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The value of Phaffia yeast as a feed ingredient for salmonid fish
Aquacultum Aquaculture
124 ( 1994) 193-200
The value of PhafJia yeast as a feed ingredient for salmonid fish Gary W. Sanderson*, Setsuko 0. Jolly Red Star Specialty Products, 433 E. Michigan Street, Milwaukee, WI 53202, USA
Abstract The yeast Phaffla rhodozyma has been developed as a natural source of astaxanthin and other nutrients for use as an ingredient in feeds for salmonid fish and other aquatic species being produced by aquaculture. This yeast has been shown to be a good source of nutritionally important substances that are characteristic of yeast in general (protein, lipids, vitamins, etc.), and it is also a source of astaxanthin. Research has indicated that astaxanthin is of importance in the nutrition of salmonid fish and certain other aquatic species, and that this carotenoid must be obtained by salmonids from dietary sources since they cannot biosynthesize this substance.
1. Introduction
Yeast products (primarily brewer’s yeast and baker’s yeast) are frequently used as feed ingredients in aquaculture because of the nutritional value of these products which include protein, lipids, B-vitamins, etc. (Mahnken, 199 1; van der Meeren, 199 1). Phu$kz rhudozyma (family Cryptococcaceae) is a species of yeast that is of particular interest to the aquaculture industry, because, in addition to the usual nutritional factors associated with yeast products, P. rhodozyma contains astaxanthin which is the most abundant carotenoid in the marine environment. Astaxanthin is the major pigment in the flesh of salmonid fish, carapace of crustacea, skin of red sea bream, and numerous other marine animals. Astaxanthin present in salmonid fish must be provided by dietary sources since these fish lack the ability to biosynthesize astaxanthin. When raised in aquaculture, salmonids must be fed diets containing a source of this carotenoid compound in order to produce their normal pigmentation. In addition to being responsible for *Corresponding
author.
0044-8486/94/$07.00 0 1994 Elsevier Science B.V. All rights reserved SSDIOO44-8486(94)00066-W
194
G. W. Sanderson, SO. Jolly /Aquaculture 124 (1994) 193-200
the characteristic color of these animals, astaxanthin plays a critical nutritional role in the life of these marine animals (Meyers and Chen, 1982). The review by Simpson et al. ( 198 1) provides much information about the chemistry of astaxanthin. Johnson and An ( 199 1) give a detailed review of the yeast P. rhodozimu as a source of astaxanthin for aquaculture. This paper reviews some recent information on the nutritional value of the yeast P. rhoduzyma and the carotenoid astaxanthin, which is a natural constituent of this particular yeast. 2. General composition of Phafia rhodoqyma and its use as a source of pigment for salmonids Phu&Gzrhodozyma contains protein, lipids, vitamins, and minerals typical of yeast in general (Tables 1 and 2). P. rhodozymu may also contain growth factors and feed attractants that have been historically associated with yeast products (Oriental Yeast, 1982; Coutteau et al., 1990). Experimental feeding trials have been carried out with rainbow trout to determine the nutritional value and efficacy of P. rhodozymu to color fish flesh. In these trials, Phuffia yeast displaced an equivalent amount of ingredients in the control feed formulation (a standard commercial feed for salmonids prepared by Zeigler Bros., Inc., Gardners, PA, USA) so that the amount of protein, lipid, and total energy was approximately unchanged. An example of the results obtained in these trials is presented in Fig. 1 (from Sedmak et al., 1992). These results show that fish consuming PhufJia yeast at up to 10% of the diet grew at the same rate as fish consuming a control diet which contained an equivalent amount of brewer’s yeast. During the 84-day trial period all groups of fish were healthy and there were no deaths. Astaxanthin from the Phufj?u yeast was incorporated into the rainbow trout flesh. As seen in Fig. 1, there was a dose-dependent, and duration-dependent, increase in the carotenoid content of the fish flesh. Commercial color level was observed in the fish fed the high dose of astaxanthin from Phuffia (Diet A) at 84 days. Even the fish fed the low dose of astaxanthin from Phu& (Diet B) had a flesh carotenoid content significantly higher than the background level of the fish fed the diet devoid of carotenoids (Diet C). Analysis of the pigment extracted from the flesh of the fish fed Phufiu yeast Table 1 Typical proximate composition Component Astaxanthin Protein Lipid Ash Moisture
of Pha@a yeast produced for use in aquaculture Content (as-is basis)
(minimum)
3000 pm 22% 23% 3% 5%
195
G. W. Sunderson. S. 0. Jolly / Aquuculiure 124 (1994) 193-200 Table 2 Amino acid, fatty acid, vitamin and mineral composition
of Phuffia yeast
Amino acid
% Crude protein basis
Fatty acid
% Total fatty acids
Vitamin
ppm
Mineral
ppm
Asp Thr Ser Glu Pro Gly Ala Cys Val Met Ile Leu Tyr Phe His Lys Arg Trp
6.3 3.9 3.1 1.9 3.6 3.6 5.6 0.8 3.7 1.1 2.9 5.1 1.9 2.8 1.7 4.1 6.3 0.7
14:o 15:o 16:0 16:ln-7 16:3n-4 16:4n- 1 17:o 180 18:ln-9 18:2n-6 18:3n-3 19:l 20:o 2210 24:0
0.32 0.39 13.18 0.48 0.85 0.33 1.24 5.62 32.43 39.65 1.26 0.25 0.62 0.75 0.76
Niacin Riboflavin Vitamin D2 Biotin Thiamine Pyridoxine Pantothenate Folic acid
1520 100 85” 1 14 12 34 6
Calcium Sodium Zinc Phosphate Magnesium Potassium Iron Heavy metals Arsenic Lead Selenium Mercury Cadmium
248 2170 14 16 200 1489 4135 31 < 10
w/
100 g From R. Gross, Technical Data for Red Star@’Ph@a Yeast, available from Red Star Specialty Products, Milwaukee, WI.
Day: 84
Diet A: 10% Phaffia yeast (67 ppm astaxanthin) Diet 8: 2.5% Phaffia yeast and 7.5% brewers yeast Diet C: 10% brewers yeast (unpigmented control)
(16 ppm astaxanthin)
Fig. 1.Weight gain and pigmentation of rainbow trout fed a commercial salmonid feed that contained varying amounts of Phuffiu yeast as the source of astaxanthin. Data from Sedmak et al., ( 1992).
196
G. W. Sanderson, S.0. Jolly / Aquaculture I24 (1994) 193-200
indicated that 95% of the flesh carotenoids were astaxanthin which is the same pigment as that which predominates in the flesh of salmonids caught in the wild (Schiedt et al., 1981; Torrissen et al., 1989).
3. Other potential benefits of astaxanthin in the culture of fish Effect on egg quality andfj, survival Based on a review of the literature, Craik ( 1985) concluded that l-3 mg of carotenoids per gram of salmonid eggs was associated with hatching percentages above 6096, whereas lower carotenoid levels produced hatching percentages below 50%. Recently, Torrissen (cited by Hardy and Stickney, 1992) has found that under certain conditions of culture, offspring of Atlantic salmon which were fed diets without astaxanthin in their “first feeding diets” suffered more than 85% mortality, whereas cohorts receiving feed containing astaxanthin survived and flourished. Watanabe et al. ( 1984) have reported that dietary astaxanthin significantly increased the normal larvae produced from red sea bream broodstock from 5 1.6% (eggs from broodstock that received no dietary carotenoids) to 9 1.2% (eggs from broodstock that received dietary astaxanthin ) . Antioxidant activity Astaxanthin has antioxidant activity as a singlet oxygen quencher and free radical scavenger (Kurashige et al., 1990; Suntory, 1990; Miki, 199 1). Astaxanthin has been shown to be more efficient than /&carotene and zeaxanthin in retarding hydroperoxidation of methyl linoleate (Terao, 1989 ). Astaxanthin has also been reported to be 200 times (molar basis) more effective than vitamin E, and 10 times (molar basis) more effective than BHA in preventing hydroperoxidation of lipids in foods, cosmetics and pharmaceutical products (Suntory, 1990). It has been suggested that the antioxidant properties of astaxanthin protect fish containing this substance against photodynamic damage and lipid peroxidation (Ranby and Rabek, 1978 ). Kurashige et al. ( 1990) have shown that astaxanthin protects biological membranes from oxidative injury by inhibiting mitochondrial lipid peroxidation. Using rat liver microsomes, Palozza and Krinsky ( 1992) have also shown that astaxanthin protects biological membranes by an antioxidant mechanism. Salmonids are coldwater fish which contain high levels of polyunsaturated lipids that are susceptible to oxidation. The above observations suggest that adequate levels of astaxanthin in the flesh of salmonid fish may offer some protection against oxidation of body lipids. Source of vitamin A (provitamin A activity) A biological function of astaxanthin as vitamin A precursors was found in vitamin-A-depleted rainbow trout (Schiedt et al., 1985 ). Katsuyama and Matsuno ( 1988 ) have reported that dihydroxy-carotenoids, such as astaxanthin, zeaxanthin, lutein and tunaxanthin, were bioconverted into vitamin A2 alcohol in Tilapia nilotica.
G. W. Sanderson. S. 0. Jolly I Aquaculture 124 (1994)
193-200
197
Effect on liver histology
Segner et al. ( 1989) studied the effect of dietary astaxanthin on the histology of the liver of Oreochromis niloticus and Colisa labiosa, two species of fish whose pigmentation is related directly to dietary carotenoid levels. This study showed that increasing astaxanthin in the diet from 0 to 132 mg/kg diet showed an improvement in the histology of the liver structure. These researchers concluded that astaxanthin has a positive nutritional function in the intermediary metabolism of these fish. Efect on survival Low levels of carotenoids in farmed tiger shrimp (Penaeus monodon) is re-
sponsible for “blue shrimp” disease (poor survival rates) which is a common problem in intensive shrimp farming operations (Howell and Matthews, 199 1) . 0
(3S,3’R)-astaxathin 0
OH
(3S,3’S)-astaxanthin Fig. 2. Structure of configurational
isomers of astaxanthin
(Andrewes and Starr, 1976).
198
G. W. Sanderson, S.O. Jolly / Aquaculture 124 (1994) 193-200 Astaxanthin
from Phaffia
...,..,.,,.,.,.synthetic Astaxanthin __________
Astaxanthin
from Wild Salmon
Retention
Time
(min)
Fig. 3. Separations and identification of astaxanthin isomers in Phafja rhodozyma, in synthetic astaxanthin, and in a wild salmon. An HPLC chromatogram using the following conditions: column, Sumipax OA-2000, mobile phase, n-hexanelmethylene dichloride/ethanol (24: 58 : 0.3); flow rate, 1.O ml/min; detection, absorption at 480 nm (modification of method by Maoka et al., 1985b, by R. Gross, Red Star Specialty Products, Milwaukee, Wisconsin, unpublished).
It was presumed that astaxanthin provided protection against the deleterious effects of the low available oxygen conditions that often occur in intensive shrimpfarming operations. Chien and Jeng ( 1992) found that kuruma prawns (Penaeus juponicus) fed astaxanthin had significantly higher survival than kuruma prawns that received similar amounts of dietary/&carotene or algal meal. Only dietary astaxanthin was effective in increasing survival compared to the control shrimps which received no dietary pigment. All shrimps that received dietary carotenoids had a significantly greater weight gain than the control shrimps.
4. Chemistry of astaxanthin and its stability in salmonid fish tissue Astaxanthin, an oxygenated carotene, has two chiral centers and may exist in three configurations: a pair of enantiomers (3&3’S) and (3R,3’R), and a mesoform (3R,3’S) (Fig. 2). The three stereo-isomers of astaxanthin are readily resolved and can be quantified using high-performance liquid chromatography (HPLC) with chiral columns (Maoka et al., 1985a,b). Astaxanthin is normally present in the flesh of wild salmonids at a concentration of about 8-37 ppm and this compound comprises over 99% of the total carotenoids present (Torrissen et al., 1989; Storebakken and No, 1992). As mentioned above, astaxanthin in salmonids is derived from the diet since salmonids cannot biosynthesize this com-
G. W. Sanderson. S.O. Jolly / Aquaculture 124 (1994) 193-200
199
pound (Simpson et al., 198 1; Torrissen et al., 1989; Storebakken and No, 1992). No epimerization occurs in the tissue of salmonid fish at the chiral centers at C3 and C-3’ in astaxanthin (Foss et al., 1984; Storebakken et al., 1985). The natural dietary sources of astaxanthin all contain astaxanthin in the enantiomeric RR or SS forms (Matsuno et al., 1984); thus wild salmonids will contain varying ratios of these two enantiomers depending on the relative amounts of each enantiomer present in the animal’s diet. Astaxanthin is naturally biosynthesized by the yeast Phaffi rhodozyma during its normal life cycle (Johnson and An, 199 1). The astaxanthin present in P. rhodozyma is in the 3R,3’Rconfiguration (Andrewes and Starr, 1976). The 3R,3’Rastaxanthin is present to some degree in all salmonid fish simply because this enantiomer is widely distributed in the natural foods consumed by salmonids in the wild (Matsuno et al., 1984; Schiedt et al., 198 1). Fig. 3 illustrates that the pigment extracted from wild salmonid fish contains the same stereoisomer of astaxanthin as Phufju as determined by HPLC with an optical resolution column that separates the three astaxanthin isomers (Maoka et al., 1985b). References Andrewes, A. and Starr, M., 1976. (3R,3’R)-Astaxanthin from the yeast Phafia rhodozyma. Phytochemistry, 15: 1009- 10 11. Chien, Y.M. and Jeng, S.C., 1992. Pigmentation of kuruma prawn, Penaeusjaponicus Bate, by various pigment sources and levels and feeding regimes. Aquaculture, 102: 333-346. Coutteau, P., Lavens, P. and Sorgeloos, P., 1990. Baker’s yeast as a potential substitute for live algae in aquaculture diets: Artemia as a case study. J. World Aquacult. Sot., 21: 1-9. Craik, J.C.A., 1985. Egg quality and pigment content in salmonid fishes. Aquaculture, 47: 61-88. Foss, P., Storebakken, T., Scheidt, K., Liaaen-Jensen, S., Austreng, E. and Streiff, K., 1984. Carotenoids in diets for salmonids. 1. Pigmentation of rainbow trout with the individual optical isomers of astaxanthin in comparison with canthaxanthin. Aquaculture, 4 1: 2 13-226. Hardy, R.W. and Stickney, R.R., 1992. Carotenoid Pigmentation of Salmon and Trout. Publication prepared March 19, 1992, and distributed privately, University of Washington, Seattle. Howell, B.K. and Matthews, A.D., 1991. The carotenoids of wild and blue disease affected farmed tiger shrimp (Penaeus monodon, Fabricus). Comp. Biochem. Physiol., 98B: 375-379. Johnson, E.A. and An, G.H., 199 1. Astaxanthin from microbial sources. Crit. Rev. Biotechnol., 1 I : 297-326. Katsuyama, M. and Matsuno, T., 1988. Carotenoid and vitamin A, and metabolism of carotenoids, /?-carotene, canthaxanthin, astaxanthin, zeaxanthin, lutein, and tunaxanthin in tilapia ( Tilapia nilotica). Comp. Biochem. Physiol., 90B: 131-139. Kurashige, M., Okimasu, E., Inoue, M. and Utsumi, K., 1990. Inhibition of oxidative injury of biological membranes by astaxanthin. Physiol. Chem. Phys. Med. NMR, 22: 27-38. Mahnken, C.V.M., 1991. Coho salmon farming in Japan. In: R.R. Stickney (Editor), Culture of Salmonid Fishes. CRC Press, Boca Raton, FL, pp. 13 1- 149. Maoka, T., Katsuyama, M.,Kaneko, N. and Matsuno T., 1985a. Stereochemical investigation of carotenoids in the antarctic krill Euphausia superba. Bul. Jpn. Sot. Sci. Fish., 5 1: 1671-1673. Maoka, T., Kumori, T. and Matsuno, T., 1985b. Direct diastereomeric resolution of carotenoids. I. 3-Hydroxy-4-oxo /J-end group. J. Chromatogr., 3 18: 122- 124. Matsuno, T., Maoka, T., Katsuyama, M., Ookubo, M., Katagiri, K. and Jimura, H., 1984. Occurrence of enantiomeric and meso-astaxanthin in aquatic animals. Bull. Jpn. Sot. Sci. Fish., 50: 15891592.
200
G. W. Sanderson, S.O. Jolly / Aquaculture 124 (1994) 193-200
Meyers, S.P. and Chen, H.M., 1982. Astaxanthin and its role in fish culture. In: R.R. Stickley and S.P. Meyers (Editors), Special Publication No. 3, World Aquaculture Society, Baton Rouge, LA, pp. 153-165. Miki, W., 199 1. Biological functions and activities of animal carotenoids. Pure Appl. Chem., 63: 14 l146. Oriental Yeast KK, 1982. Feed for rearing fish contains digested yeast body extract. Japan Patent No. JP-181447. Palozza, P. and Krinsky, L., 1992. Astaxanthin and canthaxanthin are potent antioxidants in a membrane model. Arch. Biochem. Biophys., 297: 29 l-295. Ranby, B. and Rabek, J.F. (Editors), 1978. Singlet Oxygen. Wiley, Chichester, 331 pp. Schiedt, K., Leuenberger, F. J. and Vecchi, M., 198 1. Natural occurence of enantiomeric and mesoastaxanthin. 5. Ex wild salmon (Safmo salar and Oncorhynchus). Helv. Chim. Acta, 64: 449-457. Schiedt, K., Levenberger, F.J., Vecchi, M. and Glinz, E., 1985. Absorption, retention and metabolic transformations ofcarotenoids in rainbow trout, salmon and chicken. Pure Appi. Chem., 57: 685692. Sedmak, J.J., Jolly, S.O. and Binkowski, F.P., 1992. An evaluation of Red Star@ Phafla yeast as a pigment source for salmonids. Red Star Bio Products, Technical Publication No. 1, Milwaukee, WI, 9 PP. Segner, H., Arend, P., von Poeppinghausen, K. and Schmidt, H., 1989. The effect of feeding astaxanthin to Oreochromis niloticusand Colisa lebrisa on the histology of the liver. Aquaculture, 79: 38 I 390. Simpson, K.L., Katayama, T. and Chichester, C.O., 198 1. Carotenoids in fish feeds. In: J.C. Bauernfeind (Editor), Carotenoids as Colorants and Vitamin A Precursors. Academic Press, New York, pp. 463-538. Storebakken, T. and No, H.K., 1992. Pigmentation of rainbow trout. Aquaculture, 100: 209-229. Storebakken, T., Foss, P., Austreng, E. and Liaaen-Jensen, S., 1985. Carotenoids in diets for salmonids. II. Epimerization studies with astaxanthin in Atlantic salmon. Aquaculture, 44: 259-269. Suntory Ltd., 1990. Composition containing astaxanthin as oxidation inhibitor useful as vitamin E substitute and as food additive, medicament, etc. Japan Patent JP-198947, 11 pp. Terao, O., 1989. Antioxidant activity of /?-carotene related carotenoids in solution. Lipids, 24: 659661. Torrissen, O.J., Hardy, R.W. and Shearer, K.D., 1989. Pigmentation of salmonids: Carotenoid deposition and metabolism. Rev. Aquat. Sci., 1: 209-225. van der Meeren, T., I99 1. Production of marine fish fry in Norway. World Aquacult., 22 (2): 37-40. Watanabe, T., Itoh, A., Murakami, A., Tsukashima, Y., Kitajima, C. and Fujita, S., 1984. Effect of nutritional quality of diets given to broodstock on the verge of spawning on reproduction of red sea bream. Bull. Jpn. Sot. Sci. Fish., 50: 1023-1028.
124 ( 1994) 193-200
The value of PhafJia yeast as a feed ingredient for salmonid fish Gary W. Sanderson*, Setsuko 0. Jolly Red Star Specialty Products, 433 E. Michigan Street, Milwaukee, WI 53202, USA
Abstract The yeast Phaffla rhodozyma has been developed as a natural source of astaxanthin and other nutrients for use as an ingredient in feeds for salmonid fish and other aquatic species being produced by aquaculture. This yeast has been shown to be a good source of nutritionally important substances that are characteristic of yeast in general (protein, lipids, vitamins, etc.), and it is also a source of astaxanthin. Research has indicated that astaxanthin is of importance in the nutrition of salmonid fish and certain other aquatic species, and that this carotenoid must be obtained by salmonids from dietary sources since they cannot biosynthesize this substance.
1. Introduction
Yeast products (primarily brewer’s yeast and baker’s yeast) are frequently used as feed ingredients in aquaculture because of the nutritional value of these products which include protein, lipids, B-vitamins, etc. (Mahnken, 199 1; van der Meeren, 199 1). Phu$kz rhudozyma (family Cryptococcaceae) is a species of yeast that is of particular interest to the aquaculture industry, because, in addition to the usual nutritional factors associated with yeast products, P. rhodozyma contains astaxanthin which is the most abundant carotenoid in the marine environment. Astaxanthin is the major pigment in the flesh of salmonid fish, carapace of crustacea, skin of red sea bream, and numerous other marine animals. Astaxanthin present in salmonid fish must be provided by dietary sources since these fish lack the ability to biosynthesize astaxanthin. When raised in aquaculture, salmonids must be fed diets containing a source of this carotenoid compound in order to produce their normal pigmentation. In addition to being responsible for *Corresponding
author.
0044-8486/94/$07.00 0 1994 Elsevier Science B.V. All rights reserved SSDIOO44-8486(94)00066-W
194
G. W. Sanderson, SO. Jolly /Aquaculture 124 (1994) 193-200
the characteristic color of these animals, astaxanthin plays a critical nutritional role in the life of these marine animals (Meyers and Chen, 1982). The review by Simpson et al. ( 198 1) provides much information about the chemistry of astaxanthin. Johnson and An ( 199 1) give a detailed review of the yeast P. rhodozimu as a source of astaxanthin for aquaculture. This paper reviews some recent information on the nutritional value of the yeast P. rhoduzyma and the carotenoid astaxanthin, which is a natural constituent of this particular yeast. 2. General composition of Phafia rhodoqyma and its use as a source of pigment for salmonids Phu&Gzrhodozyma contains protein, lipids, vitamins, and minerals typical of yeast in general (Tables 1 and 2). P. rhodozymu may also contain growth factors and feed attractants that have been historically associated with yeast products (Oriental Yeast, 1982; Coutteau et al., 1990). Experimental feeding trials have been carried out with rainbow trout to determine the nutritional value and efficacy of P. rhodozymu to color fish flesh. In these trials, Phuffia yeast displaced an equivalent amount of ingredients in the control feed formulation (a standard commercial feed for salmonids prepared by Zeigler Bros., Inc., Gardners, PA, USA) so that the amount of protein, lipid, and total energy was approximately unchanged. An example of the results obtained in these trials is presented in Fig. 1 (from Sedmak et al., 1992). These results show that fish consuming PhufJia yeast at up to 10% of the diet grew at the same rate as fish consuming a control diet which contained an equivalent amount of brewer’s yeast. During the 84-day trial period all groups of fish were healthy and there were no deaths. Astaxanthin from the Phufj?u yeast was incorporated into the rainbow trout flesh. As seen in Fig. 1, there was a dose-dependent, and duration-dependent, increase in the carotenoid content of the fish flesh. Commercial color level was observed in the fish fed the high dose of astaxanthin from Phuffia (Diet A) at 84 days. Even the fish fed the low dose of astaxanthin from Phu& (Diet B) had a flesh carotenoid content significantly higher than the background level of the fish fed the diet devoid of carotenoids (Diet C). Analysis of the pigment extracted from the flesh of the fish fed Phufiu yeast Table 1 Typical proximate composition Component Astaxanthin Protein Lipid Ash Moisture
of Pha@a yeast produced for use in aquaculture Content (as-is basis)
(minimum)
3000 pm 22% 23% 3% 5%
195
G. W. Sunderson. S. 0. Jolly / Aquuculiure 124 (1994) 193-200 Table 2 Amino acid, fatty acid, vitamin and mineral composition
of Phuffia yeast
Amino acid
% Crude protein basis
Fatty acid
% Total fatty acids
Vitamin
ppm
Mineral
ppm
Asp Thr Ser Glu Pro Gly Ala Cys Val Met Ile Leu Tyr Phe His Lys Arg Trp
6.3 3.9 3.1 1.9 3.6 3.6 5.6 0.8 3.7 1.1 2.9 5.1 1.9 2.8 1.7 4.1 6.3 0.7
14:o 15:o 16:0 16:ln-7 16:3n-4 16:4n- 1 17:o 180 18:ln-9 18:2n-6 18:3n-3 19:l 20:o 2210 24:0
0.32 0.39 13.18 0.48 0.85 0.33 1.24 5.62 32.43 39.65 1.26 0.25 0.62 0.75 0.76
Niacin Riboflavin Vitamin D2 Biotin Thiamine Pyridoxine Pantothenate Folic acid
1520 100 85” 1 14 12 34 6
Calcium Sodium Zinc Phosphate Magnesium Potassium Iron Heavy metals Arsenic Lead Selenium Mercury Cadmium
248 2170 14 16 200 1489 4135 31 < 10
w/
100 g From R. Gross, Technical Data for Red Star@’Ph@a Yeast, available from Red Star Specialty Products, Milwaukee, WI.
Day: 84
Diet A: 10% Phaffia yeast (67 ppm astaxanthin) Diet 8: 2.5% Phaffia yeast and 7.5% brewers yeast Diet C: 10% brewers yeast (unpigmented control)
(16 ppm astaxanthin)
Fig. 1.Weight gain and pigmentation of rainbow trout fed a commercial salmonid feed that contained varying amounts of Phuffiu yeast as the source of astaxanthin. Data from Sedmak et al., ( 1992).
196
G. W. Sanderson, S.0. Jolly / Aquaculture I24 (1994) 193-200
indicated that 95% of the flesh carotenoids were astaxanthin which is the same pigment as that which predominates in the flesh of salmonids caught in the wild (Schiedt et al., 1981; Torrissen et al., 1989).
3. Other potential benefits of astaxanthin in the culture of fish Effect on egg quality andfj, survival Based on a review of the literature, Craik ( 1985) concluded that l-3 mg of carotenoids per gram of salmonid eggs was associated with hatching percentages above 6096, whereas lower carotenoid levels produced hatching percentages below 50%. Recently, Torrissen (cited by Hardy and Stickney, 1992) has found that under certain conditions of culture, offspring of Atlantic salmon which were fed diets without astaxanthin in their “first feeding diets” suffered more than 85% mortality, whereas cohorts receiving feed containing astaxanthin survived and flourished. Watanabe et al. ( 1984) have reported that dietary astaxanthin significantly increased the normal larvae produced from red sea bream broodstock from 5 1.6% (eggs from broodstock that received no dietary carotenoids) to 9 1.2% (eggs from broodstock that received dietary astaxanthin ) . Antioxidant activity Astaxanthin has antioxidant activity as a singlet oxygen quencher and free radical scavenger (Kurashige et al., 1990; Suntory, 1990; Miki, 199 1). Astaxanthin has been shown to be more efficient than /&carotene and zeaxanthin in retarding hydroperoxidation of methyl linoleate (Terao, 1989 ). Astaxanthin has also been reported to be 200 times (molar basis) more effective than vitamin E, and 10 times (molar basis) more effective than BHA in preventing hydroperoxidation of lipids in foods, cosmetics and pharmaceutical products (Suntory, 1990). It has been suggested that the antioxidant properties of astaxanthin protect fish containing this substance against photodynamic damage and lipid peroxidation (Ranby and Rabek, 1978 ). Kurashige et al. ( 1990) have shown that astaxanthin protects biological membranes from oxidative injury by inhibiting mitochondrial lipid peroxidation. Using rat liver microsomes, Palozza and Krinsky ( 1992) have also shown that astaxanthin protects biological membranes by an antioxidant mechanism. Salmonids are coldwater fish which contain high levels of polyunsaturated lipids that are susceptible to oxidation. The above observations suggest that adequate levels of astaxanthin in the flesh of salmonid fish may offer some protection against oxidation of body lipids. Source of vitamin A (provitamin A activity) A biological function of astaxanthin as vitamin A precursors was found in vitamin-A-depleted rainbow trout (Schiedt et al., 1985 ). Katsuyama and Matsuno ( 1988 ) have reported that dihydroxy-carotenoids, such as astaxanthin, zeaxanthin, lutein and tunaxanthin, were bioconverted into vitamin A2 alcohol in Tilapia nilotica.
G. W. Sanderson. S. 0. Jolly I Aquaculture 124 (1994)
193-200
197
Effect on liver histology
Segner et al. ( 1989) studied the effect of dietary astaxanthin on the histology of the liver of Oreochromis niloticus and Colisa labiosa, two species of fish whose pigmentation is related directly to dietary carotenoid levels. This study showed that increasing astaxanthin in the diet from 0 to 132 mg/kg diet showed an improvement in the histology of the liver structure. These researchers concluded that astaxanthin has a positive nutritional function in the intermediary metabolism of these fish. Efect on survival Low levels of carotenoids in farmed tiger shrimp (Penaeus monodon) is re-
sponsible for “blue shrimp” disease (poor survival rates) which is a common problem in intensive shrimp farming operations (Howell and Matthews, 199 1) . 0
(3S,3’R)-astaxathin 0
OH
(3S,3’S)-astaxanthin Fig. 2. Structure of configurational
isomers of astaxanthin
(Andrewes and Starr, 1976).
198
G. W. Sanderson, S.O. Jolly / Aquaculture 124 (1994) 193-200 Astaxanthin
from Phaffia
...,..,.,,.,.,.synthetic Astaxanthin __________
Astaxanthin
from Wild Salmon
Retention
Time
(min)
Fig. 3. Separations and identification of astaxanthin isomers in Phafja rhodozyma, in synthetic astaxanthin, and in a wild salmon. An HPLC chromatogram using the following conditions: column, Sumipax OA-2000, mobile phase, n-hexanelmethylene dichloride/ethanol (24: 58 : 0.3); flow rate, 1.O ml/min; detection, absorption at 480 nm (modification of method by Maoka et al., 1985b, by R. Gross, Red Star Specialty Products, Milwaukee, Wisconsin, unpublished).
It was presumed that astaxanthin provided protection against the deleterious effects of the low available oxygen conditions that often occur in intensive shrimpfarming operations. Chien and Jeng ( 1992) found that kuruma prawns (Penaeus juponicus) fed astaxanthin had significantly higher survival than kuruma prawns that received similar amounts of dietary/&carotene or algal meal. Only dietary astaxanthin was effective in increasing survival compared to the control shrimps which received no dietary pigment. All shrimps that received dietary carotenoids had a significantly greater weight gain than the control shrimps.
4. Chemistry of astaxanthin and its stability in salmonid fish tissue Astaxanthin, an oxygenated carotene, has two chiral centers and may exist in three configurations: a pair of enantiomers (3&3’S) and (3R,3’R), and a mesoform (3R,3’S) (Fig. 2). The three stereo-isomers of astaxanthin are readily resolved and can be quantified using high-performance liquid chromatography (HPLC) with chiral columns (Maoka et al., 1985a,b). Astaxanthin is normally present in the flesh of wild salmonids at a concentration of about 8-37 ppm and this compound comprises over 99% of the total carotenoids present (Torrissen et al., 1989; Storebakken and No, 1992). As mentioned above, astaxanthin in salmonids is derived from the diet since salmonids cannot biosynthesize this com-
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pound (Simpson et al., 198 1; Torrissen et al., 1989; Storebakken and No, 1992). No epimerization occurs in the tissue of salmonid fish at the chiral centers at C3 and C-3’ in astaxanthin (Foss et al., 1984; Storebakken et al., 1985). The natural dietary sources of astaxanthin all contain astaxanthin in the enantiomeric RR or SS forms (Matsuno et al., 1984); thus wild salmonids will contain varying ratios of these two enantiomers depending on the relative amounts of each enantiomer present in the animal’s diet. Astaxanthin is naturally biosynthesized by the yeast Phaffi rhodozyma during its normal life cycle (Johnson and An, 199 1). The astaxanthin present in P. rhodozyma is in the 3R,3’Rconfiguration (Andrewes and Starr, 1976). The 3R,3’Rastaxanthin is present to some degree in all salmonid fish simply because this enantiomer is widely distributed in the natural foods consumed by salmonids in the wild (Matsuno et al., 1984; Schiedt et al., 198 1). Fig. 3 illustrates that the pigment extracted from wild salmonid fish contains the same stereoisomer of astaxanthin as Phufju as determined by HPLC with an optical resolution column that separates the three astaxanthin isomers (Maoka et al., 1985b). References Andrewes, A. and Starr, M., 1976. (3R,3’R)-Astaxanthin from the yeast Phafia rhodozyma. Phytochemistry, 15: 1009- 10 11. Chien, Y.M. and Jeng, S.C., 1992. Pigmentation of kuruma prawn, Penaeusjaponicus Bate, by various pigment sources and levels and feeding regimes. Aquaculture, 102: 333-346. Coutteau, P., Lavens, P. and Sorgeloos, P., 1990. Baker’s yeast as a potential substitute for live algae in aquaculture diets: Artemia as a case study. J. World Aquacult. Sot., 21: 1-9. Craik, J.C.A., 1985. Egg quality and pigment content in salmonid fishes. Aquaculture, 47: 61-88. Foss, P., Storebakken, T., Scheidt, K., Liaaen-Jensen, S., Austreng, E. and Streiff, K., 1984. Carotenoids in diets for salmonids. 1. Pigmentation of rainbow trout with the individual optical isomers of astaxanthin in comparison with canthaxanthin. Aquaculture, 4 1: 2 13-226. Hardy, R.W. and Stickney, R.R., 1992. Carotenoid Pigmentation of Salmon and Trout. Publication prepared March 19, 1992, and distributed privately, University of Washington, Seattle. Howell, B.K. and Matthews, A.D., 1991. The carotenoids of wild and blue disease affected farmed tiger shrimp (Penaeus monodon, Fabricus). Comp. Biochem. Physiol., 98B: 375-379. Johnson, E.A. and An, G.H., 199 1. Astaxanthin from microbial sources. Crit. Rev. Biotechnol., 1 I : 297-326. Katsuyama, M. and Matsuno, T., 1988. Carotenoid and vitamin A, and metabolism of carotenoids, /?-carotene, canthaxanthin, astaxanthin, zeaxanthin, lutein, and tunaxanthin in tilapia ( Tilapia nilotica). Comp. Biochem. Physiol., 90B: 131-139. Kurashige, M., Okimasu, E., Inoue, M. and Utsumi, K., 1990. Inhibition of oxidative injury of biological membranes by astaxanthin. Physiol. Chem. Phys. Med. NMR, 22: 27-38. Mahnken, C.V.M., 1991. Coho salmon farming in Japan. In: R.R. Stickney (Editor), Culture of Salmonid Fishes. CRC Press, Boca Raton, FL, pp. 13 1- 149. Maoka, T., Katsuyama, M.,Kaneko, N. and Matsuno T., 1985a. Stereochemical investigation of carotenoids in the antarctic krill Euphausia superba. Bul. Jpn. Sot. Sci. Fish., 5 1: 1671-1673. Maoka, T., Kumori, T. and Matsuno, T., 1985b. Direct diastereomeric resolution of carotenoids. I. 3-Hydroxy-4-oxo /J-end group. J. Chromatogr., 3 18: 122- 124. Matsuno, T., Maoka, T., Katsuyama, M., Ookubo, M., Katagiri, K. and Jimura, H., 1984. Occurrence of enantiomeric and meso-astaxanthin in aquatic animals. Bull. Jpn. Sot. Sci. Fish., 50: 15891592.
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Meyers, S.P. and Chen, H.M., 1982. Astaxanthin and its role in fish culture. In: R.R. Stickley and S.P. Meyers (Editors), Special Publication No. 3, World Aquaculture Society, Baton Rouge, LA, pp. 153-165. Miki, W., 199 1. Biological functions and activities of animal carotenoids. Pure Appl. Chem., 63: 14 l146. Oriental Yeast KK, 1982. Feed for rearing fish contains digested yeast body extract. Japan Patent No. JP-181447. Palozza, P. and Krinsky, L., 1992. Astaxanthin and canthaxanthin are potent antioxidants in a membrane model. Arch. Biochem. Biophys., 297: 29 l-295. Ranby, B. and Rabek, J.F. (Editors), 1978. Singlet Oxygen. Wiley, Chichester, 331 pp. Schiedt, K., Leuenberger, F. J. and Vecchi, M., 198 1. Natural occurence of enantiomeric and mesoastaxanthin. 5. Ex wild salmon (Safmo salar and Oncorhynchus). Helv. Chim. Acta, 64: 449-457. Schiedt, K., Levenberger, F.J., Vecchi, M. and Glinz, E., 1985. Absorption, retention and metabolic transformations ofcarotenoids in rainbow trout, salmon and chicken. Pure Appi. Chem., 57: 685692. Sedmak, J.J., Jolly, S.O. and Binkowski, F.P., 1992. An evaluation of Red Star@ Phafla yeast as a pigment source for salmonids. Red Star Bio Products, Technical Publication No. 1, Milwaukee, WI, 9 PP. Segner, H., Arend, P., von Poeppinghausen, K. and Schmidt, H., 1989. The effect of feeding astaxanthin to Oreochromis niloticusand Colisa lebrisa on the histology of the liver. Aquaculture, 79: 38 I 390. Simpson, K.L., Katayama, T. and Chichester, C.O., 198 1. Carotenoids in fish feeds. In: J.C. Bauernfeind (Editor), Carotenoids as Colorants and Vitamin A Precursors. Academic Press, New York, pp. 463-538. Storebakken, T. and No, H.K., 1992. Pigmentation of rainbow trout. Aquaculture, 100: 209-229. Storebakken, T., Foss, P., Austreng, E. and Liaaen-Jensen, S., 1985. Carotenoids in diets for salmonids. II. Epimerization studies with astaxanthin in Atlantic salmon. Aquaculture, 44: 259-269. Suntory Ltd., 1990. Composition containing astaxanthin as oxidation inhibitor useful as vitamin E substitute and as food additive, medicament, etc. Japan Patent JP-198947, 11 pp. Terao, O., 1989. Antioxidant activity of /?-carotene related carotenoids in solution. Lipids, 24: 659661. Torrissen, O.J., Hardy, R.W. and Shearer, K.D., 1989. Pigmentation of salmonids: Carotenoid deposition and metabolism. Rev. Aquat. Sci., 1: 209-225. van der Meeren, T., I99 1. Production of marine fish fry in Norway. World Aquacult., 22 (2): 37-40. Watanabe, T., Itoh, A., Murakami, A., Tsukashima, Y., Kitajima, C. and Fujita, S., 1984. Effect of nutritional quality of diets given to broodstock on the verge of spawning on reproduction of red sea bream. Bull. Jpn. Sot. Sci. Fish., 50: 1023-1028.