The long-term effects of ethanol on immobilized cell reactor performance using K. fragilis

The effects of ethanol on reactor performance were studied in a small, 5-cm packed height, "differential" type immobilized cell reactor. Lactose utilizing yeast cells, Kluyveromyces fragilis, were absorbed to a porous adsorbant sponge matrix in a gas continuous reactor. Step changes in the feed ethanol concentration to the column (10-130 g/L) were used to test the reactor response over extended periods of time (about 30-50 h per dosage level) followed by a return to basal zero inlet ethanol feed. Effluent cell density and effluent cell viability were measured at intervals. An inhibitory response in ethanol productivity to feed dosage ethanol levels above 20 g/L was detected almost immediately, with a near steady state response noted within 2.5 h of initiating the dosage. Feed ethanol levels above 50 g/L resulted in a subsequent gradual decrease in reactor productivity over time, which was associated with a decrease in the fraction of viable shed cells in the reactor effluent. The reactor response to a step removal of the ethanol inhibition was also monitored. Quick and complete rebounding of the fermentation rate to the original basal rate was noted following dosage concentrations of under 50 g/L ethanol. Recovery rates slowed following ethanol dosage levels above 50 g/L. Viable shed cell density improved overtime during the slow recovery periods. Growth rates (as determined by shed cell density) were more strongly inhibited than productivity. Growth responded more slowly to changes in ethanol environment as growth rates at 30 h fell to about 40% of the rates measured 7.5 h after initiation of a dosage level. It is concluded that ethanol contributions to cell injury and death (and consequent ICR performance degradation) may be more important than ethanol inhibition of productivity rates in the long-term operation of immobilized cell reactors at ethanol concentrations over 50 g/L.     

Filament formation and ethanol production by Zymomonas mobilis in adsorbed cell bioreactors

Zymomonas mobilis immobilized on microporous ion exchange resins has previously been shown to allow the attainment of high ethanol productivities in packed-bed bioreactors. The formation of bacterial filaments after several days of continuous operation, however, had resulted in excessive pressure increases across the reactor bed. The present work examines techniques for controlling filament formation by Z. mobilis in two reactor sizes (161 mL and 7.85 L) and a feed glucose concentration of 100 g/L. By controlling the fermentation temperature at 20-25 degrees C it has been possible to eliminate filament formation by Z. mobilis and to operate the larger bioreactor for 232 h with an ethanol productivity of 50 g/L h (based on total reactor volume). The rate of ethanol production has been shown to be very sensitive to temperature in the range 20-30 degrees C, and it is likely that slightly higher temperatures than those used in this study will improve ethanol productivity while still permitting long-term operation.     

Effect of particle loading on GM-CSF production by Saccharomyces cerevisiae in a three-phase fluidized bed bioreactor

Continuous production of a recombinant murine granulocyte-macrophage colony-stimulating factor (MuGM-CSF) by immobilized yeast cells, Saccharomyces cerevisiae strain XV2181 (a/a, Trp1) containing plasmid palphaADH2, in a fluidized bed bioreactor was studied at a 0.03 h(-1) dilution rate and various particle loading rates ranging from 5% to 33% (v/v). Cells were immobilized on porous glass beads fluidized in an air-lift draft tube bioreactor. A selective medium containing glucose was used to start up the reactor. After reaching a stable cell concentration, the reactor feed was switched to a rich, nonselective medium containing ethanol as the carbon source for GM-CSF production. GM-CSF production increased initially and then dropped gradually to a stable level. During the same period, the fraction of plasmid-carrying cells declined continuously to a lower level, depending on the particle loading. The relatively stable GM-CSF production, despite the large decline in the fraction of plasmid-carrying cells, was attributed to cell immobilization. As the particle loading rate increased, the plasmid stability also increased. Also, as the particle loading increased from 5% to 33%, total cell density in the bioreactor increased from 16 to 36 g/L, and reactor volumetric productivity increased from 0.36 to 1.31 mg/L.h. However, the specific productivity of plasmid-carrying cells decreased from 0.55 to 0.07 mg/L.g cell. The decreased specific productivity at higher particle loading rates was attributed to reduced growth efficiency caused by nutrient limitations at higher cell densities. Both the reactor productivity and specific cell productivity increased by two- to threefold or higher when the dilution rate was increased from 0.03 to 0.07 h(-1). (c) 1996 John Wiley & Sons, Inc.     

Continuous ethanol production from pineapple cannery waste using immobilized yeast cells

The cells of Saccharomyces cerevisiae ATCC 24553, were immobilized in k-carrageenan and packed in a tapered glass column reactor for ethanol production from pineapple cannery waste at temperature 30 degrees C and pH 4.5. The maximum productivity was 42.8 g ethanol 1(-1) h(-1) at a dilution rate of 1.5 h(-1). The volumetric ethanol productivity of the immobilized cells was ca. 11.5 times higher than the free cells. The immobilized cell reactor was operated over a period of 87 days at a dilution rate of 1.0 h(-1), without any loss in the immobilized cell activity. The maximum specific ethanol productivity and specific sugar uptake rate of the immobilized cells were 1.2 g ethanol g(-1) dry wt. cell h(-1) and 2.6 g sugar g(-1) dry wt. cell h(-1), respectively, at a dilution rate of 1.5 h(-1).     

An immobilized cell reactor with simultaneous product separation. I. Reactor design and analysis

A four-phase reactor-separator (gas, liquid, solid, and immobilized catalyst) is proposed for fermentations characterized by a volatile product and nonvolatile substrate.In this reactor, the biological catalyst is immobilized onto a solid column packing and contacted by the liquid containing the substrate.A gas phase is also moved through the column to strip the volatile product into the gas phase. The Immobilized Cell Reactor-Separator (ICRS) consists of two basic gas-liquid flow sections: a cocurrent "enricher" followed by a countercurrent-"stripper".In this article, an equilibrium stage model of the reactor is developed to determine the feasibility and important operational variables of such a reactor-separator. The ICRS concept is applied to the ethanol from whey lactose fermentation using some preliminary immobilized cell reactor performance data. A mathematical model for a steady-state population based on an adsorbed monolayer of cells is also developed for the reactor. The ICRS model demonstrated that the ICRS should give a significant increase in reactor productivity as compared to an identically sized Immobilized Cell Reactor (ICR) with no separation. The gas-phase separation of the product also allows fermentation of high inlet substrate concentrations. The model is used to determine the effects of reactor parameters on ICRS performance including temperature, pressure, gas flow rates, inlet substrate concentration, and degree of microbial product inhibition.     

Novel,linked bioreactor system for continuous production of biologics

A novel, alternative intensified cell culture process comprised of a linked bioreactor system is presented. An N-1 perfusion bioreactor maintained cells in a highly proliferative state and provided a continuous inoculum source to a second bioreactor operating as a continuous-flow stirred-tank reactor (CSTR). An initial study evaluated multiple system steady-states by varying N-1 steady-state viable cell densities, N-1 to CSTR working volume ratios, and CSTR dilution rates. After identifying near optimum system steady-state parameters yielding a relatively high volumetric productivity while efficiently consuming media, a subsequent lab-scale experiment demonstrated the startup and long-term operation of the envisioned manufacturing process for 83 days. Additionally, to compensate for the cell-specific productivity loss over time due to cell line instability, the N-1 culture was also replaced with younger generation cells, without disturbing the steady-state of the system. Using the model cell line, the system demonstrated a two-fold volumetric productivity increase over the commercial-ready, optimized fed-batch process.     

Ethanol fermentation in an immobilized cell reactor using Saccharomyces cerevisiae

Fermentation of sugar by Saccharomyces cerevisiae, for production of ethanol in an immobilized cell reactor (ICR) was successfully carried out to improve the performance of the fermentation process. The fermentation set-up was comprised of a column packed with beads of immobilized cells. The immobilization of S. cerevisiae was simply performed by the enriched cells cultured media harvested at exponential growth phase. The fixed cell loaded ICR was carried out at initial stage of operation and the cell was entrapped by calcium alginate. The production of ethanol was steady after 24 h of operation. The concentration of ethanol was affected by the media flow rates and residence time distribution from 2 to 7 h. In addition, batch fermentation was carried out with 50 g/l glucose concentration. Subsequently, the ethanol productions and the reactor productivities of batch fermentation and immobilized cells were compared. In batch fermentation, sugar consumption and ethanol production obtained were 99.6% and 12.5% v/v after 27 h while in the ICR, 88.2% and 16.7% v/v were obtained with 6 h retention time. Nearly 5% ethanol production was achieved with high glucose concentration (150 g/l) at 6 h retention time. A yield of 38% was obtained with 150 g/l glucose. The yield was improved approximately 27% on ICR and a 24 h fermentation time was reduced to 7 h. The cell growth rate was based on the Monod rate equation. The kinetic constants (K(s) and mu(m)) of batch fermentation were 2.3 g/l and 0.35 g/lh, respectively. The maximum yield of biomass on substrate (Y(X-S)) and the maximum yield of product on substrate (Y(P-S)) in batch fermentations were 50.8% and 31.2% respectively. Productivity of the ICR were 1.3, 2.3, and 2.8 g/lh for 25, 35, 50 g/l of glucose concentration, respectively. The productivity of ethanol in batch fermentation with 50 g/l glucose was calculated as 0.29 g/lh. Maximum production of ethanol in ICR when compared to batch reactor has shown to increase approximately 10-fold. The performance of the two reactors was compared and a respective rate model was proposed. The present research has shown that high sugar concentration (150 g/l) in the ICR column was successfully converted to ethanol. The achieved results in ICR with high substrate concentration are promising for scale up operation. The proposed model can be used to design a lager scale ICR column for production of high ethanol concentration.     

Minimal nutritional requirements for immobilized yeast

The effect of reduced nutritional levels (particularly nitrogen source) for immobilized K. fragilis type yeast were studied using a trickle flow, "differential" plug flow type reactor with cells immobilized by adsorption onto an absorbant packing matrix. Minimizing nutrient levels in a feed stream to an immobilized cell reactor (ICR) might have the benefits of reducing cell growth and clogging problems in the ICR, reducing feed preparation costs, as well as reducing effluent disposal costs. In this study step changes in test feed medium nutrient compositions were introduced to the ICR, followed by a return to a basal medium. Gas evolution rates were monitored and logged on a continuous basis, and effluent cell density was used as an indicator of cell growth rate of the immobilized cell mass. Startup of the reactor using a YEP medium showed a rapid buildup of cells in the reactor during the initial 110 h operation. The population density then stabilized at 1.6 x 10(11) cells/g sponge. A defined medium containing a complex mix of essential nutrients with an inorganic nitrogen source (ammonium sulfate) was able to maintain 90% of the productivity in the ICR as compared to the YEP medium, but proved unable to promote growth of the immobilized cell mass during startup. Experiments on reduced ammonium sulfate in the defined medium, and reduced yeast extract and peptone in YEP medium indicated that stable productivity could be maintained for extended periods (80 h) in the complete absence of any nutrients besides a few salts (potassium phosphate and magnesium sulfate). It was found that productivity rates dropped by 35-65% from maximal values as nitrogenous nutrients were eliminated from the test mediums, while growth rates (as determined by shed cell density from the reactor) dropped by 75-95%. Thus, nutritional deficiencies largely decoupled growth and productivity of the immobilized yeast which suggests productivity is both growth- and non-growth-associated for the immobilized cells. A yeast extract concentration of 0.375 g/L with or without 1 g/L ammonium sulfate was determined to be the minimum level which gave a sustained increase in productivity rates as compared to the nutritionally deficient salt medium. This represents a 94% reduction in complex nitrogenous nutrient levels compared to standard YEP batch medium (3 g/L YE and 3.5 g/L peptone).     

Continuous propionate production from whey permeate using a novel fibrous bed bioreactor

Continuous production of propionate from whey lactose by Propionibacterium acidipropionici immobilized in a novel fibrous bed bioreactor was studied. In conventional batch propionic acid fermentation, whey permeate without nutrient supplementation was unable to support cell growth and failed to give satisfactory fermentation results for over 7 days. However, with the fibrous bed bioreactor, a high fermentation rate and high conversion were obtained with plain whey permeate and de-lactose whey permeate. About 2% (wt/vol) propionic acid was obtained from a 4.2% lactose feed at a retention time of 35 to 45 h. The propionic acid yield was approximately 46% (wt/vol) from lactose. The optimal pH for fementation was 6.5, and lower fermentation rates and yields were obtained at lower pH values. The optimal temperature was 30 degrees C, but the temperature effect was not dramatic in the range of 25 to 35 degrees C. Addition of yeast extract and trypticase to whey permeate hastened reactor startup and increased the fermentation rate and product yields, but the addition was not required for long-term reactor performance. The improved fermentation results with the immobilized cell bioreactor can be attributed to the high cell density, approximately 50 g/L, attained in the bioreactor, Cells were immobilized by loose attachement to fiber surfaces and entrapment in the void spaces within the fibrous matrix, thus allowing constant renewal of cells. Consequently, this bioreactor was able to operate continuously for 6 months without encountering any clogging, degeneration, or contamination problems. Compared to conventional batch fermentors, the new bioreactor offers many advantages for industrial fermentation, including a more than 10-fold increase in productivity, acceptance of low-nutrient feedstocks such as whey permeate, and resistance to contamination. (c) 1994 John Wiley & Sons, Inc.     

Acetate addition to an immobilized yeast column of ethanol production

Acetate, a by-product of ethanol fermentation by Saccharomyces cerevisiae, has been shown to inhibit cell growth if present in high concentrations. Consequently, acetate has been considered undesirable in systems where the production rate depends upon steady-state growth. Acetate, however, may be desirable in some systems since it increases the specific rate of ethanol production by increasing the maintenance requirements of yeast. In immobilized cell reactors using the crosslinking method, steady state is not achieved and cell overgrowth is a problem. This article presents the results of a study aimed at taking advantage of the use of acetate, both to reduce cell overgrowth and to increase productivity. Various concentrations of acetate were added to batch and plug flow systems, while monitoring the effects on cell growth and ethanol production. The productivity was increased by as much as 50% in an immobilized cell reactor (ICR), while cell growth was greatly reduced.     

Ethanol production by immobilized whole cells ofZymomonas mobilis in a continuous flow expanded bed bioreactor and a continuous flow stirred tank bioreactor

Continuous ethanol fermentation by immobilized whole cells ofZymomonas mobilis was investigated in an expanded bed bioreactor and in a continuous stirred tank reactor at glucose concentrations of 100, 150 and 200 g L^–1. The effect of different dilution rates on ethanol production by immobilized whole cells ofZymomonas mobilis was studied in both reactors. The maximum ethanol productivity attained was 21 g L^–1 h^–1 at a dilution rate of 0.36 h^–1 with 150 g glucose L^–1 in the continuous expanded bed bioreactor. The conversion of glucose to ethanol was independent of the glucose concentration in both reactors.     

Development of a method for immobilization of non-flocculating cells in loofa (Luffa cylindrica) sponge

Both the matrix structure of loofa sponge and the flocculating property of cells were necessary for efficient immobilization. The addition of chitosan to a reactor containing a bed of loofa sponge and a Candida brassicae cell suspension induced cell flocculation and the cells were efficiently immobilized. During ethanol production by the immobilized cells, the free cell concentration in the broth was controlled at the desired level by intermittent addition of chitosan to the reactor. The immobilized cell concentration increased but their specific ethanol productivity decreased with an increase in the chitosan concentration. The maximum ethanol productivity was obtained at a low chitosan concentration of 0·03 g/litre. With this optimal concentration, the cell concentration, ethanol yield and productivity were, respectively, 2, 1·3 and 3 times higher than those of the suspension culture.     

Acetic acid production from lactose by an anaerobic thermophilic coculture immobilized in a fibrous-bed bioreactor

An anaerobic thermophilic coculture consisting of a heterofermentative bacterium (Clostridium thermolacticum) and a homoacetogen (Moorella thermoautotrophica) was developed for acetic acid production from lactose and milk permeate. The fermentation kinetics with free cells in conventional fermentors and immobilized cells in a recycle batch fibrous-bed bioreactor were studied. The optimal conditions for the cocultured fermentation were found to be 58 degrees C and pH 6.4. In the free-cell fermentation, C. thermolacticum converted lactose to acetate, ethanol, lactate, H(2) and CO(2), and the homoacetogen then converted lactate, H(2), and CO(2) to acetate. The overall acetate yield from lactose ranged from 0.46 to 0.65 g/g lactose fermented, depending on the fermentation conditions. In contrast, no ethanol was produced in the immobilized-cell fermentation, and the overall acetate yield from lactose increased to 0.8-0.96 g/g lactose fermented. The fibrous-bed bioreactor also gave a higher final acetate concentration (up to 25. 5 g/L) and reactor productivity (0.18-0.54 g/L/h) as compared to those from the free-cell fermentation (final acetate concentration, 15 g/L; productivity, 0.06-0.08 g/L/h). The superior performance of the fibrous-bed bioreactor was attributed to the high cell density (20 g/L) immobilized in the fibrous-bed and adaptation of C. thermolacticum cells to tolerate a higher acetate concentration. The effects of yeast extract and trypticase as nutrient supplements on the fermentation were also studied. For the free-cell fermentation, nutrient supplementation was necessary for the bacteria to grow in milk permeate. For the immobilized-cell fermentation, plain milk permeate gave a high acetate yield (0.96 g/g), although the reactor productivity was lower than those with nutrient supplementation. Balanced growth and fermentation activities between the two bacteria in the coculture are important to the quantitative conversion of lactose to acetic acid. Lactate and hydrogen produced by C. thermolacticum must be timely converted to acetic acid by the homoacetogen to avoid inhibition by these metabolites.     

High ethanol productivities using small Ca-alginate beads of immobilized cells of Zymomonas mobilis

Summary Zymomonas mobilis cells were immobilized into small 1 mm diameter beads of Ca-alginate in order to minimize mass transfer limitations and maximize immobilized cell activity. A combination of small bead size with a high cell concentration of 58 g dry wt. cell per lit. bead volume resulted in high ethanol productivities using a newly designed packed bed bioreactor system. Steady-state dilution rates ranging from 0.4 h^-1 to 3.9 h^-1 were run resulting in a maximum productivity of 102 g ethanol/l/h for an inlet substrate concentration of 100 g glu/l and 87% conversion. The bioreactor was run continuously at a fixed dilution rate for 384 h and short intermittent treatment of the beads with CaCl₂ temporarily increased ethanol productivity to a maximum of 116 g ethanol/l/h.     

Long-term Continuous Production of Monoclonal Antibody by Hybridoma Cells Immobilized in a Fibrous-Bed Bioreactor

The kinetics and long-term stability of continuous production of monoclonal antibody IgG2b by hybridoma HD-24 cells immobilized in a fibrous-bed bioreactor (FBB) were studied for a period of ～8 months. The cells were immobilized in the fibrous bed by surface attachment of cells and entrapment of large cell clumps in the void space of the fibrous matrix. A high viable cell density of 1.01 × 10⁸/ml was attained in the bioreactor, which was about 63 times higher than those in conventional T-flask and spinner flask cultures. The continuous FBB produced IgG at a concentration of ～0.5 g/l, with reactor productivity of ～7 mg/h·l, which was about 23 times higher than those from conventional T-flask and spinner flask cultures. The IgG concentration can be further increased to ～0.67 g/l by using higher feed (glucose and glutamine) concentrations and running the reactor at a recycle batch or fed-batch mode. The long-term performance of this bioreactor was also evaluated. For a period of 36 days monitored, the MAb produced in the continuous well-mixed bioreactor at 50 h retention time (0.02/h dilution rate) was maintained at a steady concentration level of ～0.3 g/l with less than 8% drift. At the end of the study, it was found that ～25% of the cells were strongly attached to the fiber surfaces and the other ～75% entrapped or weakly immobilized in the fibrous matrix. The strongly attached cells had a high viability of ～90%, compared to ～75% for cells weakly immobilized and only ～1.4% for freely suspended cells, suggesting that the fibrous matrix preferentially retained and protected the viable (productive) cells. The FBB thus was able to maintain its long-term productivity because nonviable and dead cells were continuously washed off from the fibrous matrix. The high MAb concentration and production rate and excellent stability for continuous long-term production obtained in this study compare favorably to other bioreactor studies reported in the literature. The reactor performance can be further improved by providing better pH and aeration controls at higher feed concentrations. The FBB is easy to operate and scale-up, and thus can be used economically for industrial production of MAb.     

Cell separator operation within temperature ranges to minimize effects on Chinese hamster ovary cell perfusion culture

A cell retention device that provides reliable high-separation efficiency with minimal negative effects on the cell culture is essential for robust perfusion culture processes. External separation devices generally expose cells to periodic variations in temperature, most commonly temperatures below 37 degrees C, while the cells are outside the bioreactor. To examine this phenomenon, aliquots of approximately 5% of a CHO cell culture were exposed to 60 s cyclic variations of temperature simulating an acoustic separator environment. It was found that, for average exposure temperatures between 31.5 and 38.5 degrees C, there were no significant impacts on the rates of growth, glucose consumption, or t-PA production, defining an acceptable range of operating temperatures. These results were subsequently confirmed in perfusion culture experiments for average exposure temperatures between 31.6 and 38.1 degrees C. A 2(5-1) central composite factorial design experiment was then performed to systematically evaluate the effects of different operating variables on the inlet and outlet temperatures of a 10L acoustic separator. The power input, ambient temperature, as well as the perfusion and recycle flow rates significantly influenced the temperature, while the cell concentration did not. An empirical model was developed that predicted the temperature changes between the inlet and the outlet of the acoustic separator within +/-0.5 degrees C. A series of perfusion experiments determined the ranges of the significant operational settings that maintained the acoustic separator inlet and outlet temperatures within the acceptable range. For example, these objectives were always met by using the manufacturer-recommended operational settings as long as the recirculation flow rate was maintained above 15 L day(-1) and the ambient temperature was near 22 degrees C.     

Use of ATP to characterize biomass viability in freely suspended and immobilized cell bioreactors

This work describes investigations into the viability of cells growing on 3,4-dichloroaniline (34DCA). Two bioreactors are employed for microbial growth, a continuous stirred tank (CST) bioreactor with a 2-L working volume, and a three-phase air lift (TPAL) bioreactor with a 3-L working volume. Experiments have been performed at several dilution rates between 0.027 and 0.115 h(-1) in the CST bioreactor and between 0.111 and 0.500 h(-1) in the TPAL bioreactor. The specific ATP concentration was calculated at each dilution rate in the suspended biomass in both bioreactors as well as in the immobilized biomass in the TPAL bioreactor. The ATP was extracted from the cells using boiling tris-EDTA buffer (pH 7.75), and the quantity determined using a firefly (bioluminescence) technique. The cultures were inspected under an electron microscope to monitor compositional changes. Results from the CST bioreactor showed that the biomass-specific ATP concentration increases from 0.44 to 1.86 mg ATP g(-1) dry weight (dw) as dilution rate increases from 0.027 to 0.115 h(-1). At this upper dilution rate the cells were washed out. The specific ATP concentration reached a limiting average value of 1.73 mg ATP g(-1) dw, which is assumed to be the quantity of ATP in 100% viable biomass. In the TPAL bioreactor, the ATP level increased with dilution rate in both the immobilized and suspended biomass. The specific ATP concentration in the immobilized biomass increased from approximately 0.051 mg ATP g(-1) dw at dilution rates between 0.111 and 0.200 h(-1) to approximately 0.119 mg ATP g(-1) dw at dilution rates between 0.300 and 0.500 h(-1). This indicates that the immobilized biomass contained a viable cell fraction of around 5%. Based on these results, kinetic data for freely suspended cells should not be applied to the modeling of immobilized cell systems on the assumption that immobilized biomass is 100% viable. (c) 1993 John Wiley & Sons, Inc.     

Ethanol production from starch in a pervaporation membrane bioreactor using Clostridium thermohydrosulfuricum

To attain both high productivity and efficient recovery of ethanol from broth, a membrane bioreactor consisting of a jar fermentor and a pervaporation system was applied to the direct production of ethanol from uncooked starch with a thermophilic anaerobic bacterium, Clostridium thermohydrosulfuricum. From four types of ethanol-selective membranes tested, microporous polytetrafluoroethylene (PTFE) membrane, the pores of which are impregnated with silicone rubber, was chosen for its large flux, high ethanol selectivity, and high stability. During fed-batch fermentation with pervaporation in the membrane bioreactor, ethanol was continuously extracted and concentrated in two traps with concentrations at 5.6%-6.2% (w/w) in trap 1 (20 degrees C) and 27%-32% (w/w) in trap 2 (liquid N(2)), while the ethanol concentration in the broth was maintained at 0.85-0.9% (w/w). Due to the low ethanol concentration in the broth, and the immobilization of bacterial cells by the membrane, the number of viable cells, and, eventually, the ethanol productivity, increased in the membrane bioreactor.     

Continuous ethanol production from Jerusalem artichoke tubers. II. Use of immobilized cells of Kluyveromyces marxianus

Kluyveromyces marxianus UCD (FST) 55-82 cells were immobilized in Na alginate beads and used in a packed-bed bioreactor system for the continuous production of ethanol from the extract of Jerusalem artichoke tubers. Volumetric ethanol productivities of 104 and 80 g ethanol/ L/h were obtained at 80 and 92% sugar utilization, respectively. The maximum volumetric ethanol productivity of the immobilized cell bioreactor system was found to be 15 times higher than that of an ordinary-stirred-tank (CST) bioreactor using cells of K. marxianus. The immobilized cell bioreactor system was operated continuously at a constant dilution rate of 0.66 h(-1) for 12 days resulting in only an 8% loss of the original immobilized cell activity, which corresponds to an estimated half-life of ca. 72 days. The maximum specific ethanol productivity and maximum specific sugar uptake rate of the immobilized cells were found to be 0.55 g ethanol/g/biomass/h and 1.21 g sugars/g biomass/h, respectively.     

Continuous ethanol production from concentrated wood hydrolysates in an internal membrane-filtration bioreactor

Continuous culture for the production of ethanol from wood hydrolysate was carried out in an internal membrane-filtration bioreactor. The hydrolysate medium was sterilized at a relatively low temperature of 60 degrees C with the intention of reducing the formation of inhibitory compounds during the sterilization. The maximum ethanol concentration and productivity obtained in this study were 76.9 g/L and 16.9 g/L-h, respectively, which were much higher than those (57.2-67 g/L and 0.3-1.0 g/L-h) obtained in batch cultures using hydrolysate media sterilized at 60 degrees C. The productivity was also found to be much higher than that (6.7 g/L-h) obtained in a continuous cell retention culture using a wood hydrolysate sterilized at 121 degrees C. These results show that the internal membrane-filtration bioreactor in combination with low-temperature sterilization could be very effective for ethanol production from wood hydrolysate.