Investigating the Vegetation–Soil Relationships on the Copper–Cobalt Rock Outcrops of Katanga (D. R. Congo), an Essential Step in a Biodiversity Conservation Plan

Plant communities of soils naturally enriched in copper and cobalt in Katanga (D. R. Congo) are critically threatened in the short term due to mining activities. For biodiversity conservation and ecosystem restoration purposes, there is an urgent need to acquire more knowledge on those plant communities including their diversity and their relationships to environmental factors. The classification of 62 vegetation plots located in 6 metal‐rich rocky hills in the Tenke Fungurume mining area resulted in 3 well‐defined steppic and steppic savanna communities. Canonical analysis showed that the community comprising the largest proportion of strictly endemic metallophytes (i.e. species that only occur on metal‐rich soils) developed in the soils with the most elevated concentrations of Cu and Co. However, contrasting species assemblages in the two other plant communities were explained by soil nutrients and percentage rocks in addition to heavy metal concentrations. The results of this study will assist with restoration efforts because they (1) provide a rigorous assessment of communities before a disturbance and (2) define essential edaphic conditions needed for the reestablishment of critical communities.     

Research Priorities for Conservation of Metallophyte Biodiversity and their Potential for Restoration and Site Remediation

Plants that have evolved to survive on metal‐rich soils—metallophytes—have key values that must drive research of their unique properties and ultimately their conservation. The ability of metallophytes to tolerate extreme metal concentrations commends them for revegetation of mines and metal‐contaminated sites. Metallophytes can also be exploited in environmental technologies, for example, phytostabilization, phytoremediation, and phytomining. Actions towards conserving metallophyte species are imperative, as metallophytes are increasingly under threat of extinction from mining activity. Although many hundreds of papers describe both the biology and applications of metallophytes, few have investigated the urgent need to conserve these unique species. This paper identifies the current state of metallophyte research, and advocates future research needs for the conservation of metallophyte biodiversity and the sustainable uses of metallophyte species in restoration, rehabilitation, contaminated site remediation, and other nascent phytotechnologies. Six fundamental questions are addressed: (1) Is enough known about the global status of metallophytes to ensure their conservation? (2) Are metallophytes threatened by the activities of the minerals industry, and can their potential for the restoration or rehabilitation of mined and disturbed land be realized? (3) What problems exist in gaining prior informed consent to access metallophyte genetic resources and how can the benefits arising from their uses be equitably shared? (4) What potential do metallophytes offer as a resource base for phytotechnologies? (5) Can genetic modification be used to “design” metallophytes to use in the remediation of contaminated land? (6) Does the prospect of using metallophytes in site remediation and restoration raise ethical issues?     

Specialized edaphic niches of threatened copper endemic plant species in the D.R. Congo: implications for ex situ conservation

Background and aims

Copper (Cu) rich soils derived from rocks of the Katangan Copperbelt in the south-eastern Democratic Republic of Congo (DRC) support a rich diversity of metallophytes including 550 heavy metal tolerant; 24 broad Cu soil endemic; and 33 strict Cu soil endemic plant species. The majority of the plant species occur on prominent Cu hills scattered along the copperbelt. Heavy metal mining on the Katangan Copperbelt has resulted in extensive degradation and destruction of the Cu hill ecosystems. As a result, approximately 80 % of the strict Cu endemic plant species are classified as threatened according to IUCN criteria and represent a conservation priority. Little is known about the soil Cu tolerance optimum of the Cu endemic plant species. The purpose of this study was to quantify the soil Cu concentration (Cu edaphic niche) of four Cu endemic plant species to inform soil propagation conditions and microhabitat site selection for planting of the species in Cu hill ecosystem restoration.

Methods

The soil Cu concentration tolerance of Cu endemic plant species was studied including Crotalaria cobalticola (CRCO); Gladiolus ledoctei (GLLE); Diplolophium marthozianum (DIMA); and Triumfetta welwitschii var. rogersii (TRWE-RO). The in situ natural habitat distributions of the Cu endemic plant species with respect to soil Cu concentration (Cu edaphic niche) was calculated by means of a generalised additive model. Additionally, the seedling emergence and growth of the four Cu endemic plant species in three soil Cu concentrations was tested ex situ and the results were compared to that of the natural habitat soil Cu concentration optimum (Cu edaphic niche).

Results

CRCO exhibited greater performance on the highest soil Cu concentration, consistent with its calculated Cu edaphic niche occurring at the highest soil Cu concentrations. In contrast, both DIMA and TRWE-RO exhibited greatest performance at the lowest soil Cu concentration, despite the calculated Cu edaphic niche occurring at moderate soil Cu concentrations. GLLE exhibited equal performances in the entire range of soil Cu concentrations.

Conclusions

These results suggest that CRCO evolved via the edaphic specialization model where it is most competitive in Cu hill habitat with the highest soil Cu concentration. In comparison, DIMA and TRWE-RO appear to have evolved via the endemism refuge model, which indicates that the species were excluded into (i.e., took refuge in) the lower plant competition Cu hill habitat due to their inability to effectively compete with higher plant competition on normal soils. The soil Cu edaphic niche determined for the four species will be useful in conservation activities including informing soil propagation conditions and microhabitat site selection for planting of the species in Cu hill ecosystem restoration.

     

May Rare Metallophytes Benefit from Disturbed Soils Following Mining Activity? The Case of the Crepidorhopalon tenuis in Katanga (D. R. Congo)

Cuprophytes are plants that mostly occur on Cu‐rich soil. In South Central Africa, these species are threatened by intensive mining exploitation destroying their habitats. Crepidorhopalon tenuis (Scrophulariaceae) is a tiny annual cuprophyte endemic to the Zambesian center of endemism and is particularly abundant in the Lubumbashi area. We investigate here the ecological niche of C. tenuis through the analyses of its abundance and distribution in relation to soil factors, plant community composition, and anthropogenic perturbations. Soil and vegetation data were collected in seven sites (five metalliferous and two nonmetalliferous). The current study shows that C. tenuis has its ecological optimum on copper‐rich soil and can be referred to as an elective pseudometallophyte. This species is rare in primary steppic savanna on natural metalliferous soil. Its frequency and abundance peak in pioneer communities on bare soil. In particular, the species showed a surprising ecological plasticity as it was able to benefit from anthropogenic disturbance and to colonize the large areas of bare, contaminated soil left over by mining activities. Our results strongly suggest that C. tenuis was a very rare species in natural metalliferous communities, restricted to patchy areas of open soil in steppic savanna. Recent anthropogenic habitats may have conservation value for some rare metallophytes with colonizing traits and low competitive ability.     

Systematics and evolutionary history of heavy metal tolerant Thlaspi caerulescens in Western Europe: evidence from genetic studies based on isozyme analysis

Thlaspi caerulescens is distributed in Europe on metalliferous and not metalliferous soils. Individuals from populations growing on heavy metal contaminated soils are well known as hyperaccumulators of zinc and cadmium. The taxonomical treatment of subspecies of Thlaspi caerulescens is unsettled. We investigated the degree of genetic variation among 28 populations of Thlaspi caerulescens from Europe with isozyme analysis to compare inter- and intrapopulational diversity. British material from heavy metal contaminated environments recognized as Thlaspi sylvestre and T. occitanicum are quite similar to each other on the level of isozyme polymophisms, but they are more closely related to populations from non-contaminated stands from Scandinavia and Middle Europe than to metallophytes distributed in Continental Europe. Our findings indicate that a taxonomical subdivision of T. caerulescens is not possible and, furthermore, heavy metal tolerance might have evolved twice in populations of Thlaspi caerulescens from different areas. The trait of zinc tolerance and hyperaccumulation is frequently found in numerous relatives of Thlaspi caerulescens, and it is suggested that this trait has been established and manifested in populations from metalliferous sites during postglacial colonization. From Scandinavia only non-metallophytes are known. These populations are very similar to each other on the isozyme level. This fits to the hypothesis that Thlaspi caerulescens was introduced to Scandinavia in recent times by human activity. Despite full self-compatibility we estimated varying outcrossing rates up to 0.88 in the metallophytes and 0.658 in the non-metallophytes depending on population size and structure.     

Ecology and Hybridization Potential of Two Sympatric Metallophytes,the Narrow Endemic Crepidorhopalon perennis (Linderniaceae) and its More Widespread Congener C. tenuis

Crepidorhopalon perennis is a metallophyte critically endangered by mining activities and currently known from only one site on copper‐rich soils in Katanga (Dem. Rep. Congo). It is closely related to the annual C. tenuis, also a rare metallophyte, but with a broader geographical range. We investigated the variation in morphometric traits and ecological niches (based on edaphic conditions and pollinator assemblages) of C. perennis and C. tenuis, to evaluate the risk of inter‐specific competition, and their potential for hybridization to ascertain if C. perennis might be at risk of genetic swamping by its more widespread congener. We examined whether species were found under sympatric or parapatric settings with opportunity for hybridization (or gene exchange). Such knowledge is essential for implementing restoration management protocols, including the introduction of C. perennis into substitution sites where C. tenuis might be already present. Fourteen morphological characters and 11 soil variables were measured and visiting pollinator species were identified at the site where the two species co‐occur. Our results show that the two species can be distinguished based on their morphological traits, show niche overlap based on edaphic properties, and share the same pollinator assemblage. In addition, no morphologically intermediate individuals could be detected, suggesting no hybridization, and that the two species may be reproductively isolated. We conclude that C. perennis conservation and restoration operations can be realized in substitution sites where C. tenuis may be present, with the need, however, to evaluate the potential effect of sharing a pollinator assemblage on reproductive success of both species.     

Studies on copper and cobalt tolerance in three closely related taxa within the genusSilene L. (Caryophyllaceae) from Zaïre

Summary Experiments were carried out on the tolerance to and uptake of copper and cobalt by three members of a phylogenetic series of taxa within the genusSilene from Zaïre which were reputed to represent a progression of increasing adaptation to metalliferous soils. Plants studied were the widespread and presumably non-tolerantSilene burchelli var.angustifolia (B), a more tolerant variant of this taxon (E) designated an ecotype, and the metallophyteS. cobalticola (C). Studies on the effects of both metals, singly and in combination, on seed germination, seedling and plant performance and yield, and metal uptake from soil cultures, confirmed in general the sequence B<E<C in relation to tolerance of both copper and cobalt. The experimental evidence also supports the ecotypic status of E and points to E and C being relatively more tolerant to copper than to cobalt.     

Soils and vegetation of central Australian sandridges. IV. Soils.

Catenary patterns on central Australian sandridges are assessed for the following soil parameters: sand depth, texture, mobility and penetrability; moisture content and characteristic; organic C content, total N and extractable P, K and Ca. There are marked dune-swale gradients in sand depth, soil texture, mobility and penetrability, moisture content at — 15 kPa, and N and organic C contents. There are dune-swale gradients in soil Ca content at a few sites only. Soil P shows significant heterogeneity but no catenary patterns, whilst K levels are homogeneous throughout. S, Mg, Fe, Mn, Cu, Zn, B, Mo, Co and CI were also measured for a number of sandridge soils but showed no significant catenary patterns and were therefore not studied in detail.     

Distribution of the endemic Balkan flora on serpentine I. – obligate serpentine endemics

Serpentine (ophiolithic) substrate covers large areas in the Balkans, more so than in any other part of Europe. These areas extend from north to south mainly in the mountainous central regions and represent specialized habitats for basiphilous-calcifugal plants. Biodiversity in the area is high, with a great number of interesting local and regional endemics. The high number of endemics indicates the importance of serpentine habitats as centres for floristic differentiation and speciation. The number of Balkan endemics growing on serpentine is c. 335 taxa (species and subspecies) of which 123 are obligate. Their distribution is presented in 50 × 50 km UTM squares as adopted in the Atlas Florae Europaeae project coordinated at Helsinki. The richest (in number of taxa) squares are situated in NW Greece (Epirus), the island of Evvia, N Albania together with SW Serbia, and N Greece (Vourinos). They indicate important centres of plant diversity in the Balkans, areas to be noted for conservation strategy. Features responsible for the distribution and abundance of these obligate serpentine endemics include: 1) edaphic isolation in relation to type of bedrock (lime, dolomite, marble, schist, etc.), 2) mountain island isolation (Smolikas, Vourinos, Ostrovica, etc.), 3) island isolation (Evvia) and 4) continuous long-term isolation without interruption or disturbance of speciation.     

Local Adaptation of Metallicolous and Non‐Metallicolous Anthyllis vulneraria Populations: Their Utilization in Soil Restoration

Restoration of metalliferous mine soils requires using plant species tolerant to high metal concentrations and adapted to nutrient‐poor soil. Legumes can increase plant productivity through N₂‐fixation, but they are often scarce in metalliferous sites. We examined survival, growth, and tolerance of four populations of a legume, Anthyllis vulneraria, from two metalliferous (MET) Zn‐Pb mine sites, Avinières (AV) ([Zn‐EDTA] = 26,000 mg/kg) and Eylie (EY) ([Zn‐EDTA] = 4,632 mg/kg), and two non‐metalliferous (NMET) sites located in the south of France with the aim to select the most appropriate populations for restoration of mined soils. In a common garden experiment, plants from each population were reciprocally grown in soil from the provenance of each population. The two NMET populations exhibited high mortality and low growth rates in soil from the mined sites. The AV MET exhibited a high growth rate in metalliferous soils, but showed high mortality in non‐metalliferous soils. The growth of the EY MET was very low in the AV‐contaminated soil, but was the highest of all populations in moderately and non‐metalliferous soils. Plants from the AV MET population showed a high growth and survival in metalliferous soil and would be appropriate in the restoration of metal‐contaminated sites (>30,000 mg Zn kg^?1). The EY MET population would be adapted to the restoration of moderate metal‐contaminated soils (<30,000 mg Zn kg^?1). Taking into account the broad distribution of A. vulneraria, these two populations could be suitable for the restoration of derelict mine sites in mediterranean and temperate regions of Europe and North America.     

Community structure on unusual habitat islands: quartz-fields in the Succulent Karoo,South Africa

Quartz fields are edaphically arid, azonal habitats occurring under different macroclimatic conditions in several arid regions of southern Africa. They are the exclusive home of 142 plant species of which ca. 70% are local or regional endemics. This paper is an analysis of the quartz-field floras and growth form-soil relationships in two quartz field regions: the Knersvlakte in the Namaqualand-Namib Domain of the Succulent Karoo, and the western Little Karoo in the Southern Karoo Domain. The Knersvlakte supported 52 quartz-field specialists of which 39 were endemic to the region. Corresponding data for the Little Karoo were 11, and 10 species, respectively. In both regions, the average canopy cover on the quartz-field relevés was ca. 8%, and more than half of this comprised contracted, succulent nanochamaephytes. Cover and vegetation stature were markedly higher on adjacent zonal habitats. Quartz fields in both regions supported a similar array of compact, subglobose and subterranean nanochamaephytes, as evidenced by convergent patterns in two distantly related genera (Argyroderma N. E. Brown and Gibbaeum (Haworth) N. E. Brown, both Mesembryanthemaceae), endemic to the Knersvlakte and largely restricted to the Little Karoo, respectively. Analyses of vegetational and edaphic data of quartz fields and adjacent, zonal habitats were carried out using multivariate direct gradient analysis (Canonical Correspondence Analysis) in order to identify those factors that control the peculiar composition of growth forms on quartz fields. The results revealed highly similar patterns of growth form composition in relation to similar edaphic gradients in both regions. In general, the soils of quartz fields were shallower compared to those of adjacent zonal habitats. In both regions, two different groups of quartz-field edaphic habitats, representing extremes of a continuum, were identified. Group 1 was characterized by high salt content, neutral to slightly acid soil pH, and low stone content. Group 2 was characterized by low salt content, low soil pH, and high stone content. Group 1 quartz fields are the most edaphically arid habitats and support the highest relative cover and diversity of subglobose and subterranean chamaephytes. The combination of reduced competition from larger growth forms, shallow soils and high soil salinity, represents a regionally unusual selective regime. Some succulent lineages in the Mesembryanthemaceae have undergone diversification which has resulted in the fine-scale discrimination of subtle edaphic gradients within the saline quartz-patch habitats. Reliable seasonal rainfall and reduced thermal stress have also played a role in the evolution of quartz patch specialists.     

Trace element budget in an African savannah ecosystem

The concentration of selected trace elements (Co, Cu, Fe, Mn, Mo, Se, and Zn) were analysed in soils, grass, bush, and tree samples from the Mole National Park, Ghana. The distribution of the essential nutrients: cobalt, copper, manganese, and selenium is controlled by bedrock geology, whereas iron, molybdenum, and zinc distribution is controlled by soil and hydrological processes. In the soils, iron, manganese, and cobalt are largely fixed in the mineral fraction while most of the copper, molybdenum, and selenium in the soils can be extracted by disodium ethylenediaminetetracetate. Copper, cobalt, and manganese appear to be preferentially concentrated in grass species while molybdenum and selenium are concentrated in browse plants. Variations in uptake exist between wet and dry seasons with all trace elements studied, except iron and manganese, showing a marked increased availability in the wet season and increased concentration in the residual fraction of the mineral and organic soils in the dry season. In the dry season the plant concentration of molybdenum and selenium decreased while copper and zine showed increased concentrations and this may be related to a lower pH of the groundwaters at this time. A budget of metal input and output in the ecosystem at Mole has been computed. From this potential dietary deficiencies in cobalt can be observed, however for other metals soil and plant concentrations are sufficient to prevent straightforward deficiencies while the concentrations of molybdenum and selenium are sufficiently low to be considered safe.     

Use of bryological communities as a diagnostic tool of heavy metal soil contamination (Cd,Pb, Zn) in northern France

Correlations between terricolous bryophyte flora and edaphic parameters (pH, organic nitrogen (ON), C/N, Cation Exchange Capacity (CEC), Organic Matter (OM) and Pb, Cd and Zn soil contents) of metalliferous wastes in northern France were performed using statistical analysis (FCA, PCA and FCMA). The significant correlation between the classification based on the bryophyte flora and that based on edaphic variables shows that bryophyte groupings have a strong bioindicative value. More information was obtained by taking into account other parameters such as physiological stages (sterility, fertility, maturation, vitality). Three bryophyte classes were distinguished according to Cd, Pb and Zn soil contents.     

Concentrations of Metals in Soils in the Neighborhood of a Hazardous Waste Incinerator: Assessment of the Temporal Trends

The concentrations of arsenic (As), beryllium (Be), cadmium (Cd), cobalt (Co), chromium (Cr), copper (Cu), mercury (Hg), manganese (Mn), nickel (Ni), lead (Pb), antimony (Sb), tin (Sn), thallium (Tl), and vanadium (V) were determined in 30 soil samples collected in April 2011 near a hazardous waste incinerator (HWI; Constantí, Catalonia, Spain), which is under regular operations since 1999. The results were compared with those of recent surveys performed in the same zone, as well as with data from a background study (1996?C1998). We also assessed the human health risks derived from metal exposure in the area under potential influence of the emissions of the HWI. Manganese was the most abundant element in soils, with a mean value of 316.4???g/g, followed by Pb and Cu (mean values, 42.5 and 38.2???g/g, respectively). In contrast, Sb, Cd, and Tl presented the lowest values (0.12, 0.27, and 0.29???g/g, respectively), while Hg was below its limit of detection. In the period 1998?C2011, only As, Cr, Sn, Tl, and V levels presented significant increases in soils. The estimated carcinogenic and noncarcinogenic risks derived from exposure to these metals from soils should not mean any special concern for the population living in the surroundings of the HWI.     

The diverse chemical forms of heavy metals in tissue extracts of some metallophytes from Shaba Province,Zaïre

Copper and cobalt were determined in tissue extracts of five metallophytes from Shaba Province, Zaïre. About 4O% of the cobalt and copper was extractable into deionized water and a further 4O% was extractable in 0.2M hydrochloric acid. It was concluded that copper and cobalt are bound to several different ligands instead of a single ligand as in the case of nickel hyperaccumulators. Proton microprobe studies on the cobalt accumulator Haumaniastrum robertii showed a strong inverse correlation between the spatial distributions of cobalt and potassium, as well as a direct relationship between cobalt and calcium. It is suggested that some of the cobalt may be immobilized with calcium in oxalate crystals.     

Trace Element Composition of Selected Fertilizers Used in Chile: Phosphorus Fertilizers as a Source of Long-Term Soil Contamination

Anthropogenic activities like agriculture have resulted in increased concentrations of some trace elements of toxicological and environmental concern in soils. Application of fertilizers has been one of the major inputs of these contaminants to agricultural soils in developing countries. Twenty-two fertilizers, including straight nitrogen (N), phosphorus (P), potassium (K), and NK fertilizers and micronutrient sources, were analyzed by inductively coupled plasma optical emission spectrometry (ICP-OES) for arsenic (As), cadmium (Cd), cobalt (Co), chromium (Cr), copper (Cu), iron (Fe), manganese (Mn), molybdenum (Mo), nickel (Ni), lead (Pb), vanadium (V), and zinc (Zn). As expected, the trace element content of fertilizers was highly variable and related to the origin of the material. Phosphorus fertilizers, especially triple superphosphate, presented the highest As, Cd, Cu, Cr, Ni, V, and Zn concentrations. In some of these fertilizers, the Cr, V, and Zn contents reached values greater than 3475 mg kg^?1 of P, and the Cd content (up to 288 mg kg^?1 of P) was several times higher than the regulatory limits of different countries. Some micronutrient sources presented the highest concentrations of Mn and Pb. In the cases of N, K, and NK fertilizers, the trace element concentration was very low, sometimes below the detection limits. In some agricultural systems the input of trace elements such as As and Cd to the soil through P fertilizers application may be higher than the outputs through plant uptake and leaching; therefore the long-term use of these fertilizers may cause the trace element concentration to increase in the plow layer of agricultural soils.     

Does phylogenetic distance aid in detecting environmental gradients related to species composition?

Questions: How should we evaluate the success of new distance measures combining community abundance and phylogenetic information? How do we interpret ordinations using these metrics? Methods: We generated synthetic data along a known environmental gradient with two hypothetical underlying phylogenetic structures: niche phylogenetically conserved or dispersed along a gradient. We also examined tree species composition associated with gradients in elevation and longitude in Oregon, USA. NMS ordinations of plots in species space from phylogenetic (PD) and Sørensen distance (SD) matrices allowed comparison of the use of PD in different scenarios. Results: PD caused plots to cluster based on the clades that they contained, reducing stress with the synthetic data but not with the real example. Phylogenetic distance highlighted clades related to gradients when these were associated. When phylogeny was not conserved along a gradient, that gradient was less strong. Regardless of phylogenetic conservation, NMS using SD consistently extracted the strongest gradients in species composition. Conclusions: The success of PD should be evaluated on how well it extracts gradients in species composition and allows community ecologists to determine which gradients are partially explained by phylogeny and not based on its ability to reduce ordination stress. PD ordinations can help community ecologists interpret niche conservation but may obscure gradients related to species composition when niches are not conserved along the gradient of interest at the scale of the study.     

Phylogenetic analysis of the endemic New Caledonian cockroach Lauraesilpha. Testing competing hypotheses of diversification

New Caledonia is a tropical hotspot of biodiversity with high rates of regional and local endemism. Despite offering an ideal setting to study the evolution of endemism, New Caledonia has received little attention compared with the other nearby hotspots, particularly New Zealand. Most studies of the Neocaledonian endemism have been carried out at the regional level, comparing the various groups and species present in New Caledonia but absent in neighboring territories. In addition, remarkably high short‐range endemism has been documented among plants, lizard and invertebrates, although these have usually been done, lacking a phylogenetic perspective. Most studies of Neocaledonian endemism have referred to the geological Gondwanan antiquity of the island and its metalliferous soils derived from ultramafic rocks. Very old clades are thought to have been maintained in refugia and diversified on the metalliferous soils. The present study documents the pattern of diversification and establishment of short‐range endemism in a phylogenetic context using the Neocaledonian cockroach genus Lauraesilpha. Mitochondrial and nuclear genes were sequenced to reconstruct phylogenetic relationships among the species of this genus. These relationships, in the light of the species distribution, do not support the hypothesis that species diversified via an adaptive radiation on metalliferous soils and are not consistent with areas of highest rainfall. Species of Lauraesilpha have similar altitudinal ranges and ecological habits and are short‐range endemics on mountains. What our analysis did reveal was that closely related species are found on nearby or contiguous mountains, and thus these formations probably played the key role establishing short‐range endemism (in association with recent climatic changes). © The Willi Hennig Society 2008.     

Does cobalt really bind tightly to hemocyanin active sites?

The analysis of Co(II)-apoHc complexes of two arthropodan species (freshwater crayfish): Orconectes limosus and Astacus astacus enabled to reach some conclusions about possible cobalt binding sites in the hemocyanin molecules. The occurrence of binding sites for Co(II) at sites other than the active center has been demonstrated. We excluded the possibility of strong binding of EDTA-non-removable cobalt ions in the binding sites occupied by copper. There were no differences between apoHc and the Co(II)apoHc complex in terms of the amount of bound Cu(I) ions and the kinetics of Cu(I) ion reconstitution.Abbreviations He hemocyanin - apoHc apohemocyanin - oxyHc oxyhemocyanin - Co-Hc hemocyanin complex with cobalt ions Offprint requests to: E. Serafln