Einosuke Tamaki

p-coumaroylputrescine, caffeoylputrescine and feruloylputrescine from callus tissue culture of Nicotiana tabacum

Abstract p-Coumaroyl-, caffeoyl-and feruloylputrescine have been isolated from callus tissue culture of Nicotiana tabacum. Proof of the structures was obtained by hydrolysis, color reactions, and comparison of i.r., u.v. and NMR spectra with those of synthetic compounds. p-Coumaroylputrescine and caffeoylputrescine are new natural products.

N-methylputrescine oxidase from tobacco roots

Abstract N -Methylputrescine oxidase, catalyzing the oxidative deamination of the primary amino group of N -methylputrescine, was demonstrated in roots of tobacco plants, and purified 150-fold. N -Methylpyrrolinium salt was identified as the reaction product by comparison with the authentic compound. The enzyme had a pH optimum at 8·0 and the K m value for N -methylputrescine was 4·5 × 10 −4 M. Putrescine and cadaverine were also oxidized. Activity of the enzyme was strongly inhibited by carbonyl reagents, thiol inhibitors and diethyldithiocarbamate, but hydrazine derivatives were without significant effect on the enzyme activity. The enzyme was localized exclusively in the roots and its activity markedly increased on decapitation of the shoots. The demonstration of N -methylputrescine oxidase, in addition to putrescine N -methyltransferase, provides evidence that the biosynthetic route of nicotine involves N -methylputrescine and 4-methylaminobutanal ( N -methylpyrrolinium salt) as intermediates.

Studies on nitrogen metabolism in tobacco plants: VI. Metabolism of glutamic acid, γ-aminobutyric acid, and proline in tobacco leaves

Abstract Metabolism of glutamic acid, γ-aminobutyric acid, and proline in tobacco leaves during the day and night has been studied by using isotopic labeled compounds. Biosynthesis of proline from glutamate was very active in the day but slow at night; this indicates that light has an important role in the synthesis. γ-Aminobutyrate was very readily transformed into organic acids, and most of the activity was found in malate, succinate, and citrate. Accordingly, in tobacco leaves γ-aminobutyrate seems to be oxidized to succinate via succinic semialdehyde and to enter the tricarboxylic acid cycle. Oxidation of proline to glutamate took place very slowly in tobacco leaves both in the day and night without any appreciable difference. Reactions of proline in the stem and roots were similar to those in leaves. Accumulation of the activity of glutamate and γ-aminobutyrate in aspartate and citrate was higher at night than in the day.

Chlorophyllase of tobacco plants. II. Enzymic phytylation of chlorophyllide and pheophorbide in vitro

Abstract The breakdown of chlorophyll by the water-soluble chlorophyllase prepared from tobacco chloroplasts was decreased by a comparatively low concentration of phytol. The same enzyme preparation could phytylate chlorophyllide to give chlorophyll. It also synthesized pheophytin from pheophorbide and phytol. Chlorophyllide and pheophorbide were prepared by a simple and quick method. They were identified by their mobility on chromatograms, HCl numbers, and absorption spectra. The possibility that chlorophyllase acts in the biosynthetic pathway of chlorophyll in higher plants as well as in vitro systems was discussed.

Phytochemical Studies on the tobacco alkaloids—x. : Degradation of the tobacco alkaloids and their optical rotatory changes in tobacco plants

Abstract Myosmine(1,2-dihydronornicotine) was isolated from Nicotiana glutinosa at the flowering stage and was proved to be a degradation product of (−)-nornicotine by feeding the latter to tobacco plants grafted onto tomato stock. 1,2-Dehydroanabasine was also found to be a degradation product of anabasine in tobacco leaves. Racemic nornicotine, anabasine and 1-(3-pyridyl)-1-ethylammoethane which have a similar 3-pyridyliminomethane skeleton were optically activated in tobacco leaves. It can be considered that the first step of the degradation of secondary amine tobacco alkaloids is a stereospecific dehydrogenation. From the findings made here and the previous observation on the optical rotation of secondary amine tobacco alkaloids, it was inferred that such secondary amine tobacco alkaloids as anabasine, anatabine and some nornicotine in the root are biosynthesized in a racemic form in tobacco plants.

Studies on nitrogen metabolism in tobacco plants. A. Part II. Diurnal variation in the amino acid composition of tobacco leaves

Abstract Nonprotein nitrogen compounds of tobacco leaves, particularly the diurnal variation in the amino acid composition, have been investigated. The main constituents of the free amino acids in the green leaves were γ-aminobutyric acid, glutamic acid, aspartic acid, proline, serine, alanine, glycine, and threonine in order of their quantities, and they composed in total more than 90% of the total nitrogen of the free amino acids. All the free amino acids except aspartic acid had their maximum contents around 2 p.m. Serine, glycine, and arginine, in particular, increased rapidly in the daytime, whereas aspartic acid had its maximum content at night. The content of γ-aminobutyric acid varies irregularly with time, but it was always predominant over the other amino acids.

Metabolism of chlorophyll in higher plants—I.: A thin-layer chromatography for the quantitative determination of phytol

Abstract Thin-layer silicic acid chromatography has been shown to be a useful technique of the separation of phytol. The procedure can be applied to the quantitative separation and estimation of phytol of various plant materials.

Metabolism of nicotinic acid in tobacco plants

Abstract 14C from nicotinic acid-6-14C which was administered to tobacco plants was incorporated into nine pyridine compounds during 3 hr incubation; two of these were identified as nicotinic acid-N-glucoside and 6-hydroxynicotinic acid. Nicotinic acid was only incorporated into intermediates of the pyridine nucleotide cycle to a limited extent the main product being the glucoside. Nicotinic acid glucoside was incorporated into nicotine with the same efficiency as nicotinic acid, but the rate of the incorporation was markedly reduced when unlabelled nicotinic acid was also fed, suggesting that the glucoside is not involved in the direct route of nicotine biosynthesis.

Changes in the amino acid composition of tobacco cells in suspension culture

Abstract Changes in the content of free and protein amino acids of tobacco cells in suspension culture during the growth were determined. The principal free amino acids throughout the culture period were glutamine, asparagine, and γ-aminobutyric acid. Glutamic acid, glycine, and lysine decreased in the lag period, followed by the increase in the log period, and then decreased towards the stationary phase. By contrast alanine, serine, valine, leucine, and proline increased in the lag period, followed by the decrease in the log period. The contents of glutamic acid and hydroxyproline in the protein fraction changed markedly during the growth. The glutamic acid content increased in the early period of the log phase and then gradually decreased whereas the hydroxyproline content decreased in the log period and then abruptly increased.

The isolation and identification of nicotianine: A new amino acid from tobacco leaves

Abstract A hitherto unknown amino acid, nicotianine, was isolated from the leaves of tobacco plants. By the use of i.r., u.v. and NMR spectrophotometry, colour reactions and synthesis the new amino acid was identified as l (+)- N -(3-amino-3-carboxypropyl)-β-carboxypyridinium betaine.