Journal of Experimental Marine Biology and Ecology
297 (2003) 107 – 118
www.elsevier.com/locate/jembe
The resistance to ammonia stress of Penaeus
monodon Fabricius juvenile fed diets
supplemented with astaxanthin
Chih-Hung Pan a, Yew-Hu Chien b,*, Brian Hunter c
a
Department of Aquaculture, National Kaohsiung Institute of Marine Science and Technology,
Kaohsiung 811, Taiwan, ROC
b
Department of Aquaculture, National Taiwan Ocean University, Keelung 202, Taiwan, ROC
c
Roche Aquaculture Centre Asia Pacific, 11/F 2535 Sukhumvit Road, Bangchak Prakanong,
Bangkok 10250, Thailand
Received 1 May 2003; received in revised form 3 July 2003; accepted 28 July 2003
Abstract
This study was aimed at determining if the increase of body astaxanthin content through dietary
supplementation in tiger prawn Penaeus monodon juvenile could enhance its antioxidant defense
capability and resistance to ammonia stress. Haemolymph total antioxidant status (TAS) and
superoxide dismutase (SOD) were chosen as parameters of shrimp antioxidant capacity. Resistance to
chemical stress was evaluated by shrimp survival rate, and haemolymph aspartate aminotransferase
(AST) and alanine aminotransferase (ALT). P. monodon 5-day postlarvae were fed diets supplemented
with 0 and 71.5 mg kg 1 astaxanthin for 8 weeks. Shrimps were then subjected to 72-h exposure of
ammonia at 0.02, 0.2, 2 and 20 mg l 1. The survival rates of the astaxanthin-fed (AX) shrimp were
higher than those of the control shrimp under all levels of ammonia except 20 mg l 1, showing that the
shrimp’s resistance to ammonia stress had been improved by dietary astaxanthin. AX shrimp had
higher TAS than control shrimp at ammonia levels higher than 0.02 mg l 1 and lower SOD at all
ammonia levels suggested that antioxidation capability had been greatly enhanced. AST in AX shrimp
was lower than that in control shrimp under all levels of ammonia stress. ALT in AX shrimp was either
lower than or equal to that in control shrimp under various levels of ammonia. Both AST and ALT
reflected that shrimp hepatopancreatic function had been improved by dietary astaxanthin. Astaxanthin
can become essential for P. monodon when the animal is under ammonia stress.
D 2003 Elsevier B.V. All rights reserved.
Keywords: Aminotransferase; Ammonia stress; Astaxanthin; Penaeus monodon; Superoxide dismutase; Total
antioxidant status
* Corresponding author. Tel.: +886-2-24622192x5204; fax: +886-2-24625393.
E-mail address: [email protected] (Y.-H. Chien).
0022-0981/$ - see front matter D 2003 Elsevier B.V. All rights reserved.
doi:10.1016/j.jembe.2003.07.002
108
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1. Introduction
While an organism is subjected to chemical, physical and biological (i.e. pathogen
infection) stress, sudden shortage of oxygen causes abnormal oxidative reactions in the
aerobic metabolic pathways, resulting in the formation of excessive amounts of singlet
oxygen (Ranby and Rabek, 1978) and other reactive oxygen species (ROS). ROS can
impair lipids, proteins, carbohydrates and nucleotides (Yu, 1994), which are important
parts of cellular constituents, including membranes, enzymes and DNA. Radical damage
can be significant because it can proceed as a chain reaction.
Carotenoids play an important role in animal health as antioxidants through inactivation
of free radicals produced from normal cellular activity and various stressors (Chew, 1995).
h-Carotene is recognized as a lipid antioxidant, i.e. a free radical trap and quencher of
singlet oxygen. Astaxanthin contains a long conjugated double bond system with relatively
unstable electron orbitals, which may help it scavenge oxygen radicals in cells (Stanier et
al., 1971). The antioxidant activity of astaxanthin was found to be approximately 10 times
stronger than h-carotene and 100 times greater than that of a-tocopherol (Shimidzu et al.,
1996). Astaxanthin also showed strong activity as an inhibitor of lipid peroxidation
mediated by active forms of oxygen and was proposed to be a ‘‘super vitamin E’’
(Miki, 1991).
Among the functions of astaxanthin in aquaculture as proposed by Torrissen (1990) and
Shimidzu et al. (1996), antioxidant properties can be closely associated with stress
resistance. Enhancement of resistance to salinity stress (Darachai et al., 1998; Merchie
et al., 1998; Chien et al., 2003), thermal stress (Chien et al., 2003) and oxygen depletion
stress (Chien et al., 1999) in penaeid shrimp postlarvae was associated with an increase in
dietary and body astaxanthin. In those studies, the close relationship between the
antioxidant properties of astaxanthin and stress resistance was indicated by increased
shrimp survival or recovery.
Total antioxidant status (TAS) is an overall indicator of the antioxidant status of an
individual. As the value increases, the antioxidant defense against free radical reaction
increases. Using TAS to detect the actual antioxidant status in crustaceans has been limited
to evaluations of the effects of astaxanthin on thermal and osmotic response (Chien et al.,
2003). No use on ammonia stress of crustaceans has been described.
Superoxide dismutase (SOD), a cytosolic enzyme that is specific for scavenging
superoxide radicals, is involved in protective mechanisms within tissue injury following
oxidative process and phagocytosis. The higher the SOD value is, the more superoxide
radicals need to be reacted. SOD analysis has been widely used in finfish. However, only
few studies of SOD in crustaceans are related to oxidative status (Bell and Smith, 1993),
immunity (Holmblad and Soderhall, 1999; Muñoz et al., 2000) and disease indication
(Neves et al., 2000).
Aspartate aminotransferase (AST) or glutamate oxalate transaminase (GOT) and
alanine aminotransferase (ALT) or glutamate pyruvate transaminase (GPT) are enzymes
involved in the transfer of amino groups from one specific amino acid to another. AST
and ALT activities are usually used as general indicators of the functioning of vertebrate
liver. High AST and ALT generally, but not definitively, indicate the weakening or
damage of normal liver function. The crustacean hepatopancreas is assumed to be
C.-H. Pan et al. / J. Exp. Mar. Biol. Ecol. 297 (2003) 107–118
109
homologous to the mammalian liver and pancreas (Gibson and Barker, 1979) and is
responsible for major metabolic events, including enzyme secretion, absorption and
storage of nutrients, molting and vitellogenesis (Chanson and Spray, 1992). As in other
animals, both ALT and AST are key enzymes for the interconversion of amino acids and
other intermediary metabolites in crustacean and are detected in the hepatopancreas,
muscle and gill (Chaplin et al., 1967). For finfish, AST and/or ALT have been used
extensively in studies that evaluate finfish response to toxins (heavy metal pollutants and
pesticides), stress caused by temperature changes, low oxygen, starvation, pH, ammonia,
nitrite, disease, health, therapeutics monitoring and nutrition. For crustaceans, AST and
ALT have been used only recently to study the effect from pesticide (Galindo-Reyes et al.,
2000), heavy metal pollution (Zhao et al., 1995) and thermal and osmotic stress (Chien et
al., 2003). This study is presumably the first attempt to relate AST and ALT to ammonia
stress in an invertebrate.
Ammonia is the most common toxicant in a culture system and toxic to fish, mollusks
and crustaceans (Colt and Armstrong, 1981). In crustaceans, it is the main end-product of
protein catabolism (Kinne, 1976) and can account for 40 – 90% of nitrogenous excretion
(Parry, 1960). Besides coming from such excretion, ammonia in a culture system is
derived from microbial metabolism of nitrogenous compounds, such as unconsumed feed,
animal feces and dead organisms under low oxygen condition (Armstrong, 1979). When
culture activity intensifies, ammonia concentrations in water increase, ammonia excretion
by aquatic organisms diminishes, levels of ammonia in blood and other tissue increase
(Colt and Armstrong, 1981) and the metabolic pattern also changes (Spotte, 1979). Shortterm exposure of fish and crustacean to high concentrations of ammonia causes increased
gill ventilation, hyperexcitability, loss of equilibrium, convulsions and then death
(Thurston et al., 1981; Maltby, 1995).
Studies on the protection of antioxidants against oxidative damage can be conducted by
pretreating the animals with antioxidants then subjecting them to oxidative stress induced
by oxidants or toxic substances (Shaikh et al., 1999). The objectives of this study were to
test the effects of astaxanthin as an antioxidant in juvenile tiger prawn Penaeus monodon
as indicated by TAS and SOD values as well as the shrimp responses, in terms of survival
rate, AST and ALT following ammonia stress.
2. Materials and methods
2.1. Rearing
Five-day old tiger prawn postlarvae, averaging 6.7 F 1 mg, were reared indoors in
two 500-l fiberglass-reinforced polyethylene tanks at a density of 500 larvae per tank.
Tanks were covered with black screen to discourage algae growth. The shrimp were fed
diets (see Chien et al., 2003) containing either 0 or 71.5 mg astaxanthin kg 1 diet at 5%
of body weight per day divided among three feedings at 08.00, 16.00 and 20.00 h. Tank
bottom debris was removed by siphon daily and about one third of the water was
replaced daily with 1 Am of filtered and ultraviolet-sterilized seawater. Experimental
water conditions: 27– 29 jC, salinity 30– 32, pH 8.2 – 8.3 and DO 5.2 –6.5 mg l 1.
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Ammonia-N and nitrite-N were monitored and kept below safe levels (Chien, 1992).
The highest concentrations observed were 0.012 and 0.004 mg l 1 for ammonia-N and
nitrite-N, respectively. The duration of feeding was 8 weeks. Before and after rearing,
the astaxanthin content of five animals from each tank was analyzed as described in
Chien et al. (2003).
2.2. Stress test
Ammonia test solutions were prepared by dissolving requisite amounts of ammonium
chloride (Merck GR grade) in saltwater. The nominal concentrations of total ammonia-N
were 0.02, 0.2, 2 and 20 mg l 1. The test was conducted using a static renewal procedure
(Hubert, 1980). The shrimp were collected at random from the holding tanks and exposed
to each test solution and a 0-mg ammonia-N l 1 control in triplicate. The trials were
conducted in 2000-ml beakers containing 1600 ml of the test solution. Each beaker
contained 20 shrimp and water was aerated continuously during the trial. Each test
solution was renewed every 24 h (Buikema et al., 1982). Experimental water conditions:
temperature 28 F 0.5 jC, dissolved oxygen 6.5 F 0.5 mg l 1, pH 8.1 F 0.1 and salinity
32.0 F 0.5. The shrimp were fed three times a day based on 6.5% of the body weight.
Observations were made at 12-h intervals to 72 h and the dead shrimp were removed.
After 72 h, the surviving shrimp were sampled and analyzed for TAS concentration and
SOD, AST and ALT activities. This protocol was conducted for control and astaxanthinfed (AX) shrimp.
2.3. Haemolymph biochemistry
Haemolymph of the post-stress shrimp was drawn from the pericardial cavity (syringe
25 G 1’’) through the intersegmental membrane between the cephalothorax and the
abdominal segment. The haemolymph sample was prepared by mixing 400 Al of isotonic
NaCl solution containing 0.94 mmol l 1 EDTA with 100 Al haemolymph immediately
after it was drawn. All assays were performed within 5 h of samples or the samples were
chilled if not immediately used for determination of TAS, SOD, AST, ALT and
haemolymph protein.
To measure haemolymph TAS and SOD, 20 and 25 Al of haemolymph sample,
respectively, were used and determined spectrophotometrically at 600 and 505 nm,
respectively, with a U-2000 spectrophotometer (Hitachi, Japan) at 37 jC using Randox
Laboratories kits (Crumlin, Antrim, UK) according to the manufacturer’s instructions.
Activities were expressed in international enzyme units (Ul 1).
AST and ALT activities were determined spectrophotometrically at 340 nm using a U2000 spectrophotometer (Hitachi, Tokyo, Japan) at 37 jC. Each used 100 Al of haemolymph sample using Randox Laboratories kits according to the manufacturer’s instructions. Activities were expressed in international enzyme units (Ul 1).
Soluble protein of 200 Al of the haemolymph sample was determined using a protein
assay kit (No.500-0006, Bio-rad laboratories, Richmond, CA., USA) and BAS (bovine
serum albumin, 66 kDa, Sigma) as a standard using a Bradford (1976) modified
method.
C.-H. Pan et al. / J. Exp. Mar. Biol. Ecol. 297 (2003) 107–118
111
Fig. 1. Survival rate, TAS, SOD, AST and ALT of astaxanthin-fed (AX) and control shrimp P. monodon exposed
to various levels of ammonia for 72 h. Values with letters ‘‘a’’, ‘‘b’’, ‘‘c’’ and ‘‘d’’ indicate significance of
differences among various ammonia levels within shrimp from the same dietary treatment. Letters ‘‘x’’ and ‘‘y’’
indicate significance of differences between AX and control shrimp exposed to the same ammonia level.
Significant level is set at p V 0.05.
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C.-H. Pan et al. / J. Exp. Mar. Biol. Ecol. 297 (2003) 107–118
2.4. Statistical analysis
One-way ANOVA, followed by Duncan’s multiple range test, was conducted to
compare the survival rate and biochemical parameters among various ammonia levels
within each shrimp group. A t-test was conducted to compare the survival rate and
biochemical parameters within each ammonia level between control and AX shrimp.
Since survival data were expressed as percentages, an arcsine square root transformation
was performed before analysis (Sokal and Rohlf, 1995; Ray et al., 1996). Correlation
analyses were used to find out the relationships between survival rate and biochemical
parameters.
3. Results
Shrimp survival rate decreased when ammonia concentration increased (Fig. 1). No
mortality occurred for both control and AX shrimp when no ammonia stress was applied.
AX shrimp had 15 –20% higher survival rates than control shrimp when subjected to 0.02–
2 mg l 1 ammonia stress for 72 h. However, when ammonia concentration increased to 20
mg l 1, no difference in survival rate was found between control and AX shrimp.
No difference in TAS was found in AX shrimp exposed to all levels of ammonia (Fig. 1).
A significant drop in TAS in control shrimp was observed when ammonia concentration
increased from 0.02 to 0.2 mg l 1. No difference in TAS was found between control shrimp
and AX shrimp at 0 and 0.02 mg l 1 ammonia concentrations. AX shrimp had significantly
higher TAS, 6.5 –8.8%, than control shrimp at 0.2 –20 mg l 1 ammonia concentration.
There was a gradual increasing trend in SOD in AX shrimp when ammonia concentration increased (Fig. 1). SOD in control shrimp jumped 18 times ((0.56 0.03)/0.03)) when
ammonia concentration increased from 0 to 0.02 mg l 1. Except at 0 mg l 1 ammonia, AX
shrimp had significantly lower SOD than control shrimp for all levels of ammonia.
Control shrimp’s AST decreased significantly when ammonia concentration increased
from 0 to 0.02 mg l 1 and remained relatively unchanged despite ammonia concentration
Table 1
Correlation matrix among survival rate and haemolymph antioxidant enzyme activities of P. monodon juveniles
fed diets supplemented with or without 71.5 mg kg 1 astaxanthin for 8 weeks (n = 30)
ALT a
ASTb
e
SUR
TASd
SODc
ASTb
a
0.5505*
0.3831*
0.5403*
Alanine Transaminase.
b
Aspartate Transaminase.
c
Superoxide Dismutase.
d
Total Antioxidative status.
e
Survival rate.
* Significant level of correlation coefficient r is at p V 0.05.
SODc
0.6071*
0.6651*
TASd
0.6754*
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113
further increased (Fig. 1). No trend in AST in AX shrimp was found when ammonia
concentration increased. Control shrimp always had higher AST than AX shrimp at all
ammonia concentrations, even at 0 mg l 1.
No trend in ALT in both control and AX shrimp was found when ammonia concentration
increased (Fig. 1). Unlike AST, ALT in AX shrimp was either lower than or equal to that in
control shrimp under various levels of ammonia. There was no difference in ALT between
control and AX shrimp when ammonia concentrations were 0.2 and 2 mg l 1.
The results of correlation analysis (Table 1) showed that TAS not only had highly
significant positive correlation with survival rates but also had correlations with the
activities of other three enzymes. SOD had a highly significant negative correlation with
survival rates and TAS. ALT and AST were highly correlated each other positively, had
negative correlations with TAS, but were not correlated with survival rates.
4. Discussion
4.1. Ammonia stress and SOD and TAS
The mechanisms of toxicity and lethal concentrations of ammonia to various fishes
and crustaceans of commercial importance are relatively well documented (Tomasso,
1994). Ammonia assumes two chemical forms in aqueous solution, the unionized form
(NH3) and the ionized form (NH4+) (Butler, 1964). Only the NH3 is toxic (Hampson,
1976), as it has high lipid solubility and is able to diffuse quite readily across cell
membranes in the direction favored by its pressure gradient (Fromm and Gillette, 1968;
Emerson et al., 1975). When NH3 concentration in water increases, the rate of diffusion
outward from the blood decreases. The result is adverse effects on membrane stability
and enzyme-catalysed reaction, an elevation of haemolymph pH, reduction in the
transport of oxygen, increase in oxygen consumption by tissues (Colt and Armstrong,
1981; Chen et al. 1991) and sometimes gill damage (Schreckenbach and Spangenberg,
1978; Tomasso, 1994), which may eventually lead to death. As ammonia toxicity
ensues, low oxygen availability may result in oxidative stress (Storey, 1996), which is
characterized by cellular damage caused by excessive reactive oxygen species (Sies,
1991). In the present study, when shrimp were exposed to ammonia stress, there would
have been the potential for generation of abnormally high levels of oxygen radicals as
shown in the increase of SOD and decrease of TAS. Few studies have been conducted
on the effects of ammonia stress on SOD and TAS in fish, mollusk and crustacean.
Elevated activities of SOD in tissues or freshwater mussels, Lamellidens marginalis,
under ammonia stress suggest increased detoxification of ammonia, superoxide anions
and peroxides which in turn enhance the tolerance and then the survivability of the
bivalve to polluted ecosystems (Chetty and Indira, 1995). While subjected to 5 min
thermal and/or osmotic stress, there was also a significant increase of SOD and decrease
of TAS in juvenile P. monodon (Chien et al., 2003). Because astaxanthin contains a long
conjugated double bond system with relatively unstable electron orbital, it may scavenge
oxygen radicals in cells (Stanier et al., 1971) and therefore reduce cellular damage and
enhance resistance.
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TAS and SOD responded to body astaxanthin and ammonia stress differently (Fig. 1).
Although SOD had a highly significant ( p V 0.01) negative correlation with TAS (Table 1),
during ammonia stress response, due to its specificity in catalyzing the dismutation of O2
into hydrogen peroxide, it did not proceed with a complete inverse (r = 0.67) relationship
with TAS, an indicator of the status of overall antioxidant defense against reactive oxygen
species and reactive oxygen intermediates. While an organism is first subjected to stress,
SOD should be able to respond accordingly and immediately to the production of
superoxide anion. Therefore, only a slight increase in ammonia concentration from 0 to
0.02 mg l 1 resulted in a jump in SOD in both shrimp; around 18 times of increase for
control shrimp and four times ((0.16 0.03)/0.03) for AX shrimp. However, there was no
change in TAS in both shrimp groups while ammonia concentration increased 0 –0.02 mg
l 1. This is because TAS indicates static potential of antioxidant defense against all
radicals, it may not have a marked change upon the stress if the production of superoxide
anion is insignificant compared to the already existing radicals.
Body astaxanthin improved and stabilized TAS and SOD, enhanced resistance against
ammonia stress and reflected higher survival rate in shrimp. As compared to control
shrimp, TAS in AX shrimp remained relatively high and constant and SOD increased
gradually but stayed low as ammonia concentration increased from 0.02 to 20 mg l 1. On
the contrary, in control shrimp, TAS decreased 8.8% ((1.48 1.35)/1.48) and SOD stayed
relatively high and fluctuated between 0.48 and 0.66 Ul 1 as ammonia concentration
increased from 0.02 to 20 mg l 1. Low SOD and high TAS favored shrimp survival as
reflected in a highly negative correlation between SOD and survival rate and a highly
positive correlation between TAS and survival rate.
4.2. ALT and AST
In freshwater teleosts, ALT and AST play an important role in ammonia detoxification
(D’Apollonia and Anderson, 1980). The physiologically toxic NH3 level was neutralized
in the organism by means of the increase of AST and ALT activity (Nemcsok et al., 1982).
The responses of AST and ALT to ammonia stresses in fish can be similar but are not
necessarily parallel to each other. Jeney et al. (1992) observed several fold increases in the
plasma activity of both AST and ALT in carp (Cyprinus carpio L.) exposed to ammonia
for 4 days. Kwon and Chang (1996) also showed that both AST and ALT increased when
black seabream Acanthopagrus schlegeli were exposed to a high level of ammonia and
both returned to the normal status during the recovery period. However, plasma AST
activity in rainbow trout (Oncorhynchus mykiss) increased after 4 days of exposure to high
ammonia concentration, but plasma ALT activity had no change (Vedel et al., 1998). In our
study, there was neither increasing nor decreasing trend in both AST and ALT in both
shrimp groups when ammonia concentration was higher than 0.02 mg l 1. However,
reduction in AST and ALT activity in both control and AX shrimp occurred when
ammonia concentration increased from 0 to 0.02 mg l 1 (Fig. 1).
ALT and AST in fish have been used as indices for the diagnosis of liver function
(Yamamoto, 1981) and damage (Oda, 1990). A study by Nakano et al. (1995) demonstrated for the first time that a dietary astaxanthin supplement had an effect on liver
function and increased defensive potential against oxidative stress of rainbow trout.
C.-H. Pan et al. / J. Exp. Mar. Biol. Ecol. 297 (2003) 107–118
115
Further study by Nakano et al. (1999) indicated that dietary red yeast, Phaffia rhodozyma,
which is rich in astaxanthin, should have a reducing effect on oxidized oil-induced
oxidative stress in rainbow trout. In their study, the levels of serum ALT and AST of fish
increased significantly after the fish were fed oxidized oil. The supply of red yeast
considerably decreased ALT and AST. Chien et al. (2003) reported the positive effect of
astaxanthin on juvenile P. monodon hepatopancreas, which was indicated by the lowering
of either AST or ALT under osmotic or thermal stress, respectively. Similar phenomenon
occurred in this study that at all ammonia concentrations, including the blank, AX shrimp,
had its AST always lower than control shrimp and its ALT lower than or equal to control
shrimp (Fig. 1). Such unparallel effects of astaxanthin on AST and ALT were also
observed by Nakano et al. (1995) who reported that AST activities of fish fed a diet
containing astaxanthin were significantly lower than those of the control fish. However,
ALT did not decrease simultaneously with AST. Although AST and ALT were highly
correlated to each other positively (Table 1), AST appeared more sensitive than ALT in
responding to stresses and astaxanthin in this and other studies.
In this study, AST and ALT had negative correlations with TAS, further indicated, AST
and ALT were directly or indirectly related to oxidant metabolites so that they could serve
as indicators of oxidative status (Chien et al., 2003). Survival rates having no correlations
with AST and ALT may indicate that shrimp’s mortality was not related to damage to the
hepatopancreas, if only it occurred.
4.3. Stress test and health
This study provided objective evidence that astaxanthin improved shrimp health in
terms of resistance against ammonia stress. Cavalli et al. (2000) concluded that short-term
ammonia toxicity tests a sensitive criterion for the evaluation of larval quality. In our study,
ammonia stress at a concentration as low as 0.02 mg l 1 for 72 h still differentiated the
shrimp supplemented with astaxanthin for 8 weeks from the ones without by their survival
rate. Mainly recognized as an antioxidant, however, astaxanthin protected shrimp from
even a slight stress in this study. Enhancement of resistance in penaeid shrimp postlarvae
to oxygen depletion stress (Chien et al., 1999), salinity stress (Darachai et al., 1998;
Merchie et al., 1998; Chien et al., 2003), thermal stress (Chien et al., 2003), and now in
this study, ammonia stress was found associated with an increase in dietary and body
astaxanthin. The lack of difference in survival rate between control and AX shrimp
exposed to 20 mg l 1 ammonia for 72 h could have been due to ammonia stress being too
intense and too long and beyond the range of physiological tolerance.
In conclusion, the enhancement of antioxidation capacity by dietary astaxanthin, and
consequently, the improvement of survival rate against ammonia stress, suggests that for
tiger prawn, astaxanthin is a survival enhancer, which can become critical particularly
when the animal is under ammonia stress.
Acknowledgement
This work was supported by the National Science Council Project No. NSC 91-2313B019-010. [SS]
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References
Armstrong, D.A., 1979. Nitrogen toxicity to crustacea and aspects of its dynamics in culture systems. In: Lewis,
D., Liang, J. (Eds.), 2nd Biennial Crustacean Health Workshop, Texas A and M Sea Grant, TAMM-SE-79114, Texas, pp. 329 – 360.
Bell, K.L., Smith, V.J., 1993. In vitro superoxide production by hyaline cells of the shore crab Carcinus maenas
(L.). Dev. Comp. Immunol. 17, 211 – 219.
Bradford, M.M., 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein using
the principle of protein-dye binding. Anal. Biochem. 72, 248 – 254.
Buikema Jr., A.L., Niedertehner, R.R., Cairns Jr., J., 1982. Biological monitoring, Part IV. Toxicity testing. Water
Res. 16, 239 – 262.
Butler, J.N., 1964. Ionic Equilibrium. Addison-Wiley Publishing, Reading, MA, USA.
Cavalli, R.O., Berghe, E.S., Lavens, P., Thuy, N.T.T., Wille, M., Sorgeloos, P., 2000. Ammonia toxicity as a
criterion for the evaluation of larval quality in the prawn Macrobrachium rosenbergii. Comp. Biochem.
Physiol., Part C 125, 333 – 343.
Chanson, M., Spray, D.C., 1992. Gating and single channel properties of gap junction channels in hepatopancreatic cells of Procambarus clarkii. Biol. Bull. 183, 341 – 342 (Marine Biological Laboratory, Woods Hole).
Chaplin, A.E., Hugginst, A.K., Aunday, K.A., 1967. The distribution of L-a-aminotransferases in Carcinus
maenas. Comp. Biochem. Physiol. 20, 195 – 198.
Chen, J.C., Nan, F.H., Kuo, C.M., 1991. Oxygen consumption and ammonia-N excretion of prawns (Penaeus
chinensis) exposed to ambient ammonia. Arch. Environ. Contam. Toxicol. 21, 377 – 382.
Chetty, A.N., Indira, K., 1995. Free radical toxicity in a fresh water bivalve, Lamellidens marginalis under
ambient ammonia stress. J. Environ. Biol. 16, 137 – 142.
Chew, B.P., 1995. Antioxidant vitamins affect food animal immunity and health. J. Nutr. 125, 1804S – 1808S.
Chien, Y.H., 1992. Water quality requirements and management for marine shrimp culture. In: Wyban, J. (Ed.),
Proceedings of the Special Session on Shrimp Farming. World Aquaculture Society, Baton Rouge, LA, USA,
pp. 144 – 155.
Chien, Y.H., Chen, I.M., Pan, C.H., Kurmaly, K., 1999. Oxygen depletion stress on mortality and lethal course of
juvenile tiger prawn Penaeus monodon fed high level of dietary astaxanthin. J. Fish. Soc. Taiwan 26, 85 – 93.
Chien, Y.H., Pan, C.H., Hunter, B., 2003. The resistance to physical stresses by Penaeus monodon juveniles fed
diets supplemented with astaxanthin. Aquaculture 216, 177 – 191.
Colt, J.E., Armstrong, D.A., 1981. Nitrogen toxicity to crustaceans, fish, and molluscs. In: Allen, L.J., Kinney,
E.C. (Eds.), Proceedings of the Bio-Engineering Symposium for Fish Culture. Fish Culture Section, American
Fisheries Society. Northeast Society of Conservation Engineers, Bethesda, MD, pp. 34 – 47.
D’Apollonia, S., Anderson, P.D., 1980. Optimal assay conditions for serum and liver glutamate oxalacetate
transaminase, glutamate pyruvate transaminase, and sorbitol dehyddrogenase from the rainbow trout, Salmo
gairdneri. Can. J. Fish. Aquat. Sci. 37, 163 – 169.
Darachai, J., Piyatiratitivorakul, S., Kittakoop, P., Nitithamyong, C., Menasveta, P., 1998. Effects of Astaxanthin
on larval growth and survival of the giant tiger prawn, Penaeus monodon. In: Flegel, T.W. (Ed.), Advances in
Shrimp Biotechnology. National Center for Genetic Engineering and Biotechnology, Bangkok, pp. 117 – 121.
Emerson, K., Russo, R.C., Lund, R.E., Thurston, R.V., 1975. Aqueous ammonia equilibrium calculations: effect
of pH and temperature. J. Fish. Res. Board Can. 32, 2379 – 2388.
Fromm, P.O., Gillette, J.R., 1968. Effect of ambient ammonia on blood ammonia and nitrogenexcretion of
rainbow trout (Salmo gairdneri). Comp. Biochem. Physiol. 26, 887 – 896.
Galindo-Reyes, J.G., Venezia, L.D., Lazcano-Alvarez, G., Rivas-Mendoza, H., 2000. Enzymatic and osmoregulative alternations in white shrimp Litopenaeus vannamei exposed to pesticides. Chemosphere 40, 233 – 237.
Gibson, R., Barker, P.L., 1979. The decapod hepatopancreas. Oceanogr. Mar. Biol. 17, 285 – 346.
Hampson, B.L., 1976. Ammonia concentration in rainbow trout rearing experiment in a closed freshwater –
seawater system. Aquaculture 9, 61 – 70.
Holmblad, T., Soderhall, K., 1999. Cell adhesion molecules and antioxidative enzymes in a crustacean; possible
role in immunity. Aquaculture 172, 111 – 123.
Hubert, J.J. (Ed.), 1980. Bioassay. Kendall Hunt Publishing, Toronto, Ontario, Canada. 164 pp.
Jeney, G., Nemcsok, J., Jeney, Z.S., Olah, J., 1992. Acute effect of sublethal ammonia concentrations on common
C.-H. Pan et al. / J. Exp. Mar. Biol. Ecol. 297 (2003) 107–118
117
carp (Cyprinus carpio L.). 2. Effect of ammonia on blood plasma transaminases GOT, GPT, G1DH enzyme
activity, and ATP value. Aquaculture 104, 149 – 156.
Kinne, O., 1976. Cultivation of marine organisms: water quality management and technology. In: Kinne, O.
(Ed.), Marine Ecology, vol. III, part 1. Wiley-Interscience, New York, pp. 79 – 300.
Kwon, J.Y., Chang, Y.J., 1996. Effects of ammonia concentration on histological and physiological status in black
seabream Acanthopagurs schlegeli. J. Korean Fish. Soc. 29, 828 – 836.
Maltby, L., 1995. Sensitivity of the crustaceans Gammarus pulex (L.) and Astllus aquaticus (L.) to short-term
exposure to hyposia and unionized ammonia: observations and possible mechanism. Water Res. 29, 781 – 787.
Merchie, G., Kontara, E., Lavens, P., Robles, R., Kurmaly, K., Sorgeloos, P., 1998. Effect of vitamin C and
astaxanthin on stress and disease resistance of postlarval tiger shrimp Penaeus monodon (Fabricius). Aquac.
Res. 29, 579 – 585.
Miki, W., 1991. Biological functions and activities of animal carotenoids. Pure and Appl. Chem. 63, 141 – 146.
Muñoz, M., Cedeno, R., Rodriguez, J., van der Knaap, W.P.W., Mialhe, E., Bachere, E., 2000. Measurement
of reactive oxygen intermediate production in haemocytes of the penaeids shrimp, Penaeus vannamei.
Aquaculture 191, 89 – 107.
Nakano, T., Tosa, M., Takeuchi, M., 1995. Improvement of biochemical features in fish health by red yeast and
synthetic astaxanthin. J. Agric. Food Chem. 43, 1570 – 1573.
Nakano, T., Kanmuri, T., Sato, M., Takeuchi, M., 1999. Effect of astaxanthin rich red yeast (Phaffia rhodozyma)
on oxidative stress in rainbow trout. Biochim. Biophys. Acta 1426, 119 – 125.
Nemcsok, J., Gyore, K., Olah, J., Boros, L., 1982. Effect of NH3 on blood glucose level, GOT, GPT, LDH
enzyme activity and respiration of three fish species. Aquacult. Hung. 3, 63 – 68.
Neves, C.A., Santos, E.A., Bainy, A.C.D., 2000. Reduced superoxide dismutase activity in Palaemonetes
argentinus (Decapoda, Palemonidae) infected by Probopyrus ringueleti (Isopoda, Bopyridae). Dis. Aquat.
Org. 39, 155 – 158.
Oda, T., 1990. The Biology of Liver University of Tokyo Press, Tokyo. 140 pp. (in Japanese).
Parry, G., 1960. Excretion. In: Waterman, T.H. (Ed.), The Physiology of Crustacea, vol. 1. Academic Press, New
York, pp. 341 – 366.
Ranby, B., Rabek, J.E., 1978. In: Ranby, B., Rabek, J.E. (Eds.), Singlet Oxygen. Wiley, Chichester, England.
331 pp.
Ray, W.M., Chien, Y.H., Chen, C.H., 1996. Comparison of probit analysis versus arcsine square root transformation on LC50 estimation. Aquac. Eng. 15, 193 – 207.
Schreckenbach, K., Spangenberg, K., 1978. pH-wert-abhängige Ammoniakvergiftung bei Fischen und Möglichkeiten ihrer Beeinflussung. Z. Binnenfisch. DDR 25, 299 – 314.
Shaikh, Z.A., Vu, T.T., Zaman, K., 1999. Oxidative stress as a mechanism of chronic cadmium-induced hepatotoxicity and renal toxicity and protection by antioxidants. Toxicol. Appl. Pharmacol. 154, 256 – 263.
Shimidzu, N., Goto, M., Miki, W., 1996. Carotenoids as singlet oxygen quenchers in marine organisms. Fish. Sci.
62, 134 – 137.
Sies, H., 1991. Oxidative stress: from basic research to clinical application. Am. J. Med. 91 (3C), 31S – 38S.
Sokal, R.R., Rohlf, F.J., 1995. Biometry, 3rd ed. Freeman, W.H., San Francisco. 887 pp.
Spotte, S., 1979. Fish and Invertebrate Culture: Water Management in Closed Systems, 2nd ed. John Wiley, New
York. 179 pp.
Stanier, R.Y., Kunizawa, M.M., Cohen-Bazire, G., 1971. Purification and property of unicellular blue-green algae
(Order Chroococales). Bacteriol. Rev. 35, 120 – 171.
Storey, K.B., 1996. Oxidative stress: animal adaptations in nature. Brasil J. Med. Biol. Res. 29, 1715 – 1733.
Thurston, R.V., Russo, R.C., Vinogradov, G.A., 1981. Ammonia toxicity to fishes: effect of pH on the toxicity of
the unionized ammonia species. Environ. Sci. Technol. 15, 837 – 840.
Tomasso, J.R., 1994. Toxicity of nitrogenous wastes to aquaculture animals. Rev. Fish. Sci. 2, 291 – 314.
Torrissen, O.J., 1990. Biological activities of carotenoids in fishes. The current status of fish nutrients in aquaculture. In: Takeda, M., Watanabe, T. (Eds.), Proceedings of The Third International Symposium on Feeding
and Nutrition in Fish. Tokyo University of Fisheries, Tokyo, Japan, pp. 387 – 399.
Vedel, N.E., Korsgaard, B., Jensen, F.B., 1998. Isolated and combined exposure to ammonia and nitrite in
rainbow trout (Oncorhynchus mykiss): effects electrolyte status, blood respiratory properties and brain glutamine/glutamate concentration. Aquat. Toxicol. 41, 325 – 342.
118
C.-H. Pan et al. / J. Exp. Mar. Biol. Ecol. 297 (2003) 107–118
Yamamoto, Y., 1981. Determination of toxicity by biochemical method. In: Egami, N. (Ed.), Fishes as Laboratory
Animals. Soft Science, Tokyo. Chap. 4, 568 pp.
Yu, B.P., 1994. Cellular defenses against damage from reactive oxygen species. Physiol. Rev. 74, 139 – 162.
Zhao, W., Wei, H., Jia, J., Lu, D., 1995. Effects of cadmium on transaminase activities and structures of tissues in
freshwater giant prawn (Macrobrachium rosenbergii). J. Fish. China 19, 21 – 27.