J. Phycol. 38, 1–8 (2002)
CARRAGEENANS BIOSYNTHESIZED BY CARPOSPOROPHYTES OF
RED SEAWEEDS GIGARTINA SKOTTSBERGII (GIGARTINACEAE) AND
GYMNOGONGRUS TORULOSUS (PHYLLOPHORACEAE) 1
José Manuel Estevez, Marina Ciancia, and Alberto Saúl Cerezo2,3
Departamento de Química Orgánica (CIHIDECAR-CONICET), Facultad de Ciencias Exactas y Naturales,
Universidad de Buenos Aires, Ciudad Universitaria - Pabellón 2, 1428 Buenos Aires, Argentina
-d-galactose or 3,6-anhydro--d-galactose units, which
are usually sulfated at specific positions. Carrageenans
have been classified in different families according to
sulfation on the 3-linked -d-galactose units (Percival
1978, McCandless and Craigie 1979, Greer and Yaphe
1984).
The life cycle of the Florideophyceae consists of three
phases: a haploid sexual phase (the gametophyte), a
parasitic diploid phase that develops directly on the female thallus (the carposporophyte), and a free living
diploid phase (the tetrasporophyte). In some cases, including the seaweeds studied here, the carposporophyte
is surrounded and protected by gametophytic tissue
(pericarp) and the whole structure is called the cystocarp. The cystocarpic plant comprises the female gametophyte and the developed cystocarp.
It has been found that in seaweeds belonging to the
Gigartinaceae and Phyllophoraceae, gametophytes and
tetrasporophytes biosynthesize different carrageenans
(Chen et al. 1973, McCandless et al. 1973, 1982, 1983,
Pickmere et al. 1973, Waaland 1975, Craigie 1990).
Carrageenans extracted from gametophytes belong to
the kappa family (the 3-linked -d-galactose units are
sulfated on C-4), whereas those obtained from tetrasporophytes are mainly lambda-carrageenans (the
3-linked -d-galactose units are sulfated on C-2) (Matulewicz et al. 1989, Stortz and Cerezo 1993). It has
been reported (Gordon-Mills and McCandless 1975),
on the basis of results from histochemical techniques,
that the carposporophytes biosynthesized lambda-carrageenans, in agreement with the diploid character of
the phase. This difference between carrageenans biosynthesized by the alternating life stages has not been
observed in seaweeds from other families (DiNinno
and McCandless 1978, Bert et al. 1989).
The different chemical structure of these two families of carrageenans gives rise to different rheological
properties: carrageenans of the kappa family form
gels at low concentrations of potassium chloride or
can be converted into gelling carrageenans by treatment with alkali, whereas lambda-carrageenans do
not gell at low concentrations of potassium chloride
even after alkaline treatment but give viscous solutions. As a consequence, they have different industrial
applications (Glicksman 1983) and, possibly, different
biological functions. The aim of this article is to determine unequivocally the type of carrageenans biosynthesized by the carposporophytes of Gigartina skottsbergii Setchell and Gardner and Gymnogongrus torulosus
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Carrageenans biosynthesized by gametophytic
and tetrasporic plants of seaweeds belonging to the
Gigartinaceae and Phyllophoraceae are different:
gametophytes produce carrageenans of the kappa
family, whereas lambda-carrageenans are extracted
from tetrasporophytes. For Gigartina skottsbergii Setchell and Gardner and Gymnogongrus torulosus Hooker
et Harvey, mature cystocarps were isolated and carrageenans were extracted. Structural determination
by methylation analysis, Fourier transform infrared
spectroscopy, and 13C-NMR spectroscopy showed
that they were kappa/iota-carrageenans. For the extract obtained from cystocarps of Gigartina skottsbergii with water at room temperature, the ratio
kappa:iota was 1:0.30 and at 90 C was 1:0.43; significant amounts of precursors were also present. The
extract obtained from cystocarps of Gymnogongrus
torulosus at 90 C showed prevalence of iota-carrageenans (ratio kappa:iota 1:1.21). These extracts are
similar to the polysaccharides produced by gametophytes of these seaweeds. For Gigartina skottsbergii, it
was possible to separate the pericarpic tissue from
the carposporophyte. Thus, they were extracted separately, and the carrageenans isolated were studied
as described before, obtaining similar conclusions.
These results clearly show that whereas the carposporophytes are located inside the cystocarp, they
produce carrageenans of the kappa family despite of
being diploid cells.
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Key index words: carposporophyte; carrageenan structure; cystocarps; gametophyte; Gigartina skottsbergii
(Gigartinaceae); Gymnogongrus torulosus (Phyllophoraceae); kappa/iota-carrageenan; lambda-carrageenan
Abbreviations: FT-IR, Fourier transform infrared;
GC, gas–liquid chromatography; GC-MS, gas–liquid
chromatography mass spectrometry
Carrageenans are sulfated galactans extracted from
red seaweeds of certain families of the order Gigartinales. They are unbranched polysaccharides comprised
of alternating 3-linked -d-galactose and 4-linked
1Received
11 June 2001. Accepted 4 January 2002.
for correspondence: e-mail [email protected].
Member of the National Research Council of Argentina
(CONICET).
2Author
3Research
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JOSÉ MANUEL ESTEVEZ ET AL.
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materials and methods
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Material. Cystocarpic plants of Gigartina skottsbergii were collected in 1995 in Bahía Camarones (44 42 S, 65 40 W)
Chubut province, Argentina. Cystocarpic plants of Gymnogongrus torulosus were collected in March 1998 in Cabo Corrientes
(38 03 S, 57 31 W), Mar del Plata, Buenos Aires province, Argentina. This material was verified by S. Fredericq (personal
communication) based on molecular sequence analysis of the
chloroplast gene rcbL. Samples were deposited in the Museo
Bernardino Rivadavia (B.A.), Buenos Aires, Argentina (collection numbers 28272 and 35707, respectively).
General. The total carbohydrate content was measured by the
phenol-sulfuric acid method (Dubois et al. 1956). Monosaccharide composition was determined by gas–liquid chromatography
(GC) of the alditol acetates obtained by reductive hydrolysis and
acetylation of the samples (Stevenson and Furneaux 1991). The
ratio d:l-galactose was estimated by the method of Cases et al.
(1995). Sulfate was determined by the method of Dodgson and
Price (1962).
Anatomical observations were made on dry material rehydrated in seawater and material preserved in 4% formalin seawater (Fig. 1). Reproductive structures were photographed using a Nikon AFX-II macroscope (Nikon, Tokyo, Japan), and
photomicrographs were taken on a Zeiss Axioplan microscope
(Zeiss, Oberkochen, Germany).
Isolation of reproductive structures. Sections of the thallus were
cut manually with a single-edged razor blade, and cystocarps
were separated from the vegetative tissues by dissection (Fig. 2).
Cystocarps of Gigartina skottsbergii were hydrated in a minimum
amount of water to facilitate the separation of the pericarp (p)
from the inner carposporophyte (c), which were lyophilized
and milled separately. Pericarps (gametophytic tissue) were
separated from the carposporophytes with the aid of a fine forceps. For Gymnogongrus torulosus, separation of the inner carposporophyte from the pericarp was not possible due to the
small size and arrangement of the carposporophyte.
Extraction. Cystocarps (1.85 g) from Gigartina skottsbergii,
previously milled, were extracted with H 2O (250 mL) at room
temperature with mechanical stirring for 24 h (Fig. 2). The residue was removed by centrifugation and the supernatant
poured into 3 volumes of iso-PrOH, where the polysaccharide
precipitated as long fibers. The liquors were decanted and the
product pressed in filter paper and dried by solvent exchange
(EtOH and Et2O) and finally in vacuo (Fig. 2, RI). The residue
(180 mg) was resuspended in H2O (9 mL) and extracted at 90
C with mechanical stirring for 4–5 h. The extract was treated as
described above (Fig. 2, HI). Pericarpic tissue (150 mg) and
carposporophytes (150 mg) were extracted separately with H 2O
at room temperature and at 90 C, as described above. For cystocarps of Gymnogongrus torulosus (103 mg), only the hot water
extraction (5 mL) was carried out (Fig. 2, HY).
Purification of HI, HIp, and HIc. The sample (10.5–24 mg)
was solubilized in 0.1 M phosphate buffer (pH 6.9, 5 mL), and
-amylase, type VIII-A from Barley Malt (Sigma, St. Louis, MO,
USA) (2–4 mg), was added. The solution was kept 24 h at room
temperature and then dialyzed against water (molecular weight
cutoff 1000) and freeze-dried.
Analysis of the residue. The fibrillar material (residue after
extraction of HIc, Fig. 2) (3 mg) was dissolved in 100% trifluoroacetic acid (37 C, 1 h) followed by dilution of the acid to
80%, heating at 100 C for 1 h, and further dilution to 2 M to
achieve the regular hydrolysis procedure (2 M trifluoroacetic
acid, for 90 min at 120 C; Morrison 1988). Hydrolyzate was derivatized to the corresponding alditol acetates. 3,6-Anhydrogalactose was analyzed directly on the residue by the resorcinol
method (Yaphe and Arsenault 1965).
GC. GC of the alditol acetates and those of the partially methylated alditol and aldononitrile acetates were carried out on a
Hewlett Packard 5890A gas–liquid chromatograph (Hewlett
Packard, Avondale, PA, USA) equipped with a flame ionization
detector and fitted with a fused silica column (0.25 mm i.d. 30 m) WCOT-coated with a 0.20-m film of SP-2330 (Supelco,
Bellefonte, PA, USA). Chromatography was carried out as described before (Estevez et al. 2000).
GC mass spectroscopy (MS). GC-MS was performed on a Shimadzu GC-17A gas–liquid chromatograph equipped the SP2330 (see above) interfaced to a GCMS-QP 5050A mass spectrometer (Shimadzu, Kyoto, Japan) working at 70 eV. Helium
was used as carrier gas.
Fourier transform infrared (FT-IR) spectroscopy. FT-IR spectra
were recorded from 4000 to 250 cm1 with a 510P Nicolet FT-IR
spectrophotometer (ThermoNicolet, Madison, WI, USA), using
films prepared by drying aqueous solutions of the polysaccharides, and 32–64 scans were taken with a resolution of 2–4 cm 1.
Methylation analysis. The polysaccharide (3–6 mg) was converted into the corresponding triethylammonium salt (Stevenson and Furneaux 1991) and methylated according to Ciucanu
and Kerek (1984) using finely powdered NaOH as base. The
methylated samples were derivatized to the alditol acetates as
described for the polysaccharides (Stevenson and Furneaux
1991). A portion of methylated sample was hydrolyzed with trifluoroacetic acid 2 M for 2 h at 120 C, and the partially methylated sugars were converted into the corresponding aldononitrile acetates (Stortz et al. 1982).
13C-NMR spectroscopy. The sample (20–30 mg) was dissolved
in H2O:D2O 1:1 solutions (1 mL), agitated 24 h at room temperature, and centrifuged. Proton decoupled 125-MHz 13CNMR spectra were recorded on a Bruker AM500 (Bruker Instruments, Billerica, MA, USA) at room temperature, with external reference of tetramethylsilane. The parameters were as
follows: pulse angle 51.4 degrees, acquisition time 0.56 s, relaxation delay 0.6 s, spectral width 29.4 kHz, and scans 19,000–
34,000. Chemical shifts were referenced to internal acetone
( 216.2 and 31.1).
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Hooker et Harvey by chemical and spectroscopic
methods.
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results
Thalli of Gigartina skottsbergii present numerous cystocarps as papillae jutting out from the surface of the
female gametophyte. Figure 1A shows a general aspect of the papillae with cystocarpic structures; Figure
1B shows pericarpic tissue and the inner carposporophyte, mainly composed of carposporangia; and Figure 1C shows the carpospororangia in detail. The cystocarps were isolated by excising them by hand and
then milling and extracting them with water at room
temperature to give extract RI. The residue was further extracted at 90 C to give extract HI (Fig. 2).
The general appearance of the branched thalli of
Gymnogongrus torulosus is presented in Figure 1E. Cystocarps are developed on terminal branches (Fig. 1F),
with small colored carposporangia (Fig. 1G). Extract
HY was obtained from the cystocarps of Gymnogongrus
torulosus by extraction at 90 C (Fig. 2). Extraction
conditions for each seaweed were chosen, taking into
account conditions used to obtain the extracts from
gametophytic samples in which the lambda structures
had been previously detected (Ciancia et al. 1993b,
1997, Estevez et al. 2001).
Yields and analyses of the extracts are shown in Table
1. For RI and HY, galactose was the major sugar component, but the percentages of 3,6-anhydrogalactose were
also important. For HI, the initial analysis showed that
the major sugar component was glucose (55.1%), although important percentages of galactose (30.4%)
and 3,6-anhydrogalactose (9.6%) and minor quantities
Q1
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CARRAGEENANS OF CARPOSPOROPHYTES
Fig. 1. (A–D) Morphology of the cystocarpic structures of Gigartina skottsbergii. (A) General aspect of mature cystocarps. Scale
bar, 300 m. (B) Light microscopy of the pericarp and the inner carposporophyte. Scale bar, 20 m. (C) Carposporangia in detail.
Scale bar, 20 m. (D) Isolated carposporophyte before carrageenan extraction. Scale bar, 300 m. (E–G) Morphology of the cystocarpic plants of Gymnogongrus torulosus. (E) General aspect of a thallus. Scale bar, 2 mm. (F) Cystocarps. Scale bar, 2 mm. (G) Inner
carposporophyte. Scale bar, 20 m.
of 3-O-methyl- and 6-O-methyl-galactose and xylose were
also detected. The high percentage of glucose arose
mainly from floridian starch, as confirmed by methylation analysis of HI (17.7% of 2,3,6-tri-O-methylglucose,
see later). Thus, the sample was treated with -amylase
and the percentage of glucose fell to 21.3%. Small
amounts of l-galactose were detected by enantiomeric
analysis of RI, HI, and HY (Cases et al. 1995), suggesting
the presence of agarans and/or d/l-hybrid chains, as
had been previously found in the system of polysaccharides synthesized by cystocarpic plants (Ciancia et al.
1993a, 1997, Estevez et al. 2001) of these seaweeds.
FT-IR spectra of extracts RI and HY (Fig. 3) showed
absorptions at 932 cm1 corresponding to the 3,6-
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JOSÉ MANUEL ESTEVEZ ET AL.
Table 1. Yields and analyses of carrageenans obtained from
cystocarps, carposporophytes (c), and pericarpic tissue (p) of
Gigartina skottsbergii (I) and cystocarps of Gymnogongrus torulosus (Y).
Fraction
1C b,c
RI
RIp
RIc
HI f,g,h
HIp f,h
HIc f,h
C1 k
HY
Yielda
SO3Na
(%)
(%)
Gal
6-O-Me-Gal
AnGal
Xyl
49.1
40.5
53.3
47.3
7.9i
12.5i
6.8i
31.1
43.7
32.1
33.5
41.4
35.4
40.8
34.6
49.4
29.9
34.4
64.0
68.6d
67.5
60.5
35.6j
53.3
59.6
60.3l
66.2m
—
—
tre
—
4.3
3.4
4.7
tr
—
32.0
25.2
30.2
23.9
30.1
32.5
23.6
38.3
30.2
1.8
2.2
1.6
4.7
1.0
1.3
1.0 14.6
8.7 21.3
6.8
4.0
8.4
3.7
1.4
tr
1.7
1.9
Monosaccharide composition (mol %)
Glc
a
Yields are given for 100 g of dry isolated algal structure.
Room temperature extract from cystocarpic plants of Gigartina
skottsbergii (included for comparison).
c 1.5% of 2-O-methylgalactose was also detected.
d 65.8% of d-galactose and 2.8% of l-galactose.
e Percentages lower than 1% are given as trace (tr).
f Treated with -amylase.
g 3.6% of 3-O-methylgalactose was also present.
h Small percentages of manose were also present.
i Calculated after deduction of the glucose content.
j 32.1% of d-galactose and 3.5% of l-galactose.
k Hot water extract from cystocarpic plants of Gymnogongrus
torulosus (included for comparison).
l 54.2% of d-galactose and 6.1% of l-galactose.
m 65.0% of d-galactose and 1.2% of l-galactose.
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b
Extraction procedure for the polysaccharides.
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Fig. 2.
indicating a higher complexity of this sample, it is also
basically a kappa/iota-carrageenan (ratio kappa:iota
1:0.43). In this sample, 1.5% of 4,6-di-O-methylgalactose was detected, indicating the presence of small
quantities of lambda structure.
HY is also a kappa/iota-carrageenan but with prevalence of the iota structure (ratio kappa:iota 1:1.21), as
was determined previously for carrageenans extracted
from gametophytes of Gymnogongrus torulosus (C1) (Estevez et al. 2001).
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anhydro ring (Stancioff and Stanley, 1969) and at 851
cm1 and 807 cm1, due to the axial sulfate group in
the 3-linked -d-galactose 4-sulfate units and to the axial sulfate group in the 4-linked 3,6-anhydro--d-galactose 2-sulfate residues, respectively. The last signal was
more important in the spectrum of HY. No absorption
around 830–820 cm1 (C2-equatorial sulfate group and
primary sulfate group) was observed (Rees 1961).
Methylation analyses of RI, HI, and HY showed they
are kappa/iota-carrageenans (Table 2). RI has a ratio
kappa:iota 1:0.30, and only trace amounts of 4,6-di-Omethylgalactose (that would correspond to -d-galactose 2-sulfate units in a lambda structure; see Discussion) were detected between the partially methylated
monosaccharides. The only unusual methylated unit
detected was 6-O-methylgalactose (3.4%), which would
correspond to disubstituted galactose units. The presence of this sugar is the only significant difference between these results and those obtained for the room
temperature extract from cystocarpic plants of Gigartina skottsbergii (Fig. 1C) (Matulewicz et al. 1989).
Methylation analysis of HI before and after treatment with -amylase gave identical results (with previous deduction of the amount of 2,3,6-tri-O-methylglucose in the former case). Although HI contains small
quantities of different methylated monosaccharides,
Fig. 3. FT-IR spectra RI, RIp, RIc, and HY. Arrows indicate
the mayor bands at 932 cm1, 851 cm1, and 807 cm1, typical
of kappa/iota-carrageenans.
5
CARRAGEENANS OF CARPOSPOROPHYTES
Table 2. Composition of partially methylated monosaccharides
produced by permethylation and hydrolysis of the carrageenans
obtained from Gigartina skottsbergii (1C, RI, and HI) and
Gymnogongrus torulosus (C1 and HY).
RI
RI p
2,3,6-Gal
2,4,6-Gal
2,6-Gal
4,6-Gal
2,4-Gal
6-Gal
3-Gal
2-Gal
2-AnGal
AnGal
tri
tr
1.6
52.2
tr
—
3.4
1.4
tr
31.9
9.5
1.7
7.4
46.6
tr
2.9
2.3
4.3
—
28.0
6.8
2.2
40.9
2.0
—
tr
2.6
2.2
37.0
13.1
RIC HI d,e HIpf
1.6
7.5
41.8
1.2
2.7
2.2
4.2
2.8
29.3
6.7
1.0
3.5
47.1
1.5
2.6
1.2
4.0
2.3
25.7
11.1
tr
3.5
48.9
n.d.j
1.0
tr
1.6
1.1
30.0
13.9
HIcf
C1 g
HY h
1.1
6.6
45.3
2.9
tr
1.6
3.3
tr
28.8
10.4
1.6
3.0
55.7
—
tr
tr
tr
tr
17.9
21.8
tr
2.5
40.3
tr
tr
—
tr
1.8
25.1
30.3
Fig. 4.
13C-NMR
spectra of (A) RI and (B) RIc.
1999). After treatment with -amylase, the glucose
content in HIc fell from 73.5% to 3.7%.
The FT-IR spectra of RIc and RIp were similar to
that of RI (Fig. 2). Methylation analyses of RIc and
RIp (Table 2) also gave results similar to those obtained from RI. The same conclusion was drawn from
HIp, HIc, and HI, where only minor differences were
observed.
The only difference between the 13C-NMR spectra
of RI (see above) and RIc (Fig. 4) is the presence in
the anomeric region of the signal at 100.8 ppm, corresponding to C-1 of an -(1→4)-glucan (floridian
starch); the other signals corresponding to this structure are also present, partially superimposed on those
of carrageenans (Seymour et al. 1976).
Analysis of the residue obtained after extraction of
the carposporophytes of Gigartina skottsbergii with hot
water (residue after extraction of HIc) indicated that
the total carbohydrate content was 53%. The major
sugar constituent was glucose, with minor amounts of
galactose, 3,6-anhydrogalactose, and mannose, suggesting that the remaining carrageenans belong also
to the kappa family, as has been shown for the skeletal
cell wall of cystocarpic Sarcothalia crispata (Bory) Leister (formerly Iridaea undulosa) (Flores et al. 1997).
For Gymnogongrus torulosus, it was not possible to separate the inner carposporophyte from the pericarpic
tissue due to their spatial arrangement (Fig. 1F) and
the small size of the carposporophytic tissue inside
the cystocarp (Fig. 1G).
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a Mol% of monosaccharide having methyl groups at the
positions indicated.
b Traces of nonmethylated galactose were detected for all the
samples.
c Room temperature extract from cystocarpic plants of
Gigartina skottsbergii (included for comparison).
d Percentages are expressed not considering 2,3,6-Glc
(17.7%, floridian starch).
e Small percentages of nonmethylated xylose and glucose
were also detected.
f Treated with -amylase before methylation.
g Hot water extract from cystocarpic plants of Gymnogongrus
torulosus (included for comparison).
h 1.7% of 2,3-Gal was also detected.
i Percentages lower than 1% are given as trace (tr).
j n.d., not determined.
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Monosaccharidea,b
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The 13C-NMR spectrum of RI (Fig. 4) is in agreement with results obtained from methylation analysis.
Signals corresponding to a kappa structure are the
most important (Usov and Shashkov 1985), although
those corresponding to iota (Usov and Shashkov 1985)
and nu diads are also obvious, the last structure being
the only precursor detected by this method (Ciancia et
al. 1993b). The absence of lambda-carrageenans was
inferred by the lack of signals in the ranges 105.0–103.2
and 67.0–62.5 ppm (signals at 103.9 and 64.5 ppm
would correspond to C-1 and C-4, respectively, of the
-d-galactose 2-sulfate units in a lambda-carrageenan
[Falshaw and Furneaux 1994, Stortz et al. 1994]).
The pericarps (p) were separated from the carposporophytes (c) of Gigartina skottsbergii and then extracted separately as described before to give RIp and
RIc at room temperature and HIp and HIc, when extracted at 90 C (Fig. 2). Figure 1D shows an isolated
carposporophyte.
Yields and analyses of these samples (Table 1)
showed similar results for the extracts obtained from
the pericarp and from the carposporphyte at room
temperature (RIp and RIc) and at 90 C (HIp and
HIc). The only significant difference is the increase in
the percentages of glucose in RIc and HIc (data not
shown); due to the higher floridian starch content
present in the carpospores, as was observed in studies
of the fine structure of carpospores of Gigartina skottsbergii (Buschmann et al. 1999), Ahnfeltiopsis durvillaei
(Bory) Silva et DeCrew (Phyllophoraceae) and A. furcellata (C. Agardh) Silva et DeCrew (Santelices et al.
discussion
The cystocarps of the Florideophyceae comprise
three compartments: the outer photosynthetic and
nonphotosynthetic tissues (haploid pericarpic tissues)
that, respectively, produce and process and store the
metabolites of photosynthesis and the parasitic developing carposporophyte (diploid tissue). Organic molecules are produced in the pericarp, transported and
6
JOSÉ MANUEL ESTEVEZ ET AL.
less 1978, Bert et al. 1989), indicated that the type of
carrageenan biosynthesized does not depend on the
cell ploidy.
Santelices et al. (1999) studied the development of
the carpospores in the coalescent process of some red
seaweeds, belonging to the Gigartinaceae and Phyllophoraceae in culture. When the naked carpospores
were released from the cystocarp, they settled on the
substratum and developed at first mitosis a new cell wall.
The spore cell wall had two well-defined components: a
thin inner layer surrounding each of the daughter
cells and a less defined outer layer surrounding the
entire sporeling. This would be the stage where the
cells start producing lambda-carrageenans.
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We are indebted to Dr. Suzanne Fredericq for the taxonomic
identification of Gymnogongrus torulosus and to Dr. Cecilia Rodriguez for the helpful discussion of the manuscript. Supported
by grants from CONICET.
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used in the development of the parasitic carposporophyte (Hommersand and Fredericq 1990).
The isolation, chemical, and spectroscopic characterization of carrageenans extracted at room temperature from the pericarp and the carposporophyte of
Gigartina skottsbergii showed that they were kappa/iotacarrageenans with similar kappa:iota ratios (1:0.24
and 1:0.23, respectively). These determinations also
indicated the presence of lesser amounts of agarans
and/or d,l-galactan hybrids. Similar determinations
carried out on the hot water extracts indicated
kappa:iota-carrageenan ratios with higher iota content (1:0.46 and 1:0.36, for the pericarp and carposporophyte, respectively) and small but significant
amounts of 6-O-methylgalactose and xylose. The cystocarp of Gymnogongrus torulosus also contained a
kappa/iota-carrageenan with similar amounts of both
structures (kappa:iota ratio 1:1.21). No evidence of
lambda-carrageenan and/or lambda structures were
found in the pericarps of Gigartina skottsbergii or in the
cystocarps of Gymnogongrus torulosus. The term lambda
structure is not used as synonym of lambda-carrageenan but naming the structural unit of this product. This structural unit can be found as constituent
of the lambda-carrageenan or, when in small or trace
amounts, interspersed in a backbone of another type
of carrageenan. Small amounts of lambda structures
were only found in the carposporophytes of Gigartina
skottsbergii. Analyses of the residue after extraction of
the carposporophytes of Gigartina skottsbergii with hot
water suggests that the small amounts of nonextracted
carrageenans were also of the kappa type and similar
to those produced by cystocarpic plants of Gigartina
skottsbergii (Matulewicz et al. 1989).
Gordon-Mills and McCandless (1975) reported
that the cell wall of carposporophytic tissues of Chondrus crispus Stackhouse stained with anti-lambda polyclonal antibodies but not with anti-kappa ones. It was
then deduced that the cells of the carposporophytes
produced lambda-carrageenan. Nevertheless, in the
same article it was also reported that the outer cell
walls of the tetrasporangia (tetrasporophytic tissue)
showed strong fluorescence when treated with both
anti-kappa and anti-lambda antibodies, suggesting a
lack of specificity of the antibodies used.
Lestang Bremond et al. (1987) reported 4.8% of
lambda-carrageenan in carrageenans extracted from
gametophytes of Chondrus crispus, but the spectrum
presented was not the one corresponding to this
product (Falshaw and Furneaux 1994, Stortz et al.
1994). On the basis of this circumstantial evidence, it
was sometimes erroneously supposed that the carposporophyte produced lambda-type carrageenans.
Recently, small amounts of lambda structure were
found in the system of carrageenans extracted from
cystocarpic plants of Gigartina skottsbergii and Gymnogongrus torulosus (Ciancia et al. 1993b, 1997, Estevez
et al. 2001). These results, together with those obtained from seaweeds in which both phases produce
only kappa/iota-carrageenans (DiNinno and McCand-
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