Species differences in adrenal lipid peroxidation: role of alpha-tocopherol

Previous reports have noted high levels of lipid peroxidation (LP) in vitro in a variety of adrenocortical preparations. However, we have observed that susceptibility to adrenal LP seems to vary considerably from species to species. The current study was done to confirm these apparent species differences in adrenal LP in vitro and to determine if they were attributable to differences in alpha-tocopherol content. Incubation of mitochondrial or microsomal preparations from guinea pig or rabbit adrenal glands with ferrous ion (Fe2+) caused a time-dependent increase in the formation of thiobarbituric acid reactive substances (TBARS) accompanied by depletion of alpha-tocopherol. By contrast, incubation of adrenal mitochondria or microsomes from rats or monkeys with Fe2+ had little or no detectable effect on TBARS and basal adrenal alpha-tocopherol levels were five to ten-fold greater than those in guinea pigs or rabbits. In addition, there was little change in alpha-tocopherol concentrations during incubation of rat or monkey adrenal tissue. Dietary alpha-tocopherol deficiency in rats reduced adrenal alpha-tocopherol to concentrations approximating those in guinea pigs. Incubation with Fe2+ induced high levels of TBARS in adrenal mitochondria and microsomes from the alpha-tocopherol deficient rats. Conversely, dietary alpha-tocopherol supplementation in rabbits increased adrenal alpha-tocopherol levels and prevented Fe2+ induced TBARS formation in mitochondria and microsomes. The results indicate that there are large species differences in adrenal susceptibility to LP in vitro and that these differences are at least partly attributable to species differences in adrenal alpha-tocopherol concentrations.     

Fatty acid profiles and lipid peroxidation of microsomes and mitochondria from liver, heart and brain of Cairina moschata

Studies were done to analyze the fatty acid composition and sensitivity to lipid peroxidation (LP) of mitochondria and microsomes from duck liver, heart and brain. The fatty acid composition of mitochondria and microsomes was tissue-dependent. In particular, arachidonic acid comprised 17.39+/-2.32, 11.75+/-3.25 and 9.70+/-0.40% of the total fatty acids in heart, liver and brain mitochondria respectively but only 13.39+/-1.31, 8.22+/-2.43 and 6.44+/-0.22% of the total fatty acids in heart, liver and brain microsomes, respectively. Docosahexahenoic acid comprised 17.02+/-0.78, 4.47+/-1.02 and 0.89+/-0.07% of the total fatty acids in brain, liver and heart mitochondria respectively but only 7.76+/-0.53, 3.27+/-0.73 and 1.97+/-0.38% of the total fatty acids in brain, liver and heart microsomes. Incubation of organelles with ascorbate-Fe(2+) at 37 degrees C caused a stimulation of LP as indicated by the increase in light emission: chemiluminescence (CL) and the decrease of arachidonic acid to: 5.17+/-1.34, 8.86+/-0.71 and 5.86+/-0.68% of the total fatty acids in heart, liver and brain mitochondria, respectively, and to 4.10+/-0.61 in liver microsomes. After LP docosahexahenoic acid decrease to 7.29+/-1.47, 1.36+/-0.18 and 0.30+/-0.11% of the total fatty acids in brain, liver and heart mitochondria. Statistically significant differences in the percent of both peroxidable fatty acids (arachidonic and docosahexaenoic acid) were not observed in heart and brain microsomes and this was coincident with absence of stimulation of LP. The results indicate a close relationship between tissue sensitivity to LP in vitro and long chain polyunsaturated fatty acid concentration. Nevertheless, any oxidative stress in vitro caused by ascorbate-Fe(2+) at 37 degrees C seems to avoid degradation of arachidonic and docosahexaenoic acids in duck liver and brain microsomes. It is possible that because of the important physiological functions of arachidonic and docosahexaenoic acids in these tissues, they are protected to maintain membrane content during oxidative stress.     

Non-enzymatic lipid peroxidation of microsomes and mitochondria from liver, heart and brain of the bird Lonchura striata: relationship with fatty acid composition

The aim of this study was to examine the fatty acid composition and non-enzymatic lipid peroxidation (LP) of mitochondria and microsomes obtained from liver, heart and brain of Lonchura striata. The percentage of total unsaturated fatty acid was approximately 30-60% in the organelles from all tissues studied. Brain mitochondria and both organelles of liver exhibited the highest percentage of polyunsaturated fatty acid (PUFA) (30 and 18%, respectively). The arachidonic acid (AA) content was 7% in mitochondria of liver and brain and 3% in heart mitochondria. The percentage of docosahexanoic acid (DHA) was 8% in brain mitochondria and approximately 2-3% in heart and liver mitochondria. The peroxidizability index (PI) of brain mitochondria and both organelles from liver was higher than that of organelles from heart and brain microsomes. Liver organelles and brain mitochondria were affected by LP, as indicated by the increase in chemiluminescence and a decrease of AA and DHA. These changes were not observed during LP of brain microsomes and both organelles from heart. These results indicate: 1) PI positively correlates with PUFA percentage and LP; 2) The resistance to LP detected in heart organelles would contribute to the cardiac protection against oxidative damage.     

Intestinal absorption and postabsorptive metabolism of linoleic acid in rats with short-term bile duct ligation

We investigated in bile duct-ligated (BDL) and sham-operated control rats whether the frequent presence of essential fatty acid deficiency in cholestatic liver disease could be related to linoleic acid malabsorption, altered linoleic acid metabolism, or both. In plasma of BDL rats, the triene-to-tetraene ratio, a biochemical marker for essential fatty acid deficiency, was increased compared with controls (0.024 +/- 0.004 vs. 0.013 +/- 0.001; P < 0.05). Net and percentage of dietary linoleic acid absorbed were decreased in BDL rats compared with control rats (1.50 +/- 0.16 mmol/day and 81.3 +/- 3.3% vs. 2.08 +/- 0.07 mmol/day and 99.2 +/- 0.1%, respectively; each P < 0.001). At 24 h after [(13)C]linoleic acid administration, BDL rats had a similar ratio of plasma [(13)C]arachidonic acid to plasma [(13)C]linoleic acid concentration compared with control rats. Delta(6)-Desaturase activity was not significantly different in hepatic microsomes from control or BDL rats. At 3 h after [(13)C]linoleic acid administration, plasma appearance of [(13)C]linoleic acid and cumulative expiration of (13)CO(2) were decreased in BDL rats, compared with controls (by 54% and 80%, respectively). The present data indicate that the impaired linoleic acid status in cholestatic liver disease is mainly due to decreased net absorption and not to quantitative alterations in postabsorptive metabolism.     

Antioxidant effect of conjugated linoleic acid and vitamin A during non enzymatic lipid peroxidation of rat liver microsomes and mitochondria

In the study reported here the effect of conjugated linoleic acid (CLA) and vitamin A on the polyunsaturated fatty acid composition, chemiluminescence and peroxidizability index of microsomes and mitochondria isolated from rat liver was analyzed. The effect of CLA on the polyunsaturated fatty acid composition of native microsomes was evidenced by an statistically significant p < 0.007 decrease of linoleic acid C18:2 n6, whereas in mitochondria it was observed a decrease p < 0.0001 of arachidonic acid C20:4 n6 when compared with vitamin A and control groups. Docosahexaenoic acid C22:6 n3 in mitochondria was reduced p < 0.04 in CLA and vitamin A groups when compared with control. After incubation of microsomes or mitochondria in an ascorbate (0.4 mM)-Fe⁺⁺ (2.15 M) system (120 min at 37°C) it was observed that the total cpm/mg protein originated from light emission: chemiluminescence was lower in liver microsomes or mitochondria obtained from CLA group (received orally: 12.5 mg/daily during 10 days) than in the vitamin A group (received intraperitoneal injection: daily 0.195 g/kg during 10 days). CLA reduced significantly maximal induced chemiluminescence in microsomes relative to vitamin A and control groups, whereas in mitochondria the effect was observed relative to control group The polyunsaturated fatty acid composition of liver microsomes or mitochondria changed by CLA and vitamin A treatment. The polyunsaturated fatty acids mainly affected when microsomes native and peroxidized from control group were compared were linoleic, linolenic and arachidonic acids, while in vitamin A group linoleic and arachidonic acid were mainly peroxidized, whereas in CLA group only arachidonic acid was altered. In mitochondria obtained from the three groups arachidonic acid and docosahexaenoic acid showed a significant decrease when native and peroxidized groups were compared. As a consequence the peroxidizability index, a parameter based on the maximal rate of oxidation of fatty acids, show significant changes in the CLA group compare vitamin A and control groups. The simultaneous analysis of peroxidizability index, chemiluminescence and fatty acid composition demonstrated that CLA is more effective than vitamin A protecting microsomes or mitochondria from peroxidative damage.     

The effect of alpha-tocopherol on lipid peroxidation of microsomes and mitochondria from rat testis

The testis is a remarkably active metabolic organ; hence it is suitable not only for studies of lipid metabolism in the organ itself but also for the study of lipid peroxidation processes in general. The content of fatty acids in testis is high with a prevalence of polyunsaturated fatty acids (PUFA) which renders this tissue very susceptible to lipid peroxidation. Studies were carried out to evaluate the effect of alpha-tocopherol in vitro on ascorbate-Fe(++) lipid peroxidation of rat testis microsomes and mitochondria. Chemiluminescence and fatty acid composition were used as an index of the oxidative destruction of lipids. Special attention was paid to the changes produced on the highly PUFA [C20:4 n6] and [C22:5 n6]. Lipid peroxidation of testis microsomes or mitochondria induced a significant decrease of both fatty acids. Total chemiluminescence was similar in both kinds of organelles when the peroxidized without (control) and with ascorbate-Fe(++) (peroxidized) groups were compared. Arachidonic acid was protected more efficiently than docosapentaenoic acid at all alpha-tocopherol concentrations tested when rat testis microsomes or mitochondria were incubated with ascorbate-Fe(++). The maximal percentage of inhibition in both organelles was approximately 70%; corresponding to an alpha-tocopherol concentration between 1 and 0.25 mM. IC50 values from the inhibition of alpha-tocopherol on the chemiluminescence were higher in microsomes (0.144 mM) than mitochondria (0.078 mM). The protective effect observed by alpha-tocopherol in rat testis mitochondria was higher compared with microsomes, associated with the higher amount of [C20:4 n6]+[C22:5 n6] in microsomes that in mitochondria. It is proposed that the vulnerability to lipid peroxidation of rat testis microsomes and mitochondria is different because of the different proportion of PUFA in these organelles The peroxidizability index (PI) was positively correlated with the level of long chain fatty acids. The results demonstrated the protective effect of alpha-tocopherol on lipid peroxidation in microsomes and mitochondria from rat testis.     

Modulation of the effects of ascorbic acid on lipid peroxidation by tocopherol in adrenocortical mitochondria

Studies were done to evaluate the role of alpha-tocopherol in modulating the effects of ascorbic acid (AA) on lipid peroxidation (LP) by adrenocortical mitochondria. In control mitochondria from the inner (zona reticularis) or outer (zona fasciculata plus zona glomerulosa) zones of the guinea pig adrenal cortex, subphysiological concentrations of AA stimulated LP but higher levels had little or no effect. However, after depletion of adrenal tocopherol, even physiological concentrations of AA exerted prooxidant effects, stimulating LP. To assess the antioxidant potency of AA, its effects to inhibit ferrous ion (Fe2+)-induced LP were determined. Mitochondria from the outer zone contained far more alpha-tocopherol than those from the inner zone and were more sensitive to the antioxidant effects of AA. After tocopherol depletion, the antioxidant potency of AA in outer zone mitochondria decreased, but there was little change in the inner zone. The results indicate that the actions of AA are determined in part by mitochondrial tocopherol content, and, as a result, vary in the different zones of the adrenal cortex.     

Purification and Properties of a Mitochondrial Lipoprotein Inhibitor of Sterol Synthesis

A lipoprotein inhibitor of hydroxymethylglutaryl CoA reductase (EC 1.1.1.34) and of cholesterol synthesis by rat liver homogenates, was isolated from the mitochondria of starved rats’ livers. The isolated lipoprotein complex contained a low molecular weight protein and fatty acids. The fatty acids identified were arachidonic, linoleic, oleic, stearic and palmitic. The saturated fatty acids and oleic acid did not inhibit. Inhibition of the enzyme was to a large extent related to the degree of fatty acid unsaturation.     

A unique cardiac cytosolic acyltransferase with preferential selectivity for fatty acids that form cyclooxygenase/lipoxygenase metabolites and reverse essential fatty acid deficiency

The rabbit heart contains a cytosolic enzyme which selectively incorporates polyunsaturated fatty acids into phosphatidylcholine. This unique acyltransferase is selective for fatty acids, thus far tested, that are substrates for cyclooxygenase or lipoxygenase (i.e., arachidonic, eicosapentaenoic, linoleic and dihomo-gamma-linoleic acids) or which reverse the symptoms of essential fatty acid deficiency (columbinic acid). On the other hand, palmitic, oleic, 5,8,11-eicosatrienoic (n-9, Mead acid), and docosatetraenoic acid (n-6, adrenic acid) were not incorporated in phospholipids by the cytosolic acyltransferase. No such fatty acid selectivity was exhibited by the cytosolic acyl-CoA synthetase or by the acyltransferase activities present in cardiac microsomes and mitochondria.     

High-cholesterol diets induce changes in lipid composition of rat erythrocyte membrane including decrease in cholesterol,increase in alpha-tocopherol and changes in fatty acids of phospholipids

Effects of high dietary cholesterol on erythrocyte membrane lipids were studied. Feeding rats with a diet containing 0.5% cholesterol and 0.15% sodium cholate for two weeks induced changes in erythrocyte membrane lipids including a decrease in cholesterol, an increase in alpha-tocopherol (alpha-Toc) and changes in the fatty acid composition of phospholipids. Oleic acid and linoleic acid increased, while arachidonic acid decreased in phosphatidylcholine. Saturated fatty acids decreased and unsaturated fatty acids increased in phosphatidylethanolamine. Almost the same changes in membrane lipids were also noted after six weeks of feeding rats with the diet. A diet containing 0.5% cholesterol but without sodium cholate caused a decrease in erythrocyte cholesterol and an increase in erythrocyte alpha-Toc after two weeks of feeding, as compared to the basal diet, indicating that high dietary cholesterol, but not sodium cholate, was responsible for these changes in the erythrocyte membrane.     

Comparative studies on lipid peroxidation of microsomes and mitochondria obtained from different rat tissues: effect of retinyl palmitate

The effect of retinyl palmitate on the polyunsaturated fatty-acid composition, chemiluminescence and peroxidizability index of microsomes and mitochondria obtained from rat liver, kidney, brain, lung and heart, was studied. After incubation of microsomes and mitochondria in an ascorbate Fe++ system (120 min at 37 degrees C) it was observed that the total cpm/mg protein originated from light emission: chemiluminescence was lower in liver microsomes, mitochondria and kidney microsomes in the vitamin A group than in the control group. In mitochondria obtained from control rats, the most sensitive fatty acids for peroxidation were arachidonic acid C20:4 n6 in liver and docosahexaenoic acid C22:6 n3 in kidney and brain. In microsomes obtained from control rats, the most sensitive fatty acids for peroxidation were linoleic acid C18:2 n6 and C20:4 n6 in liver and C22:6 n3 in kidney. Changes in the most polyunsaturated fatty acids were not observed in organelles obtained from lung and heart. As a consequence the peroxidizability index, a parameter based on the maximal rate of oxidation of fatty acids, showed significant changes in liver, kidney and brain mitochondria, while in microsomes changes were significant in liver and kidney. These changes were less pronounced in membranes derived from rats receiving vitamin A. Our results confirm and extend previous observations that indicated that vitamin A may act as an antioxidant protecting membranes from deleterious effects.     

The effect of alpha-tocopherol on the lipid peroxidation of mitochondria and microsomes obtained from rat liver and testis.

The effect of intraperitoneal administration of alpha-tocopherol (100 mg/kg wt/24 h) on ascorbate (0.4 mM) induced lipid peroxidation of mitochondria and microsomes isolated from rat liver and testis was studied. Special attention was paid to the changes produced on the highly polyunsaturated fatty acids C20:4 n6 and C22:6 n3 in liver and C20:4 n6 and C22:5 n6 in testis. The lipid peroxidation of liver mitochondria or microsomes produced a significant decrease of C20:4 n6 and C22:6 n3 in the control group, whereas changes in the fatty acid composition of the alpha-tocopherol treated group were not observed. The light emission was significantly higher in the control than in the alpha-tocopherol treated group. The lipid peroxidation of testis microsomes isolated from the alpha-tocopherol group produced a significant decrease of C20:4 n6 , C22:5 n6 and C22:6 n3, these changes were not observed in testis mitochondria. The light emission of both groups was similar. The treatment with alpha-tocopherol at the dose and times indicated showed a protector effect on the polyunsaturated fatty acids of liver mitochondria, microsomes and testis mitochondria, whereas those fatty acids situated in testis microsomes were not protected during non enzymatic ascorbate-Fe2+ lipid peroxidation. The protector effect observed by alpha-tocopherol treatment in the fatty acid composition of rat testis mitochondria but not in microsomes could be explained if we consider that the sum of C20:4 n6 + C22:5 n6 in testis microsomes is 2-fold than that present in mitochondria.     

Cytochrome P-450 lowering effect of alkyl halides,correlation with decrease in arachidonic acid

Treatment of male rats with carbon tetrachloride, bromotrichloromethane, chloroform, 1,2-dibromoethane, 1-bromo-2-chloroethane, and 1,2-dibromo-3-chloropropane results in a decrease in cytochrome P-450 content and alterations in the relative content of fatty acids in hepatic microsomes. A high correlation was found between the loss of cytochrome P-450, the decrease in arachidonic acid (r=0.93), and the increases in linoleic (r=?0.91) and oleic acids (r=?0.89).     

Effects of alpha-tocopherol and tocotrienols on blood pressure and linoleic acid metabolism in the spontaneously hypertensive rat (SHR).

Both alpha-tocopherol and a 1:1.7 mixture of alpha-tocopherol and tocotrienols at a 0.2% dietary level significantly depressed the age-related increase in the systolic blood pressure of spontaneously hypertensive rats (SHRs) after 3 weeks of feeding. The aortic production of prostacyclin was increased 1.5 times both by alpha-tocopherol and a tocotrienol mixture, suggesting a possible relevance to their hypotensive effect. These vitamins did not influence the delta 6- and delta 5-desaturase activities of liver microsomes, but fatty acid profiles of the liver phospholipids predicted a reduction of linoleic acid desaturation. These effects were in general more clear with tocotrienols than with alpha-tocopherol. Platelet aggregation by 5 microM ADP remained uninfluenced. Thus, tocotrienols may have effects on various lipid parameters somewhat different from those of alpha-tocopherol.     

Regional differences in microsomal lipid peroxidation and antioxidant levels in the guinea pig adrenal cortex

Lipid peroxidation (LP) and antioxidant levels were studied in the chromatically distinct inner (zona reticularis) and outer (zona fasciculata + zona glomerulosa) zones of the guinea pig adrenal cortex. Ferrous ion (Fe2+) produced a concentration-dependent (10(-5) to 10(-3) M) stimulation of microsomal LP in both zones, but LP, as estimated by malonaldehyde production, was far greater in the inner zone. Although cytosolic ascorbic acid content was similar in the two zones, microsomal tocopherol levels were approx 4 times greater in the outer than inner zone. Subphysiological concentrations of ascorbic acid, like Fe2+, initiated LP to a greater extent in inner than outer zone microsomes; optimal stimulation of LP by ascorbic acid occurred at concentrations of 100-200 microM in both zones. Physiological concentrations of ascorbic acid (1-5 mM), by contrast, did not initiate LP and, in fact, markedly inhibited Fe2+-induced LP in both inner and outer zone microsomal preparations. Outer zone microsomes were more sensitive to the antioxidant effects of ascorbic acid than were inner zone preparations. Addition of alpha-tocopherol to inner zone microsomal suspensions inhibited Fe2+-induced LP. The results indicate that there are regional differences in adrenocortical LP which may be caused by differences in tocopherol content. alpha-Tocopherol may serve important antioxidant functions within the adrenal cortex, thereby contributing to the functional zonation of the gland.     

Relationship between mitochondrial lipid peroxidation and alpha-tocopherol levels in the guinea-pig adrenal cortex

Lipid peroxidation in mitochondria from the functionally distinct inner (zona reticularis) and outer (zona fasciculata + zona glomerulosa) zones of the guinea-pig adrenal cortex was investigated. Ferrous ion (Fe2+)-induced lipid peroxidation was far greater in inner than outer zone mitochondria. Ascorbic acid similarly initiated lipid peroxidation to a greater extent in inner zone mitochondrial preparations. Differences in the unsaturated fatty acid content of inner and outer zone mitochondria could not account for the regional differences in lipid peroxidation. Total fatty acid concentrations were greater in the outer than in the inner zone, and the relative amounts of each fatty acid were similar in the two zones. However, mitochondrial concentrations of alpha-tocopherol, an antioxidant known to inhibit lipid peroxidation, were approx. 5-times greater in the outer than inner zone. The results demonstrate that there are regional differences in mitochondrial lipid peroxidation in the adrenal cortex which may be attributable to differences in alpha-tocopherol content. Thus, alpha-tocopherol may serve to protect outer zone mitochondrial enzymes from the consequences of lipid peroxidation and thereby contribute to some of the functional differences between the zones of the adrenal cortex.     

Effects of adrenocorticotropic hormone and dexamethasone on adrenal and hepatic alpha-tocopherol concentrations

Studies were done to determine the effects of ACTH treatment on adrenal alpha-tocopherol (alpha-T) concentrations in female rats. Administration of dexamethasone (DEX) to inhibit endogenous ACTH secretion increased whole adrenal alpha-T levels as well as the fractional amount in adrenal cytosol. Adrenal ascorbic acid (AA) concentrations were unaffected by DEX. DEX treatment also had no effect on hepatic AA content but decreased alpha-T concentrations in the liver. The subcellular distribution of alpha-T in the liver was not altered by DEX. Administration of ACTH to DEX-treated animals decreased adrenal alpha-T content and restored the pattern of subcellular distribution to that seen in controls. ACTH had no effect on hepatic alpha-T concentrations or subcellular distribution. ACTH treatment also had no effect on AA concentrations in adrenals or livers. The results demonstrate that ACTH has a role in the regulation of adrenal alpha-T but the mechanism(s) involved remain to be determined. The data also indicate that glucocorticoids such as DEX directly influence hepatic alpha-T levels independent of their effects on ACTH secretion.     

Chronologic studies of the effects of a hypoproteinic diet followed by an equilibrated diet on delta-6 and delta-5 desaturations of linoleic acid in liver microsomes in the rat

A low protein diet affects amounts of linoleic and arachidonic acids in hepatic microsomal phospholipids of growing rats. Are the changes related to modifications in microsomal delta 6- and delta 5- linoleic acid desaturase activities? Two groups of Wistar rats weighing 80 +/- 5 g at the beginning of the experiment were used: Control group (T) was fed on a 16% gluten + 4% casein diet for 53 days; Experimental group (E) was fed on a 4% gluten + 1% casein diet for 26 days (MP) then Control diet for 27 days (RE). After 2, 14 and 26 days of MP and 2, 15 and 27 days of RE, rats of each group were sacrificed. Protein and water contents of liver, quantitative fatty acid, composition of total lipids in liver and hepatic microsomes were determined. delta 6- and delta 5- linoleic acid desaturase activities were estimated from incubation of liver microsomes with [1-14C] C 18: 2 n-6 or [2(14)C] C 20: 3 n-6 respectively. The low protein diet stops practically ponderal growth. The fatty-acid compositions of microsomal total lipids of E rats were affected in comparison with values of T rats. These modifications persist after 27 days of RE. The C 20: 4 n-6/C 18: 2 n-6 ratio in microsomal total lipids was slightly different between T and E rats but increased strongly during refeeding. Same modifications take place in the fatty-acid composition of hepatic total lipids. After two days of MP, delta 6- and delta 5- desaturase activities were depressed, phenomenon that not persist in the course of MP. These enzyme activities increase to higher values than those of the T after two days of RE.     

Changes in n-6 and n-3 polyunsaturated fatty acids during lipid-peroxidation of mitochondria obtained from rat liver and several brain regions: effect of alpha-tocopherol

The effect of intraperitoneal administration of alpha-tocopherol (100 mg/kg weight/24 h) on ascorbate (0-0.4 mM) induced lipid peroxidation of mitochondria isolated from rat liver, cerebral hemispheres, brain stem and cerebellum was examined. The ascorbate induced light emission in hepatic mitochondria was nearly completely inhibited by alpha-tocopherol (control-group: 114.32+/-14.4; vitamin E-group: 17.45+/-2.84, c.p.m.x10(-4)). In brain mitochondria, 0.2 mM ascorbate produced the maximal chemiluminescence and significant differences among both groups were not observed. No significant differences in the chemiluminescence values between control and vitamin E treated groups were observed when the three brain regions were compared. The light emission produced by mitochondrial preparations was much higher in cerebral hemispheres than in brain stem and cerebellum. In liver and brain mitochondria from control group, the level of arachidonic acid (C20:4n6) and docosahexaenoic acid (C22:6n3) was profoundly affected. Docosahexaenoic in liver mitochondria from vitamin E group decreased by 30% upon treatment with ascorbic acid when compared with mitochondria lacking ascorbic acid. As a consequence of vitamin E treatment, a significant increase of C22:6n3 was detected in rat liver mitochondria (control-group: 6.42 +/-0.12; vitamin E-group: 10.52 +/-0.46). Ratios of the alpha-tocopherol concentrations in mitochondria from rats receiving vitamin E to those of control rats were as follows: liver, 7.79; cerebral hemispheres, 0.81; brain stem, 0.95; cerebellum, 1.05. In liver mitochondria, vitamin E shows a protector effect on oxidative damage. In addition, vitamin E concentration can be increased in hepatic but not in brain mitochondria. Lipid peroxidation mainly affected, arachidonic (C20:4n6) and docosahexaenoic (C22:6n3) acids.     

The effects of unsaturated fatty acids on hepatic microsomal drug metabolism and cytochrome P-450

1. The effects of unsaturated fatty acids on drug-metabolizing enzymes in vitro were measured by using rat and rabbit hepatic 9000g supernatant fractions. 2. Unsaturated fatty acids inhibited the hepatic microsomal metabolism of ;type I' drugs with inhibition increasing with unsaturation: arachidonic acid>linolenic acid>linoleic acid>oleic acid. Inhibition was independent of lipid peroxidation. Linoleic acid competitively inhibited the microsomal O-demethylation of p-nitroanisole and the N-demethylation of (+)-benzphetamine. 3. The hepatic microsomal metabolism of ;type II' substrates, aniline and (-)-amphetamine, was not affected by unsaturated fatty acids. 4. The rate of reduction of p-nitrobenzoic acid and Neoprontosil was accelerated by unsaturated fatty acids. 5. Linoleic acid up to 3.5mm did not decelerate the generation of NADPH by rat liver soluble fraction, nor the activity of NADPH-cytochrome c reductase of rat liver microsomes. Hepatic microsomal NADPH oxidase activity was slightly enhanced by added linoleic acid. 6. No measurable disappearance of exogenously added linoleic acid occurred when this fatty acid was incubated with rat liver microsomes and an NADPH source. 7. The unsaturated fatty acids used in this study produced type I spectra when added to rat liver microsomes, and affected several microsomal enzyme activities in a manner characteristic of type I ligands.