Comparative studies of carotenoids in the fauna of the Gullmar Fjord (Bohuslan,Sweden) III. Echinodermata

The author investigated the presence of various carotenoids in the Echinodermata from Gullmar Fjord (Bohuslan, Sweden) by means of columnar and thin-layer chromatography. The investigations revealed the presence of the following:

- inHenricia sanguinolenta:β-carotene, echinenone, canthaxanthin, guraxanthin, lutein-5, 6-epoxide and astaxanthin.

- inAmphiura filiformis: canthaxanthin, cryptoxanthin, lutein, lutein-5, 6-epoxide, isozeaxanthin, zeaxanthin, astaxanthin and 4-hydroxy-4-keto-β-carotene.

- inAmphipholis squamata:β-carotene, cryptoxanthin, lutein, lutein-5, 6-epoxide, astaxanthin, astaxanthin ester, asterin-acid and rubixanthin derivative.

- inOphiopholis aculeata: canthaxanthin, cryptoxanthin, isozeaxanthin, astaxanthin, astaxanthin ester, asterinacid, 4-hydroxy-4-keto-β-carotene, hydroxy rubixanthin and gazaniaxanthin-like substances.

- inOphiothrix fragilis: canthaxanthin, lutein-5, 6-epoxide, isozeaxanthin, astaxanthin, astaxanthin ester, 4-hydroxy-4-keto-β-carotene, and hydroxy rubixanthin.

- inAntedon petatus:canthaxanthin, guaraxanthin, isozeaxan-thin, zeaxanthin, astaxanthin, astaxanthin ester and 4-keto-4-ethoxy-β-carotene.

- inEchinocardium cordatum:β-carotene,γ-carotene, canthaxanthin, lutein, isozeaxanthin, zeaxanthin, astaxanthin and astaxanthin ester.

- inSpatangus purpureus: isozeaxanthin, astaxanthin, astaxanthin ester and 4-hydroxy-4-keto-β-carotene.

     

Investigations of carotenoids in some animals of the Adriatic Sea V. Echinodermata

The author investigated the carotenoids in the Echinodermata from Adriatic sea by means of columnar and thin-layer chromatography. The following carotenoids were identified:

- in Coscinasterias tenuispina: β-carotene, isocryptoxanthin lutein, lutein-5, 6-epoxide, 4-hydroxy-4-keto-β-carotene, zeaxanthin, astaxanthin and asterinacid.

- in Marthasterias glacialis: β-carotene, echinenone, cryptoxanthin, lutein, lutein 5, 6-epoxide, 4-hydroxy-4-keto-β-carotene, zeaxanthin, astaxanthin ester, astaxanthin and 3, 4-didehydro-α-carotene.

- in Paracentrotus lividus: β-carotene, echinenone, cryptothin, isocryptoxanthin, lutein, lutein-5, 6-epoxide, 4-hydroxy-4-keto-β-carotene, zeaxanthin, astaxanthin, astaxanthin ester and asterinacid.

- in Sphaerechinus granularis: ,β-carotene, echinenone, cryptoxanthin, lutein, lutein-5, 6-epoxide, astaxanthin and guaraxanthin.

     

Carotenoids in fish. XXVI. Pungitius pungitius (L.) and Gasterosteus aculeatus L. (Gasterosteidae)

The author investigated the presence of various carotenoids in the different parts of the body of Pungitius pungitius (L.) and Gasterosteus aculeatus L. by means of columnar and thin-layer chromatography. The investigations revealed the presence of the following carotenoids:

in Pungitius pungitius. α-carotene, β-carotene, β-cryptoxanthin, mutatochrome, zeaxanthin and astaxanthin;

in Gasterosteus aculeatus: β-carotene, β-cryptoxanthin, β-carotene epoxide, neothxanthin, canthaxanthin, mutatochrome, lutein, phoenicoxanthin, zeaxanthin, taraxanthin, tunaxanthin, astaxanthin, astaxanthin ester and α-doradexanthin. The total carotenoid content ranged from 2.229 to 138.504 µg/g wet weight.

     

Carotenoids in fish II. Carotenoids and vitamin A in some fishes from the coastal region of the Black Sea

The presence of various carotenoids and vitamin A in seven species of fish from the coastal region of the Black Sea was investigated by means of columnar and thinlayer chromatography. The investigations revealed the presence of the following carotenoids: Mugil auratus: ß-carotene, canthaxanthin, lutein, zeaxanthin, astaxanthin ester and astacene. Diplodus annularis: ß-carotene, canthaxanthin, tunaxanthin, lutein, zeaxanthin and astacene. Diplodus sargus: ß-carotene, tunaxanthin, lutein, taraxanthin, zeaxanthin and astaxanthin. Crenilabrus tinca: tunaxanthin, canthaxanthin, lutein, astaxanthin and astacene. Blennius sphinx: ß-carotene, χ-carotene (?), lutein, tunaxanthin, taraxanthin and astaxanthin. Blennius sanguinolentus: ß-carotene, tunaxanthin and astaxanthin (ester and free). Gobius melanostomus: ß-carotene and astacene. Some fractions were not identified. Vitamin A was found in all species investigated.     

Precence of β-Carotene and Xantophylls in two species of the genus Niphargus (Crustacea: Amphipoda)

The presence of carotenoids in the Niphargus tatrensis Wrzesniowski and Niphargus aquilex schellenbergi Karaban has been investigated. In extracts separated by means of column and thin-layer chromatography, the following carotenoids were identified:

1. in Niphargus tatrensis: β-carotene, astaxanthin ester, rubixanthin, celaxanthin and astaxanthin.
2. in Niphargus aquilex schellenbergi: cantha-xanthin, astaxanthin ester, isozeaxanthin (only from Jeker), 4-keto-4′-hydroxy-β-carotene (only from Terzietebronbos), rubixanthin, celaxanthin and astaxanthin.

It was not possible to identifity the sexth fractions.     

Metabolism of carotenoids in sea-urchin Pseudocentrotus depressus

• 1.1. Feeding experiments with β,β-carotene, canthaxanthin and astaxanthin on the sea urchin Pseudocentrotus depressus were investigated.
• 2.2. In the case of β,β-carotene group, β-carotene was accumulated, β-isocryptoxanthin appeared and β-echinenone increased 6.8 times as much as the control group. On the other hand, in canthaxanthin and astaxanthin groups, canthaxanthin and astaxanthin increased significantly, respectively. The metabolic products of these carotenoids could not be found.
• 3.3. It was concluded that β,β-carotene was bioconverted to β-echinenone via β-isocryptoxanthin in P. depressus and could not be oxidatively metabolized beyond β-echinenone.

     

The carotenoids of eggs of wild and farmed cod (Gadus morhua)

• 1.1. Eggs of wild cod, and of farmed cod fed (a) a diet supplemented with astaxanthin and (b) a diet supplemented with both astaxanthin and canthaxanthin, were analysed with respect to carotenoids.
• 2.2. The total carotenoid contents in eggs were 0.7 ppm for wild cod and 0.5 ppm for farmed cod.
• 3.3. Cod, having white flesh, deposit ketocarotenoids in the eggs, preferably astaxanthin.
• 4.4. Canthaxanthin can replace astaxanthin in the eggs, but astaxanthin appears to be deposited preferentially when both carotenoids are present in the diet.
• 5.5. The isomer distribution of (3S, 3′S):(3R, 3′S, meso):(3R, 3′R) astaxanthin in the eggs reflected the isomer composition of the diet.
• 6.6. Echinenone, 4′-hydroxyechinenone, adonixanthin and zeaxanthin encountered in cod eggs may represent reductive metabolites of canthaxanthin and astaxanthin.

     

Characteristics of carotenoid features in muscle and ovary from anadromous and river resident types of Alaskan dolly varden charr (Salvelinus malma malma)

• 1.1. The carotenoids in the muscles and eggs from two types of natural Alaskan dolly varden charr, anadromous and river resident types, were examined.
• 2.2. In the muscle from the anadromous charr, astaxanthin was the major component comprising more than 70% of the total, followed by idoxanthin and 4-keto-zeaxanthin.
• 3.3. The egg carotenoid features in the river resident charr were more complicated compared with those in the anadromous fish. A considerable proportion of unidentified carotenoids was found in the eggs from the river resident charr.
• 4.4. Idoxanthin was the main component along with considerable propotions of β-carotene tetrol and astaxanthin in the eggs from the anadromous charr, whereas zeaxanthin and lutein were detected besides idoxanthin and β-carotene tetrol in the eggs from the river resident fish.
• 5.5. The prey was considered to be responsible for the difference in the carotenoid features of the eggs from the anadromous and river resident charr.

     

Bioconversion pathway of astaxanthin into retinol2 in mature rainbow trout (Salmo Gairdneri Rich.)

• 1.1. Carotenoid and retinoid forms were analysed by HPLC in various tissues of mature rainbow trout fed for 11 months (7–9°C) with astaxanthin (50 and 100 mg/kg diet) or canthaxanthin (100 mg/kg diet).
• 2.2. Decreasing concentrations of canthaxanthin, echinenone and β-carotene, but no retinol₁, were found in the liver and skin of canthaxanthin-fed fish.
• 3.3. Higher retinol₂ concentrations were found in ovaries and testes of astaxanthin-fed fish compared to canthaxanthin and control groups.
• 4.4. A new metabolic pathway for direct conversion of astaxanthin into retinol₂ in gonads is proposed.

     

Carotenoids in fish. XXIII. Syngnathidae family

The author investigated the presence of various carotenoids in three species of the Syngnathidae family by means of columnar and thin-layer chromatography. The investigations revealed the presence of the following carotenoids: canthaxanthin, lutein, lutein epoxide, zeaxanthin, astaxanthin (free and ester form) and 4-hydroxy-4-keto--carotene. Ketocarotenoids (astaxanthin and canthaxanthin) comprised the greatest part     

Carotenoids and carotenoid metabolism in Carcinus maenas (Crustacea: Decapoda)

The carotenoid pigments of the hepatopancreas, ovaries and epidermis of Carcinus maenas were investigated. The following pigments were identified: β-carotene, δ-carotene, echinenone, isocryptoxanthin, canthaxanthin, lutein, zeaxanthin, flavoxanthin and astacene.
The relative abundance of these pigments in the three tissues and the presence of possible hydroxy and keto intermediates suggest the metabolism of astaxanthin from β-carotene. The metabolic pathway in Carcinus is discussed in relation to recent studies on other invertebrates.     

Carotenoids and carotenoid metabolism in Carcinus maenas (Crustacea: Decapoda)

The carotenoid pigments of the hepatopancreas, ovaries and epidermis of Carcinus maenas were investigated. The following pigments were identified: β-carotene, δ-carotene, echinenone, isocryptoxanthin, canthaxanthin, lutein, zeaxanthin, flavoxanthin and astacene.
The relative abundance of these pigments in the three tissues and the presence of possible hydroxy and keto intermediates suggest the metabolism of astaxanthin from β-carotene. The metabolic pathway in Carcinus is discussed in relation to recent studies on other invertebrates.     

Comparative biochemical studies of carotenoids in sea-urchins—II. The more primitive sea-urchins belonging to the orders Cidaroida,Echinothurioida, Diadematoida and Arbacioida

• 1.1. The carotenoids of seven species of more primitive sea-urchins, [orders Cidaroida (I), Echinothurioida (II), Diadematoida (III), and Arbacioida (IV)] were investigated from the comparative biochemical point of view.
• 2.2. β,β-carotene and β-echinenone have been isolated as major carotenoids in (I) and (III, IV), respectively. In (II), β,β-carotene, β-echinenone, canthaxanthin and (3S,3′S)-astaxanthin were foundto be predominant carotenoids.
• 3.3. The carotenoid patterns of (I) which is the most primitive sea-urchin from the phylogenetic point of view, and of (II) which is direct developers with non-feeding larvae, were quite different from those of the other sea-urchins showing typical development with feeding larvae.

     

Reconstruction of the astaxanthin biosynthesis pathway in rice endosperm reveals a metabolic bottleneck at the level of endogenous β-carotene hydroxylase activity

Astaxanthin is a high-value ketocarotenoid rarely found in plants. It is derived from β-carotene by the 3-hydroxylation and 4-ketolation of both ionone end groups, in reactions catalyzed by β-carotene hydroxylase and β-carotene ketolase, respectively. We investigated the feasibility of introducing an extended carotenoid biosynthesis pathway into rice endosperm to achieve the production of astaxanthin. This allowed us to identify potential metabolic bottlenecks that have thus far prevented the accumulation of this valuable compound in storage tissues such as cereal grains. Rice endosperm does not usually accumulate carotenoids because phytoene synthase, the enzyme responsible for the first committed step in the pathway, is not present in this tissue. We therefore expressed maize phytoene synthase 1 (ZmPSY1), Pantoea ananatis phytoene desaturase (PaCRTI) and a synthetic Chlamydomonas reinhardtii β-carotene ketolase (sCrBKT) in transgenic rice plants under the control of endosperm-specific promoters. The resulting grains predominantly accumulated the diketocarotenoids canthaxanthin, adonirubin and astaxanthin as well as low levels of monoketocarotenoids. The predominance of canthaxanthin and adonirubin indicated the presence of a hydroxylation bottleneck in the ketocarotenoid pathway. This final rate-limiting step must therefore be overcome to maximize the accumulation of astaxanthin, the end product of the pathway.     

The occurrence of the Entner-Doudoroff pathway in bacteria

The occurrence of the two key enzymes of the Entner-Doudoroff pathway, gluconate-6-phosphate dehydrase and 2-keto-3-deoxygluconate-6-phosphate aldolase, was determined in approximately 150 strains belonging to 37 different bacterial genera. The following results were obtained:

1. 24 out of 37 genera have at least one representative with the Entner-Doudoroff mechanism. It is thus more widespread than previously thought.
2. The Entner-Doudoroff mechanism occurs mainly in gram-negative bacteria with a DNA base composition in the range 52–70% GC. Eighty-five per cent of these organisms contain the system, while only 20% (6 strains) of the gram-negative organisms with less than 52% GC possess both enzymes.
3. This pathway is absent in all gram-positive organisms investigated except in 5 out of 12Nocardia strains.
4. Erwinia and some strains of theAchromobacter-Alcaligenes group are exceptional, since they possess only 2-keto-3-deoxygluconate-6-phosphate aldolase.

     

Investigation of seasonal variations in cow serum retinol and β -carotene by high performance liquid chromatographic method

• 1.1. High performance liquid chromatography (HPLC) was used to determine β-carotene and retinol in cow serum.
• 2.2. Two groups of state and private farm cows (Groups 1 and 2) were used to assess seasonal variation when different food sources were fed to cows on serum β -carotene and retinol concentrations.
• 3.3. Mean serum concentrations of β-carotene and retinol from October to April in both Groups 1 and 2 cows were lower (P < 0.05) than in the other months when the cows were fed various combination of maize silage, alfalfa and carrot residues and grass hay, respectively.
• 4.4. Mean serum β-carotene and retinol concentrations in June and July were higher (P < 0.05) than in other months when the cows were in pasture.
• 5.5. Mean serum β-carotene and retinol concentrations in May, August and September were lower (P < 0.05) than in June and July and higher (P < 0.05) than in other months when a lesser amount of green pasture was available to the cows.
• 6.6. There was a seasonal variation (P < 0.05) in serum β -carotene and retinol concentrations. When the carotene intake is very high, conversion of β -carotene to retinol decreases. Mean monthly serum β -carotene and retinol concentrations showed that combination of alfalfa hay and maize silage, and grass hay and carrot residues can maintain adequate serum β-carotene and retinol concentrations during the dry season.

     

Carotenoids in fish. XXIV. Sardina pilchardus Walb. (Clupeidae)

The author investigated the presence of various carotenoids in Sardina pilchardus Walb. from the coast of Southern Europe.The presence of the following carotenoids has been stated: -carotene, -carotene epoxide, -cryptoxanthin, canthaxanthin, lutein epoxide, zeaxanthin, astaxanthin (free and ester form) and mutatochrome. The dominant carotenoid in all the parts of the body was astaxanthin, especially its ester form. The total content of carotenoid ranged from 10.537 (skin and muscles) to 116.309 µg/g fresh weight (liver).     

Engineering of the carotenoid pathway in <Emphasis Type="Italic">Xanthophyllomyces dendrorhous</Emphasis> leading to the synthesis of zeaxanthin

Zeaxanthin is an essential nutrient for prevention of macular degeneration. However, it is limited in our diet. For the production of zeaxanthin, we have engineered zeaxanthin synthesis into a carotenoid mutant of Xanthophyllomyces dendrorhous which is blocked in astaxanthin synthesis and accumulates β-carotene instead. Two strategies were followed to reach high-yield zeaxanthin synthesis. Total carotenoid synthesis was increased by over-expression of genes HMGR, crtE, and crtYB encoding for limiting enzymes in the pathway leading to and into carotenoid biosynthesis. Then bacterial genes crtZ were used to extend the pathway from β-carotene to zeaxanthin in this mutant. The increase of total carotenoids and the formation of zeaxanthin is dependent on the number of gene copies of crtYB and crtZ integrated into the X. dendrorhous upon transformation. The highest zeaxanthin content around 500 μg/g dw was reached by shaking flask cultures after codon optimization of crtZ for Xanthophyllomyces. Stabilization of carotenoid and zeaxanthin formation in the final transformant in the absence of selection agents was achieved after passing through a sexual cycle and germination of basidiospores. The values for the transformant before and after stabilization were very similar resembling about 70 % of total carotenoids and corresponding to a conversion rate of 80 % for hydroxylation of β-carotene to zeaxanthin. The stabilized transformant allowed experimental small-scale fermentation yielding X. dendrorhous cells with a zeaxanthin content similar to the shaking flask cultures. Our result demonstrates the potential of X. dendrorhous for its development as a zeaxanthin producer and its suitability for large-scale fermentation.     

Some carotenoids in Chironomus annularius Meig. larvae (Diptera : Chironomidae)

Summary The author investigated the presence of various carotenoids in the body of the larvae of the Chironomus annularius MEIG. moquito (Diptera: Chironomidae) by means of columnar and thin-layer chromatography.The investigations revealed the presence of the following carotenoids in the larvae: -carotene, -carotene, canthaxanthin, cryptoxanthin, lutein, astacene and a kind of xanthophyl which was not possible to identify more closely.     

Absorption and blood clearance of labelled carotenoids ([14C]astaxanthin, [3H]Canthaxanthin and [3H]zeaxanthin) in mature female rainbow trout (Oncorhynchus mykiss)

• 1.1. Using one force-fed meal, eight mature female rainbow trout received [¹⁴C]astaxanthin ([¹⁴C]Ax) with [³H]canthaxanthin ([³H]Cx; N = 3) or with [³H]zeaxanthin ([³H]Zx; N = 5).
• 2.2. Approximately 200 μl of blood were collected via caudal puncture every 24 hr for 4 days. After 96 hr, the fish were killed and pyloric caeca (P.C.) from the duodenal intestine (D.I.) section, ileal intestine (I.I.), and posterior intestine (P.I.) were dissected out.
• 3.3. In the blood, Ax levels were higher than Cx followed by Zx levels.
• 4.4. This corresponds to their respective absorption by the trout as was confirmed by their relative concentrations in P.C., I.I. and P.I.
• 5.5. However, blood clearance was similar for all three compounds. [¹⁴C]Phoenicoxanthin ([¹⁴C]Px) was detected as a reduced metabolite of [¹⁴C]Ax in all gut sections.