Effects of cage beddings on microsomal oxidative enzymes in rat liver

The purpose of the present studies was to evaluate the effects of some commercially available cage beddings on rat liver microsomal cytochrome P-450-dependent drug-metabolizing enzyme, ethylmorphine N-demethylase, and the carcinogen-metabolizing enzyme, benzo(a)pyrene hydroxylase. Sprague-Dawley rats were housed in cages containing cedar chip, corncob or heat-treated pinewood bedding for 3 weeks. Control rats were housed in cages on wire bottom floors containing no bedding material. Rats housed in cages containing cedar chip showed 18, 46 and 49% increases in liver cytochrome P-450 content, ethylmorphine N-demethylase and benzo(a)pyrene hydroxylase activities, respectively. The liver enzyme activities of rats housed in cages containing corncob bedding were similar to those obtained with control rats. In contrast, the pinewood-bedded rats showed a 21% decrease in ethylmorphine N-demethylase activity without affecting cytochrome P-450 content and benzo(a)pyrene hydroxylase activity. Hexobarbital-induced sleep times of the variously bedded rats were similar to those of control animals. These data suggest that the commercial bedding materials differ in their abilities to affect liver microsomal enzymes. Thus, interlaboratory variability in basal enzyme activities reported in the literature may be partly due to bedding materials used in the animal's cages.     

Participation of a cytochrome P450 enzyme from the 2C subfamily in progesterone 21-hydroxylation in sheep liver.

Progesterone 21-hydroxylation in hepatic microsomes from adult male sheep is a quantitatively important metabolic pathway (0.27 +/- 0.08 nmol deoxycorticosterone formed/min/mg protein; representing 13-25% of total progesterone conversion). This study was undertaken to determine whether the ovine hepatic progesterone 21-hydroxylase may be another member of the P450 2C subfamily, normally associated with progesterone 21-hydroxylation in rodent liver. An IgG preparation raised in rabbits against purified rat liver microsomal cytochrome P450 2C6 was found to recognize a single antigen (MW 52 kDa) in sheep liver microsomes. This protein was present in sheep liver (apparent concentration 16 +/- 4 ng/micrograms microsomal protein) representing approx. 28% of the corresponding content of P450 2C6 in untreated rat liver. Preincubation of the anti-P450 2C6 IgG with hepatic microsomes was found to decrease the rate of progesterone 21-hydroxylation to 50-80% of uninhibited control. Taken together, from these findings it is apparent that a P450 enzyme, most likely from the 2C subfamily, catalyses deoxycorticosterone formation from progesterone in sheep liver and that this is a quantitatively important pathway of progesterone hydroxylation in these fractions.     

Characterization of sheep liver and lung microsomal ethylmorphine N-demethylase

Ethylmorphine N-demethylase activity of the sheep liver and lung microsomes was reconstituted in the presence of solubilized microsomal cytochrome P-450, NADPH-cytochrome c reductase and synthetic lipid, phosphatidylcholine dilauroyl. The Km of the lung microsomal ethylmorphine N-demethylase was calculated to be 4.84 mM ethylmorphine from its Lineweaver-Burk graph and lung enzyme was inhibited by its substrate, ethylmorphine, when its concn was 25 mM and above, reaching to 67% inhibition at 50 mM concn. The Lineweaver-Burk and Eadie-Hofstee plots of the liver enzyme were found to be curvilinear. From these graphs, two different Km values were calculated for the liver enzyme as 4.17 mM and 0.40 mM ethylmorphine. Ethylmorphine N-demethylase activities of both liver and lung microsomes were inhibited by NiCl2, CdCl2 and ZnSO4. Ethylalcohol inhibited N-demethylation of ethylmorphine in lung and liver microsomes. Acetone (5%) slightly enhanced the N-demethylase activity of the liver enzyme, whereas 5% acetone completely inhibited the lung enzyme. Phenylmethylsulfonyl fluoride at 0.10 mM and 0.25 mM concn had no effect on liver enzyme activity, while at these concns, it inhibited the activity of the lung enzyme by about 35%.     

Regulation of cytochrome P-450j, a high-affinity N-nitrosodimethylamine demethylase, in rat hepatic microsomes

Polyclonal antibodies were produced in rabbits against purified cytochrome P-450j isolated from isoniazid-treated adult male rats. The monospecificity of immunoadsorbed antibody to cytochrome P-450j was demonstrated by Ouchterlony double diffusion analyses, enzyme-linked immunosorbent assays, and immunoblots. Immunoquantitation results indicated that rat liver microsomal cytochrome P-450j content decreases between 3 and 6 weeks of age in both the male and female animal. Several xenobiotics, such as Aroclor 1254, mirex, and 3-methylcholanthrene, repressed cytochrome P-450j levels when administered to male rats. Isoniazid, dimethyl sulfoxide, pyrazole, 4-methylpyrazole, and ethanol were inducers of cytochrome P-450j in rat liver although these compounds showed different inducing potencies. Microsomes from adult male rats with chemically induced diabetes also contained elevated levels of cytochrome P-450j compared to untreated animals. Cytochrome P-450j levels were measurable in kidney, whereas this isozyme was barely detectable in lung, ovaries, and testes; however, extrahepatic cytochrome P-450j was inducible by isoniazid. Approximately 80-90% of microsomal N-nitrosodimethylamine demethylation was inhibited by antibody to cytochrome P-450j whether the microsomes were isolated from untreated rats or animals administered inducers or repressors of cytochrome P-450j. The residual catalytic activity resistant to antibody inhibition may be a reflection of the inaccessibility of a certain amount of cytochrome P-450j due to interference by NADPH-cytochrome P-450 reductase based on results obtained with the reconstituted system. There was a good correlation (r2 = 0.87) between cytochrome P-450j content and N-nitrosodimethylamine demethylase activity in microsomes from rats of different ages and treated with various xenobiotics. The evidence presented indicates that cytochrome P-450j is the primary, and perhaps sole, microsomal catalyst of N-nitrosodimethylamine demethylation at substrate concentrations relevant to hepatocarcinogenesis induced by N-nitrosodimethylamine.     

The effects of ascorbic acid deficiency and repletion on pulmonary, renal, and hepatic drug metabolism in the guinea pig.

The effects of ascorbic acid (AA) deficiency on microsomal and soluble (postmicrosomal supernatant) enzymes which catalyze drug metabolism were studied in the guinea pig liver, lung, and kidney, (i) Twenty-one days of AA depletion produced a 50–60% decrease in hepatic cytochrome P-450 levels, 20–30% decreases in renal levels, but no significant changes in pulmonary cytochrome P-450 content. Upon repletion of ascorbic acid, recovery to control levels occurred within 7 days. (ii) The decreases in hepatic cytochrome P-450 in scurvy were not accompanied by a corresponding increase in cytochrome P-420. (iii) Aminopyrine N-demethylation decreased by 40% in livers of deficient animals, and recovered within 3 days, but there were no corresponding changes in lungs and kidneys. (iv) There were no significant alterations of NADPH-cytochrome c reductase activity in scorbutic animals in any of the three organs. (v) Activity of “native” UDP-glucuronyl transferase was increased in liver microsomes after 21 days of deficiency, but this apparent increase was not observed when the enzyme was fully activated in vitro with UDP N-acetylglucosamine. “Native” UDP-glucuronyl transferase was increased in kidneys of deficient animals and unchanged in lungs. (vi) In the postmicrosomal supernatant, glutathione S-aryl transferase activity in deficient livers decreased tc 50% of control and did not fully recover after 14 days of ascorbic acid repletion. These changes were not seen in kidney and lung. (vii) Also in the postmicrosomal supernatant, p-aminobenzoic acid (PABA) N-acetyl transferase activity increased in the kidneys of deficient animals, but was unchanged in liver and lungs. (viii) Addition of ascorbic acid in vitro to hepatic microsomes prepared from scorbutic animals had no effect on activities of aminopyrine N-demethylase, NADPH-cytochrome c reductase, PABA N-acetyl transferase, and glutathione S-aryl transferase.     

Influence of neonatal and postnatal administration of diethylstilbestrol on hepatic monooxygenase activities and lipid peroxidation in adult rats.

Treatment of adult male and female rats with DES caused a significant reduction of cytochrome P450 concentration, ethylmorphine N-demethylation and ethoxycoumarin O-deethylation. These decreases are not modified by a neonatal treatment with this hormone. In contrast to monooxygenase activities the NADPH-induced lipid peroxidation shows only a tendency to decrease if animals received DES as adults. But a decrease of about 50% is observed after "neonatal plus treatment of adults" with DES. The role of formation of reactive oxygen species characterized by chemiluminescence methods in connection with lipid peroxidation is discussed.     

In vivo effects of 3-methylcholanthrene, phenobarbital, pyrethrum and 2,4,5-T isooctylester on liver, lung and kidney microsomal mixed-function oxidase system of guinea-pig: a comparative study

The optimum conditions (pH, microsomal protein amount and substrate concentration) of guinea-pig liver, lung and kidney microsomal aniline 4-hydroxylase, ethylmorphine N-demethylase and benzo[a]pyrene hydroxylase activities were determined. Male guinea-pigs weighing 500-700 g were administered 3-methylcholanthrene (25 mg/kg, i.p. 3 days), phenobarbital (75 mg/kg, i.p. 3 days), pyrethrum (120 mg/kg, i.p. 2 days) and 2,4,5-T isooctylester (200 mg/kg, i.p. 3 days). 3-Methylcholanthrene treatment caused significant increases in liver microsomal benzo[a]pyrene hydroxylase and kidney microsomal aniline 4-hydroxylase activities. However, with phenobarbital treatment the only significant increase was observed in liver microsomal ethylmorphine N-demethylase activity. Pyrethrum treatment decreased kidney microsomal ethylmorphine N-demethylase activity significantly. 2,4,5-T isooctylester treatment increased liver microsomal aniline 4-hydroxylase and lung microsomal ethylmorphine N-demethylase activities significantly. Liver microsomal NADPH-cytochrome c reductase activity was increased significantly by phenobarbital and pyrethrum treatment. The other treatments did not cause any significant changes in microsomal NADPH-cytochrome c reductase activities of liver, lung and kidney. Cytochrome P-450 content of guinea-pig liver microsomes were increased significantly about 2.5-fold and 2-fold by treatment with 3-methylcholanthrene and phenobarbital, respectively. 3-Methylcholanthrene also caused 1 nm spectral shift in the absorption maxima of CO difference spectrum of the dithionite-reduced liver microsomal cytochrome P-450, forming P-449.     

Electrophoretic, spectral, catalytic and immunochemical properties of highly purified cytochrome P-450 from sheep lung

1. Cytochrome P-450LgM2 was purified from sheep lung microsomes in the presence of detergents, Emulgen 913 and cholate. 2. The purification procedure involved the chromatography of the detergent solubilized microsomes on DEAE-cellulose and hydroxylapatite. 3. Cytochrome P-450LgM2 was further purified on second DEAE-cellulose and hydroxylapatite columns. 4. The specific content of the highly purified P-450LgM2 was 16-18 nmol P-450/mg protein and purified 164-fold. 5. The yield was 16% of the initial content in microsomes. 6. The SDS-polyacrylamide slab gel electrophoresis (PAGE) of the purified lung cytochrome P-450LgM2 showed one protein band having the monomer molecular weight of 49,500. 7. The absolute CO-difference spectrum of dithionate-reduced P-450LgM2 gave a peak at 451 nm. 8. When sheep lung cytochrome P-450LgM2 and P-450LM2 purified from liver of phenobarbital (PB)-induced rabbit were subjected to Western Blotting and visualized immunochemically with anti-P-450LM2, they showed identical mobilities. 9. P-450LgM2 was found to be very active in N-demethylation of benzphetamine in a reconstituted system containing purified sheep lung reductase and synthetic lipid. 10. Turnover numbers (min-1) for benzphetamine, aniline, ethylmorphine and p-nitrophenol were determined to be 273, 1.2, 15.5 and 1.05, respectively, in a reconstituted microsomal lung monooxygenase system. 11. Spectral, electrophoretic, biocatalytic and immunochemical properties of sheep lung P-450LgM2 were found to be similar to those of P-450 isozyme 2, purified from PB-treated rabbit liver and of rabbit lung microsomes.     

A conspicuous down-regulating effect of morphine on essential steroid hydroxylation reactions and certain drug N-demethylations.

This study was conducted to explore the potency of morphine to induce reductions of specific cytochrome P450 isoenzyme functions. Male Sprague-Dawley rats were treated with escalating doses (20-125 mg/kg per day) of morphine for 2 weeks in order to study the effects on the following cytochrome P450 catalyzed reactions: 16 alpha-hydroxylation of dehydroepienderosterone (DHA) and progesterone; 17 alpha- and 21-hydroxylation of progesterone; N-demethylation of ethymorphine, codeine and morphine as well as O-dealkylation of ethylmorphine and codeine. 16 alpha-Hydroxylation of DHA and progesterone and 17 alpha-hydroxylation of progesterone decreased to 18, 12 and 10% of control activities, respectively. The N-demethylation of ethylmorphine and codeine decreased to 34 and 43% of control activities, respectively. Morphine treatment had no effect on the 21-hydroxylation reactions or the O-dealkylation of ethylmorphine or codeine. A monoclonal antibody (Mab) against rat liver cytochrome P450 2 c/RLM 5 exerted a 66-73% inhibition of the N-demethylation of ethylmorphine and codeine, respectively, whereas the O-dealkylation reactions were not affected. This Mab inhibited the 16 alpha- and 17 alpha-hydroxylation of DHA and progesterone, whereas the 21-hydroxylation reactions were unaffected. The steroid hydroxylation reactions in rat adrenals were not altered upon morphine treatment. Our data suggest that a major part of the 16 alpha- and 17 alpha-steroid hydroxylations are catalyzed by the same (or closely related) cytochrome(s) P450 as the opioid N-demethylation reactions.     

Liver microsome phospholipids and cytochrome P.450 conceptration in phenobarbital treated rats fed on different diets

Liver microsomal concentration of cytochrome P.450 is increased in animals which are fed diets rich in polyunsaturated fatty acids. On the other hand, the effects of phenobarbital are more important when the dietary fat is more unsaturated. The unsaturation index in liver microsomal phosphatidylcholines depends on the unsaturation of the dietary fats. The treatment with phenobarbital constantly results in a decrease of the unsaturation index of fatty acids both in lecithins and cephalins. The importance of the liver microsomal cytochrome P.450 increase and the importance of the unsaturation index decrease in liver microsomal lecithins, both promoted by phenobarbital, are in good agreement.     

Induction of drug-metabolizing enzymes and aryl hydrocarbon hydroxylase by microscope immersion oil

Microscope immersion oil when administered intraperitoneally or applied to skin in experimental animals substantially increased liver weight, microsomal protein, NADPH-cytochrome c reductase activity, cytochrome P-450 content and the metabolism of the model substrates, ethylmorphine and benzo(a)pyrene. Immersion oil caused the induction of the polycyclic hydrocarbon type of hemoprotein, cytochrome P-448. When applied to skin, the oil also caused an 11-fold increase in benzo(a)pyrene hydroxylase activity at the skin sites.     

Partial hepatectomy of marmoset: clinical and pathological effects and utility in microsomal enzyme analysis.

Liver biopsy based on a partial hepatectomy technique (shearing) was performed in 10 common marmosets (Callithrix jacchus). This is a preliminary study to evaluate the effects of drugs on hepatic microsomal enzymes: cytochrome P-450 and T4 uridine diphosphate glucuronyl transferase (T4-UDPGT), by comparing post-treatment values with pre-treatment values individually with a limited number of animals. The effects of the biopsy on clinical findings and liver pathology were evaluated during the first 5 post-surgical weeks. Although the plasma aspartate aminotransferase (AST) activities tended to decrease from 1 to 4 weeks post-surgery, no abnormality was noted in clinical sign, body weight, the hematocrit value or other blood chemical values. At necropsy, adhesion of the sheared site of the liver to the parietal peritoneum or the small intestine was evident in 2 of the 4 marmosets. Microscopic examination revealed focal fibrosis in the liver, but it was localized around the sheared site. Based on the above results, it was concluded that liver biopsy must be performed more than one month before administration of the drug to be tested. The biopsy samples and the whole liver samples obtained at autopsy were subjected to analysis of microsomal protein content, cytochrome P-450 content and T4-UDPGT activity. In comparison with the values from the whole liver samples, those from the biopsy samples showed no significant difference. Furthermore, there was a significant correlation rather than difference between matched values. This suggested that partial hepatectomy is a useful method for obtaining pretreatment values in liver biochemistry to evaluate the effects of drug-treatment in individual animals.     

Modulation of rat liver cytochrome P450 activity by prolonged red wine consumption

Cytochrome P450-dependent oxidation of lauric acid, p-nitrophenol and ethanol by liver microsomal fractions were studied in control rats and in animals given either ethanol, red wine, or alcohol-free red wine for 10 weeks. Ethanol increased the total cytochrome P450 and the isoenzyme 2E1 content, as well as the p-nitrophenol hydroxylation and ethanol oxidation. These effects of ethanol treatment were attenuated by red wine administration. Red wine increased the total antioxidant capacity of plasma, whereas the alcohol-free red wine decreased the cytochrome P450 content and decreased the oxidation of lauric acid, p-nitrophenol and ethanol to values lower than control. It is concluded that red wine administration attenuates the ethanol-induced enhancement in liver microsomal parameters dependent on cytochrome P450 2E1 activity, an affect that seems to be accomplished by the non-alcoholic constituents of red wine known to have antioxidant properties.     

The NADPH-dependent cytochrome P-450 reduction in liver microsomes of rats of different ages with and without phenobarbital pretreatment.

The activity of NADPH-cytochrome P-450 reductase in liver microsomes of 10- to 60-day-old rats was determined. Neither the half life time of cytochrome P-450 reduction nor the absolute amount of cytochrome P-450 reduced per time unit depend on age. Phenobarbital pretreatment enhances the reduction rate in all age groups. The addition of hexobarbital or ethylmorphine to microsomal suspension accelerates the reduction of cytochrome P-450 in some age groups only. Age differences corresponding to developmental changes in drug-metabolizing activities are not detectable. The NADPH-cytochrome P-450 reductase seems to be not responsible for the age dependence of drug metabolism.     

Characterization of cytochrome P-450-dependent activities in hagfish, dogfish, perch and spectacle caiman

1. Owing to interest in the original function and evolution of cytochrome P-450 this enzyme system was studied in four species each at different levels on the phylogenetic tree, representing Agnatha, Selachi, Teleostei, and Reptilia. 2. All species contained considerable amounts of hepatic microsomal cytochrome P-450. The content in dogfish, perch and spectacle caiman was 0.2-0.3 while in the dogfish it was 0.6 nmol per mg protein. 3. Hepatic microsomal cytochrome P-450 dependent ethylmorphine demethylase, benzo(a)pyrene hydroxylase, 7-ethoxycoumarin-O-deethylase and 7-ethoxyresorufin-O-deethylase activities have been measured and the assays were carefully characterized with respect to pH-, protein content-, incubation time- and temperature-activity relationships. 4. The study revealed species-dependent differences in the liver microsomal cytochrome P-450 enzyme system, which could be due to differences in cytochrome P-450 composition or in the microenvironment around the enzyme.     

Effects of in vivo benzo(a)pyrene treatment on liver microsomal mixed-function oxidase activities of gilthead seabream (Sparus aurata)

Benzo(a)pyrene [B(a)P] treatment of gilthead seabream, 25 mg/kg, i.p. for 5 consecutive days, did not cause any significant changes in ethylmorphine N-demethylase and aniline 4-hydroxylase activities of liver microsomes. The same treatment did not alter the liver microsomal cytochrome b5 content, NADH-cytochrome b5 reductase and NADPH-cytochrome P450 reductase activities. However, benzo(a)pyrene treatment caused a 2–3-fold increase in 7-ethoxyresorufin O-deethylase (7-EROD) activity of gilthead seabream liver microsomes. Although, upon treatment, total cytochrome P450 content of liver microsomes increased about 1.7-fold in 1990 fall, no such increase was observed in spring 1991. However, a new cytochrome P450 with an apparent M_r of 58,000 was observed on SDS-PAGE of liver microsomes obtained from benzo(a)pyrene treated gilthead seabream. Besides, in vitro addition of 0.2 × 10⁻⁶ M benzo(a)pyrene to the incubation mixture inhibited 7-ethoxyresorufin O-deethylase activity by 93%. Gilthead seabream liver microsomal 7-ethoxyresorufin O-deethylase activity was characterized with respect to substrate concentration, amount of enzyme, type of buffer used, incubation period and temperature.     

Test systems for biomonitoring based on membrane-bound enzyme complexes. V. Mixed-function monooxygenase induction in the liver microsomes of Lake Baikal fish

The content of cytochrome P450 and monooxygenase activity has been studied in the liver of Baikal fishes (Coregonus automnalis, Thymallus articus, Brachymystax lenok and Cottocomphorus greminsky). The administration of 3-methylcholanthrene increases considerably the level of metabolic activity of microsomal fraction and cytochrome P450 content in liver. The data of microsomal fractions of rats and fishes liver electrophoresis have shown that xenobiotic causes the synthesis of similar according to the molecular weight forms of cytochrome P450 in these animals. The induction of microsomal monooxygenase inhibits the lipid peroxidation of microsomal fraction.     

Induction of cytochrome P450 by toluene

At least six cytochrome P450 (P450) isoenzymes, including CYP1A1/2, CYP2A1, CYP2B1/2, CYP2C6, CYP2C11 and CYP2E1, are involved in the metabolism of toluene in rat liver. Toluene exposure induces CYP1A1/2, CYP2B1/2, CYP2E1 and CYP3A1, but decreases CYP2C11/6 and CYP2A1 in adult males. Both sex and age influence the induction of P450s by toluene: in general, the inductive effect is more prominent in younger than in older animals; in males than in females. Neonatal exposure to toluene causes significant changes in liver microsomal P450 dependent monooxygenase activities during the early stage of life, whereas the enects on the rats of more than 3 weeks of age are small. Although structurally related chemicals of toluene also influence similar hepatic P450 isoenzymes, the degree of CYP2B1/2 induction increases, whilst that of CYP2E1 decreases with increasing molecular weight and aliphatic moieties. Unlike liver, exposure to toluene does not influence the distribution of pulmonary or renal microsomal P450-related enzyme activity in rats. In humans, occupational exposure to toluene is so low that it could not lead to the induction of P450. However, the induction may be seen in toluene sniffers who are exposed to high concentrations.     

Formation of cytochrome P-450 containing haem or cobalt-protoporphyrin in liver homogenates of rats treated with phenobarbital and allylisopropylacetamide.

The potent porphyrogen allylisopropylacetamide and related compounds decrease hepatic concentrations of cytochrome P-450. This decrease occurs particularly in phenobarbital-induced cytochrome P-450 and is caused by suicidal breakdown of the haem of cytochrome P-450. Quantitative rocket immunoelectrophoresis showed that the protein moiety of the major phenobarbital-inducible form of hepatic cytochrome P-450 was not diminished up to 1 h, but was markedly decreased (to 43% of that of the phenobarbital-treated control) at 20 h after allylisopropylacetamide treatment. In contrast, the concentration of total cytochrome P-450, measured spectrophotometrically, decreased to 30-40% of the control at both 1 and 20 h after allylisopropylacetamide. Cytochrome P-450-dependent demethylations of ethylmorphine and benzphetamine decreased to a similar extent. When liver homogenates from rats treated with allylisopropylacetamide 1 h before being killed were incubated with haem, functional holocytochrome P-450 could be reconstituted from the apoprotein. Incubation with haem increased spectrophotometrically measurable cytochrome P-450 to 69%, ethylmorphine demethylase to 64% and benzphetamine demethylase to 93% of the activities in rats treated with phenobarbital alone. At 20 h after allylisopropylacetamide treatment, however, little or no reconstitution of cytochrome P-450 occurred after incubation with haem. When liver homogenates were incubated with cobalt and protoporphyrin, and microsomal proteins were then subjected to polyacrylamide-gel electrophoresis, cobalt-protoporphyrin was found specifically associated with proteins of Mr 50 000-53 000. When homogenates from rats given allylisopropylacetamide for 1 h or 20 h were compared, it was found that the extent of this association was higher in livers from the rats containing more apocytochrome P-450, suggesting that cobalt-protoporphyrin had associated with the apocytochrome. The data provide insight into the association of haem with the protein moiety of cytochrome P-450 and factors affecting breakdown of this protein.     

A model for the regulation of δ-aminolaevulinate synthetase induction in rat liver

A reciprocal relationship exists between the cytochrome P-450 content and delta-aminolaevulinate synthetase activity in adult rats. In young rats the basal delta-aminolaevulinate synthetase activity is higher and the cytochrome P-450 content is lower compared with the adult rat liver. Administration of allylisopropylacetamide neither induces the enzyme nor causes degradation of cytochrome P-450 in the young rat liver, unlike adult rat liver. Allylisopropylacetamide fails to induce delta-aminolaevulinate synthetase in adrenalectomized-ovariectomized animals or intact animals pretreated with successive doses of the drug, in the absence of cortisol. The cortisol-mediated induction of the enzyme is sensitive to actinomycin D. Allylisopropylacetamide administration degrades microsomal haem but not nuclear haem. Haem does not counteract the decrease in cytochrome P-450 content caused by allylisopropylacetamide administration, but there is evidence for the formation of drug-resistant protein-bound haem in liver microsomal material under these conditions. Phenobarbital induces delta-aminolaevulinate synthetase under conditions when there is no breakdown of cytochrome P-450. On the basis of these results and those already published, a model is proposed for the regulation of delta-aminolaevulinate synthetase induction in rat liver.