 1,3-propanediol. In this work, a new Clostridium butyricum strain L4 was isolated from biogas reactor leachate after rigorous adaption and 35 subcultures under increasing stress conditions and studied for green production of 1,3-propanediol (PDO) from RCG and further process development. Evaluation of fermentative reforming kinetics was performed and the optimal reaction conditions are pH 7.0, temperature 30 °C, 2 g yeast extract/L and 15 g ammonium sulphate/L. Glycerol-glucose co-fermentation (10:1) enhanced cell growth and thus, PDO output by 11.6 g/L. In comparison to batch fermentation (24.8 g PDO/L; 0.58 mol PDO/mol glycerol) there was 2.8-fold improvement with fed-batch process resulting in accumulation of 70.1 g PDO/L (Yield = 0.65 mol PDO/mol glycerol) using the studied biocatalyst in 150 h. In order to predict yields under different operational conditions a multiple linear regression model was developed (r2 = 0.783) with six independent variables (p < 0.05), where biomass (g/L) and temperature (oC) were forecasted as top contributors to PDO yield. Finally, this biocatalyst appears as a potential candidate for industrial use due to its non-pathogenic nature, ability to grow in wide pH and temperature conditions, tolerance to high substrate and product concentration, insignificant generation of by-products and Coenzyme B12 independent biotransformation. The study can add value to bio-utilization of RCG to produce green 1,3-propanediol.