Uptake and accumulation and oxidative stress in garlic (Allium sativum L.) under lead phytotoxicity

The effects of different concentrations of Pb on growth of Allium sativum L, Pb uptake and accumulation, antioxidant enzyme activity and malondialdehyde content were investigated. The results indicated that shoot growth at high concentration of Pb (10⁻³ M) and roots growth at 10⁻³ M and 10⁻⁴ M Pb were significantly inhibited. Lead ions were accumulated mainly in the roots and only small amounts were translocated to bulbs and shoots. SOD activities in shoot and roots exposed to 10⁻³ M Pb were observed to be high. Plants exposed to 10⁻³ M Pb showed a significant increase in POD activity in roots versus the control and other Pb treatments. In roots, CAT activity and MDA concentration at 10⁻³ M Pb is high significantly. The mechanisms of Pb toxicity and tolerance in garlic are briefly discussed.     

Inhibitory effects of L- and T-type calcium antagonists on contractions of human detrusor muscle

The inhibitory and relaxant effects of the L-type calcium antagonists nifedipine, nimodipine, verapamil and diltiazem, and of the T-type calcium antagonist mibefradil, on contractions of isolated human detrusor muscle were investigated. The tissue was obtained from 10 patients undergoing cystectomy due to bladder cancer. Effects of the calcium antagonists at different concentrations on the concentration-response curves for carbachol were investigated. Furthermore, concentration-relaxation curves were performed using potassium-precontracted muscle strips. All L-type calcium antagonists suppressed the mean concentration-response curve of carbachol significantly at a concentration of 10⁻⁶ M. Mibefradil up to 10⁻⁵ M did not significantly suppress it. Nifedipine significantly reduced the carbachol-induced maximum contraction to 75% and 44%, verapamil to 75% and 67% of the appropriate control value at concentrations of 10⁻⁷ and 10⁻⁶ M, respectively. Diltiazem reduced it insignificantly to 96% and 71% at the above-mentioned concentrations. The concentration-relaxation experiments revealed following pD2-values and maximum relaxations of nifedipine, nimodipine, verapamil and diltiazem, respectively: 6.23, 6.37, 5.66, 5.81 and 85%, 83%, 82%, 90%. Maximum relaxations and pD2-values were not significantly different from each other. The lowest concentration, for which a significant effect compared to control in Student`s t-test was found, amounted to 10⁻¹⁰ M, 10⁻⁹ M, 10⁻⁷ M, 10^−6.5 M and 10⁻⁴ M for nimodipine, nifedipine, diltiazem, verapamil and mibefradil, respectively. L-type calcium antagonists are very potent relaxant agents of the human detrusor muscle in vitro.     

In vitro studies of human alcohol dehydrogenase inhibition in the process of methanol and ethylene glycol oxidation

The liver is the major organ responsible for methanol and ethylene glycol oxidation, and alcohol dehydrogenase (alcohol: NAD⁺ oxidoreductase, EC 1.1.1.1.) is the main enzyme involved. In the present study, alcohol dehydrogenase (ADH) activity was measured spectrophotometrically in vitro at physiological pH 7.4 and 37 °C using human enzyme hepatic fraction. The percentage of residual activity was calculated for four inhibitors at concentrations of 10⁻³, 10⁻⁴, and 10⁻⁵ M (pyrazole, 4-methylpyrazole, cimetidine, theophylline) and methylene blue at concentrations 10⁻⁴ and 10⁻⁵ M. Our results have shown that the best inhibitor, cimetidine, decreased oxidation of 0.1 M and 0.05 M methanol to 24 and 29% respectively at a drug concentration of 1 mM. Reaction with 0.1 M ethylene glycol as the ADH substrate was blocked by the same substances and 4-methylpyrazole was found to be a highly effective inhibitor. Received: 3 November 1997 / Accepted: 5 May 1998     

First-pass metabolism of decursin, a bioactive compound of Angelica gigas, in rats

Decursin is considered the major bioactive compound of Angelica gigas roots, a popular Oriental herb and dietary supplement. In this study, the pharmacokinetics of decursin and its active metabolite, decursinol, were evaluated after the administration of decursin in rats. The plasma concentration of decursin decreased rapidly, with an initial half-life of 0.05 h. It was not detectable at 1 h after intravenous administration at an area under the plasma concentration-time curve of 1.20 μg · mL-1·h, whereas the concentration of decursinol increased rapidly reaching a maximum concentration of 2.48 μg · mL-1 at the time to maximum plasma concentration of 0.25 h and an area under the plasma concentration-time curve of 5.23 μg · mL-1·h. Interestingly, after oral administration of decursin, only decursinol was present in plasma, suggesting an extensive hepatic first-pass metabolism of decursin. The extremely low bioavailability of decursin after its administration via the hepatic portal vein (the fraction of dose escaping first-pass elimination in the liver, FH = 0.11) is indicative of extensive hepatic first-pass metabolism of decursin, which was confirmed by a tissue distribution study. These findings suggest that decursin is not directly associated with the bioactivity of A.?gigas and that it may work as a type of natural prodrug of decursinol.     

Radical scavenging capacities of some thiazolylthiazolidine-2,4-dione derivatives

In this study, a series of 2,4-dichlorothiazolyl thiazolidine-2,4-dione (Ia-f) and 4-chloro-2-benzylsulfanylthiazolyl-thiazolidine-2,4-dione derivatives (IIa-f) were tested for their antioxidant properties by determining their effects on superoxide anion formation, and the 2,2-diphenyl-1-picrylhydrazyl (DPPH) stable free radical. Compound Ic showed the best superoxide anion scavenging activity at the 10⁻³ M and 10⁻⁴ M concentrations. The compounds (Ia-f and IIa-f) had the strong DPPH radical scavenger capacity at the 10⁻³ M concentration. Compound Ib, Id, and Ie showed similar activity to butylated hydroxytoluene (BHT) at 10⁻³ M and 10⁻⁴ M concentrations.     

Assay of nitrofuran drugs using an amperometric monoamine oxidase biosensor

A method for assaying nitrofuran drugs (furazolidone, furadonine, and furagin) using an amperometric biosensor based on a printed platinum electrode and immobilized monoamine oxidase was developed for analysis of medicinal formulations and urine. Nitrofuran derivatives were found to have inhibitory actions, albeit weaker than those of classical tricyclic antidepressants, on the catalytic activity of monoamine oxidase. This side-effect inhibitory action allowed assay of furazolidone, furadonine, and furagin with lower detection limits, which were 8.3 × 10⁻⁹, 8.5 × 10⁻⁸, and 9.4 × 10⁻¹⁰ M respectively. The results obtained here show that this side effect of furazolidone, furadonine, and furagin must be taken into consideration when these agents are prescribed to patients.     

Decursinol and decursin protect primary cultured rat cortical cells from glutamate-induced neurotoxicity

We previously reported six neuroprotective decursinol derivatives, coumarins from Angelica gigas (Umbelliferae) roots. To elucidate the action patterns of decursinol derivatives, we investigated the neuroprotective effects of decursinol and decursin, which showed highly significant activity and were major constituents of A. gigas, using primary cultures of rat cortical cells in-vitro. At concentrations of 0.1-10.0 microM, both decursinol and decursin exerted a significant neuroprotective activity pretreatment and throughout treatment. In addition, decursin had a neuroprotective impact in the post-treatment paradigm implying that decursin might possess different action mechanisms from that of decursinol in the protection of neurons against glutamate injury. Both decursinol and decursin effectively reduced the glutamate-induced increased intracellular calcium ([Ca(2+)](i)) in cortical cells, suggesting that these two coumarins may exert neuroprotection by reducing calcium influx by overactivation of glutamate receptors. This suggestion was supported by the result that decursinol and decursin protected neurons against kainic acid (KA)-induced neurotoxicity better than against that induced by N-methyl-D-aspartate (NMDA). Moreover, both decursinol and decursin significantly prevented glutamate-induced decreases in glutathione, a cellular antioxidant, and glutathione peroxidase activity. In addition, both compounds efficiently reduced the overproduction of cellular peroxide in glutamate-injured cortical cells. These results suggested that both decursinol and decursin protected primary cultured rat cortical cells against glutamate-induced oxidative stress by both reducing calcium influx and acting on the cellular antioxidative defence system. Moreover, decursin is considered to probably have a different action mechanism from that of decursinol in protecting cortical cells against glutamate injury.     

Inhibitory effect of papaverine on the contraction induced by transmural stimulation in the isolated mouse vas deferens

The effect of papaverine on the contractions induced by adrenergic neurotransmission in the isolated mouse vas deferens was investigated. Papaverine, 10⁻⁷–10⁻⁵M, showed a dose-dependent and reversible inhibition on the induced contractions. When the frequency of stimulation was varied (2.5–20.0 Hz), the inhibitory effect tended to be marked at the lower frequencies.     

Antibacterial coumarins from<Emphasis Type="Italic">Angelica gigas</Emphasis> roots

Systematic fractionation of Angelica gigas roots led to the isolation of linear furano(pyrano)coumarins such as bergapten (1), decursinol angelate (2), decursin (3), nodakenetin (4) and nodakenin (5). The antibacterial activities of those compounds against pathogenic bacteria were investigated. Among the compounds tested, decursinol angelate (2) and decursin (3) exhibited significant antibacterial activity against Bacillus subtilis with the minimum inhibitory concentrations (MICs) of 50 and 12.5 microg/mL, respectively.     

An in vitro approach for demonstrating the critical role of serum albumin in the detoxication of the carbamate carbaryl at in vivo toxicologically relevant concentrations

The hydrolysis of carbaryl by bovine serum albumin (BSA) was studied at toxicologically relevant concentrations (range 15–300 μM) in order to determine the role of this protein in the detoxication of the carbamate in vivo. The 1-naphthol released during the hydrolysis of carbaryl was monitored using gas chromatography coupled with mass spectrometry. BSA hydrolyzed carbaryl in a time-progressive way. The hydrolysis was also dependent of enzyme (1.0, 2.5, 5.0 and 7.0 mg ml⁻¹) and substrate (range between 15 and 1,000 μM) concentration. The estimated turnover number and Michaelis–Menten constant were 1.6 × 10⁻⁴ s⁻¹ and 430 μM, respectively. Thus, the second order rate constant was 0.37 M⁻¹ s⁻¹. At enzyme concentrations of 7.0 mg ml⁻¹ and substrate concentrations ranging between 50 and 300 μM about 80% of substrate was hydrolyzed in 3 h. At lower substrate concentrations (15 and 30 μM carbaryl) also significant hydrolysis was detected at the highest enzyme concentration, even when these substrate concentrations were 30 and 15 times lower than the Michaelis–Menten constant. Although the efficacy of the enzymatic hydrolysis is low, the extrapolation of our results to the physiological albumin high concentrations (around 40 mg ml⁻¹) suggests that the hydrolysis of carbaryl by serum albumins plays a critical role in the detoxication of this carbamate at in vivo toxicologically relevant concentrations. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Platelet anti-aggregatory effects of coumarins from the roots of<Emphasis Type="Italic">Angelica genuflexa</Emphasis> and<Emphasis Type="Italic">A. gigas</Emphasis>

Five coumarins, isoimperatorin (1), pabulenol (2), isooxypeucedanin (3), oxypeucedanin hydrate (4) and osthol (5) were isolated from the MeOH extract of Angelica genuflexa in the course of searching for anti-platelet and anti-coagulant components from plants. Pabulenol (2) was isolated from A. genuflexa for the first time. The five compounds isolated from A. genuflexa, together with decursinol angelate (6), decursin (7) and nodakenin (8) from A. gigas were evaluated for their effects on platelet aggregation and blood coagulation. Compounds 2, 5, 6 and 7 were observed to be either equally effective or 2-4 times more inhibitory than ASA in both arachidonic acid and U46619 (TXA2 mimetic) induced platelet aggregations.     

In Vitro inhibitory potential of decursin and decursinol angelate on the catalytic activity of cytochrome P-450 1A1/2, 2D15, and 3A12 isoforms in canine hepatic microsomes

Danggui is one of the most popular herbal medicines consumed by patients in different clinical settings in Asian countries. In this study, the two major pyranocoumarin compounds extracted from the Korean Angelica gigas root decursin (DC) and decursinol angelate (DA) were examined in vitro with regard to their abilities to inhibit hepatic CYP1A1/2, CYP2D15, and CYP3A12 catalytic activities in canine liver microsomes. The two components were capable of inhibiting CYP1A1/2, CYP2D15, and CYP3A12 catalytic activities, but the potencies varied. DC and DA selectively and noncompetitively inhibited CYP1A1/2 activity, with K _i values of 90.176 and 67.560 μM, respectively. On the other hand, they exhibited slight inhibitory effects on CYP2D15 and CYP3A12 with K _i values of 666.180 and 872.502 μM, 990.500 and 909.120 μM (1’hydroxymidazolam, MDZ1’H), and 802.800 and 853.920 μM (4-hydroxymidazolam, MDZ4H), respectively. Additionally, they showed increased inhibition after preincubation, which suggests the involvement of a mechanism-based inhibition. In sum, this in vitro data should be heeded as a signal of possible in vivo interactions. The use of human liver preparations would considerably strengthen the practical impact of the data generated from this study. The two authors contributed equally to this paper.     

Biological effects of the dihydroorotate dehydrogenase inhibitor polyporic acid, a toxic constituent of the mushroom Hapalopilus rutilans , in rats and humans

Inhibitors of dihydroorotate dehydrogenase (DHO-DH), such as brequinar or leflunomide, have been intensively tested for their antitumour and immunomodulating effects. Polyporic acid (PA) from the mushroom Hapalopilus rutilans (H. r.) also is a DHO-DH inhibitor (50% inhibitory concn., IC₅₀, 10⁻⁴–10⁻³ M). As three people had been poisoned following ingestion of H. r. we wanted to investigate the effects of PA in rats and in cell cultures. Rats given PA via probang (100–800 mg/kg) within 24 h developed strongly reduced locomotor activity, depressed visual placing response and impaired wire manoeuvre. Laboratory investigation of blood revealed hepatorenal failure, metabolic acidosis as well as hypokalaemia and hypocalcaemia. All symptoms closely paralleled the effects seen in the poisoned people. Proliferation of cultured cells (including rat brain neurons and glia, fibroblasts, tumour cells) was depressed at 10⁻⁴–10⁻³ M PA. We conclude that the intoxication of people poisoned with H. r. is due to the high content of the DHO-DH inhibitor PA. Received: 6 July 1998 / Accepted: 7 September 1998     

Urotensin II acutely increases myocardial length and distensibility: potential implications for diastolic function and ventricular remodeling

Urotensin II (U-II) is a cyclic peptide that may be involved in cardiovascular dysfunction. In the present study, the acute effects of U-II on diastolic properties of the myocardium were investigated. Increasing concentrations of U-II (10⁻⁸ to 10⁻⁶ M) were added to rabbit papillary muscles in the absence (n = 15) or presence of: (1) damaged endocardial endothelium (EE; n = 9); (2) U-II receptor antagonist, urantide (10⁻⁵ M; n = 7); (3) nitric oxide (NO) synthase inhibitor, N^G-Nitro-l-Arginine (10⁻⁵ M; n = 9); (4) cyclooxygenase inhibitor, indomethacin (10⁻⁵ M; n = 8); (5) NO synthase and cyclooxygenase inhibitors, N^G-Nitro-l-Arginine (10⁻⁵ M) and indomethacin (10⁻⁵ M), respectively, (n = 8); or (6) protein kinase C (PKC) inhibitor, chelerythrine (10⁻⁵ M; n = 9). Passive length–tension relations were constructed before and after a single concentration of U-II (10⁻⁶ M; n = 3). U-II concentration dependently decreased inotropy and increased resting muscle length (RL). At 10⁻⁶ M, active tension decreased 13.8 ± 5.4%, and RL increased to 1.007 ± 0.001 L/L _max. Correcting RL to its initial value resulted in an 18.1 ± 3.0% decrease in resting tension, indicating decreased muscle stiffness, which was also suggested by the down and rightward shift of the passive length–tension relation. This effect remained unaffected by EE damage and PKC inhibition. In contrast, the presence of urantide and NO inhibition abolished the effects of U-II on myocardial stiffness, while cyclooxygenase inhibition significantly attenuated them. U-II decreases myocardial stiffness, an effect that is mediated by the urotensin-II receptor, NO, and prostaglandins. This represents a novel mechanism of acute neurohumoral modulation of diastolic function, suggesting that U-II is an important regulator of cardiac filling. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. Presented in part at the American Heart Association Scientific Sessions conference, 2006, in Chicago, Illinois.     

Effect of heavy metal exposure on blood haemoglobin concentration and methemoglobin percentage in Lumbricus terrestris

The earthworm haemoglobin (Hb) is a large extracellular hemoprotein flowing in a closed circulatory system. In spite of the fundamental role of this respiratory pigment in earthworm physiology, little is known about its sensitivity to environmental pollutants. The aim of the present work was to investigate the possible effect of heavy metal (cadmium, copper, mercury) exposure on Hb concentration and oxidation state (methemoglobin formation) in the earthworm Lumbricus terrestris. In addition, the tissue concentration of metallothioneins, a well-known biomarker of heavy metal exposure, was determined as an indicator of metal uptake. The animals were exposed to increasing concentrations of Cd, Cu and Hg utilizing the standard acute toxicity test, “Filter paper test” for 48 h. Exposure to heavy metals (10⁻⁵–10⁻³ M for Cd, 10⁻⁴–10⁻³ M for Hg, and 10⁻⁴–10⁻² M for Cu) was found to increase haemoglobin concentration in L. terrestris, although the magnitude of such an increase was dependent on the metal. In addition, metal exposure led to the formation of methemoglobin. Compared to other known biological responses to heavy metals, such as metallothionein induction, methemoglobin increase showed a higher sensitivity and a higher percentage variation in exposed organisms, showing to be a possible suitable biomarker of exposure/effect to be included in a multi biomarker strategy in earthworm in soil monitoring assessment.     

β-secretase (BACE1) inhibitors from <Emphasis Type="Italic">Perilla frutescens</Emphasis> var. <Emphasis Type="Italic">acuta</Emphasis>

In the course of screening for anti-dementia agents from natural products, two β-secretase (BACE1) inhibitors were isolated from the methanolic extract of Perilla frutescens var. acuta and identified as luteolin (1) and rosmarinic acid (2) with IC₅₀ values of 5.0×10⁻⁷ M and 2.1×10⁻⁵ M, respectively. They inhibited BACE1 in a non-competitive manner with a substrate in Dixon plots, suggesting that they might bind to either β-secretase subsite or to another regulatory site. Ki values of 1 and 2 were 6.2×10⁻⁵ M and 3.9×10⁻⁵ M, respectively. They were less inhibitory against other enzymes such as α-secretase (TACE), acetylcholine esterase (AchE), chymotrypsin, and elastase, indicating that they were relatively specific inhibitors of BACE1.     

Effect ofAlpha-2 adrenergic agonist onBeta adrenoceptor-mediated control of blood glucose in the fasted mouse

Dose-dependent increases in blood glucose were produced by epinephrine and clonidine in fasted male mice. Isoproterenol was ineffective in increasing blood glucose at lower doses (10⁻⁸ M/kg−10⁻⁷ M/kg); with higher dose (10⁻⁶ M/kg) the glucose level was increased. The hyperglycemia induced by epinephrine was inhibited by yohimbine, prazosin and propranolol, indicating that the hyperglycemic effect of epinephrine is mediated byalpha-1,alpha-2 andbeta adrenoceptor. When clonidine (10⁻⁶ M/kg) was administered simultaneously with isoproterenol (10⁻⁶ M/kg), an enhenced hyperglycemic effect was observed. The increment produced by clonidine plus isoproterenol was higher than that by clonidine alone. These results suggest that stimulation ofalpha-2 adrenoceptor may be responsible for the exertion of the hyperglycemic effect bybeta agonists in fasted mice.     

Fluvastatin reduces endothelin secretion of cultured human umbilical vein endothelial cells

Objective: Fluvastatin is an agent of a new lipid lowering drug class, the 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors, which seems to elicit direct effects on the vasculature. Methods: The effect of fluvastatin on endothelin secretion in endothelial cell cultures from human umbilical veins was investigated. Results: Fluvastatin significantly reduced endothelin secretion by 13% at a concentration of 10⁻⁸ M, by 41% at 10⁻⁷ M and by 62% at 10⁻⁶ M. Conclusion: Since endothelin is a potent vasoconstrictor which may be associated with the aetiology of cardiovascular diseases, the reduction of its synthesis by fluvastatin may contribute to the beneficial effects of this substance on the cardiovascular system. Received: 12 April 1999 / Accepted in revised form: 23 August 1999     

Quantitative determination of decursin, decursinol angelate, and decursinol in mouse plasma and tumor tissue using liquid-liquid extraction and HPLC

The pyranocoumarin compound decursin and its isomer decursinol angelate (DA) are the major hydrophobic phytochemicals in the root of Angelica gigas Nakai (AGN, Korean Angelica), a major traditional medicinal herb. The ethanol extract of AGN and especially the purified decursin and DA have been shown to exhibit antitumor activities by our collaborative team and others. Although decursinol has been identified as a major hydrolysis metabolite of decursin and DA in vivo in previous pharmacokinetic studies with mouse and rat, other recently published results sharply disputed this conclusion. In this study, we set up a practical method for the concurrent analysis of decursin, DA, and decursinol in mouse plasma and tumor tissues by liquid-liquid extraction and HPLC-UV and applied the method to several animal experiments. Plasma or tumor homogenate was extracted directly with ethyl acetate. The extraction efficiency for decursin/DA (quantitated together) and decursinol was between 82-95?% in both mouse plasma and tumor homogenate. The lower limit of quantitation (LLOQ) was approximately 0.25?μg/mL for decursin/DA and 0.2?μg/mL for decursinol in mouse plasma. In a pilot pharmacokinetic study, male C57BL/6 mice were given a single dose of 4.8?mg decursin/DA mixture (~240?mg/kg) per mouse either by oral gavage or intraperitoneal injection. Maximum plasma concentrations for decursin/DA and decursinol were 11.2 and 79.7?μg/mL, respectively, when decursin/DA was administered via intraperitoneal injection, and 0.54 and 14.9?μg/mL via oral gavage. Decursin/DA and decursinol contents in the tumor tissues from nude mouse xenografts correlated very well with those in plasma. Overall, our results confirm the conclusion that the majority of decursin/DA hydrolyze to decursinol in rodent models with a tiny fraction remaining as the intact compounds administered.     

Valve movement response of the mussel Mytilus galloprovincialis to metals (Cu, Hg, Cd and Zn) and phosphate industry effluents from Moroccan Atlantic coast

Valve activity was measured in the Mediterranean mussel Mytilus galloprovincialis in response to sublethal concentrations of four metals (Hg, Cu, Zn and Cd) and two phosphate industry effluents from the Atlantic coast of Morocco. Valve movements were monitored using a proximity inductive sensor which could display all activity figures from full closure to wide opening of the shell valves. In a 1 h exposure experiments, all metals induced a decrease in the time of normal opening and the appearance of sequences of stress behaviour, including enhanced valve adductions and complete closure at high concentrations. Mercury (tested from 5 to 75 μg Hg l⁻¹) was the most toxic to the valve activity, with a threshold effective concentration at 10 μg Hg l⁻¹ and full valve closure occurring at 50 μg Hg l⁻¹. Copper (15–150 μg Cu l⁻¹) showed a toxic effect starting at threshold concentration of 20 μg Cu l⁻¹ and induced full valve closure at 150 μg Cu l⁻¹. Zinc (100–500 μg Zn l⁻¹) was effective in reducing the time of normal opening (threshold concentration at 100 μg Zn l⁻¹) but no complete closure was recorded in any of the tested concentrations. For cadmium (1000–5000 μg Cd l⁻¹), the valve activity was insensitive for exposures under 2000 μg Cd l⁻¹. Results for the testing of several samplings of the phosphate industry effluents (Safi and Jorf Lasfar) showed that their toxicity varied over the time. The effluent of the Jorf Lasfar plant (2–9.4%) was, however, more toxic than that of Safi (1–25%). In the light of these results, the sensitivity of the valve activity of Mytilus galloprovincialis to pollutants and its usefulness for in situ monitoring of coastal pollution in Morocco are discussed.