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C Monder

Publications and source records attributed to C Monder.

At least 73 records · Page 4Linked to original sources
PubMed1985

Extraction of 11 beta-hydroxysteroid dehydrogenase from rat liver microsomes by detergents.

In these studies our goal was to solubilize the microsomal enzyme, 11 beta-hydroxysteroid dehydrogenase (11-HSD) as the first step in its purification. Enzyme was extracted from rat liver microsomes with representative detergents (Zwittergents, Tritons, modified sterols). Oxidation-reduction (O-R) ratios of extracts varied with detergent used and ranged from 0.18 (CHAPS) to 3.8 (Zwittergent 3-14) relative to a ratio of 1.7 in intact microsomes. All detergents solubilized 11-HSD using lack of sedimentation during high speed centrifugation as criterion. With Triton DF-18 and Triton X-100, optimum extraction of 11-HSD occurred in the detergent-protein ratio range of 0.1 to 0.2 O-R ratios decreased with increased Triton X-100, but were constant as Triton DF-18 was varied. The pH optimum of enzyme extraction was 9 at a detergent-protein ratio of 0.05 and 7.5-8.0 at a ratio of 0.2. Sodium chloride increased enzyme extraction by detergents; in the absence of detergent, salt extracted protein, but not enzyme. In aqueous solution at 0 degrees C or -15 degrees C, microsomal 11-oxidation activity rose within 24 h, then decreased; reductase activity consistently decreased. Oxidation and reduction activities were inversely related in the microsomal bound enzyme. No relationship between these activities appeared in detergent-solubilized enzymes. Possible mechanisms to account for the unexpected behavior of this enzyme are discussed.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1985

Measurement of ethynylestradiol and levonorgestrel incorporated into sustained release contraceptive formulations.

High performance liquid chromatography was used to measure the concentrations of ethynylestradiol (EE) and levonorgestrel (LNG) released from contraceptive devices into aqueous medium containing the cationic detergent, benzalkonium chloride. Most of the detergent was removed after solvent extraction of the steroid, although small amounts of it remained in the steroid phase. Over the course of many injections into octasilyl (C8) or octadecylsilyl (C18) reversed-phase columns, the chromatographic profile of EE was gradually altered with multiple peaks emerging. The profile of LNG was not changed. EE chromatographed as a double peak on a fully end-capped C18 column. Rechromatography of each peak yielded a mixture of the two. Mass spectral analysis showed that the peaks differed only in the gain or loss of an equivalent of water. Introduction of a cation exchange column before the analytical column removed residual benzalkonium ions, and by thus preventing deterioration of the column, permitted EE to consistently emerge as a single, symmetrical peak.

Chromatography, High Pressure Liquid↗
PubMed1985

Evidence for independent 11-oxidase and 11-reductase activities of 11 beta-hydroxysteroid dehydrogenase: enzyme latency, phase transitions, and lipid requirements.

Experimental modification of the membrane structure of rat liver microsomes affected the behavior of the 11-oxidase and 11-reductase components of 11 beta-hydroxysteroid dehydrogenase in different ways. 1) The latency of 11-oxidase was released by detergents, phospholipases, or elevated temperature; 11-reductase activity was not increased by these manipulations. 2) 11-Reductase was rapidly inactivated at 25 C and 37 C; 11-oxidase was stable at these temperatures. 3) Arrhenius plots of microsome bound 11-reductase between 5 C and 40 C showed discontinuity at 23 C. Activation energies above and below the critical temperature were 2 kcal and 16 kcal, respectively. Solubilized 11-reductase showed no discontinuity [activation energy (Ea) = 15 kcal]. Ea for 11-oxidase was 15 kcal at all temperatures for membrane bound or solubilized enzyme, with no discontinuities. 4) Phospholipases A2 and C rapidly inactivated 11-reductase. Triton DF-18 regenerated 50% of the reductase activity of phospholipase C-treated microsomes, but had no effect on phospholipase A2-treated microsomes. Phospholipases increased 11-oxidase activity. The independent behavior of corticosteroid 11-oxidase and 11-reductase are consistent with the properties of closely associated, independent enzymes.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1984

Application of polyethyleneimine cellulose for the class separation of steroidal carboxylic acids from neutral steroids and pigments in urine.

Metabolites of corticosteroids that contain the 21-oic acid moiety are found in human urine. The acids from neutral steroids and urinary pigments have been separated by passing the mixture through a column of polyethyleneimine cellulose. The acids adhering to the column are quantitatively eluted with dilute formic acid. The purified preparation is suitable for derivatization and chromatographic analysis.

Adrenal Cortex Hormones↗
PubMed1984

Stimulation of mouse liver corticosteroid side chain isomerase by cobaltous and nickelous ions: evidence for an endogenous inhibitor of isomerase activity.

Corticosteroid side chain isomerase of mouse liver cytosol was stimulated by Co2+ and Ni2+. The magnitude of stimulation increased with incubation time. For Co2+ and Ni2+, respective enhancements were 2.8- and 4.0-fold at 15 min and 3.9- and 5.0-fold at 60 min. The relationship between steroid substrate concentration (11-deoxy-[21-3H]corticosterone) and initial velocity was consistent with a model in which the cations reacted with a cytosol inhibitor of isomerase activity. Enzyme, partially purified by ammonium sulfate fractionation and gel filtration, had a 6.8-fold increased specific activity. Co2+ and Ni2+ enhanced the activity of partially purified enzyme 1.6- and 1.9-fold. Unlike the cytosol, stimulation was achieved without lag and was not altered by prolonged incubation. Metal ion chelating agents did not have a consistent effect on the activity of the partially purified enzyme. Cyanide and alpha,alpha-dipyridyl increased, and dithizone and 8-hydroxyquinoline decreased activity. The data are not consistent with the hypothesis that side chain isomerase is a metalloenzyme. It is concluded that Co2+ and Ni2+ stimulate the enzyme by removing an endogenous inhibitor.

Animals↗
PubMed1984

The fate of corticosterone and 11-deoxycorticosterone in C57BL/6 and BALB/c strains of mice: distribution and oxidative metabolism.

The distribution kinetics and oxidative metabolism of [4-C14] corticosterone (B) and 11-deoxy-[1,2-3H] corticosterone (DOC) were compared in C57BL/6 (B6) and BALB/c (C) mice. Statistically important differences in the distribution of [14C]B and [3H]DOC occurred that were independent of strain, while other differences were strain dependent. Intestinal excretion of metabolites of B and DOC was greater in B6 mice than in C mice, and kidney excretion was greater in C mice than B6 mice. In both C and B6 mice, 3H was cleared from liver faster than 14C, with no strain differences. DOC metabolite levels exceeded B metabolite levels in small intestine and gall bladder of both strains. In most other organs, B metabolites exceeded DOC metabolites. Time average strain differences in accumulation of B and its metabolites favoring B6 were found in pancreas, brain, lung, heart, muscles, adrenals, spleen, mesentery and small intestine. Except for the organs of excretion, no strain differences were found for [3H]DOC metabolites. Sixty minutes after steroid administration, 45% of B metabolites and a third of DOC metabolites were 20-hydroxy-21-oic acids. In the intestine, accumulation of acids derived from either B or DOC was greater for B6 than C strain mice, reflecting the greater proportion of total steroid excreted in the B6 strain.

Animals↗
PubMed1984

11 beta-Hydroxysteroid dehydrogenase: fact or fancy?

Previous attempts to explain the diverse behavior of 11 beta-hydroxysteroid dehydrogenase (11-HSD) within and between species have not been successful. We now propose that 11-HSD activity is the resultant of the coordinated interaction of two enzyme types, 11-dehydrogenase and 11-reductase. We have demonstrated their separate existence by physico-chemical and kinetic methods. Based on these findings, two classes of disease in humans that have been recently described can now be characterized as being associated with a deficiency in either 11-dehydrogenase or 11-reductase.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1983

Metabolism of corticosterone in the mouse. Identification of 11 beta, 20 alpha-dihydroxy-3-oxo-4-pregnen-21-oic acid as a major metabolite.

We have shown that mouse liver contains enzymes that catalyze the conversion of the ketol side chain to the 20-hydroxy-21-oic acid side chain. In this paper, we have studied the oxidative metabolism of corticosterone to acidic end products in intact mice. A significant fraction of radioactivity from intraperitoneal injections of [4-14C]corticosterone appeared in liver and intestine within 5 min. The major steroid in liver at 5 min postinjection was found to be corticosterone, although acidic metabolites were detected. Within 30 min after injection, 11 beta, 20 alpha-dihydroxy-3-oxo-preg-4-en-21-oic acid became the dominant steroid. At 60 min, it was the major steroid isolated from liver or intestine. Several other acid metabolites were present in lesser amounts in both organs. About half of the remaining radioactive metabolites in liver and intestine were steroid acids, as determined by their reaction with diazomethane. The identification of the major steroid acid as 11 beta, 20 alpha-dihydroxy-3-oxo-pregn-4-en-21-oic acid was made by comparing the chromatographic behavior of the free acid and its methyl ester with that of authentic synthetic acid using thin layer and high performance liquid chromatography. Identity was confirmed by showing that the specific activities of the homogeneous 14C-labeled free acid remained unchanged when reanalyzed as the 21-methyl ester.

Animals↗
PubMed1983

Aldose and aldehyde reductase exhibit isocorticosteroid reductase activity.

In this paper we describe the reduction of corticosteroid metabolites containing the 17 beta-aldol side chain (isocorticosteroids) by aldose and aldehyde reductase from human tissues. Aldose reductase catalyzed the reduction of the aldehydes derived from cortisol and corticosterone at about the same rate, whereas aldehyde reductase preferentially acted on the aldehydes derived from 17-deoxycorticosteroids. At comparable rates of reduction the Michaelis constants for the best steroid aldehydes were one order of magnitude lower than for the hitherto best substrates. We propose that aldose and aldehyde reductase participate in the conversion of the corticosteroid ketol side chain to the glycol side chain via an aldol intermediate by the 'long loop' pathway proposed by Monder and Bradlow [(1977) J. Steroid Biochem. 8, 897-908].

Alcohol Oxidoreductases↗
PubMed1983

Asymmetric reduction of steroidal 20-ketones: chemical synthesis of corticosteroid derivatives containing the 20 alpha, 21-diol and 17 alpha, 20 alpha, 21-triol side chains.

A method is presented for the chemical synthesis of corticosteroid derivatives containing the 20 alpha, 21-diol and 17 alpha, 20 alpha, 21-triol side chains. The ketol side chains of cortisol, corticosterone, 11-deoxycortisol, and 11-deoxycorticosterone were reduced at C-20 with sodium borohydride in a two-phase system consisting of aqueous calcium chloride and an organic phase of chloroform or ethyl acetate. Stereoselectivity of reduction was 92% alpha-oriented for cortisol and 79% alpha-oriented for 11-deoxycortisol at -27 degrees. The 20 alpha-form diminished relative to the 20 beta-form with increasing temperature. For the 17-deoxy steroids, reduction to the 20 alpha-form was 23% for 11-deoxycorticosterone and 41% for corticosterone. The 20 alpha/20 beta ratios of 17-deoxy steroids were unchanged between 0 degree and -27 degrees. Calcium ions increased the solubility of corticosteroids in the aqueous phase. We propose that calcium ions affect the stereochemistry of reduction by forming a bidentate complex with the side chains of 17 alpha-hydroxy steroids, fixing them in an orientation favorable to 20 alpha-reduction, and by altering the phase partition of the steroids.

Borohydrides↗
PubMed1983

Genetic control of corticosteroid side-chain isomerase activity in the mouse.

The corticosteroid side-chain isomerases of mammalian liver catalyze the interconversion of the ketol and aldol side chains. In the mouse, isomerase was low in C57BL/6 (B6) mice (130 pmol/mg protein . 2 h) and high in BALB/c (C) mice (230 pmol/mg protein . 2 h). From analysis of hybrids between B6 and C and of backcrosses of these hybrids to B6, it was concluded that isomerase levels are controlled by a single autosomal gene dominant for high activity. The distribution of high and low isomerase levels in a series of CXB/By recombinant inbred strains of mice was consistent with linkage of the isomerase gene to H-2. Congenic BALB.B mice (H-2b haplotype from C57BL/10) had low isomerase activities corresponding to C57BL/10, not the high activity of the background strain BALB/c(H-2d). Similarly, BN10.D2 congenic mice (H-2d haplotype from the DBA/2 strain) had high activity characteristic of DBA/2. In the (C X B6)F1, (C X BALB.B)F1 and (B10 X B10.D2)F1 hybrids, all of which are H-2d/H-2d heterozygotes, isomerase activity was high. The association of isomerase levels with H-2 type was further confirmed in mice of the following backcrosses: (C X BALB.B)F1 X BALB.B, (C X B6)F1 X B6 and (B10 X B10.D2)F1 X B10. H-2b/H-2b homozygous segregants had consistently low activity and H-2b/H-2d heterozygous segregants had consistently high activity. It was concluded that the level of corticosteroid side-chain isomerase activity in mouse liver is controlled by a gene(s) in the region of the H-2 locus on chromosome 17.

Animals↗
PubMed1983

A comparison of the tissue distribution and metabolism of 11-deoxy-[1,2-3H]corticosterone in the BALB/c and C57BL/6 strains of mice.

Corticosteroid side-chain isomerase of mouse liver catalyzes the reversible interconversion of the ketol and aldol configurations of the corticosteroid side chain. Activity of the enzyme is under genetic control. To see if the differences in activity that were observed in vitro between inbred strains of mice were also expressed in vivo, the metabolism of 11-deoxy-[1,2-3H]corticosterone ([1,2-3H]DOC) was studied in BALB/c (C) and C57BL/6 (B6) mice. Maximum radioactivity appeared in most organs within 5-10 min after ip injection. Uptake of tracer into liver was greater for C than B6 mice. Tritium levels in blood, kidney, and pancreas were higher in C mice; levels in adrenal, abdominal fat, and mesentery were higher in B6 mice. In both strains, the concentrations of tracer in tissues, except in gastrointestinal tract, declined and reached a minimum within 60 min. Most of the radioactivity (84%) from [1,2-3H]DOC accumulated in the lumen of the intestinal tract, and few counts were found in the wall. Intestinal concentrations of 3H at different postinjection intervals were greater for B6 than C mice. In contrast, twice as much radioactivity appeared in the kidneys of C than of B6 mice. The organs of excretion (kidney, liver, gall bladder, and intestine) concentrated steroid from blood. Heart, striated muscle, and spleen excluded steroid. Four acidic metabolites of [1,2-3H]DOC were detected in liver, and two were detected in small intestine. Acids formed in liver did not accumulate, and no differences between C and B6 strains were seen. More acid metabolites accumulated in intestines of C mice than in those of B6 mice. The quantitative aspects of steroid acid formation in vivo are consistent with our previous in vitro findings that livers from C mice synthesize more pregnolic acid from DOC than do livers from B6 mice.

Acids↗
PubMed1983

Altered cortisol metabolism in cells cultured from trabecular meshwork specimens obtained from patients with primary open-angle glaucoma.

Cells cultured from trabecular meshwork specimens obtained from patients with primary open angle glaucoma (TMPOAG cells) exhibited two major differences in cortisol-metabolizing enzymes when compared with similar cells from nonglaucomatous patients. One is a marked increase (greater than 100-fold) in delta 4-reductase activity and the other is a decrease (4-fold) in 3-oxidoreductase activity. Peripheral lymphocytes from one of these patients as well as from five additional patients with POAG, did not show these abnormalities, indicating that the defects are not found in all cortisol-metabolizing cells. The abnormal metabolism of cortisol by TMPOAG cells may be of significance in the pathogenesis of POAG.

Adult↗
PubMed1982

Oxidation of cortisol to hydroxy acid metabolites by liver cytosol.

Cytosols (post-microsomal supernatants) prepared from rat, hamster and mouse livers oxidized cortisol to 11 beta, 17, 20-trihydroxy-3-oxo-pregn-4-en-21-oic acids. Mouse liver enzymes yielded over 90% 20 alpha-hydroxy epimer from cortisol, 21-dehydrocortisol (11 alpha, 17-dihydroxy-3,20-dioxo-pregn-4-en-21-aldehyde), and 20 alpha-isocortisol (11 alpha, 17, 20 alpha-trihydroxy-3-oxo-pregn-4-en-21-aldehyde). The 20 beta-epimer of isocortisol yielded both 20 alpha- and 20 beta-hydroxy acid. Rat and hamster liver cytosols converted, 21-dehydrocortisol and 20 alpha-isocortisol to both 20 alpha and 20 beta-hydroxy acids, with the former predominant. The hamster enzyme oxidized 20 beta-isocortisol mainly to the 20 beta-hydroxy acid. The results support our conclusion that both 17 alpha-hydroxy and 17-deoxy corticosteroids are oxidized to hydroxy acids by similar pathways and that isosteroids are obligatory intermediates.

Animals↗
PubMed1982

Oxidation of the 17-aldol (20 beta hydroxy-21-aldehyde) intermediate of corticosteroid metabolism to hydroxy acids by homogeneous human liver aldehyde dehydrogenases.

In human liver, the oxidation of corticosteroids to 20-hydroxy-21-oic acids proceeds via the formation and oxidation of aldol (20-hydroxy-21-aldehyde) intermediates. Human liver aldehyde dehydrogenases E1 and E2, which we have previously purified to homogeneity, catalyzed the oxidation of the aldol isomer of cortisol (isocortisol) or of 11-deoxycorticosterone (isoDOC) by E1 and E2 respectively, were identified by the criteria of chromatographic mobility, derivatization, and reverse isotope dilution of 4-14C labeled acid end products. Both enzymes showed broad substrate specificity and oxidized both 17-hydroxy and 17-deoxy steroids, though at widely varying rates. Kinetic analysis of the course of oxidation of isocortisol and isoDOC by NAD+ gave intersecting initial velocity plots that conform with a sequential mechanism. The inhibition patterns for both enzymes with thionicotinamide adenine dinucleotide or chloral hydrate were consistent with random sequential behavior.

Adrenal Cortex Hormones↗
PubMed1982

Oxidation of corticosteroids to steroidal 20-hydroxy-21-oic acids by mouse liver.

We have studied the enzyme catalyzed oxidation of 11-deoxycorticosterone to 20-hydroxy-3-oxo-4-pregnen-21-oic acid (pregnolic acid) in mouse liver. Enzyme activity, though variable, was higher than that of other species. Pregnolic acid was identified as the free acid, as the methyl ester by thin layer chromatography, and as the p-bromophenacyl ester by high performance liquid chromatography. With [4-14C, 21-3H]-DOC as substrate, exchange of tritium with water (interpreted as due to the reversible isomerization of the ketol to aldol form by the side chain) and the overall conversion of the ketol side chain to hydroxy acid was catalyzed by the post-microsomal supernatant fraction. Although we could not physically separate tritium exchange and acid production, pregnolic acid formation could be decreased or eliminated while tritium exchange was retained, consistent with our previous conclusion that isomerization to aldol was a precondition for acid formation. In preparations that made no acid, [4-14C]-DOC was recovered, depleted of tritium. The rate of exchange of [21S, 21-3H]-DOC with water was faster than [21R, 21-3H]-DOC. The stereochemistry of pregnolic acid at C-20 was 85-90% R (i.e.. 20 alpha-hydroxy-21-oic acid). The Km for isomerase with [21RS-21(3) H-DOC was 4.3 x 10(-5); Km for pregnolic acid formation was 8.0 x 10(-5) M. Corticosterone was oxidized to acid metabolites at 20% the rate of DOC.

Adrenal Cortex Hormones↗
PubMed1982

Metabolism of cortoic acids in man.

Administrated radioactive tracers of beta-cortolic or beta-cortolonic acids are excreted mostly unchanged, without significant alteration of the molecule at C-11 or C-20; a minor amount of cleavage to C-20 etianic acids occurs. Evidence was obtained for possible noncovalent complexing of the steroid acids to macromolecular components present in the enzyme preparation for hydrolysis of the urinary glucuronides; this complex can be dissociated by acidification to pH 2, and the acidic metabolites can then be readily extracted. The findings suggest that beta-cortoic acids are essentially terminal metabolites in man.

Adult↗