EPR characterization of oxide supported transition metal ions: Relevance to catalysis

EPR spectroscopy is a powerful tool to identify at a molecular level, the different steps of catalyst preparation, and of catalytic reactions:

1. Deposition of paramagnetic transition metal ions onto a support is monitored, and the coordination sphere of the metallic center is characterized by EPR.
2. The catalyst is also characterized after activation (thermal oxidation or reduction):

- the distribution among the different sites in zeolites can be determined;

- the dispersion of the active phase may be appreciated;

- the unsaturation degree of the active site may be evaluated using probe molecules such as water or¹³C enriched carbon monoxide.

1. The catalytic mechanisms can be investigated by studying the elementary steps of the catalytic reaction, as illustrated for methanol oxidation over Mo/SiO₂ catalysts whose EPR results have extended the reaction mechanism proposed on the basis of kinetic data. In addition, reaction intermediates may be isolated inquasi-in situ conditions as in the case of olefin oligomerization catalyzed by Ni/SiO₂ systems.

     

Uptake of guanine by synchronizedChlorella fusca

1. The transport of guanine in autospores of light-dark synchronizedChlorella fusca was studied using radioactive guanine in the concentration range of 4 nM to 50 μM.
2. The transport system was constitutive, it had high specificity for the permeant, and theQ ₁₀ value was in the range of 1.5 to 2.2. At concentrations lower than 0.2 μM the half saturating constant, S_0.5 was 1 μM both for cells kept in dark and cells kept in light. At higher concentrations the S_0.5 of darkened cells was about 0.23 μM, while that of illuminated cells was unchanged. Only above 0.2 μM guanine did illumination of the cells or addition of glucose increase the transport rate.
3. Guanine which had accumulated did not leak out at temperatures below 45°C or by treatment with 10 μM dinitrophenol, which completely inhibited transport. Furthermore, the accumulated guanine did not exchange with exogenous guanine.
4. The guanine accumulated, more than 10⁵-fold over the external concentration, showing that the transport, was active.
5. The initial transport rate per cell revealed annual fluctuations.

     

Habitability of the early earth: Clues from the physiology of nitrogen fixation and photosynthesis

In the absence of direct evidence concerning the nature of the early Earth environments, it is acceptable under the uniformitarian principle to attempt to define primitive habitats from modern procaryotic physiology. Combining the rock and fossil record with present phylogenetic reconstuctions, application of this paleoecological approach to the evolutionary biochemistry and physiology of nitrogen fixation and photosynthesis leads to several inferences about the nature of Archean environments:

1. To stimulate nitrogenase evolution and avoid its repression, the activity of the NH ₄ ⁺ ion was less than 10^?3, and probably lower.
2. To be consistent with a moderately protective ozone screen, while not also repressing nitrogenase activity, incursions of abiotic dissolved oxygen at levels in the range 10^?1.2?10^?3.5 PAL would have been acceptable.
3. To induce the formation and activity of RuBP carboxylase, the pCO₂ was less than 100 PAL.
4. To support Photosystem I activity, sulfide concentrations of at least 10^?4 M were present in the photic zone.
5. To avoid a too-rapid oxidation of sulfide, the pH was probably between 6–7, where H₂S exceeds HS^?.

Evolutionary ‘pressure’ to stimulate the later development of oxygenic photosynthesis (Photosystem II), would require several subsequent habitat modifications:

1. Lowering the sulfide to < 10^?4 M to inhibit Photosystem I.
2. Raising the pH above neutral (HS^? > H₂S), to mediate more rapid oxidation of HS^?.
3. Maintaining either an illumination below 300–400 lux (to avoid photosynthetic O₂ self-repression of nitrogen fixation), or an adequate local source of combined nitrogen (aNH ₄ ⁺ > 10^?4) to repress nitrogen fixation entirely.

     

Total recovery of O2 evolution and nanosecond reduction kinetics of chlorophyll-a II + (P-680+) after inhibition of water cleavage with acetate

Oxygen evolution and reduction kinetics of the photooxidized Chl-a_II ⁺ have been measured in oxygen-evolving complexes from the thermophilic cyanobacterium Synechococcus sp.

1. Incubation of PS II particles with acetate resulted in an inhibition of oxygen evolution and a retardation of the Chl-a_II ⁺=reduction kinetics from the nanosecond range to the microsecond range, indicating a modification of the donor side of photosystem II (PS II).
2. After the first two flashes given to a dark-adapted, acetate treated sample, Chl-a_II ⁺ was re-reduced with a half-life time of 160 μs by a component of the donor side of PS II. Under repetitive excitation Chl-a_II ⁺ was re-reduced in 500 μs by electron back reaction from the primary acceptor Q_A ^- (X-320^-). Obviously, in the presence of acetate only two electrons are available from the donor side.
3. Both oxygen evolution and nanosecond reduction kinetics of Chl-a_II ⁺ were restored to the control level when acetate was removed.
4. The results indicate a tight coupling between O₂ evolution and nanosecond reduction kinetics of Chl-a_II ⁺.
5. The reversible inhibition is probably due to a replacement of Cl^- by acetate within the water splitting enzyme.
6. Due to its strongly retarded kinetics, the reversibly modified system may facilitate investigations of the mechanism of the donor side.

     

The role of mitochondria in modifying the cellular inoc environment: Studies of the kinetic accumulation of calcium by rat liver mitochondria

1. The affinity of ATP-supported Ca²⁺ accumulation for both Ca²⁺ and ATP was determined from initial rate studies employing isolated rat liver mitochondria. TheK _m values for “free” Ca²⁺ and ATP were calculated to be of the order of 2 μM and 100 μM, respectively. TheK _m for ATP decreased as the Ca²⁺ concentration was increased.
2. The curve relating initial rates of Ca²⁺ accumulation to Ca²⁺ concentration was singmoidal in shape; values obtained for the Hill coefficient were in the range 1.5–1.9.
3. Concomitant with the ATP-stimulated accumulation of Ca²⁺, ATP translocation was itselt increased in the presence of Ca²⁺. This stimulation took place independently of Ca²⁺ accumulation.
4. Decreasing the pH of the incubation medium decreased the rate of Ca²⁺ accumulation. This inhibition was competitive in that the affinity of mitochondrial for Ca²⁺ could be altered. The maximal rate of accumulation did not change with change in pH.
5. The permeant anions inorganic phosphate and acetate stimulated the accumulation of Ca²⁺ in a non-competitive manner. Both theV _max and K_m varied when either of the anions were present.
6. The data are discussed in relation to the role that mitochondria play in controlling the cellular ionic environment.

     

Charakterisierung eines phagenähnlichen Partikels aus Zellen von Nitrobacter

1. Polyhedral particles were isolated from cells of Nitrobacter winogradskyi and of Nitrobacter strains K₁, K₄ and α₁. Their physical and biological properties are characterized.
2. The investigated strains contain polyhedral particles, 1000–1200 Å in size. With increasing age of the culture more particles are found in cells of Nitrobacter. Simultaneously the number of colony producing nitritoxidants decreases.
3. In strain α₁ the loss of the capability to form colonies is connected with partial lysis of the cell and release of particles.
4. A homogeneous fraction of particles was obtained by zone density gradient centrifugation in Tris-Mg-SH-buffer.
5. The polyhedral particles have a sedimentation coefficient of s _w,20 ⁰ =825S and a CsCl-buoyant density of ?²⁵ g/cm³.
6. Based on the determined properties the particles are classified as phage-like Nitrobacter particles Nb₁.

     

The use of a continuous assay system to show the stimulation of phosphoenolpyruvate production in intact mitochondria by valinomycin and by Mn2+

1. A method for the direct recording of the PEP efflux from isolated mitochondria is described.
2. This method has been used to show the stimulation of PEP efflux by externally added Mn⁺⁺ ions.
3. Valinomycin, uncoupler and oleate were also shown to stimulate PEP efflux.
4. Valinomycin caused an increase in the internal concentration of both PEP and citrate.
5. The results indicate that the major pathway of PEP synthesis in isolated mitochondria is via PEP carboxykinase and the results do not call for an unknown pathway of metabolism.
6. Two interactions between PEP and citrate are described; competition for the mitochondrial interior and the stimulation of PEP production by citrate.

     

Seasonal variation in the biochemical composition of Mytilus viridis at Ratnagiri on the West Coast of India

1. The seasonal variation in the water, protein, fat and glycogen contents of the mussel, Mytilus viridis has been studied for the year March, 1974 to March, 1975.
2. The water level increased during the monsoon season and decreased in summer.
3. The level of protein, fat and glycogen showed correlation with the reproductive cycle of the mussel.
4. The protein level was high when the mussels were mature and dropped during the breeding period.
5. During sex change from male to female in May the protein level remained high whereas during sex change from female to male in October and November it was low.
6. The fat level was high in mature mussels and declined on spawning.
7. The glycogen level was at its peak in immature mussels and low in mature.

     

Molten Earth and the origin of prebiological molecules

Evidence for the molten Earth at its accretion time has been accumulated through the geochemical investigations and the observations of the surfaces of planets by space probes such as Venera 8, Mariner 9, Surveyor, Luna, and Apollo. The primitive terrestrial atmosphere might have been derived from the volcanic gases, as suggested by Rubey, but of a higher temperature than so far assumed. A thermochemical calculation of the composition of the volcanic gas suggests the following possibilities:

1. Large amounts of H₂ and CO were present in the primitive atmosphere. This gives a theoretical basis for the HCN-production experiment by Abelson.
2. HCHO and NH₃ existed in the primitive oceans, of the amount comparable with the weight of the present biosphere.
3. Plenty of NO ₃ ⁻ , SO ₄ ⁻⁻ , and PO ₄ ⁻⁻⁻ were expected in the primitive oceans. The NO ₃ ⁻ ions might have been useful for the nitrate respiration advocated by Egami.

In an appendix, it is argued, on the basis of the observational evidence of the exospheric temperatures of planets by space probes, that a highly reducing atmosphere would (if it existed on the primitive Earth) have disappeared very quickly due to the thermal escape of hydrogen from its exosphere.     

Presence of a Na+/H+ antiporter in Methanobacterium thermoautotrophicum and its role in Na+ dependent methanogenesis

Methane formation from H₂/CO₂ by methanogenic bacteria is dependent on Na⁺ ions. In this communication it is shown with Methanobacterium thermoautotrophicum that a Na⁺/H⁺ antiporter plays a role in methane formation from H₂ and CO₂ and in the regulation of the ΔpH. This is based on the following findings:

1. Li⁺ ions, an alternative substrate of Na⁺/H⁺ antiporters, could replace Na⁺ in stimulating methanogenesis from H₂ and CO₂.
2. Harmaline, amiloride, and NH ₄ ⁺ , which are inhibitors of Na⁺/H⁺ antiporters, inhibited methanogenesis; inhibition was competitive to Na⁺ or Li⁺.
3. Addition of Na⁺ or Li⁺ rather than of other cations to cell suspensions resulted in an acidification of the suspension medium. The rate and extent of acidification was affected by those inhibitors, which inhibited methanogenesis competitively to Na⁺ or Li.
4. During methane formation from H₂ and CO₂ the generation of a ΔpH (inside alkaline) was dependent on the presence of Na⁺ or Li⁺. However, methanogenesis was also dependent on Na⁺ or Li⁺ under conditions where ΔpH was zero.
5. ATP synthesis driven by an electrogenic potassium efflux was significantly enhanced in the presence of Na⁺ or Li⁺. Na⁺ or Li⁺ were shown to prevent acidification of the cytoplasm under these conditions.

     

Hen's egg yolk alkaline phosphatase: General characterization and kinetic stuy with inhibitors

• 1.1. The non-specific hen's egg yolk alkaline phosphatase is a metalloprotein (Zn²⁺?) composed of two identical inactive subunits.
• 2.2. A second metal site preferably binds Mg²⁺ (15-fold activation). Me(II))H₂O)H⁺, a charged arginine, and tyrosine in the active site are involved in positioning and binding of the substrate and metal ion.
• 3.3. Substrate inhibition differs with pH. This may be related to the presence of two active sites in the enzyme, one in each subunit.
• 4.4. Uncompetitive inhibition with L-phenylalanine and analogues suggests a phosphorylated intermediate.
• 5.5. Inhibition is weakly competitive with P_i strong non-competitive with PPi as compared to Mg²⁺-free PP_i, and partially competitive with arsenate.
• 6.6. The purified enzyme is stabilized and activated by amines and proteins.

     

Effects of in vivo ethanol administration on Ca2+/Mg2+ ATPase and ATP-dependent Ca2+ uptake activity in synaptosomal membranes

1. Acute administration of ethanol (4 g/kg, i.p.) to mice inhibits the sequestration of calcium into endoplasmic reticulum-like organelles in synaptosomal membranes.
2. Ethanol administration inhibits both Ca²⁺-stimulated adenosine triphosphate hydrolysis and ATP-dependent calcium uptake in the vesicles at time of loss of righting reflex.
3. At recovery of righting reflex, the Ca²⁺-ATPase activity returns to normal levels, while the ATP-dependent uptake remains inhibited.
4. The effect of ethanol is specific for the sequestration (active transport) of calcium since calcium binding to synaptic membranes is not altered.
5. Alteration in mechanisms responsible for synaptosomal buffering of cytosolic Ca²⁺ levels by in vivo ethanol may contribute to altered transmitter release rates following ethanol adminstration.

     

Anaerobic respiration in latex ofHevea brasiliensis substrate and limiting factors

The site of anaerobic respiration in the latex is the serum. The main respiratory substrate is fructose. The CO₂ formation in serum is increased by additional fructose on the average about 2.5–3 times. Glucose does not influence CO₂ evolution by serum but slightly increases O₂ consumption. With respect to sugars, latex serum contains essentially only sucrose and a low amount of raffinose. During the incubation of serum sucrose is hydrolysed, the fructose component is immediately utilized in respiration and glucose accumulates. The rate of CO₂ formation in latex as influenced by fructose is negatively related to the rubber content of the latex. Latex with a high rubber content reacts only slightly or not at all on additional fructose. The main limiting factors of latex respiration and sugar utilization are the following:

1. The deficiency of substrate, due to low activity of β-fructofuranosidase.
2. The rate of glucose phosphorylation (D'Auzac, Jacob 1967).
3. Presumably the low activity of phosphoglucoisomerase.
4. The rubber content of the latex.
5. The concentration of CO₂ in latex; this factor may be important in vivo, in the laticiferous system.

     

Regulation of rat-kidney cortex fructose-1,6-bisphosphatase activity. I. Effects of fructose-2,6-bisphosphate and divalent cations

• 1.1. The native rat-kidney cortex Fructose-1,6-BPase is differentially regulated by Mg²⁺ and Mn²⁺.
• 2.2. Mg²⁺ binding to the enzyme is hyperbolic and large concentrations of the cation are non-inhibitory.
• 3.3. Mn²⁺ produces a 10-fold rise in V_max higher than Mg²⁺. [Mn²⁺]_0.5 is much larger than [Mg²⁺]_0.5. At elevated [Mn²⁺] inhibition is observed.
• 4.4. Mg²⁺ and Mn²⁺ produce antagonistic effects on the inhibition of the enzyme by high substrate.
• 5.5. Fru-2,6-P₂ inhibits the enzyme by rising the S_0.5 and favouring a sigmoidal kinetics.
• 6.6. The inhibition by Fru-2,6-P₂ is released by Mg²⁺ and more powerfully by Mn²⁺ increasing the I_0.5.

     

Functional characterisation of a purified homogeneous Photosystem II core complex with high oxygen evolution capacity from spinach

The functional properties of a purified homogeneous spinach PS II-core complex with high oxygen evolution capacity (Haag et al. 1990a) were investigated in detail by measuring thermoluminescence and oscillation patterns of flash induced oxygen evolution and fluorescence quantum yield changes. The following results were obtained:

1. Depending on the illumination conditions the PS II-core complexes exhibit several thermoluminescence bands corresponding to the A band, Q band and Z_v band in PS II membrane fragments. The lifetime of the Q band (T_max=10°C) was determined to be 8s at T=10°C. No B band corresponding to S₂Q_B ^? or S₃Q_B ^? recombination could be detected.
2. The flash induced transient fluorescence quantum yield changes exhibit a multiphasi relaxation kinetics shich reflect the reoxidation of Q _A ^? . In control samples without exogenous acceptors this process is markedly slower than in PS II membrane fragments. The reaction becomes significantly retarded by addition of 10 μM DCMU. After dark incubation in the presence of K₃[Fe(CN)₆
3. Excitation of dark-adapted samples with a train of short saturating flashes gives rise to a typical pattern dominated by a high O₂ yield due to the third flash and a highly damped period four oscillation. The decay of redox states S₂ and S₃ are dominated by short life times of 4.3 s and 1.5 s, respectively, at 20°C.

The results of the present study reveal that in purified homogeneous PS II-core complexes with high oxygen evolution isolated from higher plants by β-dodecylmaltoside solubilization the thermodynamic properties and the kinetic parameters of the redox groups leading to electron transfer from water to Q_A are well preserved. The most obvious phenomenon is a severe modification of the Q_B binding site. The implications of this finding are discussed.     

Growth of a floating aquatic weed,Salvinia under standard conditions

1. Growth of the floating aquatic weed, Salvinia, in sterile culture was exponential for at least 2 weeks under standardized conditions.
2. Increase in light intensity or in CO₂ resulted in increases in growth rate, but did not extend the exponential period of growth.
3. This aquatic plant, like many others, discriminates against calcium relative to strontium.
4. In culture Salvinia exhibited luxury consumption of N and P.
5. Because of high C/N ratios, Salvinia may not be a favorable source of animal food, but might be useful in nutrient removal schemes.
6. In sterile culture, S. molesta produced fewer leaves than S. minima, but maintained a significant increase in leaf area and dry weight. This may be correlated with the ability of the first species to rapidly spread over tropical waterways.

     

Ostrich fructose-1,6-bisphosphatase: Kinetic characterization of the liver isoenzyme

• 1.1. Purified ostrich (Struthio camelus) liver fructose-1,6-bisphosphatase exhibited an absolute requirement for Mg²⁺.
• 2.2. The enzyme catalyzed the hydrolysis of fructose-1,6-bisphosphate, sedoheptulose-l,7-bisphosphate and ribulose-l,5-bisphosphate.
• 3.3. S_0.5 for substrate was 1.4 μM.
• 4.4. AMP was a potent non-competitive inhibitor with respect to substrate (K_i of 25 μM).
• 5.5. Fructose-2,6-bisphosphate was a potent competitive inhibitor of the enzyme (K_i of 4.8 μM).

     

The reproductive cycle in a typical estuarine mollusc Nausitora hedleyi Schepman based on a study of the Gonad Index

1. Previous work on the methods employed for the determination of the breeding season of shipworms is briefly reviewed.
2. The method adopted for studying the reproductive cycle by using the “gonad index” is described.
3. The reproductive cycle of Nausitora hedleyi is described in detail based on a study of the gonad index of different sexes collected at monthly intervals from the estuarine environment of Cochin harbour.
4. The fact that breeding is restricted as marked by seasonal activity is shown from the size and activity of the gonad during the different months of the year.
5. The environment, and the hydrographic conditions prevailing in the habitat of N. hedleyi in the Cochin harbour are described.

     

Les brisures de symetrie du temps

Introduction

Atoms theory and symmetry theory dominated physics. Symmetry propagation and interactions verify the Curie principle. But its violation by symmetry breaking is spontaneous.Fragility is creative. An information breaks a generalized symmetry. Results on symmetry breakings are not valid for fuzzy symmetries. The breaking of a fuzzy symmetry leads only to a pour symmetry (Fig.1). Homogeneity breaking, and atom of time are not usual concepts. We examine in this work symmetry breakings which generate the living time.

Relativistic Time-Space Breaking

1. Medium and environment of living define ordinary referential of space and referential of time. Astronomical phenomena following classical mechanics and microphysical phenomena following quantum mechanics can be written with the same t coordinate.
2. Relativity corrections. Schrödinger's Quantum mechanics (Eq.0) approximately governs molecular systems (Relativity corrections can be expressed as physical effects in the above defined referential).
3. Time reversal symmetry. The well-known Wigner's transformation determines the microscopic reversibility.
4. The three essential particle-vacancy equilibria. This transformation is verified by all particle-vacancy reciprocity. Vacancy moves like particle but with negative moment and positive kinetic energies. Only three biochemical equilibria admit this time reversal symmetry, namely: oxydo-reduction, acido-basicity, fluidity-viscosity. In these case, reacting electron, solvated proton, water molecule are respectively antagonist of the corresponding vacancy.
5. Fuzzy character of time reversal symmetry. Dirac's equation does not admit this symmetry which only appears at the “non relativistic” limit of quantum phenomena. Hence particle-vacancy reciprocity is fuzzy according to the experimental evidence. (Laforgue et al., 1988).

Oriented Time

1. From the universal reversible time, an additional breaking generates the oriented time, both in the astronomical and in the living matter.
2. Irreversibility for the environment. We refer to Prigogine and Stengers (1988).
3. Irreversibility for the living matter. We refer to Lochak (1986). Because equation (0), above discussed, is “microreversible” the second breaking could come from an additional term vanishing in the stationary states but increasing with time in evolutionary processes.
4. Negative times. Taking into account the fuzzy character of the time reversed symmetry, the third breaking cannot suppress completely the occurrence of negative times. Reversed time is controlled by direct time. Except in the three above reported cases, time reversal symmetry is not verified by the medium. Free motion of the particle following eg.(0) or of the vacancy following time reversal reciprocal equation takes place only during short jumps from an interaction site to an other. Fig. 2 schematizes the law of motion of the electric charge corresponding to the transport by proton or by proton vacancy in an unitary field (fluctuations are neglected). The reserved jumps are estimated in the range of 10^?12s. It is not excluded that such a jump can control a direct phenomenon.
5. The living time. Biological phenomenon appears as an oriented set of events. Nevertheless latency or exaltation phases could be perceived. This modulation could be described by positive and negative times additional to the basic time. (Negative can be interpreted as above)

Living produces Time

1. That were not understandable, if time was only a frame, in which change occurs. Taking chance as frame and time as effect, we regard biological activity as integrating reversible and irreversible time. Living synchronizes internal and external time by its own effort as it results (Lestienne, 1990) from Chronobiology.
2. Time modulation. Let us consider the dy₁...dy_i...dy_p changes in the variables of the systems, dy={dy_i} has produced dt. We proof (eq.(1) to (4)) that time is modulated by a φ_(y) speed coefficient depending on the medium. t_modulated=tφ _(y) ^?1
3. The production of reversible time (e.g.acido-basicity) determines time modulation. As above reported it remains some reversibility effects (jumps of negative time) which modulate time. E.G., if an important amount of reagent is necessary to modify an acid-base equilibrium, φ_(y) is small.
4. Time modulation and activation-repression reciprocity. As well-known, long t_modulated means repression, short t_modulated means exaltation. Extrema of ? are symmetrical because particle and vacancy are reciprocal. Nevertheless reciprocity is not perfect. E.g., on fig. 3, the wet receptor determines the cell increasing, the dry receptor the cell senescence of a certain alga (Lück, 1962).
5. Irreversible time production. Medium accepts entropy. Hence it acts in the second breaking of time. Living extracts the free energy from the medium, like a dissipative structure. That insures an operative point far from the thermodynamical equilibrium.

Consumption of Time

1. The three followings correspond to the more trivial time consumption.
2. Rhythmical time. Free energy flux is favourable to the arising of order in space or time. This later gives a structure to the living time.
3. Mutual dependence of reversible time and rhythms. Time irreversible structure can be controlled by the above considered particle-vacancy equilibrium. Consequently the living time (modulated and structured) is a chemical time connected to molecular properties and to statistical thermodynamics. Practically, the connection between chronobiology and chemistry is important. The use of drugs could be interpreted as a response to an aggression against biorhythms.
4. Lifetime. The dead-birth rythm can be broken in two ways: evolution or indefinite life. This later is non exceptional for the living matter, e.g. in the vegetals where it is connected with the chlorophyllic assimilation; the time reversal significance of which is evident.
5. The plan of the alchemist. Indefinitely life has fascinated individuals. Do the human species becomes better adapted by a longer life?

Conclusions

1. Atoms of time could exist.
2. Biological time is defined by the breaking of five generalized symmetries, namely: Minkovski's space symmetry, reversibility, homogeneity, rhythmicity, generations reproduction.
3. Environment and medium determine non relativistic, oriented, structured time.
4. At the microphysical scale, a fuzzy time reversal symmetry takes place, the breaking of which is not complete. Reversible time and dominating irreversible time are integrated in living phenomena.
5. Three fundamental particle-vacancy reciprocities admit a part of reversibity. Irreversibility governs the all others phenomena.
6. Time is produced chemically.
7. A new perspective is the connection between chemical equilibria and rhythms including the time of the life.

     

Toxicity of cadmium administration to the toad and the treatment of its poisoning with EDTA

• 1.1. The effect of cadmium administration on female Bufo regularis was studied. The median lethal doses were 22, 18, 15 and 6.2 Cd²⁺/kg after 24, 48, 72 and 96 hr respectively.
• 2.2. After a single intramuscular injection of 6.2 Cd²⁺/kg (representing 96-hr ld₅₀), the results indicated that Cd²⁺ causes severe physiological abnormalities to this experimental animal.
• 3.3. The serums alanine aminotransferase (AlAt), aspartate aminotransferase (AAt), alkaline phosphatase (A1P) and lactic dehydrogenase (LDH) were elevated while the calcium serum was not influenced by Cd²⁺ throughout the experimental period
• 4.4. On the other hand, phosphorus, total protein and total bilirubin were increased.
• 5.5. EDTA treatment (0.2 mmole/kg protected female toads from mortality up to 20 mg Cd²⁺/kg. It overcame the physiological alterations that were caused by the Cd²⁺ injection.
• 6.6. This may be due to the fact that Cd²⁺ is bound to EDTA in a strong complex which is readily excreted via the kidneys.