Improved sugar co-utilisation by encapsulation of a recombinant Saccharomyces cerevisiae strain in alginate-chitosan capsules.

Johan O Westman,Mohammad J Taherzadeh,Nicklas Bonander,Carl Johan Franzén

doi:10.1186/1754-6834-7-102

Abstract

BackgroundTwo major hurdles for successful production of second-generation bioethanol are the presence of inhibitory compounds in lignocellulosic media, and the fact that Saccharomyces cerevisiae cannot naturally utilise pentoses. There are recombinant yeast strains that address both of these issues, but co-utilisation of glucose and xylose is still an issue that needs to be resolved. A non-recombinant way to increase yeast tolerance to hydrolysates is by encapsulation of the yeast. This can be explained by concentration gradients occuring in the cell pellet inside the capsule. In the current study, we hypothesised that encapsulation might also lead to improved simultaneous utilisation of hexoses and pentoses because of such sugar concentration gradients.ResultsIn silico simulations of encapsulated yeast showed that the presence of concentration gradients of inhibitors can explain the improved inhibitor tolerance of encapsulated yeast. Simulations also showed pronounced concentration gradients of sugars, which resulted in simultaneous xylose and glucose consumption and a steady state xylose consumption rate up to 220-fold higher than that found in suspension culture. To validate the results experimentally, a xylose-utilising S. cerevisiae strain, CEN.PK XXX, was constructed and encapsulated in semi-permeable alginate-chitosan liquid core gel capsules. In defined media, encapsulation not only increased the tolerance of the yeast to inhibitors, but also promoted simultaneous utilisation of glucose and xylose. Encapsulation of the yeast resulted in consumption of at least 50% more xylose compared with suspended cells over 96-hour fermentations in medium containing both sugars. The higher consumption of xylose led to final ethanol titres that were approximately 15% higher. In an inhibitory dilute acid spruce hydrolysate, freely suspended yeast cells consumed the sugars in a sequential manner after a long lag phase, whereas no lag phase was observed for the encapsulated yeast, and glucose, mannose, galactose and xylose were utilised in parallel from the beginning of the cultivation.ConclusionsEncapsulation of xylose-fermenting S. cerevisiae leads to improved simultaneous and efficient utilisation of several sugars, which are utilised sequentially by suspended cells. The greatest improvement is obtained in inhibitory media. These findings show that encapsulation is a promising option for production of second-generation bioethanol.

Highlights

Two major hurdles for successful production of second-generation bioethanol are the presence of inhibitory compounds in lignocellulosic media, and the fact that Saccharomyces cerevisiae cannot naturally utilise pentoses
Finite element modelling shows increased co-consumption due to encapsulation Mathematical modelling was implemented to visualise the effect of mass transport resistances on the concentration profiles of glucose, xylose, HMF and furfural in a capsule, and to analyse the sensitivity of the sugar consumption rates to different characteristics of the system
The base case model of the diffusion and reaction rates resulted in very low concentrations of glucose and furfural in large parts of the cell pellet (Figure 1)

Summary

Introduction

Two major hurdles for successful production of second-generation bioethanol are the presence of inhibitory compounds in lignocellulosic media, and the fact that Saccharomyces cerevisiae cannot naturally utilise pentoses. One of the main problems is the issue of inhibitors produced during the pretreatment and hydrolysis of the raw material into fermentable sugars Another problem is the fact that pentoses are not fermentable by wild-type Saccharomyces cerevisiae [2], there are a few exceptional cases of slow consumption [3]. Xylose will not be consumed in considerable amounts until the concentration of glucose is low [6,7] The reason for this is that there are no specific pentose transporters in S. cerevisiae, and pentoses are instead transported by the native hexose transporters [8,9]. These transporters have higher affinity for glucose, and there is a strong preference for glucose uptake as long as it is present

Methods

Results

Conclusion