Mannonic Acid and Bio-Ethanol Production from Konjac Using a Two-Step Bioprocess with Candida Shehatae and Gluconobacter Oxydans

Amorphophallus konjac is rich in glucomannan,which can be hydrolyzed into glucose and mannose,thereby acting as an economic raw material for the acquisition of glucose and mannose.The total sugar yield was 91.2%when konjac powder was treated with 0.75%hydrochloric acid at 121℃for 1 h.Thus,dilute acid hydrolysates of konjac powder were used as a carbon source for obtaining value-added products.Here we showed that the microbial production of ethanol and mannonic acid was obtained by employing Candida shehatae(C.shehatae)and Gluconobacter oxydans(G.oxydans).Through a step-by-step bioprocess,glucose is the first selectively converted to ethanol by C.shehatae,which enables G.oxydans-mediated biocatalysis of mannose to mannonic acid.Finally,approximately 100 g ethanol and 340 g mannonic acid were produced starting from 1 kg refined konjac powder.The results demonstrated the feasibility of this bioconversion method for producing mannonic acid starting from crude hydrolysates of konjac powder.

Investigating the Interaction Between Gluconobacter Oxydans and Bacillus M egaterium for 2-keto-L-gulonic Acid Biosynthesis in the Two-Step Vitamin C Fermentation

In the two-step vitamin C fermentation process, its precursor 2-keto-L-gulonic acid was synthesized from L-sorbose by mixed culture of Gluconobacter oxydans and Bacillus megaterium. The interaction between Gluconobacter oxydans and Bacillus megaterium remains unclear and it is a challenge to mathematically model the mixed growth of these two strains. The Monod-type equations were previously proposed to describe the coupled growth of Gluconobacter oxydans and Bacillus megaterium. However, in this study, we modeled the interaction of these two strains in a macroscopic view by introducing the population theory. Taking account of the fact that the density or concentration of Gluconobacter oxydans or Bacillus megaterium was hardly to measure accurately in the mixed culture broth, the data of concentrations of the substrate and product were used to indirectly investigate the relation between these two strains. Three batch experiments were used to validate our model. And according to the values of identified parameters, the type of interaction between Gluconobacter oxydans and Bacillus megaterium was concluded to be predation, where Gluconobacter oxydans was predator, and Bacillus megaterium was prey.

Enhanced production of L-sorbose by systematic engineering of dehydrogenases in Gluconobacter oxydans

L-Sorbose is an essential intermediate for the industrial production of vitamin C(L-ascorbic acid).However,the formation of fructose and some unknown by-products significantly reduces the conversion ratio of D-sorbitol to L-sorbose.This study aimed to identify the key D-sorbitol dehydrogenases in Gluconobacter oxydans WSH-003 by gene knockout.Then,a total of 38 dehydrogenases were knocked out in G.oxydans WSH-003,and 23 dehydrogenase-deficient strains could increase L-sorbose production.G.oxydans-30,wherein a pyrroloquinoline quinone-dependent glucose dehydrogenase was deleted,showed a significant reduction of a by-product with the extension of fermentation time.In addition,the highest conversion ratio of 99.60%was achieved in G.oxydans MD-16,in which 16 different types of dehydrogenases were inactivated consecutively.Finally,the gene vhb encoding hemoglobin was introduced into the strain.The titer of L-sorbose was 298.61 g/L in a 5-L bioreactor.The results showed that the systematic engineering of dehydrogenase could significantly enhance the production of L-sorbose.

Study on the Effect of Mutated Bacillus megaterium in Two-Stage Fermentation of Vitamin C

Bacillus megaterium as a companion strain in two-stage fermentation of vitamin C could secrete some active substances to spur growth of Gluconobacter oxydans to produce 2-KLG. In the fermenting system where Gluconobacter oxydans was combined with GB82-a mutated strain of B. megaterium by ion implantation, the amount of 2-KLG harvested was larger than that produced by the original B. megaterium BP52 being substituted for GB82. In this paper, we studied the effect of the active substances secreted by GB82 to enhance the capability of Gluconobacter oxydans to produce 2-KLG. The supernate of GB82 sampled at different cultivation times all had much more activity to spur Gluconobacter oxydans to yield 2-KLG than that of the original B. megaterium, which might be due to the genetic changes in the active components caused by ion implantation. Furthermore, the active substances of GB82's supernate would lose a part of its activity in extreme environments, which is typical of some proteins.