The enzymatic determination of bicarbonate and CO2 in reagents and buffer solutions

A method for the determination of bicarbonate in buffer solutions between pH 7.5 and 8.75 and in stock solutions of NaHCO3 is described. The HCO-3 is reacted with phosphoenolpyruvate (PEP) in the presence of PEP carboxylase (EC 4.1.1.31) and the oxaloacetate formed reduced to malate by NADH in the reaction catalyzed by malate dehydrogenase (EC 1.1.1.37). The extent of oxidation of NADH is measured spectrophotometrically. Experiments using standard solutions show that 1 mol of NADH is oxidized per mol of HCO-3 added. The method was used to establish the precautions needed to prepare buffer solutions containing less than 1% of the bicarbonate which would be present in the same buffers in equilibrium with air.     

分子氮影响下的蓝藻Anabaena7120乙炔还原

经分子氮预处理的蓝藻乙炔还原活性下降。分子氮对乙炔还原的抑制作用可因CO_2的加入而削弱,氮浓度增高时,CO_2的此种有益作用即消失。在5％ CO_2条件下:(1)光照强度大时,经分子氮预处理的蓝藻乙炔还原活性比未经处理的(氮气中)高些,弱光下则削弱,暗中削弱更大;(2)它受到光合抑制剂的抑制较大;(3)对CO_2和O_2的敏感度比未经氮预处理者小;(4)分子氢对其支持与来经氮预处理的相差甚微,但对其受O_2损伤时的保护作用则大些;(5)MSX对分子氮预处理的蓝藻乙炔还原也有明显的抑制,且比单加CO_2的大。     

Platelet activating factor (PAF) production by mouse embryos in vitro and its effect on embryonic metabolism

Factors affecting the production of platelet activating factor (PAF) by mouse embryos during culture in vitro were investigated. Detectable levels of embryo-derived PAF were produced within 1-4 hr with maximum PAF activity being observed after 6 hr of culture in vitro. The amount of PAF detected in media after 24 hr of culture of two-cell embryos was equivalent to 12.8 ng PAF/embryo. However, differences in activity were apparent with increased time in culture. Reduced synthesis of PAF during culture in vitro was supported by the observation that morulae stage embryos collected fresh from the reproductive tract displayed more PAF activity than morulae resulting from the 48 hr culture of two-cell embryos. In addition to determining production characteristics of PAF by embryos, we also show that the production of CO2 from carbon-1 position of lactate is positively correlated with the ability of embryos to develop during subsequent culture in vitro and therefore could be used as a measure of embryo viability. Furthermore, culture of embryos in media supplemented with PAF resulted in an increase in lactate utilization demonstrating a direct effect of PAF on the embryo. As PAF is produced by preimplantation embryos, an autocoid role of PAF in regulating embryo development is implicated. Therefore, the reduced production of PAF by embryos in vitro may explain the decreased viability of embryos commonly observed following their culture in vitro.     

Root respiration during grain filling in sunflower: The effects of water stress

Instantaneous rates of (soil + root) respiration were measured periodically during grain filling in sunflower crops that were i) irrigated at weekly intervals and ii) subjected to water stress for the last 25 days of the 40-day grain filling period. Daily (soil + root) respiration was calculated using instantaneous respiration rates, an empirically determined temperature response function, and diurnal records of soil temperature. Daily soil respiration was estimated using empirically determined functions linking soil respiration to soil temperature and water content. Between anthesis and maturity, daily root respiration of the irrigated crop dropped by about one half from ca. 1.8 g C m^-2 d^-1, exhibiting a strong association with daily crop gross photosynthesis. Water stress brought about a rapid decrease in root respiration, which fell to about 0.1 g C m^-2 d^-1 at maturity. Root respiration during grain filling was 46 and 30 g C m^-2 for irrigated and stressed crops, respectively.     

Monoterpene glycoside biosynthesis in detached grape berries grown in vitro

A procedure for the culture in vitro of isolated small berries of Vitis vinifera L. cv. Muscat of Alexandria in a Murashige and Skoog basal medium supplemented with N⁶-benzyladenine and indoleacetic acid is described. Berries developed well in culture during 60 days and tripled in size, but remained green and smaller than normal berries grown in vivo. Some callus formed on the distal end of the berry, and where major skin damage occurred, callus emerged from the cracked berries. In order to examine their biosynthetic competency, berries which were previously cultured in vitro for 60 days were incubated for 48 h in a Murashige and Skoog medium containing a [¹⁴C]-labelled water-soluble fraction. This fraction was isolated from grape berries located adjacent to a leaf that had been exposed to gaseous ¹⁴CO₂ in full sunlight for 5 h. The berries were then recultured for 48 h after which a glycosidic fraction was isolated on a C18 reversed phase column and further separated by thin layer chromatography (TLC). The major labelled band corresponded to the geranyl-β-rutinoside marker, indicating that grape berries have the ability to synthesize monoterpene glycosides. This band also consisted of other monoterpene glycosides as revealed by the gas chromatography-mass spectrometry (GC-MS) analysis of their aglycones (released by enzymatic hydrolysis).     

Seasonal nitrogen and carbon concentrations in white,brown and woody fine roots of sugar maple (Acer saccharum Marsh)

Small diameter (<1.0-mm) Acer saccharum Marsh roots were separated into white, brown and woody development state classes and analyzed for total N and C concentrations in April, July and October of 1988. White roots had greater concentrations of N and C than either brown or woody roots at each sampling date, and the N concentration of brown roots was consistently greater than that of woody roots. There were no temporal changes in N concentrations in any of the roots. C was slightly elevated in mid-summer in all three classes of roots. The data suggest the possible existence of an N translocation mechanism in ageing and developing fine roots. More research should be undertaken to establish the mechanisms of N loss in developing fine roots.     

Frankia populations in soils under different tree species—with special emphasis on soils under Betula pendula

The major source of substrates for microbial activity in the ectorhizosphere and on the rhizoplane are rhizodeposition products. They are composed of exudates, lysates, mucilage, secretions and dead cell material, as well as gases including respiratory CO₂. Depending on plant species, age and environmental conditions, these can account for up to 40% (or more) of the dry matter produced by plants. The microbial populations colonizing the endorhizosphere, including mycorrhizae, pathogens and symbiotic N₂-fixers have greater access to the total pool of carbon including that recently derived from photosynthesis. Utilization of rhizodeposition products induces at least a transient increase in soil biomass but a sustained increase depends on the state of the native soil biomass, the flow of other metabolites from the soil to the rhizosphere and the water relations of the soil. In addition, the phenomena of oligotrophy, cryptic growth, plasmolysis, dormancy and arrested metabolism can all influence the longevity of rhizosphere organisms. With this background, microbial growth in the rhizosphere will be discussed.     

Dissimilation curves as a simple method for the characterization of growth of plant cell suspension cultures

A simple non-invasive method for the characterization of growth of a plant cell suspension in a single culture flask is given. The dissimilation of sugars by a cell-culture causes a loss of weight of the contents of the culture flask, and can therefore be used to follow the growth in that single culture flask. Because a correction for water evaporation is necessary, accurate results can only be obtained when a stable closure is used (e.g. Silicosen T-type plugs). The dissimilation curves obtained in this way were correlated to the concentration of sugars in the medium, the dry weight and the fresh weight. From these correlations the amount of intracellularly stored carbohydrates could be estimated. Rate constants for CO₂-diffusion were determined for different types of closure. These values allowed the estimation of CO₂ levels inside the culture flasks from the dissimilation curves (CO₂ release curves). The dissimilation curves obtained using this method can easily be related to other types of growth curves. Different growth-phases can be clearly distinguished, e.g. lag-phase, exponential growth-phase and stationary-phase.     

INORGANIC CARBON TRANSPORT IN RELATION TO CULTURE AGE AND INORGANIC CARBON CONCENTRATION IN A HIGH-CALCIFYING STRAIN OF EMILIANIA HUXLEYI (PRYMNESIOPHYCEAE)1

The relationships among inorganic carbon transport, bicarbonate availability, intracellular pH, and culture age were investigated in high-calcifying cultures of Emiliania huxleyi (Lohmann) Hay & Mohler. Measurement of inorganic carbon transport by the silicone-oil centrifugation technique demonstrated that gadolinium, a potential Ca²⁺ channel inhibitor, blocked intracellular inorganic carbon uptake and photosynthetic ¹⁴CO₂₊ fixation in exponential-phase cells. In stationary-phase cells, the intracellular inorganic carbon concentration was unaffected by gadolinium. Gadolinium was also used to investigate the link between bicarbonate and Ca²⁺ transport in high-calcifying cells of E. huxleyi. Bicarbonate availability had significant and rapid effects on pH_i in exponential- but not in stationary-phase cells. 4′, 4′-Diisothiocyanostilbene-2,2′-disulfonic acid did not block bicarbonate uptake from the external medium by exponential-phase cells. Inorganic carbon utilization by exponential- and stationary-phase cells of Emiliania huxleyi was investigated using a pH drift technique in a closed system. Light-dependent alkalization of the medium by stationary-phase cells resulted in a final pH of 10.1 and was inhibited by dextran-bound sulphonamide, an inhibitor of external carbonic anhydrase. Exponential-phase cells did not generate a pH drift. Overall, the results suggest that for high-calcifying cultures of E. huxleyi the predominant pathway of inorganic carbon utilization differs in exponential and stationary phase cells of the same culture.