Effects of selected bioactive natural products on the vascular endothelium.

The endothelium, a highly active structure, regulates vascular homeostasis through the release of numerous vasoactive factors that control vascular tone and vascular smooth cell proliferation. A larger number of medicinal plants and their isolated chemical constituents have been shown to beneficially affect the endothelium. For example, flavonoids in black tea, green tea, and concord grape cause a vasodilation possibly through their antioxidant properties. Allicin, a by-product of the enzyme alliinase, has been proposed to be the main active metabolite and responsible for most of the biological activities of garlic, including a dose-dependent dilation on the isolated coronaries. Thymoquinone, the principal phytochemical compound found in the volatile oil of the black seed, and the hawthorn extract have also been shown to improve aging-related impairment of endothelium-dependent relaxations in animal models. In this review, the effect of some of the natural products, including Camellia sinensis (black tea and green tea), Vitis labrusca (concord grape), Allium sativum (garlic), and Nigella sativa (black seed) and Crataegus ssp (hawthorn extract), is explored. The molecular mechanisms behind these potential therapeutic effects are also discussed.

Effects of Lepidium sativum, Nigella sativa and Trigonella foenum-graceum on phenytoin pharmacokinetics in beagle dogs.

The present work was designed to evaluate the effect of some commonly used herbs viz. garden cress (Lepidium sativum), black seed (Nigella sativa) and fenugreek (Trigonella foenum-graceum) on the disposition of phenytoin after oral administration in a dog model. Phenytoin was given orally at a dose of 50 mg, and blood samples were obtained for the determination of drug's pharmacokinetic parameters. After a suitable washout period, animals were commenced on a specific herb treatment for one week. Garden cress treatment caused a modest increase in maximum observed concentration (Cmax ) and terminal half-life (T1/2λ ) of phenytoin with a reduction in clearance by 33%. The effect of black seed therapy was more drastic on drug elimination and to a lesser extent on its volume of distribution at steady state (Vss ) with a consequent reduction in systemic exposure measured by area under the curve (AUC0-∞ ) by about 87%. The effect of fenugreek therapy resembled, albeit to a lesser extent, that of black seed with a significant reduction in AUC0-∞ by ~72%. In addition, there was a 73% increase in Vss . Our findings suggest that the phenytoin disposition can be significantly altered by the concurrent consumption of tested herbal products.

Hg bioavailability and impact on bacterial communities in a long-term polluted soil.

Different soil samples characterised by a long-term Hg-pollution were studied for Hg total content, fractionation, phytotoxicity and influence on the bacterial community. Hg pollution ranged from 1 to 50 mg kg(-1) and most of it was speciated in scarcely soluble forms. In agreement with this, the biochemical quality indexes were investigated (biomass, enzyme activities) and the bacterial community (viable heterotrophic (VH) bacteria, functional diversity) apparently was not influenced by the degree of Hg pollution. In particular, the investigated soils exhibited a low percentage of Hg-resistant (Hg(R)) bacteria ranging from less than 0.001% to 0.25% of the VH and the addition of available Hg in the form of HgCl(2) induced an enrichment of resistant Hg(R) populations. The general biodiversity of the bacterial community was evaluated by denaturing gradient gel electrophoresis of DNA of Hg spiked soil microcosms and of control soils. Hg(R) bacteria capable to grow in a minimal medium containing HgCl(2) were also isolated and identified. MerA and merB gene PCR fragments were obtained from different Hg(R) strains and the range of similarities at the DNA level and at the deduced amino acid level showed that they carried mercuric reductase and lyase. Differently from bacteria, some influence of soil Hg content on seeds' germination and root elongation was observed for Lepidium sativum L. and Solanum lycopersicum L. In conclusion, most of the Hg in these long-term polluted soils was scarcely mobile and available and did not significantly influence the soil bacterial community. The risk of potential Hg remobilization over time, that could be naturally favoured by the activity of plant roots or other inorganic processes occurring in soil, can be extenuated since bacterial community was resistant and resilient to subsequent Hg stress.

Cardiac performance during exercise in patients with Fabry's disease.

BACKGROUND: Fatigability and dyspnoea on effort are present in many patients with Fabry's disease. We assessed the determinants of cardiac performance during exercise in patients with Fabry's disease and preserved left ventricular ejection fraction at rest. MATERIALS AND METHODS: Sixteen patients with Fabry's disease and 16 control subjects underwent radionuclide angiography at rest and during exercise, tissue Doppler echocardiography and magnetic resonance imaging at rest. RESULTS: The exercise-induced change in stroke volume was +25 +/- 14% in controls and +5.8 +/- 19% in patients with Fabry's disease (P < 0.001). In 10 patients (group 1), the stroke volume increased (+19 +/- 10%), and in 6 patients (group 2) it decreased (-16 +/- 9%) with exercise. Patients of group 2 were older, had worse renal function, higher left ventricular mass and impaired diastolic function compared to group 1. The abnormal stroke volume response to exercise in group 2 was associated with a decrease in end-diastolic volume (P < 0.001) and a lack of reduction of end-systolic volume (P < 0.01) compared with both controls and group 1. The ratio of peak early-diastolic velocity from mitral filling to peak early-diastolic mitral annulus velocity was the only independent predictor of exercise-induced change in stroke volume (B -0.44; SE 0.119; beta-0.70; P < 0.005). CONCLUSIONS: The majority of patients with Fabry's disease were able to augment stroke volume during exercise by increasing end-diastolic volume, whereas patients with more advanced cardiac involvement may experience the inability to increase cardiac output by the Frank Starling mechanism.

Impact of river overflowing on trace element contamination of volcanic soils in south Italy: part II. Soil biological and biochemical properties in relation to trace element speciation.

The effect of heavy metal contamination on biological and biochemical properties of Italian volcanic soils was evaluated in a multidisciplinary study, involving pedoenvironmental, micromorphological, physical, chemical, biological and biochemical analyses. Soils affected by recurring river overflowing, with Cr(III)-contaminated water and sediments, and a non-flooded control soil were analysed for microbial biomass, total and active fungal mycelium, enzyme activities (i.e., FDA hydrolase, dehydrogenase, beta-glucosidase, urease, arylsulphatase, acid phosphatase) and bacterial diversity (DGGE characterisation). Biological and biochemical data were related with both total and selected fractions of Cr and Cu (the latter deriving from agricultural chemical products) as well as with total and extractable organic C. The growth and activity of soil microbial community were influenced by soil organic C content rather than Cu or Cr contents. In fact, positive correlations between all studied parameters and organic C content were found. On the contrary, negative correlations were observed only between total fungal mycelium, dehydrogenase, arylsulphatase and acid phosphatase activities and only one Cr fraction (the soluble, exchangeable and carbonate bound). However, total Cr content negatively affected the eubacterial diversity but it did not determine changes in soil activity, probably because of the redundancy of functions within species of soil microbial community. On the other hand, expressing biological and biochemical parameters per unit of total organic C, Cu pollution negatively influenced microbial biomass, fungal mycelium and several enzyme activities, confirming soil organic matter is able to mask the negative effects of Cu on microbial community.

Pharmacokinetic plasma behaviors of intravenous and oral bioavailability of thymoquinone in a rabbit model.

Thymoquinone (THQ), the active constituent of Nigella sativa seeds, has demonstrated some potential pharmacological activities. The present study was designed to investigate the pharmacokinetic behavior of THQ following intravenous (IV) and oral (PO) administration using an animal model. THQ was given vascularly (5 mg/kg IV) and extravascularly (20 mg/kg PO) to Vole rabbits, and blood samples were collected at predetermined time points. The concentrations of THQ in plasma were measured by a high-performance liquid chromatography, and the pharmacokinetic parameters were determined using both compartmental and non-compartmental analyses. The calculated clearance (CL) following IV administration was 7.19 ± 0.83 ml/kg/min, and the estimated volume of distribution at steady state (V ss) was 700.90 ± 55.01 ml/kg. Whereas with PO dosing, apparent CL/F value was 12.30 ± 0.30 ml/min/kg and V ss/F was 5,109.46 ± 196.08 ml/kg. These parameters were associated with an elimination half-life (T 1/2) of 63.43 ± 10.69 and 274.61 ± 8.48 min with IV and PO dosing, respectively. The calculated absorption T 1/2 was about 217 min. Compartmental analysis revealed T 1/2α of ~8.9 min and T 1/2ß of ~86.6 min. The calculated absolute bioavailability of THQ was ~58 % with a lag time of ~23 min. The estimated THQ protein binding was >99 %. Therefore, THQ represents a compound with rapid elimination and relatively slower absorption following PO administration.

Intrarenal determinants of sodium retention in mild heart failure: effects of angiotensin-converting enzyme inhibition.

The onset and the mechanisms leading to Na+ retention in incipient congestive heart failure (CHF) have not been systematically investigated. To investigate renal Na+ handling in the early or mild stages of CHF, Na+ balance and renal clearances were assessed in 10 asymptomatic patients with idiopathic or ischemic dilated cardiomyopathy and mild heart failure (HF) off treatment (left ventricular ejection fraction, 29.7+/-2%) and in 10 matched normal subjects during a diet containing 100 mmol/d of NaCl and after 8 days of high salt intake (250 mmol/d). Six patients were studied again after 6 weeks of treatment with enalapril (5 mg/d P.O.). At the end of the high salt diet, in patients with mild HF the cumulative Na+ balance exceeded by 110 mmol that of normal subjects (F=3.86, P<.001). During high salt intake, renal plasma flow and glomerular filtration rate were similarly increased in both normal subjects and mild HF patients. In spite of comparable increases of filtered Na+ in the two groups, fractional excretion of Na+, fractional clearance of free water, and fractional excretion of K+ (indexes of distal delivery of Na+) increased in normal subjects and were reduced in patients with mild HF. During enalapril treatment, in the mild HF patients the cumulative Na+ balance was restored to normal; furthermore, enalapril significantly attenuated the abnormalities in the distal delivery of Na+. Our results indicate that a defective adaptation of Na+ reabsorption in the proximal nephron is associated with Na+ retention in response to increased salt intake in the early or mild stages of HF. These abnormalities of renal Na+ handling are largely reversed by enalapril.

Blood levels of erythropoietin in congestive heart failure and correlation with clinical, hemodynamic, and hormonal profiles.

Plasma levels of erythropoietin (mU/ml) were measured in patients with congestive heart failure (CHF) (n = 108) and in a control group of normal subjects (n = 45). In normal subjects, plasma levels of erythropoietin were 1.9 +/- 0.2. In patients with CHF, plasma levels of erythropoietin increased progressively according to New York Heart Association (NYHA) class (I: 1.4 +/- 0.2, n = 28; II: 5.4 +/- 0.8, n = 27; III: 9.6 +/- 2, n = 32; IV: 34 +/- 8, n = 21; F = 57.7, p < 0.001) and were significantly higher in NYHA classes II, III, and IV than in normal subjects. Plasma erythropoietin significantly decreased (from 43 +/- 14 to 12 +/- 3 mU/ml, p < 0.01) in patients with severe CHF (n = 9) when enalapril (20 mg/day administered orally) was added to long-term treatment for 3 weeks. Finally, in a subgroup of patients with NYHA class IV CHF (n = 9) and high plasma erythropoietin levels (37 +/- 9 mU/ml), packed red blood cell volume, assessed by the iodine-125-albumin dilution method, was higher than that in normal subjects (n = 11) (2,616 +/- 235 vs 2,028 +/- 119 ml, p < 0.05). The present study demonstrates that plasma erythropoietin levels are elevated in a large cohort of patients with CHF of varying etiology, and that this increase is related to the progression of the disease. The increase in circulating erythropoietin is associated with augmented packed red blood cell volume in patients with severe CHF. These results suggest a participation of erythropoietin in the complex neurohormonal response that occurs in CHF.

In vitro and in vivo effects of phenolic antioxidants against cisplatin-induced nephrotoxicity.

We have investigated the effect of phenolic antioxidants on cisplatin-induced cytotoxicity in vero (African Green Monkey Kidney) cells and in rat renal cortical slices in vitro, and on cisplatin-induced nephrotoxicity in rats in vivo. Incubation of cisplatin with vero cells resulted in time- and concentration-dependent cytotoxicity, as characterized by decreased tryphan blue exclusion (TBE) and increased release of lactate dehydrogenase (LDH) into the medium. Cisplatin also caused reduction of glutathione (GSH) in a concentration-dependent manner. In the rat renal cortical slices model, incubation of cisplatin for 120 min caused an increase in malondialdehyde (MDA), a decrease in GSH and inhibited p-aminohippurate (PAH) uptake in a concentration-dependent manner. Among phenolic antioxidants, isoeugenol (IG) was found to be more active against cisplatin-induced cytotoxicity in vero cells as well as in rat renal cortical slices than eugenol (EG) and dehydrozingerone (DZ). However none of the test compounds were able to arrest the reduction of the GSH content induced by cisplatin in either the vero cells or the renal cortical slice model. Administration of cisplatin (3 mg/kg) i.p. to rats resulted in significant reduction of body weight, and elevation of blood urea nitrogen (BUN) and serum creatinine. Treatment with IG 10 mg/kg i.p. 1 h before cisplatin resulted in partial but significant protection against the cisplatin-induced reduction of body weight, and elevation of BUN and serum creatinine, the protection being 34, 46, and 62%, respectively. EG and DZ (10 mg/kg, i.p.) were found to be inactive in vivo. Because IG is a potent free radical scavenger and protects against cisplatin-induced toxicitiy, the present results have many clinical implications in chemotherapy and thus warrants further investigation.

Avian B cell development: lessons from transgenic models.

The bursa of Fabricius is critical for the development of B lymphocytes in avian species. Despite considerable advances in our understanding of the molecular mechanisms by which avian antibody diversity is generated, many stages of B-cell development in the bursa and the means by which they are regulated remain unclear. Here we discuss the use of productive chicken retroviral vectors which allow gene transfer in vitro or in vivo as tools to probe the requirements for bursal B-cell development. Expression of a truncated form of bursal cell surface IgM, lacking variable region encoded determinants, is sufficient to promote the initial colonization and clonal expansion of B-cells within the bursa. Expression of this truncated IgM does not, however, protect developing bursal cells against the apoptosis that occurs within the bursa after hatch. Conversely, over-expression of the proto-oncogene bcl-2, following retroviral gene transfer, protects cells against apoptotic cell death but is not sufficient to allow B lineage progression in the absence of sIgM expression. Finally we discuss the use of regulated promoters within the retroviral gene transfer system to show that while bursal cells are susceptible to transformation by the v-rel oncogene in vitro, this oncogene preferentially targets mature peripheral cells in vivo.

Oxidative transformation of phenols in aqueous mixtures.

The transformation by an oxidoreductase (a laccase from Rhus vernificera) of a mixture of four phenols (catechol, methylcatechol, m-tyrosol and hydroxytyrosol) that simulates a typical wastewater derived from an olive oil factory was investigated. Results achieved in this study confirm that laccase-mediated transformation of phenols depends on the nature and the initial concentration of the involved phenol, the time course of the reaction, and mainly, on the complexity of the phenolic incubation mixture. Actually, the four phenols each have a completely different response to enzyme action both in terms of quantitative and kinetic transformation. For example, after 24-h incubation, methylcatechol was completely removed, whereas 30% of untransformed hydroxytyrosol and catechol and more than 65% of m-tyrosol were still present in the reaction mixture. A reduction of enzyme activity occurred for all phenols after enzymatic oxidation. No correspondence between phenol transformation and disappearance of enzymatic activity was observed, thus suggesting that different mechanisms are probably involved in the laccase-mediated transformation of the four phenols. The behavior of phenols became more complex when an increasing number of phenols was present in the reaction mixture, and even more so when different concentrations of phenols were used. Competitive effects may arise when more than one phenol is present in the reaction solution and interacts with the enzyme.

Kinetics and thermal behaviour of the structure formation process in HMP/sucrose gelation.

The concentration and temperature dependence of the gelation kinetics of high-methoxyl pectin (HMP; 60% sucrose, pH 3) was investigated using measurements of small-amplitude oscillatory shear. The rate of gelation close to the gel point can be described as a second-order rate process using the kinetic model of Ross-Murphy (Carbohydr. Polym. 1991, 14, 281) and a critical exponent close to that predicted by the percolation approach. The modulus after a long ageing time showed a power concentration dependence with an exponent around 3.1, higher than the classical square of concentration dependence, which was probably either due to the nonequilibrium state of the HMP gels even after long ageing times, or due to the proximity of the concentration range studied to the critical gelling concentration. The gelation rate of HMP/sucrose systems is strongly dependent on the temperature. An Arrhenius relationship was applied to describe this dependence. Two different processes are proposed to explain the discontinuity observed, each one having rates with different temperature dependence. The applicable kinetics at longer times are quite different, with a lower dependence on polymer concentration and ageing temperature. A non-isothermal kinetic model was used to describe the gelation process of the HMP/sucrose system during cooling.

Bacterial communities and enzyme activities of PAHs polluted soils.

Three soils (i.e. a Belgian soil, B-BT, a German soil, G, and an Italian agricultural soil, I-BT) with different properties and hydrocarbon-pollution history with regard to their potential to degrade phenanthrene were investigated. A chemical and microbiological evaluation of soils was done using measurements of routine chemical properties, bacterial counts and several enzyme activities. The three soils showed different levels of polycyclic aromatic hydrocarbons (PAHs), being their contamination strictly associated to their pollution history. High values of enzyme activities and culturable heterotrophic bacteria were detected in the soil with no or negligible presence of organic pollutants. Genetic diversity of soil samples and enrichment cultures was measured as bands on denaturing gradient gel electrophoresis (DGGE) of amplified 16S rDNA sequences from the soil and enrichment community DNAs. When analysed by Shannon index (H'), the highest genetic biodiversity (H'=2.87) was found in the Belgian soil B-BT with a medium-term exposition to PAHs and the poorest biodiversity (H'=0.85) in the German soil with a long-term exposition to alkanes and PAHs and where absence, or lower levels of enzyme activities were measured. For the Italian agricultural soil I-BT, containing negligible amounts of organic pollutants but the highest Cu content, a Shannon index=2.13 was found. The enrichment of four mixed cultures capable of degrading solid phenanthrene in batch liquid systems was also studied. Phenanthrene degradation rates in batch systems were culture-dependent, and simple (one-slope) and complex (two-slope) kinetic behaviours were observed. The presence of common bands of microbial species in the cultures and in the native soil DNA indicated that those strains could be potential in situ phenanthrene degraders. Consistent with this assumption are the decrease of PAH and phenanthrene contents of Belgian soil B-BT and the isolation of phenanthrene-degrading bacteria. From the fastest phenanthrene-degrading culture C(B-BT), representative strains were identified as Achromobacter xylosoxidans (100%), Methylobacterium sp. (99%), Rhizobium galegae (99%), Rhodococcus aetherovorans (100%), Stenotrophomonas acidaminiphila (100%), Alcaligenes sp. (99%) and Aquamicrobium defluvium (100%). DGGE-profiles of culture C(B-BT) showed bands attributable to Rhodococcus, Achromobacter, Methylobacterium rhizobium, Alcaligenes and Aquamicrobium. The isolation of Rhodococcus aetherovorans and Methylobacterium sp. can be consistent with the hypothesis that different phenanthrene-degrading strategies, cell surface properties, or the presence of xenobiotic-specific membrane carriers could play a role in the uptake/degradation of solid phenanthrene.

Enzymatic oxidative transformation of chlorophenol mixtures.

Chlorinated phenols are major industrial and agricultural xenobiotics that pollute soil and ground water. It has been shown that laccases catalyze the oxidative coupling of phenolic compounds. Therefore, the transformation of one or a mixture of several chlorinated phenols by a laccase from the fungus Trametes villosa was studied. Generally, if more than one phenol was added, the transformation of chlorinated phenols decreased, and if the concentration of the laccase was increased, the transformation of the phenols was enhanced. There were exceptions to these observations: for instance, the transformation of 0.1 mM 4-chlorophenol incubated with 1 mM 2,4-dichlorophenol in buffered salt solutions was not enhanced if the concentration of the laccase was increased from 2 to 20 DMP units/mL. The reason for the reduced transformation of chlorinated phenols in the presence of additional phenols is still unknown. However, in spite of some limitations, the application of laccase to decontaminate wastewater polluted with chlorinated phenols appears feasible.

Inhibition of steroid induced gastric ulcers in forestomachectomized albino rats by phenobarbital.

Oral administration of 2.5 mg of prednisolone/100 gm B.W. to starved forestomachectomized albino rats for three days induces 5-7 acute gastric ulcers with a severity of 3+ in 86% of the rats in the corpus comparable to human gastric ulcers with an ulcer index of 17.1. When 15 mg of Phenobarbital/100 gm B.W. was concomitantly administered with prednisolone either orally or subcutaneously prevents the formation of steroid induced ulcers probably due to their insensitization of gastric epithelium to corticoid ulcerogenic induction. This preliminary observation may have a therapeutic value as this sedative has a common usage. It is is suggested that forestomachectomized rats are best suited for ulcerogenic experiments.

Enzymes as useful tools for environmental purposes.

In the environment enzymes may play important and different roles at least in three cases: as main agents (as isolated, cell-bound or immobilized enzymes) in charge of either the transformation and/or degradation of compounds polluting the environment and the restoration of the polluted environment; as reliable and sensitive tools to detect and measure the amount and concentration of pollutants before, during and after the restoration process; as reliable, easy and sensitive indicators of quality and health status of the environment subjected to the restoration process. To our knowledge papers or reviews integrating findings on these three functions of enzymes are missing in literature. Therefore the main scope of the present paper is to briefly encompass general and specific concepts about roles of enzymes as decontaminating agents, pollutant assaying agents and indicators of environment safety. Examples chosen among those published very recently, supporting and confirming peculiarities, features, and performance of enzymatic agents will be illustrated.

Lepidium sativum but not Nigella sativa affects carbamazepine disposition in an animal model.

Efficacy and safety profile of a drug may be affected when concomitantly used with herbal medicines. The present study was conducted to investigate the effects of some commonly used herbal products viz. Nigella sativa (Black seed) and Lepidium sativum (Garden cress) on the pharmacokinetics of carbamazepine (CBZ), a narrow therapeutic index drug, in an animal model. In a control group, five rabbits received 40 mg/kg of CBZ orally and blood samples were withdrawn at different time intervals (0, 0.5, 1, 1.5, 2, 3, 4, 6, 8, 12 and 24 hrs) from a marginal ear vein. After a suitable washout period, an aqueous saline suspension of Nigella sativa (200 mg/kg) or Lepidium sativum (150 mg/kg) was given orally for eight days to the rabbits. On day eight, CBZ (40 mg/kg) was re-administrated orally and blood samples were collected using the same sampling scheme. Drug levels in plasma were determined by liquid chromatography and pharmacokinetic parameters were calculated using non-compartmental analysis. No significant difference was observed in the maximum concentration (Cmax), area under concentration curve (AUC), half-life (T1/2), clearance (Cl/F) and volume of distribution (Vz/F) of CBZ following Nigella sativa treatment. Whereas, increased Cmax, absorption rate measured by the time to Cmax (Tmax), and prolongation of the terminal elimination half-life (T1/2) were observed after the co-administration with Lepidium sativum. Findings of the present study suggest that concurrent use of Lepidium sativum alters the pharmacokinetics of CBZ in an animal model. Further confirmation of these results in humans will warrant changes in CBZ dose and/or frequency before co-administration with these herbal medicines.

Effect of pH, NaCl, CaCl2 and temperature on self-assembly of ß-lactoglobulin into nanofibrils: a central composite design study.

The ability of certain globular proteins to self-assemble into amyloid-like fibrils in vitro opens opportunities for the development of new biomaterials with unique functional properties, like highly efficient gelation and viscosity enhancement. This work explored the individual and interacting effects of pH (1 to 3), NaCl (0-100 mM), CaCl(2) (0-80 mM) and heating temperature (80 to 120 °C) on the kinetics of ß-lactoglobulin self-assembly and the morphology of resulting nanofibrils. Statistically significant (p < 0.05) interactions included CaCl(2)*temperature, NaCl*pH, CaCl(2)*pH, temperature*pH and NaCl*CaCl(2). Particularly notable was the very rapid self-assembly at pH 3 and the highly nonlinear effect of pH on self-assembly kinetics. Nanofibril morphologies ranged from long and semiflexible or curled and twisted to short and irregular. There did not seem to be a link between the kinetics of fibril formation and the morphology of fibrils, except at pH 3, where self-assembly was very rapid and fibrils were short and irregular, suggesting haphazard, uncontrolled self-assembly.