Toshiake Matsuzaki

Disaccharide esters screened for inhibition of tumor necrosis factor-α release are new anti-cancer agents

Tumor necrosis factor‐α (TNF‐α) is a proinflammatory cytokine playing a part in various pathological states. Non‐toxic inhibitors of TNF‐α release are thought to be promising agents for cancer prevention. We found that the acetone fraction of the tobacco leaf surface lipid containing glucose esters and sucrose esters inhibited both TNF‐α release from BALB/3T3 and KATO III cells induced by okadaic acid and tumor promotion by okadaic acid on mouse skin initiated with 7,12‐dimethylbenz(a)anthracene (DMBA). Next, we investigated the inhibition of TNF‐α release with synthetic disaccharide esters, such as 6,6′‐di‐0‐alkanoyl‐α,α‐trehaloses (6,6′‐diester‐trehaloses), 4,4′‐di‐0‐alkanoyl‐α,α‐trehaloses (4,4′‐diester‐trehaloses) and 6,6′‐diamino‐6,6′‐dideoxy‐N, N′‐dial‐kanoyl‐α,α‐trehaloses (6,6′‐diamide‐trehaloses) bearing fatty acids of various chain lengths, and n‐ dodecyl‐β‐D‐maltoside as a disaccharide monoester. 6,6′‐Diester‐trehaloses and 4,4′‐diester‐treha‐loses of C8 to C14 fatty acids, 6,6′‐diamide‐trehaloses of C8 to C14 fatty acids, and n‐dodecyl‐β‐D‐maltoside all inhibited TNF‐α release in a dose‐dependent manner. The IC50 values are 7.4‐14.8 μM for 6,6′‐diester‐trehaloses (C8 to C12), 14.6‐21.6 μM 4,4′‐diester‐trehaloses (C8 to C12), 2.9‐15.0 μM for 6,6′‐diamide‐trehaloses (C8 to C14) and 23 μM for dodecyl‐β‐D‐maltoside. Both 6,6′‐di‐O‐octanoyl‐α,α‐trehalose (C8, designated as SS555) and n‐dodecyl‐β‐D‐maltoside (C12) inhibited tumor promotion by okadaic acid on mouse skin initiated with DMBA. Percentages of tumor‐bearing mice in week 15 of tumor promotion were reduced from 60.0 to 13.3 with SS555, and to 46.7 with n‐dodecyl‐β‐D‐maltoside. Moreover, SS555 inhibited TNF‐α gene expression mediated through inhibition of AP‐1 activation, but not NF‐αB activation. This paper reports that diester‐trehaloses of C8 to C12 fatty acids and mimics of disaccharide monoesters such as n‐dodecyl‐β‐D‐maltoside appear to be potential cancer‐preventive agents of a new type.

Hydroxy and normal fatty acid distribution in stigmas of Nicotiana and other plants

Abstract The ω-hydroxy and normal fatty acid distribution in stigmas of 51 Nicotiana species and 25 other plant species is presented. All Nicotiana species studied had high proportions of ω-hydroxy fatty acids (C18: 1 -ω-OH and C18: 2-ω)-OH), constituting from 22 to 83% of the total fatty acid content. In most Nicotiana species, the percentage of C18: 1-ω-OH was higher than that of C18: 2-ω-OH and the percentage of C18: 1 was higher than that of C18: 2. The reverse pattern of this fatty acid composition was observed in the self-incompatible Nicotiana species. Many plant species with wet type stigma and a relatively few species with dry type stigma contained considerable amounts of C18: 1-ω-OH and C18: 2-ω-OH. The ω-hydroxy fatty acids were contained not only in the triacylglycerol and diacylglycerol fractions but also in the polar fraction of stigma lipids.

The fatty acid composition of seeds and leaves of Nicotiana species

Abstract Fatty acid analyses of seeds in 62 Nicotiana species and leaves in 56 Nicotiana species are presented. The total fatty acid content on a dry wt basis ranged from 25 to 40 %of seeds and from 2.1 to4.4% of green leaves. Linolenate was the dominant fatty acid in the leaves of all species studied, comprising 50–63% of the total fatty acid content. In seeds of most species linoleate predominated, constituting 69–79% of the total fatty acid content. Fourteen of 21 species in the section Suaveolentes and one species in the section Noctiflorae had relatively high proportions (10–38%) of linolenate. In two linolenate-rich species studied, linolenate was the major fatty acid of triacylglycerols which predominated in the seed lipids.

Sucrose esters from the surface lipids of Nicotiana cavicola

Three sucrose esters were isolated and identified for the first time from the surface lipids of Nicotiana cavicola. They contained acetic acid and such branched short-chain fatty acids as 5-methylhexanoic, 5-methylheptanoic, and 6-methylheptanoic acids. The structures of the sucrose esters were 6-O-acetyl-4-O-acyl-alpha-D-glucopyranosyl-4,6-O-diacetyl-3-O-acyl-beta- D- fructofuranoside, 4-O-acyl-alpha-D-glucopyranosyl-1,4-O-diacetyl-3-O-acyl-beta-D-fructofur anoside and 6-O-acetyl-4-O-acyl-alpha-D-glucopyranosyl-4-O-acetyl-3-O- acyl-beta-D-fructofuranoside.

New Types of Glycolipids from the Surface Lipids of Nicotiana umbratica

The surface lipids of Nicotiana umbratica contained several varieties of glycolipids. Three types of glucose esters and five types of sucrose esters were isolated and identified from this species. These glycolipids contained short-chain fatty acids such as acetic, methylpropionic, methylbutyric, 3-methylpentanoic, 4-methylpentanoic and methylhexanoic acids. The acetylated positions of each compound were determined by 2-D NMR, using the HMBC technique. The structures of the glucose esters were 1,6-di-O-acetyl-2,3-4-tri-O-acyl-alpha-D-glucopyranose (I), 6-O-acetyl-2,3,4-tri-O-acyl-alpha-D-glucopyranose (II) and 2,3,4-tri-O-acyl-alpha, beta-D-glucopyranose (III). The structures of the sucrose esters were 6-O-acetyl-2,3,4-tri-O-acyl-alpha-D-glucopyranosyl-1,3-di-O-acetyl-beta- D- fructofuranoside (IV), 2,3,4-tri-O-acyl-alpha-D-glucopyranosyl-1,3-di-O-acetyl-beta-D-fructofur anoside (V), 6-O-acetyl-2,4-di-O-acyl-alpha-D-glucopyranosyl-3-O-acetyl-beta-D- fructofuranoside (VI), 2,3,4-tri-O-acyl-alpha-D-glucopyranosyl-3-O-acetyl-beta-D-fructofuranosi de 2,3,4-tri-O-acyl-alpha-D-glucopyranosyl-3-O-acetyl-beta-D-fructofuranosi de (VII) and 2,3,4-tri-O-acyl-alpha-D-glucopyranosyl-beta-D-fructofuranoside (VIII). Among them, I, IV, V and VI were isolated for the first time from plants of the family Solanaceae. These newly isolated glycolipids were inhibitory against barnyardgrass growth.

Sucrose esters from the surface lipids of Petunia hybrida

Abstract Three sucrose ester types were isolated and identified from the leaf surface lipids of Petunia hybrida. They contained both unbranched and branched fatty acids (from C2 to C8). Further, one of these sucrose esters contained malonic acids. The structures of the sucrose esters were 2,3,4-O- tri-acyl -α- d -glucopyranosyl -1-O- malonyl -4-O- acyl -6-O- acetyl -β- d -fructofuranoside , 2,3,4-O- tri-acyl -α- d -glucopyranosyl -4-O- acyl -6-O- acetyl -β- d -fructofuranoside and 2,3,4,6-O- tetra-acyl -α- d -glucopyranosyl -β- d -fructofuranoside .

Production of somatic hybrids between Nicotiana benthamiana and N. tabacum and their resistance to aphids

Abstract In order to introduce aphid resistance into tobacco, somatic hybrids between tobacco and aphid resistant Nicotiana benthamiana were produced. The hybrids which were proved to be true hybrids on the basis of electrophoretic analysis of ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) and restriction fragment length polymorphism (RFLP) analysis showed significant resistance to aphids. In the leaf surface lipids of the hybrids, lipids specific to N. benthamiana were detected with those specific to tobacco. Five out of 22 of the hybrids were fertile in self-pollination.

Changes in glycerolipid content and fatty acid composition during tobacco seed germination.

Abstract Tobacco seeds were germinated under 12 hr photoperiod at 28° for eight days and the contents of individual glycerolipids as their constituent fatty acids, as well as the fatty acid composition, were determined at one to two day intervals. Triacylglycerols, which were by far the largest seed lipid component, decreased markedly from 32 to 8.2% of seed mass. 1,2-Diacylglycerols increased remarkably in amount at first but then decreased; 1,3-diacylglycerol and free fatty acid content also increased as did that of phospholipids. Both monogalactosyldiaclyglycerol and digalactosyldiacylglycerol, as well as chlorophyll, were detected after three days of germination and largely increased thereafter. The fatty acid composition of the triacylglycerols did not change significantly during germination. Generally, in phospholipids the proportion of oleate decreased and that of linolenate increased, while in glycolipids the percentage of linolenate increased with concomitant decreases in that of palmitate, stearate and oleate.

Changes in glycerolipids and their fatty acid composition during maturation of tobacco seeds

Abstract Triacylglycerol, which was one of the minor lipid components in immature seeds of tobacco, accumulated dramatically between 7 and 27 days after flowering and, in mature seeds at 37 days, the fatty acid methyl esters of the triacylglycerols comprised 96.3% of those of the total lipids. Diacylglycerols and sterol ester also increased significantly during seed development. Phosphatidylcholine and phosphatidylethanolamine, which were major components in immature seeds, decreased constantly with increasing maturation as well as the quantities of phosphatidylinositol and phosphatidylglycerol. Monogalactosyldiacylglycerols, digalactosyldiacylglycerols and sulfoquinovosyldiacylglycerols also decreased and disappeared in mature seeds. In the triacylglycerols the percentages of palmitate, stearate and linolenate fell with increasing seed age, while that of linoleate increased up to 75.3% in mature seeds. A similar trend was observed in the fatty acid composition in the diacylglycerols and sterol ester. Generally, in the phospholipids the proportions of linoleate and linolenate decreased with concomitant increases of stearate and oleate.

Calcium chloride as a major component contributing to aphid resistance ofNicotiana benthamiana

Substances with antiaphid activity were extracted from the leaf surface of aphid-resistantN. benthamiana and purified. Sugar esters and diacyl glycerols were isolated from the chloroform extract, but only diacyl glycerols showed significant activity. However, the major activity was found in the water extract rather than in the chloroform extract. From the water extract calcium chloride was isolated as the most abundant active substance. Only calcium chloride showed significant activity among several calcium salts and chlorides of several metals that are abundant in plants. Calcium contents per unit area of leaf surface ofN. benthamiana and aphid-resistantN. gossei were almost equal to each other and 10–100 times higher than that of aphid-susceptibleN. tabacum.