Effects of controlled gas environments in solid-substrate fermentations of rice

The gas environment is solid-substrate fermentations of rice significantly affected levels of biomass and enzyme formation by a fungal species screened for high amylase production. Constant oxygen and carbon dioxide partial pressures were maintained at various levels in fermentations by Aspergillus oryzae. Control of the gas phase was maintained by a “static” aeration system admitting oxygen on demand and stripping excess carbon dioxide during fermentation. Constant water vapor pressures were also maintained by means of saturated salt solutions. High Oxygen pressures stimulated amylase productivity significantly. On the other hand, amylase production was severely inhibited at high carbon dioxide pressures. While relatively insensitive to oxygen pressure, maximum biomass productivities were obtained at an intermediate carbon dioxide pressure. High oxygen transfer rates were obtained at elevated oxygen pressures, suggesting, in view of the stimulatory effect of oxygen on amylase production, a stringent oxygen requirement for enzyme synthesis. Solid-substrate fermentations were highly advantageous as compared with submerged cultures in similar gas environments. Not only were amylase productivities significantly higher, but the enzyme was highly concentration in the aqueous phase of the semisolid substrate particles and could be extracted in a small volume of liquid. Results of this work suggest that biomass and product formation in microbial processes may be amenable to control by the gas environment. This is believed to offer an interesting potential for optimizing selected industrial fermentation processes with respect to productivity and energy consumption.     

A comparison of carbon dioxide production and oxygen uptake in sediment cores from four south Swedish lakes

Carbon dioxide production and oxygen uptake were measured in undisturbed sediment cores taken during winter from four lakes of different trophic state. Respiration was measured at 5, 10, 15 and 20°C at high oxygen saturation (75–100%). The respiratory quotient, calculated from the mean values of carbon dioxide production and oxygen uptake at each temperature for each lake, was 0.83–0.96 with a mean value for the four lakes of 0.90. At very low oxygen saturations (<10%) carbon dioxide production was 21–42% of the production at 20°C and high oxygen saturations. The results indicate that under aerobic conditions, oxygen uptake and carbon dioxide production are closely-coupled processes in these lake sediments.     

THE EFFECT OF LOW PRESSURES ON CELL OXIDATION

1. A method is described for measuring tissue oxidation under reduced barometric pressure. 2. The oxygen uptake of yeast is diminished by low barometric pressures to a greater extent than by a reduction of the partial pressure of oxygen, to a corresponding degree, at atmospheric pressure. 3. This effect of low pressure is not observed with certain in vitro oxidation systems. 4. The anaerobic respiration (carbon dioxide production) of yeast is not at all affected by low pressures. 5. The inhibition of tissue oxidation caused by carbon monoxide is removed by lowering the pressure.     

THE RESPIRATION OF LUMINOUS BACTERIA AND THE EFFECT OF OXYGEN TENSION UPON OXYGEN CONSUMPTION

1. The respiration of luminous bacteria has been studied by colorimetric and manometric methods. 2. Limulus oxyhaemocyanin has been used as a colorimetric indicator of oxygen consumption and indicator dyes were used for colorimetric determination of carbon dioxide production. 3. The Thunberg-Winterstein microrespirometer has been used for the measurement of the rate of oxygen consumption by luminous bacteria at different partial pressures of oxygen. 4. The effect of oxygen concentration upon oxygen consumption has been followed from equilibrium with air to low pressures of oxygen. 5. Luminous bacteria consume oxygen and produce carbon dioxide independent of oxygen pressures from equilibrium with air (152 mm.) to approximately 22.80 mm. oxygen or 0.03 atmosphere. 6. Dimming of a suspension of luminous bacteria occurs when oxygen tension is lowered to approximately 2 mm. Hg (0.0026 atmosphere) and when the rate of respiration becomes diminished one-half. 7. Pure nitrogen stops respiratory activity and pure oxygen irreversibly inhibits oxygen consumption. 8. The curve for rate of oxygen consumption with oxygen concentration is similar to curves for adsorption of gasses at catalytic surfaces, and agrees with the Langmuir equation for the expression of the amount of gas adsorbed in unimolecular layer at catalytic surfaces with gas pressure. 9. A constant and maximum rate of oxygen consumption occurs in small cells when oxygen concentration becomes sufficient to entirely saturate the surface of the oxidative catalyst of the cell.     

The mode of action of insecticidal controlled atmospheres

Arthropods cope with reduced oxygen and elevated carbon dioxide atmospheres with a reduction in metabolic rate, also called metabolic arrest. The reduction in metabolism lessens the pressure on the organism to initiate anaerobic metabolism, but also leads to a reduction in ATP production. The natural permeability of cellular membranes appears to be important for the survival of the arthropod under low oxygen or high carbon dioxide atmospheres. Despite the similarities in response, arthropod mortality is generally greater in response to high carbon dioxide as apposed to low oxygen atmospheres. There appears to be a greater decrease in ATP and energy charge in arthropods exposed to high carbon dioxide as compared with low oxygen atmospheres, and this may be due to greater membrane permeability under carbon dioxide leading to an inefficient production of ATP. Reduced oxygen and elevated carbon dioxide atmospheres can have an additive effect in some cases, depending on the concentrations used. The effect of these atmospheres on arthropods depends also on temperature, species and life stage. Additional work is needed to fully understand the mode of action of controlled atmospheres on arthropod pests.     

Development of Haemonchus contortus in vitro and the stimulus from the host.

Infective larvae of Haemonchus contortus which have been exposed for about 9 hr to relatively high partial pressures of carbon dioxide at near-neutral pH, as they might be in the rumen, develop slowly to the fourth stage or not at all. But following this treatment, exposure to lower partial pressures of carbon dioxide and higher concentrations of hydrogen ions, such as might be encountered in the abomasal mucosa, brings about development at a rate comparable with that in the sheep.     

Gas Exchange of Algae: II. Effects of Oxygen, Helium, and Argon on the Photosynthesis of Chlorella pyrenoidosa

Changes in the oxygen partial pressure of air over the range of 8 to 258 mm of Hg did not adversely affect the photosynthetic capacity of Chlorella pyrenoidosa. Gas exchange and growth measurements remained constant for 3-week periods and were similar to air controls (oxygen pressure of 160 mm of Hg). Oxygen partial pressures of 532 and 745 mm of Hg had an adverse effect on algal metabolism. Carbon dioxide consumption was 24% lower in the gas mixture containing oxygen at a pressure 532 mm of Hg than in the air control, and the growth rate was slightly reduced. Oxygen at a partial pressure of 745 mm of Hg decreased the photosynthetic rate 39% and the growth rate 37% over the corresponding rates in air. The lowered metabolic rates remained constant during 14 days of measurements, and the effect was reversible after this time. Substitution of helium or argon for the nitrogen in air had no effect on oxygen production, carbon dioxide consumption, or growth rate for 3-week periods. All measurements were made at a total pressure of 760 mm of Hg, and all gas mixtures were enriched with 2% carbon dioxide. Thus, the physiological functioning and reliability of a photosynthetic gas exchanger should not be adversely affected by: (i) oxygen partial pressures ranging from 8 to 258 mm of Hg; (ii) the use of pure oxygen at reduced total pressure (155 to 258 mm of Hg) unless pressure per se affects photosynthesis, or (iii) the inclusion of helium or argon in the gas environment (up to a partial pressure of 595 mm of Hg).     

Batch and continuous cultivation of Pseudomonas fluorescens at increased pressure

The effect of increased total pressure and partial pressures of oxygen and carbon dioxide on the growth of Pseudomonas fluorescens was investigated in an airlift reactor. In batch cultivations bacterial growth was completely inhibited with air at 8 bars total pressure. The same effect was observed with aeration by pure oxygen at 1.15 bars. Carbon dioxide partial pressure did not show inhibitory effects. Continuous experiments confirm the assumption that growth inhibition at higher total pressure is caused by the increase in oxygen partial pressure. Incubation of P. fluorescens at higher oxygen partial pressure led to an increase of bacterial productivity during subsequent continuous cultivation at ambient pressure (1 bar) with air. Maximum productivity was increased by about 75% after aeration with pure oxygen. This effect is probably the result of metabolic adaption of the bacterial cells to high oxygen partial pressure.     

Carbon dioxide inhibition of yeast growth in biomass production

Saccharomyces cerevisiae was grown under aerobic and substrate-limiting conditions for efficient biomass production. Under these conditions, where the sugar substrate was fed incrementally, the growth pattern of the yeast cells was found to be uniform, as indicated by a constant respiratory quotient during the entire growing period. The effect of carbon dioxide was investigated by replacing portions of the nitrogen in the air stream with carbon dioxide, while maintaining the oxygen content at the normal 20% level, so that identical oxygen transfer rate and atmospheric pressure were maintained for all experiments with different partial pressures of carbon dioxide. Inhibition of yeast growth was negligible below 20% CO₂ in the aeration mixture. Slight inhibition was noted at the 40% CO₂ level and significant inhibition was noted above the 50% CO₂, level, corresponding to 1.6 × 10^?2M of dissolved CO₂ in the fermentor broth. High carbon dioxide content in the gas phase also inhibited the fermentation activity of baker's yeast.     

Fed-batch cultivation of Saccharomyces cerevisiae in a hyperbaric bioreactor

Fed-batch is the dominating mode of operation in high-cell-density cultures of Saccharomyces cerevisae in processes such as the production of baker's yeast and recombinant proteins, where the high oxygen demand of these cultures makes its supply an important and difficult task. The aim of this work was to study the use of hyperbaric air for oxygen mass transfer improvement on S. cerevisiae fed-batch cultivation. The effects of increased air pressure up to 1.5 MPa on cell behavior were investigated. The effects of oxygen and carbon dioxide were dissociated from the effects of total pressure by the use of pure oxygen and gas mixtures enriched with CO(2). Fed-batch experiments were performed in a stirred tank reactor with a 600 mL stainless steel vessel. An exponential feeding profile at dilution rates up to 0.1 h(-)(1) was used in order to ensure a subcritical flux of substrate and, consequently, to prevent ethanol formation due to glucose excess. The ethanol production observed at atmospheric pressure was reduced by the bioreactor pressurization up to 1.0 MPa. The maximum biomass yield, 0.5 g g(-)(1) (cell mass produced per mass of glucose consumed) was attained whenever pressure was increased gradually through time. This demonstrates the adaptive behavior of the cells to the hyperbaric conditions. This work proved that hyperbaric air up to 1.0 MPa (0.2 MPa of oxygen partial pressure) could be applied to S. cerevisiae cultivation under low glucose flux. Above that critical oxygen partial pressure value, i.e., for oxygen pressures of 0.32 and 0.5 MPa, a drastic cell growth inhibition and viability loss were observed. The increase of carbon dioxide partial pressure in the gas mixture up to 48 kPa slightly decreased the overall cell mass yield but had negligible effects on cell viability.     

Effects of carbon dioxide on ethylene production and action in intact sunflower plants

A continuous flow system was used to study the interactions between carbon dioxide and ethylene in intact sunflower (Helianthus annuus L.) plants. An increase in the concentration of carbon dioxide above the ambient level (0.033%) in the atmosphere surrounding the plants increased the rate of ethylene production, and a decrease in carbon dioxide concentration resulted in a decrease in the rate of ethylene production. The change in the rate of ethylene production was evident within the first 15 minutes of the carbon dioxide treatment. Continuous treatment with carbon dioxide was required to maintain increased rate of ethylene production. The rate of carbon dioxide fixation increased in response to high carbon dioxide treatment up to 1.0%. Further increases in carbon dioxide concentration had no additional effect on carbon dioxide fixation. Carbon dioxide concentrations higher than 0.11% induced hyponasty of the leaves whereas treatment with 1 microliter per liter ethylene induced epinasty of the leaves.     

Oxygen enriched air supply in Escherichia coli processes: production of biomass and recombinant human growth hormone

In order to investigate the impact of high oxygen and carbon dioxide concentrations, Escherichia coli was grown in batch cultivations where the air supply was enriched with either oxygen or carbon dioxide. The effect of elevated concentrations of oxygen and carbon dioxide on stochiometric and kinetic constants was studied this way. The maximum growth rate was significantly reduced, the production of acetic acid and the biomass yield coefficient on glucose increased in cultures with carbon dioxide enriched air, compared to reference cultivations and cultivations with oxygen enriched air. The application of oxygen enriched air was studied in high cell density cultivations of Escherichia coli. Two production processes were chosen to investigate the impact of oxygen enrichment. Biomass concentration, specific growth rate, yield coefficient, respiration, mixed acid fermentation products and the product yield and quality for the recombinant product were investigated. First, a process for the production of biomass was investigated. Exponential growth could proceed for a longer time and higher growth rates could be maintained with oxygen enriched air supply. However, a higher specific oxygen consumption rate per glucose was measured after the start of the oxygen enrichment, indicating higher maintenance and consequently the growth rate and yield coefficient decreased drastically in the end of the process. Second, a process for the production of recombinant human growth hormone (rhGH) was investigated. Although the glucose feed rate and all medium components were doubled, the amount of produced biomass could only be increased by 77% when oxygen enriched air (40% oxygen) supply was applied. This was due to a decreased yield coefficient of biomass per glucose. The total amount of produced product was decreased by almost 50% compared to the control, although less proteolytically degraded variants were produced.     

Nitrogen fixation and respiration by root nodules ofAlnus rubra Bong.: Effects of temperature and oxygen concentration

Summary Using a root nodule cuvette and a continuous flow gas exchange system, we simultaneously measured the rates of carbon dioxide evolution, oxygen uptake and acetylene reduction by nodules ofAlnus rubra. This system allowed us to measure the respiration rates of single nodules and to determine the effects of oxygen concentration and temperature on the energy cost of nitrogen fixation. Energy cost was virtually unchanged (2.8–3.5 moles of carbon dioxide or oxygen per mole of ethylene) from 16 to 26°C (pO₂=20 kPa) while respiration and nitrogenase activity were highly temperature dependent. At temperatures below 16°C, nitrogenase activity decreased more than did respiration and as a result, energy cost rose sharply. Acetylene reduction ceased below 8°C. Inhibition of nitrogenase activity at low temperatures was rapidly reversed upon return to higher temperatures. At high temperatures (above 30°C) nitrogenase activity declined irreversibly, while respiration and energy cost increased.Energy cost was nearly unchanged at oxygen partial pressures of 5 to 20 kPa (temperature of 20°C). Respiration and nitrogenase activity were strongly correlated with oxygen tension. Below 5 kPa, acetylene reduction and oxygen uptake decreased sharply while production of carbon dioxide increased, indicating fermentation. Fermentation alone was unable to support nitrogenase activity. Acetylene reduction was independent of oxygen concentration from 15 to 30 kPa. Nitrogenase activity decreased and energy cost rose above 30 kPa until nearly complete inactivation of nitrogenase at 70–80 kPa. Activity declined gradually, such that acetylene reduction at a constant oxygen concentration was stable, but showed further inactivation when oxygen concentration was once again increased. Alder nodules appear to consist of a large number of compartments that differ in the degree to which nitrogenase is protected from excess oxygen.Supported by United States Department of Agriculture Grant 78-59-2252-0-1-005-1     

Effect of hydrogen and carbon dioxide partial pressures on growth and sulfide production of the extremely thermophilic archaebacterium Pyrodictium brockii

The effect of hydrogen and carbon dioxide partial pressure on the growth of the extremely thermophilic archaebacterium Pyrodictium brockii at 98 degrees C was investigated. Previous work with this bacterium has been done using an 80:20 hydrogen-carbon dioxide gas phase with a total pressure of 4 atm; no attempt has been made to determine if this mixture is optimal. It was found in this study that reduced hydrogen partial pressures affected cell yield, growth rate, and sulfide production. The effect of hydrogen partial pressure on cell yield and growth rate was less dramatic when compared to the effect on sulfide production, which was not found to be growth-associated. Carbon dioxide was also found to affect growth but only at very low partial pressures. The relationship between growth rate and substrate concentration could be correlated with a Monod-type expression for either carbon dioxide or hydrogen as the limiting substrate. The results from this study indicate that a balance must be struck between cell yields and sulfide production in choosing an optimal hydrogen partial pressure for the growth of P. brockii.     

Oxygen and carbon dioxide fluxes from barley shoots depend on nitrate assimilation

A custom oxygen analyzer in conjunction with an infrared carbon dioxide analyzer and humidity sensors permitted simultaneous measurements of oxygen, carbon dioxide, and water vapor fluxes from the shoots of intact barley plants (Hordeum vulgare L. cv Steptoe). The oxygen analyzer is based on a calciazirconium sensor and can resolve concentration differences to within 2 microliters per liter against the normal background of 210,000 microliters per liter. In wild-type plants receiving ammonium as their sole nitrogen source or in nitrate reductase-deficient mutants, photosynthetic and respiratory fluxes of oxygen equaled those of carbon dioxide. By contrast, wild-type plants exposed to nitrate had unequal oxygen and carbon dioxide fluxes: oxygen evolution at high light exceeded carbon dioxide consumption by 26% and carbon dioxide evolution in the dark exceeded oxygen consumption by 25%. These results indicate that a substantial portion of photosynthetic electron transport or respiration generates reductant for nitrate assimilation rather than for carbon fixation or mitochondrial electron transport.     

Effect of elevated dissolved carbon dioxide concentrations on growth of Corynebacterium glutamicum on D-glucose and L-lactate

The effect of increased dissolved carbon dioxide concentrations on growth of Corynebacterium glutamicum was studied with continuous turbidostatic cultures. The carbon sources were either l-lactate or d-glucose. To increase the dissolved carbon dioxide concentration the carbon dioxide partial pressure of the inlet gas stream pCO2,IN was increased stepwise from 0.0003 bar (air) up to 0.79 bar, while the oxygen partial pressure of the inlet gas stream was kept constant at 0.21 bar. For each resulting carbon dioxide partial pressure pCO2 the maximum specific growth rate mu(max) was determined from the feed rate resulting from the turbidostatic control. On d-glucose and pCO2 up to 0.26 bar, mu(max) was mostly constant around 0.58 h(-1). Higher pCO2 led to a slight decrease of mu(max). On l-lactate mu(max) increased gradually with increasing carbon dioxide partial pressures from 0.37 h(-1) under aeration with air to a maximum value of 0.47 h(-1) at a pCO2 of 0.26 bar. At very high pCO2 (0.81 bar) mu(max) decreased down to 0.35 h(-1) independent of the carbon source.     

Effect of modified atmosphere composition on the metabolism of glucose by Brochothrix thermosphacta

The influence of atmosphere composition on the metabolism of Brochothrix thermosphacta was studied by analyzing the consumption of glucose and the production of ethanol, acetic and lactic acids, acetaldehyde, and diacetyl-acetoin under atmospheres containing different combinations of carbon dioxide and oxygen. When glucose was metabolized under oxygen-free atmospheres, lactic acid was one of the main end products, while under atmospheres rich in oxygen mainly acetoin-diacetyl was produced. The proportions of the total consumed glucose used for the production of acetoin (aerobic metabolism) and lactic acid (anaerobic metabolism) were used to decide whether aerobic or anaerobic metabolism predominated at a given atmosphere composition. The boundary conditions between dominantly anaerobic and aerobic metabolisms were determined by logistic regression. The metabolism of glucose by B. thermosphacta was influenced not only by the oxygen content of the atmosphere but also by the carbon dioxide content. At high CO(2) percentages, glucose metabolism remained anaerobic under greater oxygen contents.     

The Effect of Inhibitors on the Path of Carbon in Photosynthesis by Chlorella at Low Partial Pressures of CO2: I. Methylamine

Methylamine has previously been shown to uncouple phosphorylationin isolated chloroplasts. Results presented here are consistentwith its having a similar action in Chlorella. It both stimulatesoxygen production and inhibits carbon dioxide fixation. It markedlyaffects the distribution of carbon within the photosyntheticcarbon reduction cycle at low partial pressures of carbon dioxide,decreasing the amount of sugar diphosphates and increasing thatof PGA. The production of glycollate and glycine is diminishedin the presence of methylamine but there is little effect onsucrose.     

The effect of dissolved carbon dioxide on penicillin production: Mycelial morphology

The effect of carbon dioxide on the morphology of Penicillium chrysogenum was examined. The scanning electron microscopic (SEM) study indicated that the morphology of P. chrysogenum was subject to change when exposed to various dissolved CO₂ concentrations in the medium. At low influent carbon dioxide partial pressures between 0% and 8%, the predominant morphological form of P. chrysogenum was filamentous. At higher influent carbon dioxide partial pressures of 15% and 20%, the appearance of swollen and stunted hyphae predominated, and a significant quantity of spherical or yeast-like cells were observed. It was evident that for production subject to high dissolved CO₂ concentrations the inhibition of cell growth and penicillin production related strongly to the concomitant morphological changes of P. chrysogenum.     

Gas Exchange of Algae: III. Relation between the Concentration of Carbon Dioxide in the Nutrient Medium and the Oxygen Production of Chlorella pyrenoidosa

The oxygen production of a photosynthetic gas exchanger containing Chlorella pyrenoidosa (1% packed cell volume) was measured when various concentrations of carbon dioxide were present within the culture unit. The internal carbon dioxide concentrations were obtained by manipulating the entrance gas concentration and the flow rate. Carbon dioxide percentages were monitored by means of electrodes placed directly in the nutrient medium. The concentration of carbon dioxide in the nutrient medium which produced maximal photosynthesis was in the range of 1.5 to 2.5% by volume. Results were unaffected by either the level of carbon dioxide in the entrance gas or the rate of gas flow. Entrance gases containing 2% carbon dioxide flowing at 320 ml/min, 3% carbon dioxide at 135 ml/min, and 4% carbon dioxide at 55 ml/min yielded optimal carbon dioxide concentrations in the particular unit studied. By using carbon dioxide electrodes implanted directly in the gas exchanger to optimize the carbon dioxide concentration throughout the culture medium, it should be possible to design more efficient large-scale units.