Journal of Applied Phycology 15: 21–27, 2003.

21
© 2003 Kluwer Academic Publishers. Printed in the Netherlands.

Polysaccharides from Ulva pertusa (Chlorophyta) and preliminary studies

on their antihyperlipidemia activity

Yu Pengzhan 1,2, Zhang Quanbin 1,2, Li Ning 1,2, Xu Zuhong 1, Wang Yanmei 1 and Li
Zhi’en 1,*
1
Department of Chemistry, Institute of Oceanology of the Chinese Academy of Sciences, Qingdao, 266071,
P.R.China; 2The Graduate School of the Chinese Academy of Sciences, Beijing, 100039, P.R. China; *Author
for correspondence (e-mail: xzh@ms.qdio.ac.cn; phone: +86-532-2898703; fax: +86-532-2968651)
Received 17 August 2002; accepted in revised form 4 November 2002

Key words: Antihyperlipidemia effect, Hyperlipidemia, Polysaccharides, Ulva pertusa, Ulvan

Abstract

Polysaccharides from Ulva pertusa were isolated and prepared by extraction in hot water and precipitation by
ethanol. The water-soluble polysaccharides were chemically well defined, containing 47.0% total carbohydrate,
23.2% uronic acids, 17.1% sulfate groups, 1.0% N and 29.9% ash. Gas chromatography analysis demonstrated
that the neutral sugars were mainly composed of rhamnose, xylose and glucose and smaller amounts of mannose,
galactose and arabinose. The FTIR and 13C-NMR spectra indicated that basic repeating units of the polysaccha-
rides were (␤-D-GlcpA-(1-> 4)-␣-L-Rhap 3S) and (␣-L-IdopA-(1-> 4)-␣-L-Rhap 3S). Fifty ICR mice were used
to study the effect of water-soluble polysaccharides from Ulva pertusa on the level of plasma lipids, with inositol
niacinate as positive control. The results indicated that the polysaccharides significantly lowered the contents of
plasma total cholesterol, low-density lipoprotein cholesterol, triglyceride and markedly increased the contents of
serum high-density lipoprotein cholesterol, compared with the hyperlipidemia control group (p < 0.01). More-
over, administration of polysaccharides significantly decreased the atherogenic index. The present results suggest
that the polysaccharides from Ulva pertusa have great potential for preventing ischemic cardiovascular and cere-
brovascular diseases.

Abbreviations: TC – total cholesterol, LDL-C – low-density lipoprotein cholesterol, TG – triglyceride, HDL-C

– high-density lipoprotein cholesterol

Introduction Ulva pertusa is distributed in China in the inter-

Seaweed polysaccharides are highly active natural
substances having valuable applications (Roehrig
1988; Southgate 1990). Agar, carrageenan and fu-
coidan are well known to be applied extensively in
industry (Franz 1989; De Philippis and Vincenzini
1998; Tseng 2001). However, interest in the potential
of seaweed polysaccharides as drugs is more recent
(Joseph and Baker 1984). Antitumor, antivirus, anti-
hyperlipidemia and anticoagulant biological activities
have been found in seaweed polysaccharides, some of
which are exploited as new drugs (Güven et al. 1990;
De Clercq 1993; Xu Zuhong et al. 1995).
tidal zone of the Yellow Sea and the Bohai Sea. In
ancient times Ulva pertusa was not only consumed as
sea-lettuce and used as an organic fertilizer, but was
also used as a traditional herb for dissipating heat and
treating hydropic and hyperlipidemia diseases, as
documented in Chinese materia medica. There are
various reports on the chemical constituents and the
structure of ulvan extracts (Lahaye et al. 1996, 1997;
Quemener et al. 1997), but few on the biological ac-
tivities of ulvan (Kaeffer et al. 1999). Studies on the
polysaccharides from Ulva pertusa and their antihy-
perlipidemia activity were carried out in this work to
test whether these could be source of new drugs for
22
preventing ischemic cardiovascular and cerebrovas-
cular diseases.
Materials and methods
Ulva pertusa was collected in the coast of Qingdao,
in September, 2001. Epiphytes and sand were re-
moved. Algae were washed in tap water, air dried and
stored in plastic bags at room temperature in a dry,
dark place before use. The reference sugars were pur-
chased from Sigma Chemical CO. (St. Louis, Mo,
USA). Acetanilide (N: 10.36%) was provided by
Wako Pure Chemical industries LTD. (Japan).
Ulvan extraction
Two hundred grams dry samples were autoclaved for
3 h at 110 °C in 40-fold volume of water. The hot
aqueous solution was separated from the algae resi-
dues by successive filtration through gauze and sili-
ceous earth as filter aid. The solution was concen-
trated to about 2 liters under reduced pressure and
dialyzed for 48 h. The polysaccharides were precipi-
tated by the addition of 4-fold volume of 95% (v/v)
ethanol, and washed twice with absolute ethanol, then
dried at 80 °C (mean yield, 22.5 ± 0.8%, n = 5).
Composition analysis
Total carbohydrate content was estimated with the
phenol-sulfuric method (Dubois et al. 1956) using
rhamnose as standard. Uronic acids were measured by
colorimetry (Thibault 1979) using glucuronic acid as
standard. Sulfate content was determined from about
10-mg samples after 2 mol L −1 HCl hydrolysis (2 h
at 100 °C) according to the method of Kawai et al.
(1969). Ash was quantified gravimetrically after 12 h
at 550 °C and a further 4 h at 900 °C. Determination
of nitrogen performed on Perkin-Elmer 240 element
analysis instrument (Perkin-Elmer Corp., USA) was
done according to the method established by AOAC
Commission (1991: P. 396) using acetanilide as stan-
dard. The neutral sugars were determined by convert-
ing them into acetylated aldononitrile derivatives and
detected by gas chromatography. Briefly, 15 mg
polysaccharides were hydrolyzed in 2 mol L −1 tri-
fluoroacetic acid at 100 °C for 4 h. The hydrolysate
was evaporated to dryness and dissolved in 0.5 mL
pyridine. After 10 mg hydroxylammonium and 1 mg
inositol (as internal reference) were added to the so-
lution, it was allowed to react at 90 °C for 30 min.
The mixture was cooled at room temperature. After
0.8 mL acetic anhydride was added to the mixture, it
was allowed to react at 90 °C for another 30 min. Gas
chromatography runs were performed on an HP5890
instrument equipped with an AC-20 capillary column
(30 m × 0.32 mm ID) and a flame ionization detector
(Agilent Technologies Co., Ltd. USA). The carrier
gas was N 2 with a flow rate of 1.0 mL min −1. The
temperature of both oven and detector was kept at 240
°C and the injector was 200 °C. Sugar identification
were done by comparison with reference sugars.
Spectra analysis
Infrared spectrum was recorded from polysaccharides
powders in KBr pellet on a Nicolet-360 FT spectrom-
eter. 13C-NMR spectrum of polysaccharides solution
in D 2O was recorded at 35 °C on a Brucker DPX-
400M spectrometer. Chemical shifts were measured
in parts per million from the internal CDCl 3 attrib-
uted to the signal at ␦76.27.
Animals and the design of the experimental work
Fifty male ICR mice, weighing 19 ± 1 g, were ob-
tained from the Pharmacology Key Laboratory of Na-
tional Products of Yunnan Province (China). The an-
imals were housed in stainless steel cages at room
temperature (25 ± 2 °C) and 12-h light cycle. After
all the experimental mice were fed a commercial feed
diet for 3 days, they were randomly divided into five
groups (n = 10) and began to feed a cholesterol-rich
diet, whose composition was 2.0% cholesterol, 8.0%
lard, 0.3% sodium cholic acid and 89.7% commercial
chow. Three groups were given 125, 250 and 500
mg kg −1 polysaccharides respectively by intubation
for 7 days, one group was treated with inositol niaci-
nate 500 mg kg −1 as positive drug control, another
group was given physiological saline as the hyperlip-
idemia control. The treatments are shown in Table 1.
The animals were given food and distilled water
ad libitum. At the end of the experimental period, the
mice were withheld food for at least 12 h and blood
samples were collected from the eyeballs to measure
the serum TC, TG, HDL-C and LDL-C levels. Serum
TC (Dceg and Ziegenhorn 1983), TG (Nagele et al.
1984) and HDL-C (Allain et al. 1974) were measured
enzymatically and LDL-C levels were calculated
from the Friedewald formula (Friedewald et al. 1972).
 23

Table 1. The experimental groups and administration of polysaccharides or inositol niacinate for hyperlipidemia mice, whose diets were
composed of 2.0% cholesterol, 8.0% lard, 0.3% sodium cholic acid and 89.7% commercial chow.

Administration
Group Number Supplement Dose Concentration Volume
(mg Kg −1) (mg mL −1) (mL)

hyperlipidemia control 10 physiological saline – – 1

polysaccharides 10 polysaccharides 125 2.5 1
10 polysaccharides 250 5.0 1
10 polysaccharides 500 10.0 1
inositol niacinate 10 inositol niacinate 500 10.0 1

Table 2. Chemical composition of sulfated polysaccharide from

Ulva pertusa.

Component Polysaccharides (% dry weight)

Total sugars 47.0

rhamnose 13.7
xylose 6.4
glucose 4.0
mannose 0.8
arabinose 0.5
galactose 0.2
uronic acids 23.2
sulfate 17.1
nitrogen 1.0
ash 29.9

All results were compared using the unpaired Stu-

dent’s t-test and values expressed as mean ± SD. A p
value < 0.01 was considered to be the level of statis-
tical significance.

Results
The chemical composition of polysaccharides from
Ulva pertusa is given in Table 2, showing that uronic
acids, rhamnose, xylose, glucose and sulfate com-
prised their main composition, with smaller amounts
of mannose, arabinose and galactose. The polysaccha-
rides fraction also contained high amounts of ash and
water-soluble proteins. Ash was attributed to counter-
ions associated with sulfate groups and uronic acids
of the polysaccharides and the proteins may exist in
the polysaccharides as sulfated glycoproteins (Ray
and Lahaye 1995).
The infrared absorption spectrum of polysaccha-
rides from Ulva pertusa is shown in Figure 1. The
signal at 1230 cm −1 and two shoulder bands at 835
Figure 1. Infrared absorption spectrum of the polysaccharides
from Ulva pertusa.
and 788 cm −1 were indicative of the presence of sul-
fate ester substitutions (Ray and Lahaye 1995). Ab-
sorbance at 1230 cm −1 is attributed to stretching of
C-O-S. Two other important bands were assigned at
1643 and 1053 cm −1 corresponding respectively to
the stretching of C = O of uronic acids and the vibra-
tion of the C-O-C bridge of glucosides.
The 13C-NMR spectrum of the polysaccharides is
shown in Figure 2. The signals of ␦175.0 and ␦16.8
were easy to recognize, and corresponded to C-6 of
uronic acids and C-6 of rhamnose respectively (Ta-
ble 3). The rest of the signal assignments were done
24

Figure 2. 13C-NMR spectra of U. armoricana from Lahaye et al. (1999) and Ulva pertusa. Letters and numbers correspond to carbons in
chemical structures depicted on top of the spectra. Gg signals are interpreted in discussion sections and correspond to carbons in a separated
glucuronan or contiguous ␤ 1-> 4 linked D-glucuronic acids.

Table 3. 13C-NMR signal chemical shifts (ppm) of main repeating units of ulvan from Ulva pertusa: (␤-D-GlcA-(1-> 4)-␣-L-Rhap 3S) and
(␣-L-IdoA-(1-> 4)-␣-L-Rhap 3S).

Ulvan Unit 1 2 3 4 5 6

glucoronic acid 103.1 73.8 73.8 78.7 76.0 175.0

rhamnose (GlcA-1-> 4-Rhap) 97.3 68.9 78.6 77.0 67.5 16.8
iduronic acid 100.8 70.2 71.2 77.9 70.2 175.0
rhamnose (IdoA-1-> 4-Rhap) 98.0 68.9 78.6 77.0 67.5 16.8

by comparison with the previously published data for Antihyperlipidemia activity

U. armoricana (Lahaye et al. 1999). The major sig-
nals for ulvanobiuronic acid 3-sulphate type A (␤-D-
GlcpA-(1-> 4)-␣-L-Rhap 3S) and B (␣-L-IdopA-(1->
4)-␣-L-Rhap 3S) were prominently observed in the
spectrum of polysaccharides from Ulva pertusa,
which demonstrated that the basic repeating struc-
tures of the polysaccharides were the same as those
of other ulvans (Lahaye et al. 1999).
The mice fed polysaccharide-rich diets showed a sig-
nificant decrease in plasma TC, TG, LDL-C, and an
increase of HDL-C, and the values for HDL-C/TC
compared well with those of the hyperlipidemia con-
trol group (Table 4). The results indicated that the low
dose of 125 mg kg −1 had optimal effect on TG, but a
lesser impact on TC and LDL-C. However, doses of

Table 4. Effect of dietary polysaccharides from Ulva pertusa on serum lipids in mice fed high cholesterol. Mean ± SE; n = 10;
−1 −1
Group TC (mmol L ) TG (mmol L )
Hyperlipidemia control 9.41 ± 1.19 0.96 ± 0.14
Polysaccharides 9.71 ± 0.23 0.63 ⴱⴱ ± 0.09
6.67 ⴱⴱ ± 0.98 0.73 ⴱⴱ ± 0.14
7.07 ⴱⴱ ± 1.11 0.74 ⴱⴱ ± 0.10
Inositol niacinate 6.80 ⴱⴱ ± 1.43 0.64 ⴱⴱ ± 0.07
TC: total cholesterol; LDL-C: low-density lipoprotein cholesterol; TG: triglyceride; HDL-C: high-density lipoprotein cholesterol.
250 and 500 mg kg −1 had significant effects on TC,
TG, HDL-C and LDL-C. More importantly, feeding
with polysaccharides had the greatest effect on the
atherogenic index in mice. A significant trend in the
dose response to atherogenic index (P < 0.01) was
observed in mice fed with 125, 250 and 500 mg kg −1
polysaccharides as compared with the control mice.
Discussion
As expected, cell-wall polysaccharides from Ulva
pertusa are heteropolymers made of rhamnose, xy-
lose, glucose, uronic acids and sulfate. The identifi-
cation of iduronic acid is complicated as there is no
standard available. However, the chemical-enzymatic
method can be employed as a suitable method for
analysis of uronic acids (Quemener et al. 1997). In
the 13C-NMR spectrum of the polysaccharides, some
signals (refer to Gg in the Figure 2.) of 1,4-linked
␤-D-glucuronan were also observed and indicated
that the structure of contiguous ␤ 1-> 4 linked D-glu-
curonic acids or a separate glucuronan may be
present. The sulfated polysaccharides of seaweeds,
differing chemically and physicochemically from
those of land plants, may have special physiological
effects on the human body (Lahaye 1991; Lahaye and
Jegou 1993). Sulfated polysaccharides are associated
with the surfaces of animal cells and involved in bio-
logical activities such as cell recognition, cell adhe-
sion or regulation of receptor functions, which are of
interest in medicine (Ellouali et al. 1993).
Epidemiological studies showed that the chronic
presence of excessive amount of cholesterol and TG
in blood plasma was one of the highest risk factors
for developing atherosclerosis (EL-Swefy et al.
2002). The polysaccharides from Ulva pertusa
showed a high antihyperlipidemia activity in mice but
the mechanism by which ulvan regulate TC, TG,
LDL-C and HDL-C/TC blood levels in hyperlipi-
demia mice is unknown. One of interpretations is that
25
ⴱⴱ
p = < 0.01.
−1 −1
HDLC (mmol L ) LDLC (mmol L ) HDLC/TC
0.83 ± 0.36 8.39 ± 1.08 0.089 ± 0.039
1.09 ± 0.28 8.49 ± 2.37 0.121 ± 0.054
1.00 ± 0.30 5.61 ⴱⴱ ± 1.08 0.152 ⴱⴱ ± 0.054
1.49 ⴱⴱ ± 0.38 5.43 ⴱⴱ ± 1.08 0.215 ⴱⴱ ± 0.059
1.44 ⴱⴱ ± 0.42 5.63 ⴱⴱ ± 1.19 0.214 ⴱⴱ ± 0.056
metabolism was influenced by intestinal morphology
changes induced by fibre feeding (Brown et al. 1979;
Stark et al. 1996). It may be sound to interpret the
pharmacological activity of polysaccharides from
Ulva pertusa, based on followings. Lahaye et al.
(1996) described that glucuronorhamnoxyloglycans
sulfate from Ulva lactuca and ⬙green-tides⬙ were able
to gel with calcium and boric acid. The gelatin of ul-
van showed resistance to human colonic flora fermen-
tation (Bobin-Dubigeon et al. 1997). The function of
resistance may make the polysaccharides conserved
for a relatively long period in the intestine during
which time they may come to confer a selective ad-
vantage. Further approaches are required to take to
observe whether the morphological changes occur in
the intestine of hyperlipidemia mice fed with polysac-
charides from Ulva pertusa. In addition, Dvir et al.
(2000) found that bile acid excretion increased signif-
icantly when mice were fed polysaccharides from
Porphyridium sp., and plasma cholesterol level and
neutral sterol excretion were enhanced markedly. This
phenomenon may indicate another mechanism by
which polysaccharides can act as stimulators of bile
acid synthesis, with the cholesterol pools being de-
pleted by fiber feeding. Whether this kind of meta-
bolic mechanism was stimulated by polysaccharides
from Ulva pertusa will further study of their pharma-
cological properties.
In this preliminary study, polysaccharides from
Ulva pertusa showed promising in preventing athero-
sclerosis and the incidence of myocardial ischemia
and cardiac events induced by atherosclerosis. The
work is in progress that the polysaccharides, as a po-
tential source of new drugs, may provide antihyper-
lipidemia effects similar to, or better than, those of
more traditional plant fibre.
26
Acknowledgements
This work was financed in part by the Scientific and
Technical Department of Shandong Province (China).
We thank Dr. Whitton and Prof. Wang Sen for help in
checking and improving the English version. I am
also grateful to Dr. Lahaye et al. for having cited the
13
C-NMR spectrum of U. armoricana in this paper.
References
Allain C.C., Poon L.S., Chan C.S.G., Richmond W. and Fu P.C.
1974. Enzymatic determination of total serum cholesterol. Clin.
Chem. 20: 470–475.
AOAC Commission of National Import and Export Commodity
Inspection Bureau (China) 1991. Official Methods of Analysis
of the Association of Official Analytical Chemists. 15th edn.
Chinese scientific and technological Press, Beijing, P.396.
Bobin-Dubigeon C., Lahaye M. and Barry J.-L. 1997. Human co-
lonic bacterial degradation of dietary fibres from sea-lettuce
(ulva sp.). J. Sci. Food Agric. 73: 149–159.
Brown N.J., Worlding J., Rumsey R.D.E. and Read N.W. 1979.
Active and inactive forms of 3-hydroxy-3-methylglutaryl coen-
zyme A reductase in the liver of the rat. J. biol. Chem. 254:
5144–5149.
Dceg R. and Ziegenhorn J. 1983. Kinetic enzymatic method for
automated determination of total cholesterol in serum. Clin.
Chem. 29: 1798–1802.
De Clercq E. 1993. Anti-HIV activity of sulfated polysaccharides.
Front. Biomed. Biotechnol. 1: 87–100.
De Philippis R. and Vincenzini M. 1998. Exocellular polysaccha-
rides from cyanobacteria and their possible applications. FEMS
Microbiol. Rev. 22: 151–175.
Dvir I., Chayoth R., Sod-Moriah U., Shany S., Nyska A., Stark
A.H. et al. 2000. Soluble polysaccharide and biomass of red
microalga Porphyridium sp. alter intestinal morphology and re-
duce serum cholesterol in rats. Br. J. Nutrition 84: 469–476.
Dubois M., Gilles K.A., Hamilton J.K., Rebers P.A. and Smith F.
1956. Colorimetric method for determination of sugars and re-
lated substances. Analyt. Chem. 28: 350–356.
Ellouali M., Boisson-Vidal C., Durand P. and Jozefonvicz J. 1993.
Antitumor activity of low molecular weight fucans extracted
from brown seaweed Ascophyllum nodosum. Anticancer Re-
search 13: 2011–2020.
Franz G. 1989. Polysaccharides in pharmacy: current applications
and future concepts. Planta Medica 55: 493–497.
Friedewald W.T., Levy R.I. and Fredrickson D.S. 1972. Estimation
of concentration of low density lipoprotein cholesterol in
plasma without use of the putative ultracentrifuge. Clin. Chem.
18: 499–502.
Güven K.C., Güvener B. and Güler E. 1990. Pharmacological ac-
tivities of marine algae. In: Akatsuka I. (ed.), Introduction to
Applied Phycology. SPB Academic Publishing, Academic Pub-
lishing bv, The Hague, The Netherlands, pp. 67–92.
Joseph T. and Baker O.B.E. 1984. Seaweeds in pharmaceutical
studies and applications. Hydrobiologia 116/117: 29–40.
Kaeffer B., Bénard C., Lahaye M., Blottière H.M. and Cherbut C.
1999. Biological properties of ulvan, a new source of green
seaweed sulfated polysaccharides, on cultured normal and can-
cerous colonic epithelial cells. Planta Medica 65: 527–531.
Kawai Y., Seno N. and Anno K. 1969. A modified method for
chondrosulfatase assay. Analyt. Biochem. 32: 314–321.
Lahaye M. 1991. Marine algae as sources of fibers: Determination
of soluble and insoluble dietary fiber contents in some ’sea
vegetables’. J. Food Sci. Agric. 54: 587–594.
Lahaye M., Brunel M. and Bonnin E. 1997. Fine chemical struc-
ture analysis of oligosaccharides produced by an ulvan-lyase
degradation of the water-soluble cell-wall polysaccharides from
Ulva sp. (Ulvales, Chlorophyta). Carbohydr. Res. 304: 325–
333.
Lahaye M. and Jegou D. 1993. Chemical and physical-chemical
characteristics of dietary fibers from Ulva lactuca (L.) Thuret
and Enteromorpha compressa (L.) Grev. J. appl. Phycol. 5:
195–200.
Lahaye M. and Ray B. 1996. Cell wall polysaccharides from the
marine green algal Ulva ’rigida’ (Ulvales, Chlorophyta). NMR
analysis of ulvan oligosaccharides. Carbohydr. Res. 283: 161–
173.
Lahaye M., Ray B., Baumberger S., Quemener B. and Axelos
M.A.V. 1996. Chemical characterisation and gelling properties
of cell wall polysaccharides from species of Ulva (Ulvales,
Chlorophyta). Hydrobiologia 326/327: 473–480.
Lahaye M., Inizan F. and Vigouroux J. 1998. NMR analysis of the
chemical structure of ulvan and of ulvan-boron complex for-
mation. Carbohydr. Polymers 36: 239–249.
Lahaye M., Alvarez-Cable Cimadevilla E., Kuhlenkamp R.,
Quemener B., Lognoné V. and Dion P. 1999. Chemical compo-
sition and 13C-NMR spectroscopic characterisation of ulvans
from Ulva (Ulvales, Chlorophyta). J. appl. phycol. 11: 1–7.
Nagele U., Hagele E.O. and Sauer G. 1984. Reagent for the enzy-
matic determination of serum total triglycerides with improved
lipolytic efficiency. Clin. Chem. Clin. Biochem. 22: 165–174.
Piskun R.P., Pentiuk A.A., Serkova V.K., Polesia T.L. and Savits-
kaia E.A. 1997. The possible means for the pharmacological
correction of lipid metabolic disorders in atherosclerosis. Eksp.
Klin. Farmakol. 60: 78–85.
Quemener B., Lahaye M. and Bobin-Dubigeon C. 1997. Sugar de-
termination in ulvans by a chemical-enzymatic method coupled
to high performance anion exchange chromatography. J. appl.
Phycol. 9: 179–188.
Ray B. and Lahaye M. 1995. Cell-wall polysaccharides from the
marine green alga Ulva ⬙rigida⬙ (Ulvales, Chlorophyta). Chem-
ical structure of ulvan. Carbohydr. Res. 274: 313–318.
Roehrig K.L. 1988. The physiological effects of dietary fiber – a
review. Food Hydrocolloids 2: 1–18.
EL-Swefy S.E., Asker M.E., Sousou I.A. and Mohammed H.E.
2002. A novel concept to preserve the beneficial effects of hor-
mone replacement therapy in bilaterally female ovariectomized
rats: Role of lovastatin therapy. Pharmacol. Res. 45: 167–173.
Southgate D.A.T. 1990. Dietary fibre and health. In: Southgate
D.A.T., Waldron K., Johnson I.T. and Fenwick G.R. (eds), Die-
tary Fibre: Chemical and Biological Aspects. The Royal So-
ciety of Chemistry, Cambridge, pp. 10–19.
Stark A., Nyska A. and Madar Z. 1996. Metabolic and morpho-
metric changes in small and large intestine in rats fed high-fi-
bre diets. Toxicologic Pathology 24: 166–171.

Thibault J.-F. 1979. Automatisation du dosage des substances pec-
tiques par la méthode au méta-hydroxydiphényl. Lebensm.
Wiss. Technol. 12: 247–251.
Tseng C.K. 2001. Algal biotechnology industries and research ac-
tivities in China. J. appl. Phycol. 13: 375–380.
27
Xu Zuhong, Li Zhi’en, Zhang Xingjun and Guo Yucai 1995. Ap-
plication of Fucodian polysaccharides sulfate as a drug for
treating chronic renal failure. CN 98 1 20283. 7., 16 Dec 1995.