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The potential of the coelomic fluid in sea cucumber as an antioxidant

  • Johor Bahru Public Health Laboratory


In a biological system the product of free-radical induced-lipid peroxidation are conjugated dienes (CDs), lipid peroxides and malondialdehyde (MDA). Its control depends on antioxidant activities. Antioxidant defend the body system by controlling the damage caused by the free radicals. This paper describes lipid peroxidation process and antioxidant activities present in three species of holothuroid obtained from the coastal waters of Terengganu, Malaysia. Among the three species studied, the coelomic fluid of Bohadschia mamorata vitiensis contained the highest level of protein (7.24 + 0.04)g/L. The coelomic fluid of Stichopus badionotus Selenka contained the highest level of superoxide dismutase (SOD) (9.46 + 0.99)X 105 IU/g protein, MDA (16.46 + 1.28)nmol/g protein and total antioxidant activities (AOA) (58.81 + 5.70) %. In Stichopus variegatus Semper the coelomic fluid only demonstrated the highest level of CDs (94.19 + 11.34) RU/g protein compared with the other two species. This preliminary study indicated that some form of antioxidant activities are present in the coelomic fluid of holothuroids.
Mal J Nutr 5:55-59, 1999
The potential of the coelomic fluid in sea cucumber as an antioxidant
Hawa I, Zulaikah M, Jamaludin M, Zainal Abidin AA, Kaswandi MA and Ridzwan BH
Department of Biomedical Science, Faculty of Allied Health Sciences,
Universiti Kebangsaan Malaysia, Jalan Raja Muda A. Aziz, 50300 K. Lumpur, Malaysia.
In a biological system the product of free-radical induced-lipid peroxidation are
conjugated dienes (CDs), lipid peroxides and malondialdehyde (MDA). Its control
depends on antioxidant activities. Antioxidant defend the body system by controlling
the damage caused by the free radicals. This paper describes lipid peroxidation process
and antioxidant activities present in three species of holothuroid obtained from the
coastal waters of Terengganu, Malaysia. Among the three species studied, the
coelomic fluid of Bohadschia mamorata vitiensis contained the highest level of
protein (7.24 + 0.04)g/L. The coelomic fluid of Stichopus badionotus Selenka
contained the highest level of superoxide dismutase (SOD) (9.46 + 0.99)X 105 IU/g
protein, MDA (16.46 + 1.28)nmol/g protein and total antioxidant activities (AOA)
(58.81 + 5.70) %. In Stichopus variegatus Semper the coelomic fluid only
demonstrated the highest level of CDs (94.19 + 11.34) RU/g protein compared with
the other two species. This preliminary study indicated that some form of antioxidant
activities are present in the coelomic fluid of holothuroids.
In the south-east Asia regions sea
cucumbers are taken as a food supplement
(Ridzwan and Che Bashaah 1985; Ridzwan
1993). Holothuroid and its product could cure
certain ailments (Shimada 1969; Sit 1998).
Fishermen often consume the liquid portion
or the coelomic fluid of sea cucumbers in an
attempt to remain healthy for days while out
at sea. It is believed that if consumed
regularly sea cucumbers could reduce
hypertension, asthma, heal internal wounds
and cancer. This bentoic organism has a
vascular system filled with fluid believed to
contain bioactive substances important for
wound healing (Perchenik 1969).
56 Hawa I, Jamaludin M, Zainal A, et. al.
Certain extracts of sea cucumbers had
also been described to contain vitamin E, an
antioxidant (Madhavan 1998). Therefore, the
purpose of this study is to further investigate
whether other antioxidant activities are
present in the coelomic fluid of three species
of holothuroids that inhabited the Malaysian
coastal waters. The species investigated are
Bohadschia mamorata vitiensis, Stichopus
badionotus Selenka and Stichopus variegatus
Samples were collected from the coastal
waters of Terengganu, Malaysia. An incision
measuring 3-5 cm were made on the ventral
side taking care not to injure the internal
organs. The coelomic fluid were collected in
separate beakers, later transferred into test
tubes, frozen until analysed. The protein
component was measured by using the Biuret
calorimetric method (Wooton and Freeman
1982), while the MDA levels determined
using the standard procedures (Hunter and
Jamaludin 1986). The activity of SOD was
determined by a modified Flohe and Otting
(1984) method, whereas total antioxidant
activity was measured according to Stock et
al. (19874). Conjugated dienes was
determined following the Lunec and
Dormandy (1978) method.
The results of the antioxidant activities
in the coelomic fluid of the three species of
holothuroids is shown in Table 1. The
coelomic fluid of Bohadschia mamorata
vitiensis contained the highest concentration
of crude protein (7.24 + 0.04 g/L) followed
by Stichopus variegatus Semper (2.05 + 0.40
g/L) and Stichopus badionotus Selenka (1.80
+ 0.24 g/L).
Among the three species studied the
highest level of SOD activity was detected in
the coelomic fluid of Stichopus badionotus
Selenka (9.40 + 0.99)X 105 IU/g protein
followed by Stichopus variegatus Semper
(8.20 + 0.91) X 105 IU/g protein and
Bohadschia mamorata vitiensis (5.30 + 0.18)
X 105 IU/g protein.
The coelomic fluid also contained CDs,
the product of primary lipid peroxidation. It
was the highest in Stichopus variegatus
(94.19 + 11.34) RU/g protein followed by
Bohadschia mamorata vitiensis (78.71 + 1.56)
RU/g protein and Stichopus badionotus
Selenka (70.00 + 4.55) RU/g protein.
Stichopus badionatotus contained the highest
level of (16.46 +1.29) nmol/g protein,
followed by Stichopus variegatus (9.89 +
0.04) nmol/g protein and Bohadschia
mamorata vitiensis (7.41 + 0.86) nmol/g
protein. Total antioxidant activity was found
to be the highest in Stichopus badionotus
Selenka (58.81 + 5.70) %, followed by
Bohadschia mamorata vitiensis (52.20 + 5.70)
% and Stichopus variegatus Semper (47.71 +
8.95) %.
The potential of coelomic fluid in sea cucumber 57
TABLE 1. The Composition of Sea cucumber Coelomic Fluid
Species Product Amount
Stichopus badionotus Selenka Crude protein 1.80 + 0.24 g/L
Superoxide dismutase 9.40 + 0.99 X 105 IU/g protein
Malondialdehyde 16.46 + 1.29 nmol/g protein
Conjugated dienes 70.00 + 4.55 RU/g protein
Total antioxidant activity 8.81 + 5.7 %
Stichopus variegatus Semper Crude protein 2.05 + 0.40 g/L
Superoxide dismutase 8.20 + 0.91 X 105IU/g protein
Malondialdehyde 9.89 + 0.04 nmol/g protein
Conjugated dienes 94.19 + 1134 RU/g protein
Total antioxidant activity 47.71 + 8.95 %
Bohadschia mamorata vitiensis Crude protein 7.24 + 0.04 g/L
Superoxide dismutase 5.30 + 0.18 X 105IU/g protein
Malondialdehyde 7.41 + 0.86 nmol/g protein
Conjugated dienes 78.71 + 1.56 RU/g protein
Total antioxidant activity 52.20 + 5.70 %
Sea cucumbers, as an alternative source
of natural dietary antioxidants probably plays
an important role to counter the detrimental
effects of oxygen free radicals formed from
normal metabolism and external factors
(pollution, radiation). This is implicated in the
development of today’s common illness such
as cancer (gene damage), premature ageing,
cardiovascular and other degenerative
diseases. Indeed, our study revealed the
presence of a mild concentration of total
protein in the coelomic fluid of all the three
species of holothuroid. The findings,
however, are similar to the results reported by
Canicatti (1989, 1990) but on Holothuria
polii and Marthasterias glacialis.
Furthermore, our study also showed a
remarkable increase in the activity of
antioxidant enzymes, namely, SOD. This is in
agreement with our study on lipid
peroxidation of polyunsaturated fatty acids of
ox brain homogenates as a bioassay system
for auto oxidation. A similar result was
obtained when using plasma serum incubated
in tissue homogenate inhibit lipid auto
oxidation process (Barber 1961). According
to Dormandy (1969) all animal tissues have a
natural defence system to inhibit the process
of oxidation towards polyunsaturated fatty
acid either by enzymatic or non-enzymatic
reaction. The main role of antioxidants invivo
is in the prevention of the peroxidation of
PUFAs and the stabilization of the biological
However, the role of this antioxidant in
holothuroids remain to be elucidated. Most
probably, the increased production of oxygen
derived free radicals during hypoxic alteration
due to external changes
58 Hawa I, Jamaludin M, Zainal A. et. al.
occurred at a highly accelerated rate, that is,
when molecular oxygen was reintroduced to
the previously hypoxic tissues of
holothuroids. Thus, hypoxia enhanced the
damages induced by oxygen metabolites
complicated with the reintroduction of
molecular oxygen when the antioxidant
defense system of holothuroid was not
effective or overloaded. Thus, our findings
are similar to previous reports on human
plasma (Barber 1961; Vidlakova et al. 1972;
Slater 1972).
These results indicated that the
coelomic fluid of holothuroids from
Terengganu coastal areas contained
antioxidant substances. These substances
could possibly be produced during the
hypoxic re-oxygenation, during which a high
level of protection against the oxygen induced
damage of “stunned holothuroid” after being
exposed to the polluted habitat. It also appears
that all animals have some antioxidant
activities but the strength varies with species.
Therefore, holothuroids could provide a good
source of external antioxidant for human in
the future.
The study is funded by IRPA grant 09-
02-02-0021. “Identification and evaluation of
structure, bioactive agent and consumption of
local sea cucumbers”.
Barber, A.A. 1961. Inhibition of lipid
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Canicatti, C. 1989. Tthe lytic system of
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review. Boll. Zool. 55:139-144.
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coelomocyte lysate. Comp. Biochem.
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Dormandy, T.L. 1969. Biological
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Flohe, L. and Otting, F. 1984. Oxygen
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Hunter, M.I.S. and Jamaludin, M. 1986.
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The Holothuria polii amebocytes produce two molecules of about 80 KDa molecular weight, serologically identical but with different chemico‐physical characteristics. A calcium‐dependent, heat‐sensible hemolysin (hemolysin 1) appears to be released under natural conditions, while a calcium‐independent, heat‐stable hemolysin (hemolysin 2) is produced under immunological stimulation. Both are able to interact with foreign cell surfaces probably through membrane sphingomyeline, producing serious damage leading to lysis. The molecules also act as opsonic elements. In the present paper a possible phylogenetic interpretation of the origin of these hemoly‐sins is discussed on the basis of similar effects observed in invertebrates and in relation to vertebrate complement and perforin.
Lipid peroxide formation in incubated rat brain homogenates was inhibited by all vertebrate blood sera tested. Rat and chicken sera contained both dialyzable and non-dialyzable inhibitors, whereas turtle, frog, and fish sera had only nondialyzable inhibitors. Chicken, turtle, and frog sera had two electrophoretically different inhibitors; rat and fish sera had only a single one. The inhibitor in rat was associated with an iron-binding protein believed to be siderophilin, and the mechanism of its action was chelation. The inhibitors of the other sera also seemed to be associated with the iron-binding proteins. More information on these proteins in the lower vertebrates is needed before the mechanism of action of these inhibitors can be established as conclusively as in the rat. The significance of these findings with respect to understanding the differences in radiation sensitivity of tissue antioxidant activity was discussed.
1.1. Holothuria polii cell-free coelomic fluid and coelomocyte lysate possess protease activity. It depends on thermolabile enzymes influenced by pH and temperature but not affected by cations supplementation in the reaction medium.2.2. The protease is produced by coelomocytes which in vitro release it.3.3. The inhibitory results indicate that the enzyme is a serine protease of the trypsin-like type.
1. Samples and extracts from synovial effusions were examined for the presence of fluorescent degradation products of free-radical oxidation (peroxidation). 2. Two classes of fluorescent compounds were identified and their fluorescence and physical properties are described. The fluorescence in the aqueous methanol phase of the extracts had the characteristics of conjugated Schiff bases. 3. Changes in synovial fluid and synovial fluid extracts in vitro induced by u.v. irradiation, prolonged incubation and the enzymic generation of oxidizing free radicals, with or without admixed peroxidizing arachidonic acid, and in the absence and presence of antioxidants, suggest that the fluorescent material is derived from peroxidized polyunsaturated lipids in damaged synovial tissue.
Lipid peroxidation (LPX) products in plasma were elevated in Duchenne muscular dystrophy (DMD) patients (n = 9) compared with an age- and sex-matched group of controls (n = 10). Trichloroacetic acid-precipitable thiobarbituric acid-reactive substances (TBARS) were increased by 35% (p less than 0.01), conjugated dienes by 77% (p less than 0.02) and fluorescent pigments (FP) by 70% (p less than 0.001). Plasma alpha-tocopherol was significantly decreased, by 50%, in DMD compared with healthy subjects (p less than 0.002). In conjunction with the finding of others of a deficiency in DMD of the plasma lipoproteins which normally transport vitamin E to the tissues, this may contribute to the increased LPX seen in the disease especially if muscle damage also liberates free Fe2+. The major circulating antioxidant protein, caeruloplasmin (CP), was significantly increased (p less than 0.001) by 25-30% in DMD patients compared with both adult and young male controls. In DMD patients only, this increase is correlated with the concentration of TBARS and FP suggesting that increased LPX may be a trigger for CP synthesis.
Autoxidation, a destructive interaction between unsaturated fats and molecular oxygen, accounts for many industrial and natural-decay processes. The possibility that the body fats might undergo a similar kind of degradation is still largely ignored—perhaps because the irregular irreversible pattern of this type of process seems at odds with the enzyme-controlled reversible pathways of traditional biochemistry. Yet work with mitochondria and other biological preparations has shown that the processes commonly grouped together as " degeneration ", " fatigue ", and " ageing " (none of which have a basis in classical enzymology) develop in close parallel with evidence of rancidification.
Tocopherol and serum lipid peroxidation inhibitor are two different antioxidative systems. This view has been proved by the fact that no parallel changes in serum antioxidative activity and tocopherol values were found and because most of the tocopherol separates in the low-density lipoprotein fraction where in comparison with the whole serum only very low antioxidative activity is found. The importance of the protein nature of the serum lipid peroxidation inhibitor is discussed in terms of its participation in the protection of unsaturated fatty acids during their intravascular transport.
An antifungal steroid glycoside, holotoxin, has been isolated from the sea cucumber Stichopus japonicus (Selenka). In vitro, it exhibits high activity against various fungi, including vegetable pathogens, but has scarcely any activity against Gram-positive and Gram-negative bacteria and mycobacteria in vitro.