Increase of beta-endorphin secretion by syringin, an active principle of Eleutherococcus senticosus, to produce antihyperglycemic action in type 1-like diabetic rats.

We employed streptozotocin-induced diabetic rats (STZ-diabetic rats) as type 1 diabetes-like animal models to investigate the mechanism(s) of antihyperglycemic action produced by syringin, an active principle purified from the rhizome and root part S of ELEUTHEROCOCCUS SENTICOSUS (Araliaceae). Bolus intravenous (i. v.) injection of syringin dose-dependently decreased the plasma glucose of STZ-diabetic rats in 30 minutes in a way parallel to the increase of plasma beta-endorphin-like immunoreactivity (BER). Syringin enhanced BER release from the isolated adrenal medulla of STZ-diabetic rats in a concentration-dependent manner from 0.001 to 10 micromol/l. Bilateral adrenalectomy in STZ-diabetic rats eliminated the activities of syringin (1 mg/kg, i. v.) including the plasma glucose-lowering effect and the plasma BER-elevating effect. Also, syringin failed to lower plasma glucose in the presence of micro-opioid receptor antagonists and/or in the micro-opioid receptor knockout diabetic mice. In conclusion, the obtained results suggest that syringin can enhance the secretion of beta-endorphin from adrenal medulla to stimulate peripheral micro-opioid receptors resulting in a decrease of plasma glucose in diabetic rats lacking insulin.

Stimulatory effect of paeoniflorin on the release of noradrenaline from ileal synaptosomes of guinea-pig in-vitro.

The effect of paeoniflorin (an active principle of Paeoniae Radix, commonly used in traditional Chinese medicine) on the release of noradrenaline (norepineprhine) from nerve terminals was investigated using guinea-pig isolated ileal synaptosomes. Release was determined as the amount of noradrenaline, quantified by high-performance liquid chromatography-electrochemical detection, from samples incubated with paeoniflorin or vehicle. Paeoniflorin stimulated the release of noradrenaline in a concentration-dependent manner without an effect on the level of lactate dehydrogenase in the bathing medium. Tetrodotoxin abolished the action of paeoniflorin at concentrations sufficient to block sodium channels. The depolarizing effect of paeoniflorin on the membrane potential was also illustrated by a concentration-dependent increase in the fluorescence of bisoxonol. Moreover, the effect of paeoniflorin on bisoxonol fluorescence in ileal synaptosomes seems more potent than that of 4-aminopyridine. That paeoniflorin causes influx of calcium ions via the depolarization of nerve terminals could be considered. The noradrenaline-releasing action of paeoniflorin was abolished by removal of calcium chloride from the bathing medium. This action of paeoniflorin was also attenuated by Rp-cAMP atconcentrations sufficientto inhibitthe action of cyclicAMP. Therefore, paeoniflorin could induce a calcium-dependent and cyclic-AMP-related release of noradrenaline from sympathetic nerve terminals of guinea-pig ileum. Guanethidine inhibited the noradrenaline-releasing action of paeoniflorin in a concentration-dependent manner. The effect of paeoniflorin on the increase of bisoxonol fluorescence was not modified by atropine. Release of noradrenaline by paeoniflorin from noradrenergic nerve terminals was characterized. These findings suggest that paeoniflorin can stimulate tetrodotoxin-sensitive depolarization of membranes to result in a calcium-dependent and cyclic-AMP-related release of noradrenaline from noradrenergic nerve terminals.

A combined solid-state and submerged cultivation integrated with adsorptive product extraction for production of Monascus red pigments.

A process taking advantages of combined solid-state and submerged cultivation of Monascus for red pigment production and integration of a product removal unit was developed. The solid-state cultivation was carried out in a 5 l fermentor, with rice being used not only as the substrate but also the support for Monascus. The inclusion of rice submergence and integration of product separation were achieved by intermittently rinsing the rice with monosodium glutamate (MSG) solutions every 12 h followed by an adsorptive extraction of the red pigment dissolved in the rinsing solution. With this new process, the Monascus red pigment production was increased by 24% as compared with that by the plain fixed-bed cultivation.

The in vitro inhibitory effect of tannin derivatives on 3-hydroxy-3-methylglutaryl-coenzyme a reductase on vero cells.

Coronary heart disease is still the major cause of death in industrialized countries. Multiple primary or secondary interventional trials to lower serum cholesterol in humans have resulted in significant reduction of coronary events and death, one of the major reasons attributed to developing a 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitor such as pravastatin. Developing new inhibitors of cholesterol synthesis is still common in the pharmaceutical industry. Tannin comprises a large group of natural polyphenolic compounds possessing antioxidant effects. The methods for analysis of specific inhibitors of mevalonate biosynthesis have already been well established by using Vero cells, a cell line obtained from kidneys of African green monkeys. Tannin derivatives isolated from different traditional Chinese herbs were dissolved in DMSO and incubated with Vero cells with or without the addition of 1 mmol/l mevalonate or 5 mmol/l sodium acetate for 24 h in order to observe cell growth. Pravastatin, a specific HMG-CoA reductase inhibitor, was used as positive control which could inhibit Vero cells growth effectively and cell growth inhibition was reversible after adding 1 mmol/l mevalonate. More than 50 tannin derivatives were used for the study, but only two compounds - proanthrocyanidin A-2 (belonging to the flavan-3-ol group) and 1,2,3,6-tetra-O-galloyl-beta-D-glucose (belonging to the gallotannin group) - showed significant growth inhibition of Vero cells. This study showed that some isolated tannin derivatives from traditional herbs were effective HMG-CoA reductase inhibitors which might be developed into new hypocholesterolemic agents.

Palladium film decoupler for amperometric detection in electrophoresis chips.

Demonstrated in this article is that a palladium metal film can be applied to decouple the electric circuitry of electrochemical detection from that of the electrophoretic separation in an electrophoresis chip. The Pd solid-state field decoupler, as well as the working electrodes, is thermally evaporated onto the plastic chip and oriented vertically across the separation channel. After the sample zones flow over the Pd decoupler, their electrochemical response is measured at working electrodes in the downstream pathway. Because the electrodes are on the separation channel, the electrode channel alignment is no longer a problem. For a separation channel of roughly 200 microm in width and 75 microm in depth in 10 mM phosphate (pH 5.1), the noise level at the working electrode is < 15 pA at an electric field of 570 V/cm.

Caffeic acid as active principle from the fruit of Xanthium strumarium to lower plasma glucose in diabetic rats.

The antihyperglycemic effect of caffeic acid, one of the phenolic compounds contained in the fruit of Xanthium strumarium, was investigated. After an intravenous injection of caffeic acid into diabetic rats of both streptozotocin-induced and insulin-resistant models, a dose-dependent decrease of plasma glucose was observed. However, a similar effect was not produced in normal rats. An insulin-independent action of caffeic acid can thus be considered. Otherwise, this compound reduced the elevation of plasma glucose level in insulin-resistant rats receiving a glucose challenge test. Also, glucose uptake into the isolated adipocytes was raised by caffeic acid in a concentration-dependent manner. Increase of glucose utilization by caffeic acid seems to be responsible for the lowering of plasma glucose.

Antihyperglycemic action of isoferulic acid in streptozotocin-induced diabetic rats.

Wistar rats with streptozotocin-induced diabetes (STZ-diabetic rats), which is similar to human insulin-dependent diabetic mellitus (IDDM), were employed to investigate the antihyperglycemic action of isoferulic acid. A single intravenous injection of isoferulic acid decreased the plasma glucose in a dose-dependent manner in the STZ-diabetic rats. Repeated intravenous administration of STZ-diabetic rats with isoferulic acid (5.0 mg kg(-1)) also resulted in the lowering of plasma glucose after one day. Stimulatory effects of isoferulic acid on the glucose uptake and glycogen synthesis in soleus muscles isolated from STZ-diabetic rats were also obtained indicating an increase of glucose utilization following isoferulic acid treatment which was not dependent on insulin. The mRNA level of glucose transporter subtype 4 form (GLUT4) in soleus muscle was raised by isoferulic acid after repeated treatment for 1 day in STZ-diabetic rats. Similar repeated treatment with isoferulic acid reversed the elevated mRNA level of phosphoenolpyruvate carboxykinase (PEPCK) in liver of STZ-diabetic rats to the normal level. However, expression of GLUT4 and PEPCK genes in nondiabetic rats were not influenced by similar treatment with isoferulic acid. These results suggest that isoferulic acid can inhibit hepatic gluconeogenesis and/or increase the glucose utilization in peripheral tissue to lower plasma glucose in diabetic rats lacking insulin.

Paeoniflorin reverses guanethidine-induced hypotension via activation of central adenosine A1 receptors in Wistar rats.

1. Intravenous injection of paeoniflorin, a glycoside purified from the root of Paeonia lactiflora, reversed guanethidine-induced hypotension in Wistar rats. 2. Pretreatment with the adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine inhibited this effect of paeoniflorin in a dose-dependent manner. 3. The action of paeoniflorin was not modified by 8-(p-sulfophenyl)theophylline, the polar antagonist of the adenosine A1 receptor, which is not able to enter the central nervous system. 4. We conclude that paeoniflorin can reverse guanethidine-induced hypotension via activation of adenosine A1 receptors in the brain of Wistar rats.

Isoferulic acid as active principle from the rhizoma of Cimicifuga dahurica to lower plasma glucose in diabetic rats.

Isoferulic acid extracted from the rhizome of Cimicifuga dahurica Maxim. (Ranunculaceae) has been determined to have in vivo antihyperglycemic activity. An antihyperglycemic action of isoferulic acid in spontaneously diabetic rats, similar to type I diabetes, is presented.

Antioxidant and hepatoprotective activity of punicalagin and punicalin on carbon tetrachloride-induced liver damage in rats.

Punicalagin and punicalin, isolated from the leaves of Terminalia catappa L., are used to treat dermatitis and hepatitis. Both compounds have strong antioxidative activity. The antihepatotoxic activity of punicalagin and punicalin on carbon tetrachloride (CCl4)-induced toxicity in the rat liver was evaluated. Levels of serum glutamate-oxalate-transaminase and glutamate-pyruvate-trans-aminase were increased by administration of CCl4 and reduced by drug treatment. Histological changes around the liver central vein and oxidation damage induced by CCl4 also benefited from drug treatment. The results show that both punicalagin and punicalin have anti-hepatotoxic activity but that the larger dose of punicalin induced liver damage. Thus even if tannins have strong antioxidant activity at very small doses, treatment with a larger dose will induce cell damage.

Stimulatory effect of paeoniflorin on adenosine A-1 receptors to increase the translocation of protein kinase C (PKC) and glucose transporter (GLUT 4) in isolated rat white adipocytes.

In an attempt to understand the subcellular signals after activation of adenosine A-1 receptors, paeoniflorin was employed to incubate with rat white adipocytes in vitro. Translocation of protein kinase C (PKC) beta-subtype from cytosol to membrane was enhanced by an incubation with paeoniflorin in a concentration-dependent manner similar to that of porcine insulin. Also, 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) inhibited this action of paeoniflorin in a concentration-related fashion and it markedly attenuated the action of paeoniflorin at a concentrations sufficient to block the action of adenosine. Moreover, chelerythrine inhibited the paeoniflorin-stimulated translocation of PKC in a way similar to that stimulated by porcine insulin. Subcellular inhibition is considered because stimulation of porcine insulin was not modified by DPCPX at concentrations sufficient to block adenosine A-1 receptors. Similar results were also observed in adipocytes regarding the translocation of glucose transporter (GLUT4) from cytosol to membrane. Thus, we found that paeoniflorin can activate adenosine A-1 receptors to increase the translocations of PKC and GLUT4, two major signals for glucose uptake, from cytosol to membrane of the white adipocytes in rats.

Identification of Anoectochilus formosanus and Anoectochilus koshunensis species with RAPD markers.

RAPD (random amplified polymorphic DNA) markers were developed to distinguish Anoectochilus formosanus from Anoectochilus koshunensis and their putative hybrids. Morphological differentiation of these two species beyond the flowering period is difficult. RAPD markers provide a rapid and easy tool for identification of the two Anoectochilus species. In the study, forty arbitrary decamer primers were screened, and nineteen species-specific RAPD markers generated from polymerase chain reactions (PCR) with eight random primers were obtained. Nine were specific to A. formosanus and ten to A. koshunensis. Two primers, OPC-08 and OPL-07, produced two markers, one specific to A. formosanus and the other specific to A. koshunensis, which simultaneously appeared in the hybrids pattern. The RAPD markers can be applied both to identification of A. formosanus and A. koshunensis species and to assessment of the extent fo hybridization in hybrids between them. This information facilitates the breeding program process.

Medetomidine analogs as alpha 2-adrenergic ligands. 3. Synthesis and biological evaluation of a new series of medetomidine analogs and their potential binding interactions with alpha 2-adrenoceptors involving a "methyl pocket".

The synthesis and the biological evaluation of a new series of medetomidine analogs are reported. The substitution pattern at the phenyl ring of the tetralin analogs had a distinct influence on the alpha 2-adrenoceptor binding affinity. 4-Methylindan analog 6 was the most potent alpha 2-adrenoceptor binding ligand among these 4-substituted imidazoles, and its alpha 2-adrenoceptor selectivity was greater than the 5-methyl tetralin analog 4c. Ligand-pharmacophore and receptor modeling were combined to rationalize alpha 2-adrenoceptor binding data of the imidazole analogs in terms of ligand-receptor interactions. The structure-activity relationships that were apparent from this and previous studies were qualitatively rationalized by the binding site models of the alpha 2-adrenoceptor. The benzylic methyl group of medetomidine or the naphthyl analog 2a was superimposable with the alpha-methyl group of (-)-alpha-methylnorepinephrine and fit into the proposed "methyl pocket" of the alpha 2-adrenoceptor defined by the residues Leu110, Leu169, Phe391, and Thr395.

Antihyperglycemic effects of paeoniflorin and 8-debenzoylpaeoniflorin, glucosides from the root of Paeonia lactiflora.

Paeoniflorin and 8-debenzoylpaeoniflorin were isolated from the dried root of Paeonia lactiflora Pall. (Ranunculaceae). They produced a significant blood sugar lowering effect in streptozotocin-treated rats and had a maximum effect at 25 min after treatment. This hypoglycemic action was also observed in normoglycemic rats only at 1 mg/kg. The antihyperglycemic activity of 8-debenzoylpaeoniflorin seems lower than that of paeoniflorin. Plasma insulin was not changed in paeoniflorin-treated normoglycemic rats indicating an insulin-independent action. Also, this glucoside reduced the elevation of blood sugar in glucose challenged rats. Increase of glucose utilization by paeoniflorin can thus be considered. There are no previous data showing the hypoglycemic activity of paeoniflorin and/or 8-debenzoylpaeoniflorin in rats.

The effects of cantharidin analogues on xanthine oxidase.

Norcantharidin[3], the demethylated product of cantharidin[1] has been used for the treatment of hepatoma, carcinomas of esophagus and gastric cardia, leukopenia and hepatitis. Since the enzyme xanthine oxidase is involved in the diseases mentioned above, and the reactive oxygen species produced by the enzyme induces DNA damage and oxidative damage of tissues, fourteen cantharidin analogues and cantharidimide derivatives were tested for their effects on xanthine oxidase. The results showed that these compounds, listed in Figure 1, displayed very weak inhibitory effects on xanthine oxidase. Contrary to expectation, disodium cantharidate [2], Norcantharidin [3], dehydronorcantharidin [4], disodium dehydronorcantharidate [5], N-(2-pyridyl) cantharidimide [12], N-(3pyridyl) cantharidimide [13] and N-(4-pyridyl) cantharidimide [14] showed a slight stimulating effect on xanthine oxidase at several concentrations.

ADP-mimicking platelet aggregation caused by rugosin E, an ellagitannin isolated from Rosa rugosa Thunb.

Among the nine ellagitannins, rugosin E was the most potent platelet aggregating agent with an EC50 of 1.5 +/- 0.1 microM in rabbit platelets and 3.2 +/- 0.1 microM in human platelets. The aggregations caused by rugosin E and ADP were inhibited by EGTA, PGE1, mepacrine, sodium nitroprusside and neomycin, but not by indomethacin, verapamil, TMB-8, BN52021 and GR32191B. Rugosin E-induced thromboxane formation was suppressed by indomethacin, EGTA, PGE1, verapamil, mepacrine, TMB-8 and neomycin. ADP-scavenging agents, such as CP/CPK and apyrase inhibited concentration-dependently ADP (20 microM)-, but not rugosin E (5 microM)-induced platelet aggregation. In thrombin (0.1 U/ml)-treated and degranulated platelets, rugosin E and ADP still caused 63.5 +/- 3.0% and 61.2 +/- 3.5% of platelet aggregation, respectively. Selective ADP receptor antagonists, ATP and FSBA inhibited rugosin E- and ADP-induced platelet aggregations in a concentration-dependent manner. Both rugosin E and ADP did not induce platelet aggregation in ADP (1 mM)-desensitized platelets. In contrast to ADP, rugosin E did not decrease cAMP formation in washed rabbit platelets. Both rugosin E and ADP did not cause phosphoinositide breakdown in [3H]myo-inositol-labeled rabbit platelets. In fura-2/AM-load platelets, both rugosin E and ADP induced increase in intracellular calcium concentration and these responses were inhibited by ATP and PGE1. All these data suggest that rugosin E may be an ADP receptor agonist in rabbit platelets.

Superoxide anion scavenge effect of Quercus glauca Thunb. in whole blood of patients with ankylosing spondylitis.

Nine phenolic compounds, catechin (1), epicatechin (2), gallocatechin (3), epigallocatechin (4), procyanidin B-4 (5), catechin-3-O-rhamnoside (6), rutin (7), querglanin (8) and isoquerglanin (9) were isolated from oak leaves (Quercus glauca Thunb. Fagaceae), and the latter two (8, 9) were identified as new compounds. Several Quercus species have been used in folk medicine as an astringent for hemorrhoids and for treatment of inflammation, jaundice, and tumor. In this study, these compounds were tested for scavenging effects of the superoxide anion in the whole blood of patients with ankylosing spondylitis by means of an ultra-sensitive chemoluminescence (CL) analyzer and lucigenin amplification. The results showed that at a concentration of 2.3 x 10(-5) M, isoquerglanin (9) displayed the strongest inhibition activity (73.55%), followed by querglanin (8) (68.81%) and then gallocatechin (3) and epigallocatechin (4) (66.97 and 60.17% inhibition, respectively). In addition, the blood chemoluminescence (CL) level of patients with ankylosing spondylitis was inhibited by superoxide dismutase (SOD) but not by catalase, suggesting that superoxide anion is the major component of reactive oxygen species (ROS) involved in this assay system.

In vitro inhibitory effects of chebulinic acid on the contractile responses of cardiovascular muscles.

1. The effects of chebulinic acid, which has been shown to elicit blood pressure lowering effect in rats, on aortic vascular contraction as well as cardiac contraction were studied in rats. 2. Chebulinic acid had no effect on KCl-induced aortic contraction, but irreversibly inhibited the contractile responses to phenylephrine in an apparently non-competitive manner. Chebulinic acid also inhibited contractile responses of rat aorta to 5-hydroxytryptamine and angiotensin II. 3. Chebulinic acid inhibited the binding of [3H]-prazosin to dog aortic microsomal membranes in a concentration-dependent manner with an IC50 value of 0.34 mmol/L. Results of saturation binding experiments suggest a mixed mode of inhibition by chebulinic acid (i.e. a decrease in both the maximal number of binding sites and the affinity for prazosin). 4. Chebulinic acid concentration-dependently and reversibly inhibited the maximal left ventricular pressure of rat heart in a Langendorff preparation with 50% inhibition occurring at a concentration of 0.3 nmol/L. 5. We conclude that chebulinic acid exerts non-specific inhibitory actions in vascular preparations. Its inhibitory effect on cardiac contraction was reversible and three orders of magnitude more potent than that on vascular contraction. We suggest that the hypotensive effect of chebulinic acid is probably mediated via the decrease in cardiac output resulting from reduced left ventricular contraction.

Medetomidine analogs as alpha 2-adrenergic ligands. 2. Design, synthesis, and biological activity of conformationally restricted naphthalene derivatives of medetomidine.

A new series of naphthalene analogs of medetomidine have been prepared and evaluated for their alpha-adrenergic activities. The methylnaphthyl analog 5a showed significant selectivity for alpha 2-adrenoceptors and behaved as a partial alpha 1-agonist in rat aorta preparations. In contrast, the Z-ethylene analog 8c was alpha 1-selective and behaved as a potent alpha 1-antagonist. Two rigid analogs (6 and 7) exhibited large differences in binding affinities at alpha 1-VS alpha 2-receptors, indicating that the conformational flexibility of 5a is important for the fulfillment of the alpha-adrenergic activities. Molecular modeling studies began with conformational analysis of classical phenethylamines and medetomidine analogs. Superimposition of medetomidine conformations with those of phenethylamines provided a tentative explanation for the alpha 2-adrenergic activity of the new imidazoles. A common binding mode for phenethylamines and imidazoles with alpha 2-adrenoceptors is proposed. Knowledge of the biological properties of the 4-substituted imidazoles, integrated with the information derived from computer-assisted molecular modeling, has provided new insights for the structural and conformational requirements of this class as new adrenergic drugs.