细胞色素P-450模拟体系的活性中间体—亚碘酰苯铬(Ⅳ)四苯基卟啉配合物的分离、表征和氧化反应

铬(Ⅲ)四苯基卟啉-亚碘酰苯体系在温和条件下能催化氧化碳氢化合物,是细胞色素P-450很好的模拟体系。分离和表征细胞色素P-450模拟体系的活性中间体,不仅能解释细胞色素P-450模拟体系催化氧化碳氢化合物的反应机理,而且也有助于深入了解细胞色素P-450催化氧化碳氢化合物的反应机理及其活性中间体的结构。我们首先分离和表征的细胞色素P-450模拟体系的一种活性中间体-氧配位铬(V)四苯基卟啉配合物,具有氧化碳氢化合物的活性。这篇论文报道细胞色素P-450模拟体系的另一种活性中间体-亚碘酰苯铬(Ⅳ)四苯基卟啉配合物的分离,表征和对环已烯的氧化性质。

细胞色素P-450模拟体系的活性中间体——第三类型氧合铬(Ⅴ)——四苯基卟啉配合物的分离、表征和氧化反应

氧合过渡金属卟啉配合物是细胞色素P-450模拟体系的可能活性中间体。已分离和表征的钛(Ⅳ)、钒(Ⅳ)、钼(Ⅳ,Ⅴ)和铬(Ⅳ)等氧合过渡金属卟啉配合物,因其不是有效的氧化剂,所以不是细胞色素P-450模拟体系的可能活性中间体。氧合锰(Ⅳ)卟啉虽能氧化碳氢化合物,但至今还未能分离出。我们曾分离出并表征的氧合铬(Ⅳ)卟啉以及

Effect of ritonavir-induced cytochrome P450 3A4 inhibition on plasma fentanyl concentrations during patient-controlled epidural labor analgesia: a pharmacokinetic simulation.

BACKGROUND: Ritonavir inhibition of cytochrome P450 3A4 decreases the elimination clearance of fentanyl by 67%. We used a pharmacokinetic model developed from published data to simulate the effect of sample patient-controlled epidural labor analgesic regimens on plasma fentanyl concentrations in the absence and presence of ritonavir-induced cytochrome P450 3A4 inhibition. METHODS: Fentanyl absorption from the epidural space was modeled using tanks-in-series delay elements. Systemic fentanyl disposition was described using a three-compartment pharmacokinetic model. Parameters for epidural drug absorption were estimated by fitting the model to reported plasma fentanyl concentrations measured after epidural administration. The validity of the model was assessed by comparing predicted plasma concentrations after epidural administration to published data. The effect of ritonavir was modeled as a 67% decrease in fentanyl elimination clearance. Plasma fentanyl concentrations were simulated for six sample patient-controlled epidural labor analgesic regimens over 24 h using ritonavir and control models. Simulated data were analyzed to determine if plasma fentanyl concentrations producing a 50% decrease in minute ventilation (6.1 ng/mL) were achieved. RESULTS: Simulated plasma fentanyl concentrations in the ritonavir group were higher than those in the control group for all sample labor analgesic regimens. Maximum plasma fentanyl concentrations were 1.8 ng/mL and 3.4 ng/mL for the normal and ritonavir simulations, respectively, and did not reach concentrations associated with 50% decrease in minute ventilation. CONCLUSION: Our model predicts that even with maximal clinical dosing regimens of epidural fentanyl over 24 h, ritonavir-induced cytochrome P450 3A4 inhibition is unlikely to produce plasma fentanyl concentrations associated with a decrease in minute ventilation.

Automated P.falciparum detection system for post-treatment malaria diagnosis using modified annular ring ratio method

The gametocytes of the malaria parasite Plasmodium falciparum are highly resistant to antimalarial drugs. Its presence in the blood can be detected even after a successful malaria treatment. This paper explains a modified Annular Ring Ratio method which successfully locates and differentiates gametocytes of P. falciparum species in thin blood film images. The method can be used as an efficient tool for gametocyte detection for post-treatment malaria diagnosis. It also identifies the presence of any White Blood Cells (WBCs) in the image, and discards other artifacts and non infected cells. It utilizes the information based on structure, color and geometry of the cells and does not require any segmentation or non-illumination correction techniques that are commonly used for cell detection.

A dimensional analysis of the benzylic hydroxylase active site in mortierella isabellina

The spatial limits of the active site in the benzylic hydroxylase enzyme of the fungus Mortierella isabellina were investigated. Several molecular probes were used in incubation experiments to determine the acceptability of each compound by this enzyme. The yields of benzylic alcohols provided information on the acceptability of the particular compound into the active site, and the enantiomeric excess values provided information on the "fit" of acceptable substrates. Measurements of the molecular models were made using Cambridge Scientific Computing Inc. CSC Chem 3D Plus modeling program. i The dimensional limits of the aromatic binding pocket of the benzylic hydroxylase were tested using suitably substituted ethyl benzenes. Both the depth (para substituted substrates) and width (ortho and meta substituted substrates) of this region were investigated, with results demonstrating absolute spatial limits in both directions in the plane of the aromatic ring of 7.3 Angstroms for the depth and 7.1 Angstroms for the width. A minimum requirement for the height of this region has also been established at 6.2 Angstroms. The region containing the active oxygen species was also investigated, using a series of alkylphenylmethanes and fused ring systems in indan, 1,2,3,4-tetrahydronaphthalene and benzocycloheptene substrates. A maximum distance of 6.9 Angstroms (including the 1.5 Angstroms from the phenyl substituent to the active center of the heme prosthetic group of the enzyme) has been established extending directly in ii front of the aromatic binding pocket. The other dimensions in this region of the benzylic hydroxylase active site will require further investigation to establish maximum allowable values. An explanation of the stereochemical distributions in the obtained products has also been put forth that correlates well with the experimental observations.

The 21-dehydroxylation of corticosteroids: evidence for an enol intermediate and the hydroxylation of steroids by fungal cyto-chrome P450: evidence for a stepwise mechanism

Two enzyme mechanisms were examined: the 21-dehydroxylation of corticosteroids by the anaerobe Eubacterium l en tum, and the hydroxylation of steroids by fungal cytochrome P450. Deuterium labelling techniques were used to study the enzymic dehydroxylation. Corticosteroids doubly labelled (2H) at the C-21 position were incubated with a culture of Eubacterium lentum. It was found that t he enzymic dehydroxylation proceeded with the loss of one 2H f rom C-21 per molecule of substrate. The kinetic isotope ef fect f or the reaction was found to be k~kD = 2. 28. These results suggest that enzyme/substr ate binding in this case may proceed via t he enol form of the substrate. Also , it appears that this binding is, at least in part, the rate determining step of t he reaction. The hydroxylation of steroids by fungal cytochrome P450 was examined by means of a product study. Steroids with a double bond at the A8 (9), ~( lO ), or ~ (ll) position were synthesized. These steroids were then incubated with fungal strains known to use a cytochrome P450 monooxygenase to hydroxylate at positions allylic to these doubl e bonds. The products formed in these incubations indicated that the double bonds had migrated during allylic hydroxylat ion. This suggests that a carbon centred radical or ion may be an intermediate i n the cytochrome P450 cat alytic cycle.

Caracterização físico-química e ações farmacológicas hepatoprotetora, antiinflamatória, pró-angiogênica, antioxidante e anticoagulante da fração rica em fucana 0,8FRF 0,8 da alga marrom Lobophora variegata

This study examines the physical and chemical composition and the pharmacological effects of brown seaweed FRF 0.8 Lobophora variegata. Fractionation of the crude extract was done with the concentration of 0.8 volumes of acetone, obtaining the FRF 0.8. The physicochemical characterization showed that it was a fucana sulfated. Anti-inflammatory activity was assessed by paw edema model by the high rates of inhibition of the edema and the best results were in the fourth hour after induction (100 ± 1.4% at the dose of 75 mg / kg) and by the strong inhibitory activity of the enzyme myeloperoxidase (91.45% at the dose of 25 mg / kg). The hepataproteção was demonstrated by measurements of enzymatic and metabolic parameters indicative of liver damage, such as bilirubin (reduction in 68.81%, 70.68% and 68.21% for bilirubin total, direct and indirect, respectively at a dose of 75 mg / kg), ALT, AST and γ-GT (decrease of 76.93%, 44.58% and 50% respectively at a dose of 75 mg / kg) by analysis of histological slides of liver tissue, confirming that hepatoprotective effect the polymers of carbohydrates, showing a reduction in tissue damage caused by CCl4 and the inhibition of the enzyme complex of cytochrome P 450 (increasing sleep time in 54.6% and reducing the latency time in 71.43%). The effectiveness of the FRF 0.8 angiogenesis was examined in chorioallantoic membrane (CAM) of fertilized eggs, with the density of capillaries evaluated and scored, showing an effect proangigênico at all concentrations tested FRF (10 mg- 1000 mg). The FRF showed antioxidant activity on free radicals (by inhibiting Superoxide Radical in 55.62 ± 2.10%, Lipid Peroxidation in 100.15 ± 0.01%, Hydroxyl Radical in 41.84 ± 0.001% and 71.47 Peroxide in ± 2.69% at concentration of 0.62 mg / mL). The anticoagulant activity was observed with prolongation of activated partial thromboplastin time (aPTT) at 50 mg (> 240 s), showing that its action occurs in the intrinsic pathway of the coagulation cascade. Thus, our results indicate that these sulfated polysaccharides are an important pharmacological target

NMR structural study on the As-P-S glassy system

Glasses having the composition As2S3(1-x)-P2S5(x) with x ranging from 0 to 0.7 have been investigated to determine the compositional effect on properties and local structure. Glass transition temperature (T,) decreases and molar volume (V,,) increases with an increase in P content. Using P-31 NMR, we measured the strength of the P-31-P-31 magnetic dipolar interaction in the glass samples and the AsPS4 crystallized phase. Based on these data, we observed the formation of the As2P2S8 network, which reflects an increase in the average coordination number and a decrease in the degree of rigidity.

Biochemical biomarkers in Nile tilapia (Oreochromis niloticus) after short-term exposure to diesel oil, pure biodiesel and biodiesel blends

Fossil fuels such as diesel are being gradually replaced by biodiesel, a renewable energy source, cheaper and less polluting. However, little is known about the toxic effects of this new energy source on aquatic organisms. Thus, we evaluated biochemical biomarkers related to oxidative stress in Nile tilapia (Oreochromis niloticus) after two and seven exposure days to diesel and pure biodiesel (B100) and blends B5 and B20 at concentrations of 0.01 and 0.1mLL -1. The hepatic ethoxyresorufin-O-deethylase activity was highly induced in all groups, except for those animals exposed to B100. There was an increase in lipid peroxidation in liver and gills in the group exposed to the higher concentration of B5. All treatments caused a significant increase in the levels of 1-hydroxypyrene excreted in the bile after 2 and 7d, except for those fish exposed to B100. The hepatic glutathione-S-transferase increased after 7d in animals exposed to the higher concentration of diesel and in the gill of fish exposed to the higher concentration of pure diesel and B5, but decreased for the two tested concentrations of B100. Superoxide dismutase, catalase and glutathione peroxidase also presented significant changes according to the treatments for all groups, including B100. Biodiesel B20 in the conditions tested had fewer adverse effects than diesel and B5 for the Nile tilapia, and can be suggested as a less harmful fuel in substitution to diesel. However, even B100 could activate biochemical responses in fish, at the experimental conditions tested, indicating that this fuel can also represent a risk to the aquatic biota. © 2011 Elsevier Ltd.

Short communication: Acute but transient increase in serum insulin reduces messenger RNA expression of hepatic enzymes associated with progesterone catabolism in dairy cows

The objective of this experiment was to evaluate the effects of glucose infusion on serum concentrations of glucose, insulin, and progesterone (P4), as well as mRNA expression of hepatic CYP2C19 and CYP3A4 in nonlactating, ovariectomized cows in adequate nutritional status. Eight Gir × Holstein cows were maintained on a low-quality Brachiaria brizantha pasture with reduced forage availability, but they individually received, on average, 3. kg/cow daily (as fed) of a corn-based concentrate from d -28 to 0 of the experiment. All cows had an intravaginal P4-releasing device inserted on d -14, which remained in cows until the end of the experiment (d 1). On d 0, cows were randomly assigned to receive, in a crossover design containing 2 periods of 24. h each (d 0 and 1), (1) an intravenous glucose infusion (GLUC; 0.5. g of glucose/kg of BW, over a 3-h period) or (2) an intravenous saline infusion (SAL; 0.9%, over a 3-h period). Cows were fasted for 12. h before infusions, and they remained fasted during infusion and sample collections. Blood samples were collected at 0, 3, and 6. h relative to the beginning of infusions. Liver biopsies were performed concurrently with blood collections at 0 and 3. h. After the last blood collection of period 1, cows received concentrate and returned to pasture. Cows gained BW (16.5 ± 3.6. kg) and BCS (0.08 ± 0.06) from d -28 to 0. Cows receiving GLUC had greater serum glucose and insulin concentrations at 3. h compared with SAL cohorts. No treatment effects were detected for serum P4 concentrations, although mRNA expression of CYP2C19 and CYP3A4 after the infusion period was reduced for cows in the GLUC treatment compared with their cohorts in the SAL treatment. In conclusion, hepatic CYP3A4 and CYP2C19 mRNA expression can be promptly modulated by glucose infusion followed by acute increases in circulating insulin, which provides novel insight into the physiological mechanisms associating nutrition and reproductive function in dairy cows. © 2013 American Dairy Science Association.

Evaluation of the mutagenic activity of chrysin, a flavonoid inhibitor of the aromatization process

Chrysin is one of the natural flavonoids present in plants, and large amounts are present in honey and propolis. In addition to anticancer, antioxidation, and anti-inflammatory activities, chrysin has also been reported to be an inhibitor of aromatase, an enzyme converting testosterone into estrogen. The present study evaluated the mutagenicity of this flavonoid using micronucleus (MN) with HepG2 cells and Salmonella. Cell survival after exposure to different concentrations of chrysin was also determined using sulforhodamine B (SRB) colorimetric assay in HepG2 cells and the influence of this flavonoid on growth of cells in relation to the cell cycle and apoptosis. TheMN test showed that from 1 to 15 mu M of this flavonoid mutagenic activity was noted in HepG2 cells. The Salmonella assay demonstrated a positive response to the TA100 Salmonella strain in the presence or absence of S9, suggesting that this compound acted on DNA, inducing base pair substitution before or after metabolism via cytochrome P-450. The SRB assay illustrated that chrysin promoted growth inhibition of HepG2 cells in both periods studied (24 and 48 h). After 24 h of exposure it was noted that the most significant results were obtained with a concentration of 50 mu M, resulting in 83% inhibition and SubG0 percentage of 12%. After 48 h of incubation cell proliferation inhibition rates (97% at 50 mu M) were significantly higher. Our results showed that chrysin is a mutagenic and cytotoxic compound in cultured human HepG2 cells and Salmonella typhimurium. Although it is widely accepted that flavonoids are substances beneficial to health, one must evaluate the risk versus benefit relationship and concentrations of these substances to which an individual may be exposed.

Risk factors and mechanisms of hepatocarcinogenesis with special emphasis on alcohol and oxidative stress

Hepatocellular cancer is the fifth most frequent cancer in men and the eighth in women worldwide. Established risk factors are chronic hepatitis B and C infection, chronic heavy alcohol consumption, obesity and type 2 diabetes, tobacco use, use of oral contraceptives, and aflatoxin-contaminated food. Almost 90% of all hepatocellular carcinomas develop in cirrhotic livers. In Western countries, attributable risks are highest for cirrhosis due to chronic alcohol abuse and viral hepatitis B and C infection. Among those with alcoholic cirrhosis, the annual incidence of hepatocellular cancer is 1-2%. An important mechanism implicated in alcohol-related hepatocarcinogenesis is oxidative stress from alcohol metabolism, inflammation, and increased iron storage. Ethanol-induced cytochrome P-450 2E1 produces various reactive oxygen species, leading to the formation of lipid peroxides such as 4-hydroxy-nonenal. Furthermore, alcohol impairs the antioxidant defense system, resulting in mitochondrial damage and apoptosis. Chronic alcohol exposure elicits hepatocyte hyperregeneration due to the activation of survival factors and interference with retinoid metabolism. Direct DNA damage results from acetaldehyde, which can bind to DNA, inhibit DNA repair systems, and lead to the formation of carcinogenic exocyclic DNA etheno adducts. Finally, chronic alcohol abuse interferes with methyl group transfer and may thereby alter gene expression.

Effects of Sulfanilamide and Methotrexate on 13C Fluxes through the Glycine Decarboxylase/Serine Hydroxymethyltransferase Enzyme System in Arabidopsis1

In C3 plants large amounts of photorespiratory glycine (Gly) are converted to serine by the tetrahydrofolate (THF)-dependent activities of the Gly decarboxylase complex (GDC) and serine hydroxymethyltransferase (SHMT). Using 13C nuclear magnetic resonance, we monitored the flux of carbon through the GDC/SHMT enzyme system in Arabidopsis thaliana (L.) Heynh. Columbia exposed to inhibitors of THF-synthesizing enzymes. Plants exposed for 96 h to sulfanilamide, a dihydropteroate synthase inhibitor, showed little reduction in flux through GDC/SHMT. Two other sulfonamide analogs were tested with similar results, although all three analogs competitively inhibited the partially purified enzyme. However, methotrexate or aminopterin, which are confirmed inhibitors of Arabidopsis dihydrofolate reductase, decreased the flux through the GDC/SHMT system by 60% after 48 h and by 100% in 96 h. The uptake of [α-13C]Gly was not inhibited by either drug class. The specificity of methotrexate action was shown by the ability of 5-formyl-THF to restore flux through the GDC/SHMT pathway in methotrexate-inhibited plants. The experiments with sulfonamides strongly suggest that the mitochondrial THF pool has a long half-life. The studies with methotrexate support the additional, critical role of dihydrofolate reductase in recycling THF oxidized in thymidylate synthesis.

A commentary on the impacts of metals and metalloids in the environment upon the metabolism of drugs and chemicals

The salient feature of metals is that unlike organic compounds they do not degrade in the environment and barely move from one environmental matrix to another. Human interventions take these compounds from their stable and non-bioavailable geological matrix into situations of biological accessibility. Studies in the 1970s and the 1980s of metal bioavailability and impacts of metals and metalloids were driven by the process of abatement of lead in the environment. Humans have clear and identifiable sources of exposure from fuels, food and leaded water pipes to lead. Interventions started at that time have dramatically lowered human lead exposure. Attention has now shifted to other metals, in particular, cadmium, which has seen increasing use. It is generally accepted that food crops grown on cadmium containing soils or soils naturally rich in this metal are the major source of exposure to humans other than exposure from smoking of cigarettes. This mini-review gives a summary and commentary on early studies on effects of lead on haem metabolism that provide us the clue to why investigations of the impacts of other toxic heavy metals and metalloids such as cadmium and arsenic on different human cytochrome P450 forms have become of great interest at the current time. (C) 2003 Elsevier Ireland Ltd. All rights reserved.