Ethanol Inhibition Kinetics of Kluyveromyces marxianus Grown on Jerusalem Artichoke Juice

The kinetics of ethanol inhibition on cell growth and ethanol production by Kluyveromyces marxianus UCD (FST) 55-82 were studied during batch growth. The liquid medium contained 10% (wt/vol) inulin-type sugars derived from an extract of Jerusalem artichoke (Helianthus tuberosus) tubers, supplemented with small amounts of Tween 80, oleic acid, and corn steep liquor. Initial ethanol concentrations ranging from 0 to 80 g/liter in the liquid medium were used to study the inhibitory effect of ethanol on the following parameters: maximum specific growth rate (μ_max), cell and ethanol yields, and sugar utilization. It was found that as the initial ethanol concentration increased from 0 to 80 g/liter, and maximum specific growth rate of K. marxianus cells decreased from 0.42 to 0.09 h⁻¹, whereas the ethanol and cell yields and sugar utilization remained almost constant. A simple kinetic model was used to correlate the μ_max results and the rates of cell and ethanol production, and the appropriate constants were evaluated.     

Modeling alcoholic fermentation of glucose/xylose mixtures by ethanologenic Escherichia coli as a function of pH

In order to understand the effect of pH on growth and ethanol production in ethanologenic Escherichia coli, we investigated the kinetic behavior of ethanologenic E. coli during alcoholic fermentation of glucose or xylose in a controlled pH environment and the fermentation of glucose, xylose, or their mixtures without pH control. Based on the Monod equation, an unstructured and unsegregated kinetic model was proposed as a function of the pH of the fermentation medium. The pH effects on cell growth, sugar consumption, and ethanol production were taken into account in the proposed model. Both cell growth and ethanol production were found to be significantly influenced by the pH of the fermentation medium. The optimal pH range for ethanol production by ethanologenic E. coli on either glucose or xylose was 6.0–6.5. The highest value of the maximum specific growth rate (μ _m) was obtained at pH 7.0. In the kinetic model of the fermentations of the sugar mixture, two inhibition terms related to glucose concentrations were included in both the cell growth and ethanol production equations because of the strong inhibitions of glucose and glucose metabolites on xylose metabolism. A good fit was found between model predictions and experimental data for both single-sugar and mixed-sugar fermentations without pH control within the experimental domain.     

Relationships between CO(2) Exchange Rates and Activities of Pyruvate,Pi Dikinase and Ribulose Bisphosphate Carboxylase, Chlorophyll Concentration, and Cell Volume in Maize Leaves

The relationships between CO₂-exchange rate (CER), DNA and chlorophyll (Chl) concentrations, pyruvate,Pi dikinase (PPDK) and ribulose bisphosphate carboxylase (RuBPCase) activities in ten maize (Zea mays L.) genotypes were investigated. The in vivo degrees of activation of PPDK and RuBPCase were estimated to make meaningful comparisons with CER. In leaves at a photosynthetic photon flux density (PPFD) of 720 micromoles per square meter per second, in vivo PPDK degree of activation was 80% of that of PPDK fully activated in vitro, whereas RuBPCase could not be further activated in vitro, suggesting that RuBPCase was fully activated in vivo. CER varied about 50% among the genotypes tested. Significant genetic differences were observed for the average weight of a cell (estimated by gram fresh weight per milligram DNA), but this character was not correlated with CER expressed on a fresh weight basis. CER was correlated with Chl concentration, and with estimates of the in vivo degree of activation of PPDK and RuBPCase. We concluded that in maize, CER is controlled by the metabolic components of photosynthesis rather than by membrane resistances to CO₂. If the latter factor were controlling CER, then smaller cells with higher amounts of exposed cell surface area per unit cell volume would have lower resistance to CO₂ diffusion, and therefore higher CER. When data were expressed on a DNA basis (proportional to a per cell basis), results indicated that larger cells (i.e. those with higher fresh weight per milligram DNA) have a higher content of Chl, and higher PPDK and RuBPCase activities, resulting in higher CER than in smaller cells.     

Tracing carbon monoxide uptake by Clostridium ljungdahlii during ethanol fermentation using 13C-enrichment technique

Conversion of synthesis gas (CO and H₂) to ethanol can be an alternative, promising technology to produce biofuels from renewable biomass. To distinguish microbial utilization of carbon source between fructose and synthesis gas CO and to evaluate biological production of ethanol from CO, we adopted the ¹³C-enrichment of the CO substrate and hypothesized that the residual increase in δ¹³C of the cell biomass would reflect the increased contribution of ¹³C-enriched CO. Addition of synthesis gas to live culture medium for ethanol fermentation by Clostridum ljungdahlii increased the microbial growth and ethanol production. Despite the high ¹³C-enrichment in CO (99 atom % ¹³C), however, microbial δ¹³C increased relatively small compared to the microbial growth. The uptake efficiency of CO estimated using the isotope mass balance equation was also very low: 0.0014 % for the low CO and 0.0016 % for the high CO treatment. Furthermore, the fast production of ethanol in the early stage indicated that the presence of sugar in fermentation medium would limit the utilization of CO as a carbon source by C. ljungdahlii.     

Influence of Environmental Factors and Medium Composition on Vibrio gazogenes Growth and Prodigiosin Production

Vibrio gazogenes ATCC 29988 growth and prodigiosin synthesis were studied in batch culture on complex and defined media and in chemostat cultures on defined medium. In batch culture on complex medium, a maximum growth rate of 0.75 h⁻¹ and a maximum prodigiosin concentration of 80 ng of prodigiosin · mg of cell protein⁻¹ were observed. In batch culture on defined medium, maximum growth rates were lower (maximum growth rate, 0.40 h⁻¹), and maximum prodigiosin concentrations were higher (1,500 ng · mg of protein⁻¹). In batch culture on either complex or defined medium, growth was characterized by a period of logarithmic growth followed by a period of linear growth; on either medium, prodigiosin biosynthesis was maximum during linear growth. In batch culture on defined medium, the initial concentration of glucose optimal for growth and pigment production was 3.0%; higher levels of glucose suppressed synthesis of the pigment. V. gazogenes had an absolute requirement for Na⁺; optimal growth occurred in the presence of 100 mM NaCl. Increases in the concentration of Na⁺ up to 600 mM resulted in further increases in the concentration of pigment in the broth. Prodigiosin was synthesized at a maximum level in the presence of inorganic phosphate concentrations suboptimal for growth. Concentrations of KH₂PO₄ above 0.4 mM caused decreased pigment synthesis, whereas maximum cell growth occurred at 1.0 mM. Optimal growth and pigment production occurred in the presence of 8 to 16 mg of ferric ion · liter⁻¹, with higher concentrations proving inhibitory to both growth and pigment production. Both growth and pigment production were found to decrease with increased concentrations of p-aminobenzoic acid. The highest specific concentration of prodigiosin (3,480 ng · mg protein⁻¹) was observed in chemostat cultures at a dilution rate of 0.057 h⁻¹. The specific rate of prodigiosin production at this dilution rate was approximately 80% greater than that observed in batch culture on defined medium. At dilution rates greater than 0.057 h⁻¹, the concentration of cells decreased with increasing dilution rate, resulting in a profile comparable to that expected for linear growth kinetics. No explanation could be found for the linear growth profiles obtained for both batch and chemostat cultures.     

Continuous high-ethanol fermentation from cane molasses by a flocculating yeast

Repeated-batch fermentation by a flocculating fusant, Saccharomyces cerevisiae HA 2, was done in a molasses medium that contained 20% (w/v) total sugar, at 30°C in an automatically controlled fermentor, and the effects of ethanol concentration on the specific growth rate and the specific production rate of ethanol were studied. Both the specific growth rate and the specific production rate of ethanol fell with increase of ethanol concentration, and there was a linear correlation between each rate and the concentration of thanol. The maximum specific growth rate (μ_max) and the maximum specific production rate of ethanol (q_max) were 0.12 h⁻¹ and 0.1 g ethanol/10⁹ cells·h, respectively. The specific growth rate and the specific production rate of ethanol fell to zero at ethanol concentration of 89 g/l and 95 g/l, respectively. The number of viable cells, calculated from the linear inhibition equation, was 1.3 × 10⁹ cells/ml for production of 85 g/l ethanol at a dilution rate (D₁) of 0.2 h⁻¹. Based on this estimation, a laboratory-scale continuous fermentation, using two fermentors in series, was done. In the second fermentor, 85 g/l ethanol was produced at a dilution rate (D₁) of 0.2 h⁻¹ by the active feedig of the fermented mash from the first fermentor into the second fermentor by pumping (hereafter called active feeding). To maintain the number of viable cells above 10⁹ cells/ml in the second fermentor, a active feeding ratio of more than 23% was required. Under these conditions, 81 g/l ethanol was produced in the second fermentor at a dilution rate (D_t) of 0.25 h⁻¹, and the high ethanol productivity of 20.3 g/l·h could be achieved. A bench-scale continuous fermentation, using two fermentors in series, with a active feeding ratio of 25% was done. An ethanol concentration of 84 g/l in the second fermentor at a dilution rate (D_t) of 0.25 h⁻¹ was achieved, just as it was in the laboratory-scale fermentation test.     

Cross-Feeding of Lactate Between Streptococcus lactis and Bacteroides sp. Isolated from Termite Hindguts

Streptococcus lactis and Bacteroides sp., isolated from hindguts of Reticulitermes flavipes termites, were grown anaerobically in monoculture and coculture. When grown in a glucose medium, S. lactis monoculture produced lactate as the major fermentation product, with small amounts of formate, acetate, ethanol, and CO₂. In coculture, glucose was completely consumed during growth of S. lactis. Lactate, produced by S. lactis, then supported much of the growth of Bacteroides and was fermented to propionate, acetate, and CO₂. Small amounts of succinate were formed during growth of Bacteroides in the coculture, but little change in the formate or ethanol concentration was observed. Monoculture growth of Bacteroides in a tryptone-yeast extract medium revealed that incorporation of 20 to 40 mM lactate increased cell yields and production of organic acids. However, initial lactate concentrations greater than 40 mM suppressed not only growth of Bacteroides but also acidic product formation. Results suggest that cross-feeding of lactate between streptococci and bacteroides constitutes one aspect of the overall hindgut fermentation in termites.     

A new process for red pigment production by submerged culture of Monascus purpureus

Formation of red pigment by Monascus purpureus via diauxic growth on glucose and ethanol in submerged culture was optimized based on inoculum preparation and culture medium. A vegetative inoculum was prepared from spores grown on ethanol. The optimized culture medium was low in phosphates, and had an initial pH?of 5.5. The characteristics of Monascus purpureus grown on glucose and on ethanol were compared: the specific consumption rate of glucose (q_G) was higher than the specific consumption rate of ethanol (q_E), whereas the specific growth rate was greatest with ethanol. The specific production rate of red pigment (p_OD) and pigment yield (Y_OD/s) with glucose was twice that with ethanol. A novel fermentation process was developed with M. purpureus initially grown with controlled ethanol formation, and consumption of the latter during pigment formation.     

Influence of Calcium Ion on Ethanol Tolerance of Saccharomyces bayanus and Alcoholic Fermentation by Yeasts

The addition of Ca²⁺ (as CaCl₂) in optimal concentrations (0.75 to 2.0 mM) to a fermentation medium with a trace contaminating concentration of Ca²⁺ (0.025 mM) led to the rapid production of higher concentrations of ethanol by Saccharomyces cerevisiae, Saccharomyces bayanus, and Kluyveromyces marxianus. The positive effect of calcium supplementation (0.75 mM) on alcoholic fermentation by S. bayanus was explained by the increase in its ethanol tolerance. The ethanol inhibition of growth and fermentation followed the equation μ_xi = μ_oi [1 - (X/X_mi)]ⁿⁱ, where μ_oi and μ_xi are, respectively, the specific growth (i = g) and fermentation (i = f) rates in the absence or presence of a concentration (X) of added ethanol, and X_mi is the maximal concentration of ethanol which allows growth or fermentation. The toxic power is given by n_i. In Ca²⁺ - supplemented medium (0.75 mM), n_g = 0.42 for growth and n_f = 0.43 for fermentation compared with 0.52 and 0.55, respectively, in unsupplemented medium; for both media, X_mg = 10% (vol/vol) and X_mf = 13% (vol/vol). For lethal concentrations of ethanol, the specific death rates were minimal for cells that were grown and incubated with ethanol in medium with an optimal concentration of Ca²⁺, maximal for cells grown and incubated with ethanol in unsupplemented medium, and intermediate for cells grown in unsupplemented medium and incubated with ethanol in calcium-supplemented medium. The effect of Ca²⁺ on the acidification curve of energized cells in the presence of ethanol was found to be closely associated with its protective effect on growth, fermentation, and viability.     

Effect of dilution rates on the amount of wasted substrate during alcohol fermentation

Continuous ethanol production in a one-stage continuous stirred tank fermentor without recycle was carried out using a yeast strain Saccharomyces cerevisiae. Different dilution rates were used. Cell and ethanol concentrations in the culture medium decreased with increasing dilution rates, and the maximum value of 3.0 g l⁻¹h⁻¹was found at a dilution rate of 0.340 h⁻¹. Specific ethanol productivities increased as dilution rates were increased, and the highest value appeared at about the same dilution rate as that for the maximum fermentor productivity. A material balance equation, which relates total amount of spent medium to cell synsthesis, ethanol production, and overall maintenance, was introduced. The cellular yield and overall maintenance coefficients increased with increasing dilution rates. The fraction of limiting substrate utilized for overall maintenance, which includes the limiting substrate spent for purposes other than cell synthesis and ethanol production, decreased with increasing dilution rates. The non-product associated substrate utilization can be minimized if correct dilution rate is chosen.     

The kinetics of Lagenidium giganteum growth in liquid and solid cultures

AIMS: Production of the mosquito biolarvacide Lagenidium giganteum in solid culture has been proposed as an economic alternative to production in liquid culture because of observations of improved shelf life and efficacy upon storage. Understanding the differences between these production systems and estimating growth rate in solid culture are important for commercialization. In order to address these needs a logistic model was developed to describe the growth kinetics of L. giganteum produced in solid and liquid cultures. METHODS AND RESULTS: Kinetic parameters in the logistic model were estimated by nonlinear regression of CO2 evolution rate (CER) and biomass data from solid and liquid cultivation experiments. Lagenidium giganteum biomass was measured using DNA extracted directly from samples. The logistic model was fit to experimental biomass and CER data with low standard errors for parameter estimates. The model was validated in two independent experiments by examining prediction of biomass using on-line CER measurements. CONCLUSIONS: There were significant differences between maximum biomass density, maintenance coefficients, and specific growth rates for liquid and solid cultures. The maximum biomass density (mg dw ml-1) was 11 times greater for solid cultivation compared with liquid cultivation of L. giganteum; however, the maintenance coefficient (mg CO2 h-1 (mg dw)-1) was six times greater for liquid cultivation than in solid cultivation. The specific growth rate at 30 degrees C was approximately 30% greater in liquid cultivation compared with solid cultivation. Slower depletion of substrate and lower endogenous metabolism may explain the longer shelf life of L. giganteum produced in solid culture. SIGNIFICANCE AND IMPACT OF THE STUDY: A simple logistic model was developed which allows real-time estimation of L. giganteum biomass from on-line CER measurements. Parameter estimates for liquid and solid cultivation models also elucidated observations of longer shelf life for production in solid culture.     

Relationship between pH and Medium Dissolved Solids in Terms of Growth and Metabolism of Lactobacilli and Saccharomyces cerevisiae during Ethanol Production

The specific growth rates of four species of lactobacilli decreased linearly with increases in the concentration of dissolved solids (sugars) in liquid growth medium. This was most likely due to the osmotic stress exerted by the sugars on the bacteria. The reduction in growth rates corresponded to decreased lactic acid production. Medium pH was another factor studied. As the medium pH decreased from 5.5 to 4.0, there was a reduction in the specific growth rate of lactobacilli and a corresponding decrease in the lactic acid produced. In contrast, medium pH did not have any significant effect on the specific growth rate of yeast at any particular concentration of dissolved solids in the medium. However, medium pH had a significant (P < 0.001) effect on ethanol production. A medium pH of 5.5 resulted in maximal ethanol production in all media with different concentrations of dissolved solids. When the data were analyzed as a 4 (pH levels) by 4 (concentrations of dissolved solids) factorial experiment, there was no synergistic effect (P > 0.2923) observed between pH of the medium and concentration of dissolved solids of the medium in reducing bacterial growth and metabolism. The data suggest that reduction of initial medium pH to 4.0 for the control of lactobacilli during ethanol production is not a good practice as there is a reduction (P < 0.001) in the ethanol produced by the yeast at pH 4.0. Setting the mash (medium) with ≥30% (wt/vol) dissolved solids at a pH of 5.0 to 5.5 will minimize the effects of bacterial contamination and maximize ethanol production by yeast.     

Indirect method for quantification of cellular biomass in a solidscontaining medium used as pre-culture for cellulase production

A process that combines the advantages of solid state fermentation (SSF) and submerged fermentation (SmF) could increase the efficiency of cellulase production required in the cellulosic ethanol industry. Due to the difficulty of measuring cellular biomass in the presence of solids, we developed a novel methodology for indirect quantification of biomass during production of the preculture for a combined fermentation process. Cultivation of Aspergillus niger was initiated as SSF using sugar cane bagasse as a solid substrate. Experiments were conducted in the absence of bagasse to determine growth kinetic parameters. Changes in glucose and biomass concentrations were measured. and the data were used for simulation employing a simple unstructured model. Parameters were estimated by applying a combination of Simulated Annealing (SA) and Levenberg-Marquardt (LM) algorithms to search for minimization of the error between model estimates and experimental data. Growth kinetics followed the Contois model, with a maximum specific growth rate (μ_max) of 0.042/h, a yield coefficient for biomass formation (Y_x/s) of 0.30 g/g and a death constant (k_D) of 0.005/h.These parameters were used to simulate cellular growth in the solids-containing medium. The proposed model accurately described the experimental data and succeeded in simulating the cell concentration profile. The selected pre-culture conditions (24 h as SSF followed by 48 h as SmF) were applied for cellulase production using the combined fermentation process and resulted in an endoglucanase activity (1,052 ± 34 U/L) greater than that obtained using the conventional SmF procedure (824 ± 44 U/L). Besides the standardization of pre-culture conditions, this methodology could be very useful in systems where direct measurement of cell mass is not possible.     

Effect of substrate loading on hydrogen production during anaerobic fermentation by Clostridium thermocellum 27405

We have investigated hydrogen (H₂) production by the cellulose-degrading anaerobic bacterium, Clostridium thermocellum. In the following experiments, batch-fermentations were carried out with cellobiose at three different substrate concentrations to observe the effects of carbon-limited or carbon-excess conditions on the carbon flow, H₂-production, and synthesis of other fermentation end products, such as ethanol and organic acids. Rates of cell growth were unaffected by different substrate concentrations. H₂, carbon dioxide (CO₂), acetate, and ethanol were the main products of fermentation. Other significant end products detected were formate and lactate. In cultures where cell growth was severely limited due to low initial substrate concentrations, hydrogen yields of 1 mol H₂/mol of glucose were obtained. In the cultures where growth ceased due to carbon depletion, lactate and formate represented a small fraction of the total end products produced, which consisted mainly of H₂, CO₂, acetate, and ethanol throughout growth. In cultures with high initial substrate concentrations, cellobiose consumption was incomplete and cell growth was limited by factors other than carbon availability. H₂-production continued even in stationary phase and H₂/CO₂ ratios were consistently greater than 1 with a maximum of 1.2 at the stationary phase. A maximum specific H₂ production rate of 14.6 mmol g dry cell⁻¹ h⁻¹ was observed. As cells entered stationary phase, extracellular pyruvate production was observed in high substrate concentration cultures and lactate became a major end product.     

Photosynthesis and photorespiration in presenescent, senescent, and rejuvenated soybean cotyledons

Various growth and physiological parameters were measured in germinating, presenescent, and senescing soybean (Glycine max [L.] Merr.) cotyledons and in cotyledons rejuvenated by epicotyl removal 18 days after planting. The maximal measured carbon dioxide exchange rates (CER) in the cotyledons were in the range of those reported for field-grown soybean leaves. Rejuvenated cotyledons accumulated total chlorophyll in excess of the maximum observed in presenescent cotyledons. When photosynthetic rates were expressed per cotyledon, the CER in rejuvenated tissue recovered to the maximal rates observed in presenescent cotyledons. Ribulose-1,5-bisphosphate carboxylase/oxygenase in rejuvenated cotyledons also recovered to the maximal amount seen in presenescent cotyledons so that CER appeared to be a function of ribulose-1,5-bisphosphate carboxylase/oxygenase content during most of the period studied. Observations of the postillumination outburst of CO₂ and ¹⁴C label in glycine indicated that photorespiration was occurring in the cotyledons and that photorespiration relative to photosynthesis was different in rejuvenated compared with presenescent cotyledons.     

Effect of mixed carbon substrate on exopolysaccharide production of cyanobacterium Nostoc flagelliforme in mixotrophic cultures

The effects of organic carbon sources on cell growth and exopolysaccharide (EPS) production of dissociated Nostoc flagelliforme cells under mixotrophic batch culture were investigated. After 7?days of cultivation, glycerol, acetate, sucrose, and glucose increased the final cell density and final EPS concentrations, and mixotrophic growth achieved higher biomass concentrations. The increase in cell growth was particularly high when glucose was added as the sole carbon source. On the other hand, EPS production per dry cell weight was significantly enhanced by adding acetate. For more effective EPS production, the effects of the mixture of glucose and acetate were investigated. Increasing the ratio of glucose to acetate resulted in higher growth rate with BG-11 medium and higher EPS productivity with BG-11₀ medium (without NaNO₃). When the medium was supplemented with a mixture of glucose (4.0?g?L^?1) and acetate (2.0?g?L^?1), 1.79?g?L^?1 biomass with BG-11 medium and 879.6?mg?L^?1 of EPS production with BG-11₀ medium were achieved. Adopting this optimal ratio of glucose to acetate established in flask culture, the culture was also conducted in a 20-L photobioreactor with BG-11 medium for 7?days. A maximum biomass of 2.32?g?L^?1 was achieved, and the EPS production was 634.6?mg?L^?1.     

Adaptation of Saccharomyces cerevisiae cells to high ethanol concentration and changes in fatty acid composition of membrane and cell size

Background

Microorganisms can adapt to perturbations of the surrounding environment to grow. To analyze the adaptation process of the yeast Saccharomyces cerevisiae to a high ethanol concentration, repetitive cultivation was performed with a stepwise increase in the ethanol concentration in the culture medium.

Methodology/Principal Findings

First, a laboratory strain of S. cerevisiae was cultivated in medium containing a low ethanol concentration, followed by repetitive cultivations. Then, the strain repeatedly cultivated in the low ethanol concentration was transferred to medium containing a high ethanol concentration and cultivated repeatedly in the same high-ethanol-concentration medium. When subjected to a stepwise increase in ethanol concentration with the repetitive cultivations, the yeast cells adapted to the high ethanol concentration; the specific growth rate of the adapted yeast strain did not decrease during repetitive cultivation in the medium containing the same ethanol concentration, while that of the non-adapted strain decreased during repetitive cultivation. A comparison of the fatty acid composition of the cell membrane showed that the contents in oleic acid (C_18:1) in ethanol-adapted and non-adapted strains were similar, but the content of palmitic acid (C_16:0) in the ethanol-adapted strains was lower than that in the non-adapted strain in media containing ethanol. Moreover, microscopic observation showed that the mother cells of the adapted yeast were significantly larger than those of the non-adapted strain.

Conclusions

Our results suggest that activity of cell growth defined by specific growth rate of the yeast cells adapted to stepwise increase in ethanol concentration did not decrease during repetitive cultivation in high-ethanol-concentration medium. Moreover, fatty acid content of cell membrane and the size of ethanol-adapted yeast cells were changed during adaptation process. Those might be the typical phenotypes of yeast cells adapted to high ethanol concentration. In addition, the difference in sizes of the mother cell between the non-adapted and ethanol strains suggests that the cell size, cell cycle and adaptation to ethanol are thought to be closely correlated.     

Rapid ethanol fermentation of cellulose hydrolysate. II. Product and substrate inhibition and optimization of fermentor design

High concentrations of both ethanol and sugar in the fermentation broth inhibit the growth of yeast cells and the rate of product formation. Inhibitory effects of ethanol on the yeast strain Saccharomyces cerevisiae NRRL-Y-132 were studied in batch and continuous chemostat cultures. Growth was limited by either glucose or ethanol. Feed medium was supplemented with different ethanol concentrations. Ethanol was found to inhibit growth and the activity of yeast to produce ethanol in a noncompetitive manner. A linear kinetic pattern for growth and product formation was observed according to μ = μ_m (1 – P/P_m) and v = v_m (1 – P/P_m′), where μ_m is the maximum specific growth rate at P = 0 (hr^?1); P_m is the maximum specific product formation rate at P = 0 (hr^?1); P_m is the maximum ethanol concentration above which cells do not grow (g/liter); P_m′ is the maximum ethanol concentration above which cells do not produce ethanol (g/liter). Substrate inhibition studies were carried out using short-time experimental techniques under aerobic and anaerobic condition. The degree of substrate inhibition was found to be higher than that has been reported for ethanol fermentation of pure sugar. The kinetic relationships thus obtained were used to compute growth, substrate utilization, and alcohol production patterns and have been discussed with reference to batch and continuous fermentation of enzymatically produced bagasse hydrolysate.     

Effect of Nystatin on the Metabolism of Xylitol and Xylose by Pachysolen tannophilus

Ethanol production from xylitol by resting cells of Pachysolen tannophilus was increased 40-fold in the presence of nystatin, amphotericin B, and filipin, a group of antifungal agents that alter the permeability of the plasma membrane. Furthermore, these agents had little or no effect on ethanol formation from xylitol or xylose by the cell extract. During xylose metabolism, nystatin caused the intracellular xylitol to leak out into the medium at a 23-fold-faster rate but did not affect overall xylose utilization and CO₂ evolution. These observations explain the rate of xylitol utilization by cell extract being higher than that by whole cells (J. Xu and K. B. Taylor, Appl. Environ. Microbiol. 59:231-235, 1993) as well as the relative inability of P. tannophilus to utilize xylitol to support significant ethanol production and cell growth.     

Short-Term Effects of CO(2) on Gas Exchange of Leaves of Bigtooth Aspen (Populus grandidentata) in the Field

The short term effects of increased levels of CO₂ on gas exchange of leaves of bigtooth aspen (Populus grandidentata Michx.) were studied at the University of Michigan Biological Station, Pellston, MI. Leaf gas exchange was measured in situ in the upper half of the canopy, 12 to 14 meters above ground. In 1900 microliters per liter CO₂, maximum CO₂ exchange rate (CER) in saturating light was increased by 151% relative to CER in 320 microliters per liter CO₂. The temperature optimum for CER shifted from 25°C in 320 microliters per liter CO₂ to 37°C in 1900 microliters per liter CO₂. In saturating light, increasing CO₂ level over the range 60 to 1900 microliters per liter increased CER, decreased stomatal conductance, and increased leaf water use efficiency. The initial slope of the CO₂ response curve of CER was not significantly different at 20 and 30°C leaf temperatures, although the slope did decline significantly during leaf senescence. In 1900 microliters per liter CO₂, CER increased with increasing light. The light saturation point and maximum CER were higher in 30°C than in 20°C, although there was little effect of temperature in low light. The experimental results are consistent with patterns seen in laboratory studies of other C₃ species and define the parameters required by some models of aspen CER in the field.