17ß-neriifolin from unripe fruits of Cerbera manghas suppressed cell proliferation via the inhibition of HOXA9-dependent transcription and the induction of apoptosis in the human AML cell line THP-1.

Homeobox A9 (HOXA9) is a transcription factor that is overexpressed in acute myeloid leukemia (AML). It is associated with the pathogenesis and progression of AML, and is a factor responsible for a poor prognosis. Therefore, the development of HOXA9-targeting molecules may contribute to not only better understanding of the mechanism of HOXA9 regulation, but also the development of therapeutic applications. We constructed a reporter assay system using the promoter region of the KBTBD10 gene, to which HOXA9 directly binds and regulates transcription, in the human acute monocytic leukemia cell line THP-1. Using this luciferase gene assay, we screened 1120 plant extracts and a methanol extract of the unripe fruits of Cerbera manghas was found to suppress the reporter gene expression mediated by the KBTBD10 promoter. From the extract, five steroid-type compounds were identified as the active constituents: 7α-neriifolin (1), 17ß-neriifolin (2), 17α-digitoxigenin ß-D-glucosyl-(1 → 4)-α-L-thevetoside (3), 17ß-digitoxigenin ß-D-glucosyl-(1 → 4)-α-L-thevetoside (4), and acetylthevetin B (5). Among the five compounds, 17ß-neriifolin most potently inhibited HOXA9-dependent gene expression without affecting the HOXA9 mRNA levels, and suppressed cell proliferation by inducing apoptosis. The findings on the structure-activity relationships of the compounds from C. manghas may contribute to the development of small molecule inhibitors of HOXA9.

Retusone A, a Guaiane-Type Sesquiterpene Dimer from Wikstroemia retusa and Its Inhibitory Effects on Histone Acetyltransferase HBO1 Expression.

Retusone A (1), a new sesquiterpene dimer consisting of two guaiane-type sesquiterpenoids, and oleodaphnal (2) were isolated from heartwood of Wikstroemia retusa (Thymelaeaceae). The planar structure of 1 was elucidated on the basis of HRESIMS and NMR spectroscopic data, and the relative stereochemistry was established by X-ray diffraction analysis. The absolute configuration of 1 was determined by electronic circular dichroism. Compound 1 suppressed luciferase reporter gene expression driven by the HBO1 (histone acetyltransferase binding to ORC1) gene promoter in human breast cancer MCF7 cells. Compound 1 also decreased the expression of endogenous HBO1 mRNA and protein, and inhibited proliferation of the cells. These results suggest that retusone A (1), which has a unique dimeric sesquiterpenoid structure with inhibitory activity against HBO1 expression, may contribute to the development of a novel therapeutic candidate for the treatment of breast cancer.

Piperine-like alkamides from Piper nigrum induce BDNF promoter and promote neurite outgrowth in Neuro-2a cells.

Black pepper (Piper nigrum) contains a variety of alkamides. Among them, piperine has been reported to have antidepressant-like effects in chronically stressed mice, but little is known about the biological activity of other alkamides. In this study, we investigated the effects of alkamides from white pepper (P. nigrum) on neuronal cells. Twelve alkamides were isolated from white pepper MeOH extracts, and their chemical structures were identified by NMR and MS analyses. The compounds were subjected to assays using the luciferase-reporter gene under the control of the BDNF promoter or cAMP response element in mouse neuroblastoma Neuro-2a cells. In both assays, marked reporter-inducing activity was observed for piperine (1), piperettine (2) and piperylin (7), all of which have in common an (E)-5-(buta-1,3-dien-1-yl)benzo[d] [1, 3] dioxole moiety. Piperettine (2) and piperylin (7) tended to increase endogenous BDNF protein levels. Furthermore, piperylin (7) promoted retinoic acid-induced neurite outgrowth. These results suggest that piperylin (7), or analogues thereof, may have a beneficial effect on disorders associated with dysregulation of BDNF expression, such as depression.

Two Alkaloids from Bulbs of Lycoris sanguinea MAXIM. Suppress PEPCK Expression by Inhibiting the Phosphorylation of CREB.

In the fasting state, gluconeogenesis is upregulated by glucagon. Glucagon stimulates cyclic adenosine monophosphate production, which induces the expression of key enzymes for gluconeogenesis, such as cytosolic phosphoenolpyruvate carboxykinase (PEPCK-C), which are involved in gluconeogenesis through the protein kinase A/cAMP response element-binding protein (CREB) pathway. Using a luciferase reporter gene assay, a methanol extract of the bulbs of Lycoris sanguinea MAXIM. var. kiushiana Makino was found to suppress cAMP-enhanced PEPCK-C promoter activity. In addition, two alkaloids, lycoricidine and lycoricidinol, in the extract were identified as active constituents. In forskolin-stimulated human hepatoma cells, these alkaloids suppressed the expression of a reporter gene under the control of cAMP response element and also prevented increases in the endogenous levels of phosphorylated CREB and PEPCK mRNA expression. These results suggest that lycoricidine and lycoricidinol suppress PEPCK-C expression by inhibiting the phosphorylation of CREB and may thus have the potential to prevent excessive gluconeogenesis in type 2 diabetes. Copyright © 2016 John Wiley & Sons, Ltd.

A new bis-seco-abietane diterpenoid from Hyptis crenata Pohl ex Benth.

A new, highly oxidized, bis-seco-abietane diterpenoid named hyptisolide A (1) was isolated from Hyptis crenata Pohl ex Benth. Its structure and stereochemistry were elucidated on the basis of data obtained by HRESIMS, NMR, and X-ray diffraction analyses, and its absolute configuration was determined with vibrational circular dichroism spectroscopy. By reporter gene assay, 1 was demonstrated to induce cAMP-responsive element-dependent transcription in Neuro2A cells.

Phenolic diterpenes from rosemary suppress cAMP responsiveness of gluconeogenic gene promoters.

The cAMP/protein kinase A/cAMP response element (CRE)-binding protein pathway is important for various physiological aspects including regulation of gluconeogenic gene expression. Rosemary, a well-known herb, has been reported to decrease blood glucose levels. We found that methanol extracts of rosemary suppressed forskolin (FSK)-stimulated luciferase expression under the control of CRE, as well as the promoters for cytosolic phosphoenolpyruvate carboxykinase (PEPCK-C) and glucose-6-phosphatase (G6Pase) catalytic subunit genes in human hepatoma HepG2 cells. Three abietane-type diterpenes and two flavonoids were isolated from the rosemary extracts. Among these, 7-O-methylrosmanol (1) and royleanonic acid (3) effectively suppressed FSK-induced luciferase expression under the control of the CRE, PEPCK-C and G6Pase gene promoters. PEPCK-C and G6Pase, which play a key role in the homeostatic regulation of blood glucose levels, are important for managing type II diabetes mellitus. Therefore, the ability of rosemary and its components to suppress cAMP responsiveness of the PEPCK-C or G6Pase gene may contribute to its antihyperglycemic activity.

Compounds isolated from Curcuma aromatica Salisb. inhibit human P450 enzymes.

Curcuma species (Zingiberaceae) are used as both food and medicine in Asia. Ten sesquiterpenes (1-10) and two curcuminoids (11 and 12) were isolated from the rhizomes of Curcuma aromatica Salisb. and identified. The compounds were evaluated for their ability to inhibit cytochrome P450 (CYP). Among them, the sesquiterpene (4S,5S)-(+)-germacrone-4,5-epoxide (7) inhibited certain subtypes of CYP more potently than or at levels comparable to the curcuminoids curcumin (11) and demethoxycurcumin (12); 7 (IC(50) = 1.0 ± 0.2 µM) > 12 (IC(50) = 7.0 ± 1.7 µM) > 11 (IC(50) = 14.9 ± 1.4 µM) for CYP3A4 inhibition; 12 (IC(50) = 1.4 ± 0.2 µM) > 11 (IC(50) = 6.0 ± 1.4 µM) > 7 (IC(50) = 7.6 ± 2.5 µM) for CYP2C9 inhibition; and 7 (IC(50) = 33.2 ± 3.6 µM) = 12 (IC(50) = 34.0 ± 14.2 µM) > 11 (IC(50) > 100 µM) for CYP1A2 inhibition. These results suggest the possibility that Curcuma aromatica Salisb. may cause food-drug interactions via cytochrome P450 inhibition by sesquiterpene 7 and curcuminoids 11 and 12.

[Identification methods of terpenoid gum bases using TLC and GC/MS].

Simple chromatographic methods were applied to terpenoid resins used as gum bases. Five triterpenoid resins, mastic, dammar resin, olibanum, benzoin gum and elemi resin, and two diterpenoid resins, rosin and copal resin, were separated with normal-phase TLC. Characteristic patterns were observed for all resins. Different samples of the same resin gave identical patterns. The TLC method is a candidate for a simple identification test for terpenoids resins. Samples were then methyl-esterified and analyzed with GC/MS. All resins exhibited characteristic chromatograms for total ion current. Major constituents of all resins were detected. Unique constituents that can be used as indicators were found in every resin. Therefore, GC/MS of methyl-esterified terpenoid resins is a valuable identification method.

Ovafolinins A-E, five new lignans from Lyonia ovalifolia.

Five new lignans, ovafolinins A-E (1-5), were isolated from the wood of Lyonia ovalifolia (Ericaceae). The structures of 1-5 were elucidated based on 2D NMR spectroscopy, X-ray crystallography, and other chemical methods.

[Examination of original plant of Jamaica quassia extract, a natural bittering agent, based on composition of the constituents].

Jamaica quassia extract is a natural bittering agent used as a food additive in Japan. The main constituents of the extract have already been reported to be quassin and neoquassin. In this study, the differences in composition of the constituents among four Jamaican quassia extract products were analyzed by LC/MS. The results showed that the four products have similar compositions of their minor constituents, as well as their main constituents. We isolated four of the minor constituents that were commonly included in the four products, and identified them as 11-dihydro-12-norneoquassin, canthin-6-one, 4-methoxy-1-vinyl-beta-carboline and 4,9-dimethoxy-1-vinyl-beta-carboline. The List of Existing Food Additives in Japan mentions that Jamaica quassia (Picrasma excelsa) is the original plant from which Jamaica quassia extract is produced. However, we presume that Jamaica quassia extract may actually be made from appropriate plants other than Picrasma excelsa, since P. excelsa is listed as an endangered species by the International Union for Conservation of Nature and Natural Resources. We prepared hot water extracts from two other species of plants, Quassia amara (American quassia, Surinam quassia) and P. quassioides ('Nigaki' in Japanese), and investigated their constituents by LC/MS. The results showed that the compositions of the constituents in the Jamaica quassia extract products resembled those in the extract derived from Q. amara. These findings suggest that Jamaica quassia extract products are probably made from Q. amara.

[Survey of synthetic disinfectants in grapefruit seed extract and its compounded products].

Grapefruit seed extract (GSE), derived from the seeds of grapefruit (Citrus paradisi MCAF.), is listed as a natural food additive in Japan. Products containing GSE are used as disinfectants made from only natural sources, especially after Japanese researchers found that GSE prevents the growth of norovirus. On the other hand, recent overseas studies indicated that synthetic disinfectants, such as benzalkonium and benzethonium chlorides, were present in some commercial GSE products. To confirm the quality of commercial GSE products available in Japanese markets, we carried out comprehensive research to identify the major constituents of commercial GSE products which are used as food additives (13 products from 6 manufacturers), dietary supplements (5 products from 4 manufacturers), cosmetic materials (16 products from 10 manufacturers) and disinfectant or deodorant sprays (7 products from 7 manufacturers). By means of NMR and LC/MS analysis, synthetic disinfectants such as benzethonium or benzalkonium salts were detected in most of the commercial GSE products.

Antioxidant phenylpropanoid glycosides from the leaves of Wasabia japonica.

From the MeOH extract of the leaves of W. japonica, seven phenylpropanoid gentiobiosides (1-7) were isolated along with eight known phenylpropanoids (8-15). Structures of 1-7 were determined based on spectroscopic data and chemical evidence. The activity of compounds 1-15 to scavenge superoxide anion radicals was investigated using an electron spin resonance (ESR) method.

Neocrocin A: a novel crocetin glycoside with a unique system for binding sugars isolated from gardenia yellow.

A novel crocetin glycosyl ester, neocrocin A (2), was isolated from gardenia yellow. The structure of 2 was elucidated as that of an all-trans-crocetin beta-D-gentiobiosyl beta-D-glucopyranosyl-(1-->6)-D-2-deoxy-glucopyranos-2-yl diester based on chemical and spectral data. The findings provide evidence that the binding system of crocetin glycosides is not limited to the anomeric position.

Structure of a new cyclic nonapeptide, segetalin F, and vasorelaxant activity of segetalins from Vaccaria segetalis.

A new cyclic nonapeptide, segetalin F, has been isolated from the seeds of Vaccaria segetalis and the structure including absolute stereochemistry was elucidated by using 2D NMR and chemical means. A series of segetalins showed a vasorelaxant activity against norepinephrine (NE)-induced contractions of rat aorta.

Phenylpropanoid derivatives from edible canna, Canna edulis.

Two phenylpropanoid sucrose esters were isolated from dry rhizomes of Canna edulis Ker Gawl., along with a known phenylpropanoid sucrose ester and four known phenylpropanoids. On the basis of analysis of spectroscopic data and chemical evidence, these two phenylpropanoid sucrose esters were shown to be 3-O-p-coumaroyl-6-O-feruloyl-beta-D-fructofuranosyl 6-O-acetyl-alpha-D-glucopyranoside and 3,6-di-O-p-coumaroyl-beta-D-fructofuranosyl 6-O-acetyl-alpha-D-glucopyranoside.

Determination of antioxidant activity of herbs by ESR.

Water extracts of 32 herbs that are constituents of curry and curry powder were screened for superoxide anion radical (O2.-) scavenging activity. Among the screened samples, only clove, allspice, and basil were shown to decrease DMPO-O2.- adduct yields by more than 50% at 0.25 mg/mL as measured by an ESR spin trapping technique based on the HPX-XOD reaction. To study the mechanism of the O2.- scavenging activity, Km values were obtained from a Lineweaver-Burk plot for XOD in the presence of different concentrations of HPX, and the IC50 values at different DMPO concentrations were compared. Clove and basil directly eliminated O2.- like superoxide dismutase (SOD), whereas allspice reduced the amount of O2.- by inhibition of formation of O2.-.