共查询到20条相似文献,搜索用时 31 毫秒
1.
Ingegerd Kalles Kjell Wikvall 《Biochemical and biophysical research communications》1981,100(3):1361-1369
Electrophoretically homogeneous preparations of cytochrome P-450 LM4 from cholestyramine-treated rabbits catalyzed 7α-hydroxylation of cholesterol, 12α-hydroxylation of 5β-cholestane-3α,7α-diol and 25-hydroxylation of 5β-cholestane-3α,7α,12α-triol. Dithiothreitol, a disulfide reducing agent, specifically stimulated the cholesterol 7α-hydroxylase activity severalfold. The 7α-hydroxylase activity was much more sensitive to the sulfhydryl reagents p-chloromercuribenzoate, N-ethylmaleimide and iodoacetamide than the 12α- and 25-hydroxylase activities. Cholesterol 7α-hydroxylase activity, inactivated by these reagents, could be reactivated by treatment with dithiothreitol. Similar results were obtained with purified cytochrome P-450 from rat liver microsomes.The results indicate that sulfhydryl groups are more important for cholesterol 7α-hydroxylation than for other C27-steroid hydroxylations. 相似文献
2.
Henry Danielsson 《Steroids》1973,22(5):667-676
Various taurine-conjugated bile acids were fed to rats at the 1%-level in the diet for 3 or 7 days and the effect on several hydroxylations involved in the biosynthesis and metabolism of bile acids was studied. The hydroxylations studied were all catalyzed by the microsomal fraction of liver homogenate fortified with NADPH. The 7α-hydroxylation of cholesterol was inhibited by feeding taurocholic acid, taurocheno-deoxycholic acid and taurodeoxycholic acid for 3 as well as 7 days. No marked inhibition was obtained with taurohyodeoxycholic acid or taurolithocholic acid. The 12α-hydroxylation of 7α-hydroxy-4-cholesten-3-one was inhibited after 3 as well as 7 days by all bile acids except taurohyodeoxycholic acid. With this acid a marked stimulation of 12α-hydroxylation was observed. The effects of the different bile acids on the 7α-hydroxylation of taurodeoxycholic acid were not very marked. The 6β-hydroxylation of lithocholie acid and taurochenodeoxycholic acid was stimulated by taurocholic acid and taurodeoxycholic acid. The reaction was inhibited by taurochenodeoxycholic acid, at least after 7 days. Taurohyodeoxycholic acid inhibited the 6β-hydroxylation slightly and taurolithocholic acid had no effect. The results were discussed in the light of present knowledge concerning mechanisms of regulation of formation and metabolism of bile acids and it was suggested that the mechanisms may be more complex than previously thought. 相似文献
3.
12alpha-hydroxylation of 7alpha-hydroxy-4-cholesten-3-one by a reconstituted system from rat liver microsomes 总被引:2,自引:0,他引:2
Claes Bernhardsson Ingemar Björkhem Henry Danielsson Kjell Wikvall 《Biochemical and biophysical research communications》1973,54(3):1030-1038
A preparation of partially purified cytochrome P-450 from rat liver microsomes was found to catalyze 12α-hydroxylation of 7α-hydroxy-4-cholesten-3-one in the presence of NADPH and phosphatidyl choline. The reaction was stimulated two- to four-fold by addition of a preparation of cytochrome P-450 reductase. The reaction was inhibited by carbon monoxide to a considerably less extent than other hydroxylations catalyzed by the reconstituted system. In the presence of optimal concentrations of cytochrome P-450 reductase, cytochrome P-450 prepared from livers of starved rats catalyzed the 12α-hydroxylation more efficiently than cytochrome P-450 prepared from livers of normal rats or rats treated with phenobarbital. 相似文献
4.
《Biochemical medicine》1976,15(1):87-94
A partially purified cytochrome P-450 fraction was prepared from the microsomal fraction of human liver. When combined with NADPH, a synthetic phospholipid and NADPH-cytochrome P-450 reductase from rat liver, the cytochrome P-450 fraction from human liver was able to catalyze the following hydroxylations: 11- and 12-hydroxylation of laurate, 12α- and 26-hydroxylation of 5β-cholestane-3α,7α-diol, 25-hydroxylation of 5β-cholestane-3α,7α,12α-triol, and 6β-hydroxylation of androstenedione and progesterone. It was shown that the rate of 11- and 12-hydroxylation of laurate was linear with increasing amounts of cytochrome P-450 and with time in the presence of excess NADPH-cytochrome P-450 reductase and the phospholipid. In the presence of a fixed amount of cytochrome P-450 and the phospholipid, the rate of 11- and 12-hydroxylation increased with increasing concentrations of NADPH-cytochrome P-450 reductase up to a certain level and then remained constant. The requirement of the phospholipid could be increased markedly by centrifugation of the cytochrome P-450 fraction at 100,000g just prior to incubation. It is concluded that cytochrome P-450 from human liver is similar to previously studied cytochrome P-450 from rat liver with respect to catalytic properties and mechanism of reaction. 相似文献
5.
H Danielsson 《Steroids》1973,22(4):567-579
The effect of biliary obstruction in the rat on several hydroxylations involved in the formation and metabolism of bile acids was studied. The hydroxylations studied were all catalyzed by the microsomal fraction of liver homogenate fortified with NADPH. The rate of 7α-hydroxylation of cholesterol increased two- to threefold between 24 and 48 hours after ligation of the bile duct and remained at this level the next 48 hours. During the first 24 hours of obstruction the rates of 1 2α-hydroxylation of 7α-hydroxy-4-cholesten-3-one and 7α-hydroxylation of taurodeoxycholic acid decreased but returned to control levels between 24 and 48 hours after operation. The rate of 6β-hydroxylation of lithocholic acid and taurochenodeoxycholic acid increased gradually and reached a plateau between 24 and 48 hours at which time the rate was two to three times faster than in the controls. The increase in 6β-hydroxylase activity was reflected in the pattern of the bile acids excreted in urine. After 48 hours of obstruction β-muricholic acid accounted for 50% or more of the bile acids in urine. 相似文献
6.
Zufan Araya Kjell Wikvall 《Biochimica et Biophysica Acta (BBA)/Molecular and Cell Biology of Lipids》1999,1438(1):47-54
The aim of the present study was to identify the enzymes in human liver catalyzing hydroxylations of bile acids. Fourteen recombinant expressed cytochrome P450 (CYP) enzymes, human liver microsomes from different donors, and selective cytochrome P450 inhibitors were used to study the hydroxylation of taurochenodeoxycholic acid and lithocholic acid. Recombinant expressed CYP3A4 was the only enzyme that was active towards these bile acids and the enzyme catalyzed an efficient 6α-hydroxylation of both taurochenodeoxycholic acid and lithocholic acid. The Vmax for 6α-hydroxylation of taurochenodeoxycholic acid by CYP3A4 was 18.2 nmol/nmol P450/min and the apparent Km was 90 μM. Cytochrome b5 was required for maximal activity. Human liver microsomes from 10 different donors, in which different P450 marker activities had been determined, were separately incubated with taurochenodeoxycholic acid and lithocholic acid. A strong correlation was found between 6α-hydroxylation of taurochenodeoxycholic acid, CYP3A levels (r2=0.97) and testosterone 6β-hydroxylation (r2=0.9). There was also a strong correlation between 6α-hydroxylation of lithocholic acid, CYP3A levels and testosterone 6β-hydroxylation (r2=0.7). Troleandomycin, a selective inhibitor of CYP3A enzymes, inhibited 6α-hydroxylation of taurochenodeoxycholic acid almost completely at a 10 μM concentration. Other inhibitors, such as α-naphthoflavone, sulfaphenazole and tranylcypromine had very little or no effect on the activity. The apparent Km for 6α-hydroxylation of taurochenodeoxycholic by human liver microsomes was high (716 μM). This might give an explanation for the limited formation of 6α-hydroxylated bile acids in healthy humans. From the present results, it can be concluded that CYP3A4 is active in the 6α-hydroxylation of both taurochenodeoxycholic acid and lithocholic acid in human liver. 相似文献
7.
Summary Male and female rat liver were studied during post-natal development. A correlation was found between biochemically determined hydroxylations and enzymhisto-chemically determined NADPH-nitro-BT reductase and Naphthol-AS-D esterase. No correlation was found between glucose-6-phosphate dehydrogenase or iso-citric acid dehydrogenase activity and hydroxylations. The difference in hydroxylating capacity between male and female rats may be caused by the fact that the number of cells with hydroxylating activity in the liver lobule, as judged by the NADPH-nitro-BT reductase and Naphthol-AS-D esterase activity, is higher in male than in female rats.List of Abbreviations NADH
reduced nicotinamide adenine dinucleotide
- NADPH
reduced nicotinamide adenine dinucleotide phosphate
- G6PD
glucose-6-phosphate dehydrogenase
- ICD
iso-citric acid dehydrogenase
- G6Pase
glucose-6-phosphatase
- NADPH
-nitro-BT red
- NADPH
Nitro-blue tetrazolium reductase
- SDH
succinic acid dehydrogenase
- TCA
trichloracetic acid 相似文献
8.
The metabolism of estradiol 17-sulfate by subcellular localization enzymes of pheochromocytoma tissue obtained from a 41-year old female was investigated. In any incubations under the presence of NADH and NADPH, metabolites hydroxylated at the C-2, C-4, C-6 beta, C-7 alpha and C-7 beta positions were produced. These hydroxylations are considered to occur without cleavage of the sulfate group. The 2-hydroxylation at the substrate concentration of 100 microM by mitochondria, microsomes and cytosol fractions occurred at rates of 141, 222 and 167 pmol/mg protein/30 min, respectively; the corresponding rates for the 4-hydroxylation were 24, 40 and 38 pmol/mg protein/30 min. Mitochondrial 2- and 4-hydroxylations were enhanced by addition of calcium ion (Ca2+) into the incubation medium. 相似文献
9.
25-hydroxylation of C27-steroids and vitamin D3 by a constitutive cytochrome P-450 from rat liver microsomes 总被引:3,自引:0,他引:3
A constitutive cytochrome P-450 catalyzing 25-hydroxylation of C27-steroids and vitamin D3 was purified from rat liver microsomes. The enzyme fraction contained 16 nmol of cytochrome P-450/mg of protein and showed only one protein band with a minimum molecular weight of 51,000 upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The purified cytochrome P-450 catalyzed 25-hydroxylation of 5 beta-cholestane-3 alpha, 7 alpha-diol, 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha-triol, and 1 alpha-hydroxyvitamin D3 up to 50 times more efficiently, and 25-hydroxylation of vitamin D3 about 150 times more efficiently than the microsomes. The cytochrome P-450 showed no detectable 25-hydroxylase activity towards vitamin D2 and was inactive in cholesterol 7 alpha-hydroxylation as well as in 12 alpha- and 26-hydroxylations of C27-steroids. It catalyzed hydroxylations of testosterone and demethylation of ethylmorphine at the same rates as, or lower rates than, microsomes. The 25-hydroxylation of 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha-triol and vitamin D3 with the purified cytochrome P-450 was not stimulated by addition of phospholipid or cytochrome b5 to the reconstituted system. Emulgen inhibited 25-hydroxylase activity towards both substrates. The possibility that 25-hydroxylation of C27-steroids and vitamin D3 is catalyzed by the same species of cytochrome P-450 is discussed. 相似文献
10.
Huan-Chen Li Chitra Mani David Kupfer 《Journal of biochemical and molecular toxicology》1993,8(4):195-206
Methoxychlor, a currently used pesticide, is demethylated and hydroxylated by several hepatic microsomal cytochrome P450 enzymes. Also, methoxychlor undergoes metabolic activation, yielding a reactive intermediate (M*) that binds irreversibly and apparently covalently to microsomal proteins. The study investigated whether methoxychlor could inhibit or inactivate certain liver microsomal P450 enzymes. The regioselective and stereoselective hydrox-ylation of testosterone and the 2-hydroxylation of estradiol (E2) were utilized as markers of the P450 enzymes inhibited by methoxychlor. Both reversible and time-dependent inhibition were examined. Coincubation of methoxychlor and testosterone with liver microsomes from phenobarbital treated (PB-microsomes) male rats, yielded marked diminution of 2α- and 16α-testosterone hydroxylation, indicating strong inhibition of P4502C11 (P450h). Methoxychlor moderately inhibited 2β-, 7α-, 15α-, 15β-, and 16β-hydroxylation and androstenedi-one formation. There was only a weak inhibition of 6β-ydroxylation of testosterone. The methox-ychlor-mediated inhibition of 6β-hydroxylation was competitive. By contrast, when methoxychlor was permitted to be metabolized by PB-microsomes or by liver microsomes from pregnenolone-16α-car-bonitrile treated rats (PCN-microsomes) prior to addition of testosterone, a pronounced time-dependent inhibition of 6β-hydroxylation was observed, suggesting that methoxychlor inactivates the P450 3A isozyme(s). The di-demethylated methoxychlor (bis-OH-M) and the tris-hydroxy (ca-techol) methoxychlor metabolite (tris-OH-M) inhibited 6β-hydroxylation in PB-microsomes competitively and noncompetitively, respectively; however, these methoxychlor metabolites did not exhibit a time-dependent inhibition. Methoxychlor inhibited competitively the formation of 7α-hydroxytestosterone (7α-OH-T) and 16α-hydroxy-testosterone (16α-OH-T) but exhibited little or no time-dependent inhibition of generation of these metabolites, indicating that P450s 2A1, 2B1/B2, and 2C11 were inhibited but not inactivated. Methoxychlor inhibited in a time-dependent fashion the 2-hydroxylation of E2 in PB-microsomes. However, bis-OH-M exhibited solely reversible inhibition of the 2-hydroxylation, supporting our conclusion that the inactivation of P450s does not involve participation of the demethylated metabolites. Both competitive inhibition and time-dependent inactivation of human liver P450 3A (6β-hydroxylase) by methoxychlor, was observed. As with rat liver microsomes, the human 6β-hydroxylase was inhibited by bis-OH-M and tris-OH-M competitively and noncompetitively, respectively. Testosterone and estradiol strongly inhibited the irreversible binding of methoxychlor to microsomal proteins. This might explain the “clean” competitive inhibition by methoxychlor of the 6β-OH-T formation when the compounds were coin-cubated. Glutathione (GSH) has been shown to interfere with the irreversible binding of methoxychlor to PB-microsomal proteins. The finding that the coincubation of GSH with methoxychlor partially diminishes the time-dependent inhibition of 6β-hydroxylation provides supportive evidence that the inactivation of P450 3A isozymes by methoxychlor is related to the formation of M*. 相似文献
11.
An NADH cytochrome c reductase has been identified in plasma membrane fractions from neutrophils in addition to the superoxide producing NADPH oxidase which has been extensively studied by other investigators. Activation of neutrophils resulted in increased enzyme activities but to different degrees; the NADH cytochrome c reductase increased 2 fold in specific activity and the NADPH oxidase 30 fold. Treatment of the plasma membrane fraction with sonication and differential centrifugation yielded a particulate fraction (R2) with a 2 fold increase in specific activities of both enzymes and concentrations of cytochrome b and FAD. The cytochrome b in the preparation was not reduced under anaerobic conditions by either NADH or NADPH. Treatment of preparations of R2 with deoxycholate or potassium thiocyanate separated the two enzymes yielding particulate preparations with only NADPH oxidase or NADH cytochrome c reductase activity, respectively. 相似文献
12.
The peptidase activity of leukotriene A4 hydrolase purified from human leukocytes has been characterized, utilizing synthetic amides as substrates. The enzyme was stimulated by several monovalent anions. Thiocyanate ions were most effective followed by chloride and bromide ions. In phosphate buffer alone the peptidase activity towards alanine-4-nitroanilide was barely detectable and addition of 100 mM NaCl increased the specific activity more than 20-fold. Increasing the concentration of NaCl (or NaSCN) did not significantly affect the apparent Km for the substrate alanine-4-nitroanilide, but resulted in a dose dependent increase of Vmax. The stimulatory effect of these anions on the reaction velocities appeared to obey saturation kinetics and thus indicated the presence of an anion binding site. Apparent affinity constants for chloride and thiocyanate ions were calculated to 100 and 50 mM, respectively. In contrast to the effect on the peptidase activity, no chloride-stimulation could be detected of the epoxide hydrolase activity of this enzyme, i.e., the conversion of leukotriene A4 into leukotriene B4. In conclusion, the results indicate that under physiological conditions, chloride ions may selectively stimulate the peptidase activity of LTA4 hydrolase. Also, the differences in chloride concentrations between cellular compartments suggest that a possible proteolytic function of the enzyme may be limited to the extracellular space. 相似文献
13.
Effects of carbon monoxide, nitrogen, ferricytochrome c and p-hydroxymercuribenzoate were studied on cholesterol 7α-hydroxylase activity of swine hepatic microsomes. The results suggest that a microsomal electron transport system is involved in hepatic microsomal cholesterol 7α-hydroxylation in swine.Cholesterol 7α-hydroxylation is inhibited by superoxide dismutase in the standard assay system containing a NADPH generating system. Superoxide dismutase also inhibited cholesterol 7α-hydroxylation in the system where superoxides were generated by enzymatic or nonenzymatic means in the absence of NADPH-generating system. The current study suggests that superoxide anion may be an important factor in the cholesterol 7α-hydroxylation of swine. 相似文献
14.
The oxidation of reduced nicotinamide nucleotides by hydrogen peroxide in the presence of lactoperoxidase and thiocyanate, iodide or bromide 总被引:1,自引:0,他引:1
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Lactoperoxidase (EC 1.11.1.7) catalysed the oxidation of NADH by hydrogen peroxide in the presence of either thiocyanate, iodide or bromide. In the presence of thiocyanate, net oxidation of thiocyanate occurred simultaneously with the oxidation of NADH, but in the presence of iodide or bromide, only the oxidation of NADH occurred to a significant extent. In the presence of thiocyanate or bromide, NADH was oxidized to NAD(+) but in the presence of iodide, an oxidation product with spectral and chemical properties distinct from NAD(+) was formed. Thiocyanate, iodide and bromide appeared to function in the oxidation of NADH by themselves being oxidized to products which in turn oxidized NADH, rather than by activating the enzyme. Iodine, which oxidized NADH non-enzymically, appeared to be an intermediate in the oxidation of NADH in the presence of iodide. NADPH was oxidized similarly under the same conditions. An assessment was made of the rates of these oxidation reactions, together with the rates of other lactoperoxidase-catalysed reactions, at physiological concentrations of thiocyanate, iodide and bromide. The results indicated that in milk and saliva the oxidation of thiocyanate to a bacterial inhibitor was likely to predominate over the oxidation of NADH. 相似文献
15.
The Malpighian tubules of Musca domestica secrete a fluid with a high concentration of potassium and low concentrations of sodium, calcium, magnesium and chloride compared with the isolating medium.Low secretion rates are produced by low medium potassium concentrations (< 7 mM), with low sodium concentrations (up to 5 mM) increasing secretion; higher potassium concentrations produce higher secretion rates whilst higher sodium concentrations have no further effect. Calcium and magnesium are essential for secretion.The rate of tubule secretion is inversely proportional to the osmotic pressure of the isolating medium and the osmotic pressure of the secreted fluid is slightly hyper-osmotic to the medium over a range of medium osmotic pressures.The metabolic inhibitors cyanide, iodoacetate and 2,4-dinitrophenol inhibit secretion: Cu2+ ions, arsenate and ouabain have no effect whereas ethacrynic acid abolishes secretion. 5-hydroxytryptamine, cycle AMP and theophylline have no effect on secretion. Sodium thiocyanate stimulates fluid secretion and increases the osmotic pressure and the concentration of sodium and chloride, but not potassium, in the secreted fluid. 相似文献
16.
A novel ADP-ribosyltransferase is present in the cytosolic fraction of various cells. The kinetic behavior and physical properties of this enzyme's activity are clearly distinguished from other known cytosolic ADP-ribosyltransferases. Agents that release nitric oxide, such as sodium nitroprusside, greatly stimulated this activity, although this effect was dependent on the presence of intact thiol groups. Dithiothreitol, reduced glutathione, or cysteine was needed for activation of the enzyme, while N-ethylmaleimide inhibited enzyme activity. High concentrations of phosphate had a slight stimulatory effect, while high concentrations of sodium chloride and thiocyanate were inhibitory. ATP also inhibited this activity. This cytosolic ADP-ribosyltransferase is clearly distinguished from other known and characterized cytosolic transferases. Its activation by biologically active nitric oxide suggests an important role for this enzymatic activity. 相似文献
17.
Clyde Eyster 《Plant and Soil》1959,11(3):207-214
Summary The growth ofChlorella pyrenoidosa in a special medium based on the critical concentrations of nutrients for autotrophic growth has been shown to be stimulated more by chloride with the sodium salt than with the potassium salt, more by sulphate in the presence of sodium than in the presence of potassium, and to be inhibited by sodium dihydrogen phosphate and not by potassium dihydrogen phosphate. Furthermore, it has been found that calcium reversed the growth inhibition produced by sodium dihydrogen phosphate, and that strontium only partially substituted for calcium. 相似文献
18.
MORENO PAO LINI LAURA POZZETTI ANDREA SAPONE ALESSANDRO CAMERINO GIORGIO CANTELLI-FORTI 《Biomarkers》1998,3(3):191-203
In this work the modulation of the regio- and stereo-selective hydroxylation of testosterone by vinclozolin was studied in evaluating cocarcinogenic properties. Changes of cytochrome P450-(CYP)-catalysed drug metabolism was investigated in liver, kidney and lung microsomes of Swiss Albino CD1 mice of both sexes after single (625 or 1250 mg kg-1 b.w.) or repeated (daily 750 mg kg-1 b.w. for 3 days) i.p. administrations. Treatment of mice with a single dose of vinclozolin caused in a dose-dependent fashion from 2 1 to 14 1-fold increase in the 7-, 6- and 2-hydroxylations of testosterone in liver. Lower increase in extrahepatic tissues ranging from 2 3 to 8 1-fold for testosterone 6-, 16 -, 2- and 2- hydroxylase activity in the kidney or from 2 2 to 5 1-fold for 6-, 16 -, 16 - and 2- hydroxylase activity in the lung were observed. Repeated treatment with this fungicide did not substantially modify the extent and pattern of induction, the liver being the only tissue responsive (up to 7 6-fold increase, 7-hydroxylation) in both male and female. In the kidney (7-, 6-, 16 -, 2-, 7-hydroxylations) and lung (6-, 7-, 6-, 16 -, 16 - and 2- hydroxylations), a typical sex-dependent induction (up to 9 0-fold, 16 -hydroxylation in the lung, female) was achieved. In general, however, vinclozolin has a complex pattern of induction and suppression of CYP-dependent enzymes, as exemplified from the reduced expression of some hydroxylations depending upon dose, sex and organ considered. For example, after a single administration, 16 -hydroxylation was suppressed in liver (up to 78% loss in male, higher dose), whereas 16 -hydroxylation was reduced in kidney up to 50% in both sexes (at the higher dose). Glutathione S-transferase activity, measured as index of post-oxidative reactions, was markedly increased by vinclozolin in the liver (up to 5 2-fold, female) and kidney (up to 3 9-fold, female) but not in the lung. Because both phase I and phase II reactions were enhanced by vinclozolin treatment in liver and kidney, the ratio between activation/detoxification mechanisms was slightly affected. Conversely, this ratio was shifted toward activating mechanisms in the lung, sustaining, in part, the expression of certain type of tumours tissue-dependent. Taken together, these findings seem to indicate the cotoxic, cocarcinogenic and promoting potential of this fungicide. 相似文献
19.
MORENO PAOLINI LAURA POZZETTI ANDREA SAPONE ALESSANDRO CAMERINO GIORGIO CANTELLI-FORTI 《Biomarkers》2013,18(3):191-203
In this work the modulation of the regio- and stereo-selective hydroxylation of testosterone by vinclozolin was studied in evaluating cocarcinogenic properties. Changes of cytochrome P450-(CYP)-catalysed drug metabolism was investigated in liver, kidney and lung microsomes of Swiss Albino CD1 mice of both sexes after single (625 or 1250 mg kg-1 b.w.) or repeated (daily 750 mg kg-1 b.w. for 3 days) i.p. administrations. Treatment of mice with a single dose of vinclozolin caused in a dose-dependent fashion from 2 1 to 14 1-fold increase in the 7-, 6- and 2-hydroxylations of testosterone in liver. Lower increase in extrahepatic tissues ranging from 2 3 to 8 1-fold for testosterone 6-, 16 -, 2- and 2- hydroxylase activity in the kidney or from 2 2 to 5 1-fold for 6-, 16 -, 16 - and 2- hydroxylase activity in the lung were observed. Repeated treatment with this fungicide did not substantially modify the extent and pattern of induction, the liver being the only tissue responsive (up to 7 6-fold increase, 7-hydroxylation) in both male and female. In the kidney (7-, 6-, 16 -, 2-, 7-hydroxylations) and lung (6-, 7-, 6-, 16 -, 16 - and 2- hydroxylations), a typical sex-dependent induction (up to 9 0-fold, 16 -hydroxylation in the lung, female) was achieved. In general, however, vinclozolin has a complex pattern of induction and suppression of CYP-dependent enzymes, as exemplified from the reduced expression of some hydroxylations depending upon dose, sex and organ considered. For example, after a single administration, 16 -hydroxylation was suppressed in liver (up to 78% loss in male, higher dose), whereas 16 -hydroxylation was reduced in kidney up to 50% in both sexes (at the higher dose). Glutathione S-transferase activity, measured as index of post-oxidative reactions, was markedly increased by vinclozolin in the liver (up to 5 2-fold, female) and kidney (up to 3 9-fold, female) but not in the lung. Because both phase I and phase II reactions were enhanced by vinclozolin treatment in liver and kidney, the ratio between activation/detoxification mechanisms was slightly affected. Conversely, this ratio was shifted toward activating mechanisms in the lung, sustaining, in part, the expression of certain type of tumours tissue-dependent. Taken together, these findings seem to indicate the cotoxic, cocarcinogenic and promoting potential of this fungicide. 相似文献
20.
delta 4-3-Ketosteroid 5 beta-reductase was purified about 230-fold from 100,000 X g supernatant of rat liver homogenate using 7 alpha-hydroxy-4-cholesten-3-one as substrate throughout. The purified enzyme was electrophoretically homogeneous, and its molecular weight determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was 37,000 and that determined by gel filtration chromatography on calibrated Sephadex G-100 column was 37,200. The absorption spectrum of the purified enzyme showed only a peak at 276 nm due to aromatic amino acids, precluding the presence of a prosthetic group such as flavine in the molecule. The enzyme is highly labile in a low buffer concentration, but is markedly stabilized in the presence of 20% glycerol in 10 mM phosphate buffer. Higher buffer concentration such as 300 mM potassium phosphate buffer was also effective to prevent deterioration in the absence of glycerol, but the effect was somewhat lower compared to glycerol. The purified enzyme showed the activity toward a variety of substrates including testosterone, cortisol, cortisone, progesterone, 4-androstene-3,17-dione, 7 alpha-hydroxy-4-cholesten-3-one, and 7 alpha,12 alpha-dihydroxy-4-cholesten-3-one. The optimal pH for the 5 beta-reduction of 7 alpha-hydroxy-4-cholesten-3-one was 7.4, and the cofactor required for the reaction was NADPH, while NADH revealed no effect. The enzyme activity was inhibited by p-chloromercuribenzoate, but its inhibition was prevented by the presence of a reduced form of glutathione. 相似文献