Wataru Miki

Comparison of carotenoids in the ovaries of marine fish and shellfish

1. The following carotenoids were isolated and identified: astaxanthin diester, tunaxanthin monoester, astaxanthin monoester, tunaxanthin, astaxanthin, doradexanthin, lutein, zeaxanthin, idoxanthin, triol and tetrol from nine species of fish; astaxanthin diester, astaxanthin monester, astaxanthin, doradexanthin, zeaxanthin, idoxanthin and tetrol from four species of crustacean, astaxanthin, pectenolone, pectenoxanthin, pectenol and tetrol from four species of scallop. 2. Tunaxanthin monoester and astaxanthin diester were major carotenoids in skipjack and Pacific cod, respectively. 3. The concentration of carotenoids ranged 0.065-1.95, 1.30-5.91 and 1.56-7.15 mg per 100 g ovary for fish, crustacean and scallop, respectively. 4. The species- and tissue-specificity of ovarian carotenoids and their possible role are discussed.

Elucidation of a Carotenoid Biosynthesis Gene Cluster Encoding a Novel Enzyme, 2,2′-β-Hydroxylase, from Brevundimonas sp. Strain SD212 and Combinatorial Biosynthesis of New or Rare Xanthophylls

ABSTRACT A carotenoid biosynthesis gene cluster mediating the production of 2-hydroxyastaxanthin was isolated from the marine bacterium Brevundimonas sp. strain SD212 by using a common crtI sequence as the probe DNA. A sequence analysis revealed this cluster to contain 12 open reading frames (ORFs), including the 7 known genes, crtW, crtY, crtI, crtB, crtE, idi, and crtZ. The individual ORFs were functionally analyzed by complementation studies using Escherichia coli that accumulated various carotenoid precursors due to the presence of other bacterial crt genes. In addition to functionally identifying the known crt genes, we found that one (ORF11, named crtG) coded for a novel enzyme, carotenoid 2,2′-β-hydroxylase, which showed intriguingly partial homology with animal sterol-C5-desaturase. When this crtG gene was introduced into E. coli accumulating zeaxanthin and canthaxanthin, the resulting transformants produced their 2-hydroxylated and 2,2′-dihydroxylated products which were structurally novel or rare xanthophylls, as determined by their nuclear magnetic resonance and high-performance liquid chromatography/photodiode array detector/atmospheric pressure chemical ionization mass spectrometry spectral data. The new carotenoid produced was suggested to have a strong inhibitory effect on lipid peroxidation.

Characterization of β-Carotene Ketolases, CrtW, from Marine Bacteria by Complementation Analysis in Escherichia coli

A complementation analysis was performed in Escherichiacoli to evaluate the efficiency of β-carotene ketolases (CrtW) from the marine bacteria Brevundimonas sp. SD212, Paracoccus sp. PC1 (Alcaligenes PC-1), and Paracoccus sp. N81106 (Agrobacteriumaurantiacum), for astaxanthin production. Each crtW gene was expressed in Escherichiacoli synthesizing zeaxanthin due to the presence of plasmid pACCAR25ΔcrtX. Carotenoids that accumulated in the resulting E.coli transformants were examined by chromatographic and spectroscopic analyses. The transformant carrying the Paracoccus sp. PC1 or N81106 crtW gene accumulated high levels of adonixanthin, which is the final astaxanthin precursor for CrtW, and astaxanthin, while the E.coli transformant with crtW from Brevundimonas sp. SD212 did not accumulate any adonixanthin and produced a high level of astaxanthin. These results show efficient conversion by CrtW of Brevundimonas sp. SD212 from adonixanthin to astaxanthin, which is a new-found characteristic of a bacterial CrtW enzyme. The phylogenetic positions between CrtW of the two genera, Brevundimonas and Paracoccus, are distant, although they fall into α-Proteobacteria.

Isolation of Marine Bacteria by In Situ Culture on Media-Supplemented Polyurethane Foam

Polyurethane foam (PUF) supplemented with various agar media was used in situ to trap marine bacteria and it consequently provided a substrate on which they could be cultivated while exposed to natural seawater in the coral reef area. The bacterial population on the PUF blocks was analyzed by denaturing gradient gel electrophoresis (DGGE) of polymerase chain reaction (PCR)-amplified 16S rDNA fragments. Changing the composition of the cultivation medium in the PUF blocks and selecting different sampling sites resulted in different bacteria being detected on the PUF blocks. For example, iron-utilizing (IU) bacteria, siderophore-producing (SP) bacteria, and petroleum-degrading (PD) bacteria were isolated from PUF blocks and it was discovered that IU and SP contained iron and PD contained hydrocarbon. This method opens up the possibility for isolating novel and useful marine bacteria.

Origin of tunaxanthin in the integument of yellowtail (Seriola quinqueradiata)

Abstract 1. 1. Bioconversion of astaxanthin and lutein was examined by feeding experiments of yellowtail. 2. 2. Carotenoid contents in the integuments of the fish fed the diets supplemented with astaxanthin or lutein were markedly increased. 3. 3. In every test fish, the same nine carotenoids, tunaxanthins C, B and A, 3′-epilutein, lutein, zeaxanthin, diatoxanthin, cynthiaxanthin and β-carotene triol were detected. 4. 4. Astaxanthin was presumably metabolized to tunaxanthins C or B, via β-carotene triol, zeaxanthin and 3′-epilutein. Lutein seemed to be metabolized to tunaxanthins C or B, via 3′-epilutein. 5. 5. Sources of tunaxanthin in the integuments of marine fish were estimated to be astaxanthin, lutein and probably zeaxanthin all of which are very common carotenoids in marine organisms.

Pigmentation of juvenile coho salmon with carotenoid oil extracted from Antarctic krill

Abstract Pigmentation of the flesh of juvenile coho salmon ( Oncorhynchus kisutch ) in fresh water was investigated by feeding diets supplemented with an oil extracted from Antarctic krill ( Euphausia superba ). The oil contained ( 3R,3′R )-astaxanthin diester as a main carotenoid. Small fish (80 g) fed the supplemented diet showed very little flesh pigmentation. However, marked pigmentation was observed when fish with an initial weight of approximately 180 g were reared on diets containing 7.2 mg astaxanthin 100 g diet for 4 weeks. The carotenoid content of flesh was about 0.2 mg 100 g and it consisted almost exclusively of astaxanthin. Fish fed a diet containing krill oil for 8 weeks retained most of the carotenoids laid down in the flesh throughout a following 24-week period on a non-carotenoid diet. Structural isomer composition of astaxanthin in the flesh of fish fed a diet containing 14.4 mg astaxanthin 100 g was 93, 4–5 and 2–3% for all- trans -, 9- cis - and 13- cis -astaxanthin at the end of the experiment, respectively. Optical isomer composition of the all- trans -astaxanthin fraction was 74, 10 and 16% for ( 3R,3′R )-, ( 3S,3′R ) meso - and ( 3S,3′S )-astaxanthin, respectively. No isomerization of astaxanthin deposited in the flesh was found.

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.

Decomposition Reaction of Sesamin in Supercritical Water

The methylenedioxyphenyl moiety in the structure of sesamin and episesamin was changed into the catechol moieties, (1R,2S,5R,6S)-6-(3,4-dihydroxyphenyl)-2-(3,4-methylenedioxyphenyl)-3,7-dioxabicyclo[3,3,0]octane, (1R,2R,5R,6S)-2-(3,4-dihydroxyphenyl)-6-(3,4-methylenedioxyphenyl)-3,7-dioxabicyclo[3,3,0]octane, (1R,2R,5R,6S)-6-(3,4-dihydroxyphenyl)-2-(3,4-methylenedioxyphenyl)-3,7-dioxabicyclo[3,3,0]octane, (1R,2S,5R,6S)-2,6-bis(3,4-dihydroxyphenyl)-3,7-dioxabicyclo[3,3,0]octane, and (1R,2R,5R,6S)-2,6-bis(3,4-dihydroxyphenyl)-3,7-dioxabicyclo[3,3,0]octane, in supercritical water. These products had same structures as the sesamin metabolites which act as antioxidants in the liver. These features suggested the direct preparation of antioxidants from sesamin by a one-step reaction using supercritical water.