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Effects of pigments from different sources on colour changes and growth of red Oreochromis niloticus
Aquaculture, 79 (1989) 375-380 Elsevier Science Publishers B.V., Amsterdam -
375 Printed in The Netherlands
Effects of Pigments from Different Sources on Colour Changes and Growth of Red Oreochromk nil0 ticus ‘MALI BOONYARATPALIN
and ‘NANTHIYA
UNPRASERT
‘National Institute of Coastal Aquaculture, Amphur Muang, Songkhla 900 00 (Thailand) ‘National Inland Fisheries Institute, Bankhen, Bangkok 10900 (Thailand)
ABSTRACT Boonyaratpalin, M. and Unprasert, N., 1989. Effect of pigments from different sources on colour changes and growth of red Oreochromis niloticus. Aquaculture, 79: 375-380. The investigation was aimed at determining the effect of different pigment sources on colour development, growth, feed conversion and survival of red tilapia. Four pigment sources were used: spirulina, Toreador marigold petal meal, shrimp head meal and turmeric. These pigments were added to the basal diet at lo%, 5%, 15% and 5%, respectively, and fed to red tilapia for 8 weeks. Fish fed with spirulina, marigold petal meal, shrimp head meal, turmeric and basal diet alone showed single red, golden flame, flame, orange and orange coloration, respectively. The snout and base of the fins of fish fed marigold petal meal developed temple yellow coloration. Growth was stimulated by shrimp head meal and inhibited by turmeric. Feed conversion was not significantly different from the basal diet except for the diet with turmeric added. Survival rate was essentially equal for all test diets and not significantly different from the basal diet.
INTRODUCTION
Since 1985 Oreochromis niloticus, commonly known as red tilapia, has become very popular mainly because of its bright red colour. Market demand, coupled with moderately high price, has induced fish farmers to culture red tilapia. It has been established conclusively that fish do not possess the ability to synthesize carotenoids. The carotenoid pigmentation pattern of fish results from the pigments present in the diet and accumulated in the tissues (Goodwin, 1951,1954; Fox, 1957; Hata and Hata, 1973; Steven, 1984). Several studies have shown that fish can be pigmented by including crustaceans and crustacean processing wastes in their feeds (Saito and Regier, 1971; Ibrahim et al., 1984) although conflicting results have been found with differ-
0044~8486/89/$03.50
0 1989 Elsevier Science Publishers B.V.
316
ent methods of processing shrimp waste (Boonyaratpalin and Phromkunthong, 1986). Plant sources have also been utilized for inducing pigmentation in fish. For example, spirulina algae have been used as a source of carotenoid pigments for rainbow trout and fancy carp (Choubert, 1979; Boonyaratpalin and Phromkunthong, 1986), and marigold petal meal was used for the tiger barb (Boonyaratpalin and Lovell, 1977). The present investigation was conducted (1) to determine the effect of pigment sources on colour development, growth, feed conversion and survival of red tilapia, (2) to explore the possibility of making tilapia colour more attractive to buyers, and (3) to determine whether diet substitution could reduce feed cost and make red tilapia culture less expensive. MATERIAL AND METHODS
The fish were kept in 250-l glass aquaria (50 x 100 X 50 cm) each with an individual air supply. Well water was stored and aerated in a fiberglass tank for a day before supplied to the aquaria. Fingerlings of red tilapia (NIFI’s strain) were used. All the fish were fed the basal diet (diet 1) for 2 weeks, prior to distribution in the experimental aquaria. The fish were also treated against parasites for 1 day in formaldehyde solution (35 mg/l). Test diets, formulated as shown in Table 1, were isonitrogenous (30% crude protein), isocaloric (3700 kcal/kg) and had equal ash values (7.3% ), but varied in protein and pigment sources. Four pigment sources were used: spirulina algae, marigold petal meal, raw shrimp head meal and turmeric at 10,5,15 and 5% (dry weight basis in the diets), respectively. Diets referred to as 2,3,4 and 5 were adjusted by substituting spirulina for fish meal and soybean meal, marigold petal meal for rice bran, shrimp head meal for peanut and mung bean meal, and turmeric for rice bran in the basal diet (diet 1) . In each case diets were balanced by removing an appropriate quantity of other ingredients with similar protein level to the pigmenting ingredient chosen. Spirulina was obtained from the Siam Algae Co. Ltd. as a dry meal. Marigold petals, from flowers of the Toreador strain, were purchased from the Agricultural Marketing Organization. Petals were dried in an oven at 60’ C and ground. Fresh shrimp head meal from the tiger shrimp (Perzaeus monodon) was obtained from a cold storage plant near the Fish Marketing Organization. The shrimp head meal was kept in the freezer until used. Turmeric was purchased in the local vegetable market. The dried ingredients were mixed in a Hobart mixer for 10 min, then 300 ml of water or wet ingredients such as ground shrimp waste or turmeric was added to each feed mixture and mixed for another 10 min. The mixture was extruded through a 3-mm-diameter die in a meat grinder. The spaghetti-like feed was
377 TABLE 1 Composition of the experimental diets for pigmentation and growth of red tilapia Ingredient
Experimental diet
Fish meal Mung bean meal Soybean meal Peanut meal Rice bran Spirohna Marigold petal meal Shrimp head meal Turmeric Bone meal Vitamin and mineral mix” Basfin (binder)
1
2
3
4
5
10.34 26.14 10.34 26.14 26.14
3.82 27.15 3.82 27.15 25.70 10.00
11.69 23.57 11.69 23.57 21.41 5.00
12.00 15.47 12.00 15.47 29.16
11.69 23.57 11.69 23.57 21.41 _ _ _
15.00
0.50 0.40
1.46 0.50 0.40
2.17 0.50 0.40
0.50 0.40
5.00 2.17 0.50 0.40
“Vitamin and mineral mix (mg/kg diet): thiamine 10; riboflavin 20; pyridoxine 10; folic acid 5; pantothenate 40; choline chloride 1000; ascorbic acid 300; niacin 150; vitamin B,, 0.002; vitamin A 4; vitamin E 50; menadione-Na-bisulfate 80; inositol400; biotin 2; ethoxyquin 200; MnS04- Hz0 250; KI 13; CuS04.5H,0 10; CoClz 1; ZnS01*7Hz0 140; ferric citrate 200; MgSO, 1250; cellulose 865.
cut into 0.5l.O-cm used. *
long pellets, sealed in plastic bags and stored at - 8°C until
Feeding trials
Two hundred and twenty-five fish weighing 7.7-8.3 g each were allotted randomly to 15 still-water aquaria, and fed in triplicate groups. The weight increase in each’aquarium was measured every 2 weeks and the feed conversion was calculated accordingly. The fish were fed twice daily at the rate of 8, 7, 6 and 5% of biomass for weeks O-2, 2-4, 4-6 and 6-8, respectively. Water was changed daily (80% of the volume) and the aquaria were cleaned every 2 weeks while fish weight was measured. At the end of the feeding trial, the fish in each aquarium were weighed and counted, and pigmentation was assessed visually using a colour chart. The data for each experiment were subjected to analysis of variance and Duncan’s multiple range test to determine whether differences among means were statistically significant (Steel and Torrie, 1969).
378 RESULTS AND DISCUSSION
The present work was carried out to determine whether carotenoids of spirulina, marigold petal meal, shrimp head meal or turmeric could induce pigmentation to make red tilapia more colourful. The first visual observations of a difference in the colour pattern on the body, fins and tail of red tilapia were at 2 weeks for the spirulina and marigold petal meal groups, and at 4 weeks for the groups given shrimp head meal. The colour pattern became brighter and darker with time. Results of the 8-week feeding trial indicated that spirulina, marigold petal meal and shrimp head meal had an appreciable influence on the pigmentation in red tilapia whereas the group given turmeric was the same colour as the group on the basal diet. The average colour score (Table 2) of red tilapia given spirulina was highest, followed by the group given marigold petal meal and shrimp head meal, which both gave the same score. The colour pattern developed differently for the different pigment sources. Fish fed spirulina, marigold petal meal, shrimp head meal, turmeric and basal diet showed single red, golden flame, flame, orange and orange coloration, respectively. The marigold petal meal diet gave a temple yellow coloration at the snout and at the base of the fins while other pigment sources did not have such effects. However, a sheen of the scales was observed in the fish fed the shrimp head diet. The rate of colour manifestation seemed to depend on the amount and nature of the carotenoid pigments in each pigment source. Average weight (Fig. 1) and percentage weight gain (Table 3 ) calculated for the total biomass in each group were significantly affected by raw shrimp head meal and turmeric. Shrimp head meal had a positive effect on growth while turmeric had a negative effect. Spirulina and marigold petal meal had no effect on growth relative to the basal diet. The average weights for the groups fed TABLE 2 Average colour scores by comparison with a colour chart for red Oreochronis niloticm fed different test diets for 0 to 8 weeks Diet no.
( 1) Basal diet (2 ) Basal diet + spirulina (3) Basal diet+marigold petal meal (4) Basal diet + shrimp head meal (5) Basal diet + turmeric
Weeks 0
2
4
6
8
1 1 1
1.7 + 1.0 3.7 rt 3.0 2.5& 1.8
2.0 * 1.1 4.0 + 3.3 3.5k2.7
3.0 f 2.5 4.2k3.5 3.6k3.0
3.422.1 5.Ok3.4 4.2 + 2.3
1 1
2.2 + 1.8 1.7 + 1.0
3.0 * 2.2 2.0+ 1.1
3.4f2.8 3.0f2.2
4.2 + 2.8 3.2 + 2.1
379 500
/ 450
O-O
basal diet
+-t
basal + spirulina
/' /
2 + Y
400
?
350
\
o-o
basal + marigold petal meal
/'
M
zM *d s
300
& 2 ? 4
250
200
150
100
I 0
2
4
(week)
6
Fig. 1. Average weight of red Oreochromis niloticus fed on different test diets for 8 weeks.
TABLE 3 Percentage weight gain (of total biomass) and feed conversion of red tilapia fed five different test diets for 8 weeks Diet no.
Percentage weight gain
Feed conversion (feed/gain)
(1) (2) (3) (4) (5 )
225 + 23b 237221b 218f25b 295 * 45 137 + 18”
2.7 + 0.5” 2.3 Z!I0.5” 2.6 I!Z0.4” 2.2kO.3” 3.9* 1.1s
Basal diet Basal diet+spirulina Basal diet + marigold petal meal Basal diet + shrimp head meal Basal diet + turmeric
Values within the same column having different letters are significantly different at P-C 0.05.
diets 1,2,3,4 and 5 for 8 weeks were 398 + 28,396 2 20,384 + 28,487 ? 66, and 284 + 18 g ( 2 SEM). Shrimp head meal had a positive effect on growth, as found by other workers (New, 1976), while turmeric depressed growth rate;
380
the reason for the latter may be the presence of an unknown toxin but this remains to be proved. The feed conversion ratio for the turmeric diet was significantly higher than for the other test diets (Table 3). The shrimp head meal diet gave the lowest feed conversion ratio but was not significantly different from spirulina, marigold petal meal or basal diet. The experimental diets did not significantly affect red tilapia survival rate (between 98 + 4 and 100% ) . CONCLUSION
Spirulina may serve as a source of pigmentation but at increased cost. Marigold petal meal has limited nutritional value and serves only as a pigmentation source. The cost is relatively high and the value of the red colour to the consumer would have to justify the expense. Shrimp head meal has significant nutritional value and also serves as a pigmentation source. The cost is reasonable. Turmeric has no nutritional or pigmentation value. Hence, where shrimp head meal is available it should be included in the feed for red tilapia and probably, for cost reasons, also in the feed of normal tilapia.
REFERENCES Boonyaratpalin, M. and Lovell, R.T., 1977. Diet preparation for aquarium fishes. Aquaculture, 12: 53-62. Boonyaratpalin, M. and Phromkunthong, W., 1986. Effects of carotenoidpigments from different sources on colour changes of fancy carp, Cyprinus carpio Linn. Songkhlanakarin J. Sci. Technol., 8 (1): 11-20. Choubert, G., Jr., 1979. Tentative utilization of spirulin algae as a source of carotenoid pigments for rainbow trout. Aquaculture, 18: 135-143. Fox, D.L., 1957. The pigment of fishes. In: M.E. Brown (Editor), Physiology of Fish. Academic Press, New York, NY, pp. 367-385. Goodwin, T.W., 1951. Carotenoids in fish. In: The biochemistry of fish. Biochem. Sot. Symp., 6: 63-82. Goodwin, T.W., 1954. Carotenoids - their Comparative Biochemistry. Chem. Publ. Co., New York, NY, 356 pp. Hata, M. and Hata, M., 1973. Studies on astaxanthin formation in some freshwater fishes. Tohoku J. Agric. Res., 24(4): 192-196. Ibrahim, A., Shimizu, C. and Kono, M., 1984. Pigmentation of cultured red sea bream, C~FYSOphrys major, using astaxanthin from Antarctic krill, Euphausia superba, and a mysid, Neomysis sp. Aquaculture, 38: 45-57. New, M.B., 1976. A review of dietary studies with shrimp and prawns. Aquaculture, 9: 101-144. Saito, A. and Regier, L.W., 1971. Pigmentation of brook trout (Salvelinus fontinalis) by feeding dried crustacean wastes. J. Fish. Res. Board Can., 28: 509-512. Steel, R.G.D. and Torrie, T.H., 1960. Principles and Procedures of Statistics. McGraw-Hill Book Co., New York, NY, 481 pp. Steven, D.M., 1984. Studies on animal carotenoide. I. Carotenoids of the brown trout (Salmo trutta Linn.). J. Exp. Biol., 25: 369-387.
375 Printed in The Netherlands
Effects of Pigments from Different Sources on Colour Changes and Growth of Red Oreochromk nil0 ticus ‘MALI BOONYARATPALIN
and ‘NANTHIYA
UNPRASERT
‘National Institute of Coastal Aquaculture, Amphur Muang, Songkhla 900 00 (Thailand) ‘National Inland Fisheries Institute, Bankhen, Bangkok 10900 (Thailand)
ABSTRACT Boonyaratpalin, M. and Unprasert, N., 1989. Effect of pigments from different sources on colour changes and growth of red Oreochromis niloticus. Aquaculture, 79: 375-380. The investigation was aimed at determining the effect of different pigment sources on colour development, growth, feed conversion and survival of red tilapia. Four pigment sources were used: spirulina, Toreador marigold petal meal, shrimp head meal and turmeric. These pigments were added to the basal diet at lo%, 5%, 15% and 5%, respectively, and fed to red tilapia for 8 weeks. Fish fed with spirulina, marigold petal meal, shrimp head meal, turmeric and basal diet alone showed single red, golden flame, flame, orange and orange coloration, respectively. The snout and base of the fins of fish fed marigold petal meal developed temple yellow coloration. Growth was stimulated by shrimp head meal and inhibited by turmeric. Feed conversion was not significantly different from the basal diet except for the diet with turmeric added. Survival rate was essentially equal for all test diets and not significantly different from the basal diet.
INTRODUCTION
Since 1985 Oreochromis niloticus, commonly known as red tilapia, has become very popular mainly because of its bright red colour. Market demand, coupled with moderately high price, has induced fish farmers to culture red tilapia. It has been established conclusively that fish do not possess the ability to synthesize carotenoids. The carotenoid pigmentation pattern of fish results from the pigments present in the diet and accumulated in the tissues (Goodwin, 1951,1954; Fox, 1957; Hata and Hata, 1973; Steven, 1984). Several studies have shown that fish can be pigmented by including crustaceans and crustacean processing wastes in their feeds (Saito and Regier, 1971; Ibrahim et al., 1984) although conflicting results have been found with differ-
0044~8486/89/$03.50
0 1989 Elsevier Science Publishers B.V.
316
ent methods of processing shrimp waste (Boonyaratpalin and Phromkunthong, 1986). Plant sources have also been utilized for inducing pigmentation in fish. For example, spirulina algae have been used as a source of carotenoid pigments for rainbow trout and fancy carp (Choubert, 1979; Boonyaratpalin and Phromkunthong, 1986), and marigold petal meal was used for the tiger barb (Boonyaratpalin and Lovell, 1977). The present investigation was conducted (1) to determine the effect of pigment sources on colour development, growth, feed conversion and survival of red tilapia, (2) to explore the possibility of making tilapia colour more attractive to buyers, and (3) to determine whether diet substitution could reduce feed cost and make red tilapia culture less expensive. MATERIAL AND METHODS
The fish were kept in 250-l glass aquaria (50 x 100 X 50 cm) each with an individual air supply. Well water was stored and aerated in a fiberglass tank for a day before supplied to the aquaria. Fingerlings of red tilapia (NIFI’s strain) were used. All the fish were fed the basal diet (diet 1) for 2 weeks, prior to distribution in the experimental aquaria. The fish were also treated against parasites for 1 day in formaldehyde solution (35 mg/l). Test diets, formulated as shown in Table 1, were isonitrogenous (30% crude protein), isocaloric (3700 kcal/kg) and had equal ash values (7.3% ), but varied in protein and pigment sources. Four pigment sources were used: spirulina algae, marigold petal meal, raw shrimp head meal and turmeric at 10,5,15 and 5% (dry weight basis in the diets), respectively. Diets referred to as 2,3,4 and 5 were adjusted by substituting spirulina for fish meal and soybean meal, marigold petal meal for rice bran, shrimp head meal for peanut and mung bean meal, and turmeric for rice bran in the basal diet (diet 1) . In each case diets were balanced by removing an appropriate quantity of other ingredients with similar protein level to the pigmenting ingredient chosen. Spirulina was obtained from the Siam Algae Co. Ltd. as a dry meal. Marigold petals, from flowers of the Toreador strain, were purchased from the Agricultural Marketing Organization. Petals were dried in an oven at 60’ C and ground. Fresh shrimp head meal from the tiger shrimp (Perzaeus monodon) was obtained from a cold storage plant near the Fish Marketing Organization. The shrimp head meal was kept in the freezer until used. Turmeric was purchased in the local vegetable market. The dried ingredients were mixed in a Hobart mixer for 10 min, then 300 ml of water or wet ingredients such as ground shrimp waste or turmeric was added to each feed mixture and mixed for another 10 min. The mixture was extruded through a 3-mm-diameter die in a meat grinder. The spaghetti-like feed was
377 TABLE 1 Composition of the experimental diets for pigmentation and growth of red tilapia Ingredient
Experimental diet
Fish meal Mung bean meal Soybean meal Peanut meal Rice bran Spirohna Marigold petal meal Shrimp head meal Turmeric Bone meal Vitamin and mineral mix” Basfin (binder)
1
2
3
4
5
10.34 26.14 10.34 26.14 26.14
3.82 27.15 3.82 27.15 25.70 10.00
11.69 23.57 11.69 23.57 21.41 5.00
12.00 15.47 12.00 15.47 29.16
11.69 23.57 11.69 23.57 21.41 _ _ _
15.00
0.50 0.40
1.46 0.50 0.40
2.17 0.50 0.40
0.50 0.40
5.00 2.17 0.50 0.40
“Vitamin and mineral mix (mg/kg diet): thiamine 10; riboflavin 20; pyridoxine 10; folic acid 5; pantothenate 40; choline chloride 1000; ascorbic acid 300; niacin 150; vitamin B,, 0.002; vitamin A 4; vitamin E 50; menadione-Na-bisulfate 80; inositol400; biotin 2; ethoxyquin 200; MnS04- Hz0 250; KI 13; CuS04.5H,0 10; CoClz 1; ZnS01*7Hz0 140; ferric citrate 200; MgSO, 1250; cellulose 865.
cut into 0.5l.O-cm used. *
long pellets, sealed in plastic bags and stored at - 8°C until
Feeding trials
Two hundred and twenty-five fish weighing 7.7-8.3 g each were allotted randomly to 15 still-water aquaria, and fed in triplicate groups. The weight increase in each’aquarium was measured every 2 weeks and the feed conversion was calculated accordingly. The fish were fed twice daily at the rate of 8, 7, 6 and 5% of biomass for weeks O-2, 2-4, 4-6 and 6-8, respectively. Water was changed daily (80% of the volume) and the aquaria were cleaned every 2 weeks while fish weight was measured. At the end of the feeding trial, the fish in each aquarium were weighed and counted, and pigmentation was assessed visually using a colour chart. The data for each experiment were subjected to analysis of variance and Duncan’s multiple range test to determine whether differences among means were statistically significant (Steel and Torrie, 1969).
378 RESULTS AND DISCUSSION
The present work was carried out to determine whether carotenoids of spirulina, marigold petal meal, shrimp head meal or turmeric could induce pigmentation to make red tilapia more colourful. The first visual observations of a difference in the colour pattern on the body, fins and tail of red tilapia were at 2 weeks for the spirulina and marigold petal meal groups, and at 4 weeks for the groups given shrimp head meal. The colour pattern became brighter and darker with time. Results of the 8-week feeding trial indicated that spirulina, marigold petal meal and shrimp head meal had an appreciable influence on the pigmentation in red tilapia whereas the group given turmeric was the same colour as the group on the basal diet. The average colour score (Table 2) of red tilapia given spirulina was highest, followed by the group given marigold petal meal and shrimp head meal, which both gave the same score. The colour pattern developed differently for the different pigment sources. Fish fed spirulina, marigold petal meal, shrimp head meal, turmeric and basal diet showed single red, golden flame, flame, orange and orange coloration, respectively. The marigold petal meal diet gave a temple yellow coloration at the snout and at the base of the fins while other pigment sources did not have such effects. However, a sheen of the scales was observed in the fish fed the shrimp head diet. The rate of colour manifestation seemed to depend on the amount and nature of the carotenoid pigments in each pigment source. Average weight (Fig. 1) and percentage weight gain (Table 3 ) calculated for the total biomass in each group were significantly affected by raw shrimp head meal and turmeric. Shrimp head meal had a positive effect on growth while turmeric had a negative effect. Spirulina and marigold petal meal had no effect on growth relative to the basal diet. The average weights for the groups fed TABLE 2 Average colour scores by comparison with a colour chart for red Oreochronis niloticm fed different test diets for 0 to 8 weeks Diet no.
( 1) Basal diet (2 ) Basal diet + spirulina (3) Basal diet+marigold petal meal (4) Basal diet + shrimp head meal (5) Basal diet + turmeric
Weeks 0
2
4
6
8
1 1 1
1.7 + 1.0 3.7 rt 3.0 2.5& 1.8
2.0 * 1.1 4.0 + 3.3 3.5k2.7
3.0 f 2.5 4.2k3.5 3.6k3.0
3.422.1 5.Ok3.4 4.2 + 2.3
1 1
2.2 + 1.8 1.7 + 1.0
3.0 * 2.2 2.0+ 1.1
3.4f2.8 3.0f2.2
4.2 + 2.8 3.2 + 2.1
379 500
/ 450
O-O
basal diet
+-t
basal + spirulina
/' /
2 + Y
400
?
350
\
o-o
basal + marigold petal meal
/'
M
zM *d s
300
& 2 ? 4
250
200
150
100
I 0
2
4
(week)
6
Fig. 1. Average weight of red Oreochromis niloticus fed on different test diets for 8 weeks.
TABLE 3 Percentage weight gain (of total biomass) and feed conversion of red tilapia fed five different test diets for 8 weeks Diet no.
Percentage weight gain
Feed conversion (feed/gain)
(1) (2) (3) (4) (5 )
225 + 23b 237221b 218f25b 295 * 45 137 + 18”
2.7 + 0.5” 2.3 Z!I0.5” 2.6 I!Z0.4” 2.2kO.3” 3.9* 1.1s
Basal diet Basal diet+spirulina Basal diet + marigold petal meal Basal diet + shrimp head meal Basal diet + turmeric
Values within the same column having different letters are significantly different at P-C 0.05.
diets 1,2,3,4 and 5 for 8 weeks were 398 + 28,396 2 20,384 + 28,487 ? 66, and 284 + 18 g ( 2 SEM). Shrimp head meal had a positive effect on growth, as found by other workers (New, 1976), while turmeric depressed growth rate;
380
the reason for the latter may be the presence of an unknown toxin but this remains to be proved. The feed conversion ratio for the turmeric diet was significantly higher than for the other test diets (Table 3). The shrimp head meal diet gave the lowest feed conversion ratio but was not significantly different from spirulina, marigold petal meal or basal diet. The experimental diets did not significantly affect red tilapia survival rate (between 98 + 4 and 100% ) . CONCLUSION
Spirulina may serve as a source of pigmentation but at increased cost. Marigold petal meal has limited nutritional value and serves only as a pigmentation source. The cost is relatively high and the value of the red colour to the consumer would have to justify the expense. Shrimp head meal has significant nutritional value and also serves as a pigmentation source. The cost is reasonable. Turmeric has no nutritional or pigmentation value. Hence, where shrimp head meal is available it should be included in the feed for red tilapia and probably, for cost reasons, also in the feed of normal tilapia.
REFERENCES Boonyaratpalin, M. and Lovell, R.T., 1977. Diet preparation for aquarium fishes. Aquaculture, 12: 53-62. Boonyaratpalin, M. and Phromkunthong, W., 1986. Effects of carotenoidpigments from different sources on colour changes of fancy carp, Cyprinus carpio Linn. Songkhlanakarin J. Sci. Technol., 8 (1): 11-20. Choubert, G., Jr., 1979. Tentative utilization of spirulin algae as a source of carotenoid pigments for rainbow trout. Aquaculture, 18: 135-143. Fox, D.L., 1957. The pigment of fishes. In: M.E. Brown (Editor), Physiology of Fish. Academic Press, New York, NY, pp. 367-385. Goodwin, T.W., 1951. Carotenoids in fish. In: The biochemistry of fish. Biochem. Sot. Symp., 6: 63-82. Goodwin, T.W., 1954. Carotenoids - their Comparative Biochemistry. Chem. Publ. Co., New York, NY, 356 pp. Hata, M. and Hata, M., 1973. Studies on astaxanthin formation in some freshwater fishes. Tohoku J. Agric. Res., 24(4): 192-196. Ibrahim, A., Shimizu, C. and Kono, M., 1984. Pigmentation of cultured red sea bream, C~FYSOphrys major, using astaxanthin from Antarctic krill, Euphausia superba, and a mysid, Neomysis sp. Aquaculture, 38: 45-57. New, M.B., 1976. A review of dietary studies with shrimp and prawns. Aquaculture, 9: 101-144. Saito, A. and Regier, L.W., 1971. Pigmentation of brook trout (Salvelinus fontinalis) by feeding dried crustacean wastes. J. Fish. Res. Board Can., 28: 509-512. Steel, R.G.D. and Torrie, T.H., 1960. Principles and Procedures of Statistics. McGraw-Hill Book Co., New York, NY, 481 pp. Steven, D.M., 1984. Studies on animal carotenoide. I. Carotenoids of the brown trout (Salmo trutta Linn.). J. Exp. Biol., 25: 369-387.