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1.
Biopharm Drug Dispos ; 20(7): 319-26, 1999 Oct.
Article in English | MEDLINE | ID: mdl-10760839

ABSTRACT

CP-191,166 is an orally active, non-peptide angiotensin II (AII) receptor antagonist developed for the treatment of hypertension and congestive heart failure (CHF). In this study, the intravenous (iv) and oral (po) single dose pharmacokinetics (PK), oral multiple dose PK and P450-mediated metabolism of CP-191,166 were determined in rats and dogs. CP-191,166 was administered in both single and multiple (22-29 day) doses to Sprague-Dawley rats (3 mg/kg iv and 5, 10, 25 and 200 mg/kg po) and to beagle dogs (5 mg/kg iv and 5, 15 and 50 mg/kg po). Blood samples were collected between 0 and 48 h and plasma CP-191,166 concentrations were determined using high performance liquid chromatography (HPLC) with ultraviolet (UV) detection. The in vitro metabolism of CP-191,166 was also evaluated with rat and dog liver microsomes. The results of these studies suggest that in both species, there may be saturable clearance occurring with higher doses, T(max) was at or near the earliest sample time point for all doses, suggesting that the drug was rapidly absorbed, and CP-191,166 was eliminated with t(1/2) values of 8-9 h. No rat or dog microsomal metabolism was observed, suggesting that metabolites detected in vivo in dogs were non-P450-mediated.


Subject(s)
Angiotensin II/metabolism , Angiotensin Receptor Antagonists , Antihypertensive Agents/metabolism , Antihypertensive Agents/pharmacokinetics , Imidazoles/metabolism , Imidazoles/pharmacokinetics , Pyridines/metabolism , Pyridines/pharmacokinetics , Animals , Antihypertensive Agents/administration & dosage , Area Under Curve , Biotransformation , Chromatography, High Pressure Liquid , Dogs , Half-Life , Imidazoles/administration & dosage , In Vitro Techniques , Male , Microsomes, Liver/metabolism , Pyridines/administration & dosage , Rats , Rats, Sprague-Dawley , Species Specificity , Spectrophotometry, Ultraviolet
2.
Drug Metab Dispos ; 26(3): 272-7, 1998 Mar.
Article in English | MEDLINE | ID: mdl-9492392

ABSTRACT

The tissue distribution of two 14C drugs were quantitatively compared using the techniques of whole body autoradioluminography (WBAL) and radiometry. Quantitative analysis of tissue radioactivity in whole body cryosections was accomplished from storage phosphor images using the MicroComputer Imaging Device. After obtaining whole body sections from four frozen rats and three frozen ferrets, each WBAL-sectioned specimen was partially thawed before obtaining tissue samples for radiometric analysis. For all tissues examined, concentrations of radioactivity determined by WBAL were comparable with those determined by dissection and liquid scintillation analysis (DLSA), except for renal tissue obtained from different kidneys of the same ferret and for rat ocular tissues. A 2-fold difference was observed between WBAL and DLSA evaluations of radioactivity in the contralateral kidneys of one ferret. DLSA evaluation only provided an assessment of total radioactivity in the eye, whereas WBAL evaluation determined the selective distribution of radioactivity to ocular tissues. Resolution in DLSA evaluation of ocular tissues was restricted by limitations of the dissection procedure. These results indicated that the quantitation of tissue radioactivity by WBAL was as precise as DLSA evaluation, and WBAL also provided results to the quantitative distribution of radioactivity to localized sites in organs not feasible by DLSA.


Subject(s)
Carbon Radioisotopes/analysis , Luminescent Measurements , Radiometry , Xenobiotics/analysis , Administration, Oral , Animals , Calibration , Cryoultramicrotomy , Female , Ferrets , Image Processing, Computer-Assisted , Male , Rats , Rats, Inbred Strains , Whole-Body Counting
3.
J Chromatogr B Biomed Sci Appl ; 696(2): 243-51, 1997 Aug 29.
Article in English | MEDLINE | ID: mdl-9323544

ABSTRACT

Simple and precise high-performance liquid chromatographic (HPLC) assays were developed and validated for the determination of a renin inhibitor (RI), 4-cyclohexyl-2-hydroxy-3- (3-methylsulfanyl-2-[2-[(morpholine-4-carbonyl)amino]- 3-phenylpropionylamino]propionylamino)butyric acid isopropyl ester (CP-80,794, I), and its hydrolytic cleavage metabolite, 2-[(morpholine-4-carbonyl)amino]-3-phenylpropionic acid (CP-84,364, II) in dog and human plasma. The internal standard for I, CP-83,092 (III, a carbon-substituted derivative of I) and analyte were extracted by liquid-liquid extraction using n-butyl chloride, cleaved to form a fragment containing a primary amine, and subsequently fluorescently derivatized for measurement by HPLC. Samples were analyzed by reversed-phase HPLC using a Waters C18 column with fluorescence detection at 390 nm excitation/440 nm emission. The quantitation limit of I was 10 ng/ml and the calibration curve was linear over the range of 0.01-1.0 microgram/ml (r2 > 0.99). In dog and human plasma, intra- and inter-assay precision ranged from 2.3 to 16% and 3.0 to 18%, respectively. The average recoveries were similar (> or = 63%) for both I and III and the upper limit of quantification of I can be as high as 4 micrograms/ml. The internal standard for II, CP-96,452 (IV, a methoxy derivative of II) and analyte were extracted by anion-exchange solid-phase extraction and subsequent liquid-liquid extraction using methyl tert.-butyl ether. Samples were analyzed by reversed-phase HPLC using a Waters C18 column with ultraviolet detection at 214 nm. The quantitation limit of II was 20 ng/ml and the calibration curve was linear over the range of 0.02-2.0 micrograms/ml (r2 > 0.99). In dog and human plasma, intra- and inter-assay precision ranged from 2.6 to 13.0% and 1.8 to 20.0%, respectively. The average recoveries were similar (> or = 75%) for both II and IV and the upper limit of quantification of II can be as high as 20 micrograms/ml. The methods described have been successfully applied to the quantification of I and II in about 5000 dog and human plasma samples over a 2 year period.


Subject(s)
Chromatography, High Pressure Liquid/methods , Dipeptides/blood , Morpholines/blood , Phenylalanine/analogs & derivatives , Renin/antagonists & inhibitors , Animals , Dogs , Humans , Phenylalanine/blood , Reference Standards , Reproducibility of Results , Sensitivity and Specificity , Spectrometry, Fluorescence
4.
J Pharmacol Exp Ther ; 272(2): 953-62, 1995 Feb.
Article in English | MEDLINE | ID: mdl-7853213

ABSTRACT

A novel method is described for quantitative whole-body autoradioluminography using [14C]-radioactive standards prepared from rat red blood cells. MicroComputer Imaging Device model 2 (MCID) and ImageQuant (IQ) imaging systems were evaluated for imaging performance and autoradioluminography quantitation. Weighted linear regression analysis resulted in linearity over five orders of magnitude with a lower limit of quantitation of 2.7 nCi/g. Using IQ, 16 days were necessary for image analysis and data processing of 30 whole-body cryosections and 1080 standards. MCID reduced the image and data processing of the same cryosections and standards to only 4 days. Embedding a series of radioactive standards with each specimen in the same carboxymethyl cellulose block provided an effective method of assessing intrasection and intersection variations in thickness of whole-body cryosections. These results demonstrated that autoradioluminography provided a sensitive, accurate, precise and reproducible method for the quantitative measurement of the tissue distribution of [14C]-radiolabeled xenobiotics in whole-body cryosections. Evaluating the biodistribution of [14C]-xenobiotics by autoradioluminography, not only provides pharmacokinetic data required for predicting the potential tissue deposition of an absorbed dose of radioactivity in man, but also allows for visual and quantitative evaluation of radioactivity in small anatomical structures that otherwise could not be detected or measured by conventional tissue combustion technology.


Subject(s)
Cryoultramicrotomy , Xenobiotics/pharmacokinetics , Animals , Autoradiography , Carbon Radioisotopes , Image Processing, Computer-Assisted , Male , Rats , Rats, Sprague-Dawley , Tissue Distribution
5.
J Cardiovasc Pharmacol ; 20(1): 75-82, 1992 Jul.
Article in English | MEDLINE | ID: mdl-1383634

ABSTRACT

The effects of coadministration of a renin inhibitor, CP-80,794, and an angiotensin converting enzyme inhibitor, captopril, on blood pressure of sodium-depleted guinea pigs was studied. Dose-response curves for CP-80,794 (0.3-3.0 mg/kg i.v.) and captopril (0.03-1.0 mg/kg i.v.) were obtained either alone or in the presence of a submaximal dose of the other inhibitor. The hypotensive response calculated for each compound individually was subtracted from the combined dose response. The results showed that statistically significant synergy with captopril and CP-80,794 occurred when the area rather than the peak drop or duration of change in blood pressure was measured. The degree of the synergy indicated that to achieve the same reduction in blood pressure, the dose of each drug, below the high end of its response range, could be decreased approximately sixfold when administered in combination. It was determined that the plasma pharmacokinetics of CP-80,794 were not altered during coadministration, as plasma concentrations of CP-80,794 were similar in the presence and absence of 0.1 mg/kg i.v. of captopril. These results indicate that by inhibiting sequential enzymes in the renin-angiotensin system, synergistic effects can be produced. The relative safety of each inhibitor could be improved by large reductions in dose when used concurrently.


Subject(s)
Blood Pressure/drug effects , Captopril/pharmacology , Dipeptides/pharmacology , Morpholines/pharmacology , Renin/antagonists & inhibitors , Analysis of Variance , Animals , Captopril/administration & dosage , Dipeptides/administration & dosage , Dipeptides/pharmacokinetics , Dose-Response Relationship, Drug , Drug Synergism , Guinea Pigs , Male , Morpholines/administration & dosage , Morpholines/pharmacokinetics
6.
Am J Vet Res ; 53(1): 129-33, 1992 Jan.
Article in English | MEDLINE | ID: mdl-1539904

ABSTRACT

The effect of feeding monensin, with or without dry hay plus wilted forage, on ruminal formation of 3-methylindole (3MI) was investigated in pastured cattle. Eighty-two cows were allotted to 3 groups. Cows of group-1 served as controls and were given a daily energy supplement (1 kg/head) without monensin for 1 day before and for 7 days after being allowed access to lush pasture. Cows of groups 2 and 3 were given the same daily energy supplement, which also contained monensin (200 mg/kg of supplement). Cows of group 3 also were fed dry hay for 5 days before the start of the study and continued to be given supplemental hay for 4 days after being allowed access to lush pasture containing a layer of wilted forage. Ruminal 3MI and indole concentrations increased on day 1 after all groups were allowed access to lush pasture. By day 7, 3MI concentration in all cows had decreased to pregrazing concentration. Indole concentration did not reach pregrazing concentration until day 10 for cows of groups 1 and 2. Group-3 cows had pregrazing indole concentration on day 7. Ruminal indole concentration did not differ (P greater than 0.05) between groups 1 and 2. Ruminal indole concentration was lower (P less than 0.01) in group-3 cows on all sample collection days, except day 10, compared with that in the other groups. Monensin reduced (P less than 0.01) 3MI formation on days 1 and 7 in group-2 cows, compared with group-1 cows. Group-3 cows had lower 3MI concentration than did group-1 cows (P less than 0.01) on days -1, 1, 4, and 7.(ABSTRACT TRUNCATED AT 250 WORDS)


Subject(s)
Animal Feed , Cattle Diseases/prevention & control , Monensin/therapeutic use , Rumen/metabolism , Skatole/metabolism , Acute Disease , Animals , Cattle , Digestion , Fatty Acids, Volatile/analysis , Female , Hydrogen-Ion Concentration , Indoles/analysis , Poaceae , Pulmonary Edema/prevention & control , Pulmonary Edema/veterinary , Pulmonary Emphysema/prevention & control , Pulmonary Emphysema/veterinary , Random Allocation , Rumen/chemistry , Skatole/analysis
7.
Eur J Pharmacol ; 173(1): 53-64, 1989 Nov 28.
Article in English | MEDLINE | ID: mdl-2606156

ABSTRACT

9-Amino-1,2,3,4-tetrahydroacridine (THA) has been reported to cause improvement in patients with senile dementia of the Alzheimer's type. We have examined some effects of THA in vitro and in vivo to define its mechanism of action. In vitro, THA inhibits acetylcholinesterase (AChE) (IC50 = 223 nM) and blocks [3H]AFDX-116 (M2) and [3H]telenzepine (M1) binding (IC50 s of 1.5 and 9.1 microM respectively). In vivo levels of THA were 10-fold higher in brain than plasma following 3.2 mg/kg i.p., a dose which was found to be active in reversing amnesia induced by scopolamine assessed in T-maze tests in rats and passive avoidance tests in mice. Additionally, these brain concentrations were above the IC50 of THA for AChE inhibition. THA (5.6-17.8 mg/kg i.p.) also elevated acetylcholine levels in the rat CNS. THA-induced side effects were blocked by the central muscarinic antagonist, scopolamine, but not by the peripheral antagonists methscopolamine and glycopyrrolate, nor by nicotinic antagonists. We conclude that brain AChE inhibition by THA is sufficient to explain its purported therapeutic activity in Alzheimer's disease and that its favorable brain/plasma distribution in vivo may account for its central cholinergic action without inducing the severe peripheral cholinergic effects typically seen with other AChE inhibitors.


Subject(s)
Aminoacridines/metabolism , Behavior, Animal/drug effects , Brain Chemistry/drug effects , Brain/metabolism , Tacrine/metabolism , Acetylcholinesterase/metabolism , Animals , Avoidance Learning/drug effects , Brain/enzymology , Cholinesterase Inhibitors , Chromatography, High Pressure Liquid , Eating/drug effects , Humans , In Vitro Techniques , Male , Memory/drug effects , Mice , Monoamine Oxidase/metabolism , Pain/physiopathology , Radioligand Assay , Rats , Rats, Inbred Strains , Spectrophotometry, Ultraviolet , Tacrine/pharmacokinetics , Tacrine/pharmacology
9.
Drug Metab Dispos ; 13(6): 690-4, 1985.
Article in English | MEDLINE | ID: mdl-2867872

ABSTRACT

Goat lung microsomes were incubated with glutathione (GSH) and 3-methylindole (3MI) to produce an adduct between GSH and an electrophilic metabolite of 3MI. The GSH-3MI adduct was purified by reverse-phase HPLC, and its structure elucidated by UV and NMR spectrometry and by thermospray LC/MS. The adduct was shown to be 3-[(glutathion-S-yl)-methyl]indole. Since nucleophilic GSH adds to the methyl position of 3MI without the incorporation of oxygen into the molecule, an epoxide metabolite is probably not the electrophilic intermediate. More likely, an imine methide intermediate, resulting from nitrogen oxidation and hydrogen abstraction from the methyl group by cytochrome P-450 monooxygenases, is the electrophilic intermediate. The imine methide electrophile is therefore proposed to be the toxic intermediate in 3MI-mediated pulmonary toxicity.


Subject(s)
Glutathione/metabolism , Imines/metabolism , Indoles/metabolism , Skatole/metabolism , Animals , Biotransformation , Chemical Phenomena , Chemistry, Physical , Chromatography, Liquid , Goats , In Vitro Techniques , Magnetic Resonance Spectroscopy , Mass Spectrometry , Microsomes, Liver/metabolism , Spectrophotometry, Ultraviolet
10.
Toxicol Lett ; 28(2-3): 79-87, 1985 Nov.
Article in English | MEDLINE | ID: mdl-4071564

ABSTRACT

Lung and liver microsomes of several species were evaluated for potential to form activated metabolites of 3-methylindole (3MI). Microsomes were incubated with [14C]3MI and glutathione (GSH). Electrophilic 3MI metabolites were trapped and quantitated as GSH adducts by HPLC, and by determining the amounts of activated intermediates which became covalently bound to microsomal protein. The highest rates of 3MI-GSH adduct formation by the lung were detected in microsomes of the goat, followed in decreasing order by pulmonary microsomes from the horse, monkey, mouse, and rat, respectively. In contrast, hepatic 3MI-GSH adduct production was highest in microsomes from the rat, followed by mouse, monkey, goat, and horse microsomes, respectively. These results suggest that the species and organ-selective toxicity of 3MI are primarily caused by differences in rates of oxidative metabolism of 3MI to an electrophilic intermediate.


Subject(s)
Glutathione/metabolism , Indoles/metabolism , Skatole/metabolism , Animals , Carbon Radioisotopes , Chlorocebus aethiops , Female , Goats , Horses , In Vitro Techniques , Lung/metabolism , Male , Mice , Microsomes/metabolism , Microsomes, Liver/metabolism , Organ Specificity , Rats , Rats, Inbred Strains , Species Specificity
11.
Toxicol Appl Pharmacol ; 81(1): 75-84, 1985 Oct.
Article in English | MEDLINE | ID: mdl-4049423

ABSTRACT

Incubations with goat lung and liver microsomes were conducted to trap with exogenous glutathione (GSH) the electrophilic intermediate produced via cytochrome P-450-dependent metabolic activation of 3-methylindole (3MI). Microsomal incubation mixtures with [14C]3MI, a NADPH-generating system, and [3H]GSH produced a dual-labeled adduct which was isolated by reverse-phase high-performance liquid chromatography. Reactive 3MI intermediates were also trapped with cysteine. Adduct formation increased in proportion to the concentration of either thiol. Covalent binding of activated 3MI metabolites to microsomal protein was inversely related to adduct production. There were both qualitative and quantitative differences in the formation of GSH adducts by lung and liver microsomes. In the presence of 2 mM GSH, the adduct was produced at a rate of 1.8 nmol/mg protein/min by lung microsomes but only at 0.1 nmol/mg protein/min by hepatic microsomes. The addition of cytosolic fractions containing glutathione S-transferase activity increased GSH adduct formation by approximately 30%. These results support the view that electrophilic 3MI intermediates are trapped by conjugation with GSH, and that organ-selective toxicity is primarily due to much faster rates of cytochrome P-450 oxidation of 3MI in the lung than in the liver.


Subject(s)
Alkylating Agents/metabolism , Glutathione/metabolism , Indoles/metabolism , Lung/metabolism , Microsomes, Liver/metabolism , Skatole/metabolism , Animals , Biotransformation , Cytochrome P-450 Enzyme System/pharmacology , Cytosol/metabolism , Female , Goats , In Vitro Techniques , Protein Binding
12.
J Anim Sci ; 60(1): 226-31, 1985 Jan.
Article in English | MEDLINE | ID: mdl-3972743

ABSTRACT

A series of in vitro and in vivo trials was conducted to determine if continuous monensin feeding for up to 56 d would reduce ruminal conversion of L-tryptophan (TRP) to 3-methylindole (3MI). Fourteen mature beef cows were adapted to a maintenance diet for 3 wk. In trial I, the sampling time to optimize 3MI production was determined. Trials II through IV were to determine the duration of efficacy of monensin on reducing 3MI concentrations in vitro and in vivo. During trials II, III and IV one-half of the cows were fed 200 mg monensin X head-1 X d-1 for 21, 36 and 55 d, respectively, while the remaining cows served as controls. All cows were fed the control diet for 21 d between each trial. Volatile fatty acid (VFA) concentrations and in vitro conversion of TRP to 3MI were determined in ruminal fluid samples collected during trials I through IV. On d 28 of trial IV, all cows were given an oral dose of .35 g TRP/kg of body weight to induce acute bovine pulmonary edema and emphysema (ABPE). Ruminal concentrations of 3MI and indole were measured at intervals for 96 h. Results of trial I demonstrated that ruminal fluid collected 15 h postfeeding produced the highest in vitro conversion of TRP to 3MI. Therefore, ruminal fluid samples were collected at that time in trials II, III and IV. In vitro conversion of TRP to 3MI was lower (P less than .01) in samples from monensin-treated cows (12.1%) compared with controls (25.6%). Monensin reduced 3MI production for 55 d, the longest time tested in these experiments.(ABSTRACT TRUNCATED AT 250 WORDS)


Subject(s)
Cattle/metabolism , Furans/pharmacology , Indoles/metabolism , Monensin/pharmacology , Rumen/metabolism , Skatole/metabolism , Animals , Cattle Diseases/chemically induced , Depression, Chemical , Fatty Acids, Volatile/metabolism , Female , Food Additives , In Vitro Techniques , Pulmonary Edema/chemically induced , Pulmonary Edema/veterinary , Pulmonary Emphysema/chemically induced , Pulmonary Emphysema/veterinary , Skatole/toxicity , Tryptophan/metabolism
13.
J Anim Sci ; 60(1): 232-8, 1985 Jan.
Article in English | MEDLINE | ID: mdl-3972744

ABSTRACT

A study was conducted to determine the dose of lasalocid that would effectively reduce ruminal conversion of tryptophan (TRP) to 3-methylindole (3MI) and prevent the development of acute bovine pulmonary edema and emphysema (ABPE). After adaptation to a maintenance diet for 3 wk, 20 mature beef cows were randomly divided into four groups of five cows each and fed 0, 200, 400 or 600 mg lasalocid X head-1 X d-1 in .5 kg ground barley for the 12-d experimental period. In vitro conversion of TRP to 3MI and indole by ruminal fluid and volatile fatty acid (VFA) concentrations were determined on d 0, 2, 4, 6 and 12. On d 6, an oral dose of .35 g TRP/kg body weight was given to induce ABPE, and ruminal production of 3MI and indole was determined at intervals thereafter. Formation of 3MI was sharply reduced (P less than .01) both in vitro and in vivo by lasalocid treatment at 200 mg X head-1 X d-1. Further suppression of 3MI production occurred as the lasalocid dose was increased (P less than .05). Linear (P less than .0001) and quadratic (P less than .002) components were determined for the relationship between lasalocid dose and 3MI production. Indole formation was variable, but tended to increase (P less than .05) with increasing lasalocid dose. Cows that received no lasalocid developed moderate to severe clinical signs of ABPE and three cows died of acute lung disease. Lasalocid treatment at all levels prevented ABPE. Lasalocid decreased ruminal acetate and butyrate, and increased propionate concentration (P less than .01).(ABSTRACT TRUNCATED AT 250 WORDS)


Subject(s)
Cattle Diseases/prevention & control , Indoles/metabolism , Lasalocid/pharmacology , Pulmonary Edema/veterinary , Pulmonary Emphysema/veterinary , Rumen/metabolism , Skatole/metabolism , Animals , Cattle , Depression, Chemical , Fatty Acids, Volatile/metabolism , Female , Food Additives , Lasalocid/administration & dosage , Pulmonary Edema/prevention & control , Pulmonary Emphysema/prevention & control , Tryptophan/metabolism
14.
Toxicol Lett ; 24(1): 25-32, 1985 Jan.
Article in English | MEDLINE | ID: mdl-3975927

ABSTRACT

Autoradiographs of horse-lung explants incubated with [3H]3-methylindole (3MI) showed 8 times greater labeling per area to bronchiolar epithelial cells than to the interalveolar septa. Incubations of horse-lung microsomes with [14C]3MI resulted in alkylation of microsomal proteins, which could be reduced by exogenous glutathione. An apparent covalent adduct of glutathione and 3MI was isolated from these incubations. These results suggest that the target cells of 3MI-induced injury in the horse, the bronchiolar epithelial cells, are alkylated by an electrophilic 3MI intermediate.


Subject(s)
Bronchi/cytology , Indoles/metabolism , Skatole/metabolism , Alkylation , Animals , Autoradiography , Bronchi/pathology , Chromatography, High Pressure Liquid , Epithelium/metabolism , Glutathione/metabolism , Horses , Microsomes/metabolism
15.
Xenobiotica ; 14(7): 561-4, 1984 Jul.
Article in English | MEDLINE | ID: mdl-6506769

ABSTRACT

[methyl-14C]-3-Methylindole (3MI) was incubated with goat-lung microsomes, an NADPH-generating system and glutathione. An adduct between an oxidative metabolite of 3MI and glutathione was formed only when the complete system was employed. The adduct, which was detected by u.v. absorbance and scintillation counting of h.p.l.c. fractions, was purified to homogeneity by reverse-phase h.p.l.c. The ability of 3MI to bind to microsomal protein was reduced to 52% and 46% of controls when 2 mM and 4 mM glutathione, respectively, were included in the incubations. These results suggest the involvement of an electrophilic metabolite as the toxic intermediate in 3MI-mediated pulmonary oedema.


Subject(s)
Indoles/metabolism , Lung/metabolism , Microsomes/metabolism , Pulmonary Edema/veterinary , Skatole/metabolism , Animal Diseases/chemically induced , Animals , Cattle , Glutathione/metabolism , Goats , NADP/metabolism , Pulmonary Edema/chemically induced , Sheep , Skatole/toxicity
16.
Proc Soc Exp Biol Med ; 176(1): 48-53, 1984 May.
Article in English | MEDLINE | ID: mdl-6709651

ABSTRACT

Covalent binding of 3-[14C]methylindole (3[14C]MI) in crude microsomal preparations of goat lung, liver, and kidney was measured to determine if a reactive intermediate was formed during the in vitro metabolism of 3-methylindole (3MI). The bound radioactivity was highest in lung compared to liver and kidney. The amount of bound radioactivity per nanomole of cytochrome P-450 was approximately 10 times higher in the lung compared to liver. No detectable bound radioactivity was found when 3-[3H]methyloxindole was used as the substrate. Cofactor requirements and the effects of inhibitors indicate that a mixed function oxidase (MFO) system is involved in formation of a reactive intermediate. Inhibitors and conjugating agents that are known to reduce the severity of 3MI-induced lung injury such as piperonyl butoxide (MFO inhibitor) and glutathione (conjugating agent) significantly decreased the in vitro binding of 3[14C]MI. The results indicate that a reactive intermediate is produced during the metabolism of 3MI by the MFO system. The organ specificity in binding suggests that covalent binding by lung microsomes may be related to the mechanism of 3MI-induced lung injury.


Subject(s)
Goats/metabolism , Indoles/metabolism , Skatole/metabolism , Animals , Cytochrome P-450 Enzyme System/metabolism , Kidney/metabolism , Liver/metabolism , Lung/metabolism , Microsomes/metabolism , Mixed Function Oxygenases/antagonists & inhibitors , Mixed Function Oxygenases/metabolism
17.
Am J Vet Res ; 45(7): 1389-92, 1984 Jul.
Article in English | MEDLINE | ID: mdl-24049904

ABSTRACT

The metabolism of 3-methylindole (3MI), a ruminal degradation product of L-tryptophan, results in acute bovine pulmonary edema and emphysema. The effect of feeding an energy or protein supplement containing monensin on ruminal 3MI formation in pastured beef cattle was investigated. A luxuriant pasture of orchard grass was established in a field that was seeded 1 year before the start of the grazing period. This 4-ha pasture was cut, fertilized, divided into 2 equal plots, and then irrigated during a 22-day growth period. All cows were fed a restricted quantity of low-quality alfalfa hay for 33 days before the grazing period. Two experiments were conducted, using 38 cows (30 of the cows were used in experiment I and all 38 cows were used in experiment II). Cows in each experiment were randomly allotted to 2 groups. One group was designated in each experiment as the control group. The control group for experiment I was fed an energy supplement. The control group for experiment II was fed a protein supplement. The 2nd group in each experiment was given the same supplement as the respective control group with 200 mg of monensin added/! kg of feed. Supplements were fed on days - 1, 0, 1, 2, 3, 4, 5, 6, and 7 of each experimental period. Supplements were fed twice daily to provide 1 kg of supplement/cow. Cows were given access to orchard grass pasture on day 0 of each experiment. Ruminal fluid was collected daily for analysis of 3MI, indole, and volatile fatty acids. Ruminal fluid pH was recorded immediately after collection. Ruminal pH of all cows decreased from 7.3 to 6.2 during the first few days of grazing the orchard grass. Ruminal pH then gradually increased toward neutrality by experimental day 10. Significantly (P < 0.01) higher molar percentages of pro-pionate and lower (P < 0.01) molar percentages of acetate and butyrate were observed in the 2 groups fed the supplements with added monensin. These changes in propionate and acetate remained different (P < 0.01) from those of the controls for 10 days (or 3 days after the last monensin feeding). Compared with pregrazing ruminal concentrations of 3MI, the 3MI values were elevated (P < 0.01) by day 1 in all groups, except in the monensin-treated cows of experiment I. In experiment I, 3MI concentrations were highest on experimental days 5 and 10 in control and monensin-treated cows, respectively. In experiment II, 3MI concentrations peaked on day 4 for the control cows and day 6 for the monensin-treated cows. Monensin supplementation reduced (P < 0.05) 3MI formation on days 1 through 5 in experiment I and on days 1 through 3 in experiment II. Formation of 3MI was increased in ruminal fluid of all cows after an abrupt change to the pasture forage, but the rate of 3MI production was slower, and a lower peak concentration of 3MI was reached in cows fed monensin than was observed in the controls. These results indicate that monensin administration in either an energy or protein supplement effectively reduced ruminal 3MI formation in pasture-fed cattle.


Subject(s)
Cattle/physiology , Dietary Proteins/administration & dosage , Dietary Supplements , Monensin/pharmacology , Rumen/metabolism , Animal Feed/analysis , Animal Nutritional Physiological Phenomena , Animals , Diet/veterinary , Dietary Proteins/analysis , Energy Intake , Female , Monensin/chemistry , Rumen/chemistry , Skatole/pharmacology
18.
Life Sci ; 32(5): 449-58, 1983 Jan 31.
Article in English | MEDLINE | ID: mdl-6823204

ABSTRACT

A study was conducted to investigate the relationship between glutathione (GSH) status, in-vivo metabolite covalent binding and 3-methylindole (3MI)-induced lung injury in goats. Cysteine or diethylmaleate pretreatments were given to sustain or deplete GSH, respectively, prior to intravenous 14C-3MI administration. Cysteine pretreatment prolonged survival times, decreased (P less than .05) covalent lung injury. Diethylmaleate pretreatment shortened survival times, increased (P less than .05) covalent binding and enhanced lung injury. Covalent binding was higher (P less than .05) in lung compared to liver and kidney. 3-Methylindole alone depleted GSH in 4 hours to 36, 66, and 75% of controls in these tissues, respectively. The relationship between tissue GSH, covalent binding and toxicity supports the hypothesis that 3MI-induced pneumotoxicity results from the formation of activated intermediates and that GSH plays a role in detoxication of these 3MI metabolites.


Subject(s)
Glutathione/metabolism , Indoles/toxicity , Lung/pathology , Skatole/toxicity , Animals , Cysteine/pharmacology , Goats , Lung/drug effects , Lung/metabolism , Maleates/pharmacology , Skatole/metabolism
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