Showing papers by "Bärbel Hahn-Hägerdal published in 1984"

Enzymatic hydrolysis of sodium-hydroxide-pretreated sallow in an ultrafiltration membrane reactor.

Abstract: An ultrafiltration membrane reactor was used to investigate the recovery of biocatalysts during enzymatic hydrolysis of pretreated sallow. Product inhibition could be eliminated by continuous removal of products through the ultrafiltration membrane, thus retaining the macromolecular substrate and enzymes. In this way, the degree of conversion was improved from 40% in a batch hydrolysis to 95% (within 20 h), and the initial hydrolysis rate was increased up to seven times. The recovery studies were focused on mechanical deactivation and irreversible adsorption on to the nonconvertible fraction of the substrate. Cellulase deactivation during mechanical agitation was not significant, and the loss of activity was attributed mainly to strong adsorption of the enzymes onto undigested material. This process was studied in semicontinuous hydrolyses, where fresh substrate was added intermittently. The amount of reducing sugars produced in this experiment was 25.7 g/g enzyme, compared to 4.7 g/g enzyme in a batch hydrolysis.

An enzyme coimmobilized with a microorganism: The conversion of cellobiose to ethanol using β‐glucosidase and Saccharomyces cerevisiae in calcium alginate gels

Abstract: The efficiency of ..beta..-glucosidase and Saccharomyces cerevisiae in directly converting cellobiose to ethanol was studied for various combinations of the two catalytic species, both free and immobilized, in order to elucidate the advantages of using a coimmobilized system. The coimmobilized preparation was superior to a combination of separately immobilized biocatalysts. However, in this preparation, one-half the enzyme activity was lost within a week when incubated at the operational temperature in the absence of substrate. In continuous experiments, an 80% conversion of cellobiose to ethanol was obtained using the coimmobilized preparation, compared to 40% using separately immobilized biocatalysts when applying a dilution rate of 0.1 h/sup -1/ in a packed bed reactor. The immobilized biocatalysts showed no decline in productivity during two weeks of continous operation.

Metabolic Behavior of Immobilized Cells—Effects of Some Microenvironmental Factorsa

Abstract: Variations in microenvironmental conditions have been demonstrated to be important on the subcellular level. Model studies on immobilized enzymes have helped to clarify the picture. However, when cells have been immobilized, changed metabolic behavior has been reported in several cases, but very little is known about the mechanisms behind these changes. The new environment created in the immobilization process differs from that in free solution, for example, by the presence of high concentrations of polymers and by impeded diffusion of reactants (nutrients). The presence of polymer can be important in the sense that these molecules can directly interact with the cells, but probably the most overlooked effect is that polymers cause a change in water activity of the medium. When setting up a system to study the effects of the microenvironment on the metabolic behavior of cells, it may be useful to simplify the system. Suspensions of cells in water solutions of polymers represent such a simplified model. In the present study, polyethylene glycol (PEG) was used to study the effects of a decreased water activity on cell metabolism, thus simulating the behavior of immobilized cells. In order to prepare suspensions with a defined water activity, methods for calculating the reduction in water activity for buffered polymer-rich media have been developed.’ The water activity is defined as:

Solid Superacids for Hydrolyzing Oligo‐ and Polysaccharidesa

Abstract: The current interest in utilizing carbohydrates as raw materials for producing chemicals and fuels via various fermentation routes has focused interest on cheap and efficient hydrolysis processes for the production of fermentable sugars. The hydrolysis of cellulose, especially, has received much attention, since cellulose is the most abundant source of renewable carbohydrates.' The present technology essentially offers two ways for cellulose hydrolysis: acid and enzymatic. The former is fast but has the drawback of low yields. Furthermore, the corrosion problem has not yet been fully solved. The latter can give theoretical yields, but is slow and subject to severe product inhibition, particularly when the intermediate cellobiose accumulates. This paper evaluates a third possibility: the utilization of solid superacids, perfluorinated resin-sulfonic acid catalysts, NAFIONR 501, generously supplied by E. I. du Pont de Nemours & Company, Wilmington, DE. Solid superacids have been successfully utilized in a number of reactions such as alkylations2 and esterifi~ations.~ Their utilization in hydrolysis resembles an acid hydrolysis. but the fact that they are solid makes it possible to remove them as well as to operate the hydrolysis process continuously in a plug-flow-type reactor. The hydrolysis of cellulose, starch, and cellobiose were first compared in a batch reaction (FIG. 1). The production of glucose from cellobiose and starch followed zero-order kinetics. No tendency for product inhibition was observed. However, cellulose was barely hydrolyzed. The difference in the results between starch and cellulose is probably due to the fact that starch is gelatinized a t the experimental temperature used, 9SoC, thus opening up the molecules to the catalyst more than for cellulose. In this way, steric hindrances imposed by both substrate and catalyst appear to prevent the hydrolysis of cellulose with solid superacids. The manufacturer recommends that the solid superacids be used in the presence of sodium chloride in order to enhance the reaction rate. In repeated batch hydrolysis experiments, it was found that the presence of sodium chloride released hydrogen ions into the medium, the effect being that the superacids lost their catalytic activity after

Shifting Product Formation from Xylitol to Ethanol in Pentose Fermentation with Candida tropicalis by Alteringa

Abstract: One problem in the utilization of lignocellulosic biomass for the production of liquid fuels such as ethanol is the fermentation of the hydrolyzed pentosan fraction, which can constitute as much as 37% of the raw material‘ and is composed mainly of the pentose, xylose. Several solutions for this problem have been suggested: the enzymatic conversion of xylose to xylulose followed by fermentation with Succharomyces cerevisiae,2 the use of thermophilic anaerobic bacteria such as Thermoanaerobacter ethanolicus’ which can directly ferment xylose to ethanol, or the use of several yeast strains, for example, Cundidu tropicalis, which under “semiaerobic” conditions are also capable of carrying out this p r o c e ~ s . ~ Each of these routes, however, has a flaw of one kind or another. For example, xylose isomerase is of bacterial origin and has a pH and temperature optimum different from yeast, making a one-step process difficult. T. ethanolicus is substrate-inhibited, resulting in product concentrations too low for economical use. Finally, the xylose-fermenting yeasts tend to produce xylitol as a major by-product, thus not utilizing xylose efficiently. This paper deals with the latter problem. Using Candida tropicalis ATCC 321 13 (generously supplied by AC Biotechnics, Arlov, Sweden), an attempt was made to shift product formation from xylitol to ethanol by altering aeration levels, with the use of the respiratory inhibitor, azide, and by decreasing the water activity in the medium. “Semiaerobic” (or oxygen-limited) conditions have been found to enhance ethanol production from xylose in some yeast strains.’ However, xylitol production is also enhanced in an oxygen-limited culture.6 Azide has previously been shown to enhance ethanol production in S. cerevisiue’ and the same was observed with a decreased water activity.* TABLE 1 summarizes the results with C. tropicalis. In agreement with previous reports, the yield of ethanol is highest under “semiaerobic” conditions, 50% higher than under aerobic conditions. However, the xylitol production increases as well under these conditions. The semiaerobic conditions were chosen for further experiments with azide and with decreased water activities because C. tropicalis functions poorly anaerobically (little or no growth and poor utilization of xylose). Azide was tested at two concentrations. 0.2 m M and 4.6 m M (TABLE 1). At the higher concentration, cell growth was inhibited and only 30% of the available xylose

Aqueous Two-Phase Systems for Producing α-Amylase Using Bacillus subtilisa

Abstract: Since water is biocompatible, aqueous two-phase systems are suitable for use with biological materials. These systems have been used successfully for large-scale separations and purifications of intracellular enzymes.',* It is also possible to culture living microbial cells,'-s as well as to perform enzymatic conversions6 in aqueous two-phase systems where the product is later isolated from one of the two phases. The present communication describes the utilization of an aqueous two-phase system for the production of a-amylase using Bacillus subtilis. FIGURE 1 shows a photograph of B. subtilis cells partitioned in a droplet of the dextran-rich bottom phase in a poly(ethy1ene glycol) (PEG)/dextran phase system. All cells were found in the bottom phase, which facilitates the upgrading of a product (a-amylase) from the cell-free top phase. For the production of a-amylase, an aqueous two-phase system was selected in which the partition coefficient, K (enzymatic activity in the top phase over enzymatic activity in the bottom phase) for n-amylase was 0.5. The composition of the phase system was: