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In this study, we aimed to evaluate the effect of canthaxanthin (CX) and iodine (I) on the production of laying hens, on counteracting debilitation of the vitelline membrane, and on inhibiting Salmonella growth in eggs stored at 30°C. Three hundred hens were reared in cages. Birds were divided into six feeding groups (10 hens × 5 repetitions) that were administered 0, 3 or 6 ppm of CX and 1 or 10 ppm of I with their diets. Laying rate, egg weights, and feed conversion ratios were controlled. The quality of fresh eggs was assessed in wks 25-26, 48-50 and 62-63 of hens lives. An additional batch of eggs was incubated at the temperature of 30°C, and egg quality changes were monitored on days 3, 6 and 9 of storage. Additionally, eggs collected from four experimental groups of hens whose diets had been iodated with 1 or 10 ppm of I and supplemented with 0 or 6 ppm of CX were infected under laboratory conditions with Salmonella, and incubated for 5 and 10 d. The laying rate, egg weights, and feed conversion ratio were significantly improved. Dietary inclusion of CX contributed to a higher resistance of the vitelline membrane of egg yolks, but only for fresh eggs. Vitelline membrane degradation during egg storage at 30°C was significantly counteracted by dietary inclusion of I at a dose of 10 ppm. The same I dose resulted in the complete inhibition of Salmonella growth until day 10 of incubation, but exclusively for eggs collected from 40-week-old hens. Dietary supplementation with 10 ppm of I was found to impart high level of resistance to the vitelline membrane against the growth of Salmonella in case of eggs stored at 30°C; therefore, I was found to be more beneficial by ensuring longer preservation than that of CX. However, dietary supplementation with CX was found to increase the resistance of vitelline membrane in fresh eggs.

Three experiments were conducted to investigate the effects of maternal dietary canthaxanthin (CX) and 25-hydroxycholecalciferol (25-OH-D3) supplementation on antioxidant status and calcium-phosphate metabolism of progeny ducks. Cherry Valley duck breeders (38 wk old) were fed either a control diet or the same diet plus CX (6 mg/kg) and 25-OH-D3 (0.069 mg/kg) for 32 weeks. Experiments 1, 2, and 3 were conducted with progeny ducks hatched from eggs laid by duck breeder hens at 54, 62, and 70 wk of age, respectively. Progeny ducks from both maternal treatments were fed with the same NRC (1994) vitamin regimen starter (1 to 14 d) and finisher (15 to 35 d) diets in experiments 1 and 2, and fed with the same high vitamin regimen starter (1 to 14 d) and finisher (15 to 35 d) diets in experiments 3. High vitamin regimen had higher levels of all vitamins, except biotin, than the NRC (1994) vitamin regimen. In experiment 1, maternal CX and 25-OH-D3 increased (P < 0.05) shank pigmentation and tibiotarsus ash and tended to decrease (P < 0.1) liver total superoxide dismutase activity (T-SOD) of one-day-old progeny ducks; and increased (P < 0.05) shank pigmentation, decreased (P = 0.05) liver protein carbonyl, and tended to increase (P < 0.1) liver total antioxidant capacity (T-AOC) of 14-day-old progeny ducks. In experiment 2, maternal CX and 25-OH-D3 increased (P < 0.05) shank pigmentation and liver T-AOC and decreased (P < 0.05) liver protein carbonyl of one-day-old progeny ducks, but increased (P < 0.05) the serum phosphate level of 14-day-old progeny ducks. In experiment 3, maternal CX and 25-OH-D3 increased (P < 0.05) shank pigmentation of one-, 14-, and 35-day-old progeny ducks and tended to increase (P < 0.1) liver T-SOD and tibiotarsus ash, but decrease (P < 0.1) liver malondialdehyde of one-day-old progeny ducks. It can be concluded that progeny dietary high vitamin regimen could partially prevent maternal CX-derived progeny shank pigmentation from bleaching. Maternal CX- and 25-OH-D3-derived effects are influenced by the hen's age and progeny's dietary vitamin regimen.

A meta-analysis was conducted to evaluate the effect of the carotenoid canthaxanthin on production performance in layer hens. The data set contained 576 performance measurements from 34 trials. The trials were all conducted according to a similar experimental protocol from 1997 to 2012. The age of the animals ranged from 21 to 65 weeks. The experimental diets were predominantly wheat/SBM based, fed in mash form ad libitum. Depending on the trial and the treatment group considered, canthaxanthin supplementation was in the range of 0 to 8 mg/kg of feed.Using a linear mixed model regression applied to all 34 trials simultaneously, significant dose-dependent increases were found in egg yolk mass (+0.53% per ppm of canthaxanthin inclusion in the feed, P < 0.001), in egg mass (+0.47% per ppm, P = 0.0132), egg weight (+0.17% per ppm, P = 0.046), and in feed intake (+0.32% per ppm, P = 0.0054). A numerical increase was found in egg production (+0.28%, P = 0.14). The FCR decreased numerically (-0.24% per ppm, P = 0.36). The deposition of canthaxanthin in the egg yolk was 2.25 ppm per ppm of canthaxanthin in the feed (P < 0.001). It is concluded that in addition to egg yolk pigmentation, antioxidant effect, enhanced reproduction, and immune-modulation, canthaxanthin can significantly increase egg mass, thereby enhancing the productivity of the flock. Mechanisms relating to carotenoid metabolism and functions in avian species are a new research area that will require further investigation to explain the observed effects., (© 2017 Poultry Science Association Inc.)

This study was designed to evaluate the effect of corn or sorghum diet and canthaxanthin (CX) supplementation on performance in broiler breeders. A total of 440 females with similar body weight (BW) (3.71 ± 0.14 kg) and 60 roosters were placed in an open-sided house with 20 pens, randomly distributed in a factorial arrangement (2 × 2). There were 4 diets of 2 ingredients; corn (CO) or sorghum (SO) and 2 levels of CX; 6 mg/kg (CX) and 0 mg/kg (NCX) totaling 5 replicate pens of 22 females and 3 males each, from 42 to 65 wk, divided in 2 periods (from wk 42 to 53 and wk 54 to 65). Birds' BW was measured every 28 d and mortality rate was calculated at the end of trial. Egg production (%), egg specific gravity (g/cm3), egg weight (g), yolk weight (%), albumen weight (%), eggshell weight (%) and yolk colorimetric score were measured weekly. Incubation parameters were recorded in 12 incubations to evaluate hatching eggs, hatching (%), hatchability (%), fertility (%), weight of the chicks born and their quality. The BW, mortality, percentage of yolk and albumen weight, fertility and some incubation parameters were not affected (P > 0.05) by diets used. An increase in the egg production, hatching eggs, chicks born and first quality chick by hen at the second period were observed in CX breeder's diets (P = 0.0066; P = 0.0060; P = 0.0368; P = 0.0326). Egg specific gravity and eggshell weight were improved at the first period by SO+CX diet (P = 0.0138; P = 0.0209) and the same effect to egg weight, but at the second period (P = 0.0251). The CX was well absorbed from the diet and effectively transferred to the egg yolk, thereby increasing egg yolk pigmentation in the both periods (P < 0.0001). The CX supplementation in broiler breeder diets improved the productive and reproductive performance (laying% and hatchable eggs) at the second period, also to the both periods improved the egg yolk pigmentation., (© 2017 Poultry Science Association Inc.)

This study investigated the effects of maternal canthaxanthin (CX, 6 mg/kg) and 25-hydroxycholecalciferol (25-OH-D 3 , 0.069 mg/kg) supplementation on the performance of Cherry Valley ducklings under two different vitamin regimens. A total of 780 duck breeder females and 156 males were randomly allotted to two diets with or without the addition of the mixture of CX and 25-OH-D 3 (CX+25-OH-D 3 ) for 32 weeks. Ducklings (males and females separately) hatched from eggs laid at 24 weeks of the duck breeder trial were fed with a NRC vitamin regimen, and ducklings (males and females separately) hatched from eggs laid at 32 weeks of the duck breeder trial were fed with a HIGH vitamin regimen (had higher levels of all vitamins except biotin than NRC vitamin regimen), for 14 days. The results showed that, maternal CX+25-OH-D 3 supplementation increased the shank pigmentation for 7-days post hatch in ducklings under a NRC vitamin regimen, and for 14-days post hatch in ducklings under a HIGH vitamin regimen. Growth performance, antioxidant status and serum phosphorus of ducklings under a NRC vitamin regimen were increased by maternal CX+25-OH-D 3 supplementation; however, these positive effects were not observed in ducklings under a HIGH vitamin regimen. Males revealed increased growth performance in ducklings under both NRC and HIGH vitamin regimens. Sexual differences in shank pigmentation, antioxidant status, tibia strength and serum phosphorus were not consistent as they were dependent on maternal CX+25-OH-D 3 status or dietary vitamin regimens. Data suggest that maternal CX+25-OH-D 3 supplementation is important for starter ducklings under a NRC vitamin regimen, but not HIGH vitamin regimen., (Journal of Animal Physiology and Animal Nutrition © 2016 Blackwell Verlag GmbH.)

The objective of this study was to examine the effects of canthaxanthin (Cx) treatment during in vitro maturation (IVM) of porcine oocytes on embryonic development after parthenogenetic activation (PA) and somatic cell nuclear transfer (SCNT), on intracellular glutathione (GSH) and reactive oxygen species (ROS) levels in mature oocytes, and on gene expression in both PA- and SCNT-derived blastocysts. To determine the optimal effective concentration of Cx, porcine oocytes were cultured in IVM medium supplemented with various concentrations (0, 20, 40 and 80 μM) of Cx for 22 hr. Compared to other groups, supplementation with 40 μM Cx significantly improved blastocyst formation rates after PA (p < .05), but no significant differences were observed among groups in total blastocyst cell numbers. Subsequently, oocytes were cultured in IVM medium supplemented with or without 40 μM Cx. Oocytes treated with 40 μM Cx showed significantly increased cleavage and blastocyst formation rates after SCNT compared to the control group (p < .05). Moreover, significantly increased intracellular GSH and reduced ROS levels were observed in the Cx-treated group (p < .05). In addition, both PA- and SCNT-derived blastocysts from the 40 μM Cx-treated group showed significantly increased mRNA expression of Bcl2 and Oct4 and decreased Caspase3 expression level (p < .05), when compared with the control group. PA-derived blastocysts from the 40 μM Cx-treated group also exhibited significantly decreased expression of Bax (p < .05). Our results demonstrated that treatment with 40 μM Cx during IVM improves the developmental competence of PA and SCNT embryos. Improvement of embryo development by Cx is most likely due to increased intracellular GSH synthesis, which reduces ROS levels in oocytes, and it may also positively regulate apoptosis- and development-related genes., (© 2016 Blackwell Verlag GmbH.)

Background: Natural xanthophylls, canthaxanthin and astaxanthin are known to exhibit anticancer activity. However, the dietary effects of canthaxanthin and astaxanthin on N-methyl-N-nitrosourea (MNU)-induced mammary cancer remain controversial, and their mechanisms of action have not been clearly identified., Materials and Methods: Three-week-old female Sprague-Dawley rats were fed a xanthophyll-free (basal diet) diet or experimental diets containing canthaxanthin or astaxanthin (0.04% and 0.4%) for 5 weeks (until 8 weeks of age), after which all rats were provided the basal diet (n=15 each). Rats were administered MNU at 6 weeks of age, and the incidence of mammary tumors at 20 weeks of age was compared. The expression of adiponectin in mammary adipose tissues taken at 7 weeks of age was also compared., Results: Compared to the basal diet group, the 0.4% (but not the 0.04%) astaxanthin diet significantly reduced the incidence of palpable mammary carcinoma (92% vs. 42%; p<0.05), while the low and high canthaxanthin diets produced no significant inhibition. Adiponectin immunoblotting showed significantly higher expression in the 0.4% astaxanthin diet group, while the other groups were similar to the basal diet group., Conclusion: High concentrations of astaxanthin suppress MNU-induced mammary carcinoma. Changes in adiponectin may be involved in the mechanism of action., (Copyright © 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.)

Effect of feeding partially saturated canthaxanthin (PSC), purified from Aspergillus carbonarius mutant, was studied using four groups of female albino rats (n=6) for 4 weeks. While the control group received basal diet ad libitum, Groups I, II and III were fed with basal diet containing 50, 100 and 250 ppm PSC, respectively. PSC feeding did not cause any significant changes in food intake and there was no gain in body weight either. PSC included in the diet significantly decreased cholesterol in blood. There was 44.75% and 60.54% decrease in LDL-cholesterol in rats fed with 50 and 100 ppm carotenoid. Hepatic ascorbic acid content increased by 44.59% in rats fed with 50 ppm PSC. Dietary PSC at 250 ppm lowered lipid peroxides by 19.49%. Activities of antioxidant enzymes, glutathione transferase and catalase were significantly higher in serum and liver of PSC fed rats compared to the controls. The results suggested that PSC feeding can induce hypocholesterolmic and antioxidant properties in rats., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Background: The storage life of frozen salmonids is often limited primarily by oxidation and flesh discolouration due to carotenoid degradation. The objective of this research was to determine the carotenoid changes and therefore the muscle colour modifications during 6 months of frozen storage (-18 °C) of whole rainbow trout fed astaxanthin (100 ppm) or canthaxanthin (80 ppm), kept under two different packagings: plastic film and cardboard box., Results: After 6 months of frozen storage, the carotenoid type effect was seen for dry matter while there was no packaging material effect on carotenoid, total lipids, and TBARS contents of trout fillets. The time under frozen storage had an effect on carotenoid and TBARS fillet concentration. The carotenoid-type effect was noted for the four colour parameters (lightness difference, chroma difference, hue angle difference, and total colour difference) of rainbow trout fillet, while the packaging material effect was observed only for chroma. Frozen storage time had an effect on the four colour parameters of rainbow trout fillet., Conclusion: Carotenoid and packaging material effects were more marked for colour parameters than for biochemical parameters. In this study, as fish were frozen and stored as whole fish, fish skin provided good protection against oxidation., (Copyright © 2011 Society of Chemical Industry.)

This study investigated the effects of beta-carotene and canthaxanthin on lipid peroxidation and antioxidative enzyme activities in rats fed a high-cholesterol, high-fat diet. Wistar rats were divided into six groups. Negative control group (group NC) received a high-fat (150 g/kg) diet; cholesterol control group (group CC) received a high-cholesterol (10 g/kg), high-fat diet. The other four groups were fed a high-cholesterol, high-fat diet supplemented with crystal beta-carotene (group BC), beta-carotene beadlet (group BB), canthaxanthin beadlet (group CX) or alpha-tocopherol (group AT). Blood and livers were collected for analysis after 6 weeks of feeding. Group BB had significantly lower hepatic thiobarbituric acid reactive substance (TBARS) and conjugated diene concentrations, whereas group CX had a significantly lower plasma TBARS concentration than did group CC. In erythrocytes, glutathione peroxidase activities were significantly greater in groups BC, BB and CX than in group CC. Moreover, compared with group CC, catalase activities were significantly greater in groups BB and CX, and superoxide dismutase (SOD) activity was significantly greater in group BB. In livers, SOD activities were significantly greater in groups BC, BB and CX, and glutathione reductase activities were significantly greater in groups BB and CX than in group CC. Compared with group CC, hepatic retinol and alpha-tocopherol concentrations were significantly greater in groups BC, BB and CX, whereas plasma and hepatic cholesterol concentrations were significantly lower in group BC. These findings suggest that beta-carotene and canthaxanthin altered the pro-oxidation and antioxidation balance and suppressed cholesterol-induced oxidative stress via modulation of antioxidant system and cholesterol metabolism.

This study was designed to assess the effect of dietary soybean phospholipids on canthaxanthin transport by serum lipoproteins and canthaxanthin muscle deposition in trout. Three groups of 12 immature trout in triplicate with a mean body weight of 130 g were fed with three experimental diets containing (1) canthaxanthin plus lecithin plus fish oil, (2) canthaxanthin plus lecithin, and (3) canthaxanthin alone, for 12 days. The two major lipoprotein classes in rainbow trout are high-density lipoproteins, which transport principally carotenoids present in the serum, and low-density lipoproteins, which are responsible for the transport of cholesterol, both independently of the administered diet. In addition, very low density lipoproteins are responsible for triglyceride transport in serum. Nevertheless, the amount of canthaxanthin in the serum increased when carotenoid was associated with phospholipids plus fish oil. When canthaxanthin is transported by lecithin plus fish oil, the amount of phospholipids, cantaxanthin, and cholesterol deposited in muscle increased but not significantly. The highest apparent canthaxanthin digestibility coefficient was obtained when canthaxanthin was carried by lecithin plus fish oil. The administration of canthaxanthin carried by phospholipids improved its accumulation in the muscle of rainbow trout. This accumulation could be enhanced if the time of administration of canthaxanthin is increased.

A comparative investigation of tissue carotenoid distribution between rainbow trout, Oncorhynchus mykiss, and Atlantic salmon, Salmo salar, was undertaken to identify the relative efficiency of utilization of astaxanthin and canthaxanthin. Higher apparent digestibility coefficients (ADCs) (96% in trout vs. 28-31% in salmon; P<0.05), and pigment retention efficiencies (11.5-12.5% in trout vs. 5.5% in salmon; P<0.05), for both astaxanthin and canthaxanthin, were observed for rainbow trout. Astaxanthin deposition was higher than canthaxanthin in rainbow trout, while the reverse was true for Atlantic salmon, suggesting species-specificity in carotenoid utilization. The white muscle (95% in trout vs. 93% in salmon) and kidneys (0.5% in trout vs. 0.2% in salmon) represented higher proportions of the total body carotenoid pool in rainbow trout than in Atlantic salmon (P<0.05), whereas the liver was a more important storage organ in Atlantic salmon (2-6% in salmon vs. 0.2% in trout; P<0.05). The liver and kidney appeared to be important sites of carotenoid catabolism based on the relative proportion of the peak chromatogram of the fed carotenoid in both species, with the pyloric caecae and hind gut being more important in Atlantic salmon than in the rainbow trout. Liver catabolism is suspected to be a critical determinant in carotenoid clearance, with higher catabolism expected in Atlantic salmon than in rainbow trout.

Broiler carcass skin color is important in the United States and Mexico. This study evaluated the use of natural and synthetic pigments in broiler diets at commercial levels. Birds were fed natural or synthetic pigments at low or high levels, simulating US and Mexican commercial practices. Skin color was measured during live production (3 to 7 wk of age) and after slaughter and chilling. The natural pigments had consistently greater skin b* values (yellowness) than the synthetic pigments. The high levels produced greater skin b* values than the low levels, regardless of source. The synthetic pigments had a slower increase in skin b* but reached the same level as the natural low by 7 wk. There was no difference in skin a* values (redness) due to pigment source or level or the age of the bird. By 7 wk, all pigment sources approached plateau levels in the blood, but the synthetic pigment diet produced higher blood levels of yellow and red pigments than the natural pigment diets. Processing intensified skin yellowness and reduced skin redness. These data suggest that although synthetic pigments might have been absorbed better than natural ones, natural pigments were more efficient at increasing skin yellowness and there were only small differences between high and low levels for each pigment source. This finding may allow reduction in pigment use and feed cost to achieve the same skin acceptance by the consumer.

1. Effects of canthaxanthin supplementation of the maternal diet on the antioxidant system of the developing chick were investigated. 2. Three hundred and twenty female broiler breeder birds were housed in one of 4 controlled environment rooms with 3 replicates for all treatments, with the exception of the control treatment of which there were 4 replicates. All birds received one of 5 diets: control low xanthophyll diet, or the same diet supplemented with 3, 6, 12 or 24 mg/kg canthaxanthin in the form of Carophyll Red. At 30 weeks of age 60 eggs from each of the 5 groups were incubated. At d 16 of the embryo development, at d 1 and d 7 posthatch tissue samples were collected and analysed by HPLC-based methods. 3. Canthaxanthin accumulation in the egg yolk was proportional to dietary content. Furthermore, at 12 to 24 mg/kg canthaxanthin was associated with an increase in gamma-tocopherol concentration in the egg yolk. Canthaxanthin was transferred from the egg yolk to the developing embryo and, as a result, its concentration in the liver of the embryo at 16 and in 1-d-old chicks was increased. Even at d 7 posthatch canthaxanthin concentration in the chicken liver was elevated. 4. Canthaxanthin supplementation of the maternal diet at 12 mg/kg was associated with an increased alpha-tocopherol concentration in the liver of 1-d-old chicks and resulted in decreased tissue susceptibility to lipid peroxidation. 5. Canthaxanthin supplementation at 6 to 24 mg/kg was also associated with a delay in alpha-tocopherol depletion from the liver for 7-d posthatch. As a result of the increased canthaxanthin and vitamin E concentrations in the liver of 7-d-old chicks, tissue susceptibility to lipid peroxidation decreased. 6. The results support an idea that dietary carotenoids can modulate antioxidant systems of the developing chicken.

In this study the effect of luminal lipid on the absorption of canthaxanthin (CTX) was investigated using the lymph duct cannulated rat. Treatments were emulsions designed to deliver increasing amounts of olive oil (10, 30, 50, 70, or 90 mg/h) and CTX (12.5 nmol/h). Emulsions were continuously infused into the duodenum for 12 h, and lymph was collected during the final 6 h of infusion for analysis. As the amount of lipid in the emulsion increased, a linear increase in the absorption of CTX was observed. The recovery of CTX in the lymph when infused with 10 mg/h olive oil was 14.2 +/- 1.2% and with 90 mg/h was 26.9 +/- 5.7%. The efficiency of CTX absorption nearly doubled by increasing the amount of lipid infused with CTX. The correlation between lipid load and CTX absorbed was r= 0.85. We conclude that luminal lipid load affects CTX absorption.

A 2 x 2 x 2 factorial experiment was planned to study the influence of dietary fat source (linseed oil or sunflower oil) and dietary doses of alpha-tocopheryl acetate (alpha-TA) (0 or 200 mg/kg of feed) and canthaxanthin (CX) (0 or 5 mg/kg of feed) on fatty acid (FA) composition and lipid oxidation of fresh and spray-dried eggs. Dietary supplementation with alpha-TA and CX modified the levels of certain long-chain polyunsaturated FA (PUFA). Lipid oxidation in fresh eggs and spray-dried eggs at 0, 6, and 12 mo of storage was measured by the lipid hydroperoxide (LHP) and TBA values. The LHP and TBA values were up to 10 times higher in spray-dried eggs than in fresh eggs. The evolution of LHP and TBA values in spray-dried eggs showed that omega3 FA-enriched eggs were more susceptible to lipid oxidation than those enriched with omega6 FA. The omega-TA supplementation increased the lipid stability of enriched eggs and was very effective throughout the storage of spray-dried eggs. On the other hand, CX supplementation did not prevent lipid oxidation in PUFA-enriched eggs. Moreover, no synergistic effect between both compounds was detected.

Purpose: To establish the threshold level of canthaxanthin crystals in the retina of cynomolgus monkeys. To correlate the spatial distribution of all-trans canthaxanthin and its metabolites with the grade of crystals., Methods: Monkeys were orally administered 0, 0.2, 0.6, 1.8, 5.4, 16.2, and 48.6 mg/kg body wt canthaxanthin daily for 2.5 to 3 years. A second group of monkeys were administered 200 and 500 mg/kg body wt/d for 5 years. Ophthalmoscopy, electroretinography (ERG), retina and carotenoid analysis were performed as previously reported., Results: Crystals in the retina periphery were observed by ophthalmoscopy preterminally only in the extreme high doses of 200 to 500 mg/kg body wt/d. There were no adverse effects on visual functions as measured by ERG. Crystals in the peripheral retina, and/or in the macula, were detected microscopically in all canthaxanthin treated groups except at the lowest dose of 0.2 mg/kg body wt/d. The grade of crystals increased up to a dose of 16.2 mg/kg body wt/d. Dose-dependent increases in canthaxanthin content also were noted in the retina, the liver, and in plasma. All-trans canthaxanthin was the major compound in the peripheral and paracentral retina of very highly dosed animals, where its concentration correlated largely with the grade of inclusions. In the macula, 4'-OH-echinenone was the dominant canthaxanthin metabolite., Conclusions: The grade of crystals in monkey retinas was dose dependent with a threshold level at 0.6 mg canthaxanthin/kg body wt/d. It correlated in the retinal periphery with the concentrations of all-trans-canthaxanthin and in the macula with its metabolites.

The 4,4'-diketo-beta-carotene, canthaxanthin, alters tocopherol status when fed to Balb/c mice, suggesting an involvement of carotenoids in the modulation of oxidative stress in vivo. We investigated further the modifications induced by an oral administration of canthaxanthin on lipid peroxidation, antioxidant enzymes and iron status in liver of Balb/c mice. Female 6-wk-old Balb/c mice were randomly divided into two groups (n = 10/group). The control group (C) received olive oil alone (vehicle) and the canthaxanthin-treated group (Cx) received canthaxanthin at a dose of 14 microg/(g body wt.d). The 15-d canthaxanthin treatment resulted in carotenoid incorporation but did not modify lipid peroxidation as measured by endogenous production of malondialdehyde (MDA). However, glutathione peroxidase activity was 35% lower (P<0.01) and catalase (59%, P<0.005) and manganese superoxide dismutase (MnSOD) (28%, P<0.05) activities were higher in canthaxanthin-treated mice than in controls. Moreover, carotenoid feeding caused a significant (P<0.05) overexpression of the MnSOD gene; mRNA levels of the enzyme were greater in treated mice than in controls. Concomitantly, a 27% (P<0.05) greater iron concentration was found in liver from canthaxanthin-treated mice compared with controls. These findings support the hypothesis that canthaxanthin alters the protective ability of tissues against oxidative stress in vivo.

The in vivo modulatory effect of beta-carotene, astaxanthin and canthaxanthin on lymphocyte function was investigated. Female BALB/c mice (8 wk old) were fed a basal diet containing 0, 0.1% or 0.4% beta-carotene, astaxanthin or canthaxanthin for 0, 2 or 4 wk (n = 8/diet/period). Splenic lymphocytes were isolated and mitogen-stimulated proliferation, IL-2 production and lymphocyte cytotoxicity were assessed. Body weight and feed intake were not different among dietary treatments. Plasma carotenoids were undetectable in unsupplemented mice but concentrations of the respective carotenoids were elevated in mice fed 0.1 or 0.4% beta-carotene (0.22 and 0.39 mumol/L), astaxanthin (16.4 and 50.2 mumol/L) and canthaxanthin (5.00 and 7.02 mumol/L) respectively. Mice fed both dietary levels of beta-carotene and astaxanthin had enhanced phytohemagglutinin-induced lymphoblastogenesis compared to unsupplemented mice (P < 0.03). No treatment difference was detected with concanavalin A- or lipopolysaccharide-induced lympho-proliferation nor with IL-2 production (P < 0.05). Astaxanthin (0.1%) also enhanced lymphocyte cytotoxic activity (P < 0.08). In contrast, canthaxanthin did not significantly influence any of the lymphocyte functions measured. Results indicate that beta-carotene and astaxanthin but not canthaxanthin exert enhanced splenic lymphocyte function in mice.

The anticancer activities of beta-carotene, astaxanthin and canthaxanthin against the growth of mammary tumors were studied in female eight-wk-old BALB/c mice. The mice were fed a synthetic diet containing 0, 0.1 or 0.4% beta-carotene, astaxanthin or canthaxanthin. After 3 weeks, all mice were inoculated with 1 x 10(6) WAZ-2T tumor cells into the mammary fat pad. All animals were killed on 45 d after inoculation with the tumor cells. No carotenoids were detectable in the plasma or tumor tissues of unsupplemented mice. Concentrations of plasma astaxanthin (20 to 28 mumol/L) were greater (P < 0.05) than that of beta-carotene (0.1 to 0.2 mumol/L) and canthaxanthin (3 to 6 mmol/L). However, in tumor tissues, the concentration of canthaxanthin (4.9 to 6.0 nmol/g) was higher than that of beta-carotene (0.2 to 0.5 nmol/g) and astaxanthin (1.2 to 2.7 nmol/g). In general, all three carotenoids decreased mammary tumor volume. Mammary tumor growth inhibition by astaxanthin was dose-dependent and was higher than that of canthaxanthin and beta-carotene. Mice fed 0.4% beta-carotene or canthaxanthin did not show further increases in tumor growth inhibition compared to those fed 0.1% of each carotenoid. Lipid peroxidation activity in tumors was lower (P < 0.05) in mice fed 0.4% astaxanthin, but not in those fed beta-carotene and canthaxanthin. Therefore, beta-carotene, canthaxanthin and especially astaxanthin inhibit the growth of mammary tumors in mice; their anti-tumor activity is also influenced by the supplemental dose.

The effect of 16 d intake of 300 mg carotenoids/kg diet (beta-carotene (beta C), bixin (BX), lycopene (LY), lutein (LU), canthaxanthin (CX) or astaxanthin (AX) on xenobiotic metabolizing enzymes in the liver, lung, kidney and small intestine of male Wistar rats was assessed. A control group received the basal diet (AIN-76) without carotenoids and a positive control group for enzyme induction received 3-methylcholanthrene (3-MC) at 666 mg/kg diet. Cytochrome P450 activity was assessed using the substrates ethoxyresorufin for P450 1A1, methoxyresorufin for P450 1A2, pentoxyresorufin for P450 2B1/2 and benzyloxyresorufin for P450 types 1A1/2, 2B1/2 and 3A. Glutathione-S-transferase (EC 2.5.1.18) and reduced glutathione status were assessed. Carotenoid uptake by the tissues was also determined. 3-MC and the carotenoids BX, CX and AX led to significant increases compared with control in liver, lung and kidney ethoxyresorufin-O-deethylation. Methoxyresorufin-O-demethylation activity was significantly increased in liver and lung by BX, CX and AX but only CX and AX significantly increased activity in kidney. Pentoxyresorufin-O-depentylation and benzyloxyresorufin-O-dearylation increased in liver of 3-MC-, BX-, CX- and AX-treated rats, but to a much lesser degree than for the other two substrates. Benzyloxyresorufin-O-dearylation in lung was significantly decreased by all carotenoids. Activities of any of the measured enzymes in the small intestine were undetectable in all treatment groups except the 3-MC group. Glutathione status was unaffected by any of the treatments. This is the first study identifying the carotenoids BX, CX and AX as inducers of rat lung and kidney xenobiotic metabolizing enzymes.

The effects of oral doses of canthaxanthin on tissue distribution of alpha- and gamma-tocopherols were investigated in three experiments in male and female Balb/c mice. Mice were assigned to receive canthaxanthin [7 or 14 microg/(g body weight.d)] or placebo (olive oil) by gavage for different periods of time (0, 1, 2, 4 and 6 wk). A 2 wk-treatment with canthaxanthin resulted in incorporation of the carotenoid in all tissues analyzed, including liver, spleen, kidney, lung and heart. In liver, the maximum accumulation of the carotenoid was reached after 2 wk of dosing in female mice and after 6 wk in male mice. Canthaxanthin incorporation was accompanied by changes in alpha- and gamma-tocopherol concentrations in plasma and tissues. These included the following: 1) a significant increase (P < 0.001) in alpha-tocopherol concentration in spleen (21 and 27% in male and female mice, respectively) after 2 wk and in liver ( approximately 50% in both male and female mice) after 6 wk; 2) a significant decrease in gamma-tocopherol concentration in plasma (P < 0.05) and tissues (P < 0.001) after 2 wk of treatment. In female mice, this decrease was 55% in plasma, 43% in liver, 44% in kidney, 71% in lung and 70% in heart. In male mice, the decrease was observed only in plasma (30%), kidney (54%) and heart (46%). In liver, the decrease in gamma-tocopherol concentration was both dose- and time-dependent and significantly (P < 0.001) greater in female than in male mice. We conclude that dietary administration of canthaxanthin modifies tocopherol status in murine tissues.

We report the case of a 4-year-old girl, who had been suffering for 2 years from a recurrent, painful crisis affecting both hands, following sun exposure. There were no obvious cutaneous lesions, which initially caused us to consider a diagnosis of a psychiatric disorder. However, the diagnosis of erythropoietic protoporphyria was then established by the demonstration of elevated levels of free protoporphyrin in erythrocytes. The present case illustrates the effectiveness of beta-carotene associated with canthaxanthin in erythropoietic protoporphyria.

We investigated the plasma appearance of beta-carotene and canthaxanthin, an oxycarotenoid, in normolipidemic premenopausal women (n = 9) who ingested beta-carotene alone, canthaxanthin alone, and a combined dose. Blood samples were taken hourly for 12 h; additional blood samples were collected over 528 h. In a subset of the women (n = 5), plasma lipoproteins were separated into chylomicrons, very-low-density-lipoproteins (VLDL) subfractions, intermediate-density lipoproteins (IDLs), and low-density lipoproteins (LDLs). The appearance of beta-carotene in plasma was biphasic, with a minor peak at 5 h followed by a sustained peak at 24-48 h. The plasma appearance of canthaxanthin was monophasic, with a rapid increase to the final hourly measurement at 12 h and a steady decrease from the next measurement at 24 h. At 6 h, 23.4 +/- 2.9% of the increase in plasma canthaxanthin was associated with LDL, in contrast with 2.4 +/- 1.4% of the increase in plasma beta-carotene (P < 0.005). Ingestion of a combined dose of beta-carotene and canthaxanthin inhibited the appearance of canthaxanthin in plasma, chylomicrons, and each VLDL subfraction (P < 0.05), but did not significantly affect the rapid accumulation of canthaxanthin in LDL within 10 h. In contrast, ingestion of the combined dose did not significantly affect the appearance of beta-carotene in plasma or plasma lipoproteins. These findings suggest distinct mechanisms of incorporation into lipoproteins and specific interactions of beta-carotene and canthaxanthin during intestinal absorption in humans.

In 2 x 3 factorial experiments, 240 broiler chicks were fed diets containing 0, 0.01, and 0.02% beta-carotene or canthaxanthin with or without 5 ppm aflatoxin to determine the effects of these two carotenoids on the health and well-being of broilers subjected to aflatoxin poisoning. Neither beta-carotene nor canthaxanthin was effective at overcoming the growth-depressing effects of aflatoxin. Relative liver weights were significantly higher in broilers receiving dietary aflatoxin in the presence of beta-carotene but not canthaxanthin. beta-Carotene and canthaxanthin had no effect on antibody production against infectious bursal disease (IBD). Interestingly, secondary antibody production against IBD was enhanced by the presence of aflatoxin in the diet. Canthaxanthin significantly increased the concentrations of cholesterol, total protein, uric acid, and triglyceride, all of which were significantly depressed by aflatoxin. beta-Carotene did not effect any of the measured blood analytes. There was a significant interaction between canthaxanthin and aflatoxin with respect to creatine kinase activity. Creatine kinase activity decreased as dietary canthaxanthin increased in the presence of aflatoxin. The data suggest that beta-carotene is not effective at ameliorating aflatoxicosis in broiler chickens but that canthaxanthin may be somewhat effective with respect to certain clinical blood chemistry indicators.

The assays of several phase I and phase II xenobiotic-metabolizing enzyme activities, as well as CYP1A immunoblot analysis, were performed in liver microsomes and cytosol of male C57BL/6 mice (Ah receptor-responsive), of male DBA/2 mice (Ah receptor-low responsive) and of female Ah receptor gene knockout mice that were fed diets containing 300 mg/kg of a nonprovitamin A carotenoid, canthaxanthin, or a provitamin A carotenoid, beta-apo-8'-carotenal for 14 days, or which were injected i.p. with 3-methylcholanthrene. Previous studies have shown that some carotenoids, such as canthaxanthin and beta-apo-8'-carotenal, are strong inducers of liver CYP1A1 and 1A2 when given to rats. In this work, only canthaxanthin induced both CYP1A1 and 1A2 in C57BL/6 mice, whereas beta-apo-8'-carotenal induced only CYP1A2 in this strain. Neither of the two carotenoids modified CYP1A1/2 protein contents or enzyme activities in Ah receptor-low responsive DBA/2 or in Ah receptor gene knockout mice. Cytosol prepared from C57BL/6 mice liver tissue was incubated with [3H] 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the presence of canthaxanthin or beta-apo-8'-carotenal and analyzed by sucrose density gradient sedimentation: neither of the carotenoids, even when present in large excess, competed with TCDD for the TCDD binding site of the cytosolic Ah receptor of C57BL/6 mice. In brief, the carotenoids canthaxanthin or beta-apo-8'-carotenal induced Cyp1a genes in mice through an Ah receptor-dependent pathway, but did not bind to the Ah receptor.

Purpose: To reproduce and investigate in a primate animal model the phenomenon of the red carotenoid canthaxanthin (beta, beta-carotene-4'4'-dione) to induce crystal-like retinal deposits as they have been observed in the ocular fundus of humans after high canthaxanthin intake (i.e., more than 30 mg/day)., Methods: Groups of four cynomolgus monkeys (Macaca fascicularis) per gender and dose were administered 5.4, 16.2, or 48.6 mg canthaxanthin/kg body weight daily by oral gavage for 2.5 years. Eight control animals received placebo. In vivo ophthalmoscopy was performed at intervals of 3 months along with electroretinography after 12 and 24 months and retinal biomicroscopy just before the monkeys were killed. Retinal wholemounts or frozen sections were investigated postmortem by polarization, bright field, and differential interference contrast microscopy. Retinal and preterminal plasma canthaxanthin concentrations were determined by high-performance liquid chromatography (HPLC)., Results: By ophthalmoscopy and retinal biomicroscopy in vivo, no crystals or other light-reflecting particles were observed in the central paramacular retina. However, postmortem polarization microscopy of all 24 canthaxanthin-treated animals showed a circular zone in the peripheral retina containing birefringent, polymorphous red, orange, or white inclusions. The density of these inclusions was diminished within 1 to 8 mm posterior to the ora serrata. These inclusions were located mainly in the inner retinal layers, that is the nerve fiber layer and ganglion cell layer, inner plexiform layer, and inner nuclear layer. Twelve of the 24 canthaxanthin-treated animals showed yellow, golden birefringent inclusions in the macula. Retinas of placebo-treated monkeys were free of birefringent, crystal-like inclusions. The HPLC confirmed the presence of all-trans canthaxanthin, and 4-OH-echinenone and isozeaxanthin as well, in the retinas of all canthaxanthin-treated animals. Neither electroretinography nor histopathology indicated any adverse effects of the canthaxanthin-induced retinal inclusions seen in this study., Conclusions: A high intake of canthaxanthin for 2.5 years led to the deposition of crystal-like birefringent inclusions in the inner layers of the peripheral retina and, to some extent, the central retina of cynomolgus monkeys. The presence of these deposits did not interfere with morphology nor with retinal function.

The carotenoids beta-carotene and canthaxanthin and the retinoid 13-cis-retinoic acid (13-RA) have inhibited oral carcinogenesis in the hamster cheek pouch (16 wks, 3 times/wk at 1.4 mg/kg) induced by an 0.5% solution of 7, 12-dimethylbenz[a]anthracene (DMBA). However, 13-RA at a higher dose (> 2.0 mg/kg per treatment) increased squamous cell carcinoma growth (Eur J Cancer Clin Oncol 24, 839-850, 1988). 13-RA, beta-carotene, and canthaxanthin administered to 60 hamsters (16 wks, 3 times/wk, 10 mg/kg) altered neovascularization characterized by immunohistochemistry for transforming growth factor-alpha (TGF-alpha) and factor VIII. 13-RA + DMBA resulted in more smaller-sized tumors, with a reduced volume and tumor burden (tumor controls, 185.9; 13-RA + DMBA, 151.0). The carotenoids reduced the number and the sizes of the carcinomas formed (beta-carotene, 60 tumors, 142.3 x 10(3) mm3; canthaxanthin, 30 tumors, 116.1 x 10(3) mm3). Factor VIII and TGF-alpha were expressed in high intensity at cancer sites of the 13-RA + DMBA and DMBA groups with > 50 and > 10 cells, respectively, per x 400 field. In contrast, beta-carotene- and canthaxanthin + DMBA-treated pouches showed > 20 and 5 cells, respectively, per x 400 field for factor VIII and TGF-alpha. These results suggest that 13-RA treatment may increase vascular growth, but the carotenoids also produced enhanced levels of endothelial cell growth and TGF-alpha compared with the untreated mucosa. The carotenoids may enhance tumor growth under the appropriate carcinogenic environment.

The antitumor effect of canthaxanthin in BALB/c mice bearing a transplantable thymoma was investigated. Male or female mice received two different doses of canthaxanthin (7 or 14 micrograms/g body wt/day) starting 15 days before tumor inoculation (7 x 10(7) cells i.p.). Canthaxanthin treatment delayed the appearance of macroscopic ascites and prolonged animal survival. This effect was dose dependent and more evident in females than in males. It appeared only when the carotenoid was administered before tumor transplantation. The antitumor efficacy of the carotenoid was related to its tissue incorporation. Canthaxanthin was incorporated in a dose-dependent manner in liver and thymoma cells and to a larger extent in females than in males. Our study shows the antitumor efficacy of canthaxanthin in vivo against a transplantable murine thymoma and points out the importance of dose, administration timing, and sex in the antitumor efficacy of this compound.

The ability of dietary supplementation with carotenoids to protect chick tissues against oxidative stress in vitro was examined. Male Leghorn chicks were fed on diets supplemented (100 mg supplement/kg diet) with either beta-carotene, zeaxanthin (beta,beta-carotene-3,3'-diol), canthaxanthin (beta,beta-carotene-4,4'-dione) or alpha-tocopherol, or on a control diet, from 1 d old until 37 d of age. Tissues (liver, heart, skeletal muscle and plasma) were removed and assayed for total carotenoids and alpha-tocopherol content and portions subjected to oxidative stress by incubation of homogenates with cumene hydroperoxide and FeSo4. Animals receiving zeaxanthin and canthaxanthin had significantly greater carotenoid concentrations in liver, heart, muscle and plasma compared with untreated controls (P < 0.05); animals fed on diets supplemented with beta-carotene, or alpha-tocopherol did not have significantly different tissue carotenoid contents compared with untreated controls. alpha-Tocopherol supplementation elevated alpha-tocopherol levels in all tissues examined (P < 0.05). Supplementation with carotenoids did not affect tissue alpha-tocopherol levels, but beta-carotene lowered plasma alpha-tocopherol levels by 50% (P < 0.05). Incubation of plasma or tissue homogenates with oxidant stressors induced lipid peroxidation (production of thiobarbituric-acid reactive substances) in all tissues. Animals given alpha-tocopherol, beta-carotene or zeaxanthin had a reduced susceptibility to oxidant stress in liver compared with unsupplemented controls (P < 0.05), and alpha-tocopherol-supplemented animals had reduced susceptibility in skeletal muscle compared with tocopherol-supplemented animals had reduced susceptibility in skeletal muscle compared with unsupplemented controls (P < 0.05). Canthaxanthin supplementation did not influence the susceptibility to oxidant stress in any tissue examined. These results suggest that zeaxanthin, a carotenoid present in animal and human diets, may have significant activity as an antioxidant against oxidative stress in tissues.

Using mouse skin papilloma as a model system, we examined whether the antitumorigenic activity of carotenoids was related to their provitamin A activity. Oral administration of canthaxanthin (CX) or beta-carotene at 200 mg/kg/day for 14 days significantly reduced the cumulative size of papillomas induced on the skin by 9,10-dimethyl-1,2-benzanthracene (p < 0.05), after the accumulation of these carotenoids in the tumors. The levels of a protooncogene, c-myc, were simultaneously suppressed in papillomas in carotenoid-treated mice. Because CX cannot be converted metabolically to retinoids, these results suggested that CX directly inhibited the growth of papillomas. Neither the accumulation of retinoids nor the expression of a retinoic acid-inducible gene, retinoic acid receptor-beta, was found in papillomas of CX- and beta-carotene-treated mice, suggesting that, like CX, beta-carotene might exert the tumor-suppressing effect without being converted to retinoids. Thus a certain antitumorigenic activity of carotenoids appears not necessarily to require their provitamin A activity.

To test whether beta-carotene or canthaxanthin can modulate the initiation of liver preneoplasia by diethylnitrosamine (DEN) in a sequential protocol of hepatocarcinogenesis, for three weeks male weanling rats were fed diets containing beta-carotene or canthaxanthin (300 mg/kg diet) or excess vitamin A (70,000 IU/kg diet) or were given beta-carotene by injection (9 injections at 10 mg/kg body wt ip). On Day 15, all rats were injected with 200 mg DEN/kg body wt ip; later they were submitted to 2-acetylaminofluorene treatment and to two-thirds hepatectomy, then to phenobarbital treatment, after which gamma-glutamyltranspeptidase- and placental glutathione-S-transferase-positive liver foci were histologically detected. Neither beta-carotene (fed or injected), canthaxanthin, nor an excess of dietary vitamin A had an influence on the number and size of preneoplastic liver foci, despite a significant incorporation and persistence in liver of both carotenoids, especially canthaxanthin, and of supplemental vitamin A. These results are in conflict with another report in which beta-carotene, given to rats during the initiation phase, was found to strongly inhibit DEN-induced hepatocarcinogenesis.

The modulating effects of dietary feeding of two xanthophylls, astaxanthin (AX) and canthaxanthin (CX) during the postinitiation phase on colon carcinogenesis initiated with azoxymethane (AOM) were investigated in male F344 rats. Animals were initiated with AOM by weekly s.c. injections of 15 mg/kg body wt for 3 weeks and then they were fed the diets containing AX or CX at concentrations of 100 and 500 p.p.m. for 34 weeks. The others contained the groups of rats treated with AX or CX alone and untreated. At the end of the study (week 37), the incidence and multiplicity of neoplasms (adenoma and adenocarcinoma) in the large intestine of rats initiated with AOM and followed by AX or CX containing diet at a high dose (500 p.p.m.) were significantly smaller than those of rats given AOM alone (P < 0.001). In addition, AX or CX feeding significantly inhibited the development of aberrant crypt foci induced by AOM. Dietary exposure to AX or CX also decreased cell proliferation activity as revealed by measuring 5'-bromodeoxyuridine-labeling index as crypt cells, colonic mucosal ornithine decarboxylase activity and blood polyamine levels. These results indicate that AX and CX are possible chemopreventers for carcinogenesis of colon in addition to urinary bladder and oral cavity and such effects may be partly due to suppression of cell proliferation.

Preruminant calves, maintained in a monogastric state by feeding an all-liquid diet, were used to compare the serum appearance and lipoprotein transport of five different carotenoids over 144 h. Thirty newborn calves were fed milk replacer for 1 wk and then randomly assigned to six groups (n = 5), with each group receiving a single 20-mg oral dose of beta-carotene in water-soluble beadlets, canthaxanthin in water-soluble beadlets, lutein in oil, lycopene in oil, crystalline alpha-carotene in oil or crystalline beta-carotene in oil as part of a morning meal. Serial blood samples were taken by jugular puncture for up to 1 wk post-dosing. Lipoprotein separation and analysis were completed with selected animals. All carotenoids were absorbed, but in variable amounts. At peak serum carotenoids levels, HDL contained 70-90% of the carotenoids. Canthaxanthin and lutein peaked earlier in serum (8 and 12 h) than did the less polar lycopene, alpha-carotene, and beta-carotene (16, 24 and 24 h). Canthaxanthin and lutein were also cleared more quickly from the serum. Serum concentrations of alpha-carotene and lycopene displayed slower disappearance rates than did beta-carotene. The peak serum level (nmol/L +/- SEM) of canthaxanthin (392 +/- 136) was lower than that of beta-carotene (1245 +/- 425), and carotenoids levels of calves receiving these commercial beadlet sources were higher than the serum levels of calves receiving beta-carotene (45 +/- 17.5), alpha-carotene (42 +/- 18.0), lutein (51 +/- 9.5) and lycopene (18 +/- 4.6), which were fed in oil.(ABSTRACT TRUNCATED AT 250 WORDS)

Hydroa vacciniforme is a very rare photodermatosis, commencing in childhood and belonging to the category of idiopathic burns. The inconstancy of the therapeutic results raises the possibility of a clinical syndrome that can have different origins. We present a case that responded very well to carotenoids.

Objective: This study investigated effects of ingestion of a combined dose of beta-carotene and canthaxanthin on their individual pharmacokinetics in serum., Methods: During three 5-day study periods, two subjects ingested either a 25 mg dose of beta-carotene, a 25 mg dose of canthaxanthin, or a combined dose of 25 mg each of beta-carotene and canthaxanthin. Pharmacokinetics of the individual and combined doses were compared within subjects., Results: Ingestion of a concurrent beta-carotene dose reduced the peak serum canthaxanthin concentration by 38.8 +/- 6.5%, and the 24- and 72-hour areas under the serum canthaxanthin concentration-time curves by 38.1 +/- 6.4 and 34.4 +/- 7.4%, respectively. The suggested antagonism between beta-carotene and canthaxanthin was not reciprocal; beta-carotene inhibited the appearance of canthaxanthin in serum but canthaxanthin did not inhibit the appearance of beta-carotene., Conclusion: Our data suggest that ingestion of a combined pharmacologic dose of beta-carotene and canthaxanthin reduces the bioavailability of the canthaxanthin dose.

The purpose of this paper is to address the very important problem of accurate statistical analysis of certain types of cancer inhibition/promotion (IP) experiments. These experiments are routinely used by the National Cancer Institute to test the effects of potential chemopreventative agents. The statistical analysis is difficult since there is Type I censoring. In the IP experiments under investigation, laboratory animals (rats) are injected with a single dose of either a direct or indirect acting carcinogen. In the mammary tumor system, animals in the control group generally develop 5-7 tumors and typical experiments are usually terminated after 4-6 months. Animals are sacrificed at the end of the experiment and all observed tumors are confirmed. The two most common response variables are the number of observed tumors per animal and the rate of tumor development. The difficulty in analyzing these experiments occurs because experiments are terminated before all induced tumors have been observed. Fewer observed tumors in one group compared to another could be the result of a decreased number of induced tumors, a decrease in growth rate, or a combination of both. It is essential for the experimenter to distinguish between these two different biological actions. Present statistical techniques do not account for this confounding and since they rely primarily on nonparametric procedures, do not present an accurate description of potential IP agents. In this paper we introduce a parametric procedure that explicitly acknowledges the confounding present in experiments of this nature. The analysis is based on the comparison of the mean number of tumors per group (lambda) and the mean time to tumor appearance (mu). A longer mean time to development is believed to indicate a slower tumor growth rate. Hypothesis tests are developed to determine if there is an overall experiment effect, to isolate which groups are contributing to an observed experiment effect, and to isolate factors (tumor number and/or growth rate) contributing to an observed group difference. Confidence regions for (lambda, mu) are also generated. This analysis leads to a better understanding of how potential IP agents function.

Interactive effects of an oral dose of equal quantities of beta-carotene and either canthaxanthin or lycopene on serum and tissue beta-carotene accumulations were investigated in domestic ferrets. Like humans, ferrets absorb a substantial portion of ingested beta-carotene intact and accumulate it in tissues. After the ferrets ingested a low carotenoid purified diet for 13 d, they were randomly assigned to one of two groups of six animals. One group was dosed with beta-carotene (10 mg/kg body weight) and the other with beta-carotene and either canthaxanthin (Experiment 1) or lycopene (Experiment 2) (10 mg/kg body weight for each). In Experiment 1, ferrets that received a combined dose of beta-carotene and canthaxanthin had serum beta-carotene concentrations that were significantly lower at 8, 12 and 24 h post-dosing (P < 0.05), compared with those that received an individual dose of beta-carotene; liver, adrenal and kidney beta-carotene concentrations were also significantly reduced. In Experiment 2, ferrets that received a combined dose of lycopene and beta-carotene had lower serum and tissue beta-carotene concentrations than in those that received beta-carotene alone; the differences were not statistically significant with the exception of serum beta-carotene concentrations at 24 h post-dosing. The results suggest that, at the doses given, a concurrent oral canthaxanthin dose has a specific antagonistic effect on the bioavailability of a beta-carotene dose in ferrets.

The supplementation with beta-carotene and canthaxanthin, two carotenoids with and without pro-vitamin A activity, reduced in vitro the number of micronuclei induced in human cultured lymphocytes by the chemotherapeutic radiomimetic drug bleomycin. The genotoxic activity of this substance is supposed to be mediated by a free radical mechanism. The reduction of micronucleated cells was in correlation with donors' carotenoid blood levels. It has been observed that 20 weeks are needed, following the end of carotenoid supplementation, in order to return to the high levels of micronuclei induced by bleomycin. It is suggested that this protective effect is due to the antioxidant properties of carotenoids acting against free radical-mediated genotoxic damage.

In a study of the effects of carotenoids, canthaxanthin (CA), beta-apo-8'-carotenal (BA), or beta-carotene in an extract of Spirulina-Dunaliella algae (AE) was fed at 0%, 0.1%, or 0.2% in a choline-deficient (CD) diet. In each of eight groups, 10 adult male Fischer 344 rats were fed diets with designated carotenoid sources and levels or a choline-sufficient diet for 12 weeks. Carotenoids altered some of the changes induced by the CD diet. Increases in enlargement of fatty livers and low plasma cholesterol levels occurred in rats fed 0.2% BA. Plasma retinol was further reduced 35% by BA or AE. BA and AE increased liver total vitamin A about 80% and 305%, respectively. Liver lipid peroxidation was enhanced and plasma alpha-tocopherol was reduced further by 1.0% AE. AE, BA, and CA (mg/g fat) depressed liver alpha-tocopherol about 49%, 67%, and 78%, respectively. The decreased liver alpha-tocopherol was concurrent with an increase in carotenoid stores of CA > BA > AE. Histopathological examination of sections of liver tissue by light microscopy showed fatty and cirrhotic changes in all rats fed CD diets. Histochemical evaluation based on a semiquantitative assay revealed a marked increase in peroxisome enzyme activity in the livers of all CD rats. None of the carotenoids appeared to have any effect on the development of morphological changes in the liver. Although carotenoids can function as antioxidants, they did not prevent changes observed in rats fed CD diets.

After long-term treatment with high dosages, canthaxanthin causes a characteristic retinopathy with circular, macula surrounding crystals. As changes in retinal functionning disappear relatively easily after withdrawal of the drug, the crystals dissolve rather slowly--over about several years. Five patients showing a profound crystalline retinopathy were re-examined with an average of 69.7 months after withdrawal of the canthaxanthin-containing drug. Three of the patients were treated for erythropoetic protoporphyria (EPP) with Phenoro (2/5 beta-carotene, 3/5 canthaxanthin), two sisters took a canthaxanthin-containing formulation (1/8 beta-carotene, 7/8 canthaxanthin) for cosmetic reasons. Two female patients complained about an increased glare sensitivity, which was explainable for one of them with a subcapsular cataract. The retinal crystals decreased quite differently. Minor deffects of the retinal pigment epithelium remained unchanged in two patients. They increased slightly in the female patient with the smallest crystal formation but highest plasma cholesterol. Shortly after withdrawal of the drugs usually an increase of a-wave amplituded of the electroretinograms was found. The a-waves returned to normal and the b-wave amplitudes showed an increase up to the final control paralleling the reduction of the retinal crystals. A- and b-wave peak latencies which were prolonged under treatment returned to normal.

The use of HPLC has established that chickens possess unexpected metabolic abilities to acylate, deacylate, reduce, and oxidize carotenoids. The use of HPLC permits more consistent and more economic pigmentation of carcasses and of egg yolks. Hopefully, the use of HPLC will raise pigmentation from an art to a science. Apparently, HPLC will be an essential tool in terms of future efforts to understand and master the process of poultry pigmentation.

The effect of dietary canthaxanthin, retinyl palmitate, or their combination on the tumor-infiltrating T-lymphocyte response (T-TIL) in de novo murine ultraviolet type B irradiation-induced tumors was investigated to elucidate potential mechanisms of action of these compounds. We found that dietary canthaxanthin greatly reduced the number of tumor-infiltrating helper/inducer, suppressor/cytotoxic, and interleukin-2 receptor-positive T lymphocytes and also observed a concomitant statistically significant increase in tumour incidence in canthaxanthin-fed animals. The addition of retinyl palmitate to the canthaxanthin diet ameliorated this negative effect on TIL and the development of skin tumors. We conclude that dietary retinyl palmitate and canthaxanthin can modulate the host T-cell immune response within a growing tumor and may affect tumorigenicity.

beta-Carotene and excess vitamin A have been shown to reduce plasma alpha-tocopherol when fed to young rats. The present study assessed the effects of beta-carotene, excess vitamin A and canthaxanthin (4,4'-diketo-beta-carotene) on carotenoid, alpha-tocopherol and iron status in adult retired breeder rats. Male 8- to 10-mo-old rats (10/group) were fed varying levels of vitamin A as retinyl palmitate, beta-carotene and canthaxanthin ad libitum for 8 wk. The AIN-76A diet was modified to contain 16% (wt/wt) fat and 50% carbohydrate (control) plus beta-carotene or canthaxanthin at 0, 0.048 (BC1 or CX1) and 0.2% (BC2 or CX2) of the diet. These compounds were fed with and without excess retinyl palmitate (RP, 220 mg/kg). Higher relative liver weights were observed in CX- and RP-fed groups. Plasma retinyl esters were detected in all RP-fed groups. Plasma retinyl palmitate was 1.6- and 1.5-fold higher in RP-BC and RP-CX groups, respectively, than in the RP groups. Plasma and liver beta-carotene and canthaxanthin were 11-54% and 26-74% lower, respectively, with excess retinyl palmitate feeding. Feeding canthaxanthin and retinyl palmitate but not beta-carotene, resulted in lower levels of plasma alpha-tocopherol. Liver non-heme iron levels were also lower in CX-fed rats irrespective of retinyl palmitate feeding. These results extend to adult rats previous findings that excess retinyl palmitate alters vitamin E and carotenoid status prior to the manifestation of clinical signs of hypervitaminosis A. Additionally, canthaxanthin feeding lowers alpha-tocopherol and iron status in adult rats.

A method, using two different systems, is described for the high-performance liquid chromatographic analysis of retinol, retinal, retinoic acid, retinyl acetate, retinyl palmitate, alpha-, beta- and gamma-carotene, beta-apo-6'-, beta-apo-8', beta-apo-10'- and beta-apo-12'-carotenal, ethyl beta-apo-8'-carotenoate, alpha-tocopherol and alpha-tocopheryl acetate. The first system consists of a laboratory-packed Hypersil-ODS 3-microns column and a mobile phase of acetonitrile-methylene chloride-methanol-water (70:10:15:5, v/v). The second system consists of a laboratory-packed Hypersil-ODS 3-microns column and a mobile phase of acetonitrile-methylene chloride-methanol-water (70:10:15:5, v/v). The second system consists of a laboratory-packed Nucleosil C18 3-microns column and a mobile phase of acetonitrile-0.1 M ammonium acetate (80:20, v/v). The detection limits in standard solutions were 10 ng/ml for retinoids and carotenoids and 60 ng/ml for the E vitamers. Analysis of the tissues and plasma of rats, after 2 weeks on a diet supplemented with either beta-carotene or canthaxanthin (both 2 mg/g), led to the conclusion that the rats were able both to transport and store beta-carotene and canthaxanthin and to convert beta-carotene to retinol. Incubation of cytosol preparations from the mucosa of the small intestine of rat with 1 microgram of beta-carotene resulted in the formation of 10-20 ng of retinal within 1 h.