Bacterial Degradation of EDTA

Degradation of EDTA (ethylenediaminetetraacetic acid) or metal–EDTA complexes by cell suspensions of the bacterial strain DSM 9103 was studied. The activity of EDTA degradation was the highest in the phase of active cell growth and decreased considerably in the stationary phase, after substrate depletion in the medium. Exponential-phase cells were incubated in HEPES buffer (pH 7.0) with 1 mM of uncomplexed EDTA or EDTA complexes with Mg²⁺, Ca²⁺, Mn²⁺, Pb²⁺, Co²⁺, Cd²⁺, Zn²⁺, Cu²⁺, or Fe³⁺. The metal–EDTA complexes (Me–EDTA) studied could be divided into three groups according to their degradability. EDTA complexes with stability constants K below 10¹⁶ (log K < 16), such as Mg–EDTA, Ca–EDTA, and Mn–EDTA, as well as uncomplexed EDTA, were degraded by the cell suspensions at a constant rate to completion within 5–10 h of incubation. Me–EDTA complexes with log K above 16 (Zn–EDTA, Co–EDTA, Pb–EDTA, and Cu–EDTA) were not completely degraded during a 24-h incubation, which was possibly due to the toxic effect of the metal ions released. No degradation of Cd–EDTA or Fe(III)–EDTA by cell suspensions of strain DSM 9103 was observed under the conditions studied.     

Using a chelate-buffered nutrient solution to establish the critical solution activity of Mn2+ required by barley (Hordeum vulgare L.)

Relatively little is known about the responses of plants to micronutrients when these nutrients are maintained at the very low levels found in soils of low fertility. We have determined the requirement of barley (Hordeum vulgare L. cv Herta) for ionic Mn²⁺ in plant culture solutions using the chelating agent HEDTA as a buffer for micronutrient metal ions. The chemical activity of Mn²⁺ was varied approximately 10,000-fold from log(Mn²⁺)=–10.8 to –6.8 (pMn 10.8 to pMn 6.8), while holding constant the activities of the other micronutrient cations. Growth, appearance, and composition of Herta barley indicated that log(Mn²⁺) of approximately –8.3 would permit optimal dry matter production and normal plant development. The specific accumulation rate of Mn by 15 to 23 day old seedlings was a linear function of the Mn²⁺ activity in solution. At log(Mn²⁺) of about –9.8 or below, barley seedlings were unable to accumulate significant amounts of Mn, and at some harvests, suffered a net loss of Mn to solution. Seedlings younger than 11 days old were ineffective accumulators of several cations, including Mn, Fe, Zn, Cu, Mg, and Ca. Differences in Mn availability did not influence uptake of other cations, except that Cu uptake by roots increased with increasing Mn uptake.Abbreviations MES 2-(N-morpholino)-ethanesulphonic acid - HEDTA N-(2-hydroxyethyl)ethylene-dinitrilotriacetic acid - DTPA diethylenetrinitrilopentaacetic acid     

Superoxide dismutase mimetic activity of a cyclic tetrameric Schiff base N-coordinated Cu(II) complex

A pulse radiolytic study using the cyclic tetrameric Schiff base N-coordinated copper complex Cu(TAAB)²⁺ has been performed. The reaction of the Cu(TAAB)²⁺ complex with superoxide revealed pseudo first-order characteristics with the rate constant of k ₂ = (2.9 ± 0.5) × 10⁸ mol^–1 s^–1 dm³. The complex survive presence of competing serum albumin in physiological concentrations. The complex stability constant K = 1.15 × 10¹⁸ (log K = 18.06) is two orders of magnitude higher than that of Cu(II)-serum albumin (log K = 16.2). Transient changes of the stability during the oxidation/reduction process and in the presence of 600 /mol l^–1 albumin did not affect significantly either the electronic absorption of the complex or its catalytic activity.     

Effect of Glycine and Strychnine on Cl–-Activated Mg2+-ATPase from Bream Brain Microsomes (Abramis bramaL.)

The effect of glycine and strychnine on Mg²⁺-ATPase from the microsomal fraction of the bream (Abramis bramaL.) brain was studied. The glycine in the concentration range 10^–7–10^–4M activates the enzyme. The effect of glycine on Mg²⁺-ATPase is obviated by 100 M strychnine. The strychnine in the concentration range 5–90 M activates the basal Mg²⁺-ATPase but decreases the effect of the enzyme activation by 10^–4M glycine. The effect of Cl^–on Mg²⁺-ATPase depends on the substrate concentration (Mg²⁺-ATP) and is not observed in the presence of 100 M strychnine. A receptor-dependent pathway of glycine and strychnine action on Cl^–-activated Mg²⁺-ATPase from bream brain microsomes is proposed.     

Amperometric estimation of BOD by using living immobilized yeasts

Summary A microbial electrode consisting of immobilized living whole cells of yeasts, porous membrane and an oxygen electrode was prepared for continuous estimation of biochemical oxygen demand (BOD). Immobilized Trichosporon cutaneum was employed for the microbial electrode sensor for BOD. When a sample solution containing the equivalent amount of glucose and glutamic acid was injected into the sensor system, the current of the electrode decreased markedly with time until steady state was reached. The response time was within 18 min. A linear relationship was observed between the current decrease and the concentration below 41 mg l ^– of glucose and 41 mg l ^– glutamic acid (5-day BOD 60 mg l ^–). The current decrease was reproducible within ± 6% of the relative error when a sample solution containing 27 mg l ^– of glucose and 27 mg l ^– of glutamic acid (5-day BOD 40 mg l ^–) was employed. The microbial electrode sensor was applied to untreated waste waters from a fermentation factory. Good comparative results were obtained between BOD estimated by the microbial electrode and that determined by the conventional 5-day method (regression coefficient was 1.2). Furthermore, the effect of various compounds on BOD estimation was also examined. The current output of the microbial electrode sensor was almost constant for 17 d and 400 tests.     

Regucalcin modulates hormonal effect on (Ca2+−Mg2+)-ATPase activity in rat liver plasma membranes

The interaction of various hormones and regucalcin on (Ca²⁺–Mg²⁺)-ATPase activity in rat liver plasma membranes was investigated. The presence of epinephrine (10^–6–10^–4 M), and insulin (10^–8–10^– M) in the reaction mixture produced a significant increase in (Ca²⁺–Mg²⁺)-ATPase activity, while the enzyme activity was decreased significantly by calcitonin, (3×10^–8–3×10^–6 M). These hormonal effects, except for calcitonin, were clearly inhibited by the presence of vanadate (10^–4 M) which can inhibit the Ca²⁺-dependent phosphorylation of enzyme. Meanwhile, regucalcin (0.25 and 0.50 M), isolated from rat liver cytosol, elevated significantly (Ca²⁺–Mg²⁺)-ATPase activity in the plasma membranes, although this elevation was not inhibited by vanadate (10^–4 M). the epinephrine (10^–5 M) or phenylephrine (10^–4 M)-induced increase in (Ca²⁺–Mg²⁺)-ATPase activity was disappeared in the presence of regucalcin; in this case the effect of regucalcin was also weakened. However, the inhibitory effect of calcitonin (3×10^–6 M) was not weakened by the presence of regucalcin (0.5 M). Moreover, GTP (10^–5 and 10^–4 M)-induced increase in (Ca²⁺–Mg²⁺)-ATPase activity was not seen in the presence of regucalcin (0.25 M). The present finding suggests that the activating mechanism of regucalcin on (Ca²⁺–Mg²⁺)-ATPase is not involved on GTP-binding protein which modulates the receptor-mediated hormonal effect in rat liver plasma membranes.     

Nickel hyperaccumulation in shoot cultures of Alyssum markgrafii

Shoot cultures of Alyssum markgrafii O.E. Shulz, endemic nickel hyperaccumulating species of central Balkan, were established and maintained on Murashige and Skoog medium supplemented with 0.2 mg dm^–3 benzyladenine (BA). Nickel in form of NiCl₂ . 6 H₂O was supplemented at 22 different concentrations ranging from 0.0001 to 15 mM but none of them was lethal to cultures. High Ni²⁺ concentrations (10 mM or more) arrested shoot growth which, upon transfer to Ni-free medium, commenced via axillary bud proliferation. Shoots that developed from axillary buds through the subculture manifested increased tolerance to Ni²⁺ expressed as shoot elongation. Shoot multiplication and dry biomass production decreased with increase of Ni²⁺ in medium. Only the accumulation of Ni²⁺ in tissues increased with Ni²⁺ content of the medium. Apart from shoot cultures, high Ni²⁺ accumulation was registered in undifferentiated callus cultured on medium with 0.5 mg dm^–3 BA and 0.5 mg dm^–3 naphthylacetic acid. Highest content of accumulated Ni was 2.37 g g^–1 (d.m.) in shoots and 2.65 g g^–1 (d.m.) in callus, both measured on medium with 15 mM Ni²⁺.     

Effects of Ca2+ and vitamin E on posttranslational regulation of acetylcholine receptor expression in the somata of identified molluscan neurons

The effects of Ca²⁺ and vitamin E (-tocopherol) on acetylcholine (Ach)-induced Cl^– currents in LP11 and RBc4 neurons of the snail Helix pomatia have been studied. Injection of Ca²⁺ into the cells and application of vitamin E (10^–5 mole/liter) induced the appearance of potentiation of Ach-induced currents in membrane parts more remote from the axon than the Ach-sensitive regions in the control. The Hill coefficient (n) for such Ach receptors was equal to 0.8, unlike 1.8 for Ach receptors active in the control. Arachidonic acid (10^–5 mole/liter) and phorbol ester TPA (10^–6 mole/liter) inhibited Ach responses, while oleoylacetyglycerol (10^–6 mole/liter) produced no effect. Calmidazolium (10^–6 mole/liter) decreased the effects of Ca²⁺ and vitamin E on Ach responses, while nordihydroquiaretic acid (5 · 10^–6 mole/liter) enhanced the modulating effect of vitamin E and weakened that of arachidonic acid. It is suggested that the expression of Ach receptors activated by Ca²⁺ and vitamin E is mediated through posttranslational mechanisms, since cycloheximide and actinomycin D, inhibitors of protein synthesis, did not influence the effects of C²⁺ and vitamin E. The mechanisms responsible for the stimulating effects of Ca²⁺ and vitamin E are discussed.Translated from Neirofiziologiya, Vol. 25, No. 1, pp. 31–39, January–February, 1993.     

Micropropagation of Capparis decidua (Forsk.) Edgew. — a tree of arid horticulture

A method for micropropagation of mature trees of Capparis decidua was developed. Multiple shoots were obtained from nodal explants on Murashige and Skoog's (1962) medium+0.1mgl^–1 NAA+5.0mgl^–1BAP+additives (50mgl^–1 ascorbic acid and25 mgl^–1 each of adenine sulphate, L-arginine and citric acid) at 28 ± 2°C, 12 h/dphotoperiod and 35–40 mol m^-2s^–1 photon flux density. The shoots were multiplied by (i) subculture of nodal shoot segments onto MS +0.1 mgl^-–1 IAA+1.0mgl^–1 BAPH+additives, and (ii) repeated transfer of original explant onto MS+ 0.1mgl^–1 IAA+mg l^–1 BAP+additives, at intervals of 3 weeks. Sixty to 70% of the shoots rooted when pulse treated with 100 mg l^–1 IBA in half strength MS liquid medium for 4h, and then transferred onto hormone-free half-strength agar-gelled MS basal saltmedium. Incubation in dark at 33 ± 2°C for 6d favoured root induction. In vitro hardened plants were transferred to pots.Abbreviations IAA Indole-3-aceticacid - IBA Indole-3-butyric acid - NAA -naphthaleneacetic acid - BAP 6-benzylaminopurine - Kn 6-furfurylaminopurine - 2-ip Isopentenyl adenine - B₅ Gamborg et al. (1968) medium - MS Murashige and Skoog's (1962) medium - WP Woody plant medium (Lloyd and McCown 1981)     

Ecophysiological characterization of carnivorous plant roots: Oxygen fluxes,respiration, and water exudation

Various ecophysiological investigations on carnivorous plants in wet soils are presented. Radial oxygen loss from roots of Droseraceae to an anoxic medium was relatively low 0.02 – 0.07 mol(O₂) m^{– 2} s^–1 in the apical zone, while values of about one order of magnitude greater were found in both Sarracenia rubra roots and Genlisea violacea traps. Aerobic respiration rates were in the range of 1.6 – 5.6 mol kg^–1 (f.m.) s^–1 for apical root segments of seven carnivorous plant species and 0.4 – 1.1 mol kg^–1 (f.m.) s^–1 for Genlisea traps. The rate of anaerobic fermentation in roots of two Drosera species was only 5 – 14 % of the aerobic respiration. Neither 0.2 mM NaN₃ nor 0.5 mM KCN influenced respiration rate of roots and traps. In all species, the proportion of cyanide-resistant respiration was high and amounted to 65 – 89 % of the total value. Mean rates of water exudation from excised roots of 12 species ranged between 0.4 – 336 mm 3 kg^–1 (f.m.) s^–1 with the highest values being found in the Droseraceae. Exudation from roots was insensitive to respiration inhibitors. No significant difference was found between exudation rates from roots growing in situ in anoxic soil and those kept in an aerated aquatic medium. Carnivorous plant roots appear to be physiologically very active and well adapted to endure permanent soil anoxia.     

The Ca2+-induced leak current in Xenopus oocytes is indeed mediated through a Cl− channel

Defolliculated oocytes of Xenopus laevis responded to removal of external divalent cations with large depolarizations and, when voltage clamped, with huge currents. Single channel analysis revealed a Cl^– channel with a slope conductance of about 90 pS at positive membrane potentials with at least four substates. Single channel amplitudes and mean channel currents had a reversal potential of approximately –15 mV as predicted by the Nernst equation for a channel perfectly selective for Cl^–. Readdition of Ca²⁺ immediately inactivated the channel and restored the former membrane potential or clamp current. The inward currents were mediated by a Ca²⁺ inactivated Cl^– channel (CaIC). The inhibitory potency of Ca²⁺ was a function of the external Ca²⁺ concentration with a half maximal blocker concentration of about 20 m.These channels were inhibited by the Cl^– channel blockers flufenamic acid, niflumic acid and diphenylamine-2-carboxylate (DPC). In contrast, 4,4-acetamido-4-isothiocyanatostilbene-2,2-disulfonicacid (SITS), another Cl^– channel blocker, led to activation of this Cl^– channel. Like other Cl^– channels, the CaIC was activated by cytosolic cAMP. Extracellular ATP inhibited the channel while ADP was without any effect. Injection of phorbol 12-myristate 13-acetate (PMA), a protein kinase C activating phorbol ester, stimulated the Cl^– current. Cytochalasin D, an actin filament disrupting compound, reversibly decreased the clamp current demonstrating an influence of the cytoskeleton.The results indicate that removal of divalent cations activates Cl^– channels in Xenopus oocytes which share several features with Cl^– channels of the CLC family. The former so-called leak current of oocytes under divalent cation-free conditions is nothing else than an activation of Cl^– channels.The microelectrode measurements are part of the PhD thesis of K. Liebold; the patch clamp contributions are part of the PhD thesis of F.W. Reifarth. This study was supported by the Deutsche Forschungsgemeinschaft (We1858/2-l) and by Sonderforschungsbereich 249.     

Stimulatory effect of hormonal signaling factors on (Ca2+−Mg2+)-ATPase activity in rat liver plasma membranes: Cross talk with regucalcin

The effect of hormonal signaling factors on (Ca²⁺–Mg²⁺)-ATPase activity in rat liver plasma membranes was investigated. The presence of inositol-glycan (10^–7–10^–5M), dibutyryl cAMP (10^–4 and 10^–3M) or inositol 1,4,5-trisphosphate (IP₃; 10^–6 and 10^–5 M) in the enzyme reaction mixture produced a significant increase in (Ca²⁺–Mg²⁺)-ATPase activity. These effects were completely inhibited by the presence of vanadate (10^–4 M), an inhibitor of the enzyme phosphorylation, and N-ethylmaleimide (5×10^–3 M), a SH group modifying reagent. Meanwhile, regucalcin, a Ca²⁺-binding protein isolated from rat liver cytosol, increased the enzyme activity by binding to the SH groups of (Ca²⁺–Mg²⁺)-ATPase in liver plasma membranes. The presence of regucalcin (0.25 M) with an effective concentration completely inhibited the effect of inositol-glycan (10^–5 M) to increase (Ca²⁺–Mg²⁺)-ATPase activity, while the effect of dibutyryl cAMP (10^–3M) or IP₃ (10^–5M) was not altered. The inositol-glycan effect was not modulated by the presence of dibutyryl cAMP or IP₃. Now, the preincubation of the plasma membranes with regucalcin did not modify the effect of inositol-glycan on the enzyme activity, suggesting that regucalcin competes with inositol-glycan for the binding to the plasma membranes. The present results suggest that there may be a cross talk with regucalcin and hormonal signaling factors in the regulation of (Ca²⁺–Mg²⁺)-ATPase activity in liver plasma membranes.     

Efficiency of soil and fertilizer nitrogen of a sod–potato system in the humid,acid and cool environment

It was hypothesized that soil N variability, and fertilization and cropping management affect potato (Solanum tuberosum L.) growth and fertilizer N efficiency. Following a 20-year sod breakup on a loamy soil in eastern Quebec, Canada (46°37 N, 71°47 W), we conducted a 3-year (1993–1995) study to investigate the effects of soil pool N and fertilizer N management on non-irrigated potato (cv. Superior) tuber yield, fertilizer N recovery (NRE), and residual N distribution in soils under humid, cool and acid pedoclimatic conditions. The fertilizer N treatments consisted of a control, side-dress at rates of 70, 105 and 140 kg ha^–1, and split applications (at seeding and bloom) at rates of 70+70, 105+70 and 140+70 kg ha^–1, respectively. Soil acidity was corrected with limestone following the plow down of the sod. Years of cropping, main effect of N treatment, and year and fertilizer N interaction were significant on total and marketable tuber yields and N uptake, which were significantly related to soil N, and root growth. Apparent NRE ranged between 29 and 70%, depending on years and N rates. Total tuber yield, N uptake, soil N use and NRE were significantly higher in the first (sod–potato) year, but decreased by 41.8, 22.7, 21.4 and 14.7%, respectively, in the third (sod–potato–potato–potato) year. Initial soil N pool was declined by 75% following the 3-year cropping. In 2–3 years, the side-dress N (140 kg ha^–1) increased significantly tuber yields (11.4–19.8%) compared to the split N (70+70 kg ha^–1). Higher split N had no effect on tuber yield and N uptake but increased residual N at harvest. Unused fertilizer N was strongly linked (R ²=0.98) to fertilizer N rates. Time factor and N treatment had significant effects (P<0.0001) on loss of N to below the root zone. Smaller scale rate and timing of split N need to be further determined. Increasing fertilizer N use efficiency could be expected with sod breakup and 75% of regional recommendation rate under humid, cool and acid pedoclimatic conditions.     

Studies on octylphenoxy surfactants: X. Effect of oxyethylene chain length on ethylene production by cowpea and mung bean and comparison with linear alcohol hydrophobes

The effects of ethoxy (EO) chain length on surfactant-induced ethylene production for selected octylphenoxy (OP) and linear alcohol (LA) surfactants were established using primary leaves of cowpea (Vigna unguiculata (L.) Walp. subsp. unguiculata Dixielee) seedlings. OP surfactant-induced ethylene production was concentration dependent and decreased log linearly with increasing EO content. C_12–15 LA-induced ethylene production decreased log linearly with increasing EO content at 0.1%; however, at 1.0% the relationship was curvilinear with maximum response at 7 EO. Relationships for the C_9–11 and C₉ LA series were nonlinear with greatest biological activity at intermediate (8–12) EO content. Short EO chain length OP surfactants were only slightly water soluble, and induced low levels of ethylene production and phytotoxicity. Addition of OP+1EO to a long chain, water soluble, non-ethylene inducing surfactant (OP+40EO) solution significantly increased ethylene production by OP+1EO in cowpea. A similar response was found for surfactant-induced phytotoxicity and EO chain length as between ethylene production and EO content. Similar EO chain length and ethylene production relationships were found for germinating mung bean (Vigna radiata (L.) R. Wilcz) seeds as for ethylene production and phytotoxicity in cowpea. Radicle growth was markedly inhibited by OP surfactants with an EO chain length of 10 or less and, in some cases, radicles were irreversibly damaged by ethylene inducing surfactants.     

Synthesis of carotenoids by<Emphasis Type="Italic"> Rhodotorula rubra</Emphasis> GED8 co-cultured with yogurt starter cultures in whey ultrafiltrate

Two cultures, a yeast (Rhodorula rubra GED8) and a yogurt starter (Lactobacillus bulgaricus 2–11+Streptococcus thermophilus 15HA), were selected for associated growth in whey ultrafiltrate (WU) and active synthesis of carotenoids. In associated cultivation with the yogurt culture L bulgaricus 2–11+S. thermophilus 15HA under intensive aeration (1.3 l^–1min^–1 air-flow rate) in WU (45 g lactose l^–1), initial pH 5.5, 30 °C, the lactose-negative strain R. rubra GED8 synthesized large amounts of carotenoids (13.09 mg l^–1 culture fluid). The carotenoid yield was approximately two-fold higher in association with a mixed yogurt culture than in association with pure yogurt bacteria. The major carotenoid pigments comprising the total carotenoids were -carotene (50%), torulene (12.3%) and torularhodin (35.2%). Carotenoids with a high -carotene content were produced by the microbial association 36 h earlier than by Rhodotorula yeast species. No significant differences were notd in the ratio between the pigments synthesized by R. rubra GED8+L. bulgaricus 2–11, R. rubra GED8+S. thermophilus 15HA, and R.rubra GED8+yogurt culture, despite the fact that the total carotenoid concentrations were lower in the mixed cultures with pure yogurt bacteria.     

Mechanisms of cytoplasmic calcium signalling in cerebellar bergmann glial cells

Mechanisms underlying intracellular calcium signals in Bergmann glial cells evoked by various neurotransmitters were investigated in experiments on cerebellar slices acutely isolated from 30-day-old mice. [Ca²⁺] _in values were measured by means of a Ca²⁺-sensitive fluorescent probe fura-2. Extracellular application of ATP (10–100 µM), histamine (10–100 µM), or noradrenaline (or adrenaline, 0.1–10.0 µM) caused a temporary increase in cytoplasmic Ca²⁺ concentrations. The effect persisted in Ca²⁺-free extracellular solution and was blocked with thapsigargin (500 nM) or a specific blocker of the inositol-1,4,5-trisphosphate-sensitive intracellular channels heparin. Based on the pharmacological analysis, we postulate the involvement of P₂ purinoreceptors, ₁-adrenoreceptors, and H₁ histamine receptors in an agonist-activated increase in [Ca²⁺] _in in Bergmann glia. Thus, ATP, monoamines, or histamine induce calcium signal generation in Bergmann glial cells via activation of Ca²⁺ release from the inositol-1,4,5-trisphosphate-sensitive internal stores.Neirofiziologiya/Neurophysiology, Vol. 26, No. 6, pp. 417–419, November–December, 1994.     

Redox potentials of the blue copper sites of bilirubin oxidases

The redox potentials of the multicopper redox enzyme bilirubin oxidase (BOD) from two organisms were determined by mediated and direct spectroelectrochemistry. The potential of the T1 site of BOD from the fungus Myrothecium verrucaria was close to 670 mV, whereas that from Trachyderma tsunodae was > 650 mV vs. NHE. For the first time, direct electron transfer was observed between gold electrodes and BODs. The redox potentials of the T2 sites of both BODs were near 390 mV vs. NHE, consistent with previous finding for laccase and suggesting that the redox potentials of the T2 copper sites of most blue multicopper oxidases are similar, about 400 mV.     

Influence of inorganic microelements on the production of camptothecin with suspension cultures of <Emphasis Type="Italic">Camptotheca acuminata</Emphasis>

The effects of eight microelements (I^–, BO₃ ^3–, MoO₄ ^2–, Co²⁺, Cu²⁺, Mn²⁺, Fe²⁺, Zn²⁺) on the biosynthesis of camptothecin and the growth of suspension cultures of Camptotheca acuminata were studied. The increase of I^– to 25 M l^–1, Cu²⁺ to 1 M l^–1, Co²⁺ to 2 M l^–1 and MoO₄ ^2– to 10 M l^–1 in Murashige and Skoog (MS) medium resulted in 1.66, 2.84, 2.53 and 2.04 times higher of camptothecin yield than that in standard MS medium respectively. Combined treatment of I^– (25 M l^–1), Cu²⁺ (1 M l^–1), Co²⁺ (2 M l^–1) and MoO₄ ^2– (10 M l^–1) lead to improve cell dry weight, camptothecin content, and camptothecin yield to 30.56 g l^–1, 0.0299%, and 9.15 mg l^–1, respectively, which were 20.2, 208.9 and 273.8% increment respectively when compared with those of control.     

Response of corn (Zea mays L.) to manganese application on Atlantic Coastal Plain soils

Field research was conducted on four Atlantic Coastal Plain soils in the United States to evaluate response of corn (Zea mays L.) plants to Mn application. The soils under study were classified as either Aeric or Typic Ochraquults. Manganese application increased corn grain yields by an average of 1195 kg ha^–1 on the four soils. The average grain yields on the soils were 7955 kg ha^–1 for the control and 9150 kg ha^–1 for the +Mn treatment. A Mitscherlich plant growth model was used to establish relationships between percent maximum grain yield and Mn concentration in the ear leaf at early silk (r=0.87, =0.01) and in the mature grain (r=0.58, =0.01). Based on 90% of maximum yield as the definition of the critical deficiency level, the critical Mn deficiency levels calculated with parameters from the Mitscherlich model were 10.6 mg kg^–1 in the ear leaf and 4.9 mg kg^–1 in the grain.     

Characterization of a secreted cholesterol oxidase from Rhodococcus sp.GKI (CIP 105335)

Extracellular cholesterol oxidase (COX) (EC 1.1.3.6) was produced by Rhodococcus sp. GK1 cells grown in a defined mineral salt medium containing a mixture of phytosterols (sitosterol, campesterol, stigmasterol) as the sole source of carbon and energy. In the same time, the sterols acted as enzyme inducers. The medium was enriched with yeast extract in order to stimulate enzyme secretion. COX was purified from the culture supernatants by affinity-like chromatography on a column packed with kieselguhr and cholesterol. Enzyme bound onto the column was eluted with 0.05 M phosphate buffer pH 7.0 containing Triton X-100 at 0.1% (w/v). Some properties of the purified COX were determined. Its specific activity at pH 7.0 and 30 °C, was around 5.5 units mg^–1. The molecular mass of the enzyme, as estimated by SDS-PAGE, was 59 kDa. Its isoelectrofocusing point was around pH 8.9. The C-5 double bond and the alkyl chain moiety in sterol molecules were necessary for an adequate oxidation of the sterol 3-ol. Enzyme inhibition by the ions (0.1 mM): AsO₂ ^–, Ba²⁺, Co²⁺, Cd²⁺, Cu²⁺, N₃ ^–, Ni²⁺, and Pb²⁺ was negligible (around 10%). However, COX inhibition by 0.1 mM of either Zn²⁺, 2-[(ethylmercurio)-thio]benzoic acid, or Hg²⁺ was 18%, 22% and 93% respectively. Inhibition of activity by Hg²⁺ was significant, even at 1 M. The purified COX (0.1–0.15 mg ml^–1 in 0.05 M phosphate pH 7.0) was relatively heat-stable at temperatures up to 50 °C. At this temperature, the half-life of its activity was around 70 min. However, 90% of the enzyme initial activity was lost by 20 min incubation at 60 °C. The aminoacid sequence of the COX N-terminal segment was: H2N–Ala–Pro–Pro–Val–Ala–Ser–X–Arg–Tyr–X–(Phe)– (X might be 2 Cys residues).