Investigating the potassium interactions with the palytoxin induced channels in Na+/K+ pump

K(+) has been appointed as the main physiological inhibitor of the palytoxin (PTX) effect on the Na(+)/K(+) pump. This toxin acts opening monovalent cationic channels through the Na(+)/K(+) pump. We investigate, by means of computational modeling, the kinetic mechanisms related with K(+) interacting with the complex PTX-Na(+)/K(+) pump. First, a reaction model, with structure similar to Albers-Post model, describing the functional cycle of the pump, was proposed for describing K(+) interference on the complex PTX-Na(+)/K(+) pump in the presence of intracellular ATP. A mathematic model was derived from the reaction model and it was possible to solve numerically the associated differential equations and to simulate experimental maneuvers about the PTX induced currents in the presence of K(+) in the intra- and extracellular space as well as ATP in the intracellular. After the model adjusting to the experimental data, a Monte Carlo method for sensitivity analysis was used to analyze how each reaction parameter acts during each experimental maneuver involving PTX. For ATP and K(+) concentrations conditions, the simulations suggest that the enzyme substate with ATP bound to its high-affinity sites is the main substate for the PTX binding. The activation rate of the induced current is limited by the K(+) deocclusion from the PTX-Na(+)/K(+) pump complex. The K(+) occlusion in the PTX induced channels in the enzymes with ATP bound to its low-affinity sites is the main mechanism responsible for the reduction of the enzyme affinity to PTX.     

Experimental validation of theoretical potassium and sodium cation affinities of amides by mass spectrometric kinetic method measurements

In this study the theoretical Gaussian-2 K(+)/Na(+) binding affinities (enthalpies) at 0 K (in kJ mol(-1)) for six amides in the order: formamide (109.2/138.5) < N-methylformamide (117.7/148.6) < acetamide (118.7/149.5) < N,N-dimethylformamide (123.9/156.4) < N-methylacetamide (125.6/157.7) < N,N-dimethylacetamide (129.2/162.6), reported previously (Siu et al., J. Chem. Phys. 2001; 114: 7045-7051), were validated experimentally by mass spectrometric kinetic method measurements. By monitoring the collision-induced dissociation (CID) of K(+)/Na(+)-bound heterodimers of the amides, the relative affinities were shown to be accurate to within +/-2 kJ mol(-1). With these six theoretical K(+)/Na(+) binding affinities as reference values, the absolute K(+)/Na(+) affinities of imidazole, 1-methylimidazole, pyridazine and 1,2-dimethoxyethane were determined by the extended kinetic method, and found to be consistent (to within +/-9 kJ mol(-1)) with literature experimental values obtained by threshold-CID, equilibrium high-pressure mass spectrometry, and Fourier transform ion cyclotron resonance/ligand-exchange equilibrium methods. A self-consistent resolution is proposed for the inconsistencies in the relative order of K(+)/Na(+) affinities of amides reported in the literature. These two sets of validated K(+) and Na(+) affinity values are useful as reference values in kinetic method measurements of K(+)/Na(+) affinity of model biological ligands, such as the K(+) affinities of aliphatic amino acids.     

Chemical synthesis of the cardiotonic steroid glycosides and related natural products

The active components from the extracts of Digitalis, cardiotonic steroid glycosides, have been ingested by humans for more than 200 years as a medicinal therapy for heart failure and abnormal heart rhythms. The positive inotropic activity of the cardiotonic steroids that mediates clinically useful physiological effects in patients has been attributed largely to a high affinity inhibitory interaction with the extracellular surface of the membrane-bound sodium pump (Na(+)/K(+)-ATPase). However, previously unrecognized intracellular signaling pathways continue to be uncovered. This Review examines both partial and de novo synthetic approaches to the medicinally important and structurally captivating cardenolide and bufadienolide steroid families, with an emphasis on the stereocontrolled construction of the pharmacophoric aglycone (genin) framework.     

Insights into the interactions between a drug and a membrane protein target by fluorine cross-polarization magic angle spinning NMR

The fluorinated anti-psychotic drug trifluoperazine (TFP) has been shown to be a K(+)-competitive inhibitor of gastric H(+)/K(+)-ATPase, a membrane-embedded therapeutic target for peptic ulcer disease. This paper describes how variable contact time (19)F cross-polarization magic angle spinning (VCT-CP/MAS) NMR has been used to probe the inhibitory interactions between TFP and H(+)/K(+)-ATPase in native gastric membranes. The (19)F CP/MAS spectra for TFP in H(+)/K(+)-ATPase enriched (GI) gastric membranes and in control membranes containing less than 5 nmol of the protein indicated that the drug associates with the membranes independently of the presence of H(+)/K(+)-ATPase. The (19)F peak intensities in the VCT-CP/MAS experiment confirmed that TFP undergoes slow dissociation (k(off) < 100 s(-1)) from binding sites in GI membranes, and more rapid dissociation (k(off) < 100 s(-1)) from control membranes. The spectra showed that up to 40% of bound TFP was displaced from GI membranes by 100 mM K(+) and by the K(+)-competitive inhibitor TMPIP, but TFP was not displaced from the control membranes. Hence the spectra of TFP in GI membranes represent the drug bound to the K(+)-competitive inhibitory site of H(+)/K(+)-ATPase and to other non-specific sites. The affinity of TFP for the K(+)-competitive site (K(D) = 4 mM) was determined from a binding curve of (19)F peak intensity versus TFP concentration after correction for non-specific binding. The K(D) was much higher than the IC(50) for ATPase inhibition (8 microM), which suggests that the substantial non-specific binding of TFP to the membranes contributes to ATPase inhibition. This novel approach to probing ligand binding can be applied to a wide range of membrane-embedded pharmaceutical targets, such as G-protein coupled receptors and ion channels, regardless of the size of the protein or strength of binding.     

Ultrasound effects on the antioxidative defense systems of Porphyridium cruentum

Ultrasound is a special physical stimulus that has a variety of biological effects. This study provides a first systemic investigation on the ultrasound-induced oxidation and protection actions of the antioxidant defense system in Porphyridium cruentum. The hydroxyl radical and superoxide anion radical scavenging ability of the cells and the electrolyte leakage of the cell membrane were examined. The change of glutathione and carotenoids produced with/without ultrasonic processing were measured; the enzyme activities of superoxide dismutase, catalase, and membrane bound ATPases (Na(+)/K(+)-ATPase, Ca(2+)/Mg(2+)-ATPase) were evaluated for either ultrasound-treated or untreated P. cruentum. The hydroxyl radical and superoxide anion radical scavenging ability of ultrasound-treated P. cruentum increase 49.8 and 76.0%, respectively, of which the electrolyte leakage and malonyldialdehyde accumulation are also found increased 48.6 and 48.0%, respectively, indicating a state of oxidative stress. A significant enhancement of the activities of superoxide dismutase by 53.5%, catalase, membrane bound ATPases (Na(+)/K(+)-ATPase, Ca(2+)/Mg(2+)-ATPase increased by 67.7 and 69.3%, respectively), and the increment of glutathione and carotenoids production are also observed. These results suggested that oxidative stress manifested by elevated reactive oxygen species levels and malonyldialdehyde contents might be resulted from the biophysical responses of P. cruentum to the physical stimuli, and most likely the enhanced antioxidation ability of the algal cells stimuli by ultrasonic comes from the enhancement of enzymatic and nonenzymatic preventive substances as observed in this work.     

Mitochondrial localization and photodamage during photodynamic therapy with tetraphenylporphines

The subcellular localization sites of TPPS4 and TPPS1 and the subsequent cellular site damage during photodynamic therapy were investigated in CT-26 colon carcinoma cells using spectroscopic and electron microscopy techniques. The association of both porphyrins with the mitochondria was investigated and the implications of this association on cellular functions were determined. Spectrofluorescence measurements showed that TPPS4 favors an aqueous environment, while TPPS1 interacts with lipophilic complexes. The subcellular localization sites of each sensitizer were determined using spectral imaging. Mitochondrial-CFP transfected cells treated with porphyrins revealed localization of TPPS1 in the peri-nuclear region, while TPPS4 localized in the mitochondria, inducing structural damage and swelling upon irradiation, as shown by transmission electron microscopy. TPPS4 fluorescence was detected in isolated mitochondria following irradiation. The photodamage induced a 38% reduction in mitochondrial activity, a 30% decrease in cellular ATP and a reduction in Na(+)/K(+)-ATPase activity. As a result, cytosolic concentrations of Na(+) and Ca(2+) increased, and the level of K(+) decreased. In contrast, the lipophilic TPPS1 did not affect mitochondrial structure or function and ATP content remained unchanged. We conclude that TPPS4 induces mitochondrial structural and functional photodamage resulting in an altered cytoplasmic ion concentration, while TPPS1 has no effect on the mitochondria.     

Chlorpromazine binding to Na+, K+-ATPase and photolabeling: involvement of the ouabain site monitored by fluorescence

This work reports the results of ultraviolet irradiation on the interaction of the phototoxic antipsychotic drug chlorpromazine (CPZ) with the sodium pump Na+, K+-ATPase. The study was performed by monitoring the fluorescence modifications of CPZ itself and of the specific probe anthroylouabain (AO). CPZ association with Na+, K+-ATPase was found to modify the kinetics of CPZ-photodegradation. It was demonstrated that UV irradiation produces a stable fluorescent photoproduct of CPZ covalently bound to Na+, K+-ATPase. The fluorescent probe AO, which specifically binds to the extracellular ouabain site of the pump, was used to localize the CPZ binding site. UV-irradiation of AO-labeled Na+, K+-ATPase treated with CPZ at concentration about 20 microM produced dose-dependent modifications of the AO fluorescence, e.g. increased quantum yield and blue shift. The results demonstrated that CPZ binds near the ouabain site. The photo-induced reaction of CPZ with AO-labeled Na+, K+-ATPase protected the ouabain site from the aqueous environment. It was also found that UV irradiation of CPZ-treated enzyme obstructs the binding of AO, which suggested occlusion of the ouabain site. This effect can be evaluated for a potential use of CPZ in photochemotherapy.     

Synthesis and characterization of some chromogenic crown ethers as potential optical sensors for potassium ions

The complexing abilities of a series of chromogenic crown ethers for potassium and sodium ions have been investigated, by spectrophotometry for the reactions in solution, and by diffuse reflectance spectroscopy when the crown ethers were immobilized. The binding coefficients of the reagents increased with increasing negative charge in the cation-binding site and with increasing extent of chelation. Centimolar K(+) concentrations could be determined with the immobilized reagents, with a K(+)/Na(+) selectivity ratio of approximately 10.     

Competition between pi and non-pi cation-binding sites in aromatic amino acids: a theoretical study of alkali metal cation (Li+, Na+, K+)-phenylalanine complexes

To understand the cation-pi interaction in aromatic amino acids and peptides, the binding of M(+) (where M(+) = Li(+), Na(+), and K(+)) to phenylalanine (Phe) is studied at the best level of density functional theory reported so far. The different modes of M(+) binding show the same order of binding affinity (Li(+)>Na(+)>K(+)), in the approximate ratio of 2.2:1.5:1.0. The most stable binding mode is one in which the M(+) is stabilized by a tridentate interaction between the cation and the carbonyl oxygen (O[double bond]C), amino nitrogen (--NH(2)), and aromatic pi ring; the absolute Li(+), Na(+), and K(+) affinities are estimated theoretically to be 275, 201, and 141 kJ mol(-1), respectively. Factors affecting the relative stabilities of various M(+)-Phe binding modes and conformers have been identified, with ion-dipole interaction playing an important role. We found that the trend of pi and non-pi cation bonding distances (Na(+)-pi>Na(+)-N>Na(+)-O and K(+)-pi>K(+)-N>K(+)-O) in our theoretical Na(+)/K(+)-Phe structures are in agreement with the reported X-ray crystal structures of model synthetic receptors (sodium and potassium bound lariat ether complexes), even though the average alkali metal cation-pi distance found in the crystal structures is longer. This difference between the solid and the gas-phase structures can be reconciled by taking the higher coordination number of the cations in the lariat ether complexes into account.     

Europium tetracycline as a luminescent probe for nucleoside phosphates and its application to the determination of kinase activity

The determination of enzyme activities and the screening of enzyme regulators is a major task in clinical chemistry and drug development. A broad variety of enzymatic reactions is associated with the consumption of adenosine triphosphate (ATP), including, in particular, phosphorylation reactions catalyzed by kinases, formation of adenosine cyclic monophosphate (cAMP) by adenylate cyclases, and ATP decomposition by ATPase. We have studied the effect of a series of adenosine (ATP, ADP, AMP, cAMP) and guanosine (GTP, GDP) phosphoric esters, and of pyrophosphate (PP) on the fluorescence emission of the europium tetracycline (EuTC) complex. We found that these compounds have strongly different quenching effects on the luminescence emission of EuTC. The triphosphates ATP and GTP behave as strong quenchers in reducing the fluorescence intensity of EuTC to 25 % of its initial value by formation of a ternary 1:1:1 complex. All other phosphate esters showed a weak quenching effect only. The applicability of this fluorescent probe to the determination of the activity of phosphorylation enzymes is demonstrated by means of creatine kinase as a model for non-membrane-bound kinases. In contrast to other methods, this approach does not require the use of radioactively labeled ATP substrates, additional enzymes, or of rather complex immunoassays.     

Studies on cardiac ingredients of plants. IX. Chemical transformation of proscillaridin by utilizing its 1,4-cycloadducts as key compounds and biological activities of their derivatives.

Three aromatic compounds (2-4) possessing a carbomethoxyl group or a dimethoxyphthaloyl group, prepared by the Diels-Alder reaction of the cardiac glycoside, proscillaridin (1), with dimethyl acetylenedicarboxylate and methyl propiolate, were transformed into alcohols, carboxylic acids and amides. The biological activities of the resulting derivatives were evaluated by the use of Na+, K(+)-adenosine triphosphatase (Na+,K(+)-ATPase) from dog kidney and isolated guinea-pig papillary muscle. Although the biological activities of the resulting derivatives were less potent than that of 1, a para-substituted benzylalcohol (5), methylbenzamides (9a and 10a), and ethylbenzamides (9b and 10b) inhibited the activity of Na+,K(+)-ATPase almost as potently as naturally occurring cardiac glycosides such as digoxin and digitoxin.     

Protective effect of Lagenaria siceraria (Mol) against membrane-bound enzyme alterations in isoproterenol-induced cardiac damage in rats

This study was aimed at evaluating the preventive role of the ethanolic extract of Lagenaria siceraria (Mol) fruit on membrane-bound enzymes, such as sodium potassium-dependent adenosine triphosphatase (Na(+)/K(+) ATPase), calcium-dependent adenosine triphosphatase (Ca(2+) ATPase) and magnesium-dependent adenosine triphosphatase (Mg(2+) ATPase) on isoproterenol (ISO)-induced myocardial infarction (MI) in rats. Male albino Wistar rats were pretreated with the ethanolic extract of L. siceraria (Mol) fruit (125, 250 and 500?mg?kg(-1) body weight) for a period of 30 days. After the treatment period, ISO (85mg?kg(-1) body weight) was subcutaneously injected into rats at 24-h intervals for 2 days. ISO-induced rats showed a significant (p?相似文献   

Cation-pi interactions and the gas-phase thermochemistry of the Na(+)/phenylalanine complex

The complex of Na(+) with phenylalanine (Phe) is a prototype for the participation of cation-pi interactions in metal-ion binding to biological molecules. A recent comparison of this complex with the Na(+)/alanine (Na(+)/Ala) counterpart suggested only a small contribution of the phenyl ring interaction to binding, casting doubt on the extent of the cation-pi effect. The present work reexamines this thermochemistry using ligand-exchange equilibrium measurements in the Fourier transform ion cyclotron resonance (FT-ICR) ion trapping mass spectrometer. An increment of 7 +/- 2 kcal mol(-1) was found in the Ala/Phe comparison of binding enthalpies, confirming the importance of cation-pi binding enhancement in the Phe case. Absolute Na(+) binding enthalpies of 38 +/- 2 and 45 +/- 2 kcal mol(-1) were assigned for Ala and Phe, respectively, using pyridine as the thermochemical reference ligand. All of these results were supported by quantum calculations using both density functional and Hartree-Fock/MP2 methods, improved in several respects over previous calculations. Alanine methyl ester (AlaMe) was also observed, and found to have an Na(+) ion affinity larger by 2.3 kcal mol(-1) than Ala. New, lower energy conformations of neutral Phe were discovered in the computations.     

Inhibition of gastric H+, K(+)-ATPase activity by compounds from medicinal plants

H+, K(+)-ATPase enzyme is a therapeutic target for the treatment of gastric disturbances. Several medicinal plants and isolated compounds inhibit the acid gastric secretion through interaction with the proton pump. In order to add new properties to some natural constituents, five compounds, a benzylated derivative of vincoside, a diterpene (abietic acid) and three alkaloids (cephaeline, vinblastine and vindoline), were tested for their activities on gastric H+, K(+)-ATPase isolated from rabbit stomach. All the compounds inhibited H+, K(+)-ATPase activity with varied potency. The IC50 value for benzylvincoside was 121 (50-293) microM, and for abietic acid 177 (148-211) microM. The alkaloids cephaeline, vinblastine and vindoline inhibited the H+, K(+)-ATPase activity with IC50 values of 194, 761 and 846 microM, respectively. The results suggest that benzylvincoside, abietic acid and cephaeline can be important sources for the development of anti-secretor agents.     

Polyionic charge density plays a key role in differential recognition of mobile ions by biopolymers

We address the question of what are the molecular mechanisms providing discrimination between seemingly similar counterions binding to various biomolecular surfaces. In the case of protein association with Na (+) and K (+) ions, recent works proposed that specificity of carboxylate functional groups interacting with these mobile ions rationalizes the observed ionic discrimination. We probe in this work whether similar arguments may be used to explain higher propensity of Na (+) ions to associate with DNA compared with K (+) ions, which was suggested by our simulations and some experiments. By comparing our extensive molecular dynamics simulations of Na (+) and K (+) distributions around a 16-base-pair DNA oligomer, [(CGAGGTTTAAACCTCG)] 2, with additional simulations where DNA is replaced by a "soup" of monomers (dimethylphosphate anion), we conclude that DNA specificity toward Na (+)/K (+) is not determined by the underlying functional group specificity. Instead, the collective effect of DNA charges drives larger Na (+) association. To gain additional microscopic insights into the mechanisms of specificity on ionic associations in these systems, we carried out energetic analysis of the association between Na (+) and K (+) with chloride and dimethylphosphate anions. The insights gained from our computational work shed light on a number of experiments on electrolyte solutions of monovalent salts and DNA.     

Dissection of the difference between the group I metal ions in inhibiting GSK3β: a computational study

Glycogen synthase kinase 3β (GSK3β) is a serine/threonine kinase that requires two cofactor Mg(2+) ions for catalysis in regulating many important cellular signals. Experimentally, Li(+) is a competitive inhibitor of GSK3β relative to Mg(2+), while this mechanism is not experienced with other group I metal ions. Herein, we use native Mg(2)(2+)-Mg(1)(2+) GSK3β and its Mg(2)(2+)-M(1)(+) (M = Li, Na, K, and Rb) derivatives to investigate the effect of metal ion substitution on the mechanism of inhibition through two-layer ONIOM-based quantum mechanics/molecular mechanics (QM/MM) calculations and molecular dynamics (MD) simulations. The results of ONIOM calculations elucidate that the interaction of Na(+), K(+), and Rb(+) with ATP is weaker compared to that of Mg(2+) and Li(+) with ATP, and the critical triphosphate moiety of ATP undergoes a large conformational change in the Na(+), K(+), and Rb(+) substituted systems. As a result, the three metal ions (Na(+), K(+), and Rb(+)) are not stable and depart from the active site, while Mg(2+) and Li(+) can stabilize in the active site, evident in MD simulations. Comparisons of Mg(2)(2+)-Mg(1)(2+) and Mg(2)(2+)-Li(1)(+) systems reveal that the inline phosphor-transfer of ATP and the two conserved hydrogen bonds between Lys85 and ATP, together with the electrostatic potential at the Li(1)(+) site, are disrupted in the Mg(2)(2+)-Li(1)(+) system. These computational results highlight the possible mechanism why Li(+) inhibits GSK3β.     

Dissociative ionization of Na2 via repulsive Rydberg states: elucidating femtosecond dynamics with nanosecond lasers

We have studied the dissociative ionization behavior of Na2 molecules using two-color, three photon optical-optical double resonance enhanced excitation via the A(1)Sigma(u)(+) and the 2(1)Pi(g) states. Excess energy ranges from about 150 to about 1500 cm(-1) above threshold for dissociative ionization into ground-state Na and Na(+). Slow atomic Na(+) fragments and Na2(+) molecular ions are detected using a linear time-of-flight spectrometer operated in low field extraction, core sampling mode. To explain the observed energy dependence of the Na(+)/Na2(+) branching ratio, we introduce a semiclassical model for the underlying decay dynamics. Franck-Condon overlap densities for bound-free transitions starting in 2(1)Pi(g) vibrational levels indicate that atomic Na(+) fragments are primarily produced via Rydberg states, with principal quantum number n between 5 and 12, converging to the repulsive 1(2)Sigma(u)(+) first excited-state potential of Na2(+). Dynamics along these Rydberg curves involves competition between electronic (autoionizing) and nuclear (dissociative) degrees of freedom. Within the model, the autoionization lifetime tau auto is the only one free parameter available to fit calculated Na(+)/Na2(+) branching ratios as a function of excess energy to the observed values. The lifetime is assumed to be the same multiple c of the Bohr period of each Rydberg potential. A chi(2)-minimization procedure yields, for the range of principal quantum numbers involved, a most likely value of c = 1.5 +/- 0.3, implying that on average the Rydberg electron completes only 1 to 2 orbits before interaction with the excited core electron leads to autoionization.     

Correlation of Solution and Gas Phase Complexation Assessed by Electrospray Ionization Mass Spectrometry: Application to One-, Two-, and Three-Ring Macrocycles

Electrospray ionization mass spectrometry (ESI-MS) was used to probe multiple cation complexation by C(12)H(25)(CH(2))(12)(CH(2))(12)C(12)H(25), 2, and <18N>CH(2)C(6)H(4)CH(2), 3. Complexation of two cations (2Na(+), 2 K(+), or Na(+) and K(+)) by 3 and three cations by 2 (3 Na(+), 3 K(+), and mixtures) as well as mixed proton-metallic cation complexes of both were observed. The K(+)/Na(+) cation-binding selectivity of 18-crown-6 was studied by ESI-MS of a methanol solution, and the selectivity profile was favorably compared with data obtained previously by ion-selective electrode techniques in the same solvent.     

Studies on cardiac ingredients of plants. VII: Chemical transformation of proscillaridin by means of the Diels-Alder reaction and biological activities of its derivatives.

The Diels-Alder reactions of a cardiac glycoside, proscillaridin (1), with some dienophiles were investigated. The reaction of 1 with alkenes such as methyl vinyl ketone and methyl acrylate afforded 3-oxo-2-oxabicyclo[2.2.2]oct-7-enes (2-5) and para-substituted benzene derivatives (6 and 7), while 1 reacted with alkynes (3-butyn-2-one, methyl propiolate) to yield para- or meta-substituted benzene derivatives (6-9). The biological activities of the resulting derivatives were evaluated by the use of isolated guinea-pig papillary muscle preparations and Na+,K(+)-adenosine triphosphatase (ATPase) preparation from dog kidney. Among the proscillaridin derivatives, compounds 4 and 7 moderately inhibited Na+,K(+)-ATPase activity. Furthermore, the concentration range of 7 over which its positive inotropic effect on guinea-pig papillary muscle preparations, increased from 5% to 95% of maximum was broader than that of 1, i.e., concentration dependency was maintained over a greater range of concentration.     

Influence of kappa-carrageenan gel structures on the diffusion of probe molecules determined by transmission electron microscopy and NMR diffusometry

The influence of the microstructures of different kappa-carrageenan gels on the self-diffusion behavior of poly(ethylene glycol) (PEG) has been determined by nuclear magnetic resonance (NMR) diffusometry and transmission electron microscopy (TEM). It was found that the diffusion behavior was determined mainly by the void size, which in turn was defined by the state of aggregation of the kappa-carrageenan. The kappa-carrageenan concentration was held constant at 1 w/w%, and the aggregation was controlled by the amount of potassium and/or sodium chloride and, for samples containing potassium, also by the cooling rate. Gels containing potassium formed microstructures where kappa-carrageenan strands are rather evenly distributed over the image size, while sodium gels formed dense biopolymer clusters interspersed with large openings. In a gel with small void sizes, relatively slow diffusion was found for all PEG sizes investigated. Extended studies of the self-diffusion behavior of the 634 g mol(-)(1) PEG showed that there is a strong time dependence in the measured PEG diffusion. An asymptotic lower time limit of the diffusion coefficient was found in all gels when the diffusion observation time was increased. According to the ratio, D/D(0), where D(0) is the diffusion coefficient in D(2)O and D is the diffusion coefficient in the gels, the gels could be divided into three classes: small, medium, and large voids. For quenched kappa-carrageenan solutions with salt concentrations of 20 mM K(+), 100 mM K(+), or 20 mM K(+)/200 mM Na(+) as well as slowly cooled solutions with only 20 mM K(+), D/D(0) ratios between 0.18 and 0.29 were obtained. By quenching a kappa-carrageenan solution with 100 mM K(+), the D/D(0) was 0.5, while D/D(0) ratios between 0.9 and 1 were obtained in a quenched solution with 250 mM Na(+) and slowly cooled samples with 20 mM K(+)/200 mM Na(+) or 250 mM Na(+).