Efficient bioconversion of crude glycerol from biodiesel to optically pured-lactate by metabolically engineered Escherichia coli

Abstract

Biodiesel has attracted considerable attention as one of the best choices among alternative and renewable fuels. Large quantities of crude glycerol are produced as a main co-product with increasing biodiesel production. Currently, the problem of waste glycerol utilization needs to be crucially addressed, not only for environmental protection but also for the economy of the biodiesel industry. In this paper, the use of crude glycerol for the production of D-lactate by engineered Escherichia coli was investigated. Engineered E. coli B0013-070 with a homolactic pathway for D-lactate synthesis by elimination of byproduct pathways (ethanol, succinate, formate and acetate) could convert 20 g L −1 of crude glycerol to 13.6 g L −1 of D-lactate with a yield of 0.67 g g −1 glycerol. Overexpression of D-lactate dehydrogenase by a low-copy vector in E. coli B0013-070 resulted in the increased production and yield of D-lactate, in which 14.5 g L −1 of D-lactate was produced with a yield of 0.72 g g −1 glycerol from crude glycerol. The effect of temperature on D-lactate fermentation by the engineered strain E. coli B0013-070-pTHldhA was also investigated, and 34 °C and 40 °C were found to be the optimal temperatures for cell growth and lactate production, respectively. The engineered strain B0013-070-pTHldhA produced 100.3 g L −1 of D-lactate with 99.97% optical purity from 531.5 g of crude glycerol with an overall productivity of 2.78 g L −1 h −1 and a yield of 75.4 g per 100 g glycerol (0.77 mol mol −1 ) using two phase fermentation combined with a temperature shifting strategy in a 7 L bioreactor. In summary, this paper shows that crude glycerol could be directly converted to D-lactate without any prior purification. † Electronic supplementary information (ESI) available. See

Metabolic engineering as a tool for enhanced lactic acid production

“…The laboratory model Escherichia coli has been used in a number of applications due to its ease of growth and well established methods of molecular and genetic manipulations [15][16][17][18]. However, E. coli requires metabolic engineering to direct pyruvate toward the single fermentation product of lactic acid ( Figure 2).…”

Synthesis of lactic acid from glycerol using a Pd/C catalyst

“…Chemical syntheses do not present pronounced problems, as there are several efficient methods to obtain the 2-hydroxypropanoic acid compared with the biotechnological ones [2,[5][6][7][8]. Among them, selective oxidation of glycerol to lactic acid at aqueous alkaline conditions at relatively low temperatures is feasible to resolve the timeconsuming of the process.…”

“…Conventionally available methods to produce lactic acid include carbohydrate fermentation, solar cells, and use of catalytic supports, sensors and fuel cells [1][2][3][4][5][6]. However, none of the abovementioned methods have met the environmental and technical goals due to potentially environmental impact, the relatively high cost of the raw materials and also the small-scale production [3].…”

Microbial production of lactic acid

“…Overexpression of ldhA increased final concentration and yield. Importantly, a two-phase approach allows different environmental conditions: for example, E. coli growth is favored at 34-37°C, while anaerobic conversion of glycerol into lactic acid favors 40°C (Chen et al 2014). The use of crude glycerol compared to purified glycerol does not appear to impact the production of lactic acid (Chen et al 2014), although methanol present as high as 28 % (w/w) is largely lost on autoclaving (Pyle et al 2008).…”

“…Moreover, the success of the conversion depends mostly on the fermentation, and in particular the ability of strains to convert completely high concentrations of glycerol. Another approach involves the typical two-phase process: using a twophase approach without overexpression of genes involved in glycerol uptake, E. coli was able to generate 99 g D-lactate l -1 in 36 h at a yield of 0.64 g/ g Chen et al 2014). Overexpression of ldhA increased final concentration and yield.…”