Carbon dioxide of air inhibits the formation of silver nanoparticles initiated by proteins in polyacrylamide gel and in solution

It was shown that the staining of proteins in polyacrylamide gel by silver is inhibited by contact with air of the ammonia complex with silver ions used at the first stage of detection. It was proved by experiments on the reduction of silver by ethanolamine from a complex with ethanolamine and by formaldehyde from a complex with ammonia that the formation of silver nanoparticles initiated by proteins is inhibited by air carbon dioxide. The participation of carbon dioxide in this process is discussed. It was found that even the breathing of an experimenter can induce variations in carbon dioxide concentration sufficient to adversely affect the reproducibility of the silver staining techniques. It was concluded that, for stable staining of proteins by silver in polyacrylamide gel, it is necessary to maintain a low concentration of carbon dioxide in air over the detection solutions.     

Trapping radioactive carbon dioxide during cellular metabolic assays under standard culture conditions: description of a unique gas-capturing device

Measurement of carbon dioxide levels has been employed to follow cellular metabolic reactions for quite some time. By radio-labeling substrate molecules and evaluating the radioactivity levels of the carbon dioxide released, insight into metabolic pathways can be gleaned. Currently, no carbon dioxide capturing device is available that can be used with large volume cell monolayers growing under standard conditions within a regular commercially available culture flask. In this note we describe a simple device for collecting radio-labeled carbon dioxide from a standard culture flask. The device is independent of the culture flask, but can be attached for metabolic measurements allowing cells to be grown under standard conditions prior to study. The presented design permits convenient transfer of the device between flasks without contaminating or disturbing cells growing within the flasks. Data are presented demonstrating the reproducibility of measurements made with multiple devices with different substrate concentrations and varying periods of time, ranging up to 3 h.     

Carbon dioxide injection method for enhancing hydrogenotrophic denitrification of secondary wastewater effluent in fixed bed reactor

This study presents an optimal injection method for using carbon dioxide as a carbon source for the hydrogenotrophic denitrification of secondary wastewater effluent in a laboratory-scale fixed bed reactor (FBR). The FBR was operated under three conditions: a continuous CO₂ supply, periodic CO₂ supply, and without a CO₂ supply. The continuous operation of the FBR without carbon dioxide injection resulted in an increase in pH to 10 and a noticeable level of nitrite accumulation. The continuous co-injection of carbon dioxide and hydrogen gas decreased the pH to a range of 6 ～ 8, but the denitrification efficiency decreased to 29%. The co-injection of carbon dioxide decreased the maximum dissolved hydrogen concentration and hydrogen mass transfer rate by 25 and 61%, respectively. Compared to the continuous injection method, a periodic injection of carbon dioxide increased the denitrification efficiency from 28.6 to 85% as the hydrogen flow rate and hydraulic retention time (HRT) increased. With the periodic injection of carbon dioxide, the nitrite accumulation appeared to be insignificant as the hydrogen flow rate increased.     

Induction of Calcium Carbonate by Bacillus cereus

The purpose of this research was to study how the bacteria Bacillus cereus (DCB1) utilizes calcium ions in a culture medium with carbon dioxide (CO₂) to yield calcium carbonate (CaCO₃). The bacteria strain DCB1 was a dominant strain isolated from dolomitic surfaces in areas of Karst topographies. The experimental method was as follows: a modified beef extract-peptone medium (beef extract 3.0 g, peptone 10 g, NaCl 5.0 g, CaCl₂ 2.0 g, glass powder 2.0 g, distilled water 1 L, and a pH between 6.5 and 7.5) was inoculated with B. cereus to attempt to induce the synthesis of CaCO₃. The sample was then processed by centrifugation every 24 h during the 7-day cultivation period. The pH, carbonic anhydrase (CA) activity, and the concentrations of both HCO^- ₃ and Ca²⁺ in the supernatant fluid were measured. Subsequently, precipitation in the culture medium was analyzed to confirm, or otherwise, the presence and if present, the formation, of CaCO₃. Methods used included X-ray diffraction (XRD), Scanning Electron Microscopy (SEM), Transmission Electron Microscopy (TEM), and Energy Dispersive Spectroscopy (EDS). Meanwhile, the carbon source in the carbonate was classified by its isotope composition. Results showed that B. cereus can improve its pH value in this culture medium; concentrations of HCO^- ₃ and Ca²⁺ showed a significant decline over the duration of the cultivation period. CA activity reached its maximum during the second day; XRD, SEM, TEM, and isotope analysis all revealed the presence of CaCO₃ as a precipitate. Additionally, these results did not occur in an aseptic control group: no detectable level of CaCO₃ was produced therein. In conclusion: B. cereus can metabolize active materials, such as secretase, by its own growth and metabolism, and can either utilize atmospheric CO₂, or respire, to induce CaCO₃ production. Experimental evidence is offered for a concomitant CO₂ reduction and CaCO₃ induction by microorganisms.     

Carbon Dioxide as an Essential Requirement for Cultured Sycamore Cells

Carbon dioxide (optimum concentration c. 1.0%) is essential to the initiation of the growth in suspension culture or on agar plates of cultured sycamore cells. By effective flushing of the cultures with CO₂-free air it is possible to demonstrate this requirement with initial cell densities up to 50 × 10³ cells ml^?1. This growth-promoting activity of carbon dioxide is not related to any effect it may have on the pH of the culture medium. The cells fix applied carbon dioxide into organic and amino acids but attempts to replace the carbon dioxide requirement by non-toxic levels of organic or amino acids have not been successful.     

Studying the effect of elevated CO2 in the open in a naturally enriched environment in Central Italy

A gas vents area was recently localized in Central Italy. The gas emitted from the vents is composed by 92% of carbon dioxide and this produces an anomaly in the composition of the atmosphere over an area of about 2 ha. Atmospheric carbon dioxide concentration was measured by means of an infrared gas analyzer and diffusion tubes in several points and for some days within the area. Measurements revealed that the site can be at least divided into three sub-areas having increasing CO₂ concentration in the air. A preliminary analysis of natural vegetation in the area was conducted by counting stomatal and epidermal cells number and measuring guard cell size on leaves of several oak trees growing both near and far away from the vents. This analysis suggested that elevated CO₂ may have reduced the size of guard cells leaving stomatal density and stomatal index unaltered.     

Open-dish incubator for live cell imaging with an inverted microscope

Here we describe the design and fabrication of an inexpensive cell culture incubator for the stage of an inverted light microscope for use in live cell imaging. This device maintains the temperature of the cell culture at 37 degrees C with great stability and, after reaching equilibrium, provides focal stability of an image for 20-25 min with oil-immersion lenses. We describe two versions of the incubator: one for use with standard 60-mm plastic culture dishes, and the other version for imaging of cells on glass coverslips. Either can be made for less than $400. Most components are widely available commercially, and it requires only simple wiring and 3 h to assemble. Although the device is generally useful for live cell imaging on an inverted microscope, it is particularly suitable for work in which instruments are introduced into the culture, such as electrophysiology or micromanipulation. The design is based on the principle that control performance is limited by the lag time between detection and response. The key element of the design is a heated, temperature-controlled aluminum ring serving as a mini-incubator surrounding the culture vessel. For this reason, we call our design a "ringcubator."     

Comparative growth characteristics of vero cells on gas-permeable and conventional supports

We have evaluated the gas-permeable membrane, etched FEP-teflon, and compared it with other cell supports for tissue culture, in particular Falcon plastic and glass. VERO cells grown on FEP-teflon in 5% CO₂, 95% air reach 1.7 times the density/unit surface area as on other cell surfaces. Cells propagated on FEP-teflon tend to form multiple layers and maintain maximum cell density for periods unusually long for VERO. pH regulation of atmospheric pCO₂, to maintain stable medium pH, increases the maximum cell density on Falcon plastic by 20%, and allows growth on FEP-teflon at serum levels 1.0%. Cells on FEP-teflon eventually form multilayers at these low serum levels. Comparison of the cell morphology in confluent sheets on glass and the top layer of multilayers on FEP-teflon show no significant differences in overall morphology. However, the submerged cells in piled-up foci appear denuded of the small microvilli typical of this cell line. The growth characteristics of FEP-teflon appear related to the permeability of this material to oxygen and/or carbon dioxide. We discuss our findings in terms of the influence of pericellular pO₂, pCO₂ and pH on serum requirements and contact inhibition.     

The Kinetics of Extracellular Glycollate Production by Chlorella pyrenoidosa

Extracellular glycollate is liberated by Chlorella pyrenoidosaduring growth in medium bubbled with air or 3 per cent carbondioxide in air. With air the rate of release of glycollate percell decreases, with 3 per cent carbon dioxide it increases,with increase in cell number. Glycollate is released duringshort-term experiments when C. pyrenoidasa, grown under lowlight and high carbon dioxide, is transferred suddenly to highlight and low carbon dioxide. No other combination of thesefactors produces a comparable release of glycollate. The quantityof glycollate released in short-term experiments increases exponentiallywith the relative growth-rate of the culture from which thecells are derived. A crucial condition for maximum glycollaterelease is that growth of the culture prior to the experimentshould not be limited by carbon-dixoide concentration. The effectof pH is related to its effect on growth-rate; i.e. C. pyrenoidosahas a lower relative growth-rate at pH 8.3 and produces correspondinglyless glycollate than faster growing cultures at pH 6.4. Duringshort-term experiments under high light and low carbon dioxidethe rate of glycollate release drops after 50–100 minutessuggesting exhaustion of the glycollate precursor.     

An Automated System for Monitoring and Regulating the pH of Bicarbonate Buffers

The bicarbonate buffer is considered as the most biorelevant buffer system for the simulation of intestinal conditions. However, its use in dissolution testing of solid oral dosage forms is very limited. The reason for this is the thermodynamic instability of the solution containing hydrogen carbonate ions and carbonic acid. The spontaneous loss of carbon dioxide (CO₂) from the solution results in an uncontrolled increase of the pH. In order to maintain the pH on the desired level, either a CO₂ loss must be completely avoided or the escaped CO₂ has to be replaced by quantitative substitution, i.e. feeding the solution with the respective amount of gas, which re-acidifies the buffer after dissociation. The present work aimed at the development of a device enabling an automatic pH monitoring and regulation of hydrogen carbonate buffers during dissolution tests.     

Open top chambers for exposing plant canopies to elevated CO2 concentration and for measuring net gas exchange

Open top chamber design and function are reviewed. All of the chambers described maintain CO₂ concentrations measured at a central location within ±30 ppm of a desired target when averaged over the growing season, but the spatial and temporal range within any chamber may be closer to 100 ppm. Compared with unchambered companion plots, open top chambers modify the microenvironment in the following ways: temperatures are increased up to 3°C depending on the chamber design and location of the measurement; light intensity is typically diminished by as much as 20%; wind velocity is lower and constant; and relative humidity is higher. The chamber environment may significantly alter plant growth when compared with unchambered controls, but the chamber effect on growth has not been clearly attributed to a single or even a few environmental factors.A method for modifying an open top chamber for tracking gas exchange between natural vegetation and the ambient air is described. This modification consists of the addition of a top with exit chimney to reduce dilution of chamber CO₂ by external ambient air, is quickly made and permits estimation of the effects of elevated CO₂ and water vapor exchange.The relatively simple design and construction of open top chambers make them the most likely method to be used in the near future for long-term elevated CO₂ exposure of small trees, crops and grassland ecosystems. Improvements in the basic geometry to improve control of temperature, reduce the variation of CO₂ concentrations, and increase the turbulence and wind speed in the canopy boundary layer are desirable objectives. Similarly, modifications for measuring water vapor and carbon dioxide gas exchange will extend the usefulness of open top chambers to include non-destructive monitoring of the responses of ecosystems to rising atmospheric CO₂.     

Carbon dioxide assimilation from air and water by duckweed Spirodela polyrrhiza (L.) Schleid

Carbon dioxide assimilation by duckweed, S. polyrrhiza, was measured using a glass assimilation box and ¹⁴C-NaHCO₃, under different pH conditions of water. S. polyrrhiza assimilates carbon dioxide from both air and water. The carbon assimilation from air is comparable to the assimilation from water under normal pH conditions.     

Will ocean acidification affect marine microbes?

The pH of the surface ocean is changing as a result of increases in atmospheric carbon dioxide (CO₂), and there are concerns about potential impacts of lower pH and associated alterations in seawater carbonate chemistry on the biogeochemical processes in the ocean. However, it is important to place these changes within the context of pH in the present-day ocean, which is not constant; it varies systematically with season, depth and along productivity gradients. Yet this natural variability in pH has rarely been considered in assessments of the effect of ocean acidification on marine microbes. Surface pH can change as a consequence of microbial utilization and production of carbon dioxide, and to a lesser extent other microbially mediated processes such as nitrification. Useful comparisons can be made with microbes in other aquatic environments that readily accommodate very large and rapid pH change. For example, in many freshwater lakes, pH changes that are orders of magnitude greater than those projected for the twenty second century oceans can occur over periods of hours. Marine and freshwater assemblages have always experienced variable pH conditions. Therefore, an appropriate null hypothesis may be, until evidence is obtained to the contrary, that major biogeochemical processes in the oceans other than calcification will not be fundamentally different under future higher CO₂/lower pH conditions.     

Influence of carbon dioxide solubility on the accuracy of measurements of carbon dioxide production rate by gas balance technique

The total amount of carbon dioxide (CD) produced by a microbial culture is evolved via an outlet gas stream, outflowing broth, and can lead to a rise of the CD content in broth. To analyze relations among the three amounts, the equilibrium is considered between different ionic forms of carbon dioxide at different pH values as well as between the gaseous and disolved CO₂. Dependences of the equilibrium constants on temperature are found by a thermodynamic analysis. Numerical estimation of an error of the CD production rate measurement, originating from the neglect of dissolved CD, is developed. The gas analysis technique alone provides a sufficient accuracy at pH lower than 6. At higher pH the error can be estimated using equations presented below.     

RESPONSES OF THE ACIDOPHILIC ALGA EUGLENA MUTABILIS (EUGLENOPHYCEAE) TO CARBON ENRICHMENT AT pH 31

A defined medium (MAM) simulating acid mine drainage waters was developed which supported reproducible growth rates of three axenic strains of Euglena mutabilis Schmitz. Growth responses to various pHs and carbon sources were examined under defined culture conditions. A lab strain and two 5eld isolates, tested over pH range 1.5-9.0, grew best under acidic conditions (pH < 5.5) with highest growth rates at pH 3-4. Photoauxotrophic growth rates of all strains at pH 3 were improved significantly over unstirred batch controls by bubbling with air and even more by enrichment with 5% CO₂ in air. These results confirmed inorganic carbon limitation in batch culture. Organic carbon substrates were tested as possible carbon supplements in batch culture at pH 3. None of the strains survived in the dark on any of the twenty organic sources added. In the light, the lab strain exhibited some photoheterotrophic growth potential on glucose, sucrose, ethanol, and amino acids but growth was inhibited by acetate. Field strains showed little or no growth improvement with any organic substrate addition. Under simultaneous enrichment with acetate and 5% CO₂ acetate continued to be inhibitory. Simultaneous enrichment with glucose and 5% CO₂ gave higher yields of the lab strain than with CO₂ alone but did not enhance growth of the field strain. We conclude that E. mutabilis is an acidophilic photoauxotroph which appears unable to use organic carbon supplements for growth even under conditions of carbon limitation.     

The use of acetic acid as a source of carbon by cultured Chondrus crispus (Gigartinales,Rhodophyta) stackhouse

When growing seaweeds in tanks, pH and carbon source supply have to be controlled in order to maximize photosynthesis. pH can be controlled either by adding various inorganic acids which requires the extra addition of carbon, or by combining pH control and carbon source with for instance CO₂ or an organic acid such as acetic acid (CH₃COOH). We have found comparable productivity of Chondrus using CO₂ or CH₃COOH in tank culture with an increase in production of 25.0 and 27.5%, respectively, over the control. Laboratory experiments showed that acetic acid enabled us to maintain a steady state total inorganic carbon in the medium, the algae displaying an active photosynthesis. Experiments using labelled acetic acid CH₃-¹⁴COOH showed that the acid molecule or at least the -¹⁴COOH group is taken up by Chondrus although the mechanism was not elucidated. Preliminary extractions with hot ethanol showed that 67.9% of the label was solubilized from labelled tissue. Few counts were found in the carrageenans (< 1 %) and between 25.6 and 45.1% were found in the cellulosic residues. Acetic acid is suggested as an interesting means of regulating the pH and adding carbon in macrophyte culture.     

Hemoglobin synthesis in isolated erythroid colonies from the chick embryo.

Primary cultures derived from mechanically dissociated definitive streak chick blastoderms were grown in a warm air stream on the stage of inverted phase microscope, through which in vitro erythroid development could be observed. Proerythroid cells divide three or four times in 48 hr to give rise to erythroid colonies ranging from 10 to 1000 cells, depending on the size of the blastoderm fragments from which they were derived.Erythroid cell development follows a similar course in cultures grown in a carbon dioxide incubator. Colonies consisting of about 50 cells, derived from blastoderm fragments containing 5 to 10 cells, were isolated and labeled with [³H]leucine, and their labeled hemoglobins were analyzed by isoelectric focusing. Both early hemoglobins (E,M,P,P′, and P″) and late hemoglobins (A and D) are made in colonies derived from single blastoderm fragments. The ratio of late to early hemoglobins is about 1.7 in all colonies analyzed. The implications of this finding for the clonal model of erythroid development are discussed.     

Pheromone monitoring in the field using single sensillum recording

In order to obtain direct information of stimulus dynamics perceived by a male moth under field conditions a portable device was constructed which enables continuous recording of responses from individual pheromone receptors. The device is suitable for tip recording by means of micro-knives as well as for recording with tungsten electrodes making it applicable for a wide range of insects. A micro thermistor air velocity sensor is placed within 2 mm from the preparation to record the momentary air flow. The signal conditioning electronic circuits are battery powered, and the signals can be stored on a portable tape recorder. Field recordings were made from individual male antennal pheromone receptors ofAegeria myopaeformis andAdoxophyes orana. In all recordings the instantaneous firing frequency of the receptor cells was strongly modulated by the air velocity. Analysis of the data may provide information about the average pheromone concentration and the fine structure of pheromone plumes under various conditions.     

Effect of CO2 enhancement in an outdoor algal production system usingTetraselmis

One of the objectives of microalgal culture is to provide reliable production technology for important live aquaculture feed organisms. Presented here are the results of experiments designed to provide a better understanding of the relationship between inorganic carbon availability and algal production.Our results suggest that through additions of CO₂ gas we were able to maintain sufficient dissolved carbon to stabilize outdoor algal cultures. Increases in the rate of addition of CO₂ increased levels of dissolved CO₂, total dissolved inorganic carbon (CO₂), and decreased pH in the growth medium. This translated into improved buffering capacity of the culture medium and higher growth rate. A minimum of 2.4 mM CO₂ was found necessary to maintain a maximal growth rate of 0.7 doublings/day. We also found that the increased productivity more than offsets the cost of adding the CO₂.