Non-esterified Astaxanthin Research Articles

Docosahexaenoic acid-acylated astaxanthin ester exhibits superior performance over non-esterified astaxanthin in preventing behavioral deficits coupled with apoptosis in MPTP-induced mice with Parkinson's disease.

Non-esterified astaxanthin (AST) has been reported to exhibit protective effects from Parkinson's disease (PD). Notably, DHA-acylated astaxanthin ester (DHA-AST) is widely distributed in the seafood. However, whether DHA-AST has an effect on PD, and the differences between DHA-AST, non-esterified AST and the combination of non-esterified AST (AST) with DHA (DHA + AST) is unclear. In the present study, mice with PD, induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), were employed to investigate the effects of DHA-AST, AST and DHA + AST on Parkinson's disease. The rotarod test results showed that DHA-AST significantly suppressed the PD development in MPTP-induced mice, and was better than the effects of AST and DHA + AST. Further mechanistic studies indicated that all three astaxanthin supplements could inhibit oxidative stress in the brain. It was noted that DHA-AST had the best ability to suppress the apoptosis of dopaminergic neurons via the mitochondria-mediated pathway and JNK and P38 MAPK pathway in the brain among the three treated groups. DHA-AST was superior to AST in preventing behavioral deficits coupled with apoptosis rather than oxidative stress, and might provide a valuable reference for the prevention and treatment of neurodegenerative diseases.

Engineered maize as a source of astaxanthin: processing and application as fish feed.

Astaxanthin from a transgenic maize line was evaluated as feed supplement source conferring effective pigmentation of rainbow trout flesh. An extraction procedure using ethanol together with the addition of vegetal oil was established. This resulted in an oily astaxanthin preparation which was not sufficiently concentrated for direct application to the feed. Therefore, a concentration process involving multiple phase partitioning steps was implemented to remove 90% of the oil. The resulting astaxanthin raw material contained non-esterified astaxanthin with 12% 4-keto zeaxanthin and 2% zeaxanthin as additional carotenoids. Isomeric analysis confirmed the exclusive presence of the 3S, 3'S astaxanthin enantiomer. The geometrical isomers were 89% all-E, 8% 13-Z and 3% 9-Z. The incorporation of the oily astaxanthin preparation into trout feed was performed to deliver 7mg/kg astaxanthin in the final feed formulation for the first 3.5weeks and 72mg/kg for the final 3.5weeks of the feeding trial. The resulting pigmentation of the trout fillets was determined by hue values with a colour meter and further confirmed by astaxanthin quantification. Pigmentation properties of the maize-produced natural astaxanthin incorporated to 3.5µg/gdw in the trout fillet resembles that of chemically synthesized astaxanthin. By comparing the relative carotenoid compositions in feed, flesh and feces, a preferential uptake of zeaxanthin and 4-keto zeaxanthin over astaxanthin was observed.

The major egg reserve protein from the invasive apple snail Pomacea maculata is a complex carotenoprotein related to those of Pomacea canaliculata and Pomacea scalaris

Snails from the genus Pomacea lay conspicuous masses of brightly colored eggs above the water. Coloration is given by carotenoproteins that also which play important roles in protection against sun radiation, stabilizing and transporting antioxidant molecules and helping to protect embryos from desiccation and predators. They seem a key acquisition, but have been little studied. Here we report the characteristics of the major carotenoprotein from Pomacea maculata and the first comparison among these egg proteins. This particle, hereafter PmPV1, represents ~52% of perivitellin fluid protein. It is a glyco-lipo-carotenoprotein responsible for the bright reddish egg coloration. With VHDL characteristics, PmPV1 apparent molecular mass is 294kDa, composed of five non-covalently bound subunits of pI 4.7-9.8 and masses between 26 and 36kDa whose N-terminal sequences were obtained. It is a glyco-lipo-carotenoprotein scarcely lipidated (<1%) but highly glycosilated (13% by wt). Lipids include phospholipids, free fatty acids and carotenoids; mannose and galactose predominate over other monosaccharides. Main carotenoids are esterified and non-esterified astaxanthin (71 and 25%, respectively). Carotenoid removal does not seem to affect the structural characteristics of the oligomer, while deglycosilation reduces subunit number from five to a single one. The carotenoid-protein association protected the former against oxidation. PmPV1 cross reacts with polyclonal antibodies against the PcOvo, the major carotenoprotein from Pomacea canaliculata. The characterization of PmPV1 allows the first comparisons among snail carotenoproteins and further highlights the importance of these perivitellins in the reproductive strategy of Pomacea.

Effect of an oral astaxanthin prodrug (CDX-085) on lipoprotein levels and progression of atherosclerosis in LDLR−/− and ApoE−/− mice

Oxidative stress and inflammation are key promoters of atherosclerosis and myocardial damage. When orally administered, the novel astaxanthin prodrug CDX-085 delivers high levels of the xanthophyll antioxidant astaxanthin that protects LDL from oxidation and reduces primary thrombosis. In this study, we analyzed whether delivery of astaxanthin from administration of the CDX-085 prodrug reduces plasma lipoprotein levels and the progression of atherosclerosis in low-density lipoprotein receptor negative (LDLR−/−) and apolipoprotein E deficient (ApoE−/−) mice. MethodsRelative circulating levels of astaxanthin derived from CDX-085 administration compared to administration of pure astaxanthin was initially evaluated in a canine model. In mouse Study #1, 16 wild-type and 16 LDLR−/− mice on 0.5% cholesterol diet supplemented with either 0.0%, 0.08%, 0.2% and 0.4% CDX-085 were used to assess plasma levels and lipoprotein biodistribution measured by FPLC after 4 weeks treatment. In Study #2, 36 male LDLR−/− mice were randomized to a 0.5% cholesterol chow diet (CHOW group, n=12) or 0.5% cholesterol chow fortified with 0.08% CDX-085 (n=12) or 0.5% cholesterol chow with 0.4% CDX-085 (n=12) for 12 weeks. In Study #3, 34 male ApoE−/− mice were randomized in the same fashion as the Study #2 and fed similar diets for 9 weeks. ResultsCDX-085 administration was shown to result in significantly higher levels of circulating astaxanthin (p<0.001 ANOVA) over a 72h period compared to pure, non-esterified astaxanthin in a single-dose pharmacokinetic study in beagles. In Study #1, plasma astaxanthin levels were 5–9-fold higher in LDLR−/− mice compared to wild-type mice. Astaxanthin was highly distributed among all lipoprotein fractions, generally reflecting cholesterol content of lipoproteins. In Study #2, administration of CDX-085 resulted in significantly lower total cholesterol levels (528±68mg/dL vs. 550±67mg/dL vs. 602±80mg/dL, p=0.047) and aortic arch atherosclerosis (9.0±4.2% vs. 9.8±3.5% vs. 13.2±3.6%, p=0.023) in the 0.4% CDX-085 group compared to the 0.08% CDX-085 and CHOW groups, respectively. In ApoE−/− mice, a 72% reduction in triglycerides in the 0.4% CDX-085 group and 50% reduction in the 0.08% CDX-085 groups was noted compared to CHOW group (final levels 17±11mg/dL vs. 30±15mg/dL vs. 60±32mg/dL, respectively, p=0.001). ConclusionOral administration of the novel astaxanthin prodrug CDX-085 shows that it distributes among lipoproteins. CDX-085 lowers total cholesterol and aortic arch atherosclerosis in LDLR−/− mice and triglyceride levels in ApoE−/− mice and shows promise for further evaluation in human studies.

Effect of background colour on the distribution of astaxanthin in black tiger prawn ( Penaeus monodon): Effective method for improvement of cooked colour

The colouration of prawns is dependent upon the presence of carotenoid pigments (predominantly astaxanthin) and has a significant impact on their market value. In this study we observed that visual appearance of colour in black tiger prawns ( Penaeus monodon), when assessed using a subjective commercial grading scale, did not always correlate well with total carotenoid content. We also observed that visual appearance was affected by the background colour of the tanks in which the prawns were growing. When prawns were grown in black or white tanks for 28 days, even though they had similar mean total astaxanthin contents (29–33 μg/g prawn tail), those grown in black tanks were much more orange/red when cooked, compared with those from the white tanks. The pigments were mainly located in the cephalothorax, abdominal epidermal layer and abdominal exoskeleton. Light microscopy showed an even distribution of pigment in the epidermal layer of more coloured prawns compared with concentrated pigment areas in the lighter-coloured prawns. Non-esterified astaxanthin was the major carotenoid present in all body locations in prawns from black tanks (50%) with the remainder being made up equally of astaxanthin mono- and di-esters. However, for prawns from white tanks, non-esterified astaxanthin accounted for just 12–13%, with the mono-ester reaching about 60% of the total present. In a separate experiment, we demonstrated that changes in background colour caused a rapid change in visual colour. When prawns were moved from a white to a black tank, there was a significant increase in grade score within 1 h without any change in astaxanthin content. Colour improvement continued to increase over 168 h. Moving prawns from a black to a white tank resulted in a reduction in grade score but the change was much less rapid. This work suggests that, provided prawns have an adequate content of astaxanthin in their diets, it is possible to improve their overall appearance (raw and cooked) by manipulating background colour about the time of harvesting.

Disodium Disuccinate Astaxanthin (CardaxTM): Antioxidant and Antiinflammatory Cardioprotection

Disodium disuccinate astaxanthin (Cardax), DDA) has cardioprotective effects in the rat, rabbit, and canine models of experimental infarction. It is highly effective by parenteral administration in subchronic and acute dosing regimens. Unpublished data in rats suggest that oral cardioprotection is also readily achievable. DDA-induced myocardial salvage in the canine can reach 100% with a 4-day subchronic dosing regimen. At a single i.v. dose DDA is cardioprotective, when given 2 h before experimental coronary occlusion, but the protection is on the average two-thirds of that achieved with the subchronic regimen in dogs. In conscious animals DDA has no effects on hemodynamic parameters. The primary mechanism of cardioprotection appears to be antioxidant activity involving direct scavenging of superoxide anion, the lynchpin radical in ischemia-reperfusion injury. In addition, modulation of serum complement activity, as well as the reduction in the levels of C-reactive protein (CRP) and the membrane attack complex (MAC) in infarcted tissue suggest a significant antiinflammatory component in the mechanism of cardioprotective action of DDA. Stoichiometric binding of the meso-form of the compound to human serum albumin (HSA) has been demonstrated in vitro. This binding capacity overcomes the supramolecular assembly of the compound in aqueous solution, which by itself improves the stability and shelf life of aqueous formulations. Non-esterified astaxanthin readily enters cardiac tissue after either oral or parenteral administration, providing a reservoir of a cardioprotective agent with a significant half-life due to favorable ADME in mammals. Due to the well-documented safety profile of non-esterified astaxanthin in humans, disodium disuccinate astaxanthin may well find clinical utility in cardiovascular indications in humans following successful completion of preclinical and clinical pharmacology and toxicology studies.

Acute and chronic administration of disodium disuccinate astaxanthin (CardaxTM) produces marked cardioprotection in dog hearts

Previous results from our laboratory have shown that a novel carotenoid derivative (disodium disuccinate astaxanthin; Cardax) produced dose-related reductions in myocardial infarct size (IS) in Sprague-Dawley rats when it was administered at any of three doses (25, 50 and 75 mg/kg, iv) on four consecutive days, followed by the acute infarct size study on day 5. Maximum salvage occurred at the highest dose (75 mg/kg) tested, and was shown as a 56% reduction in IS. In the present follow-up study, we used a more relevant large animal model, the dog, and looked at the effect of administering Cardax iv either acutely 2 h prior to occlusion (N = 8) or for 4 days at 50 mg/kg iv as previously done in the rat model (N = 6). The results were compared to a saline vehicle-treated group (N = 10). In all groups, dogs were subjected to 60 min of left anterior descending (LAD) coronary artery occlusion and 3 h of reperfusion. IS was determined using a triphenyltetrazolium chloride (TTZ) histochemical stain and was expressed as a percent of the area at risk (IS/AAR). IS/AAR was 20.9 +/- 1.6 % (mean +/- S.E.M.) in controls and was reduced to 11.0 +/- 1.7% (47.3% salvage; p < 0.01) in dogs treated only once iv at 2 h prior to occlusion, and 6.6 +/- 2.8% (68.4% salvage; p < 0.001) in dogs treated for 4 days. In the chronic treatment group, two of the three dogs with plasma concentrations of non-esterified astaxanthin above 1 microM had 0% IS/AAR (100% cardioprotection). These results suggest that Cardax has marked cardioprotective properties in both rodents and canines. Thus, Cardax may be a novel and powerful new means to prevent myocardial injury and/or necrosis associated with elective and/or urgent cardiac surgical interventions such as coronary angioplasty and stenting, as well as coronary artery bypass surgery (CABG).

Direct superoxide anion scavenging by a disodium disuccinate astaxanthin derivative: relative efficacy of individual stereoisomers versus the statistical mixture of stereoisomers by electron paramagnetic resonance imaging

Carotenoids are a related group of greater than 600 natural compounds, irrespective of geometric- and stereoisomers, with demonstrated antioxidant efficacy. The carotenoids are broadly divided into “carotenes,” or non-oxygen substituted hydrocarbon carotenoids, and “xanthophylls,” oxygen-substituted carotenoids. The natural compounds are excellent singlet oxygen quenchers as well as lipid peroxidation chain-breakers; this dual antioxidant capacity is generally attributed to the activity of the polyene chain, and increases with the number of conjugated double bonds along the polyene chain length. However, the poor aqueous solubility of most carotenes and the vast majority of xanthophylls limits their use as aqueous-phase singlet oxygen quenchers and direct radical scavengers. A variety of introduction vehicles (e.g., organic solvents, cyclodextrins) have been used to introduce the insoluble carotenoids into aqueous test systems. Hawaii Biotech, Inc. (HBI) successfully synthesized a novel carotenoid derivative, the disodium disuccinate derivative of astaxanthin (3,3 ′-dihydroxy-β,β-carotene-4,4 ′-dione) in all- trans (all- E) form. The novel derivative is a water-dispersible symmetric chiral molecule with two chiral centers, yielding four stereoisomeric forms: 3 R,3 ′ R and 3 S,3 ′ S (enantiomers), and the diastereomeric meso forms (3 R,3 ′ S and 3 ′ R,3 S). The individual stereoisomers were synthesized at high purity (>90% by HPLC) and compared directly for efficacy with the statistical mixture of stereoisomers obtained from the synthesis from the commercial source of astaxanthin (1:2:1 ratio of 3 S,3 ′ S, meso, and 3 R,3 ′ R, respectively). Direct scavenging of superoxide anion was evaluated in a standard in vitro isolated human neutrophil assay by electron paramagnetic resonance (EPR) imaging, employing the spin-trap DEPMPO. Each novel derivative was tested in pure aqueous formulation and in ethanolic formulation shown to completely disaggregate the compounds in solution. In each case, the ethanolic formulation was a more potent scavenging vehicle. No significant differences in scavenging efficiency were noted among the individual stereoisomers and the statistical mixture of stereoisomers, suggesting that the polyene chain alone was responsible for superoxide scavenging. Dose-ranging revealed that the statistical mixture of stereoisomers of the novel derivative, at millimolar (mM) concentrations, could nearly completely eliminate the superoxide anion signal generated in the activated human neutrophil assay. All ethanolic formulations of the novel derivatives exhibited increased scavenging efficiency over equimolar concentrations of non-esterified astaxanthin delivered in a dimethyl sulfoxide (DMSO) vehicle. These novel compounds will likely find utility in applications requiring aqueous delivery of a highly potent direct radical scavenger.

Carotenoids and their derivatives in organs of the maturing female crayfish Cherax quadricarinatus

Carotenoids were separated by high-performance liquid chromatography from organ homogenates of pre-vitellogenic (oocyte diameter ∼300μm) and late-vitellogenic (oocyte diameter >1mm) Cherax quadricarinatus females. Carotenoids were present predominantly in the cuticle and in the late-vitellogenic ovary. Smaller amounts were present in the hepatopancreas, and almost no carotenoids were found in the muscular tissue. The cuticle contained mostly esterified astaxanthins. The ovaries contained mostly non-esterified astaxanthin. Pre-vitellogenic ovaries were also relatively rich in lutein, whereas late-vitellogenic ovaries were relatively rich in β-carotene. The hepatopancreas contained mostly β-carotene. Possible roles for the different carotenoids in the cuticle and ovary of the crayfish are discussed.

Metabolism of dietary carotenoids in eggs of red sea bream

Abstract 1. 1. Transfer of dietary carotenoids into eggs was examined by a feeding experiment of red sea bream broodstock. 2. 2. Levels of carotenoids incorporated in the eggs were approximately 60 μg/100 g irrespective of amounts and kinds of carotenoids supplemented to the diets. 3. 3. Only keto and non-esterified carotenoids were detected in the eggs. 4. 4. Canthaxanthin was transferred into eggs but not β-carotene. Astaxanthin esters were converted largely to non-esterified astaxanthin and partly to doradexanthin. It was presumed that idoxanthin present in the eggs was derived from non-esterified astaxanthin in the diets.