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Naoki Yamauchi and Alley E. Watada

Degradation of chlorophyll in spinach (Spinacia olearacea L. cv. Hybrid 612) appeared to be regulated through the peroxidase-hydrogen peroxide pathway, which opens the porphyrin ring, thus resulting in a colorless compound. This conclusion was arrived at from the analysis of chlorophylls (Chls) and their metabolizes by HPLC and of enzyme activities catalyzing the degradative reactions. Chls decreased at 25C but not at 1C. The chlorophyll oxidase pathway was not active, as noted by the lack of accumulation of a reaction product named Chl a-1. Lipid peroxidation increased with storage, but the products of the reaction. did not degrade chlorophyll, as noted by the lack of increase in Chl a-1. Chlorophyllase activity increased, but chlorophyllide, the expected product of the reaction, changed minimally during senescence. Ethylene at 10 ppm did not alter the pathway that degraded chlorophyll in spinach.

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Naoki Yamauchi and Alley E. Watada

Chlorophylls and xanthophylls were monitored in broccoli (Brassica oleracea L. var. italica Plen.) florets stored in air, air + 10 ppm ethylene, or 10% CO2 + 1% O2 controlled atmosphere (CA) at 15 °C. Chlorophylls a and b, as measured with high-performance liquid chromatography, decreased in florets held in air. The decrease was accelerated by ethylene treatment and suppressed in CA. Chlorophyllide a and pheophorbide a were present in fresh broccoli florets, but the levels decreased significantly in all treatments during storage. The oxidized product of chlorophyll a, 132-hydroxychlorophyll a, did not accumulate. Xanthophylls decreased, but new pigments, suggested to be esterified xanthophylls, formed with yellowing in stored florets. The chlorophyll degradative pathway in broccoli florets was not altered by ethylene or CA and differed from that reported for parsley (Petroselium crisum Nym.) and spinach (Spinacia oleracea L.) leaves.

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Naoki Yamauchi and Alley E. Watada

Pigments in stored parsley leaves (Petroselinum crispum Nym.) were monitored to determine if degradative products of chlorophyll (chl) differed while under different types of atmosphere. The leaves were stored in a closed container under a stream of humidified air at 20C with or without 10 ppm ethylene and with or without 10 percent oxygen and 10 percent carbon dioxide. Analysis of pigments with HPLC showed that chl a and b decreased sharply with or without ethylene and the decrease was considerably less under CA. Chlorophyll a-1, the oxidized form of chl a, was initially low, and the level decreased slightly with all of the storage conditions. Chlorophyllide was also low, but it increased slightly during storage. Xanthophyll derivatives, which appeared to be the esterified xanthophylls, increased slightly during storage. These results indicate that chl degradation in stored parsley leaves was hastened by ethylene or suppressed by CA condition and the pathway of chl degradation did not appear to be altered by the different storage atmospheres.

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Naoki Yamauchi, Xiao-Ming Xia, and Fumio Hashinaga

Effects of flavonoid pigments on chlorophyll (Chl) degradation by Chl peroxidase in the flavedo of Wase satsuma mandarin (Citrus unshiu Marc. var. praecox Tanaka) fruits were studied. Chl was degraded when hydrogen peroxide was added in a reaction mixture containing Chl and a phosphate buffer extract from the flavedo. Chlorophyllide, which was formed by the action of chlorophyllase in the extract, was also degraded. The flavonoid contents decreased with the Chl degradation in the reaction mixture. Analysis of the flavonoid with HPLC showed that hesperidin and narirutin were contained in the flavedo as a major flavonoid, and that the former decreased significantly and the latter showed almost no change with the Chl degradation in the reaction mixture. In the ethylene-treated fruits, the hesperidin content in the flavedo also decreased with the degreening of stored fruits, suggesting that the flavonoid oxidation by Chl peroxidase could be involved in the Chl degradation.

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Shigenori Yaguchi, Tetsuya Nakajima, Toshihisa Sumi, Naoki Yamauchi, and Masayoshi Shigyo

Eight kinds of japanese bunching onion [Allium fistulosum L. (genomes FF)]-shallot [Allium cepa L. Aggregatum group (genomes AA)] monosomic addition line [MAL (FF+1A-FF+8A)] were used to study the effects of single alien chromosomes from A. cepa on the production of carbohydrates in the leaf tissues of A. fistulosum. Carbohydrate contents in green leaf blades of these MALs were measured during alternate months from May 2005 to Mar. 2006. The determination of soluble sugar content from leaf blades of each MAL and A. fistulosum revealed that nonreducing sugars (sucrose and fructan) accumulated in winter leaf blades. Reducing sugar (fructose) in the leaf blades of each MAL was lower than A. fistulosum in almost every time period. In the leaf blades of FF+4A, high fructan accumulation was observed from Nov. 2005 to Mar. 2006. A series of determinations on the pectin content showed that amounts of NaOH-soluble pectin and HCl-soluble pectin remained at low levels at all time periods. High pectin accumulations in FF+7A and FF+8A occurred in September and slightly decreased in November. A decrease in hexametaphosphoric acid-soluble pectin content was associated with the maturity of MALs from autumn to winter, whereas the water-soluble pectin content increased. The pectin methylesterase and polygalactronase genes of shallot were assigned to chromosome 7A and 4A, respectively. These results demonstrate that important genes related to pectin metabolism in shallot are located on chromosomes 4A, 7A, and 8A of shallot.

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Shinichi Masuzaki, Naoyuki Araki, Naoki Yamauchi, Naoko Yamane, Tadayuki Wako, Akio Kojima, and Masayoshi Shigyo

Bulb onion (Allium cepa L.) has a very large genome composed of a high proportion of repetitive DNAs. Genetic analyses of repetitive sequences may reveal microsatellites in order to increase the number of genetic markers in onion. Thirty microsatellites were previously isolated from an onion genomic library (Fischer and Bachmann, 2000). A complete set of Japanese bunching onion (A. fistulosum) – shallot (A. cepa Aggregatum group) monosomic addition lines were used to assign these microsatellites to the chromosomes of A. cepa. Simplified PCR conditions for each microsatellite were determined and 28 of the 30 primer pairs amplified DNA fragments, of which 21 microsatellite markers were assigned to chromosomes of A. cepa. Subsequent mapping of these microsatellites will enable us to establish the chromosomal distribution of these markers.

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Shigenori Yaguchi, Masanori Atarashi, Masatoshi Iwai, Shin-ichi Masuzaki, Naoki Yamauchi, and Masayoshi Shigyo

Eight Allium fistulosum L.–Allium cepa L. Aggregatum group (shallot) monosomic addition lines (2n = 17, FF+1A–FF+8A) have been useful in revealing the effects of single alien chromosomes from A. cepa on the production of l-ascorbic acid in the leaf tissue of A. fistulosum. In this study, the determination of ascorbic acid content revealed that the incorporation of alien chromosome 1A into a diploid background of A. fistulosum increased the internal ascorbic acid content of the leaf blade tissue. We produced a 1A disomic addition in the tetraploid of A. fistulosum (2n = 34) and demonstrated high-frequency transfer of the alien chromosome in crosses with A. fistulosum. Five plants of the 1A disomic additions were regenerated via apical meristem culture of the FF+1A on a Murashige and Skoog medium containing colchicine. These 1A disomic additions showed partial fertility for female and male gametes. Most of the progenies from selfing of the 1A disomic additions and reciprocal crossing with A. fistulosum possessed chromosome 1A. Interestingly, 64% (18 of 28) of the plants obtained from the reciprocal crosses were 1A monosomic additions in a triploid background of A. fistulosum. These monosomic additions were more vigorous and vitamin C-rich than euploid plants of A. fistulosum.