Aquaculture 250 (2005) 765 – 774

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Effect of dietary taurine levels on growth and feeding behavior

of juvenile Japanese flounder Paralichthys olivaceus
Shin-Kwon Kima, Toshio Takeuchia,*, Masahito Yokoyamab, Yuko Muratac,
Masaki Kaneniwad, Yoshitaka Sakakurae
a
Graduate School of Marine Science and Technology, Tokyo University of Marine Science and Technology, Minato, Tokyo 108-8477, Japan
b
National Research Institute of Aquaculture, Tamaki, Mie 519-0423, Japan
c
National Research Institute of Fisheries Science, Yokohama, Kanagawa 236-8648, Japan
d
Japan International Research Center for Agriculture Sciences, Tsukuba, Ibaraki 305-8686, Japan
e
Faculty of Fisheries, Nagasaki University, Nagasaki 852-8521, Japan
Received 20 January 2005; received in revised form 22 April 2005; accepted 26 April 2005

Abstract

This study was conducted to investigate the effect of dietary taurine levels on growth and feeding behavior of juvenile
Japanese flounder. Three different taurine level diets were prepared by supplementation of taurine (T-0%, 0.5% and 1.5%) to the
basal diet. Fish meal washed with 70% ethanol to remove taurine was used as the sole protein source. Feeding experiments were
carried out twice at 20 8C by using different size of fish (average body weight: 0.3 g in Experiment I and 3.7 g in Experiment
II). The feeding behavior of fish was observed throughout the experimental period. At the end of experiments, fish were killed
for amino acids analysis.
The final average body weight and feed efficiency of juvenile Japanese flounder fed the T-1.5% diet was significantly higher
than those of fish fed the T-0% diet in Experiments I and II. Abnormal feeding behavior such as multiple feeding while
swimming in the water column was observed in the T-0% group in Experiment I. These findings indicate that taurine is essential
for normal growth and development of normal feeding behavior of juvenile Japanese flounder.
D 2005 Elsevier B.V. All rights reserved.

Keywords: Feeding behavior; Growth; Japanese flounder; Juvenile; Taurine

1. Introduction senting one of the most important species of marine

In Japan, 20 million hatchery-reared juveniles of
the Japanese flounder are released each year, repre-
* Corresponding author. Tel./fax: +81 3 5463 0545.
E-mail address: take@s.kaiyodai.ac.jp (T. Takeuchi).
0044-8486/$ - see front matter D 2005 Elsevier B.V. All rights reserved.
doi:10.1016/j.aquaculture.2005.04.073
stock enhancement in this country (Masuda and Tsu-
kamoto, 1998; Tanaka et al., 1998). However in most
cases, lower survival has been observed than expected.
Furuta (1996, 1998) suggested that mortality due to
predation is higher in hatchery-reared juveniles than in
wild juveniles. In the natural environment, mysids
766 S.-K. Kim et al. / Aquaculture 250 (2005) 765–774
were formed to the most important food organisms for
juvenile flounder after settlement (Hirota et al., 1990),
and the abundance of mysids affected the growth of
flounder juveniles in the nursery grounds (Tanaka,
1993; Furuta, 1996). Live mysids were found to be a
higher quality of food than formula feed for juvenile
flounder (Sekai et al., 1997). Taurine is the most
abundant free amino acid in mysids (Park et al.,
2000). Mysids contain about 30 mg/g of taurine.
This is a very large amount (up to 4- to 10-fold the
amount) compared with rotifer, Artemia or artificial
diets (Park et al., 2000).
Taurine (2-aminoethane sulfonic acid) is not incor-
porated into protein and exists as a free amino acid in
tissues of mammals. Taurine is synthesized from me-
thionine via cystine by a series of enzymatic reactions.
The various potential pathways of taurine synthesis
are reviewed elsewhere (Oja and Kontro, 1983). The
predominant route of synthesis varies among species
and depends on the type of tissue. However, the
enzyme cysteine sulfinate decarboxylase appears to
be the rate-limiting step in taurine biosynthesis in
many mammalian species (Jacobsen and Smith,
1968). In mammals, taurine is present in high concen-
tration in plasma and cells, plays an important role in
several biological processes such as the development
of the central nervous system (CNS) and the retina,
calcium modulation, membrane stabilization, bile
acid-conjugation, reproduction, and immunity (Stur-
man, 1988, 1993; Huxtable, 1992; Sturman and Ches-
ney, 1995). A dietary deficiency of taurine in cats
results in malfunctioning of a number of systems,
including the visual system resulting in decreases in
visual acuity, electroretinogram, and visual evoked
responses along with degeneration of the retina and
tapetum lucidum, the biological mirror behind the
retina (Sturman et al., 1982; Jacobson et al., 1987;
Morris et al., 1990). In addition spontaneous resorp-
tion, abortion, and stillbirths are frequent, and surviv-
ing offspring have abnormal development (Sturman et
al., 1987; Palackal et al., 1988; Sturman and Messing,
1991). Cats have low activity of cysteine sulfinate
decarboxylase, the rate-limiting enzyme for taurine
biosynthesis, and are, therefore, dependent on a die-
tary source of taurine (Knopf et al., 1978; De La Rosa
and Stipanuk, 1985; Sturman et al., 1986). Human
infants, children, and adults who were fed totally with
parenteral nutrition (via intravenous administered
fluids) had reduced concentrations of taurine in their
plasma, since such solutions did not contain taurine
(Geggel et al., 1985; Vinton et al., 1987; Zelikovic et
al., 1990). The role of taurine in mammals has been
extensively studied and reviewed in Sturman and
Chesney (1995).
In fish, it is known that cysteine sulfinate decar-
boxylase activity differs among fish species and he-
patic cysteine sulfinate decarboxylase activity of
flounder is low (Yokoyama et al., 2001). Park et al.
(2001) reported that juvenile flounder fed a taurine-
supplemented fish meal diet showed significantly bet-
ter growth than those fed a non-supplemented fish
meal diet. We also demonstrated that jack mackerel
meal contained about 2.3 mg/g of taurine (Kim et al.,
2003). This study was conducted to investigate the
effects of taurine deficiency and determine the effect
of dietary taurine on growth and feeding behavior of
juvenile Japanese flounder.
2. Materials and methods
2.1. Experimental diets
The formulation and percentage of crude protein
and lipid in the experimental diets are shown in
Table 1. Jack mackerel meal contained about 6
mg/g of taurine. So the fish meal was washed 3
times with 70% ethanol to remove the taurine. The
washing treatment, likely removed other compo-
nents, such as FAA, IMP (inosine 5V-monopho-
sphate) and inosine were analyzed in both washed
fish meal and non-washed fish meal using amino
acid analysis and high performance liquid chroma-
tography (HPLC) (Tsuchimoto et al., 1985). Based
on these analyses, attractants were added to diets to
adjust their contents to the level of non-washed fish
meal (Table 2). Three experimental diets were pre-
pared using the washed-fish meal as the primary
protein source and supplemented with 0%, 0.5%
and 1.5% taurine (Wako Pure Chemical Industries,
Ltd., Osaka, Japan). All ingredients were mixed
with distilled water, to make a mash, pelleted with
a press machine and freeze dried for 24 h. The
content (mg/g) of free amino acids in the diets are
shown in Table 3. Crude protein and lipid content
were adjusted to 55% and 10% in the experimental
 S.-K. Kim et al. / Aquaculture 250 (2005) 765–774 767

Table 1 Table 3
Composition and crude protein and lipid contents of the experimen- Free amino acid composition (dry basis mg/g diet, mean F S.D.,
tal diets n = 5) of the experimental diets
Ingredients Ta-0% T-0.5% T-1.5% Amino acid T-0% T-0.5% T-1.5%
Jack mackerel meal (washed) 74.0 74.0 74.0 Essential amino acid
Attractantb 0.8 0.8 0.8 Arginine 0.09 F 0.00 0.09 F 0.00 0.09 F 0.00
Taurine 0.0 0.5 1.5 Lysine 0.15 F 0.00 0.15 F 0.00 0.14 F 0.00
a-Starch 8.0 8.0 8.0 Histidine 4.66 F 0.07 4.22 F 0.47 4.86 F 0.63
Dextrin 5.0 5.0 5.0 Phenylalanine 0.06 F 0.00 0.05 F 0.00 0.06 F 0.00
Cellulose 1.9 1.4 0.4 Tyrosine 0.00 F 0.00 0.00 F 0.00 0.00 F 0.00
n 3 HUFAc 4.2 4.2 4.2 Leucine 0.12 F 0.01 0.11 F 0.01 0.10 F 0.00
Mineral mixtured 4.0 4.0 4.0 Isoleucine 0.11 F 0.00 0.11 F 0.01 0.10 F 0.01
Vitamin mixturee 1.5 1.5 1.5 Methionine 0.14 F 0.00 0.12 F 0.01 0.13 F 0.01
Choline chloride 0.5 0.5 0.5 Valine 0.25 F 0.01 0.26 F 0.00 0.26 F 0.00
Vitamin Ef 0.1 0.1 0.1 Threonine 0.08 F 0.00 0.08 F 0.02 0.08 F 0.00

Chemical analysis Non-essential amino acid

Crude protein (%) 54.5 F 0.1 55.8 F 0.2 55.4 F 1.3 Taurine 0.56 F 0.00 5.56 F 0.66 16.00 F 2.92
Crude lipid (%) 9.8 F 0.1 10.5 F 0.1 9.8 F 0.1 Cystathionine 0.23 F 0.00 0.25 F 0.01 0.24 F 0.02
a
T means taurine supplemented experimental diet. Alanine 1.22 F 0.01 1.25 F 0.05 1.18 F 0.02
b
See Table 2. Glycine 0.35 F 0.00 0.35 F 0.00 0.35 F 0.01
c
n 3 HUFA: n 3 highly unsaturated fatty acids (EPA, 7.6%; Glutamic acid 0.65 F 0.04 0.63 F 0.02 0.64 F 0.01
DHA, 38.5%.Nippon Chemical Feed Co., Ltd., Chiba, Japan). Serine 0.05 F 0.00 0.05 F 0.00 0.05 F 0.00
d
Mineral mixture ingredient (g/100 g): NaCl 1.0 g, MgSO4d 7H2O Aspartic acid 0.06 F 0.00 0.07 F 0.00 0.07 F 0.00
15.0 g, NaH2PO4d 2H2O 25.0 g, KH2PO4 32.0 g, Ca(H2PO4)2d H2O Proline 0.22 F 0.01 0.25 F 0.05 0.26 F 0.01
20.0 g, Fe-citrate 2.5 g, Ca-lactate 3.5g, Trace element mixture 1.0 g.
Trace element mixture ingredient (mg/100 mg): ZnSO4d 7H2O 35.3
mg, MnSO4d 4H2O 16.2 mg, CuSO4d 5H2O 3.1 mg, KIO3 0.3 mg,
CoCl2d 6H2O 0.1 mg, Cellulose 45 mg. diets. Taurine was measured at a level of 0.6 mg/g
e
Vitamin mixture ingredient (mg/118 g): Vitamin B1 900 mg, in T-0% diet, 5.6 mg/g in T-0.5% diet and 16 mg/g
Vitamin B2 1500 mg, Vitamin B6 600 mg, Vitamin B12 1.5 mg, l- in T-1.5% diet, respectively.
Ascorbic acid 75  103 mg, Niacin 6  103 mg, Ca-pantotenate
1500 mg, Inositol 30  103 mg, Biotin 90 mg, Folic acid 225 mg,
p-Aminobenzoic acid 750 mg, Vitamin K3 750 mg, Vitamin A 2.2. Experimental fish
600 000 IU, Vitamin D3 600 000 IU.
f
dl-a-Tocopheryl acetate, purity 50% (Nippon Roche, Tokyo, Two feeding experiments were conducted at the
Japan). Where appropriate, data are the mean F S.D. (n = 5). National Research Institute of Fisheries Science,
Yokohama, Japan. Juvenile Japanese flounder (Para-
Table 2 lichthys olivaceus) were obtained from the Nisshin
Attractant composition used in the experimental diets
Marin Tech, Aichi, Japan. Before starting the experi-
Attractant ment, juvenile Japanese flounder were fed a commer-
Trace free amino acids cial diet for 5 days in Experiment I (Higashimaru,
Histidine 375.2 Kagoshima, Japan) and for 33 days in Experiment II
Alanine 105.8
Glycine 25.2
(Nippon Formula Feed, Ehime, Japan). Experiment I
Glutamic acid 45.1 was conducted using two replicate tanks per dietary
Proline 25.9 treatment. Each tank conformed 30 fish (average body
weight 0.3 g) (60-l aquaria, 60  35  30 cm3). In
Nucleotide-related compounds Experiment II, replicates of 10 fish (average body
Inosine 135.4
IMPa 87.4
weight 3.7 g) were placed randomly into each of six
Total 800 mg/100 g in diet 60-l aquaria. Fish in two tanks in each experiment
All attractants are produced from Wako Pure Chemical Industries,
were fed one of the three experimental diets, T-0%, T-
Ltd., Osaka, Japan. 0.5%, T-1.5%, respectively. Fish were fed three times
a
IMP means inosine 5V-monophosphate. a day to satiation for 4 weeks at a water temperature of
768 S.-K. Kim et al. / Aquaculture 250 (2005) 765–774

20 8C. At the end of the feeding trial, fish were
weighed and stored at 80 8C for free amino acid
analysis, and three fish from each aquaria were fed the
respective experimental diet for an additional 2 days.
These fish were placed in separate plastic flasks con-
taining 1-l of water at 20 8C and starved for 24 h.
After 24 h, water samples from each plastic flask were
collected to determine taurine excretion using the
method of Yokoyama and Nakazoe (1991). In these
experiments, the trials were conducted for 4 weeks,
because in a preliminary experiment more than 50%
of the fish fed T-0% died after 5–6 weeks.
2.3. Free amino acids analysis
Frozen fish from each group were dissected and the
brain, liver, eyes and muscle were used for free amino
acid (FAA) analysis. Other frozen fish were used for
FAA analysis of whole body. The whole body and
tissues (n = 5 in Experiment I; n = 3 in Experiment II)
were homogenized with 2% sulfosalicylic acid and
centrifuged at 2300g for 15 min at 5 8C. For deter-
mination of taurine excretion, water samples were
deproteinized by the addition of 10% sulfosalicylic
acid, followed by centrifugation at 2300g for 15 min
at 5 8C. Free amino acid levels were determined
individually with an automatic amino acid analyzer
(Model L-8500A, Hitachi, Tokyo, Japan). Statistical
analysis of growth performances and FAA concentra-
tion in fish fed the three T-0%, T-0.5% and T-1.5%
diets are performed using one-way ANOVA and
Tukey’s multiple-range test.
2.4. Feeding behavior observation
Aquaria were used for 4 weeks in Experiment I and
were set up indoors under 12L:12D light conditions.
Filtered seawater was added to a depth of 40 cm and
the water in the aquaria was exchanged at 850 ml/min.
Fish feeding behavior was recorded for about 7 min
each morning for four days using a CCD camera
(Matsushita Electric Industrial Co., Ltd., Osaka,
Japan). The video recordings were analyzed to deter-
mine the effects of the different taurine diets on
feeding behavior using the classification method of
Miyazaki et al. (2000). Feeding behavior was identi-
fied as one of four specific patterns (A, B, C and D)
as: fish consumed the diet and then returned immedi-
ately to the initial position (A); fish returned in the
same direction but not close to the initial position (B);
fish returned in a different direction after feeding (C);
fish fed several times in a single off-bottom behavior
(D). Feeding behavior was observed 30 times in T-0%
group, 28 times in T-0.5% group and 30 times in
T1.5% group, respectively. Differences in the feeding
behavior duration in the three experimental diets
groups were compared by the v 2 test.
3. Results
3.1. Effects of dietary taurine on growth
The growth of juvenile Japanese flounder was
improved with taurine supplementation (Table 4). In

Table 4
Results of the 4-week feeding trial in Experiments I and II1
Kind of diets Taurine content Mean body weight (g) Percent Feed* Mortality
in diet (%) Initial Final gain (%) efficiency (%)
Experiment I
T-0% 0.06 0.3 F 0.1 1.0 F 0.4c 294c 0.74c 1.6
T-0.5% 0.56 0.3 F 0.1 2.0 F 0.8b 656b 0.99b 3.3
T-1.5% 1.60 0.3 F 0.1 2.4 F 0.9a 818a 1.24a 3.3

Experiment II
T-0% 0.06 3.7 F 0.2 11.3 F 2.8b 306b 1.69c 0
T-1.5% 0.56 3.7 F 0.2 14.5 F 2.4a 393a 1.86b 5
a, b, c
Means with different superscripts within the same column are significantly different ( P b 0.05).
1
Water temperature 20.0 F 0.2 8C.
* Feed efficiency ratio formula: weight gain/the dry weight of diet consumed.
 S.-K. Kim et al. / Aquaculture 250 (2005) 765–774 769

Experiment I, the final average body weights of juve- T-0% (1.0 g). The final body weight, percent gain and
nile flounder fed the T-1.5% (2.4 g) and T-0.5% (2.0 feed efficiency ratio were affected by the taurine
g) diets were significantly higher than that of fish fed content of the experimental diets. In Experiment II,

Brain Liver
25 0.8 40 1.5

Cystathionine content (µmol/g)

Cystathionine content (µmol/g)
a a
20
Taurine content (µmol/g)

Taurine content (µmol/g)

0.6 30
1.0
15 A b
b 0.4 20 A
10
0.5
B 0.2 10 B
5 B
C
c c
0 0.0 0 0.0
T-0% T-0.5% T-1.5% T-0% T-0.5% T-1.5%

Muscle Eyes
60 8 0.8
a
Cystathionine content (µmol/g)

Cystathionine content (µmol/g)

12 a
Taurine content (µmol/g)

Taurine content (µmol/g)

6 0.6
40 A
8
B 4 A b 0.4

20 b B
4
2 C 0.2

c C c
0 0 0 0.0
T-0% T-0.5% T-1.5% T-0% T-0.5% T-1.5%

Whole body
40 4
Cystathionine content (µmol/g)

35
a
Taurine
Taurine content (µmol/g)

30 3
25 Cystathionine
A
20 2
A
15 b

10 1
B
5 c
0 0
Initial T-0% T-0.5% T-1.5%

Fig. 1. Taurine and cystathionine content in the brain, liver, muscle, eyes, whole body and initial whole body of juvenile Japanese flounder in
Experiment I. Means (n = 5) with different superscripts within the same column are significantly different ( P b 0.05).
770 S.-K. Kim et al. / Aquaculture 250 (2005) 765–774

Brain Liver
20 1.2 30 1.0

Cystathionine content (µmol/g)

Cystathionine content (µmol/g)

a 1.0 25 a
0.8
Taurine content (µmol/g)

Taurine content (µmol/g)

15 b
0.8 20 b
0.6
10 0.6 15
0.4
0.4 10
c
5
0.2
0.2 5
c
0 0.0 0 0.0
Initial T-0% T-0.5% T-1.5% Initial T-0% T-0.5% T-1.5%

Muscle Eyes
40 8 12 0.6
a
Cystathionine content (µmol/g)

Cystathionine content (µmol/g)

10 a
Taurine content (µmol/g)

Taurine content (µmol/g)

30 6
8 bA 0.4
A
20 4 6 B
bB
4 0.2
c C
10 2
2
c
C
0 0 0 0.0
Initial T-0% T-0.5% T-1.5% Initial T-0% T-0.5% T-1.5%

Whole body
40 5
Cystathionine content (µmol/g)

a
4
Taurine content (µmol/g)

30 Taurine

3 Cystathionine
A
20
b 2
B
10
1
c
C
0 0
Initial T-0% T-0.5% T-1.5%

Fig. 2. Taurine and cystathionine content in the brain, liver, muscle, eyes and whole body of initial and final juvenile Japanese flounder in
Experiment II. Means (n = 5) with different superscripts within the same column are significantly different ( P b 0.05).
 S.-K. Kim et al. / Aquaculture 250 (2005) 765–774 771

Table 5 Experiment I are shown in Fig. 1. The taurine content

Taurine and NH3 excretion of Japanese floundera of whole body and tissues of the T-1.5% group were
Kind of diets 17-fold in brain, 16-fold in liver, 34-fold in muscle,
T-0% T-0.5% T-1.0% 16-fold in eyes and 17-fold in the whole body higher
Experiment I than those of the T-0% group. The cystathionine
Taurine ND ND ND content in the whole body and tissues of juvenile
NH3 9.3 F 2.7 8.7 F 2.4 8.4 F 2.7 Japanese flounder decreased with increasing dietary
Experiment II
taurine level.
Taurine ND ND ND
7.9 F 2.9 8.2 F 2.5 8.5 F 2.4 3.2.2. Experiment II
Data are the mean F S.D. (n = 3). The taurine content of whole body and tissues of
ND, not detected. the T-1.5% group were 3-fold in brain, 17-fold in
a
Amol/g body weight per 24 h.
liver, 13-fold in muscle, 3-fold in eyes and 12-fold
the final body weight of juvenile flounder was 14.5 g in whole body higher than those of the T-0% group
in the T-0.5% and T-1.5% groups, respectively. The (Fig. 2). The cystathionine content in the whole body
weight gain of juvenile flounder fed the T-0% diet was and muscle of juvenile Japanese flounder decreased
significantly lower than that of the other experimental with increasing dietary taurine level. Dietary taurine
groups. Feed efficiency ratio of the T-0% group was content had no apparent effect on the content of
significantly lower compared to the T-0.5% and T- cystathionine in brain, liver and eyes. In Experiments
1.5% groups. In Experiment II, the growth of juvenile I and II, taurine excretion was not detected in among
flounder was not significantly different between the T- group (Table 5).
0.5% and T-1.5% groups.
3.3. Effects of dietary taurine on feeding behavior in
3.2. Effects of dietary taurine on taurine accumulation Experiment I
in tissues, whole body and on excretion
The patterns of feeding behavior are shown in Fig.
3.2.1. Experiment I 3. The frequencies of types A and B were very high in
The taurine and cystathionine content in brain, the T-0.5% and T-1.5% groups. The frequencies of
liver, muscle, eyes and whole body of fish from types C and D were very high in the T-0% group. The

100%

80%
Frequency (%)

C+D
60%

A+B
40%

20%

0%
TAU-0% TAU-0.5% TAU-1.5%

Fig. 3. Composition of the feeding patterns observed in 7 min. Types A, B, C and D in Experiment I. See explanation of feeding types in
Miyazaki et al. (2000).
772 S.-K. Kim et al. / Aquaculture 250 (2005) 765–774
Fig. 4. The feeding action of juvenile Japanese flounder in T-0%
and T-1.5% groups of Experiment I.
frequency of feeding behavior types A and B was
80% in the T-1.5% group, while in the that of the T-
0% group types A and B was only 20% (Fig. 4).
4. Discussion
In this study, taurine supplementation was found to
be related to growth and feed efficiency ratio of
juvenile Japanese flounder, indicating that the dietary
taurine requirements change ontogenetically. In Ex-
periment I, weight gain and feed efficiency ratio were
improved with the increase on dietary taurine. In
Experiment II, weight gain was not significantly dif-
ferent between the T-1.5% and the T-0.5% group, but
feeding performances were higher than those of the T-
0% group. This result indicates that the early stage
juvenile Japanese flounder of Experiment I (initial
size 0.3 F 0.1 g) require taurine in their diets but
this requirement is reduced for the larger juvenile of
Experiment II (initial size 3.7 F 0.2 g). The beneficial
effects of dietary taurine have been demonstrated in
many species. Conceicao et al. (1997) suggested a
correlation between taurine levels and the growth of
turbot larvae. Park et al. (2000) reported that juvenile
Japanese flounder fed a mysid meal diet contained
higher levels of taurine (above 20 mg/g) than juvenile
Japanese flounder fed a fish meal diet. Park et al.
(2001) also suggested that the taurine content in the
diet affects growth and metabolism of sulfur contain-
ing amino acids in juvenile Japanese flounder, and
concluded that the taurine requirement is 15–20 mg/g
in the diet. Kim et al. (2003) also reported that taurine
has an important role during the juvenile period (ini-
tial size 0.4 g) of Japanese flounder, but did not
significantly improve growth of fingerling fish (initial
size 14.7 g). This indicates that taurine is a condition-
ally essential amino acid during the ontogenetic de-
velopment of juvenile Japanese flounder.
The taurine content of whole body and tissues
increased with an increase in dietary taurine. In con-
trast, the concentration of cystathionine in the whole
body and tissues decreased with an increase of dietary
taurine. Cystathionine is an intermediate in the trans-
sulfuration pathway from methionine to taurine (Fin-
kelstein and Martin, 1986). Yokoyama et al. (2001)
reported that the activity of cysteine sulfinate decar-
boxylase differs among species and the activity in
flounder is very low. Therefore cystathionine might
accumulate in the whole body and tissues of juvenile
flounder in fish fed a diet without taurine supplemen-
tation and the accumulated cystathionine of these fish
might be used for taurine biosynthesis.
Taurine is synthesized from methionine via cystine
by a series of enzymatic reactions. The enzyme cyste-
ine sulfinate decarboxylase appears to be the rate-lim-
iting step in taurine biosynthesis in many mammalian
species (Jacobsen and Smith, 1968). The activities of
cysteine sulfinate decarboxylase in fetal liver and brain
of humans, monkeys, rabbits, rats, guinea pigs and cats
are lower than in adult tissues (Sturman and Hayes,
1980). This may explain the difference in growth of the
different size fish used in Experiments I and II.
In Experiment I, the T-1.5% group showed 80% of
type A and B feeding patterns, however, the T-0%
juvenile group showed only 20%. This is similar to
results of Furuta (1996, 1998) using live mysids and a
commercial diet. Furuta (1992) reported that fish fed
 S.-K. Kim et al. / Aquaculture 250 (2005) 765–774
live mysids exhibited type A feeding behavior which
was speculated to be important in avoiding predation.
Miyazaki et al. (2000) reported that this swim-up
feeding behavior was divided into the following four
phases: aim, creep, attack and return. As flounder eyes
are on the upper body side surface, they may not be
able to recognize bottom-dwelling predators during the
creep phase. Therefore, flounder juveniles are more
vulnerable to predation when they are in the creep
phase of their feeding action. This suggests that mul-
tiple feeding behavior (type D) is very dangerous and a
behavior liable to predation. This means that taurine is
an essential amino acid for normal feeding behavior
avoided predation in juvenile Japanese flounder.
The taurine content of brain and eyes depended on
the taurine level in the diet. The osmoregulatory role
of taurine may be important in the central nervous
system. In many tissues in marine animals (flounder
and skate erythrocyte, skeletal muscle of smooth dog-
fish and the brain of numerous species), taurine com-
prises more than 50% of the free amino acid pool
(Lombardini et al., 1979). During acclimatization of
marine species to dilute seawater, plasma osmolarity
declines and several tissues lose taurine to maintain a
constant cell volume (Forster and Goldstein, 1979).
Because these animals ingest other marine species
containing large quantities of taurine and because
they are losing intracellular taurine, this excess taurine
must be excreted. Fish actually have a net renal
tubular secretion of taurine into their urine (King et
al., 1982). Chesney et al. (1998) reported that taurine
is an important osmolyte in the brain and the renal
medulla of infants. In these tissues, taurine is a pri-
mary factor in the cell volume regulatory process, in
which brain or renal cells swell or shrink in response
to osmolar changes. Infants depleted of taurine may
be able to respond to hyper- or hyponatremic stress
with massive changes in neuronal volume, which has
obvious clinical significance (Chesney et al., 1998).
Omura and Yoshimura (1999) suggested that the
abundant taurine localization in the retinal photorecep-
tor and neural layers of juvenile flounder may be
involved in the protection of the photoreceptor outer
segment, the regulation of neural transmission, and
photoreceptor differentiation during the developmental
stages. Heller-Stilb et al. (2002) reported that taurine is
involved in cell volume homeostasis, antioxidant, pro-
tein stabilization, and stress responses. High levels of
773
intracellular taurine are maintained by a Na+-depen-
dent taurine transporter in the plasma membrane. They
generated a mouse model with a disrupted gene coding
for the taurine transporter. These mice show markedly
decreased taurine levels in a variety of tissues, reduced
fertility, and loss of vision due to severe retinal degen-
eration. These findings suggest that taurine is very
important for the development and maintenance of
normal retinal function and morphology.
In conclusion, dietary taurine supplementation was
related to the growth performance and feeding behav-
ior of juvenile Japanese flounder. Taurine content of
whole body and tissues increased with an increase in
dietary taurine. This result indicates that the early
stage juvenile Japanese flounder require taurine in
their diets. This is the first report of taurine deficiency
in juvenile Japanese flounder. Further studies are
necessary to understand the physiological roles of
taurine during ontogenetic development and develop-
ment of feeding behavior of this species.
Acknowledgment
This research was financially supported by the
Sasakawa Scientific Research Grant from the Japan
Science Society.
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