Benny Palmqvist

The impact of pH coupled to process design for the conversion of the energy crop Arundo donax to ethanol was assessed in the present study under industrially relevant solids loadings. Two main process strategies were investigated, i.e. the traditional simultaneous saccharification and co-fermentation (SSCF) and a HYBRID design, where a long high temperature enzymatic hydrolysis step was carried out prior to continued low temperature SSCF, keeping the same total reaction time. Since acetic acid was identified as the major inhibitor in the slurry, the scenarios were investigated under different fermentation pH in order to alleviate the inhibitory effect on, in particular, xylose conversion. The results show that, regardless of fermentation pH, a higher glucan conversion could be achieved with the HYBRID approach compared to SSCF. Furthermore, it was found that increasing the pH from 5.0 to 5.5 for the fermentation phase had a large positive effect on xylose consumption for both process designs, although the SSCF design was more favored. With the high sugar concentrations available at the start of fermentation during the HYBRID design, the ethanol yield was reduced in favor of cell growth and glycerol production. This finding was confirmed in shake flask fermentations where an increase in pH enhanced both glucose and xylose consumption, but also cell growth and cell yield with the overall effect being a reduced ethanol yield. In conclusion this resulted in similar overall ethanol yields at the different pH values for the HYBRID design, despite the improved xylose uptake, whereas a significant increase in overall ethanol yield was found with the SSCF design.

Mixing is an energy demanding process which has been previously shown to affect enzymatic hydrolysis. Concentrated biomass slurries are associated with high and non-Newtonian viscosities and mixing in these systems is a complex task. Poor mixing can lead to mass and/or heat transfer problems as well as inhomogeneous enzyme distribution, both of which can cause possible yield reduction. Furthermore the stirring energy dissipation may impact the particle size which in turn may affect the enzymatic hydrolysis. The objective of the current work was to specifically quantify the effects of mixing on particle-size distribution (PSD) and relate this to changes in the enzymatic hydrolysis. Two rather different materials were investigated, namely pretreated Norway spruce and giant reed.

A common trend in the research on 2nd generation bioethanol is the focus on intensifying the process and increasing the concentration of water insoluble solids (WIS) throughout the process. However, increasing the WIS content is not without problems. For example, the viscosity of pretreated lignocellulosic materials is known to increase drastically with increasing WIS content. Further, at elevated viscosities, problems arise related to poor mixing of the material, such as poor distribution of the enzymes and/or difficulties with temperature and pH control, which results in possible yield reduction. Achieving good mixing is unfortunately not without cost, since the power requirements needed to operate the impeller at high viscosities can be substantial. This highly important scale-up problem can easily be overlooked. In this work, we monitor the impeller torque (and hence power input) in a stirred tank reactor throughout high solid enzymatic hydrolysis (< 20% WIS) of steam-pretreated Arundo donax and spruce. Two different process modes were evaluated, where either the impeller speed or the impeller power input was kept constant. Results from hydrolysis experiments at a fixed impeller speed of 10 rpm show that a very rapid decrease in impeller torque is experienced during hydrolysis of pretreated arundo (i.e. it loses its fiber network strength), whereas the fiber strength is retained for a longer time within the spruce material. This translates into a relatively low, rather WIS independent, energy input for arundo whereas the stirring power demand for spruce is substantially larger and quite WIS dependent. By operating the impeller at a constant power input (instead of a constant impeller speed) it is shown that power input greatly affects the glucose yield of pretreated spruce whereas the hydrolysis of arundo seems unaffected. The results clearly highlight the large differences between the arundo and spruce materials, both in terms of needed energy input, and glucose yields. The impact of power input on glucose yield is furthermore shown to vary significantly between the materials, with spruce being very affected while arundo is not. These findings emphasize the need for substrate specific process solutions, where a short pre-hydrolysis (or viscosity reduction) might be favorable for arundo whereas fed-batch might be a better solution for spruce.

When scaling up lignocellulose-based ethanol production, the desire to increase the final ethanol titer after fermentation can introduce problems. A high concentration of water-insoluble solids (WIS) is needed in the enzymatic hydrolysis step, resulting in increased viscosity, which can cause mass and heat transfer problems because of poor mixing of the material. In the present study, the effects of mixing on the enzymatic hydrolysis of steam-pretreated spruce were investigated using a stirred tank reactor operated with different impeller speeds and enzyme loadings. In addition, the results were related to the power input needed to operate the impeller at different speeds, taking into account the changes in rheology throughout the process.

Simultaneous saccharification and co-fermentation (SSCF) has been recognized as a feasible option for ethanol production from xylose-rich lignocellulosic materials. To reach high ethanol concentration in the broth, a high content of water-insoluble solids (WIS) is needed, which creates mixing problems and, furthermore, may decrease xylose uptake. Feeding of substrate has already been proven to give a higher xylose conversion than a batch SSCF. In the current work, enzyme feeding, in addition to substrate feeding, was investigated as a means of enabling a higher WIS content with a high xylose conversion in SSCF of a xylose-rich material. A recombinant xylose-fermenting strain of Saccharomyces cerevisiae (TMB3400) was used for this purpose in fed-batch SSCF experiments of steam-pretreated wheat straw. By using both enzyme and substrate feeding, the xylose conversion in SSCF could be increased from 40% to 50% in comparison to substrate feeding only. In addition, by this design of the feeding strategy, it was possible to process a WIS content corresponding to 11% in SSCF and obtain an ethanol yield on fermentable sugars of 0.35 g g-1. A combination of enzyme and substrate feeding was shown to enhance xylose uptake by yeast and increase overall ethanol yield in SSCF. This is conceptually important for the design of novel SSCF processes aiming at high-ethanol titers. Substrate feeding prevents viscosity from becoming too high and thereby allows a higher total amount of WIS to be added in the process. The enzyme feeding, furthermore, enables keeping the glucose concentration low, which kinetically favors xylose uptake and results in a higher xylose conversion.