Metal–Halogen Biomaterials
Robert M. S. Schofield
T
he jaws, tarsal claws, stings, and other
“tools” of a large fraction of arthropods,
some worms, and members of other phyla
contain extraordinary amounts of heavy metals (e.g., zinc, manganese, copper) and halogens
(bromine, chlorine). Although the measured concentrations reach 25% of dry mass, the tissue is not
filled with a biomineral, like calcified tissues are.
Instead, metal–halogen biomaterials appear to be
part of a distinctly different system that is widely
found among small organisms. Very little is known
about the form and function of these biomaterials
or their role in the behavior, ecology, and evolution
of invertebrates.
Occurrence: Enrichment Patterns, Enriched
Structures, and Phylogenetic Distribution
Metal–halogen enrichment is often present in
complex distribution patterns involving more than
one metal. The spider chelicera in Fig. 1 demonstrates a typical metal–halogen enrichment pattern
seen even in marine worms: zinc along with chlorine
in one region (the fang, in this case), and manganese
along with calcium in a second, more proximal
region (the marginal teeth). Near the tip of the spider fang in Fig. 1, the measured zinc concentration
reached 15(+/–3)% of dry mass, and the Cl concentration 3(+/–1)%. In the marginal teeth, manganese
concentrations reached 3.7(+/–0.7)% (Schofield
1990; Schofield and Lefevre 1992, 1993).
The structures in which metal–halogen biomaterials have been found are not all mouthparts, but
they all come into direct contact with the environment. Fig. 2 demonstrates metal–halogen biomaterials in the tarsal claws of scorpions. In our work
at the University of Oregon, we have found zinc in
the contact regions of structures, such as mandibles,
chelicerae, pedipalps, forcipules, jaws, paragnaths,
tarsal claws, pedal spurs, stings, and stylets. Our
findings, along with those of other researchers, are
reviewed in Schofield (2001). In each of these structures, metals were localized in regions susceptible
to abrasion and mechanical force by contact with
the environment. Metal–halogen biomaterials have
not been found in structures such as joints, which
might need hardening but do not usually come into
contact with the environment.
Which organisms use these materials? The
strongest predictor of whether an organism contains
metal–halogen biomaterials is not its behavior or
habitat, but whether or not other members of its
family also use metal–halogen biomaterials. AssociaAmerican Entomologist • Volume 51, Number 1
Fig. 1. Zinc and chlorine in the fang, manganese and calcium in
the marginal teeth of a garden spider, Araneus diadematus. The
large image is a Scanning Transmission Ion Microscopy image in
which lighter shades indicate greater projected mass. The smaller
images are Proton Induced X-ray Emission images showing the
origin of chlorine, calcium, manganese, and zinc X-rays. Frame
size: 1 mm × 1mm. Source: Schofield 2001.
tions with feeding, such as herbivory, or with habitat
have been suggested, but these have not withstood
the expanding catalogue of species; for example, zinc
and manganese enrichment have been found in all of
the 14 examined species in the ant family (Hillerton
and Vincent 1982; Schofield 1990, 2001, unpublished
data), which range widely in both habitat and in feeding behavior, from carnivory to herbivory (Schofield
1990). On the other hand, phylogenetic surveys
(Hillerton and Vincent 1982; Hillerton et al. 1984;
Schofield 1990, 2001; Fontaine et al. 1991; Quicke
et al. 1998; Schofield 2001) have only rarely found
metal distribution differences between the species
within any single arthropod family.
Metal–halogen biomaterials are widely distributed, especially among arthropods. High concentrations of zinc have been found in the “tools”
of at least 136 species in five orders of insects, 4
species in one order of centipedes, 30 species in
six orders of arachnids 12 species in one order of
polychaete worms, and in 2 species in one order
Fig. 2. Metal–halogen bio-materials
at the tips of tarsal claws from two
species of scorpion. The member
of the buthid family, Centruroides
exilicauda, has large quantities
of manganese but little zinc in
the lateral claws. In contrast,
Vaejovis confusus has moderate
amounts of manganese and large
quantities of zinc at the tips of the
medial as well as the lateral claws.
The former distribution pattern
is characteristic of all examined
buthids, while the latter is characteristic of all examined non-buthid
scorpions. Frame size: 1 mm ×
1mm. Source: Schofield 2001.
45
proximal manganese and calcium;
• a similar time course of incorporation in the
distantly related ants and scorpions; and
• similar ultrastructure, such as the nano-scale
canals that seem to be associated with zinc
incorporation.
Fig. 3. Zinc is
incorporated late in
cuticle development
of the scorpion
Vaejovis spinigerus.
Three specimens
from littermates of
different ages are
shown in each image:
clockwise from the
top left in each image,
38, 90, and 160 h
post-ecdysis. The zinc
content of the various
enriched structures is
very low at 38 h and
nearly at adult levels
by 160 h. Frame edge:
1.1, 0.55, and 0.82 mm
for the pedipalp, tarsal
claw, and chelicera
images respectively.
Source: Schofield et
al. 2003
of nemertean worms (a table of the distribution
in the higher taxa is included in Schofield 2001).
Other heavy metals, manganese, iron, and copper,
as well as the halogens chlorine and bromine have
been found in >1% levels in association with zinc
or by themselves in comparable numbers of species.
We have found metal–halogen biomaterials in more
than half of the families that we have examined.
Although this is not an unbiased survey, it suggests
that metal–halogen biomaterials are extremely
common and widespread.
Development and Ultrastructure
Zinc is incorporated into the cuticle very late, after the cuticle is formed and sclerotized (Schofield et
al. 2003). In the ant Tapinoma sessile, zinc incorporation began about 125 to 150 h after pre-ecdysial
tanning (and mainly after eclosion), and in second
instars of the scorpion Vaejovis spinigerus (Fig. 3),
it began between ≈150 and 190 h after pre-ecdysial
tanning (and >50 h after ecdysis). The zinc was
distributed uniformly throughout the metal-bearing
cuticle in the ant and the scorpion. Any electron
density associated with a separate zinc-containing
phase was limited in size to less than a few nanometers, the resolution of our technique.
How is zinc incorporated homogeneously into
cuticle at such a late stage? We found that the
metal-bearing portion of the cuticle was filled with
nanometer-scale canals in the scorpions and the
ants. These canals were not present in adjacent nonmetal–bearing cuticle or in cuticle of other toothlike
features that were not metal-enriched. Near the
boundary of the zinc-enriched regions, the cuticle
immediately surrounding the nanometer-scale and
larger canals was electron-dense and contained the
highest concentrations of zinc. This suggests that
zinc was incorporated through these canals.
Evolution
Several pieces of evidence suggest that metal–halogen biomaterials evolved very early, possibly before
the divergence of annelids and arthropods:
• the widespread phylogenetic distribution discussed
earlier;
• the repeated element distribution patterns, found
even in worm jaws: distal zinc and chlorine,
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Chemistry
One of the most interesting questions about
metal halogen biomaterials is their chemical form.
Biological metals are present as single ions bound
to protein side chains, or are bound in a biomineral, such as the iron-containing magnetite and
goethite in the radular teeth of certain mollusks.
The concentrations of heavy metals in cuticular
structures are difficult to reconcile with either of
these molecular forms.
The 25% zinc concentration in scorpion cuticle
seems too high to be accounted for by binding of
individual ions to protein side-chains. Hillerton and
Vincent (1982) presented such a hypothesis, suggesting that zinc in the mandibles of insects might
increase the number of secondary bonds in the
cuticle and thereby increase the density and fracture
toughness of these structures. However, we have
estimated the maximum possible zinc concentrations for two organic binding mechanisms (binding
to amino acid side chains of cuticular proteins, or
binding to catecholate ligands, which accumulate
during cuticle sclerotization), and it was found that
both gave maximum zinc concentrations <8% of
dry mass, well under the concentrations in many
organisms (Schofield 1990, 2001).
On the other hand, in addition to not producing
mineral diffraction patterns, the concentrations of
zinc in cuticular “tools” are very low compared
with cation concentrations in biomineralized
structures. There would not be enough of a putative zinc biomineral to fill the tissue to the extent
that the mechanical properties of the tissue would
approach those of the pure mineral, as in calcified
tissue. In addition, structural biominerals typically occur as large inclusions of pure mineral in
an organic matrix. As mentioned before, no such
inclusions are evident in the examined metal–halogen biomaterials.
Perhaps the answer lies in a combination of
these two binding modes. We are testing the
hypothesis that the zinc is bound in biomineral
nanoclusters attached to proteins.
Mechanical Properties
The functions of the enriched structures suggest
that metal–halogen biomaterials enhance mechanical properties. We used atomic force microscopy to
measure the hardness of the mandibular teeth of
the leaf cutter ant Atta sexdens during the period
of zinc incorporation (Schofield et al. 2002). As
with the Tapinoma sp., mandibular teeth of Atta
contain very little zinc at the time of eclosion. We
found that the hardness of the mandibular teeth
increases almost three-fold as the zinc is incorporated, whereas the hardness of the zinc-free
off-tooth region does not increase.
American Entomologist • Spring 2005
Fig. 4. Metal–halogen enriched cuticle appears more resistant to
chipping than calcified cuticle. A cheliped claw of the shore crab,
Pachygrapsus crassipes, before (left), and after (right) bead-blasting.
The dark region at the tip of the pedipalp claw is enriched with bromine.
Source: Schofield and Nesson, unpublished data.
Whereas zinc-enriched cuticle is harder than
zinc-free cuticle, biomineralized cuticle is often
even harder. Crustaceans such as isopods and
certain crabs contain calcium biominerals and
metal–halogen biomaterials. The metal–halogen
biomaterials are found at the tips of structures such
as claws, and the rest of the cuticle is calcified and
harder. The tips of these structures are particularly
susceptible to wear and fracture. Fig. 4 shows that
the dark metal–halogen biomaterial at the tips of a
crab cheliped claw does not chip away as fast as the
lighter, calcified cuticle when the claw is subjected
to bead-blasting. Although hardness is probably
desirable at the tip, the metal–halogen biomaterial may provide a better balance of hardness and
resistance to wear and fracture.
Simple models suggest that wear and fracture
resistance become increasingly important in smaller
structures. This may explain the prevalence of
metal–halogen biomaterials over biomineralization
in the mechanical structures of small organisms.
Summary
It is likely that metal–halogen biomaterials
affect the behavior and ecology of the large fraction of arthropods in which they are found. For
example, leaf cutter ants may delay leaf cutting
until zinc has hardened their mandibles (Schofield
et al. 2002). In addition, an unknown and interesting biochemistry is associated with the binding
of so much metal in the cuticle. This chemistry
improves mechanical properties and may lead to
new biomimetic man-made polymers. Finally, the
study of metal–halogen biomaterials should help
us better understand the balance of mechanical
properties needed in smaller organisms.
Zool. J. Linn. Soc. 124: 387–396.
Schofield, R. M. S. 1990. X-ray microanalytic concentration
measurements in unsectioned specimens: a technique
and its application to zinc, manganese, and iron enriched
mechanical structures of organisms from three phyla.
Ph.D. dissertation, University of Oregon.
Schofield, R. M. S. 2001. Metals in cuticular structures, pp.
In P. Brownell and G. Polis [Eds.] Scorpion biology and
research. Oxford University Press, Oxford, U.K.
Schofield, R. M. S., and H. W. Lefevre. 1992. PIXE–STIM
microtomography: zinc and manganese concentrations
in a scorpion stinger. Nucl. Instrum. Meth. Phys. Res.
B72: 104–110.
Schofield, R. M. S., and H. W. Lefevre. 1993. Analysis
of unsectioned specimens: 2D and tomographic PIXE
with STIM. Nucl. Instrum. Meth. Phys. Res. B77:
217–224.
Schofield, R. M. S., M. H. Nesson, and K. A. Richardson.
2003. Zinc is incorporated into cuticular “tools”
after ecdysis: The time course of zinc accumulation
in “tools” and whole bodies of an ant and a scorpion.
J. Insect Physiol. 49: 31–44.
Schofield, R. M. S., M. H. Nesson, and K. A.Richardson.
2002. Tooth hardness increases with zinc-content in
mandibles of young adult leaf-cutter ants. Naturwissenschaften 89: 579–583.
Robert M. S. Schofield University of Oregon
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References Cited
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