The degree of carotenoid esterification influences the absorption of astaxanthin in rainbow trout, Oncorhynchus mykiss (Walbaum)

The absorption of astaxanthin from diets (30 mg kg^?1 inclusion) supplemented with either unesterified astaxanthin; isolated astaxanthin monoesters, diesters or a cell‐free carotenoid extract from Haematococcus pluvialis were studied in rainbow trout (>200 g). No significant differences (P > 0.05) were recorded in the apparent digestibility coefficients (ADC) (≈60–65%) between astaxanthin sources. However, following consumption of a single meal, peak serum astaxanthin levels at 32 h (≈1.0–1.6 μg mL^?1) were significantly higher (P < 0.05) in fish fed unesterified astaxanthin and astaxanthin monoester, compared to fish fed astaxanthin diester and the cell free extract. However, no significant differences (P > 0.05) were recorded in serum astaxanthin uptake rates between sources of astaxanthin. Results suggest that the extent of carotenoid esterification negatively influences the peak serum levels of astaxanthin in rainbow trout.     

Comparison of diets for the tropical spiny lobster Panulirus ornatus: astaxanthin-supplemented feeds and mussel flesh

The fast‐growing tropical lobster Panulirus ornatus is a good aquaculture candidate generating increased research to develop potential feeds. We conducted a 12‐week experiment, assessing growth, survival and tissue carotenoid levels of juvenile P. ornatus. Lobsters were fed either pelleted feeds supplemented with astaxanthin and containing 30, 60, 90 or 120 mg total carotenoid kg^?1; or one of two fresh mussel reference feeds – blue Mytilus edulis and green‐lipped Perna canaliculus. There was no clear dose response, in terms of growth rate, to increasing dietary astaxanthin content; mussel‐fed lobsters had inferior growth rates. Twelve‐week survival was unaffected by treatment. Whole lobster carotenoid (4.7, 16.7, 27.8 and 32.8 mg kg^?1, dry matter basis) increased with increasing dietary astaxanthin; pre‐treatment carotenoid was 22.2 mg kg^?1. Apparent total carotenoid content of the mussel‐fed lobsters was unexpectedly high because of interference by other pigments. High‐performance liquid chromatographic analysis of free astaxanthin levels varied from a pre‐treatment value of 7.3 mg kg^?1 to 2.0, 7.6, 12.5 and 23.6 mg kg^?1 with increasing dietary astaxanthin, and 3.5 (green‐lip) and 5.9 (blue) mg kg^?1 for the mussel‐fed lobsters. Although dietary astaxanthin, over the investigated range, did not affect growth rate or survival, there was a dose–response increase in tissue carotenoid content and darkening of the exoskeleton pigmentation, which may have important implications for immunocompetency and marketing. These implications are discussed in the context of pelleted feed development for this species.     

Enhancement of non‐specific immune responses in common carp,Cyprinus carpio,by dietary carotenoids obtained from shrimp exoskeleton

The immunostimulatory role of carotenoid extract from shrimp processing discards was evaluated by feeding common carp fingerlings with a diet containing carotenoid extract for 21 weeks at 100 and 200 mg kg^?1 astaxanthin levels. Haemoglobin content was significantly (P < 0.05) higher in the blood of fish fed with carotenoid diet (>8.1 g dL^?1) compared with that from fish fed with carotenoid deficient diet (6.86 g dL^?1) and also leukocyte counts were higher (P < 0.05). No differences (P > 0.05) were observed in total serum protein, globulin level and albumin‐globulin ratio, but albumin content was higher (P < 0.05). Respiratory burst activity was significantly (P < 0.05) higher, the serum lysozyme activity almost doubled and the serum bactericidal activity was significantly increased when the fish were fed with diet containing 200 mg kg^?1 of astaxanthin but no significant differences were observed in serum trypsin inhibitory activity. There was a significant (P < 0.05) increase in leukocyte myeloperoxidase activity due to dietary carotenoids. Challenging fish with Aeromonas hydrophila after the feeding period resulted in 50% mortality in the control group while in the group fed with diet containing 100 mg kg^?1 astaxanthin, the mortality rate was 15%. No mortality and even symptoms of infection was not observed in the group fed with diet containing 200 mg kg^?1 of astaxanthin. The study indicated that carotenoid extract from shrimp processing discards can effectively be used as immunostimulants in aquaculture of carps and dietary carotenoids were found to enhance various immune defence mechanisms and also provide protection against the infection of pathogen A. hydrophila.     

Effect of dietary bile extracts on serum response of astaxanthin in rainbow trout (Oncorhynchus mykiss): a preliminary study

Effects of porcine bile extracts added at three different dietary concentrations 0, 10 and 20 g kg^?1 were studied on astaxanthin serum concentration in rainbow trout (mean weight 200 ± 7 g). Astaxanthin from micro‐algae Haematococcus pluvialis and synthetic astaxanthin (CAROPHYLL® pink) were incorporated in diets of rainbow trout at a rate of 100 mg astaxanthin kg^?1 of feed. Fish were hand fed twice a day. After 5 days of feeding there was a significant effect of the pigment source on the ratio (total blood astaxanthin per unit body weight to cumulative astaxanthin intake per unit body weight). Trout receiving synthetic astaxanthin showed a significantly (P < 0.05) higher ratio than trout fed algal astaxanthin. Increasing dietary bile extract did not lead to produce any effect on this ratio. The power of the statistical analysis is discussed. Therefore, the interaction (pigment source × dietary bile concentration) showed no more effect.     

Effect of esterification on the absorption of astaxanthin in rainbow trout, Oncorhynchus mykiss (Walbaum)

Gastrointestinal and serum absorption of astaxanthin was studied in rainbow trout, Oncorhynchus mykiss (Walbaum) (217 ± 2 g) fed diets supplemented with either esterified astaxanthin (from Haematococcus pluvialis) or free astaxanthin (synthetic, as 8% w/w beadlets) at similar levels (50 mg kg^?1). After 56 days of feeding, there was a significant difference (P = 0.0582) between steady‐state serum astaxanthin concentrations for fish fed free (2.0 ± 0.3 μg mL^?1) or esterified astaxanthin (1.3 ± 0.1 μg mL^?1) at the 90% confidence level. However, following ingestion of a single meal supplemented with free or esterified astaxanthin, the rates of astaxanthin absorption into serum were not significantly different (P > 0.1) (0.8 ± 0.2 µg mL^?1 h^?1 and 1.0 ± 0.4 µg mL^?1 h^?1 respectively). In fish fed both free or esterified astaxanthin, higher absorption (P < 0.05) of astaxanthin by the ileal (0.8 ± 0.14 μg g^?1 and 0.9 ± 0.15 μg g^?1 respectively) compared with the posterior (0.2 ± 0.01 μg g^?1 and 0.3 ± 0.14 μg g^?1 respectively) intestine was recorded. This confirmed the role of the anterior intestine in carotenoid absorption. Non‐detectable levels of esters in digesta taken from the hind intestine suggest the anterior intestine is also the primary region for ester hydrolysis.     

Natural bixin as a potential carotenoid for enhancing pigmentation and colour in goldfish (Carassius auratus)

Discovering natural carotenoids for colour enhancement and health benefits of fish is important to develop new feed formulations. We have purified natural bixin from achiote seeds and evaluated the effect of colour enhancing and pigmentation in goldfish. Varying levels of bixin‐based diets were prepared with 420 g kg^?1 of crude protein and 120 g kg^?1 of lipid content. Our results clearly showed that bixin (0.05, 0.10, 0.20 and 0.60 g kg^?1) based diets significantly (P < 0.05) enhanced the skin and fin colour at 30 and 60 days compared to diet without bixin. Interestingly, diet which contains 0.20 g kg^?1 bixin and commercial feed (with astaxanthin) had similar effect on carotenoid deposition in skin. Moreover, total carotenoid deposition in fin was higher than in skin of all bixin‐containing diets. However, 0.60 g kg^?1 bixin‐containing diet had lower specific growth rate (1.01 ± 0.01) and higher feed conversion ratio (2.05 ± 0.19) compared to the control group. The present results demonstrate that achiote bixin can be successfully used as an alternative natural carotenoid source against synthetic astaxanthin in fish feed. Our data indicate that 0.20 g kg^?1 is a suitable dietary level of bixin to ensure strong pigmentation, acceptable growth and feed utilization in goldfish.     

Utilization of carotenoids from various sources by rainbow trout: muscle colour,carotenoid digestibility and retention

Rainbow trout (Oncorhynchus mykiss) with a mean (sd) weight of 120 (2) g were fed diets supplemented with astaxanthin extracted from the yeast Phaffia rhodozyma (OY1 = 50 mg carotenoids kg^–1 feed, OY2 = 100 mg carotenoids kg^–1 feed), astaxanthin (AX = 100 mg astaxanthin kg^–1 feed) and canthaxanthin (CX = 100 mg canthaxanthin kg^–1 feed) for 4 weeks. Muscle analyses at the end of the experiment indicated a significantly higher carotenoid concentration in the AX group, while CX and OY1 groups were similar in spite of the differences in dietary concentration. The measure of total muscle colour difference (E^* _ab) between initial samples and 4 week ones was higher for the AX fish group but showed no significant difference between OY1, OY2, and CX. The hue and the reflectance ratio (R₆₅₀:R₅₁₀) of fish muscle increased in proportion to carotenoid intake. Digestibility (ADC) of yeast astaxanthin in OY1 and OY2 groups was significantly higher than that in the AX group. Canthaxanthin ADC was about one sixth of that of astaxanthin (AX group). Carotenoid retention in the muscle, expressed as a percentage of carotenoid intake, was higher for the AX group than that recorded for OY1 and OY2. According to ADC, carotenoid retention showed a marked lower value for the CX group. Muscle retentions were similar for astaxanthins from both sources.     

Diets supplemented with microalgal biomass: effects on growth,survival and colouration of ornamental fish Hyphessobrycon eques (Steindacher 1882)

This study evaluated the effects of the addition of microalgae (Ankistrodesmus gracilis and Haematococcus pluvialis) to the fish diet in improving the growth and optimal pigmentation (red carotenoid) of Hyphessobrycon eques. The basal mixed diets consisted of a formulated diet, supplemented with dried microalgae biomass of A. gracilis (1.5 g kg^?1) and H. pluvialis (1.5 g kg^?1). The live food diets contained zooplankton was cultured in open ponds, associated with microalgae. All the microalgae were cultured in the laboratory. No mortality was observed with any experimental diets. Fish performance results showed significant differences (P < 0.05) between the basal diet (BD) and the live food diet. The higher weight and total length were observed with mixed diets (BD + H. pluvialis and BD + A. gracilis). The mixed diets promoted more intense values of chroma (Cab*), lightness (L*) and redness (a*) to H. eques. Diaphanosoma birgei (Cladocera) represented more than 32% of zooplankton ingested by ornamental fish in live food (zooplankton and zooplankton + microalgae), and Argyrodiaptomus furcatus (Copepoda) was the species most ingested by H. eques in live food dietary treatment zooplankton. The feeding behaviour observed in the laboratory as well as the food preferences of H. eques was dependent on the zooplankton composition present in the used open ponds. This study showed that diets with microalgae and zooplankton were able to enhance the pigmentation of H. eques, being a good tool to benefit the culture management of this species.     

Dietary astaxanthin levels affect colour,growth, carotenoid digestibility and the accumulation of specific carotenoid esters in the Giant Tiger Shrimp,Penaeus monodon

The carotenoid astaxanthin (Axn) plays a vital role in shrimp pigmentation, with direct influence on product quality, and forms a significant cost component of shrimp aquaculture feeds. However, the effects of dietary Axn on other measures of shrimp physiological performance are varied, and the efficiency of carotenoid uptake from the diet and deposition in shrimp tissues is poorly defined. This study fed juvenile shrimp (Penaeus monodon) diets that contained 0, 25, 50 or 100 mg kg^?1 Axn for 6 weeks. Shrimp fed carotenoid‐free diets had significantly reduced colour and growth than those fed carotenoids, but survival was unaffected. Carotenoid digestibility improved as dietary carotenoid levels increased, and was 98.5% in shrimp fed 100 mg kg^?1 Axn. After 6 weeks, whole body carotenoid levels were significantly depleted in 0 or 25 mg kg^?1 fed shrimp, compared with those fed 50 or 100 mg kg^?1 or compared with initial shrimp. This study also showed that Axn monoesters were enriched with saturated fatty acids, whereas Axn diesters were enriched with monounsaturated and polyunsaturated fatty acids. Combined, these studies demonstrate that a total dietary carotenoid intake of between 25 and 50 mg kg^?1 Axn is required for normal shrimp growth and health in P. monodon. Evidence suggests that there is a functional role for the accumulation of carotenoids and the formation of specific Axn fatty acid esters, and these may be linked to the metabolism, storage, mobilization or deposition of Axn within various tissues.     

Given the same dietary carotenoid inclusion, Atlantic salmon, Salmo salar (L.) display higher blood levels of canthaxanthin than astaxanthin

Atlantic salmon, Salmo salar, fitted with permanent dorsal aorta cannulae were fed diets containing either 0, 30, 60 mg kg^?1 or combinations of astaxanthin and canthaxanthin, with the aim of comparing the uptake efficiencies to blood of the two pigments and evaluating possible interactions during absorption when formulated in the same diet. Given either astaxanthin or canthaxanthin in separate diets, at dietary levels of <30 mg kg^?1, an identical linear relationship (R² = 0.97) between dietary levels and blood concentrations was observed for both carotenoids. At dietary astaxanthin inclusions above 30 mg kg^?1, blood astaxanthin concentration approached saturation at an average level of 1.2 ± 0.04 μg mL^?1 (arithmetic mean ± SD), whereas blood levels of canthaxanthin continued to increase linearly throughout the inclusion range tested (0–60 mg kg^?1). When both carotenoids were presented in the same diet, a reduction in the absorption efficiency of both pigments was observed (P < 0.05). This manifested itself as a lower level in blood than the level observed when each carotenoid was administered separately. The negative interaction was most prominent for astaxanthin, the maximum average blood saturation level of which fell (P < 0.05) to 0.73 ± 0.03 μg mL^?1 (arithmetic mean ± SD). Our data support the conclusion that at higher dietary inclusions, canthaxanthin is more efficiently absorbed from the digestive tract into the blood of S. salar than astaxanthin.     

Effect of carotenoids and background colour on the skin pigmentation of Australian snapper Pagrus auratus (Bloch & Schneider, 1801)

Three 2‐factor experiments were conducted to determine the effects of background colour and synthetic carotenoids on the skin colour of Australian snapper Pagrus auratus. Initially, we evaluated the effects on skin colour of supplementing diets for 50 days with 60 mg kg^?1 of either astaxanthin (LP; Lucantin^® Pink), canthaxanthin (LR; Lucantin^® Red), apocarotenoic acid ethyl ester (LY; Lucantin^® Yellow), selected combinations of the above or no carotenoids and holding snapper (mean weight=88 g) in either white or black cages. In a second experiment, all snapper (mean weight=142 g) from Experiment 1 were transferred from black to white, or white to white cages to measure the short‐term effects of cage colour on skin L^*, a^* and b^* colour values. Skin colour was measured after 7 and 14 days, and total carotenoid concentrations were determined after 14 days. Cage colour was the dominant factor affecting the skin lightness of snapper with fish from white cages much lighter than fish from black cages. Diets containing astaxanthin conferred greatest skin pigmentation and there were no differences in redness (a^*) and yellowness (b^*) values between snapper fed 30 or 60 mg astaxanthin kg^?1. Snapper fed astaxanthin in white cages displayed greater skin yellowness than those in black cages. Transferring snapper from black to white cages increased skin lightness but was not as effective as growing snapper in white cages for the entire duration. Snapper fed astaxanthin diets and transferred from black to white cages were less yellow than those transferred from white to white cages despite the improvement in skin lightness (L^*), and the total carotenoid concentration of the skin of fish fed astaxanthin diets was lower in white cages. Diets containing canthaxanthin led to a low level of deposition in the skin while apocarotenoic acid ethyl ester did not alter total skin carotenoid content or skin colour values in snapper. In a third experiment, we examined the effects of dietary astaxanthin (diets had 60 mg astaxanthin kg^?1 or no added carotenoids) and cage colour (black, white, red or blue) on skin colour of snapper (mean weight=88 g) after 50 days. Snapper fed the astaxanthin diet were more yellow when held in red or white cages compared with fish held in black or blue cages despite similar feed intake and growth. The skin lightness (L^* values) was correlated with cage L^* values, with the lightest fish obtained from white cages. The results of this study suggest that snapper should be fed 30 mg astaxanthin kg^?1 in white cages for 50 days to increase lightness and the red colouration prized in Australian markets.     

Pigmentation of gilthead seabream, Sparus aurata (L. 1875), using Chlorella vulgaris (Chlorophyta, Volvocales) microalga

A feeding experiment was conducted over 9 weeks with seven groups of 30 (fish per group) unpigmented gilthead seabream, Sparus aurata (L. 1875) (initial mean weight = 145.2 ± 12.3 g). Three experimental diets were prepared by adding to a basal diet free of carotenoid (final pigment content of around 40 mg per kg feed): (i) a biomass of the carotenogenic Chlorella vulgaris (Chlorophyta, Volvocales); (ii) a synthetic astaxanthin; and (iii) a mixture (1:1) of microalgal biomass and synthetic astaxanthin. At 3‐week intervals, five fish were sampled from each tank for total carotenoids analysis in skin and muscle. The carotenoid pigments (total amount = 0.4%) identified in the carotenogenic alga were lutein (0.3%), β‐carotene (1.2%), canthaxanthin (36.2%), astaxanthin, free and esterified forms (55.0%), and other pigments (7.3%). Carotenoid pigments were significantly deposited in the four skin zones studied during the feeding trial: the forefront between the eyes, the opercule, along the dorsal fin and in the abdominal area. In the muscle, regardless of the astaxanthin source, the amount of carotenoids measured was very low (less than 1 mg kg^?1) and differences not significant. Moreover, no muscle pigmentation was evident, and there was no variation in the amount of carotenoid analysed in skin tissue, through the trial, for each treatment. It was concluded that supplementing the feed with C. vulgaris would be an acceptable practice in aquaculture to improve the market appeal of the gilthead seabream.     

Changes of carotenoids contents and analysis of astaxanthin geometrical isomerization in Haematococcus pluvialis under outdoor high light conditions

In this study, carotenoid contents of Haematococcus pluvialis during outdoor high light cultivation were measured, moreover, changes of astaxanthin geometrical isomers and biotic factors which may affect isomerization were investigated. During the incubation, contents of both astaxanthin and its precursors (zeaxanthin and canthaxanthin) increased over time, whereas the relative content of lutein decreased. Contents of all astaxanthin isomers increased, while the proportion of different astaxanthin geometrical isomers fluctuated during the incubation. All‐trans‐astaxanthin of total astaxanthin (T/A) was higher during the astaxanthin accumulation phase than that in the cell transformation phase, which was in contrast to results of 13‐cis‐astaxanthin of total astaxanthin (13C/A) and 9‐cis‐astaxanthin of total astaxanthin (9C/A). Farnesyl diphosphate synthase (trans (2E,6E)‐FPPS, EC2.5.1.10) and geranylgeranyl diphosphate synthase (GGPS, EC2.5.1.29), the key proteins involved in geometrical isomerization, decreased during the astaxanthin accumulation phase. Moreover, the presence of cis (2Z,6E)‐FPPS was firstly confirmed in H. pluvialis by HPLC‐MS/MS shotgun method. The results indicated the biotic factor (trans‐FPPS, cis‐FPPS and GGPS) may play an important role in astaxanthin geometrical isomerization of H. pluvialis, but not a crucial role. This study would help in optimizing the regulation of astaxanthin geometrical isomers in H. pluvialis, with great significance in theory and production.     

Effects of dietary astaxanthin concentration and feeding period on the skin pigmentation of Australian snapper Pagrus auratus (Bloch & Schneider, 1801)

A single‐factor experiment was conducted to investigate the effects of dietary astaxanthin concentration on the skin colour of snapper. Snapper (mean weight=129 g) were held in white cages and fed one of seven dietary levels of unesterified astaxanthin (0, 13, 26, 39, 52, 65 or 78 mg astaxanthin kg^?1) for 63 days. Treatments comprised four replicate cages, each containing five fish. The skin colour of all fish was quantified using the CIE L^*, a^*, b^* colour scale after 21, 42 and 63 days. In addition, total carotenoid concentrations of the skin of two fish cage^?1 were determined after 63 days. Supplementing diets with astaxanthin strongly affected redness (a^*) and yellowness (b^*) values of the skin at all sampling times. After 21 days, the a^* values increased linearly as the dietary astaxanthin concentration was increased before a plateau was attained between 39 and 78 mg kg^?1. The b^* values similarly increased above basal levels in all astaxanthin diets. By 42 days, a^* and b^* values increased in magnitude while a plateau remained between 39 and 78 mg kg^?1. After 63 days, there were no further increases in measured colour values, suggesting that maximum pigmentation was imparted in the skin of snapper fed diets >39 mg kg^?1 after 42 days. Similarly, there were no differences in total carotenoid concentrations of the skin of snapper fed diets >39 mg kg^?1 after 63 days. The plateaus that occurred in a^* and b^* values, while still increasing in magnitude between 21 and 42 days, indicate that the rate of astaxanthin deposition in snapper is limited and astaxanthin in diets containing >39 mg astaxanthin kg^?1 is not efficiently utilized. Astaxanthin retention after 63 days was greatest from the 13 mg kg^?1 diet; however, skin pigmentation was not adequate. An astaxanthin concentration of 39 mg kg^?1 provided the second greatest retention in the skin while obtaining maximum pigmentation. To efficiently maximize skin pigmentation, snapper growers should commence feeding diets containing a minimum of 39 mg unesterified astaxanthin kg^?1 at least 42 days before sale.     

Dietary esterified astaxanthin effects on color, carotenoid concentrations, and compositions of clown anemonefish, Amphiprion ocellaris, skin

This study examined the effects of dietary esterified astaxanthin concentration on coloration, accumulation of carotenoids, and the composition of carotenoids over time in the skin of Amphiprion ocellaris. Juveniles of 30 days-post-hatch were fed 40, 60, 80, or 160 mg esterified astaxanthin per kg diet (mg kg^?1) for 90 days. Skin coloration was analyzed using the hue, saturation, and luminosity model. Increased astaxanthin concentrations and duration on diet lead to improvements in skin color, that is, lower hues (~27–29 to ~14–17; redder fish), higher saturation (~77 to ~87 %), and lower luminosity (~43 to ~35 %). Fish fed 80 and 160 mg kg^?1 astaxanthin feed showed significant coloration improvements over fish fed lower astaxanthin feeds. Increasing both dietary astaxanthin concentration and time on the feed resulted in significant increases in total skin carotenoid concentration (0.033–0.099 μg mm^?2). Furthermore, there was a significant linear relationship between hue and total skin carotenoid concentration. Compositionally, free astaxanthin and 4-hydroxyzeaxanthin were the major skin carotenoids. 4-hydroxyzeaxanthin was previously unreported for A. ocellaris. Carotenoid composition was affected by duration on diet. Fraction 4-hydroxyzeaxanthin increased by ~15 %, while free astaxanthin decreased equivalently. The transition from 4-hydroxyzeaxanthin to free astaxanthin appears to follow a reductive pathway. Results suggest that managing coloration in the production of A. ocellaris juveniles requires manipulation of both dietary astaxanthin concentration and period of exposure to astaxanthin containing diet. In order to achieve more orange–red-colored fish, feeding 80–160 mg kg^?1 esterified astaxanthin for an extended duration is recommended.     

The pigmenting effect of different carotenoids on fancy carp (Cyprinus carpio)

The optimal concentration of a panel of individual and combined carotenoid sources on skin pigmentation in fancy carp was investigated by nine experimental diets that were formulated and supplemented with astaxanthin at 25 mg kg^?1, lutein at 25 and 50 mg kg^?1, β‐carotene at 25, 50 and 75 mg kg^?1, and lutein combined with β‐carotene at 25 : 25 and 50 : 50 mg kg^?1, while a diet without supplemented carotenoid served as a control. The results showed that serum TC of fish fed diets containing supplemented with lutein plus β‐carotene at 25 : 25; 50 : 50 mg kg^?1 and lutein 50 mg kg^?1 diet were higher than the other treatments (P ≤ 0.05). Serum TC of the respective treatments was 6.2 ± 2.0, 7.8 ± 3.3 and 7.3 ± 1.9 μg mL^?1 serum, respectively. Fish fed diets combined with lutein and β‐carotene at 25 : 25, 50 : 50 mg kg^?1 and lutein 50 mg kg^?1 diet had serum astaxanthin concentrations similar to fish fed the diet with astaxanthin alone at 25 mg kg^?1. Serum astaxanthin concentrations was 0.7 ± 0.01, 0.9 ± 0.01, 0.4 ± 0.02 and 1.7 ± 0.18 μg mL^?1 serum, respectively. The chromaticity of fish body skin of red and white position was assessed by colourimetry using the CIE L*a*b (CIELAB) system. Pigmentation response of skin redness of fancy carp fed with diets combined with lutein and β‐carotene at 25 : 25, 50 : 50 mg kg^?1 and lutein 50 mg kg^?1 were higher than other treatments (P ≤ 0.05) but they were similar to fish fed with 25 mg kg^?1 astaxanthin diet. The redness (a* values) of fish fed diets with diets combined with lutein and β‐carotene at 25 : 25, 50 : 50 mg kg^?1 and lutein 50 mg kg^?1 were 28.3 ± 0.53, 29.9 ± 1.38, 28.8 ± 3.95 and 28.5 ± 2.49, respectively. After 3 weeks of feeding the experimental diets, the fish fed on a diet without carotenoid supplement for one week demonstrated that the same three groups still retained their redness and had an overall tendency to improve skin colouring. Finally, concentrations 50 mg kg^?1 of lutein, or the combination of lutein and β‐carotene at 25 : 25 mg kg^?1 showed the highest efficiency for improving skin pigmentation and redness of skin.     

Dietary astaxanthin improved the body pigmentation and antioxidant function,but not the growth of discus fish (Symphysodon spp.)

To assess the effects of dietary astaxanthin on the growth and body colour of red discus fish (Symphysodon spp.), synthetic astaxanthin was added into the basal diet (beef heart hamburger) with the levels of 0 (control diet), 50, 100, 200, 300 and 400 mg kg^?1 respectively. The six experimental diets were fed to discus fish with an initial body weight of 10.3 ± 0.8 g for 8 weeks. The results showed that the supplementation of 50–200 mg kg^?1 astaxanthin had no significant effects on growth performance of discus fish, but the high supplementation of astaxanthin (300 or 400 mg kg^?1) significantly reduced the weight gain and increased the feed coefficient ratio (P < 0.05). After 4 or 8 weeks of feeding, the L* (lightness) values in astaxanthin‐supplemented groups were significantly lower, while a* (redness), b* (yellowness) and skin astaxanthin contents were significantly higher than the control group (P < 0.05). When the astaxanthin supplementation reached 200 mg kg^?1, skin redness and astaxanthin contents remained relatively stable. When b* was relatively stable, the supplemental astaxanthin was 300 (4 weeks) and 50 mg kg^?1 (8 weeks) respectively. With the supplemental astaxanthin increasing, the astaxanthin retention rate significantly decreased and hepatic total antioxidant capacity was strengthened. The dietary astaxanthin also significantly increased the reduced glutathione level (P < 0.05) when the astaxanthin inclusion was higher than 50 mg kg^?1. The above results showed that dietary astaxanthin could effectively improve the skin pigmentation of red discus fish in 4 weeks and the supplementation level was suggested to be 200 mg kg^?1.     

Effects of astaxanthin on Brachionus manjavacas (Rotifera) population growth

Astaxanthin is a red secondary carotenoid and powerful antioxidant that is used in aquaculture to enhance colour and improve fish health. Brachionid rotifers are often used as a live feed for larval fish, but do not contain endogenous carotenoids. However, they can be enriched with astaxanthin through their diet and transfer it to larval predators. When supplemented with 2 μg/ml astaxanthin oleoresin extracted from the green alga Haematococcus pluvialis, Brachionus manjavacas rotifer cultures reached significantly higher population densities and maintained them for longer. Furthermore, data are presented that exposure to oleoresin or pure astaxanthin enhances rotifer resistance to oxidative stress, a common cause for the collapse of rotifer mass cultures. Astaxanthin can be visualized in the gut of the rotifers, allowing the time course of uptake to be estimated by image analysis. Using this method, it was found that accumulation of astaxanthin in the rotifer gut saturates after 1.5 hr of exposure. The bioactive dose of astaxanthin oleoresin for rotifers was found to be 1–20 μg/ml. Astaxanthin concentration in rotifer tissues was measured using absorbance spectrophotometry. It was found that treating rotifers with 20 μg/ml for 24 hr; the concentration of astaxanthin absorbed into rotifer tissues was 2.6 mg/g. Overall, these experiments indicate that astaxanthin extracted from H. pluvialis can be used to improve the productivity and stability of rotifer mass cultures by increasing oxidative stress resistance and enhance the nutritional content of rotifers for larval fish.     

Effects of Haematococcus pluvialis supplementation on antioxidant system and metabolism in rainbow trout (Oncorhynchus mykiss)

Effects of commercial source for astaxanthin (Haematococcus pluvialis) (H.p) on antioxidant power, specific marker enzymes, and some metabolites were examined in rainbow trout (Oncorhynchus mykiss). Fish were fed on diets containing 1, 3, and 10 g microalga kg⁻¹ feed for 30 days. Serum total antioxidant activity and lipid peroxidation product, indicated by malondialdehyde (MDA), significantly enhanced with different doses of administration, indicating the elevated antioxidant status in all treatment groups. In group fed with high dose of alga, significantly elevated aspartate aminotransferase activity (AST) was noted, indicating damage of normal liver function in this group. Alkaline phosphatase (ALP) and alanine aminotransferase (ALT) were not affected in all groups. Although serum total protein remained unaffected, serum glucose level was decreased significantly in lower doses of administration. Furthermore, triglyceride and cholesterol levels showed significant decrease in 3 g kg⁻¹ microalga group by modulation of lipid metabolism in this group. On the other hand, in highest dose, significant increase in lipids was observed, indicating the slight dysfunction in lipid metabolism in this treatment group. The present study suggests that Haematococcus pluvialis especially in dose of 3 g kg⁻¹ feed administration may effectively enhance the antioxidant system and some biochemical parameters in rainbow trout.     

Effect of carotenoids on body colour of discus fish (Symphysodon aequifasciatus axelrodi Schultz, 1960)

This study assessed the effects of three kinds of carotenoids on the body colour of solid red discus fish (Symphysodon aequifasciatus axelrodi Schultz, 1960). Astaxanthin, xanthophylls and canthaxanthin were added into the beef heart diet at the level of 350 mg kg^?1 respectively. In the astaxanthin group (group A), the carotenoid concentration (CC) in the skin and dorsal fin reached saturation levels on days 40 and 20 respectively. However, CC consistently increased in the muscle. In the xanthophyll group (group B), CC in the skin increased through day 20; CC in the dorsal fin increased from days 10 to 20. In the canthaxanthin group (group C), CC in the skin increased during the first 20 days, reaching saturation levels on day 10 in the dorsal fin and muscle. On day 50, CC in the skin and muscle of group A was significantly higher than that of groups B or C. There were no significant differences in dorsal fin CC among the groups; however, CC in group C reached saturation levels in the shortest time. Therefore, astaxanthin was the most effective pigment for the skin and muscle; xanthophyll was the most effective pigment for the dorsal fin.