[Leaching of copper ore of the Udokanskoe deposit at low temperatures by an association of acidophilic chemolithotrophic microorganisms].

Pure cultures of indigenous microorganisms Acidithiobacillus ferrooxidans strain TFUd, Leptospirillum ferrooxidans strain LUd, and Sulfobacillus thermotolerans strain SUd have been isolated from the oxidation zone of sulfide copper ore of the Udokanskoe deposit. Regimes of bacterial-chemical leaching of ore have been studied over a temperature range from -10 to +20 degrees C. Effects of pH, temperature, and the presence of microorganisms on the extraction of copper have been shown. Bacterial leaching has been detected only at positive values of temperature, and has been much more active at +20 than at +4 degrees C. The process of leaching was more active when the ore contained more hydrophilic and oxidized minerals. The possibility of copper ore leaching of the Udokanskoe deposit using sulfuric acid with pH 0.4 at negative values of temperature and applying acidophilic chemolithotrophic microorganisms at positive values of temperature and low pH values was shown.

The primary structure and characteristics of the ISAfe600, an insertion sequence from Acidithiobacillus ferrooxidans strains.

A new IS-like element (604 bp) was revealed in the genome of several Acidithiobacillus ferrooxidans strains isolated from diverse biotopes. It includes 26-bp imperfectly matched terminal inverted repeats (TIRs), similar in structure to the TIRs of ISAfel insertion element. The 60-bp DNA fragment adjacent to the right TIR (TIRR) exhibits pronounced homology with the similarly located DNA fragments in ISAfel and IST445, as well as with the internal fragment of ISAfel encoding the transposase gene (nucleotides from 254 to 311 bp). The central section of ISAfe600 is unique and exhibits no homology with any prokaryotic DNA. A duplication of 8 bp of the target DNA was found in the ISAfe600 insertion site. One to four copies of ISAfe600 were revealed by Southern hybridization in the genome of A. ferrooxidans strains studied. The number of ISAfe600 copies varies depending on the growth conditions (energy substrate). Since the open reading frames big enough to encode transposase are not presert in the structure of ISAfe600, it may be a deficient IS element; its translocation is possibly achieved under control of the ISAfel transposase.

[Isolation of an aboriginal bacterial community capable of utilizing cyanide, thiocyanate, and ammonia from metallurgical plant wastewater].

An aboriginal bacterial community capable of degrading cyanide (10 mg/l) and thiocyanate (2 g/l) and eliminating ammonia (120 mg/l) had been isolated from recycled water samples after blast-furnace gas purification of a metallurgical plant wastewater. It was shown that the optimal conditions for this bacterial community were as follows: temperature, 34 degrees C; pH, 8.8-9.0; available organic matter concentration (glucose equivalent), 5 g/l; and dissolved O2 concentration, 8-10 mg/l. This aboriginal community was formed by the bacteria belonging to the genus Pseudomonas.

[Genotypic and phenotypic polymorphism of environmental strains of the moderately thermophilic bacterium Sulfobacillus sibiricus].

Five cultures of moderately thermophilic spore-forming acidophilic chemolithotrophic bacteria were isolated from the zones of spontaneous heating of pyrrhotine-containing pyrite-arsenopyrite gold-arsenic sulfide ores in an operating open pit (strains B1, B2, B3, OFO, and SSO). Analysis of the chromosomal DNA structure revealed differences between these cultures at the strain level (apart from B3 and SSO, which had identical restriction profiles). All the strains had a similar G + C DNA molar content (47.4-48.3%). The level of DNA reassociation was 85 to 95%. The similarity between the DNA of the type strain Sulfobacillus sibiricus N1 isolated from arsenopyrite ore concentrate and that of these strains (83-93%) indicates that they belong to the same species. The strains had similar values of pH and temperature optimal for growth on ferrous iron (1.6-2.0 and 45-55 degrees C, respectively). They were mixotrophs; Fe(II), S0, and sulfide minerals along with organic compounds were used as energy sources and electron donors. However, the kinetic parameters of growth and substrate oxidation varied from strain to strain. Genetic variety of the strains from diverse ecosystems and environments is possibly the result of the different rates of microevolution processes.

[The dependence of intracellular ATP level on the nutrition mode of the acidophilic bacteria Sulfobacillus thermotolerans and Alicyclobacillus tolerans].

The dynamics of the ATP pool in the aerobic spore-forming acidothermophilic mixotrophic bacteria Sulfobacillus thermotolerans Kr1T and Alicyclobacillus tolerans K1T were studied in the course of their chemolithoheterotrophic, chemoorganoheterotrophic, and chemolithoautotrophic growth. It was established that, during mixotrophic growth, the maximum ATP concentrations in the cells of S. thermotolerans Kr1 and A. tolerans K1 were 3.8 and 0.6 nmol/mg protein, respectively. The ATP concentrations in sulfobacilli and alicyclobacilli during organotrophic growth were 2.2 and 3.1 nmol/mg protein, respectively. In the cells of the obligately heterotrophic bacterium Alicyclobacillus cycloheptanicus 4006T, the maximum ATP concentration was several times higher and reached 12.3 nmol/mg protein. During lithotrophic growth, the maximum values of the ATP concentration in the cells of S. thermotolerans Kr1 and A. tolerans K1 were 0.3 and <0.1 nmol/mg protein, respectively; in the cells of the autotrophic bacterium Acidithiobacillus ferrooxidans TFBk, the ATP content was about 60-300 times higher (17.0 nmol/mg protein). It is concluded that low ATP content is among the possible causes of growth cessation of S. thermotolerans Kr1 and A. tolerans K1 under auto- and heterotrophic conditions after several culture transfers.

[Lithotrophic microorganisms of the oxidative cycles of sulfur and iron].

The review deals with sulfur bacteria (the first chemolithotrophs ever studied) and with the acidophilic bacteria of sulfur and iron cycles which were investigated as a result of Winogradsky's discovery. The diversity of these organisms and the factors and mechanism of its origin are emphasized; their metabolic functions and nutritional regulation are discussed.

[Mechanism of cyanide and thiocyanate decomposition by an association of Pseudomonas putida and Pseudomonas stutzeri strains].

The intermediate and terminal products of cyanide and thiocyanate decomposition by individual strains of the genus Pseudomonas, P. putida strain 21 and P. stutzeri strain 18, and by their association were analyzed. The activity of the enzymes of nitrogen and sulfur metabolism in these strains was compared with that of the collection strains P. putida VKM B-2187T and P. stutzeri VKM B-975T. Upon the introduction of CN- and SCN- into cell suspensions of strains 18 and 21 in phosphate buffer (pH 8.8), the production of NH4+ was observed. Due to the high rate of their utilization, NH3, NH4+, and CNO- were absent from the culture liquids of P. putida strain 21 and P. stutzeri strain 18 grown with CN- or SCN-. Both Pseudomonas strains decomposed SCN- via cyanate production. The cyanase activity was 0.75 micromol/(min mg protein) for P. putida strain 21 and 1.26 micromol/(min mg protein) for P. stutzeri strain 18. The cyanase activity was present in the cells grown with SCN- but absent in cells grown with NH4+. Strain 21 of P. putida was a more active CN- decomposer than strain 18 of P. stutzeri. Ammonium and CO2 were the terminal nitrogen and carbon products of CN- and SCN- decomposition. The terminal sulfur products of SCN- decomposition by P. stutzeri strain 18 and P. putida strain 21 were thiosulfate and tetrathionate, respectively. The strains utilized the toxic compounds in the anabolism only, as sources of nitrogen (CN- and SCN-) and sulfur (SCN-). The pathway of thiocyanate decomposition by the association of bacteria of the genus Pseudomonas is proposed based on the results obtained.

[Dependence of the genotypic characteristics of Acidithiobacillus ferrooxidans on the physical, chemical, and electrophysical properties of pyrites].

Comparison of Acidithiobacillus ferrooxidans strains TFV-1 and TFBk with respect to their capacity to oxidize pyrite 1, with hole-type (p-type) conductivity, or pyrite 2, with an electron-type (n-type) conductivity, showed that, at a pulp density of 1%, both before and after its adaptation to the pyrites, strain TFBk, isolated from a substrate with a more complex mineral composition, grew faster and oxidized the pyrites of both conductivity types more efficiently than strain TFV-1, which was isolated from a mineralogically simple ore. At a pulp density of 3-5%, the oxidation of pyrite 1 by strain TFV-1 and both of the pyrites by strain TFBk began only after an artificial increase in Eh to 600 mV. If the pulp density was increased gradually, strain TFBk could oxidize the pyrites at its higher values than strain TFV-1, with the rate of pyrite 2 oxidation being higher than that of pyrite 1. During chemical oxidation of both of the pyrites, an increase was observed in the absolute values of the coefficients of thermoelectromotive force (KTEMF); during bacterial-chemical oxidation, the KTEMF of pyrite 1 changed insignificantly, whereas the KTEMF of pyrite 2 decreased.

[Dependence of the genotypic characteristics of Acidithiobacillus ferrooxidans on the physical, chemical, and electrophysical properties of pyrites].

This study focused on the effect of physical, chemical, and electrophysical properties of two pyrites, pyrite 1, which had hole-type (p-type) conductivity, and pyrite 2, with electron-type (n-type) conductivity, on the genotypic characteristics of Acidithiobacillus ferrooxidans strains TFV-1 and TFBk, which were isolated from different substrates. After the adaptation of the strains to the pyrites at a pulp density of 1%, pulsed-field electrophoresis revealed changes in the chromosomal DNA of strain TFV-1 adapted to pyrite 1 and strain TFBk adapted to either of the pyrite types. In pyrite-adapted strain TFBk, the plasmid composition was the same as after growth on a medium containing ferrous iron, whereas, in strain TFV-1, changes in plasmid sizes or both in plasmid sizes and plasmid number occurred. After an increase in the density of the pyrite 2 pulp from 1 to 10%, the plasmid number increased from three to four, and, after an increase in the density of the pyrite 1 pulp from 1 to 7%, the plasmid number increased from two to six.

[Interaction of chromosomal and plasmid DNA in Acidithiobacillus ferrooxidans strains adapted to different oxidation substrates]. / Vzaimodeistvie khromosomnoi i plazmidnoi DNK u shtammov Acidithiobacillus ferrooxidans, adaptirovannykh k raznym substratam okisleniia.

Restriction analysis of plasmids pTFK1 and pTFK2 of the Acidithiobacillus ferrooxidans strain TFBk was carried out, and the sizes of these plasmids were determined (13.5 and 30 kb, respectively). A macrorestriction map was built for plasmid pTFK1. DNA-DNA hybridization revealed that the plasmids contained homologous nucleotide sequences. Plasmid pTFK2 labeled with 32P was used as a probe for Southern hybridization with blots of XbaI-generated fragments of the chromosomal DNA of A. ferrooxidans strains grown on a medium containing Fe2+ or adapted to different oxidation substrates. Low-intensity hybridization signals were observed for many fragments of the chromosomal DNA of the strains studied. In the process of adaptation to new oxidation substrates, the localization of bands producing the low-intensity hybridization signals changed in a number of cases. Certain fragments of the chromosomal DNA of the strains adapted to different oxidation substrates produced strong hybridization signals with pTFK2. The data obtained are discussed in terms of the possible role of IST elements and plasmids in the adaptation of A. ferrooxidans to new energy substrates, microevolution, and strain polymorphism.

[Activity of the enzymes of carbon metabolism in Sulfobacillus sibiricus under various conditions of cultivation]. / Aktivnost' fermentov uglerodnogo obmena u Sulfobacillus sibiricus v razlichnykh usloviiakh kul'tivirovaniia.

The thermoacidophilic iron-oxidizing chemolithotroph Sulfobacillus sibiricus N1T is characterized by steady growth and amplified cell yield when grown in vigorously aerated medium containing Fe2+, glucose, and yeast extract as energy sources. In this case, carbon dioxide, glucose, and yeast extract are used as carbon sources. Glucose is assimilated through the fructose-bisphosphate pathway and the pentose-phosphate pathway. Glyoxylate bypass does not function in S. sibiricus, and the tricarboxylic acid cycle is disrupted at the level of 2-oxoglutarate dehydrogenase. The presence of ribulose-bisphosphate carboxylase indicates that carbon dioxide fixation proceeds through the Calvin cycle. The activity of ribulose-bisphosphate carboxylase is highest in autotrophically grown cells. The cells also contain pyruvate carboxylase, phosphoenolpyruvate carboxylase, phosphoenolpyruvate carboxykinase, and phosphoenolpyruvate carboxytransphosphorylase.

[Plasmid profiles of Acidithiobacillus ferrooxidans strains adapted to different oxidation substrates]. / Plazmidnye profili shtammov Acidithiobacillus ferrooxidans, adaptirovannykh k raznym substratam okisleniia.

Plasmid profiles were studied in five Acidithiobacillus ferrooxidans strains of various origin cultivated on medium with Fe2+, as well as adapted to such oxidation substrates as S0, FeS2, and sulfide concentrate. The method used revealed plasmids in all A. ferrooxidans strains grown on medium with Fe2+. One plasmid was found in strain TFL-2, two plasmids, in strains TFO, TFBk, and TFV-1, and three plasmids were detected in strain TFN-d. The adaptation of strain TFN-d to sulfide concentrate and the adaptation of strain TFV-1 to S0, FeS2, or sulfide concentrate resulted in a change in the number of plasmids occurring in cells. In cells of strain TFN-d adapted to sulfide concentrate, the number of plasmids decreased from three to two. The number of plasmids in cells of strain TFV-1 adapted to different substrates varied from three to six depending on the energy source present in the medium: three plasmids were found after growth on FeS2, four after growth on S0, and six after growth on sulfide concentrate. The possible role of plasmids in the adaptation of A. ferrooxidans to new energy substrates and in the regulation of the intensity of their oxidation is discussed.

[Sulfobacillus sibiricus sp. nov., a new moderately thermophilic bacterium]. / Novaia umerenno termofil'naia bacteria Sulfobacillus sibiricus sp. nov.

In the course of pilot industrial testing of a biohydrometallurgical technology for processing goldarsenic concentrate obtained from the Nezhdaninskoe ore deposit (East Siberia, Sakha, Yakutiya), a new gram-positive rod-shaped spore-forming moderately thermophilic bacterium (designated as strain N1) oxidizing Fe2+, S0, and sulfide minerals in the presence of yeast extract (0.02%) was isolated from a dense pulp. Physiologically, strain N1 differs from previously described species of the genus Sulfobacillus in having a somewhat higher optimal growth temperature (55 degrees C). Unlike the type strain of S. thermosulfidooxidans, strain N1 could grow on medium with 1 mM thiosulfate or sodium tetrathionate as a source of energy only within several passages and failed to grow, in the absence of an inorganic energy source, on media with sucrose, fructose, glucose, reduced glutathione, alanine, cysteine, sorbitol, sodium acetate, or pyruvate. The G + C content of the DNA of strain N1 was 48.2 mol %. The strain showed 42% homology after DNA-DNA hybridization with the type strain of S. thermosulfidooxidans and 10% homology with the type strain of S. acidophilus. The isolate differed from previously studied strains of S. thermosulfidooxidans in the structure of its chromosomal DNA (determined by the method of pulsed-field gel electrophoresis) which remained stable as growth conditions were changed. According to the results of the 16S rRNA gene analysis, the new strain forms a single cluster with the bacteria of the species Sulfobacillus thermosulfidooxidans (sequence similarity of 97.9-98.6%). Based on these genetic and physiological features, strain N1 is described as a new species Sulfobacillus sibiricus sp. nov.

[Characteristics of the restriction profile of chromosomal DNA in strains of Acidithiobacillus ferroxidans, adapted to various oxidation substrates]. / Kharakteristika restriktsionnykh profilei khromosomnoi DNK u shtammov Acidithiobacillus ferrooxidans, adaptirovannykh k raznym substratam okisleniia.

Restriction profiles of chromosomal DNA were studied in different Acidithiobacillus ferrooxidans strains grown on medium with Fe2+ and further adapted to another oxidation substrate (S0, FeS2, or sulfide ore concentrates). The restriction endonuclease XbaI digested the chromosomal DNA from different strains into different numbers of fragments of various sizes. Adaptation of two strains (TFBk and TFN-d) to new oxidation substrates resulted in structural changes in XbaI-restriction patterns of their chromosomal DNA. Such changes in the DNA restriction patterns occurred in strain TFBk after the adaptation to precyanidated gravitational pyrite-arsenopyrite concentrate (no. 1) from the Nezhdaninskoe deposit or to copper-containing ore from the Udokanskoe deposit and also in strain TFN-d adapted to untreated pyrite-arsenopyrite concentrate (no. 2) from the Nezhdaninskoe deposit. No changes in the number or size of the XbaI-restriction patterns of chromosomal DNA were revealed in either strain TFBk cultivated on media with pyrite from the Angren and Tulun deposits or in strains TFN-d and TFO grown on media with S0 and pyrite. Neither were changes observed in the XbaI-restriction patterns of the DNA from strain TFV-1, isolated from the copper ore of the Volkovskoe deposit, when Fe2+ was substituted with alternative substrates--S0, pyrite or concentrate no. 2 from the ore of Nezhdaninskoe deposit. In strain TFO, no differences in the XbaI-restriction patterns of the chromosomal DNA were revealed between the culture grown on medium containing concentrate no. 2 or the concentrate of surface-lying ore from Olimpiadinskoe deposit and the culture grown on medium with Fe2+. When strain TFO was cultivated on the ore concentrate from deeper horizons of the Olimpiadinskoe deposit, which are characterized by lower oxidation degree and high antimony content, mutant TFO-2 differing from the parent strain in the chromosomal DNA structure was isolated. The correlation between the lability of chromosomal DNA structure in A. ferrooxidans strains and the physical and chemical peculiarities of the isolation substrate and habitat is discussed.

[Strain polymorphism of the plasmid profiles in Acidithiobacillus ferrooxidans]. / Shtammovyi polimorfizm plazmidnykh profilei u Acidithiobacillus ferrooxidans.

Plasmid profiles were studied in 27 Acidithiobacillus ferrooxidans strains isolated from different geographic zones and substrates differing in the composition of the main sulfide minerals, and also in experimentally obtained strains with acquired enhanced resistance to the ions of heavy metals (Fe, Ni, Cu, Zn, As). In 16 out of 20 strains isolated from different substrates, one to four 2- to 20-kb and larger plasmids were revealed. Plasmids were found in all five strains isolated from gold-containing pyrite-arsenopyrite ores and concentrates, in nine of 11 strains isolated from the ores and concentrates containing nonferrous metals, and in two of four strains isolated from the oxidation substrates of simple composition (mine waters, pyritized coals, active sludge). Changes in the plasmid profiles in some A. ferrooxidans strains (TFZ, TFI-Fe, TFV-1-Cu) with experimentally enhanced resistance to Zn2+, Fe3+, and Cu2+, respectively, were noted as compared with the initial strains. After 30 passages on S0-containing medium, strain TFBk showed changes in the copy number of plasmids. The role of plasmids in the processes of oxidation of energy substrates and in the acquired enhanced resistance to the heavy metal ions is discussed.

[Effect of the medium composition and cultivation conditions on sporulation in chemolithotrophic bacteria]. / Vliianie sostava sredy i uslovii kul'tivirovaniia na sporoobrazovanie khemolitotrofnykh bakterii.

The possibility of controlling endospore formation by changing cultivation conditions was for the first time shown in acidophilic chemolithotrophic bacteria Sulfobacillus thermosulfidooxidans type strain 1269 and the thermotolerant strain K1 formerly described as "S. thermosulfidooxidans subsp. thermotolerans". Suppression of sporulation occurred when these strains were cultured in Manning's liquid medium with yeast extract. This medium was optimized by gradually reducing the concentrations of ferrous iron salts (the source of energy), phosphorous, nitrogen, and yeast extract and simultaneously increasing the concentrations of calcium, magnesium, and manganese (the elements important for sporogenesis) to attain higher yields of endospores by strains 1269 and K1. As a result, a new medium A was proposed, in which the life cycle of the strains studied culminated in sporulation at a level of 45 and 60%, respectively, of the total cell number. In a series of additional tests, the growth temperature and medium pH were adjusted to obtain the maximum yield of endospores. The optimal ranges found were 40-50 degrees C and pH 1.8-2.2 for strain 1269 and 35-40 degrees C and pH 2.5-2.7 for strain K1. An even higher yield of endospores, amounting to 55 and 75% for strains 1269 and K1, respectively, was obtained when the above growth conditions were combined (growth on medium A at optimal temperatures and pH). Our results suggest a new approach to optimizing sporulation by acidophilic chemolithotrophs, which consists in limiting the energy and nutrient sources and using temperature and pH values within the tolerance bounds of these cultures but outside their growth of optimum ranges.

[The enzyme of carbon metabolism in the thermotolerant sulfobacillus strain K1]. / Fermenty metabolizma ugleroda u termotolerantnoi sul'fobatsilly, shtamm K1.

To determine enzymatic activities in the thermotolerant strain K1 (formerly "Sulfobacillus thermosulfidooxidans subsp. thermotolerans"), it was grown in a mineral medium with (1) thiosulfate and Fe2+ or pyrite (autotrophic conditions), (2) Fe2+, thiosulfate, and yeast extract or glucose (mixotrophic conditions), and (3) yeast extract (heterotrophic conditions). Cells grown mixo-, hetero-, and autotrophically were found to contain enzymes of the tricarboxylic acid (TCA) cycle, as well as malate synthase, an enzyme of the glyoxylate cycle. Cells grown organotrophically in a medium with yeast extract exhibited the activity of the key enzymes of the Embden-Meyerhof-Parnas and Entner-Doudoroff pathways. An increased content of carbon dioxide (up to 5 vol%) in the auto- and mixotrophic media enhanced the activity of the enzymes involved in the terminal reactions of the TCA cycle and the enzymes of the pentose phosphate pathway. Carbon dioxide was fixed in the Calvin cycle. The highest activity of ribulose bisphosphate carboxylase was detected in cells grown autotrophically at the atmospheric content of CO2 in the air used for aeration of the growth medium. The activities of pyruvate carboxylase, phosphoenolpyruvate carboxylase, phosphoenolpyruvate carboxykinase, and phosphoenolpyruvate carboxytransphosphorylase decreased with the increasing content of CO2 in the medium.

[Phenotypic features of Ferroplasma acidiphilum strains Yt and Y-2]. / Fenotipicheskie osobennosti shtammov Yt i Y-2 Ferroplasma acidiphilum.

Earlier, we described a new family of mesophilic, strictly autotrophic Fe(2+)-oxidizing archaebacteria, Ferroplasmaceae, which belongs to the order Thermoplasmales and includes the genus Ferroplasma and species F. acidiphilum (strain YT) [1]. The present work is concerned with a comparative study of phenotypic characteristics of the type strain YT and a new strain, F. acidiphilum Y-2, isolated from dense pulps produced during oxidation of arsenogold concentrates from the Bakyrchikskoe (Kazakhstan) and Olimpiadinskoe (Krasnoyarsk Krai) ore deposits, respectively. The G + C content of DNA from strains YT and Y-2 comprised 35.1 and 35.2 mol%, respectively; the level of DNA-DNA homology between the strains was 84%. Restriction profiles of chromosomal DNA from both strains exhibited a similarity coefficient of 0.87. Genotypic characteristics of these strains indicate their affiliation to the same species. The cells of both strains are polymorphic and lack cell walls. Strains of F. acidiphilum oxidized ferrous oxide and pyrite as the sole source of energy and fixed carbon dioxide as the sole carbon source. Strains required yeast extract as a growth factor. Optimum pH for cell growth ranged from 1.7 to 1.8; the temperature optima for the growth of strains YT and Y-2 were 34-36 and 40-42 degrees C, respectively. Comparative analysis of total lipids revealed their close similarity in the strains; two glycophospholipids comprised 90% of total lipids: lipid I, beta-D-glucopyranosylcaldarchaetidylglycerol (about 55%), and lipid II, trihexosylcaldarchaetidylglycerol (26%), whose isopranyl chains contained no cyclopentane rings. The carbohydrate fraction of lipid I hydrolysate contained only D-glucose, whereas hydrolysate of lipid II contained both D-glucose and D-galactose in a molar ratio of 2:1. Thus, it was established that the intraspecific phylogenetic divergence within F. acidiphilum is manifested in two the strains by different temperature optima against the background of similarity in other phenotypic properties.

[The treatment industrial sewage of nonferrous heavy metals using biosorbents]. / Ochistka promyshlennykh stochnykh vod ot tiazhelykh tsvetnykh metallov s pomoshch'iu biosorbentov.

Producers of the antibiotics neomycin and lincomycin were most potent in sorbing nonferrous metals; they removed 99% Zn and Cd and 95% Ni. The degree of metal sorption increased with an increase in solution pH and calcium content in the biomass. Dynamic studies of biosorbents prepared from microbial biomass showed that neomycin industrial waste products are highly efficient in treating galvanic washing water. The total dynamic exchange capacities by nickel and cadmium were 6.85 and 7.16 mg/ml, respectively.