Nariyuki Ishikura

A survey of anthocyanins in fruits of some angiosperms, II

Anthocyanins in the fruits of 41 species belonging to 25 families were investigated paper-chromatographically. Fifteen kinds of anthocyanins, in addition to the previous findings, were newly identified. In addition to cyanidin 3-monoglucoside as the most common anthocyanin, the 3-rutinoside and the 3-sambubioside were found in the fruits with high frequency.

Anthocyanins and other phenolics in autumn leaves

Abstract Anthocyanins and other phenolics in young, mature and autumn red leaves of Prunus, Rhus, Euonymus, Parthenocissus and Acer have been studied. In general, autumnal red leaves contained chrysanthemin, except that Rhus succedanea contained, in addition, a small amount of peonidin 3-monoglucoside. In three Acer species and one Prunus species, red colour of young leaves was due to a mixture of 3-mono-glucoside and 3-rutinoside of cyanidin, and some glycosides of peonidin were formed transiently in young leaves of R. succedanea . Hydrolysed leaf-extracts contained ellagic and gallic acids, kaempferol and quercetin.

An investigation of the intracellular site of anthocyanoplasts using isolated protoplasts and vacuoles

Microscopic observations made during preparation of protoplasts and vacuoles from red radish seedling hypocotyl (Raphanus sativus L.) show that anthocyanoplasts, the strongly pigmented bodies present in the pigmented cells of the hypodermis, begin as apparently membranous vesicles in the cytoplasm made visible by the deposition and accumulation of anthocyanins, but only rarely appear in the isolated vacuole. Isolation of protoplasts and vacuoles was also achieved from mung bean seedling hypocotyl (Vigna radiata L Wilczek), red cabbage leaf (Brassica oleracea L.) and Prunus x yedoensis Matsum callus. Anthocyanoplasts were usually in the vacuole, although sometimes in the cytoplasm, of the mung bean and cabbage, but were never seen in vacuoles of Prunus callus.

Biosynthesis of gallic and ellagic acids with14C-labeled compounds inAcer andRhus leaves

The biosynthetic pathway for gallic and ellagic acids in young, mature and autumn leaves ofAcer buergerianum andRhus succedanea was examined by tracer experiments, and also by isotope competition, withd-shikimic acid-14C,l-phenylalanine-U-14C,l-phenyllactic acid-U-14C, gallic acid-G-14C and their unlabeled compounds. In young leaves of both plants, the incorporation rate of labeled shikimic acid into gallic acid was significantly higher than that of labeled phenylalanine, whereas in the mature and autumn leaves the latter was a good precursor rather than the former for the gallic acid biosynthesis. Therefore, two pathways for gallic acid formation, through β-oxidation of phenylpropanoid and through dehydrogenation of shikimic acid, could be operating inAcer andRhus leaves, and the preferential pathway is altered by leaf age. In both plants, the incorporation rate of labeled phenyllactic acid during a 24 hr metabolic period was almost the same as that of labeled phenylalanine. The incorporation ofd-skikimic acid-G-14C,l-phenylalanine-U-14C andl-phenyllactic acid-U-14C into ellagic acid was very similar to the case of the radioactive gallic acid formation. Furthermore, regardless of the presence of unlabeled shikimic acid and/or phenylalanine, incorporation of the radioactivity of labeled gallic acid into ellagic acid occurred at a very high rate, suggesting the reciprocal radical reaction of gallic acid for the ellagic acid formation. The incorporation of labeled compounds into ellagitanins was also examined and their biosynthesis discussed further.

A stable reddish purple anthocyanin in the leaf of Gynura aurantiaca cv. 'Purple Passion'

The anthocyanin (GAA) in the epidermis and hair of the leaf ofGynura aurantiaca cv. ‘Purple Passion’ was isolated and identified as cyanidin tetra-glucoside acylated by three molecules of caffeic acid and one molecule of malonic acid. GAA was also isolated from the lower epidermis of the leaf ofG. bicolor DC. GAA showed a very stable reddish purple color from weakly acid to neutral pH region, but the color of the deacylated compound disappeared rapidly in the same region. This indicated that the attached organic acids must play an essential role in the stabilization of the color.Comparison of the profiles of the visible absorption spectra of the intact epidermal peels and cells ofG. aurantiaca andG. bicolor with those of GAA dissolved in various pH solutions suggested that the pH of the epidermal vacuole containing GAA was nearly 4.3.GAA was indistinguishable from the anthocyanin (rubrocinerarin) which we had previously isolated from the purplish red flowers ofSenecio cruentus DC. by means of UV-Vis, NMR and Mass spectra.

Seasonal changes in contents of phenolic compounds and sugar inRhus, Euonymus andAcer leaves with special reference to anthocyanin formation in autumn

Seasonal variation in sugar, total phenol and flavanol contents was examined inRhus, Euonymus andAcer leaves. In all plant leaves, the total phenol and flavanol content per leaf increased rapidly at the early growth stages but thereafter the content was kept rather constant. Later on, sugar content increased to a high level, and the autumnal reddening began. An excessive accumulation of sugar just before the reddening indicated that the accumulation related to the anthocyanin formation. The incorporation of radioactivity into anthocyanin in autumn leaves from glucose-[U-14C] and phenylalanine-[U-14C] was also observed.

3-Desoxyanthocyanin and other phenolics in the water fernAzolla

The phenolic compounds ofAzolla imbricata andA. japonica have been examined in the present study. Both species were found to contain luteolinidin 5-glucoside and several phenolic compounds, particularly chlorogenic acid, aesculetin, caffeic acid 3,4-diglucoside and 6-(3′-glucosylcaffeoyl)-aesculetin. In addition, glucose esters ofp-coumaric acid and glucose, 1,6-diester of caffeic and chlorogenic acids were present in a small amount. The acid- and alkali-hydrolyzates ofAzolla plants yielded caffeic acid and aesculetin present at the level of about 0.047% and 0.012% in fresh plants, and a large part of the caffeic acid seems to be present as the ester.

Distribution pattern of anthocyanins in the polygonaceae

A survey of anthocyanins in the flowers and other organs of thirty-three species of three genera belonging to the Polygonaceae has been carried out. There are thirteen anthocyans. Cyanidin glycosides, especially the 3-glycoside (chrysanthemin), are present most commonly and peonidin glycosides including the arabinosylglucoside are found with low frequency. The distribution of malvidin 3,5-diglucoside (malvin) is confined to the species belonging to the sectionEchinocaulon of the genusPolygonum. It is noted that cyanidin itself occurs in the stems ofPolygonum perfoliatum andP. senticosum.

An enzyme from Eucalyptus which converts cinnamoyl triacetic acid into pinosylvin

Abstract An enzyme catalysing the cyclization of cinnamoyl triacetic acid to form pinosylvin was isolated from Eucalyptus sideroxylon leaves and purified by Sephades column chromatography. One enzyme preparation showed a 132-fold increase in specific activity and was fairly stable when stored as a freeze-dried preparation at 2–4°. The optimum pH of the reaction was at 8·2. Mg 2+ at 2–10 mM concentration increased the rate of pinosylvin formation two and a half times, and Mn 2+ and boron also stimulated the reaction rate to some extent. On the other hand, the enzyme was little affected by reagents such as p -chloromercuribenzoate but was strongly inhibited by KCN and cysteine, and also by mercury and iodine at high concentration. The function of this enzyme in plants is discussed.

Multiple forms of flavonol O-glucosyltransferases in young leaves of Euonymus alatus F. Ciliato-dentatus

Abstract Two O -glucosyltransferases (F3GT and F7GT) catalysing the transfer of d -glucose from UDP- d -glucose to the 3- and 7-positions of the flavonol nucleus, respectively, were isolated from young leaves of Euonymus alatus f. ciliato-dentatus . F3GT was further separated into two isozymes (F3GT1 and F3GT2) by ion-exchange chromatography and chromatofocusing with recovery of ca a 1 : 1 ratio in activity. F3GT1, F3GT2 and F7GT were purified by ca 50-,95- and 136-fold, respectively, by precipitation with ammonium sulphate followed by passage through a series of columns. All three enzymes had a pH optimum of 7.5 and an M r of 48 × 103· Mg 2+ had a slight stimulatory effect on F3GT1 and F3GT2 activities, but F7GT had no Mg 2+ requirement. All three enzymes were strongly inhibited by 1 mM each of Zn 2+ , Cu 2+ , N-ethylmaleimide, p -chloromercuribenzoate and phenylmercuriacetate; and they were stimulated by 14 mM 2-mercaptoethanol (2-ME). F3GT1, F3GT2 and F7GT had an isoelectric point (pI) at pH 5.40, 4.92 and 4.45, respectively. F3GT1 and F3GT2 both mediated the transfer of d -glucose exclusively to the 3-hydroxyl group of kaempferol and some other flavonols, but neither 7- O -glucosides nor 3- O -glucosides were able to accept d -glucose. On the other hand, F7GT mediated the transfer of d -glucose exclusively to the 7-hydroxyl group of kaempferol and some other flavonols, and in addition, the 3- O -glucosides of kaempferol and quercetin were able to accept d -glucose, although less efficiently.