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: [email protected]; 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-

tidal zone of the Yellow Sea and the Bohai Sea. In
Seaweed polysaccharides are highly active natural ancient times Ulva pertusa was not only consumed as
substances having valuable applications (Roehrig sea-lettuce and used as an organic fertilizer, but was
1988; Southgate 1990). Agar, carrageenan and fu- also used as a traditional herb for dissipating heat and
coidan are well known to be applied extensively in treating hydropic and hyperlipidemia diseases, as
industry (Franz 1989; De Philippis and Vincenzini documented in Chinese materia medica. There are
1998; Tseng 2001). However, interest in the potential various reports on the chemical constituents and the
of seaweed polysaccharides as drugs is more recent structure of ulvan extracts (Lahaye et al. 1996, 1997;
(Joseph and Baker 1984). Antitumor, antivirus, anti- Quemener et al. 1997), but few on the biological ac-
hyperlipidemia and anticoagulant biological activities tivities of ulvan (Kaeffer et al. 1999). Studies on the
have been found in seaweed polysaccharides, some of polysaccharides from Ulva pertusa and their antihy-
which are exploited as new drugs (Güven et al. 1990; perlipidemia activity were carried out in this work to
De Clercq 1993; Xu Zuhong et al. 1995). test whether these could be source of new drugs for
22

preventing ischemic cardiovascular and cerebrovas- lution, it was allowed to react at 90 °C for 30 min.
cular diseases. 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
Materials and methods chromatography runs were performed on an HP5890
instrument equipped with an AC-20 capillary column
Ulva pertusa was collected in the coast of Qingdao, (30 m × 0.32 mm ID) and a flame ionization detector
in September, 2001. Epiphytes and sand were re- (Agilent Technologies Co., Ltd. USA). The carrier
moved. Algae were washed in tap water, air dried and gas was N 2 with a flow rate of 1.0 mL min −1. The
stored in plastic bags at room temperature in a dry, temperature of both oven and detector was kept at 240
dark place before use. The reference sugars were pur- °C and the injector was 200 °C. Sugar identification
chased from Sigma Chemical CO. (St. Louis, Mo, were done by comparison with reference sugars.
USA). Acetanilide (N: 10.36%) was provided by
Wako Pure Chemical industries LTD. (Japan). Spectra analysis

Ulvan extraction Infrared spectrum was recorded from polysaccharides

powders in KBr pellet on a Nicolet-360 FT spectrom-
Two hundred grams dry samples were autoclaved for eter. 13C-NMR spectrum of polysaccharides solution
3 h at 110 °C in 40-fold volume of water. The hot in D 2O was recorded at 35 °C on a Brucker DPX-
aqueous solution was separated from the algae resi- 400M spectrometer. Chemical shifts were measured
dues by successive filtration through gauze and sili- in parts per million from the internal CDCl 3 attrib-
ceous earth as filter aid. The solution was concen- uted to the signal at ␦76.27.
trated to about 2 liters under reduced pressure and
dialyzed for 48 h. The polysaccharides were precipi- Animals and the design of the experimental work
tated by the addition of 4-fold volume of 95% (v/v)
ethanol, and washed twice with absolute ethanol, then Fifty male ICR mice, weighing 19 ± 1 g, were ob-
dried at 80 °C (mean yield, 22.5 ± 0.8%, n = 5). tained from the Pharmacology Key Laboratory of Na-
tional Products of Yunnan Province (China). The an-
Composition analysis imals were housed in stainless steel cages at room
temperature (25 ± 2 °C) and 12-h light cycle. After
Total carbohydrate content was estimated with the all the experimental mice were fed a commercial feed
phenol-sulfuric method (Dubois et al. 1956) using diet for 3 days, they were randomly divided into five
rhamnose as standard. Uronic acids were measured by groups (n = 10) and began to feed a cholesterol-rich
colorimetry (Thibault 1979) using glucuronic acid as diet, whose composition was 2.0% cholesterol, 8.0%
standard. Sulfate content was determined from about lard, 0.3% sodium cholic acid and 89.7% commercial
10-mg samples after 2 mol L −1 HCl hydrolysis (2 h chow. Three groups were given 125, 250 and 500
at 100 °C) according to the method of Kawai et al. mg kg −1 polysaccharides respectively by intubation
(1969). Ash was quantified gravimetrically after 12 h for 7 days, one group was treated with inositol niaci-
at 550 °C and a further 4 h at 900 °C. Determination nate 500 mg kg −1 as positive drug control, another
of nitrogen performed on Perkin-Elmer 240 element group was given physiological saline as the hyperlip-
analysis instrument (Perkin-Elmer Corp., USA) was idemia control. The treatments are shown in Table 1.
done according to the method established by AOAC The animals were given food and distilled water
Commission (1991: P. 396) using acetanilide as stan- ad libitum. At the end of the experimental period, the
dard. The neutral sugars were determined by convert- mice were withheld food for at least 12 h and blood
ing them into acetylated aldononitrile derivatives and samples were collected from the eyeballs to measure
detected by gas chromatography. Briefly, 15 mg the serum TC, TG, HDL-C and LDL-C levels. Serum
polysaccharides were hydrolyzed in 2 mol L −1 tri- TC (Dceg and Ziegenhorn 1983), TG (Nagele et al.
fluoroacetic acid at 100 °C for 4 h. The hydrolysate 1984) and HDL-C (Allain et al. 1974) were measured
was evaporated to dryness and dissolved in 0.5 mL enzymatically and LDL-C levels were calculated
pyridine. After 10 mg hydroxylammonium and 1 mg from the Friedewald formula (Friedewald et al. 1972).
inositol (as internal reference) were added to the so-
 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
Figure 1. Infrared absorption spectrum of the polysaccharides
The chemical composition of polysaccharides from from Ulva pertusa.
Ulva pertusa is given in Table 2, showing that uronic
acids, rhamnose, xylose, glucose and sulfate com- and 788 cm −1 were indicative of the presence of sul-
prised their main composition, with smaller amounts fate ester substitutions (Ray and Lahaye 1995). Ab-
of mannose, arabinose and galactose. The polysaccha- sorbance at 1230 cm −1 is attributed to stretching of
rides fraction also contained high amounts of ash and C-O-S. Two other important bands were assigned at
water-soluble proteins. Ash was attributed to counter- 1643 and 1053 cm −1 corresponding respectively to
ions associated with sulfate groups and uronic acids the stretching of C = O of uronic acids and the vibra-
of the polysaccharides and the proteins may exist in tion of the C-O-C bridge of glucosides.
the polysaccharides as sulfated glycoproteins (Ray The 13C-NMR spectrum of the polysaccharides is
and Lahaye 1995). shown in Figure 2. The signals of ␦175.0 and ␦16.8
The infrared absorption spectrum of polysaccha- were easy to recognize, and corresponded to C-6 of
rides from Ulva pertusa is shown in Figure 1. The uronic acids and C-6 of rhamnose respectively (Ta-
signal at 1230 cm −1 and two shoulder bands at 835 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- The mice fed polysaccharide-rich diets showed a sig-
GlcpA-(1-> 4)-␣-L-Rhap 3S) and B (␣-L-IdopA-(1-> nificant decrease in plasma TC, TG, LDL-C, and an
4)-␣-L-Rhap 3S) were prominently observed in the increase of HDL-C, and the values for HDL-C/TC
spectrum of polysaccharides from Ulva pertusa, compared well with those of the hyperlipidemia con-
which demonstrated that the basic repeating struc- trol group (Table 4). The results indicated that the low
tures of the polysaccharides were the same as those dose of 125 mg kg −1 had optimal effect on TG, but a
of other ulvans (Lahaye et al. 1999). lesser impact on TC and LDL-C. However, doses of
 25
ⴱⴱ
Table 4. Effect of dietary polysaccharides from Ulva pertusa on serum lipids in mice fed high cholesterol. Mean ± SE; n = 10; p = < 0.01.
−1 −1 −1 −1
Group TC (mmol L ) TG (mmol L ) HDLC (mmol L ) LDLC (mmol L ) HDLC/TC

Hyperlipidemia control 9.41 ± 1.19 0.96 ± 0.14 0.83 ± 0.36 8.39 ± 1.08 0.089 ± 0.039
Polysaccharides 9.71 ± 0.23 0.63 ⴱⴱ ± 0.09 1.09 ± 0.28 8.49 ± 2.37 0.121 ± 0.054
6.67 ⴱⴱ ± 0.98 0.73 ⴱⴱ ± 0.14 1.00 ± 0.30 5.61 ⴱⴱ ± 1.08 0.152 ⴱⴱ ± 0.054
7.07 ⴱⴱ ± 1.11 0.74 ⴱⴱ ± 0.10 1.49 ⴱⴱ ± 0.38 5.43 ⴱⴱ ± 1.08 0.215 ⴱⴱ ± 0.059
Inositol niacinate 6.80 ⴱⴱ ± 1.43 0.64 ⴱⴱ ± 0.07 1.44 ⴱⴱ ± 0.42 5.63 ⴱⴱ ± 1.19 0.214 ⴱⴱ ± 0.056
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, metabolism was influenced by intestinal morphology
TG, HDL-C and LDL-C. More importantly, feeding changes induced by fibre feeding (Brown et al. 1979;
with polysaccharides had the greatest effect on the Stark et al. 1996). It may be sound to interpret the
atherogenic index in mice. A significant trend in the pharmacological activity of polysaccharides from
dose response to atherogenic index (P < 0.01) was Ulva pertusa, based on followings. Lahaye et al.
observed in mice fed with 125, 250 and 500 mg kg −1 (1996) described that glucuronorhamnoxyloglycans
polysaccharides as compared with the control mice. 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-
Discussion tation (Bobin-Dubigeon et al. 1997). The function of
resistance may make the polysaccharides conserved
As expected, cell-wall polysaccharides from Ulva for a relatively long period in the intestine during
pertusa are heteropolymers made of rhamnose, xy- which time they may come to confer a selective ad-
lose, glucose, uronic acids and sulfate. The identifi- vantage. Further approaches are required to take to
cation of iduronic acid is complicated as there is no observe whether the morphological changes occur in
standard available. However, the chemical-enzymatic the intestine of hyperlipidemia mice fed with polysac-
method can be employed as a suitable method for charides from Ulva pertusa. In addition, Dvir et al.
analysis of uronic acids (Quemener et al. 1997). In (2000) found that bile acid excretion increased signif-
the 13C-NMR spectrum of the polysaccharides, some icantly when mice were fed polysaccharides from
signals (refer to Gg in the Figure 2.) of 1,4-linked Porphyridium sp., and plasma cholesterol level and
␤-D-glucuronan were also observed and indicated neutral sterol excretion were enhanced markedly. This
that the structure of contiguous ␤ 1-> 4 linked D-glu- phenomenon may indicate another mechanism by
curonic acids or a separate glucuronan may be which polysaccharides can act as stimulators of bile
present. The sulfated polysaccharides of seaweeds, acid synthesis, with the cholesterol pools being de-
differing chemically and physicochemically from pleted by fiber feeding. Whether this kind of meta-
those of land plants, may have special physiological bolic mechanism was stimulated by polysaccharides
effects on the human body (Lahaye 1991; Lahaye and from Ulva pertusa will further study of their pharma-
Jegou 1993). Sulfated polysaccharides are associated cological properties.
with the surfaces of animal cells and involved in bio- In this preliminary study, polysaccharides from
logical activities such as cell recognition, cell adhe- Ulva pertusa showed promising in preventing athero-
sion or regulation of receptor functions, which are of sclerosis and the incidence of myocardial ischemia
interest in medicine (Ellouali et al. 1993). and cardiac events induced by atherosclerosis. The
Epidemiological studies showed that the chronic work is in progress that the polysaccharides, as a po-
presence of excessive amount of cholesterol and TG tential source of new drugs, may provide antihyper-
in blood plasma was one of the highest risk factors lipidemia effects similar to, or better than, those of
for developing atherosclerosis (EL-Swefy et al. more traditional plant fibre.
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
26

Acknowledgements 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-
This work was financed in part by the Scientific and cerous colonic epithelial cells. Planta Medica 65: 527–531.
Technical Department of Shandong Province (China). Kawai Y., Seno N. and Anno K. 1969. A modified method for
We thank Dr. Whitton and Prof. Wang Sen for help in chondrosulfatase assay. Analyt. Biochem. 32: 314–321.
checking and improving the English version. I am Lahaye M. 1991. Marine algae as sources of fibers: Determination
also grateful to Dr. Lahaye et al. for having cited the of soluble and insoluble dietary fiber contents in some ’sea
13 vegetables’. J. Food Sci. Agric. 54: 587–594.
C-NMR spectrum of U. armoricana in this paper. 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
References Ulva sp. (Ulvales, Chlorophyta). Carbohydr. Res. 304: 325–
333.
Lahaye M. and Jegou D. 1993. Chemical and physical-chemical
Allain C.C., Poon L.S., Chan C.S.G., Richmond W. and Fu P.C.
1974. Enzymatic determination of total serum cholesterol. Clin. characteristics of dietary fibers from Ulva lactuca (L.) Thuret
and Enteromorpha compressa (L.) Grev. J. appl. Phycol. 5:
Chem. 20: 470–475.
AOAC Commission of National Import and Export Commodity 195–200.
Lahaye M. and Ray B. 1996. Cell wall polysaccharides from the
Inspection Bureau (China) 1991. Official Methods of Analysis
of the Association of Official Analytical Chemists. 15th edn. marine green algal Ulva ’rigida’ (Ulvales, Chlorophyta). NMR
analysis of ulvan oligosaccharides. Carbohydr. Res. 283: 161–
Chinese scientific and technological Press, Beijing, P.396.
173.
Bobin-Dubigeon C., Lahaye M. and Barry J.-L. 1997. Human co-
lonic bacterial degradation of dietary fibres from sea-lettuce Lahaye M., Ray B., Baumberger S., Quemener B. and Axelos
M.A.V. 1996. Chemical characterisation and gelling properties
(ulva sp.). J. Sci. Food Agric. 73: 149–159.
Brown N.J., Worlding J., Rumsey R.D.E. and Read N.W. 1979. of cell wall polysaccharides from species of Ulva (Ulvales,
Chlorophyta). Hydrobiologia 326/327: 473–480.
Active and inactive forms of 3-hydroxy-3-methylglutaryl coen-
zyme A reductase in the liver of the rat. J. biol. Chem. 254: Lahaye M., Inizan F. and Vigouroux J. 1998. NMR analysis of the
5144–5149. chemical structure of ulvan and of ulvan-boron complex for-
Dceg R. and Ziegenhorn J. 1983. Kinetic enzymatic method for mation. Carbohydr. Polymers 36: 239–249.
automated determination of total cholesterol in serum. Clin. Lahaye M., Alvarez-Cable Cimadevilla E., Kuhlenkamp R.,
Quemener B., Lognoné V. and Dion P. 1999. Chemical compo-
Chem. 29: 1798–1802.
De Clercq E. 1993. Anti-HIV activity of sulfated polysaccharides. sition and 13C-NMR spectroscopic characterisation of ulvans
from Ulva (Ulvales, Chlorophyta). J. appl. phycol. 11: 1–7.
Front. Biomed. Biotechnol. 1: 87–100.
Nagele U., Hagele E.O. and Sauer G. 1984. Reagent for the enzy-
De Philippis R. and Vincenzini M. 1998. Exocellular polysaccha-
matic determination of serum total triglycerides with improved
rides from cyanobacteria and their possible applications. FEMS
lipolytic efficiency. Clin. Chem. Clin. Biochem. 22: 165–174.
Microbiol. Rev. 22: 151–175.
Piskun R.P., Pentiuk A.A., Serkova V.K., Polesia T.L. and Savits-
Dvir I., Chayoth R., Sod-Moriah U., Shany S., Nyska A., Stark
kaia E.A. 1997. The possible means for the pharmacological
A.H. et al. 2000. Soluble polysaccharide and biomass of red
correction of lipid metabolic disorders in atherosclerosis. Eksp.
microalga Porphyridium sp. alter intestinal morphology and re-
Klin. Farmakol. 60: 78–85.
duce serum cholesterol in rats. Br. J. Nutrition 84: 469–476.
Quemener B., Lahaye M. and Bobin-Dubigeon C. 1997. Sugar de-
Dubois M., Gilles K.A., Hamilton J.K., Rebers P.A. and Smith F.
termination in ulvans by a chemical-enzymatic method coupled
1956. Colorimetric method for determination of sugars and re-
to high performance anion exchange chromatography. J. appl.
lated substances. Analyt. Chem. 28: 350–356.
Phycol. 9: 179–188.
Ellouali M., Boisson-Vidal C., Durand P. and Jozefonvicz J. 1993.
Ray B. and Lahaye M. 1995. Cell-wall polysaccharides from the
Antitumor activity of low molecular weight fucans extracted
marine green alga Ulva ⬙rigida⬙ (Ulvales, Chlorophyta). Chem-
from brown seaweed Ascophyllum nodosum. Anticancer Re-
ical structure of ulvan. Carbohydr. Res. 274: 313–318.
search 13: 2011–2020.
Roehrig K.L. 1988. The physiological effects of dietary fiber – a
Franz G. 1989. Polysaccharides in pharmacy: current applications
review. Food Hydrocolloids 2: 1–18.
and future concepts. Planta Medica 55: 493–497.
EL-Swefy S.E., Asker M.E., Sousou I.A. and Mohammed H.E.
Friedewald W.T., Levy R.I. and Fredrickson D.S. 1972. Estimation
2002. A novel concept to preserve the beneficial effects of hor-
of concentration of low density lipoprotein cholesterol in
mone replacement therapy in bilaterally female ovariectomized
plasma without use of the putative ultracentrifuge. Clin. Chem.
rats: Role of lovastatin therapy. Pharmacol. Res. 45: 167–173.
18: 499–502.
Southgate D.A.T. 1990. Dietary fibre and health. In: Southgate
Güven K.C., Güvener B. and Güler E. 1990. Pharmacological ac-
D.A.T., Waldron K., Johnson I.T. and Fenwick G.R. (eds), Die-
tivities of marine algae. In: Akatsuka I. (ed.), Introduction to
tary Fibre: Chemical and Biological Aspects. The Royal So-
Applied Phycology. SPB Academic Publishing, Academic Pub-
ciety of Chemistry, Cambridge, pp. 10–19.
lishing bv, The Hague, The Netherlands, pp. 67–92.
Stark A., Nyska A. and Madar Z. 1996. Metabolic and morpho-
Joseph T. and Baker O.B.E. 1984. Seaweeds in pharmaceutical
metric changes in small and large intestine in rats fed high-fi-
studies and applications. Hydrobiologia 116/117: 29–40.
bre diets. Toxicologic Pathology 24: 166–171.
 27

Thibault J.-F. 1979. Automatisation du dosage des substances pec- Xu Zuhong, Li Zhi’en, Zhang Xingjun and Guo Yucai 1995. Ap-
tiques par la méthode au méta-hydroxydiphényl. Lebensm. plication of Fucodian polysaccharides sulfate as a drug for
Wiss. Technol. 12: 247–251. treating chronic renal failure. CN 98 1 20283. 7., 16 Dec 1995.
Tseng C.K. 2001. Algal biotechnology industries and research ac-
tivities in China. J. appl. Phycol. 13: 375–380.