NPH075.

fm Page 111 Tuesday, February 27, 2001 10:27 AM

Research

Intracellular calcium oxalate crystal structure in

Dracaena sanderiana
Blackwell Science Ltd

Svoboda V. Pennisi1, Dennis B. McConnell2, Laurie B. Gower3, Michael E. Kane2 and T. Lucansky3
1

University of Georgia, Horticulture Department, Coastal Plains Experimental Station, Tifton, GA 31793, USA; 2University of Florida, Environmental

Horticulture Department, 1519 Fifield Hall, Gainesville, FL 326110670, USA; 3University of Florida, Materials Science and Engineering Department, 210
Rhines Hall, Gainesville, FL 32611 6400; 3University of Florida, Botany Department, 3191 McCarty Hall, Gainesville, FL 32611 6400, USA

Summary
Author for correspondence:
Dennis B. McConnell
Tel: +1 352 392 7932
Fax: +1 352 392 3870
Email: DBM@GNV.IFAS.UFL.EDU
Received: 27 April 2000
Accepted: 7 November 2000
Florida Agricultural Experiment Station
journal series no. R-07536

The chemistry, crystallography and ultrastructure of intracellular calcium oxalate

deposits in the angiosperm, Dracaena sanderiana are reported here.
Crystalline deposits extracted from mature leaves and leaf primordia of D. sanderiana
were studied by scanning electron microscopy and X-ray powder diffractometry
techniques, and compared with X-ray standards for calcium monohydrate and calcium
oxalate dihydrate.
Intracellular calcium oxalate deposits were of two types; calcium oxalate monohydrate raphides or solitary calcium oxalate dihydrate crystals. Raphide-containing
cells exhibited lamellate sheaths around the chamber walls, mucilage-like materials
surrounding the developing crystal chambers, and paracrystalline bodies with closely
spaced subunits within the chambers. The intracellular calcium oxalate dihydrate
crystals usually displayed typical tetragonal-dipyramidal morphology, but development of some unusual crystal faces occasionally occurred.
Two intracellular hydrate forms of calcium oxalate (monohydrate and dihydrate)
exist in D. sanderiana. The elaboration of crystal vacuoles derived from rough endoplasmic reticulum and modified crystals with energetically unfavourable faces
suggest that precipitation of calcium oxalate dihydrate in D. sanderiana cells might
be biologically controlled.
Key words: Plant crystals, biomineralization, calcium oxalate, raphides, crystal
idioblast.
New Phytologist (2001) 150: 111120

Introduction
The most common calcium oxalate (CO) hydrates found in
plants are calcium oxalate monohydrate (COM) and calcium
oxalate dihydrate (COD). Angiosperms typically deposit CO
crystals inside cell vacuoles of highly specialized cells. The
crystal-containing cell is usually conspicuously larger than
surrounding cells and is termed crystal idioblast. Horner &
Wagner (1995) proposed two general systems based on the
presence or absence of membranes and associated subcellular
structures. System I was exemplified by druses in Capsicum
and Vitis, raphides in Psychotria, and crystal sand in Beta.
System I crystal idioblasts presented cytoplasmic spherosomes,
vacuolar organic paracrystalline bodies, membrane complexes,

New Phytologist (2001) 150: 111 120 www.newphytologist.com

plasmalemmasomes, and crystal chambers. The vacuolar

paracrystalline bodies exhibited subunits with large periodicity
and were linked to a membrane network, which formed the
crystal chambers. System I crystal idioblasts were observed
only in dicotyledonous species. System II was exemplified by
the monocotyledonous raphide idioblasts in Typha, Vanilla
and Yucca. System II lacked vacuolar membrane complexes,
and paracrystalline bodies displayed closely spaced subunits.
Mucilage-like material was present around developing crystal
chambers and lamellate sheaths were observed around chamber walls (Wattendorff, 1976; Horner & Wagner, 1995).
In reports where conclusive X-ray diffraction analysis was
used, the crystalline matter in plants was shown to be COM.
Reported occurrences of COD crystals are scarce, and conclusive

111

NPH075.fm Page 112 Tuesday, February 27, 2001 10:27 AM

112 Research

reports of their chemical identity are even scarcer. In most

instances, crystal morphology has been the only definitive
feature used in analysis. Confirmed reports on intracellular
COD in plants determined by X-ray diffraction include:
Capsicum annuum (Solanaceae), solitary prisms (Wagner, 1983)
and druses (Horner & Wagner, 1992); Begonia sp. (Begoniaceae),
solitary prisms (Horner & Zindler-Frank, 1981) and Begonia
maculata , B. manicata , B. metallica , solitary prisms and druses
(Al-Rais et al., 1971); Coleus sp. (Labiatae), solitary prisms and
druses (Al-Rais et al., 1971) and Beta vulgaris (Chenopodiaceae),
solitary prisms and cylindrics (Al-Rais et al., 1971); and, Echinomastus intertextus, Echinocactus horizonthalonius, Escobaria
tuberculosa (Cactaceae), druses (Rivera, 1973). Reports on intracellular COD in plants using crystal morphology as the only
criterion include: Telfairia sp., solitary prisms (Okoli & McEuen,
1986); Acacia senegal, solitary prisms (Parameswaran & Schultze,
1973); and, Aglaonema modestum, Hydrosome rivieri, solitary
prisms (Genua & Hillson, 1985).
In addition to periplasmic COM deposits (Pennisi et al.,
2000), D. sanderiana forms crystals in intracellular locations.
Their chemical, crystallographic, and ultrastructural aspects
are the focus of this paper.

Materials and Methods

Intracellular crystal extraction and processing
Individual raphides were extracted by pressing freshly cut
leaves of Dracaena sanderiana hort Sander ex M.T. Mast.
(Dracaenaceae) onto circular glass coverslips and glass slides.
Samples were analysed with a scanning electron microcope and
X-ray powder diffractometer as outlined below. Intracellular
deposits other than raphides were obtained from two sources,
mature and immature leaves. Pieces from mature leaves and
basal portions of leaf primordia (510 mm in length) were
placed in a maceration solution containing cellulase (1.0% w/v),
hemicellulase (1.0% w/v), and pectinase (0.1% w/v) (Protoplast
Isolation Enzyme Solution I, Sigma (Sigma-Aldrich Company,
St. Louis, MO, USA) ) for 24 h. Basal portions were used to
minimize contamination from cuticular periplasmic COM
crystals (Pennisi et al., 2000). These portions were cut under
an optical microscope equipped with polarizing optics to
observe the cuticular crystals. The maceration procedure
reduced all internal tissue to individual cells and the epidermis
to a long tube (due to the shape of monocotyledonous leaf
primordia). The epidermis was removed, and the cell suspension
was pipetted onto glass slides and circular glass coverslips. The
glass slides and circular glass coverslips were examined with an
optical microscope and any raphide contamination removed.
The suspension was flooded with water, causing protoplast
swelling and cell rupture. Intracellular crystals were freed and
settled to the bottom of the suspension. Excess water and
cellular debris were drawn off with filter paper. Three water
rinses were followed by three 100% ethanol rinses. Coverslips

were prepared for SEM, and glass slides were processed for
X-ray diffraction as outlined elsewhere (Pennisi et al., 2000).
Results were compared with American Society for Testing
Materials (ASTM) X-ray standards for calcium oxalate monohydrate (whewellite) and calcium oxalate dihydrate (weddellite).
ASTM data were obtained from the Joint Committee on Powder
Diffraction Standards ( JCPDS) International Centre for
Diffraction Data 1996.
Light and transmission electron microscopy
Procedures for light microscopy (LM) and TEM are described
in detail elsewhere (Pennisi et al., 2000).

Results
Calcium oxalate monohydrate raphides
D. sanderiana leaf primordia develop intracellular crystals,
each containing a centrally located bundle of numerous individual crystals termed raphides (Fig. 1a c). High birefringence
(Fig. 1b) and X-ray diffraction data (Table 1) confirmed that
the raphides were composed of COM. The raphides are 80
100 m long with sharp pointed ends (Fig. 1d) and irregular
edges (Fig. 1e). Raphide bundles contain 100150 individual
crystals (Fig. 1b). Ultrastructurally, the raphide idioblasts exhibit
several distinctive characteristics. Paracrystalline bodies with
closely spaced subunits were observed (Fig. 2a). Individual
raphides are located randomly in the cell vacuole and measured
approx. 1 m in transverse section. All raphides are embedded
in a mucilagenous matrix different from the surrounding
cytoplasm (Fig. 2b). Individual raphides are orientated randomly
lengthwise with respect to one another, with large spaces
between individual crystals (Fig. 2bc). The most striking
feature of D. sanderiana raphide bundles are the crystal chambers
(Fig. 2cf ). Each crystal is surrounded by a lamellate crystal
chamber that is not connected to neighbouring chambers and
is distinct from the mucilagenous matrix of the raphide
bundle. The chambers have double membrane walls, and looplike lamellate extensions along their wide ends (in transverse
section) (Fig. 2df ). The length of the lamellate extensions
ranges from 0.5 m to 1.6 m, and some extensions appeared
to end blindly without completing a full loop (Fig. 2f ). The
blind ends probably reflect the plane of sectioning. Some
sections show loop-like extensions connected to only one side
of the chamber wall (Fig. 2f ).
Calcium oxalate dihydrate crystals
Numerous tetragonal crystals are present in immature leaf
primordial cells observed under polarized light (Fig. 3a).
When mature leaf mesophyll cells are isolated by maceration
variously sized rod-like as well as some prismatic crystals are
evident (Fig. 3b). The rod-like deposits are small ( 45 m),

www.newphytologist.com New Phytologist (2001) 150: 111 120

NPH075.fm Page 113 Tuesday, February 27, 2001 10:27 AM

Research

Fig. 1 Light microscopy (LM) and scanning

electron microscopy (SEM) micrographs of
raphide idioblasts and isolated crystals in
immature Dracaena sanderiana leaves.
(a) A single raphide idioblast isolated from
macerated primordial tissue. (bc)
Photographs taken between crossed polars.
(b) Raphide bundles in the base of a leaf
primordium showing high birefringence
typical of calcium oxalate monohydrate
(COM). Bar, 50 m (c) Two isolated individual
raphide crystals recognized by their lengthwidth and pointed ends. The lower crystal is
orientated so that maximum brightness is
observed, while the top crystal is in partial
extinction position. Periplasmic cuticular
COM crystals also are present. Crystal
birefringence is a function of orientation in
polarized light. Bar, 10 m. (de) Single
isolated raphide with an irregular outline near
its end (arrow). (d) Bar, 20 m (e) Bar, 1 m.

Table 1 Comparison of American Society for Testing Minerals

(ASTM) data of calcium oxalate monohydrate and intracellular
raphides extracted from the mesophyll of Dracaena sanderiana
ASTM whewellitex
CaC2O4H2O

Dracaena sanderiana

D, y

I/I0z

D,

I/I0

*5.93w
5.79
*3.65
3.01
*2.97
2.92
2.84
2.49
2.36
2.08
1.98
1.85
1.82

100
30
70
10
45
10
10
18
30
14
10
6
6

5.93
5.79
3.66

2.88

2.49
2.32

1.96
1.84
1.82

100
25
54.8

43

10
12.8

8.4
2.8
2.8

ASTM data were obtained from Joint Committee on Powder

Diffraction Standards (JCPDS) International Centre for Diffraction
Data 1996. yD is the wavelength spacings in ngstroms. zI/Io is relative
intensity of diffraction response compared to the primary peak. wThe
three major peaks are indicated by an asterisk (*) in each analysis.

New Phytologist (2001) 150: 111 120 www.newphytologist.com

and some exhibit twinning (growth of two crystals joined by

a common plane) (Fig. 3cd). Their birefringence was lower
than COM crystals and their morphology differs from COM.
Both crystal morphology and X-ray diffraction data (Table 2)
confirmed that the intracellular crystals were COD. Leaf primordial cells contain vacuoles with somewhat angular outlines
(Fig. 3e f ), which closely match the prismatic shape of COD
crystals (Fig. 3f ). The vacuoles appear to be connected to flat
sheets of rough ER (RER) (Fig. 3e).
SEM revealed that the prismatic crystals vary in size from
2 to 4.5 m (Fig. 4). These crystals are characterized by a fourfold axis of symmetry (Fig. 4ac). Various faces are expressed in
these prismatic crystals. The {101} faces enclose a tetragonal
pyramid at both ends of the crystals, and the {100} faces
enclose the parallel sides of the tetragonal prism. The final crystal
form is a combination of tetragonal bipyramid and tetragonal
prism (Fig. 4b). Rotational twins (the common plane between
the crystals is derived by rotation) are common (Fig. 4c). Some
crystals show a high degree of defects, including holes on the
pyramidal surface (Fig. 4c). Some had {111} and {110} crystal
faces (Fig. 4de), which differ from typical COD crystals
including the presence of pinacoid plane {001} (Fig. 4dh).
Some rounded crystal corners are also evident (Fig. 4gh).

113

NPH075.fm Page 114 Tuesday, February 27, 2001 10:27 AM

114 Research

www.newphytologist.com New Phytologist (2001) 150: 111 120

NPH075.fm Page 115 Tuesday, February 27, 2001 10:27 AM

Research

Fig. 3 Light microscopy (LM) of calcium

oxalate dihydrate (COD) crystals, and
transmission electron microscopy (TEM)
micrographs of leaf mesophyll cells of
Dracaena sanderiana. (a) This cell in the leaf
primordium is completely filled with prismatic
crystals (arrow) of low birefringence.
(b) Isolated mature cell with numerous
intracellular rod-like crystals (right pointing
arrows) and a single bipyramid crystal (left
pointing arrow). Bars, 10 m (cd) Isolated
rod-like crystals with cross-polarized light.
In (d), a first order red -plate gives a
magenta-coloured background. The dark
rod-like crystals and one of the crystals
forming a cross (indicates twinning, see
Fig. 5g) are in the extinct ion position (arrow).
Bars, 5 m (ef) Vacuoles with pronounced
angular outlines (stars) in epidermal (e) and
mesophyll (f) cells. CW, cell wall. Vacuolar
membranes are continuous with two profiles
of RER (arrows). Bars, 1 m. CW indicates the
position of the cell wall in the figure.

Fig. 2 Transmission electron microscopy (TEM) micrographs of raphide idioblasts in Dracaena sanderiana leaves. Abbreviations: CW, cell wall;
CYT, cytoplasm; PB, paracrystalline body; RH, raphide hole; MB, mucilagenous body, matrix; V, vacuole. (a) Paracrystalline body with closely
spaced subunits. (b) In ultrathin sections crystals are not visible since they are not infiltrated by the resin and fall out during sectioning; however,
their original locations remain visible as white holes. The boundary between the cytoplasm and mucilagenous body is arrowed. (c) Portion of
raphide bundle in a developing leaf cell. Note the larger spaces between the individual raphides compared with (f), and the irregular orientation
of crystals with respect to each other. The loop-like crystal chamber extensions do not appear to be connected with each other, and do not
extend to the edge of the mucilagenous body (down arrow). Note also the blind end of the crystal chamber extension (up arrow). (d) The edges
of the chamber walls can be discerned (right pointing and down black arrows) and in some places appear to connect to the dark extensions at
the corners as well as some of the sidewalls (left pointing black arrow). Some crystal chamber extensions are symmetrical (up white arrow), while
other are asymmetrical (left pointing white arrow). Bar, 1 m (e) Joining point of the lamellate extensions (arrow). Bar, 100 nm (f) Two blind
ends (black arrows) of the loop-like extensions. Within this section one lamellate part of the loop appears not connected to the other lamellate
part (white arrow). Bar, 100 nm. Abbreviations used in the figure are CYT, cytoplasm; RH, raphide hole; and MB, mucilagenous body, matrix.

New Phytologist (2001) 150: 111 120 www.newphytologist.com

115

NPH075.fm Page 116 Tuesday, February 27, 2001 10:27 AM

116 Research
Table 2 Comparison of American Society for Testing Minerals
(ASTM) data of calcium oxalate dihydrate and intracellular crystals
extracted from the mesophyll of Dracaena sanderiana
ASTM weddellitex
CaC2O42H2O

Dracaena sanderiana

D, y

I/I0z

D,

I/I0

*6.18w
*4.42
*3.78
2.41
2.24
1.90

100
30
65
16
25
16

6.21
5.99
3.72
2.41
2.25

100
25
16.5
20.3
13.9

x
ASTM data were obtained from Joint Committee on Powder
Diffraction Standards (JCPDS) International Centre for Diffraction
Data 1996. yD is the wavelength spacings in ngstroms. zI/Io is relative
intensity of diffraction response compared to the primary peak.
w
The three major peaks are indicated by an asterisk (*) in each
analysis.

Discussion
Calcium oxalate monohydrate raphides
The raphides in D. sanderiana are composed of COM, which
is consistent with previous reports of raphides in D. fragrans
(Scurfield and Mitchell, 1973). Mature raphides bundles in
D. sanderiana exhibit characteristics typical of System II
crystal idioblasts as defined by Horner & Wagner (1995). This
system is exemplified by the monocotyledonous raphide idioblasts
in Typha, Vanilla and Yucca, and typified by lamellate sheaths
around the chamber walls, mucilage-like material surrounding
the developing crystal chambers, and paracrystalline bodies
with closely spaced subunits (Horner & Whitmoyer, 1972;
Wattendorff, 1976; Tilton & Horner, 1980). The loop-like
extensions of the crystal chambers in D. sanderiana are very similar
to chamber wall extensions of Agave raphides (Wattendorff,
1976). However, in Agave these structures are larger, display
multiple lamellae and form symmetrical, closed loops, while
in D. sanderiana some of the raphide chamber extensions are
single lamellae, and are less symmetrically orientated than the
loop-like extensions in Agave raphide chambers. Unlike the
crystal lamellae in Typha, which are continuous with lamellae
from neighbouring crystals (Horner et al., 1981), the chamber

lamellae in D. sanderiana do not appear to anastomose with

other crystal chambers and do not show any discernible
continuity with the vacuolar membrane (tonoplast). The
paracrystalline body (Fig. 3a) is an enigmatic structure, which
is rarely observed, and no satisfactory explanation of its nature
and function has been determined (Barnabas & Arnott, 1990).
It has been hypothesized as a raphide precursor (Horner &
Wagner, 1995).
Development of raphide crystal idioblasts has been extensively documented (Franceschi & Horner, 1980). Theories
concerning the development of these crystal morphologies are
numerous, and some theories have implicated macromolecules
(i.e. proteins and complex polysaccharides) (Webb et al., 1995;
Webb, 1999). Crystal chambers may act as molds and control
both the shape and size of the crystals within them (Arnott,
1976). A wide variety of additives have altered COM morphology in vitro and produced crystals resembling some of those
found in plants (Cody & Horner, 1984; Cody & Cody, 1987;
Stevens et al., 1999). Arnotts suggestion that the crystal chambers may act as molds controlling the crystal shape has been
extended by one hypothesis of how macromolecules (acidic
proteins) can affect the mineral phase via a polymer-induced
liquid precursor (PILP) process (Gower & Odom, 2000).
Nonequilibrium crystal morphologies were generated in a
solution crystallization of calcium carbonates in the presence
of polyaspartic acid. This strongly acidic polypeptide induced
a liquid phase separation, in which droplets of a liquid precursor to the mineral accumulated in the form of mineral
films and coatings. As the precursor is a liquid, it can fill a space,
and the final mineral retains the shape of the precursor molded
to form the unusual morphologies (Gower & Odom, 2000).
The PILP process has recently been demonstrated in experiments with CO (Malpass & Gower, 1999).
Calcium Oxalate Dihydrate crystals
Intracellular COD crystals in D. sanderiana exhibit two types
of morphology. One is typical of the tetragonal-bipyramidal
class, with expression of {101} faces enclosing two tetragonal
pyramids at both crystal ends. The other crystal morphology
is atypical of the tetragonal class. Development of some
unexpected {100} faces enclosing the tetragonal prism was
observed. This crystal form (combination of a tetragonal
dipyramid and a tetragonal prism) has been documented in
Begonia (Horner & Zindler-Frank, 1981) and Capsicum

Fig. 4 Scanning electron microscopy (SEM) micrographs of intracellular crystals isolated from leaf primordia of Dracaena sanderiana showing
typical (ac) and atypical (dh) calcium oxalate dihydrate (COD) morphology. (a) This COD crystal has one four-fold axis of symmetry resulting
in tetragonal pyramids at both crystal ends. Arrow indicates the plane (100) enclosing a prism. (b) This twinned COD is a rotational combination,
showing both the tetragonal bipyramid (black arrows) and the tetragonal prism (white arrow). (c) An interpenetrant twinned crystal shows a
high degree of defects, including holes on the pyramidal surface (arrow). (df) Crystals display {111} faces (black stars) and {110} faces, which
are inconsistent with the typical bipyramidal COD morphology (compare with (a)). The pinacoid {001} is present. (g) The {101} faces (arrows)
are small compared to the typical COD morphology (compare with (a)). The crystals also exhibit somewhat rounded corners (white stars). Bars,
1 m.

www.newphytologist.com New Phytologist (2001) 150: 111 120

NPH075.fm Page 117 Tuesday, February 27, 2001 10:27 AM

Research

New Phytologist (2001) 150: 111 120 www.newphytologist.com

117

NPH075.fm Page 118 Tuesday, February 27, 2001 10:27 AM

118 Research

Fig. 5 Schematic illustration showing

hypothetical growth modifications of crystal
faces in intracellular calcium oxalate dihydrate
(COD) crystals in Dracaena sanderiana.
(a) Typical tetragonal COD bipyramids grown
in vitro with no additives. (b) Development
of tetragonal prisms {100} in intracellular
COD crystals in D. sanderiana (compare with
Fig. 3b). (c) Development of {001} pinacoids.
(d) Growth of the {001} pinacoid planes is
stabilized resulting in large {001} faces and
truncated {101} faces (compare with Fig. 4d).
(e) Additional habit modifications with
development of {111} and {110} faces
(compare with Fig. 4). (f) Same as (e) viewed
down the c axis. (g) Twinning of COD crystals
resulting in appearance of crosses (compare
with Fig. 4cd). Note the large area of the
{100} faces compared to {101} faces.

(Wagner, 1983; Horner & Wagner, 1992). To the best of our

knowledge, prior to this study, detailed identification of COD
crystal faces has not been attempted nor have the unusual
crystal morphologies in D. sanderiana COD crystals resulting
from expression of {001}, {111}, and {110} faces been
previously documented.
Fig. 5 represents a hypothetical sequence, which helps
explain the changes in crystal COD morphology, which were
observed in D. sanderiana intracellular crystals. All variations
observed in D. sanderiana COD crystals (Fig. 4bh) can be
derived from the typical morphology in (a) by the development of some additional faces (bg). The typical tetragonal
bipyramid of synthetic COD crystals (a) is achieved by development of {101} faces. However, when the {100} face develops,

a new crystal shape (b) emerges (compare with Fig. 4a).

Furthermore, development of the terminal pinacoid plane
{100} and the {111} planes (f ) results in yet another crystal
shape similar to those in Fig. 4(d,f,h). Development of a set of
{110} planes (e) results in a shape similar to the crystals in
Fig. 4(g h). The rod-like crystals observed with light microscopy (Fig. 3bd) are similar to the twinned COD crystals
illustrated by Frey-Wyssling (1981). Their morphology could
be explained by the relative expression of {100} faces compared to the typical {101} faces (g ). The planes observed in D.
sanderiana COD crystals are not consistent with the synthetic
COD morphology. Instead, they are unstable, high-energy
crystal faces, commonly developed in crystals precipitated in
vitro in the presence of various solution constituents (Addadi

www.newphytologist.com New Phytologist (2001) 150: 111 120

NPH075.fm Page 119 Tuesday, February 27, 2001 10:27 AM

Research

& Weiner, 1989). Development of less stable crystal faces can

occur due to interactions with solution constituents (i.e.
acidic macromolecules, including proteins extracted from
biominerals). Crystal habit modifications of COM have been
achieved by the growth of crystals in the presence of citrate
and phosphocitrate (Sikes & Wierzbicki, 1996). In addition,
alterations of the less common COD crystals have been
brought about by growth of crystals in solutions with ,dicarboxylic acids (Stevens et al., 1999). Suberic acid was also
suggested to have a structural motif, which matched the {110}
face of COD thus stabilizing expression of these high-energy
planes.
The intracellular vacuoles observed in D. sanderiana with
their angular outlines may have contained the COD crystals
(Fig. 3ef ). Crystals are not preserved in ultrastructural preparations, but one indication of their presence is the spaces they
previously occupied. If association between the vacuoles and
the crystals is accepted, several implications follow. Since the
vacuoles were attached to parent RER membranes, the COD
precipitation inside D. sanderiana cells may be controlled by
the elaboration of RER crystal vacuoles. Further support of
this hypothesis is that crystal morphology is modified by the
development of unstable crystal faces, presumably through
interactions with impurities in the vacuolar medium. The
defects present in some crystals (holes and cracks on the crystal
surface, Fig. 4c) are also typical of biogenically precipitated
minerals, as are rounded crystal corners (Fig. 4gh), which are
attributed to non-specific interactions with impurities (Addadi
& Weiner, 1989). Our extraction procedure involved enzymatic
tissue digestion to release the crystals, therefore these defects
were not likely artifacts. Space delineation is one of the most
distinctive features of biologically controlled biomineralization, and lipid membranes are the most common way of sealing off a predetermined compartment (Lowenstam & Weiner,
1989). This sealing off process allows selective uptake of ions
and provides a means to control concentration and composition of the initial solution from which the mineral forms.
Our study of crystalline deposits in D. sanderiana leads to
the following conclusions. There is definitive evidence for two
hydrate forms of CO, COM and COD, in the same plant
species. Three distinctive crystal morphologies exist, periplasmic
COM crystals, intracellular COM raphides, and intracellular
COD crystals. The factors controlling CO phase and morphology
in D. sanderiana remain to be determined, but the constancy
of CO forms in tissue-specific locations seems to indicate
a highly developed phytosystem for biologically controlled
biomineralization. D. sanderiana has proven to be an excellent
example of a phytosystem with highly controlled deposition
of biogenic CO hydrates.

Acknowledgements
The authors thank Drs Karen Koch and Bart Schutzman for
review of the manuscript and instructive criticism.

New Phytologist (2001) 150: 111 120 www.newphytologist.com

References
Addadi L, Weiner S. 1989. Stereochemical and structural relations between
macromolecules and crystals in biomineralization. In: Mann S, Webb J,
Williams RJP, eds. Biomineralization chemical and biochemical perspectives.
New York, USA: VCH Publishers, 135156.
Al-Rais AH, Myers A, Watson L. 1971. The isolation and properties of
oxalate crystals from plants. Annals of Botany 35: 1213 1218.
Arnott HJ. 1976. Calcification in higher plants. In: Watabe N, Wilbur KM,
eds. The mechanisms of mineralization in the invertebrates and plants.
Columbia, USA: University of South Carolina Press, 55 78.
Barnabas AD, Arnott HJ. 1990. Calcium oxalate crystal formation in
the bean (Phaseolus vulgaris L.) seed coat. Botanical Gazette 151: 331
341.
Cody AM, Cody RD. 1987. Contact and penetration twinning of calcium
oxalate monohydrate (CaC2O4H2O). Journal of crystal Growth 83:
485498.
Cody AM, Horner HT. 1984. Crystallographic analysis of crystal images in
scanning electron micrographs and their application to phytocrystalline
studies. Scanning Electron Microscopy 3: 14511460.
Franceschi VR, Horner HT Jr. 1980. Calcium oxalate crystals in plants.
Botanical Review 46: 361427.
Frey-Wyssling A. 1981. Crystallography of the two hydrates of crystalline
calcium oxalate crystals in plants. American Journal of Botany 68: 130141.
Genua JM, Hillson CJ. 1985. The occurrence, type, and location of calcium
oxalate crystals in the leaves of fourteen species of Araceae. Annals of Botany
56: 351361.
Gower LA, Odom DJ. 2000. Deposition of calcium carbonate films by a
polymer-induced liquid precursor process. Journal of Crystal Growth 210:
719734.
Horner HT, Kausch AP, Wagner BL. 1981. Growth and change of
raphide and druse calcium oxalate crystals as a function of intracellular
development in Typha angustifolia L. (Typhaceae) and Capsisum annuum
L. (Solanaceae). Scanning Electron Microscopy 3: 251262.
Horner HT, Wagner BL. 1992. Association of four different calcium crystals
in the anther connective tissue and hypodermal stomium of Capsicum
annuum (Solanaceae) during microsporogenesis. American Journal of
Botany 79: 531541.
Horner HT, Wagner BL. 1995. Calcium oxalate formation in higher plants.
In: Khan SR, ed. Calcium Oxalate in biological systems. Boca Raton, FL,
USA: CRC Press, 5375.
Horner Jr HT, Whitmoyer RE. 1972. Raphide crystal cell development
in leaves of Psychotria punctata (Rubiaceae). Journal of Cell Science 2:
339355.
Horner HT, Zindler-Frank E. 1981. Histochemical, spectroscopic, and
X-ray diffraction identifications of the two hydration forms of calcium
oxalate crystals in three legumes and Begonia. Canadian Journal of
Botany 60: 10211027.
Lowenstam HA, Weiner S. 1989. On Biomineralization. New York, USA:
Oxford University Press.
Malpass CA, Gower LA. 1999. Can a polymer-induced liquid precursor
(PILP) process be elicited in calcium oxalates? [Abs.] 1999. FASEB
Summer conference on Calcium Oxalate in biological systems, Copper
Mountain, CO, USA.
Okoli BE, McEuen AR. 1986. Calcium-containing crystals in Telfairia
Hooker (Cucurbitaceae). New Phytolologist 102: 199 207.
Parameswaran N, Schultze R. 1973. Fine structure of chambered
crystalliferous cells in the bark of Acacia senegal. Z. pflanzenphysiol. bd.
71: 9093.
Pennisi SV, McConnell DB, Gower LB, Kane ME, Lucansky T. 2000.
Periplasmic cuticular calcium oxalate crystal deposition in Pracaena
sanderiana. New Phytologist 149: 209218.
Rivera ER. 1973. Echinomastus intertextus: an ultrastructural, physiological
and biochemical study. PhD Dissertation. University of Texas at Austin

119

NPH075.fm Page 120 Tuesday, February 27, 2001 10:27 AM

120 Research
(Libr. Congr. Card, Mic. 74 5315). University Microfilms, Ann Arbor,
MI (Diss. Abstract. Int. 34: 09-B).
Scurfield FLS, Mitchell AJ. 1973. Crystals in woody stems. Botanical
Journal of the Linnean Society. 66: 277289.
Sikes CS, Wierzbicki A. 1996. Polyamino acids as antiscalants,
dispersants, antifreezes, and absorbent gelling materials. In: Mann S,
ed. Biomimetic materials chemistry. New York, USA: VCH Publishers,
249 278.
Stevens C, Heywood BR, Johnson V. 1999. Crystallization of calcium
oxalate in the presence of ,-dicarboxylic acids. [Abs.] 1999. FASEB
Summer conference on calcium oxalate in biological systems. Copper
Mountain, CO, USA.
Tilton VR, Horner HT. 1980. Calcium oxalate raphide crystals and

crystalliferous idioblasts in the carpels of Ornithogalum caudatum. Annals

of Botany 46: 533539.
Wagner BL. 1983. Genesis of the vacuolar apparatus responsible for the
druse formation in Capsicum annuum L. (Solanaceae) anthers. Scanning
Electron Microscopy 2: 905912.
Wattendorff J. 1976. A third type of raphide crystal in the Plant Kingdom:
six-sided raphides with laminated sheaths in Agave americana L. Planta
130: 303311.
Webb MA. 1999. Cell-mediated crystallization of calcium oxalate in plants.
Plant Cell 11: 751761.
Webb MA, Cavaletto JM, Carpita NC, Lopez LE, Arnott HT. 1995. The
intravacuolar organic matrix associated with calcium oxalate crystals in
leaves of Vitis. Plant Journal 7: 633648.

www.newphytologist.com New Phytologist (2001) 150: 111 120