Pollen germination and tube growth: dependent on carbon dioxide and independent of ethylene

The influence of ethylene and CO₂ on pollen germination and tube growth was investigated employing ventilated culture systems. Ethylene had no effect on pollen germinability or tube growth. Germinating pollen did not produce a detectable amount of ethylene (less than 0.1 nl/g·hr). Supplementing the cultures with CO₂ caused a marked increase in germination and tube growth. The half-maximal response for germination was less than 0.5%. CO₂ levels ranging from 1.08 to 2.22% were found in the internal cavity of lily styles. CO₂ derived from stylar metabolism may, therefore, modulate pollen tube growth thus integrating the events leading to fertilization.     

Effects of ethylene and carbon dioxide on the germination of osmotically inhibited lettuce seed

Lettuce seeds (Lactuca sativa L.) used in this study germinated 98% at 25 C in light or dark. Their germination was completely inhibited by 0.20 m NaCl, 0.35 m mannitol, or polyethylene glycol 6000 (−7 bars) under continuous light when germination tests were made in Petri dishes. Approximately 50% germination occurred in sealed flasks due to endogenously produced C₂H₄ and CO₂. Removal of either or both gases prevented germination. In the presence of endogenous CO₂, addition of C₂H₄ (0.5 to 16 microliters/liter) stimulated 95 to 100% germination (after 5 days) only in the light, but the rate of germination was dependent on C₂H₄ concentration. At 16 microliters/liter C₂H₄, full germination occurred within 72 hours. Addition of up to 3.2% CO₂ had no adverse effect on the C₂H₄ action. Higher concentrations or the complete absence of CO₂ reduced both rate and total germination. CO₂ alone was ineffective.     

Role of carbon dioxide in germination of spores of Streptomyces viridochromogenes

CO₂ in required continuously during germination of Streptomyces viridochromogenes spores. Spores incubated in a defined germination medium in the absence of CO₂ remain phase bright and do not release spore carbon. In the presence of CO₂, the spores initiate germination accompanied by loss of refractility and spore carbon. The CO₂ requirement is replaced by oxaloacetate or a mixture of tricarboxylic acid cycle (TCA) intermediates. Labeled CO₂ is taken up by germinating spores, and is incorporated into protein and RNA. TCA cycle intermediates and related amino acids contain most of the acid-soluble label following short term exposures of germinating spores to ¹⁴CO₂. TCA cycle inhibitors repress germination and ¹⁴CO₂ uptake whereas folic acid antagonists do not. The results indicate that CO₂ is incorporated into oxaloacetate which is converted to biosynthetic intermediates required for germination. Operation of the TCA cycle appears to be essential for spore germination. The conclusion is reached that CO₂ is required during germination in order to maintain the cycle by an anaplerotic reaction.Abbreviations SN sucrose-nitrate medium - TX buffer Trisbuffer pH 7.3 containing-Triton X-100 - DGM defined germination medium - TX salts TX buffer plus Mg and Ca ions - TA trichloroacctic acid - TCA tricarboxylic acid     

Effect of oxygen and carbon dioxide on germination and growth ofRhizopus oligosporus on model media and soya beans

 The microcolony technique enables the effects of several atmospheric conditions on fungal growth to be studied by measuring the radius of the colony, while excluding effects of those conditions on germination of the sporangiospores. Various concentrations of oxygen and carbon dioxide in the gas environment were found to influence growth of Rhizopus oligosporus on malt extract/soya peptone/agar. The maximum radial growth rate was 1.48 mm/h and the maximum specific growth rate was 0.109 h^-1 at 30 °C. Oxygen became limiting below 1% (v/v), but growth remained possible at levels of 0.001% oxygen. Carbon dioxide stimulated growth at limiting oxygen levels. The specific growth rate increased from 0.043 h^-1 at 0.5% (v/v) oxygen and 0% (v/v) carbon dioxide to 0.096 h^-1 at 0.5% (v/v) oxygen and 5% (v/v) carbon dioxide. A mixture of 0.5% (v/v) oxygen and 35% (v/v) carbon dioxide inhibited growth. Delay of sporangiospore germination due to low (less than 0.001%) amounts of oxygen was not observed with the techniques used. Fungal activity in a rotating drum fermentor was more strongly affected by low levels of oxygen than was biomass formation on model media. High concentrations of carbon dioxide inhibited growth in the rotating drum fermentor at non-limiting levels of oxygen. It is concluded that aeration and heat removal are both essential aspects of optimization of large-scale solid-substrate bioreactors with Rh. oligosporus. Received: 5 August 1994/Received revision: 14 November 1994/Accepted: 5 December 1994     

Effect of potassium deficiency on carbon dioxide,cation, and phosphate content of muscle,with a note on the carbon dioxide content of human muscle

Albino rats weighing 160 to 175 gm. were fed a complete synthetic diet containing 0.003 per cent potassium and 0.7 per cent sodium for 40 days. Controls were given the same diet plus adequate added potassium. 1. Data from analyses of serum and skeletal muscle showed (a) a fall in serum chloride concentration and an increase in serum carbon dioxide concentration and pH in the potassium-deficient rats; (b) increases of sodium, magnesium, and calcium and a decrease of potassium in the muscle of the potassium-deficient rats; (c) no change of muscle chloride or carbon dioxide concentrations in the potassium-deficient rats. (2) Application of the Wallace-Hastings calculations to these data revealed (a) intracellular pH of the skeletal muscle of the normal rat to be 6.98 +/- 0.08; (b) an increase in serum partial pressure of carbon dioxide (pCO(2)) in potassium deficiency, together with increases in concentrations of [H(2)CO(2)] and [HCO(3) (-)] per kg. extracellular water and [H(2)CO(3)] per kg. cell water; (c) a decrease in values for [CO(2)] and [HCO(3) (-)] per kg. intracellular water; (d) a fall of intracellular pH in potassium deficiency to 6.42 +/- 0.05. (3) Analyses of sacrospinalis muscle from five men undergoing operation for ruptured intervertebral disc showed a mean value of 9.46 +/- 1.31 mM carbon dioxide per kg. blood-free tissue. Some problems of interpretation of data are briefly discussed.     

Protein purification with vapor-phase carbon dioxide

Gaseous CO2 was used as an antisolvent to induce the fractional precipitation of alkaline phosphatase, insulin, lysozyme, ribonuclease, trypsin, and their mixtures from dimethylsulfoxide (DMSO). Compressed CO2 was added continuously and isothermally to stationary DMSO solutions (gaseous antisolvent, GAS). Dissolution of CO2 was accompanied by a pronounced, pressure-dependent volumetric expansion of DMSO and a consequent reduction in solvent strength of DMSO towards dissolved proteins. View cell experiments were conducted to determine the pressures at which various proteins precipitate from DMSO. The solubility of each protein in CO2-expanded DMSO was different, illustrating the potential to separate and purify proteins using gaseous antisolvents. Polyacrylamide gel electrophoresis in sodium dodecyl sulfate (SDS-PAGE) was used to quantify the separation of lysozyme from ribonuclease, alkaline phosphatase from insulin, and trypsin from catalase. Lysozyme biological activity assays were also performed to determine the composition of precipitates from DMSO initially containing lysozyme and ribonuclease. SDS-PAGE characterizations suggest that the composition and purity of solid-phase precipitated from a solution containing multiple proteins may be accurately controlled through the antisolvent's pressure. Insulin, lysozyme, ribonuclease, and trypsin precipitates recovered substantial amounts of biological activity upon redissolution in aqueous media. Alkaline phosphatase, however, was irreversibly denaturated. Vapor-phase antisolvents, which are easily separated and recovered from proteins and liquid solvents upon depressurization, appear to be a reliable and effective means of selectively precipitating proteins.     

Inhibition of germination and sporulation (conidiation) in Neurospora sitophila Shear & Dodge by carbon dioxide

A complete inhibition of germination and conidiation was observed inNeurospora sitophila when maintained under a pure carbon dioxide atmosphere. Experiments were performed to explain the relatively poor conidiation of this species of mold under standard laboratory experimental conditions.     

Interactive effects of carbon dioxide, temperature, and ultraviolet-B radiation on soybean (Glycine max L.) flower and pollen morphology, pollen production, germination, and tube lengths

Plant reproduction is highly vulnerable to global climate change components such as carbon dioxide concentration ([CO(2)]), temperature (T), and ultraviolet-B (UV-B) radiation. The objectives of this study were to determine the effects of season-long exposure to treatments of [CO(2)] at 360 (control) and 720 micromol mol(-1) (+CO(2)), temperature at 30/22 degrees C (control) and 38/30 degrees C (+T) and UV-B radiation 0 (control) and 10 kJ m(-2) d(-1) (+UV-B) on flower and pollen morphology, pollen production, germination, and tube lengths of six soybean genotypes (D 88-5320, D 90-9216, Stalwart III, PI 471938, DG 5630RR, and DP 4933RR) in sunlit, controlled environment chambers. The control treatment had 360 micromol mol(-1) [CO(2)] at 30/22 degrees C and 0 kJ UV-B. Plants grown either at +UV-B or +T, alone or in combination, produced smaller flowers with shorter standard petal and staminal column lengths. Flowers so produced had less pollen with poor pollen germination and shorter tube lengths. Pollen produced by the flowers of these plants appeared shrivelled without apertures and with disturbed exine ornamentation even at +CO(2) conditions. The damaging effects of +T and +UV-B were not ameliorated by +CO(2) conditions. Based on the total stress response index (TSRI), pooled individual component responses over all the treatments, the genotypes were classified as tolerant (DG 5630RR, D 88-5320: TSRI >-790), intermediate (D 90-9216, PI 471938: TSRI <-790 to >-1026), and sensitive (Stalwart III, DP 4933RR: TSRI <-1026). The differences in sensitivity identified among genotypes imply the options for selecting genotypes with tolerance to environmental stresses projected to occur in the future climates.     

End-tidal carbon dioxide tension reflects arterial carbon dioxide tension in the heat-stressed human with and without simulated hemorrhage

End-tidal carbon dioxide tension (Pet(CO(2))) is reduced during an orthostatic challenge, during heat stress, and during a combination of these two conditions. The importance of these changes is dependent on Pet(CO(2)) being an accurate surrogate for arterial carbon dioxide tension (Pa(CO(2))), the latter being the physiologically relevant variable. This study tested the hypothesis that Pet(CO(2)) provides an accurate assessment of Pa(CO(2)) during the aforementioned conditions. Comparisons between these measures were made: 1) after two levels of heat stress (N = 11); 2) during combined heat stress and simulated hemorrhage [via lower-body negative pressure (LBNP), N = 8]; and 3) during an end-tidal clamping protocol to attenuate heat stress-induced reductions in Pet(CO(2)) (N = 7). Pet(CO(2)) and Pa(CO(2)) decreased during heat stress (P < 0.001); however, there was no group difference between Pa(CO(2)) and Pet(CO(2)) (P = 0.36) nor was there a significant interaction between thermal condition and measurement technique (P = 0.06). To verify that this nonsignificant trend for the interaction was not due to a type II error, Pet(CO(2)) and Pa(CO(2)) at three distinct thermal conditions were also compared using paired t-tests, revealing no difference between Pa(CO(2)) and Pet(CO(2)) while normothermic (P = 0.14) and following a 1.0 ± 0.2°C (P = 0.21) and 1.4 ± 0.2°C (P = 0.28) increase in internal temperature. During LBNP while heat stressed, measures of Pet(CO(2)) and Pa(CO(2)) were similar (P = 0.61). Likewise, during the end-tidal carbon dioxide clamping protocol, the increases in Pet(CO(2)) (7.5 ± 2.8 mmHg) and Pa(CO(2)) (6.6 ± 3.4 mmHg) were similar (P = 0.31). These data indicate that mean Pet(CO(2)) reflects mean Pa(CO(2)) during the evaluated conditions.     

Precipitation of porcine insulin with carbon dioxide

Recent works have pointed to the use of volatile electrolytes such as carbon dioxide (CO₂) dissolved in aqueous solutions as a promising alternative to the precipitating agents conventionally used for protein recovery in the food and pharmaceutical industries. In this work we investigated experimental and theoretical aspects of the precipitation of porcine insulin, a biomolecule of pharmaceutical interest, using CO₂ as an acid‐precipitating agent. The solubility of porcine insulin in NaHCO₃ solutions in pressurized CO₂ was determined as a function of temperature and pressure, with a minimum being observed close to the protein isoelectric point. A thermodynamic model was developed and successfully utilized to correlate the experimental data. Insulin was considered a polyelectrolyte in the model and its self‐association reactions were also taken into account. The biological activity of insulin was maintained after precipitation with CO₂, although some activity can be lost if foam is formed in the depressurization step. Biotechnol. Bioeng. 2009;103: 909–919. © 2009 Wiley Periodicals, Inc.     

Euthanasia of neonatal mice with carbon dioxide

Exposure to carbon dioxide (CO2) is the most prevalent method used to euthanize rodents in biomedical research. The purpose of this study was to determine the time of CO2 exposure required to euthanize neonatal mice (0 to 10 days old). Multiple groups of mice were exposed to 100% CO2 for time periods between 5 and 60 min. Mice were placed in room air for 10 or 20 min after CO2 exposure, to allow for the chance of recovery. If mice recovered at one time point, a longer exposure was examined. Inbred and outbred mice were compared. Results of the study indicated that time to death varied with the age of the animals and could be as long as 50 min on the day of birth and differed between inbred and outbred mice. Institutions euthanizing neonatal mice with CO2 may wish to adjust their CO2 exposure time periods according to the age of the mice and their genetic background.     

Predicting the impact of increasing carbon dioxide concentration and temperature on seed germination and seedling establishment of African grasses in Brazilian Cerrado

Global climate changes and biological invasions are environmental disturbances that may interact synergistically, causing loss of biodiversity. As the early stages of development are the most sensitive and easily affected by these constraints, this study investigated the effects of increased carbon dioxide (CO₂) and temperature, as forecasted for 2100, on seed germination and early development of three species of invasive African grasses that have gradually replaced landscapes of the Brazilian Cerrado biome. It was observed that these parameters affected percentage and rate of germination in Urochloa brizantha, rate of germination and mean germination time in Urochloa decumbens and accelerated autotrophy acquisition in U. brizantha, U. decumbens and Megathyrsus maximus. Regarding root elongation, all species showed changes in total length, absolute and relative growth rate, but at different stages of development or time intervals, with increased temperature being more significant than increased CO₂, probably due to seed reserves still being the main carbon sources at this stage. Taken together, the results indicate that the effects of CO₂ and increased temperature are species specific and highlight the greatest potential of U. brizantha to germinate, and of U. decumbens for seedling establishment under these environmental changes.