Characterisation and selectivity of divalent metal ions binding by citrus and sugar-beet pectins

A scale of selectivity for the binding of calcium and some heavy metal ions by citrus and sugar-beet pectins was set up by pH-measurements. The same order of selectivity was found for the two pectins, decreasing as follows: Cu²⁺ Pb²⁺ Zn²⁺ > Cd²⁺ Ni²⁺ ≥ Ca²⁺. Binding isotherms for Ca²⁺, Cu²⁺, Ni²⁺, Pb²⁺ and Zn²⁺ ions have shown a greater binding level when the ionic strength decreased and when the pectin concentration increased in the presence of 0.1 M NaNO₃. By comparing binding isotherms, the same order of selectivity was found as by pH-studies. Scatchard plots and Hill index evaluation showed for all ions and all pectins anticooperative interactions in water. In the presence of 0.1 M NaNO₃, citrus pectins displayed cooperative interactions for all metal ions. In contrast, for sugar-beet pectins, cooperative interactions only occured with Cu²⁺ and Pb²⁺. With Ca²⁺, Ni²⁺ and Zn²⁺ sugar-beet pectins displayed Scatchard plots which could not be distinguished from an anticooperative binding. This difference of behaviour could be related to the presence of acetyl groups decreasing the affinity of Me²⁺ for sugar-beet pectins.     

Influence of co-cations on biosorption of lead and zinc – a comparative evaluation in binary and multimetal systems

The influence of co-cations (cadmium, copper, cobalt and nickel) on lead and zinc biosorption by Streptoverticillium cinnamoneum and Penicillium chrysogenum in binary and multimetal systems was evaluated. The metal sorption capacity of S. cinnamoneum was higher than P. chrysogenum for all the metals tested. Both the biomasses exhibited preferential uptake of lead in a multimetal situation. Even though mutual inhibition was seen for all binary systems containing zinc, systems containing lead exhibited unequal inhibition. The extent of metal sorption was dependent on metal chemistry, affinity for binding sites and the type of metal binding. In multimetal systems, S. cinnamoneum and P. chrysogenum exhibited preferential sorption orders: Pb²⁺ > Zn²⁺=Cu²⁺ > Cd²⁺ > Ni²⁺ > Co²⁺ and Pb²⁺ > Cu²⁺ > Zn²⁺ > Cd²⁺ > Ni²⁺ > Co²⁺. The order of metal biosorption in a multimetal system could be predicted well on the basis of Langmuir parameters evaluated in binary metal systems.     

Studies on the exchange of G-actin-bound calcium with bivalent cations

1. The possibility of the replacement of G-actin-bound calcium by various bivalent cations has been investigated. After the reaction with all cations studied, with the exception of Cu²⁺, action remains active, i.e., contains bound ATP and polymerizes in 0.1 M KCl.

2. The amount of G-actin-bound calcium, as well as the sum of bivalent cation after replacement, not removable by short-time Dowex-50 treatment, accounts to about 1 mole per 50000 g of G-actin.

3. The rate of exchange is of the same order for bivalent cations studied, including calcium.

4. G-actin-bound Ca²⁺ is fully replaced, besides free Ca²⁺, by free Mn²⁺ and Cd²⁺. The replacement with Mg²⁺, Co²⁺, Ni²⁺ and Zn²⁺ is not complete, and there is practically no reaction with Ba²⁺ and Sr²⁺.

5. Assuming the affinity constant of Ca²⁺ as 1, the following affinity constants for other bivalent cations were obtained: Mn²⁺, 0.90; Cd²⁺, 1.07; Mg²⁺, 0.27; Zn²⁺, 0.22; Co²⁺, 0.18; Ni²⁺, 0.08.

6. The results obtained show that there exists a close correlation between the ionic radius of a particular bivalent cation, and its ability to replace bound Ca²⁺.     

Metal ion-coordinating properties of imidazole and derivatives in aqueous solution: interrelation between complex stability and ligand basicity

The stability constants of the 1:1 complexes formed between Mg²⁺, Ca²⁺, Sr²⁺, Ba²⁺, Mn²⁺, Co²⁺, Ni²⁺, Cu²⁺, Zn²⁺ or Cd²⁺ (M²⁺) and the simple, sterically unhindered imidazole-type ligands, imidazole, 1-methylimidazole, 5-chloro-1-methylimidazole, N-(2,3,5,6-tetrafluorophenyl)imidazole or 4′-(imidazol-1-yl)acetophenone (L), were determined by potentiometric pH titrations in aqueous solution (25°C; I = 0.5 M, NaNO₃). The construction of log K_ML^M versus pK_HL^H plots results in straight lines; the equations for the least-squares lines are calculated and listed. These data allow calculation of the expected stability constant for a complex of any imidazole-type ligand, provided its pK_HL^H value (in the pK_a range 4–8) is known. For the stabilities of Fe²⁺ complexes with imidazole-type ligands an estimation procedure is provided. It is shown further that the complex formation between 1-methylbenzimidazole (MBI) and Mn²⁺, Ni²⁺, Cu²⁺ or Zn²⁺ is s sterically hindered, i.e. the data points for these M(MBI)²⁺ complexes do not fall on the straight lines defined by the imidazole-type ligands.     

Inactivation of chloroplast photosynthetic electron-transport activity by Ni

Ni²⁺ inhibits electron-transport activity of isolated barley chloroplasts and this inhibition of electron transport by Ni²⁺ is distinctly different from other heavy metal ion (e.g., Pb²⁺, Cd²⁺, Zn²⁺)-induced inhibition of chloroplast function. Ni²⁺ inactivates Photosystem II (PS II) activity at a lower concentration than that required for the same extent of inhibition of Photosystem I (PS I)-mediated electron flow. Ni²⁺ induces changes in chlorophyll a (Chl a) emission characteristics and brings about a lowering of the Chl a fluorescence yield, and this lowering of Chl a fluorescence intensity is not relieved by the exogenously supplied electron donor NH₂OH which donates electrons very close to the PS II reaction centres. Immobilization of the chloroplast membrane structure with glutaraldehyde fails to arrest the Ni²⁺-induced loss of PS II activity. Also, Ni²⁺-treated chloroplasts do not regain the ability to photoreduce 2,6-dichlorophenolindophenol even after washing of chloroplasts with buffer. These results indicate that unlike Zn²⁺ or Pb²⁺, Ni²⁺ induces alterations in the chloroplast photosynthetic apparatus resulting in an irreversible loss of electron-transport activity.     

Purification and chemical characterization of thermostable amylases produced by Bacillus stearothermophilus

The amylases produced by a Bacillus stearothermophilus were purified through a series of four steps. Two separable enzyme fractions having starch hydrolysing activity were eluted from a DEAE-cellulose column by NaCl gradient elution. The homogeneity of the purified enzymes was checked on polyacrylamide gel electrophoresis. The product formation studies indicated that fraction I was an -amylase whereas fraction II was a β-amylase. The molecular weights were determined to be 48 000 and 57 000 and the carbohydrate moiety was found to be 13.2 and 0.8% for - and β-amylase, respectively. The protein digest of these enzymes indicated a total number of 15 amino acids with aspartic and glutamic acid showing the highest value. The purified amylase showed maximal activity at 80°C and pH 6.9. Fe³⁺, Cd²⁺, Pb²⁺, Hg²⁺, Ni²⁺ and Ag¹⁺ were potent inhibitors whereas Zn²⁺, Mg²⁺, Mn²⁺ and Al³⁺ were mild inhibitors. Ca²⁺, Ba²⁺, Sr²⁺ and K⁺ stimulated amylase activity in the order of Ca²⁺ > Ba²⁺ > Sr²⁺ > K⁺. PCMB, EDTA and sodium iodoacetate were inhibitory whereas glutathione (GSH) and cysteine afforded protection of enzyme activity. EDTA showed dose-dependent noncompetitive inhibition of both - as well as β-amylase activities. EDTA inhibition was reversed by the addition of Ca²⁺ and PCMB inhibition by the addition of glutathione (reduced). The K_m for - and β-amylases were found to be 1.05 and 1.25 mg starch per ml, respectively.     

Stability constants of metal complexes of phosphonoacetic acid

The antiviral drug, phosphonoacetic acid (PAA), forms stable complexes with Mg²⁺, Ca²⁺, Cu²⁺ and Zn²⁺. Stability constants of these complexes were determined in aqueous solution (0.15 M in KNO₃, 37°) by potentiometric titration. Mixed ligand complex formation of Cu²⁺ and Zn²⁺ with PAA and glycinate ion, and with PAA and histidinate ion, was studied. In a theoretical model for blood plasma, PAA affects the distribution of Mg²⁺ and, to a lesser extent, Ca²⁺.     

Biosorption of lead, cadmium and mercury by immobilized Microcystis aeruginosa in a column

Immobilized Microcystis aeruginosa in a flow-through sorption column was evaluated for the potential to remove Pb²⁺, Cd²⁺ and Hg²⁺ from aqueous solutions. M. aeruginosa showed high affinity for the three heavy metals with removal efficiency of 90% for Cd²⁺ and Hg²⁺, and 80% for Pb²⁺ at saturation conditions. Competitive sorption experiments conducted in paired and ternary systems indicated that Pb²⁺ was sequestered preferentially over Cd²⁺ and Hg²⁺. The presence of Cd²⁺ interfered only slightly with the uptake of Hg²⁺, as Pb²⁺ and Hg²⁺ did with Cd²⁺. In contrast, Hg²⁺ sorption was affected by Pb²⁺ to a great extent. Desorption with 1 M HCl was completed within 25 min with high efficiency and effectiveness for the three metals. The results of this study indicate that M. aeruginosa is to be a potential biosorbent material except when Hg²⁺ and Pb²⁺ are in the same contaminated solution.     

改性粉煤灰对废水中镉的吸附作用

以工业固废粉煤灰(FA)为原料,将其与NaOH和Ca(OH)₂混合后,在250 ℃下焙烧1.5 h,制得改性粉煤灰(MFA).利用扫描电镜(SEM)、能量色散X射线光谱仪(EDS)、X-射线衍射仪(XRD)、全自动比表面积分析仪(BET)和傅里叶红外光谱仪(FTIR)等技术分析MFA的理化性质.BET分析表明,MFA的比表面积比FA增大了20.6倍.SEM分析表明,FA表面玻璃相溶解,表面粗糙,具有多孔结构.FTIR分析表明,-OH官能团在Cd²⁺的吸附过程中具有重要作用.静态吸附试验研究表明,当MFA的投加量为0.2 g、Cd²⁺初始浓度为100 mg·L^-1、溶液pH为7、25 ℃下吸附90 min时,Cd²⁺的去除率达到97.3%;共存阳离子(K⁺、Na⁺、Mg²⁺和Ca²⁺)均会抑制MFA对Cd²⁺的吸附,其中Ca²⁺的抑制作用最强.Langmuir等温吸附模型和准二级吸附动力学方程能较好地描述Cd²⁺在MFA上的吸附行为,且最大吸附量为55.77 mg·g^-1.热力学研究表明,MFA对Cd²⁺的吸附是一个自发的吸热过程.MFA吸附能力优于FA,在废水处理方面有一定的应用前景.     

A new macrocyclic tetraamine, 2,4-dinitrophenylcyclen (= 1-(2,4-dinitrophenyl)-1,4,7,10-tetraazacyclododecane):synthesis, cation reporter properties, and recognition of 1-methylthymine by its zinc(II) complex

A new functional macrocyclic ligand, 2,4-dinitrophenylcyclen (= 1-(2,4-dinitrophenyl)-1,4,7,10-tetraazacyclododecane), has been synthesized and isolated as its trihydrochloric acid salt (L·3HCl). The protonation constants (log K_n) for three secondary nitrogens of L were determined by potentiometric pH titration to be 10.10, 7.33 and <2 with I = 0.10 (NaNO₃) at 25°C. The 2,4-dinitrophenylaniline chromophore was proven to be a good reporter signaling proton- and metal-binding events in the macrocyclic cavity. The UV absorption band (λ_max 370 nm, 8200) of the 2,4-dinitrophenylaniline moiety at pH ≥ 9 becomes quenched as pH is lowered (to pH 3.1, where the major species is L·2H⁺), due to the strong protonation effect extended to the aniline moiety within the macrocyclic cavity. This is in sharp contrast to the pH-independent UV absorption (λ_max 390 nm, 14 000) of a reference compound, N,N-diethyl-2,4-dinitroaniline. The UV absorption band of L is shifted to lower wavelengths with Zn²⁺ (λ_max 320 nm), Cd²⁺ (λ_max 316 nm) and Pb²⁺ (λ_max 317 nm), while it almost disappears with Cu²⁺ and Ni²⁺. The 1:1 Zn²⁺ and Cu²⁺ complexes with L were isolated and characterized. The Zn²⁺ complex recognizes 1-methylthymine anion (MT⁻) in aqueous solution at physiological pH to yield a stable ternary complex ZnL-MT⁻. The X-ray crystal structure of ZnL-MT⁻ showed that Zn²⁺ is four-coordinate with three secondary nitrogens of L and the deprotonated imide anion that is cofacial to the 2,4-dinitrophenyl ring.     

The use of chemically modified and unmodified cassava waste for the removal of Cd, Cu and Zn ions from aqueous solution

The use of different chemically modified cassava waste biomass for the enhancement of the adsorption of three metal ions Cd, Cu and Zn from aqueous solution is reported in this paper. Treating with different concentrations of thioglycollic acid modified the cassava waste biomass.

The sorption rates of the three metals were 0.2303 min⁻¹ (Cd²⁺), 0.0051 min⁻¹ (Cu²⁺), 0.0040 min⁻¹ (Zn²⁺) and 0.109 min⁻¹ (Cd²⁺), 0.0069 min⁻¹ (Cu²⁺), 0.0367 min⁻¹ (Zn²⁺) for 0.5 and 1.00 M chemically modified levels, respectively. The adsorption rates were quite rapid and within 20–30 min of mixing, about 60–80% of these ions were removed from the solutions by the biomass and that chemically modifying the binding groups in the biomass enhanced its adsorption capacity towards the three metals. The results further showed that increased concentration of modifying reagent led to increased incorporation, or availability of more binding groups, in the biomass matrix, resulting in improved adsorptivity of the cassava waste biomass. The binding capacity study showed that the cassava waste, which is a serious environmental nuisance, due to foul odour released during decomposition, has the ability to adsorb trace metals from solutions.     

不同炭化温度制备的蚕丝被废弃物生物炭对重金属Cd2+的吸附性能

以蚕丝被废弃物为原料,在300、500和700 ℃高温缺氧条件下热解炭化制备成3种生物炭(BC300、BC500和BC700).利用扫描电镜(SEM)、傅里叶红外光谱仪(FT-IR)、X-射线衍射仪(XRD)、比表面积分析仪等对其理化性质进行表征,并研究了不同温度下制备的生物炭对溶液中Cd²⁺的吸附特性.结果表明: 随着炭化温度上升,BET比表面积、pH、灰分均增大,生物炭表面形态结构越来越不规则.XRD结果显示:不同温度下获得的生物炭中均含有一定量的方解石,FT-IR光谱图上的峰主要为-OH和方解石典型的吸收峰;pH对生物炭吸附Cd²⁺的影响不大;Langmuir方程能更好地拟合3种生物炭对Cd²⁺的吸附等温过程,其最大吸附量分别为25.61、52.41和91.07 mg·g^-1.3种生物炭对Cd²⁺吸附过程均更符合准二级动力学方程,且BC700对Cd²⁺的吸附效果最佳.进一步研究离子浓度及阳离子共存对BC700吸附Cd²⁺的影响,结果显示: NaCl浓度越高,对Cd²⁺的吸附抑制越明显;共存阳离子中,Ca²⁺和Mg²⁺对Cd²⁺的吸附抑制更明显,而K⁺几乎无影响.因此,以蚕丝被废弃物制备的生物炭作为去除水体中Cd²⁺的吸附剂具有较强的应用潜力.     

Detection of heavy metal toxicity using cardiac cell-based biosensor

Biosensors incorporating mammalian cells have a distinct advantage of responding in a manner which offers insight into the physiological effect of an analyte. To investigate the potential applications of cell-based biosensors on heavy metal toxicity detection, a novel biosensor for monitoring electrophysiological activity was developed by light-addressable potentiometric sensor (LAPS). Extracellular field potentials of spontaneously beating cardiomyocytes could be recorded by LAPS in the range of 20 μV to nearly 40 μV with frequency of 0.5–3 Hz. After exposed to different heavy metal ions (Hg²⁺, Pb²⁺, Cd²⁺, Fe³⁺, Cu²⁺, Zn²⁺; in concentration of 10 μM), cardiomyocytes demonstrated characteristic changes in terms of beating frequency, amplitude and duration under the different toxic effects of ions in less than 15 min. This study suggests that, with the physiological monitoring, it is possible to use the cardiac cell-based biosensor to study acute and eventually chronic toxicities induced by heavy metal ions in a long-term and no-invasive way.     

Growth rate based dose-response relationships and EC-values of ten heavy metals using the duckweed growth inhibition test (ISO 20079) with Lemna minor L. clone St

The duckweed Lemna minor L. clone St was used to investigate the effect of 10 heavy metals under the standardised test conditions of the ISO protocol 20079. By using growth rates derived from frond number (FN), fresh weight (FW), dry weight (DW), chlorophyll and carotenoid (Car) contents, concentration–response curves for all heavy metals and all growth parameters were classified. In addition, all data were fitted to obtain the inhibitions of growth rates (E_rC_x) at the level of 10%, 20% and 50% (E_rC₁₀, E_rC₂₀ and E_rC₅₀, respectively) then used to evaluate the phytotoxicity of the different heavy metals. On the basis of the E_rC₅₀ values (average ranking of all five growth parameters), the following series of phytotoxicity was detected by using molar concentrations: Ag⁺>Cd²⁺>Hg²⁺>Tl⁺>Cu²⁺>Ni²⁺>Zn²⁺>Co²⁺>Cr(VI)>As(III)>As(V).     

Effects of divalent metal ions on chlorophyll a fluorescence in isolated spinach chloroplasts

The effects of divalent metal ions on the yields of chlorophyll a fluorescence were investigated in isolated spinach chloroplasts at room and liquid nitrogen temperatures. Mg²⁺, Ca²⁺, Sr²⁺, Ba²⁺ and Mn²⁺ increased the yields of fluorescence emission at 684 and 695 nm from pigment system II and decreased that at 735 nm from pigment system I. Al³⁺ showed similar but less significant effects on the fluorescence yields. Zn²⁺ and Cd²⁺ showed no significant effect on the fluorescence yields at concentrations lower than 5 mM.

In accordance with the results of our previous study concerning the effects of Mg²⁺ on the excitation transfer in the chloroplasts, it was concluded that ions of alkaline earths and manganese suppress the excitation transfer from bulk chlorophylla of pigment system II to that of pigment system I.     

Molecular mechanism of action of Pb and Zn on water oxidizing complex of photosystem II

Pb²⁺ and Zn²⁺ inhibition of photosystem II (PSII) activity was reported to be mediated via displacement of native inorganic cofactors (Cl⁻, Ca²⁺ and Mn²⁺) from the oxygen evolving complex, OEC [Rashid and Popovic (1990) FEBS Lett. 271, 181–184; Rashid et al. (1991) Photosynth. Res. 30, 123–130]. Since the binding sites of these cofactors are protected by a shield of three extrinsic polypeptides (17, 23 and 33 kDa), we investigated whether these metal ions affect the extrinsic polypeptide shield of OEC. By immunoblotting with antibodies recognizing the 23 and 33 kDa polypeptides, we showed that both the metal ions significantly dissociated the 23 kDa (+17 kDa) polypeptide, and partially dissociated the 33 kDa. Ca²⁺, one of the important inorganic cofactors of oxygen evolution, strongly prevented the dissociating action of Pb²⁺ but did not prevent the action of Zn²⁺. The probable molecular mechanism of action of Pb²⁺ and Zn²⁺ on PSII OEC is discussed.     

Trypsin immobilization by direct adsorption on metal ion chelated macroporous chitosan-silica gel beads

Silica gel bead coated with macroporous chitosan layer (CTS-SiO₂) was prepared, and the metal immobilized affinity chromatographic (IMAC) adsorbents could be obtained by chelating Cu²⁺, Zn²⁺, Ni²⁺ ions, respectively on CTS-SiO₂, and trypsin could be adsorbed on the IMAC adsorbent through metal–protein interaction forces. Batch adsorption experiments show that adsorption capacity for trypsin on these IMAC adsorbent variated with change of pH. The maximal adsorption reached when the solution was in near neutral pH in all three IMAC adsorbents. Adsorption isothermal curve indicated that maximal adsorption capacity could be found in the Cu²⁺-CTS-SiO₂ with the value of 4980 ± 125 IU g⁻¹ of the adsorbent, while the maximal adsorption capacity for trypsin on Zn²⁺ and Ni²⁺ loaded adsorbent was 3762 ± 68 IU g⁻¹ and 2636 ± 53 IU g⁻¹, respectively. Trypsin immobilized on the IMAC beads could not be desorbed by water, buffer and salt solution if the pH was kept in the range of 5–10, and could be easily desorbed from the IMAC beads by acidic solution and metal chelating species such as EDTA and imidazole. The effect of chelated metal ions species on CTS-SiO₂ beads on the activity and stability of immobilized trypsin was also evaluated and discussed. Trypsin adsorbed on Zn-IMAC beads retained highest amount of activity, about 78% of total activity could be retained. Although the Cu-IMAC showed highest affinity for trypsin, only 25.4% of the calculated activity was found on the beads, while the activity recovery found on Ni-IMAC beads was about 37.1%. A remarkable difference on stability of trypsin immobilized on three kinds of metal ion chelated beads during storage period was also found. Activity of trypsin on Cu-IMAC decreased to 24% of its initial activity after 1-week storage at 4 °C, while about 80% activity was retained on both Ni-IMAC and Zn-IMAC beads. Trypsin immobilized on Zn-CTS-SiO₂ could effectively digest BSA revealed by HPLC peptide mapping.     

Binding of ions to chitosan—selectivity studies

Selectivity coefficients for binding of negative and positive ions to chitosans of different chemical composition have been determined by equilibrium dialysis. Chitosans with different fraction of acetylated units (F_A of 0.01 and 0.49) generally behaved similarly in their selectivity towards both negative and positive ions. No selectivity was found in the binding of chloride and nitrate ions, while chitosan showed a strong selectivity towards molybdate polyoxyanions, with selectivity coefficients around 100. Chitosan showed a strong selectivity towards copper (Cu²⁺) compared to the metal ions zinc (Zn²⁺), cadmium (Cd²⁺) and nickel (Ni²⁺), with selectivity coefficients from 10 to 1000, while little or no selectivity could be detected with the other metal ions. Ionic strength and pH did not influence the selectivity coefficients of the chitosans towards the metal ions.     

Binding of divalent copper and calcium ions to poly I

The effect ot Cu²⁺ and Ca²⁺ ions, on the ultraviolet differential (UVD) spectra of single-stranded poly I was studied and the coordination (Δε_b) and conformation (Δε_c) conponents of the spectra calculated The comparison of Δε_b and the UVD spectrum of protonated IMP leads to the conclusion that N(7) ot inosine-5'-monophosphate (IMP) is a coordinating site tor Ca²⁺ and Cu²⁺ ions on the polymer bases. The binding ot Ca²⁺ and Cu²⁺ ions causes differently directed displacements of the four absorption bands of poly I, which are observed in the wavenumber range (50-34) × 10³ cm⁻¹ The calculation of concentration dependencies tor the association constants (K“) ot Ca²⁺ and Cu²⁺ ions binding to poly I bases shows that the binding is cooperative The K“ values for the poly I + Ca²⁺ complex are two orders of magnitude lower than those for the poly 1 + Cu²⁺ complex At low ion concentrations, binding to the poly I phosphates predominates and increases the degree of the polynucleotide helicity. At higher concentrations the spectra are mainly affected by the ion binding to bases, which results in melting of the helical parts of poly I At Ca²⁺ concentrations exceeding 10⁻³ M light-scattering aggregates are formed. The degree of monomer order in them is close to that observed in multistranded helices of poly I     

Effect of metallic cations and pH on reassembly of glial fibrillary acidic protein

Glial fibrillary acidic protein (GFAP), which was purified from acetone powder of the bovine spinal cord, was reassembled in 0.1 M imidazole HCl buffer containing metallic cations, Ca²⁺, Mg²⁺, Na⁺ or K⁺ at physiological or more acidic pH. An electron microscopy revealed reassembled glial filaments at pH 6.8 without any cations but amorphous aggregates at pH 6.3 which were readily observed as a white precipitate by the naked eye. Under more alkaline pH (pH 7.4) only rod-shaped short filaments were formed. In the presence of mM concentrations of Ca²⁺ or Mg²⁺, thick bundles of glial filaments, detectable by light microscopy, were formed at acidic pH. At pH 7.4 long reassembled filaments could be formed in the buffer containing divalent cations. Na⁺ (0.1 M) made filament-like structures of GFAP but they are rather random compared to the filaments promoted by the divalent cations. K⁺ made only amorphous aggregation of the short filaments. These findings indicate that the reassembly of GFAP at physiological pH requires essentially divalent cations but not ionic strength.