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D M Ziegler

Publications and source records attributed to D M Ziegler.

At least 37 records · Page 2Linked to original sources
PubMed1991

Estimation of flavin-containing monooxygenase activities in crude tissue preparations by thiourea-dependent oxidation of thiocholine.

The activity of flavin-containing monooxygenases in microsomes and whole homogenates is readily estimated by following the thiourea-dependent oxidation of thiocholine. NADPH- and oxygen-dependent flavin-containing monooxygenases catalyze the oxidation of thiourea to formamidine sulfenic acid, which oxidizes thiocholine to thiocholine disulfide. The latter reaction is quite rapid and never rate limiting even at concentrations of thiocholine below 30 microM. The loss of thiocholine in deproteinized aliquots of the reaction medium is measured colorimetrically with the thiol reagent, DTNB [5,5'-dithiobis(2-nitrobenzoate)]. In the absence of thiourea, thiocholine is not oxidized and its disulfide is not reduced at a detectable rate even in reactions containing 4-5 mg of liver or kidney homogenate protein per milliliter. In all tissues where both can be measured, rates of thiocholine oxidation and N,N-dimethylaniline N-oxygenation were virtually identical, which suggests that both activities are catalyzed by the same monooxygenase.

Aniline Compounds↗
PubMed1991

Effects of dietary protein-energy interrelationships on Holstein steer performance and ruminal bacterial fermentation in continuous culture.

In vivo and in vitro 3 x 2 factorial experiments were conducted concurrently to evaluate the incorporation of 0, 15, or 30% sugar beet pulp (SBP) as an energy source in diets fed to growing Holstein steers with either soybean meal (SBM) or alcohol-treated, defatted soybean flakes (ATSBF) as primary supplemental protein sources. Three groups of 42 Holstein steers each were fed six different diets from 54 kg initial BW to 320 kg in three experimental periods. There were no overall SBP level x protein source interactions (P greater than .05). Beet pulp level tended to decrease ADG (linear, P = .05) and increase feed/gain (linear, P less than .05) and DMI (quadratic, P less than .05). Each grower diet was used in a substrate for ruminal microbial metabolism in six dual-flow, continuous-culture fermenters. Organic matter and carbohydrate digestion were similar (P greater than .05) among diets. Increasing dietary levels of SBP caused a concomitant increase (P less than .05) in acetate and decrease (P less than .05) in butyrate and isobutyrate concentrations. Beet pulp level x protein source interactions (P less than .05) were observed for CP degradation, ammonia and nonammonia N, and dietary N flow. Crude protein degradation was higher (P less than .05) for the 0% SBP with SBM diet (81.3%) than for the 30% SBP with ATSBF diet (64.4%). Efficiency of bacterial synthesis was similar (P greater than .05) among diets. Results indicated that SBP is an effective dietary energy source for high-energy grower diets at 15 or 30% of the DM but may cause a decrease in some performance traits. There were no nutritional benefits of using ATSBF vs SBM as the supplemental N source.

Animal Feed↗
PubMed1990

The flavin-containing monooxygenase expressed in pig liver: primary sequence, distribution, and evidence for a single gene.

The primary sequence of the flavin-containing monooxygenase expressed in pig liver has been derived from the nucleotide sequence of cloned cDNA. The derived sequence is composed of 532 amino acids and represents a protein having a molecular weight of 58,952. The complete sequence was obtained from a single clone containing 2070 bases. A second clone, obtained from an independent library, yielded an identical sequence for the 1374 bases present. The amino acid composition compiled from the derived sequence is very similar to that obtained previously from the purified protein. In addition, a 10 amino acid sequence in a peptide formed from the purified protein by digestion with V8 protease exactly matches the derived sequence for residues 309-318. The flavin-containing monooxygenase expressed in pig liver is also expressed in pig lung and kidney as determined by analysis of both microsomal proteins and mRNA. The ratio of mRNA to protein for the enzyme in kidney is about 5 times greater than the same ratio for liver and about twice the ratio for lung. The reasons for these differences are not understood. Southern analysis of genomic DNA indicates that there is a single gene encoding the flavin-containing monooxygenase expressed in pig liver. Therefore, the broad activity of this enzyme in liver appears to be the result of the catalytic diversity of a single protein.

Amino Acid Sequence↗
PubMed1990

Flavin-containing monooxygenases: enzymes adapted for multisubstrate specificity.

Unlike all other oxidases, microsomal flavin-containing monooxygenases (FMO) discriminate between essential and foreign compounds by excluding the former rather than selectively binding the latter. As Daniel Ziegler describes here, xenobiotics that readily cross cell membranes can enter the catalytic cavity, whereas charged groups on essential metabolites that prevent their passive diffusion out of the cell also block their access to FMO. FMO appears to be ideally adapted to catalyse the detoxification of structurally diverse soft nucleophiles (e.g. alkaloids with basic side-chains and organic sulfur xenobiotics) so abundant in food derived from plants.

Animals↗
PubMed1990

Evaluation of various nitrogen supplements in starter diets for growing Holstein steers and their effects on ruminal bacterial fermentation in continuous culture.

Concurrent in vivo and in vitro studies were conducted to evaluate urea (U), soybean meal (SBM), ground soybeans (RAW), extruded soybeans (ES) or extruded soybeans plus urea (ES + U) as primary supplemental N sources in starter diets for Holstein steers. Three groups of 48 Holstein steers each were fed five different starter diets to 181 kg BW in three experimental periods over 2 yr. Average daily gains were similar (P greater than .05) for steers fed ES + U (1.12 kg), ES (1.08 kg) and SBM (1.09 kg) but lower (P less than .05) for those fed U (1.00 kg) or RAW (.97 kg) diets. Feed/gain was similar (P greater than .05) for ES-fed steers vs those fed other diets except U. From 181 to 477 kg, all steers were fed the same diet. Steers fed the RAW starter diet had the lowest (P less than .05) ADG for the entire period. The starter diets were used as substrates for ruminal microbial metabolism in eight dual-flow continuous culture fermenters. True OM digestion was higher and NDF and ADF digestion was lower (P less than .05) for the ES + U diet than for the ES diet. Dietary protein degradation was lowest (P less than .05) for the ES diet (64.4%). Total bacterial N flow was higher (P less than .05) with the ES + U, SBM and U diets than with the ES diet. Lysine flow was higher (P less than .05) for the ES + U diet than for all other diets except ES. Results of these experiments indicate that ES as a protected ruminal escape N source with or without added urea did not improve steer performance above that obtained from SBM in starter diets.

Animal Feed↗
PubMed1989

Nitroglycerin and isosorbide dinitrate stimulation of glutathione disulfide efflux from perfused rat liver.

Nitroglycerin (GTN) and isosorbide dinitrate (ISD) are metabolized by glutathione S-transferase to nitrite with production of GSSG from GSH. Infusion of organic nitrates into perfused rat liver led to efflux of GSSG in the bile and nitrite in the perfusate. Biliary GSSG increased more rapidly than did nitrite release as GTN infusion rate was increased, indicating that GSSG reducing capacity was being exceeded. Rapid GTN-induced oxidation of GSH may be the mechanism of tissue GSH depletion by GTN and other alkylnitrates. Such depletion of glutathione may reduce nitrite production from organic nitrates and underlie tolerance to these drugs.

Animals↗
PubMed1989

Tissue and biliary glutathione disulfide in the perfused vitamin E-deficient rat liver.

Vitamin E and glutathione protect against oxidative damage in vivo. In this study the relationship between these two defenses has been examined in the isolated perfused rat liver. The activities of glutathione reductase and glutathione S-transferase were unaffected by vitamin E deficiency, while glutathione peroxidase activity was decreased slightly. The glutathione redox status of vitamin E-deficient and control livers was assessed. GSSG was slightly higher in vitamin E-deficient livers (70 +/- 5 nmol GSH equivalents/g liver) than in controls (56 +/- 3 nmol GSH equivalents/g liver) under basal conditions. However, biliary GSSG release was 41% lower in vitamin E-deficient livers (0.46 +/- 0.08 nmol GSH equivalents/g liver.min) than in controls (0.78 +/- 0.23 nmol GSH equivalents/g liver.min). Inhibition of GSSG reduction by BCNU raised liver and biliary GSSG by a similar amount in vitamin E-deficient and control livers. Thus biliary GSSG efflux, a frequently used index of oxidant stress, is not increased in vitamin E-deficient perfused livers compared with control. Therefore, in the perfused rat liver model, no evidence was obtained that vitamin E deficiency activates the hepatic glutathione system.

Animals↗
PubMed1988

Estimation of lipid concentrations on thin-layer plates by densitometry of transparent copies.

A relatively simple procedure for the quantitative estimation of phospholipids resolved on thin-layer plates has been developed. After resolution in an appropriate solvent, the lipids are visualized by staining with iodine, ninhydrin, or molybdate and then photocopied onto transparent sheets with a standard office copy machine. The density of each spot on the photocopy, measured with a simple silicon cell area densitometer, is a direct function of each lipid applied to the plate over at least a six- to eightfold range in concentration. Under controlled conditions the staining and photocopying steps are quite reproducible. Known concentrations of the choline, ethanolamine, serine, and inositol derivatives of L-alpha-phosphatidic acid applied either separately or as mixtures can be determined essentially quantitatively (100 +/- 5%) by this procedure following their resolution on the thin-layer plates.

Animals↗
PubMed1988

N-methylation: potential mechanism for metabolic activation of carcinogenic primary arylamines.

Two amine N-methyltransferases isolated from rabbit liver catalyze S-adenosylmethionine-dependent N-methylation of benzidine and 4-aminobiphenyl but not of 4-aminoazobenzene or 2-aminobiphenyl. The enzymatic reaction products were analyzed and found to be identical to synthetic N-methylbenzidine and N-methyl-4-aminobiphenyl. N-Methylation may be a critical step in the metabolic activation of primary arylamines because N-methylarylamines, unlike primary arylamines, are readily N-oxygenated by the NADPH- and oxygen-dependent microsomal flavin-containing monooxygenase. Kinetic studies carried out with the purified porcine liver monooxygenase demonstrate that, while activity with primary arylamines could not be detected, N-methyl derivatives of benzidine, 4-aminoazobenzene, and 4-aminobiphenyl are substrates. Products formed from N-methyl-4-aminobiphenyl had the properties of the hydroxylamine and/or nitrone in that the enzyme- and time-dependent incubation product(s) reduced Fe3+ to Fe2+, and formaldehyde was formed during the course of the reaction. These data suggest that N-methyl-4-aminobiphenyl is oxidized to N-hydroxy-N-methyl-4-aminobiphenyl, which can undergo further oxidation to a nitrone that hydrolyzes to formaldehyde and N-hydroxy-4-aminobiphenyl.

Aniline Compounds↗
PubMed1988

Stereoselectivity in the N'-oxidation of nicotine isomers by flavin-containing monooxygenase.

N'-Oxidation of nicotine isomers by porcine liver flavin-containing monooxygenase shows a clear stereoselectivity in the formation of the diastereomeric N'-oxides. (S)-(-)-Nicotine exhibited no stereoselectivity in the formation of cis-1'R,2'S- and trans-1'S,2'S-products, whereas with (R)-(+)-nicotine, only the trans-1'R,2'R-N'-oxide was formed. The concentration of each isomer required for half maximal activity differs significantly, and access of (S)-(-)-nicotine to the active site appears to be more restricted than for (R)-(+)-nicotine as judged from the observed Km values (Km = 181 and 70 microM, respectively, for the (S)-(-)- and (R)-(+)-isomers). These results indicate that a region adjacent to the active site may sterically prohibit binding of (R)-(+)-nicotine when the N'-methyl and pyridyl groups are in a cis-orientation. N-Methylnicotinium ion (both R- and S-isomers) is not a substrate for either porcine flavin monooxygenase, guinea pig liver microsomes, or ram seminal vesicular microsomes.

Animals↗
PubMed1987

Studies on substrate specificity of the hog liver flavin-containing monooxygenase. Anionic organic sulfur compounds.

The influence of anionic groups on interaction of nucleophilic sulfur compounds with the purified hog liver flavin-containing monooxygenase was evaluated from kinetic constants obtained with various dithiobenzoates, thiolbenzoates, and thiolalkylcarboxylic acids. All compounds tested bearing a single negative charge localized on sulfur were excellent substrates but derivatives with a carboxylic acid group one or two carbons removed from the heteroatom exhibited low or no substrate activity. The effect of a carboxylic acid group more distal from sulfur appeared to depend on steric factors that are not well defined. For instance, none of the carboxylic acids (C2-C8) bearing a single thiol on the terminal carbon were oxygenated at detectable rates, whereas dihydrolipoic acid appeared to be a substrate although the concentration required for half-maximal activity was quite high (approximately 2 mM). Lipoic acid was a much better substrate (Km = 0.12 mM), and kinetic constants obtained with lipoic acid analogues suggest that position of the negative charge relative to the dithiolane ring is critical, since increasing the length of the side chain increased the Km. None of the alicyclic disulfides or sulfides containing one or more carboxylic acid groups showed detectable substrate activity. However, the more lipophilic sulfur-containing fatty acids inhibited the enzyme which may mask their potential substrate activity.

Animals↗