Inhibitory Constituents Research Articles

5 Alpha-Reductase and Aromatase Inhibitory Constituents from Brassica rapa L. Pollen

In the screening of biologically active constituents from Brassica rapa pollen, the supercritical CO(2) fluid extract (SFE-CO(2)) showed potent 5 alpha-reductase and aromatase inhibiting activity. The SFE-CO(2) extract was separated by various chromatographic methods to give two new phytosterol derivatives, 24-methylenecholesterol linolenate (1) and cycloeucalenol linolenate (2), as well as eight known compounds, 24-methylenecholesterol palmitate (3), cycloeucalenol (4), pollinastanol (5), 24-methylenecholesterol (6), linolenic acid (7), palmitic acid (8), monolinolein (9) and monopalmitin (10), compounds 7 and 9 showed potent 5 alpha-reductase inhibitory activity; compounds 1-6 and 10 showed potent aromatase inhibitory activity.

Topoisomerase I and II inhibitory constituents from the bark of Tilia amurensis

Two coumarins (1 and 6), one flavan-3-ol (2), one fatty acid (3), and two lignan glycosides (4 and 5) were isolated from the EtOAc and CH(2)Cl(2) extract of the bark of Tilia amurensis. Their chemical structures were identified by comparing their physicochemical and spectral data with those of published in literatures. Compounds 4, 5, and 6 were isolated from Tilia genus for the first time. Compounds 2 and 3 showed potent inhibitory activity against both DNA topoisomerase I (IC(50) values; 49 microM and 4 microM, respectively, with 18 microM of positive control compound, comptothecin) and DNA topoisomerase II (IC(50) values; 13 microM and 3 microM, respectively, with 50 microM of positive control compound, etoposide). However, all compounds did not showed cytotoxicity against the human colon adenocarcinoma cell line (HT-29), the human breast adenocarcinoma cell line (MCF-7), and human liver hepatoblastoma cell line (HepG-2).

The in vitro inhibition of human neutrophil elastase activity by some Yemeni medicinal plants

15 extracts of different polarities (dichloromethane, methanol and aqueous extracts) from 5 Yemeni medicinal plants (Aspilia helianthoides, (Schumach. & Thonn.) Oliv. & Hiern subsp. ciliate (Schumach.) C.D. Adams leaves; Ceropegia rupicola, Defl. var. rupicola whole plant; Kniphofia sumarae, Defler whole plant; Pavetta longiflora, Vahl. subsp. longifloraleaves; and Plectranthus cf barbatus (Thulin & Gifri) leaves) were tested for their inhibitory effects against the enzymatic activity of human neutrophil elastase (HNE) (EC 3.4.21.37) in an in vitro human neutrophil elastase inhibition assay. 14 extracts at various concentrations were found able to inhibit the activity of HNE. Among the plants tested,A. helianthoides was the most active inhibitor of HNE. The dichloromethane extracts of all tested plants, exhibited more inhibitory effect on HNE activity than the methanolic and aqueous extracts. The dichloromethane extract of A. helianthoides showed the most active inhibitory effect on the HNE activity (IC50 = 0.4μg/ml). Within the HNE inhibitory active methanolic extracts, those of A. helianthoides and P. cf barbatus were the most active inhibitors with IC50 of about 3μg/ml. These results provide some scientific justification for the use of A. helianthoides in traditional medicine and indicate the presence of HNE inhibitory constituents in the active tested plants.

Inhibitory Constituents of Lipopolysaccharide-Induced Nitric Oxide Production in BV2 Microglia isolated fromAmomum tsao-ko

A methanolic extract of the fruits of AMOMUM TSAO-KO (Zingiberaceae) significantly attenuated nitric oxide production in lipopolysaccharide-simulated BV2 microglia. Two new bicyclic nonanes characterized as 6,7-dihydroxy-indan-4-carbaldehyde ( 1) and 6-hydroxy-indan-4-carbaldehyde ( 2) were isolated with the eleven known compounds 6,7-dihydroxy-3,7-dimethyloct-2-enoic acid ( 3), tsaokoin ( 4), isotsaokoin ( 5), 8-oxogeraniol ( 6), P-menth-1-ene-5,6-diol ( 7), 3alpha-hydroxycarvotagenone ( 8), tsaokoarylone ( 9), 1,7-bis(4-hydroxy-3-methoxyphenyl)-4,6-heptadien-3one ( 10), (+)-hannokinol ( 11), MESO-hannokinol ( 12) and hannokinin ( 13), from the fruits of A. TSAO-KO using bioactivity-guided fractionation. All thirteen compounds significantly inhibited lipopolysaccharide-induced nitric oxide production in BV2 microglial cells at concentrations ranging from 1 microM to 100 microM.

Lipoxygenase inhibitory constituents from rhubarb

Phytochemical study on the ethanol extract of rhubarb led to the isolation of fifteen compounds, including five anthraquinones: chrysophanol (1), physcion (2), emodin (7), chrysophanol-8-O-beta-D: -glucopyranoside (9) and emodin-8-O-beta-D: -glucopyranoside (15), and ten stilbenes: desoxyrhaponticin (3), rhaponticin (4), resveratrol (5), desoxyrhapotigenin (6), rhapontigenin (8), piceatannol-3'-O-beta-D: -glucopyranoside (10), piceid (11), epsilon-viniferin (12), ampelopsin B (13) and isorhaponticin (14). Their structures were identified by comparing the physicochemical data with those of published papers. Among the isolated compounds, stilbene derivatives (3-6, 8 and 10-14) showed remarkable inhibitory effect on lipoxygenase with IC(50) values ranging from 6.7 to 74.1 microM. The inhibition kinetics analyzed by Lineweaver-Burk plots found that they were competitive inhibitors with the linoleic acid at the active site of lipoxygenase. In addition, stilbenes exhibited significantly free radical scavenging activity against ABTS(.+) with trolox equivalent activity capacity (TEAC) values ranging from 1.16 to 4.64. Whereas, anthraquinone derivatives (1-2, 7, 9 and 15) neither inhibited lipoxygenase nor scavenged free radical ABTS(.+). These results indicated that stilbene derivatives were considerate to be mainly lipoxygenase inhibitor and free radical scavenger constituents of rhubarb.

Inhibitory constituents of aldose reductase in the fruiting body of Phellinus linteus

Aldose reductase (alditol/NADP+ oxidoreductase, AR), the first enzyme in the polyol pathway, catalyzes the reduction of the aldehyde form of D‐glucose to D‐sorbitol with concomitant conversion of NADPH to NADP+. It is generally accepted that this polyol pathway plays an important role in the development of some degenerative complications of diabetes such as retinopathy, neuropathy, and nephropathy. In an effort to characterize active principles for diabetic complication from medicinal mushroom, AR inhibitors were isolated from the fruiting body of Phellinus linteus and identified as hispidin, phelligridimer A, davallialactone, methyldavallialactone, hypholomine B, interfungins A, and inoscavin A (11), together with protocatechuic acid (1), protocatechualdehyde, caffeic acid, and ellagic acid. Their structures were elucidated by spectroscopic analyses. Among them, davallialactone, hypholomine B, and ellagic acid exhibited potent rat lens AR and human recombinant AR inhibitory activity with IC50 values of 0.33, 0.82, 0.63 ¥ìM and 0.56, 1.28, 1.37 ¥ìM, respectively. Supported by Brain Korea 21.

A Survey of Watermelon Germplasm for Inhibitory Seed Exudates

Watermelon [Citrullus lanatus v. lanatus (Thunb.) Matsum & Nakai] seed exudates are inhibitory to germination and seedling growth of other plant species. A miniature bioassay experiment that measured proso millet (Panicum miliaceum L.) radicle growth was used to assess the inhibition caused by seed exudates of 125 genotypes of watermelon and related Citrullus species. Exudates of most genotypes were not inhibitory; however, exudates of 53 accessions reduced radicle growth in comparison with the control. In subsequent proso millet radicle growth experiments, genotypes were found to vary in inhibitory potential, and concentration response curves generated using filtered, pasteurized exudates were different among genotypes. Filter-sterilized seed exudates of Citrullus accessions also varied in the level of inhibition in a bioassay that measured their effect on sporangia formation by the watermelon pathogen, Phytophthora capsici. These observations suggest that constituents in Citrullus seed exudates affect organisms in the spermosphere and that the inhibitory potential of seed exudates varies among genotypes. Differences in concentration response curves in the millet bioassay and differences in the relative inhibition of genotypes in the millet and fungus bioassays indicate that the inhibitory constituents in seed exudates vary among genotypes.

Inhibitory Constituents of Aldose Reductase in the Fruiting Body of Phellinus linteus

In an effort to characterize active principles for diabetic complication from medicinal mushroom, aldose reductase inhibitors were isolated from the fruiting body of Phellinus linteus and identified as hispidin (5), phelligridimer A (6), davallialactone (7), methyldavallialactone (8), hypholomine B (9), interfungins A (10), and inoscavin A (11), together with protocatechuic acid (1), protocatechualdehyde (2), caffeic acid (3), and ellagic acid (4). Their structures were elucidated by spectroscopic analyses. Among them, davallialactone (7), hypholomine B (9), and ellagic acid (4) exhibited potent rat lens aldose reductase and human recombinant aldose reductase inhibitory activity with IC50 values of 0.33, 0.82, 0.63 microM and 0.56, 1.28, 1.37 microM, respectively.

The In Vitro Inhibition of Human Neutrophil Elastase Activity by some Yemeni Medicinal Plants

15 extracts of different polarities (dichloromethane, methanol and aqueous extracts) from 5 Yemeni medicinal plants (Aspilia helianthoides, (Schumach. & Thonn.) Oliv. & Hiern subsp. ciliate (Schumach.) C.D. Adams leaves; Ceropegia rupicola, Defl. var. rupicola whole plant; Kniphofia sumarae, Defler whole plant; Pavetta longiflora, Vahl. subsp. longifloraleaves; and Plectranthus cf barbatus (Thulin & Gifri) leaves) were tested for their inhibitory effects against the enzymatic activity of human neutrophil elastase (HNE) (EC 3.4.21.37) in an in vitro human neutrophil elastase inhibition assay. 14 extracts at various concentrations were found able to inhibit the activity of HNE. Among the plants tested,A. helianthoides was the most active inhibitor of HNE. The dichloromethane extracts of all tested plants, exhibited more inhibitory effect on HNE activity than the methanolic and aqueous extracts. The dichloromethane extract of A. helianthoides showed the most active inhibitory effect on the HNE activity (IC50 = 0.4μg/ml). Within the HNE inhibitory active methanolic extracts, those of A. helianthoides and P. cf barbatus were the most active inhibitors with IC50 of about 3μg/ml. These results provide some scientific justification for the use of A. helianthoides in traditional medicine and indicate the presence of HNE inhibitory constituents in the active tested plants.

Cytotoxic and DNA topoisomerases I and II inhibitory constituents from the roots ofAralia cordata

Bioactivity-guided fractionation, based on the DNA topoisomerase inhibitory activity, lead to the isolation of five compounds (1-5) from the methylene chloride extract of the roots of Aralia cordata Thunb. (Araliaceae). These compounds were identified as ent-pimara-8(14), 15-dien-19-oic acid (1), ent-pimara-8(14), 15-dien-18-oic acid (2), 16alpha-hydrogen-17-isovaleryloxy-ent-kauran-19-oic acid (3), 16alpha-hydroxy-17-isovaleryloxy-ent-kauran-19-oic acid (4) and dehydrofalcarindiol-8-acetate (5) from their spectral data. Compound 3 was isolated for the first time from this plant, and also showed the strongest inhibition of both DNA topoisomerase I and II activities, with 53 and 96% inhibitions, respectively, at a concentration of 20 microM. However, all the compounds exhibited either weak or no cytotoxicities against the human colon carcinoma cell line (HT-29), the human breast carcinoma cell line (MCF-7) and human hepato blastoma cell line (HepG-2).

Mangiferin Identified in a Screening Study Guided by Neuraminidase Inhibitory Activity

A screening study on neuraminidase inhibitory constituents was carried out, and activity-guided fractionations of three plants, Gouania obtusifolia, Zizyphus cambodiana, and Mangifera odorata, led to the isolation of eleven compounds (1-11). Mangiferin was identified as a significant neuraminidase inhibitor.

Repair of injured fiber tracts in the mammalian central nervous system

When fiber tracts are interrupted after an injury of the brain or spinal cord, the distal parts of the fibers degenerate and regeneration from the proximal stump is absent. Therefore, functional deficits as typically observed e.g. in paraplegia persist throughout life. Recent studies show that an initial attempt for regeneration is blocked by specific neurite growth inhibitory proteins and glycoproteins present in central nervous system (CNS) myelin and in the lesion scar. Antibodies that neutralize one of the most potent inhibitory constituents, Nogo-A, allow long-distance re-growth of injured axons in the rat and macaque spinal cord after injury, as well as enhanced compensatory growth of spared fiber tracts. These animals show a high degree of functional recovery for locomotion or hand function following large, incomplete spinal cord transections. Reagents to inactivate other inhibitory molecules and combined treatments for reduction of inhibition in myelin and scar as well as growth promoting factors will be applied in the future.

Presence of cholinergic and calcium antagonist constituents in Saussurea lappa explains its use in constipation and spasm

This study was carried out to provide a scientific basis for the traditional use of Saussurea lappa, in constipation and spasms. Isolated tissue preparations were used to see if the aqueous-methanol crude extract of the S. lappa root (Sl.Cr) contains gut stimulatory and inhibitory constituents. In isolated guinea-pig ileum, a quiescent preparation, Sl.Cr caused a concentration-dependent (0.3-5.0 mg/mL) spasmogenic effect, with the maximum effect reaching 91% of the acetylcholine maximum. A further increase in concentration caused a declining effect, indicating the presence of spasmolytic constituent(s). The spasmolytic effect was more marked in the spontaneously contracting rabbit jejunum and in the atropinized preparations. The spasmolytic effect was mediated through calcium channel blocking (CCB) activity, as evident by its inhibitory effect against high K(+) (80 mm)-induced contraction and displacement of the Ca(++) concentration-response curves to the right. These data indicate that the crude extract of Sl.Cr contains gut stimulatory constituent(s) of cholinergic-type providing a scientific basis for its use in constipation. The presence of spasmolytic constituents of CCB-type more evident in the spontaneous contracting gut preparation may explain its use in spasms.

Kakkalide and Irisolidone: HMG-CoA Reductase Inhibitors Isolated from the Flower of Pueraria thunbergiana

As part of our search for anti-arteriosclerosis agents from traditional Chinese medicines, the 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase (HCR)-inhibitory constituent, kakkalide, was isolated from the flower of Pueraria thunbergiana (PT, family Leguminosae). The antihyperlipidemic effects of kakkalide and its metabolite, irisolidone, which may be a bioactive form in vivo and potently inhibit the HCR activity, were investigated in vivo. Both the oral and interperitoneal administrations of kakkalide and irisolidone, with the exception of intraperitoneally treated kakkalide, potently lowered the serum levels of total cholesterol (TC) and triglyceride (TG) in Trition WR1339-induced hyperlipidemic mice. The oral administrations of kakkalide and irisolidone in hyperlipidemic mice induced, by the long-term feeding of a high fat diet, also potently reduced the serum levels of TC and TG and epididymal fat pad weight. These findings suggest that PT can improve hyperlipidemia, and the hypolipidemic effect may be due to HMG-CoA reductase.

Chymotrypsin inhibitory constituents from Haloxylon recurvum

Haloxylase, the triglyceride (1), unsaturated fatty acid (2), and saturated fatty acid methyl ester (3) have been isolated from the chloroform-soluble fraction of Haloxylon recurvum. Their structures have been elucidated through spectroscopic studies and chemical transformation. All these compounds showed significant inhibitory activity against the enzyme chymotrypsin in a concentration dependent fashion.

Screening of some Yemeni medicinal plants for inhibitory activity against neutral endopeptidase

Due to the physiological importance of neutral endopeptidase (NEP) in the modulation of nociceptive and pressor responses as well as intestinal secretory mechanism, there is great interest in searching for inhibitors of NEP as novel analgesic, anti-hypertensive, and antidiarrheal agents [1, 2, 3]. The purpose of our study was therefore to screen 15 extracts of different polarity (dichloromethane, methanol, and aqueous extracts) from 5 Yemeni medicinal plants (Aspilia helianthoides leaves, Ceropegia rupicola whole plant, Kniphofia sumarae whole plant, Pavetta longiflora leaves, and Plectranthus cf barbatus leaves) for inhibitory effect against NEP activity. Enzyme activity was determined according to Bormann and Melzig [4]. A series of concentrations (200, 100, 50, 25, 10, and 1µg/ml) of each extract were tested. The remaining activity of the enzyme was calculated as a percentage to the control without inhibitor, considering the influence of the solvent (DMSO as solvent for the dichloromethane and methanol extracts), and test extract. Three independent tests with triplicate parallel samples were performed. All values were expressed as mean±standard deviations. Wilcoxon's U-test was used to test the significance. The IC50 values were obtained from dose-effect curves by linear regression. Out of 15 extracts, 4 extracts (methanol extracts of Ceropegia rupicola, IC50=111µg/ml, Kniphofia sumarae, IC50=141µg/ml and Plectranthus cf barbatus, IC50=139µg/ml and the aqueous extract of Pavetta longiflora IC50=144µg/ml) were found able to inhibit the enzymatic activity of NEP. Although the active plants are used traditionally for other purposes e.g. skin and inflammatory diseases and as haemostatic in Yemen [5], the presence of NEP inhibitory constituents in those plants is supported by reports of related species used in other localities as analgesic, antidiarrheal, and antihypertensive agents, and for the treatment of congestive heart failure (6, 7,8).

Bioactive Compounds from Tobacco as Inhibitors of Neuronal Nitric Oxide Synthase

Tobacco contains thousands of bioactive compounds which may act as inhibitors of specific enzymes. Neuronal nitric oxide synthase (nNOS), a flavoenzyme, becomes dysregulated during ischemia. Thus, selective nNOS inhibitors are needed to improve treatment for stroke and head injury. Studies were initiated using 2,3,6-trimethyl-1,4-napthoquinone (TMN), a tobacco-derived flavoprotein inhibitor (Khalil et al., 2003), and the structurally similar 2-methyl-1,4-naphthoquinone (menadione) to evaluate their effects on nNOS activity. Both compounds inhibited the conversion of [14C] L-arginine to [14C] L-citrulline as well as NADPH oxidation in concentration-dependent manners. In parallel studies, several tobacco varieties were fractionated based on lipophilicity and tested for nNOS inhibition. Inhibitory potency varied between fractions in the rank order: hexane > methanol > aqueous. The highest potency hexane extract (TI-1565) was sub-fractionated to characterize the compound(s) causing nNOS inhibition. Results indicate that: the nNOS inhibitory constituent(s) from TI-1565 is lipophilic, non-polar, non-volatile and may contain copper, and multiple compounds possessing nNOS inhibitory activity are present in tobacco to different extents depending on variety as well as method of extraction. (Supported by KSEF 02-239-RDE-002 & NIEHS ES-011982 to RTM)

Cytotoxic and COX-2 inhibitory constituents from the aerial parts ofAralia cordata

Three diterpenes (1, 8, and 9), three triterpenes (3, 4, and 7), one saponin (11), four sterols (2, 5, 6, and 12), and one cerebroside (10) were isolated from the EtOH extract of the aerial parts of Aralia cordata by repeated silica gel column chromatography. Their chemical structures were identified by comparing their physicochemical and spectral data with those published in literatures. All isolated compounds were evaluated for their cytotoxicity against L1210, K562, and LLC tumor cell lines using MTT assay. Of which, 3beta,5alpha-dihydroxy-6beta-methoxyergosta-7,22-diene (6) showed a potent cytotoxicity against all cell lines with IC50 values of 11.7, 11.9, and 15.1 microM, respectively, while compounds 1, 5, and 11 showed a moderate or weak cytotoxicity. These isolates were also examined for their inhibitory activity against COX-1 and COX-2. Although most compounds, except for 2, 10, and 12, showed a strong inhibitory activity against COX-1, they exhibited a moderate or weak inhibitory activity against COX-2.

HPLC assisted chemobiological standardization of α-glucosidase-I enzyme inhibitory constituents from Piper longum Linn-An Indian medicinal plant

Formulations of traditional medicines are usually made up of complex mixture of herbs. However, effective quality control methods in order to select right quality materials are lacking. Though Piper longum is a widely used herb in several Ayurvedic formulations prescribed for various diseases, there is no analytical method in the literature so far which can help in selecting the right quality material with proper proportions of the active ingredients (α-glucosidase-I enzyme inhibitory principles). We employed a systematic bioassay guided fractionation method and isolated pipataline, pellitorine, sesamin, brachystamide B and guineensine as active principles. A reversed-phase high-performance liquid chromatography method was developed to quantify these active principles in the plant material, which can serve as an effective quality control tool. The separation was carried out using a Discovery HS F5 C-18 (ODS) column and the solvent system used was a gradient comprising of ( A) acetonitrile and ( B) water with a flow rate of 1 ml/min. The detection was performed using a PDA detector. Regression equation pertaining to all the bioactive isolates revealed a linear relationship ( r 2 > 0.9995). The detection limits (S/N = 3) ranged from 0.005 to 0.001 μg/ml. Of all the active isolates, sesamin was identified to be present in maximum quantities (0.91%) where as brachystamide B was found in minimum quantity (0.01%).

Comparison of the Lithogenic Risk Factors for First Time and Recurrent Stone-formers

Purpose: The lithogenic risk factors were compared between the first time stone patients and recurrent stone patients according to age and gender. Materials and Methods: We performed stone metabolic studies on first time stone formers (67 men and 42 women) and the recurrent stone formers (40 men and 20 women). We analyzed the groups' excretion differences for the lithogenic and inhibitory constituents such as calcium, uric acid, oxalate, sodium and citrate; we measured volume from the 24-hour urine samples and calcium, uric acid, sodium, potassium, chloride and phosphate from the serum samples. Hypercalciuria, hyperoxaluria, hypocitraturia and a low 24-hour urine volume (<1,500ml) were compared between the two groups according to age and gender. Results: Hypocitraturia was the most common metabolic abnormality in all the groups. The incidence of hypocitraturia was higher in the recurrent stone formers (50.0%) than in the first time stone formers (48.6%), but this was not statistically significant. A low urine volume was shown to have more significant association (p<0.05) for recurrent stone formers (33.3%) compared to the first time stone formers (18.3%). Conclusions: Hypocitraturia was the most common lithogenic risk factor for stone patients. In the recurrent stone formers, a low urine volume is the risk factor that differentiates them from the first time stone formers. (Korean J Urol 2006;47:1093-1098) ꠏꠏꠏꠏꠏꠏꠏꠏꠏꠏꠏꠏꠏꠏꠏꠏꠏꠏꠏꠏ