Dihydroxyacetone production from glycerol using Gluconobacter oxydans: Study of medium composition and operational conditions in shaken flasks

Abstract

The production of dihydroxyacetone from glycerol employing aerobic cultures of Gluconobacter oxydans is studied. Dihydroxyacetone is one of the most important value‐added products obtained from glycerol, a by‐product of biodiesel production. The effect of organic nitrogen source and initial substrate concentrations has been studied together with the possibility of product inhibition. Afterward, the influence of the main operating conditions (temperature, shaking speed, and initial biomass concentration) on in vivo glycerol dehydrogenase activity has also been considered. The results show no evidence of glycerol inhibition, but an important product inhibition was detected, which has been taken into account in a kinetic model for enzymatic activity description. In terms of operating conditions, pH was found to exert a great impact on glycerol conversion, being necessary to keep it above 4 to ensure complete glycerol conversion. The minimum temperature that maximized enzymatic activity was found to be 30°C. In addition, a surprising decoupling between biomass concentration and dihydroxyacetone production rate was observed when adding increasing nitrogen source concentrations at a fixed shaking speed. Glycerol dehydrogenase activity remains constant despite the increase in biomass concentration, contrary to what would be expected. This fact revealed the existence of a rate limiting factor, identified subsequently as oxygen transfer rate depending on the biomass concentration.

Kinetic Modelling of the Coproduction Process of Fumaric and Malic Acids by Rhizopus arrhizus NRRL 1526

“…Due to the morphological characteristics of the used fungus, which develops hyphae, biomass concentration cannot be measured by turbidity techniques, such as in bacterial bioprocesses [24][25][26], so biomass quantification must be done by dry weight cell determination. To this end, a working protocol called "one flask, one sample" was developed.…”

“…A first question to be answered concerns the number of previous stages that must be done previously to the production of the acid. On certain bacterial processes, it is known that the use of two previous stages (called inoculum and pre-inoculum) can be beneficial for reaching a proper metabolic state and, as a consequence, the desired product concentration is maximized [24][25][26]. In Figure 5, it can be seen how this procedure is not applicable to this fungal process.…”

Kinetic Modeling of Dihydroxyacetone Production from Glycerol by Gluconobacter oxydans ATCC 621 Resting Cells: Effect of Fluid Dynamics Conditions

“…The most widely employed microorganism to produce DHA from glycerol as raw material is Gluconobacter oxydans, although there are several different works using other microorganisms [5][6][7][8][9][10][11][12][13][14][15][16]. G. oxydans is a Gram-negative aerobic bacterium, whose respiratory metabolism allows it to partially oxidize different organic compounds [17].…”

“…Other authors also describe a strong substrate and product inhibition on DHA production [12,13,15,22]. Furthermore, Dikshit et al employed various different kinetic models in order to describe DHA production considering the effect of substrate inhibition [23] and substrate and product combined inhibition [24].…”

“…In order to achieve this objective, several experiments were carried out both in Erlenmeyer flasks and STBR, varying the agitation speed. Operating conditions were fixed, taking into account the results achieved in previous works [15,16]. Throughout the study, resting cells were employed in operation mode.…”