Dietary Intake of Medium- and Long-chain Triacylglycerols Prevents the Progression of Hyperglycemia in Diabetic ob/ob Mice.

We examined the effects of the dietary intake of medium- and long-chain triacylglycerol (MLCT) on hyperglycemia in diabetic ob/ob mice. Six-week-old male ob/ob mice were fed a diet containing longchain triacylglycerol (LCT) or MLCT for 3 wks. During the dietary treatment, we determined the plasma glucose and insulin concentrations in the fed state once a week. Whereas the body weights did not differ between the two groups, the total intra-abdominal fat mass was significantly higher in the MLCT group compared to the LCT group. The plasma glucose levels in the freely fed state gradually increased during the 3-wk dietary treatment in the LCT but not MLCT group, although the daily food intake did not differ between the two groups. In the fed state, the MLCT group's plasma glucose was significantly lower and their insulin concentrations were significantly higher than those observed in the LCT group (p<0.01). Plasma glucose concentrations at the end of dietary treatment (3rd wk) were negatively correlated with plasma insulin concentrations (p<0.05) and tended to be inversely related to total intra-abdominal fat mass (p=0.08). These results suggest that the dietary intake of MLCT may delay the progression of hyperglycemia in ob/ob mice, possibly through the stimulation of glucose uptake in intra-abdominal fat tissue caused by enhanced insulin secretion.

Medium-chain triacylglycerol suppresses the decrease of plasma albumin level through the insulin-Akt-mTOR pathway in the livers of malnourished rats.

Recent studies have shown that medium-chain triacylglycerol (MCT) improved serum albumin concentration in elderly people with protein-energy malnutrition (PEM) and in malnourished rats. However, the mechanism for this effect has not been clarified. Dietary MCT promotes insulin secretion from the pancreas, and insulin activates mammalian target of rapamycin (mTOR) complex 1 (mTORC1) via the activation of phosphoinositide 3-kinase (PI3K) and its downstream effecter, Akt. mTORC1 promotes mRNA translation through S6K and 4E-BP1. Therefore, we hypothesized that dietary MCT elevates albumin synthesis through promotion of insulin-Akt-mTOR transduction in the liver. To test this hypothesis, we measured phosphorylated Akt, mTOR and albumin in the livers of malnourished rats. In the present study we examined rats fed low-protein diets containing either MCT or long-chain triacylglycerol (LCT) with energy restriction. The plasma and liver albumin levels were significantly higher in the MCT-fed group than in the LCT-fed group. In addition, plasma insulin concentration, liver phosphorylated Akt/Akt and phosphorylated mTOR/mTOR levels were significantly higher in the MCT-fed group than in the LCT-fed group. These results suggest that one of the mechanisms for the albumin improvement effect of dietary MCT is the promotion of albumin synthesis through the insulin-Akt-mTOR signaling pathway of the liver.

Dietary intake of medium- and long-chain triacylglycerols ameliorates insulin resistance in rats fed a high-fat diet.

OBJECTIVE: Excessive accumulation of visceral fat is strongly associated with insulin resistance. The present investigation examined the effects of dietary intake of medium- and long-chain triacylglycerols (MLCTs), which have been shown to induce significantly lower visceral fat accumulation in rats and humans, on high-fat diet-induced obesity and insulin resistance in rats. These effects were then compared with those observed in long-chain triacylglycerol (LCT)-fed rats. METHODS: After an 8-wk feeding of a high-fat diet, which induced severe whole-body insulin resistance, male Sprague-Dawley rats were fed a standard diet containing LCTs or MLCTs for 6 wk. After the dietary treatment, an oral glucose tolerance test was performed. RESULTS: Although body weight and total intra-abdominal fat mass did not differ between the two groups, mesenteric fat weight in the MLCT-fed group was significantly lower than that in the LCT group (P < 0.05). The increase in plasma insulin concentrations, but not in glucose, after glucose administration (area under the curve) was significantly smaller in the MLCT group than in the LCT group (P < 0.01) and was significantly associated with mesenteric fat weight (P < 0.05). MLCT-fed rats had significantly higher plasma adiponectin concentrations compared with LCT rats (P < 0.05). Adiponectin concentrations were negatively correlated with the area under the curve for plasma insulin (P < 0.05) and tended to be inversely related to mesenteric fat weight (P = 0.08). CONCLUSION: These results suggest that dietary intake of MLCTs may improve insulin resistance in rats fed a high-fat diet, at least in part through increased adiponectin concentrations caused by a lower mesenteric fat mass.

Compensatory expression of MRP3 in the livers of MRP2-deficient EHBRs is promoted by DHA intake.

We have hypothesized a suppressive mechanism against dietary docosahexaenoic acid (22:6n-3; DHA)-induced tissue lipid peroxidation, in which the degradation products, including their conjugates, are excreted into the urine by xenobiotic or organic anion transporters. In this study, we employed parent-strain Sprague-Dawley rats (SDRs), together with their mutant strain, Eisai hyperbilirubinuria rats (EHBRs). EHBRs are deficient in multidrug resistance-associated protein (MRP) 2, and show defective urinary excretion of numerous xenobiotics and organic anions. Both strains of rats were fed a diet containing DHA at 8.4% of total energy for 31 d. In the livers of the DHA-fed rats, the level of free malondialdehyde (MDA) + 4-hydroxy-2-alkenals (HAE) fell, and conversely glutathione S-transferase (GST) activity increased in MRP2-deficient EHBRs as compared to the SDRs, suggesting that the glutathione (GSH)-conjugation reaction for the aldehydes generated on DHA intake was accelerated in the MRP2-deficient EHBRs. Since the gene expression of liver MRP3 in the MRP2-deficient EHBRs was amplified to compensate for DHA intake, it is thought that the transport of MRP3 substrates into the bloodstream, rather than MRP2-mediated excretion of its substrates into the bile, was promoted. Indeed, excretion of mercapturic acid (acetylcysteine conjugates derived metabolically from the conjugate of each aldehyde with GSH) into the urine increased significantly in MRP2-deficient EHBRs fed DHA.

Effect of life-long dietary n-6/n-3 fatty acid ratio on life span, serum lipids and serum glucose in Wistar rats.

Types of dietary lipid affect the life span of rats. In this study, we investigated the influence of the life-long dietary n-6/n-3 ratio on life span and serum lipid and glucose levels. A semi-purified diet adjusted to a constant saturated : monounsaturated : polyunsaturated fatty acid ratio and an n-6/n-3 ratio of 1 (R1), 4 (R4) or 16 (R16) was fed to rats (n=33) from 4 wk of age until death. There were no significant differences in the food intake or body weight, nor were there survival curve or mean life span variations among the 3 groups. The serum cholesterol levels after feeding the test diet for 6 and 12 mo were significantly lower in the R1 group than in the other groups, and the serum triacylglycerol levels were significantly lower than those in the R16 group. However, no significant differences were noted in the serum cholesterol or triacylglycerol level after feeding for 18 mo among the 3 groups. A significantly higher serum glucose level was noted in the R1 group only at 18 mo of test diet ingestion, compared to that in the R4 group. The results suggest that the influence of the dietary n-6/n-3 ratio on the serum lipid and glucose levels varies, depending on the duration and life stage of feeding. Our findings further suggest that the life span of Wistar rats is not affected even if the ratio of dietary n-6/n-3 changes from 1 to 16.

Effect of dietary n-6/n-3 ratio on liver n-6/n-3 ratio and peroxisomal beta-oxidation activity in rats.

The optimal dietary n-6/n-3 ratio has not been fully elucidated. To investigate the influence of the dietary n-6/n-3 ratio on this ratio in the body and on liver beta-oxidation peroxisomal activity, rats were fed diets containing fat at an n-6/n-3 ratio of 1 to 16 for 4 weeks. To investigate whether elevation of the liver peroxisomal beta-oxidation activity increases the n-6/n-3 ratio in the body, rats were fed a diet containing a peroxisome-activating agent, bezafibrate, for 2 weeks, and its influence on the liver n-6/n-3 ratio was examined. The slope of the regression line between the dietary and liver total lipid n-6/n-3 ratios was significantly smaller when the dietary n-6/n-3 ratio was 4 or greater than when it was smaller than 4. Peroxisomal beta-oxidation and acyl CoA oxidase activities were significantly lower in rats fed a diet with an n-6/n-3 ratio of 16 than in those fed a diet with a ratio of 1. The peroxisomal beta-oxidation activity in the bezafibrate-supplemented group was significantly higher than that in the control group. The serum and liver total lipid n-6/n-3 ratios were significantly higher in the 0.015% bezafibrate-supplemented group than in the control group. These findings suggest that the liver n-6/n-3 ratio might be controlled via peroxisomal beta-oxidation in rats.

The application of medium-chain fatty acids: edible oil with a suppressing effect on body fat accumulation.

The bulk of fatty acids found in our diets consists of long-chain fatty acids (LCFA), which are molecules containing 12 or more carbon atoms. In contrast, medium-chain fatty acids (MCFA) are composed of 8-10 carbon atoms, and are found in palm kernel oil, among other types of foods. MCFA have attracted attention as being part of a healthy diet, because they are absorbed directly into the portal vein, transported rapidly to the liver for beta-oxidation, and thus increase diet-induced thermogenesis. In contrast, long-chain triacylglycerols are absorbed via the intestinal lymphatic ducts and transported by chylomicrons through the thoracic duct into the systemic circulation. Because medium-chain triacylglycerols (MCT) containing solely MCFA have a few disadvantages when used for deep frying, we have developed a new kind of triacylglycerol product: medium- and long-chain triacylglycerol (MLCT). MLCT is produced by lipase-catalyzed enzymatic transesterification. Long-term clinical trials have demonstrated that MLCT and MCT result in less body fat accumulation in humans. MLCT oil has been approved as FOSHU (Food for Specified Health Use) for use as cooking oil with a suppressing effect on body fat accumulation.

Alpha-linolenic acid-rich flaxseed oil ingestion increases plasma adiponectin level in rats.

Adiponectin, an adipose-specific secretory protein, exhibits antidiabetic and antiatherogenic properties. The effect of alpha-linolenic acid (ALA) on adiponectin has not been revealed. ALA is included abundantly in vegetable oils such as flaxseed oil. In this study, we attempted to clarify the effect of ALA-rich flaxseed oil (FSO) intake on the adiponectin level in rats. Seven-week-old male Sprague-Dawley rats were fed test diets containing high oleic safflower oil (HOSO) or FSO for 4 weeks. After the experimental period, the plasma adiponectin concentration in the FSO-fed group was higher than that in the HOSO-fed group. The adiponectin content of perirenal adipose tissue in the FSO-fed group was also significantly higher than that in the HOSO-fed group. However, the adiponectin mRNA level in the perirenal adipose tissue did not differ significantly between the HOSO-fed and FSO-fed groups. In this study, we clarified the effect of the ALA-rich FSO ingestion on the plasma adiponectin concentration in rats. It was suggested that the ALA-rich FSO intake might exhibit beneficial effects through an increase of the adiponectin level.

Lowering systolic blood pressure and increases in vasodilator levels in SHR with oral alpha-linolenic acid administration.

The present study attempted to clarify the antihypertensive effect and its mechanism when alpha-linolenic acid (ALA) is administered orally. For this purpose, 1 mL of flaxseed oil, which is rich in ALA, and high oleic safflower oil was administered orally to spontaneously hypertensive rats (SHR) of a control and an ALA group on days 1 and 5. Systolic blood pressure was measured on day 1, and blood and liver were collected on day 5. Four hours after the oral administration on day 1, systolic blood pressure of the ALA group was lower than that of the control group. Levels of plasma vasodilators, such as prostaglandin I(2) metabolite, nitric oxide metabolites, and bradykinin, in the ALA group were significantly higher than those in the control group, but levels of vasoconstrictors, such as angiotensin II and thromboxane A(2) metabolite, did not differ significantly. It is known that bradykinin induces prostaglandin I(2) and nitric oxide. The present study shows that ALA reduced the systolic blood pressure of SHR, and its mechanism may be related to increases of prostaglandin I(2) and nitric oxide through bradykinin stimulation.

Effect of dietary lard containing higher alpha-linolenic acid on plasma triacylglycerol in rats.

It is well known that the consumption of n-3 polyunsaturated fatty acid (PUFA) decreases the plasma triacylglycerol (TAG) level. The technology of elevating the content of n-3 PUFAs in pig meat has already reached a practical level. In this study, the effects of dietary lard containing higher alpha-linolenic acid (LNA) on plasma TAG were compared with those of normal lard in rats. The rats were fed a diet containing either 10% normal lard or a high linolenic lard for 4 weeks. The plasma and liver TAG levels in the high linolenic lard group were significantly lower than those in the normal lard group. The activity of the fatty acid synthase (FAS) of the liver in the high linolenic lard group was significantly lower than that in the normal lard group. The contents of n-3 PUFAs in hepatic total lipid, TAG fraction, and the phospholipids (PLs) fraction increased in the high linolenic lard group. The results indicate that the high linolenic lard suppressed hepatic FAS activity compared with the control lard, resulting in a lower concentration of plasma TAG. These results also suggest that pig meat containing high LNA may be more nourishing than normal pig meat.

Antihypertensive effect and safety of dietary alpha-linolenic acid in subjects with high-normal blood pressure and mild hypertension.

We investigated the antihypertensive effect and safety of alpha-linolenic acid (ALA) in human subjects. In Experiment 1, subjects with high-normal blood pressure and mild hypertension ingested bread containing 14 g of common blended oil (control oil) or ALA-enriched oil for 12 weeks. The test oil contained 2.6g/14 g of ALA. The subjects ingested strictly controlled meals during the study period. Systolic blood pressure was significantly lower in the ALA group than in the control group after ingestion of the test diet for 4, 8 and 12 weeks. Diastolic blood pressure was significantly lower in the ALA group than in the control group after ingestion of the test diet for 12 weeks. In Experiment 2, we evaluated the safety of high intake of ALA (7.8 g/d), particularly its effects on oxidation in the body and blood coagulation. Normotensive, high-normotensive and mildly hypertensive subjects ate bread that contained 42 g of the control oil or the test oil for 4 weeks. No significant difference was noted in the lipid peroxide level, high-sensitive C-reactive protein level, plasma prothrombin time or activated partial thromboplastin time between the two groups. No abnormal changes were noted after test diet ingestion on blood test or urinalysis, and no adverse event considered to have been induced by the test oil was observed in Experiment 1 and 2. These results suggest that ALA have an antihypertensive effect with no adverse effect in subjects with high-normal blood pressure and mild hypertension.

Effect of Docosahexaenoic Acid Ingestion on Temporal Change in Urinary Excretion of Mercapturic Acid in ODS Rats.

We hypothesized a suppressive mechanism for docosahexaenoic acid (22:6n-3; DHA)-induced tissue lipid peroxidation in which the degradation products, especially aldehydic compounds, are conjugated with glutathione through catalysis by glutathione S-transferases, and then excreted into urine as mercapturic acids. In the present study, ascorbic acid-requiring ODS rats were fed a diet containing DHA (3.6% of total energy) for 31 days. Lipid peroxides including degradation products and their scavengers in the liver and kidney were determined, and the temporal change in the urinary excretion of mercapturic acids was also measured. The activity of aldehyde dehydrogenase, which catalyzes the oxidation and detoxification of aldehydes, tended to be higher in the liver of DHA-fed rats. The levels of lipid peroxides as measured by thiobarbituric acid-reactive substances and aldehydic compounds were higher and that of alpha-tocopherol was lower in the liver, and the pattern of temporal changes in the urinary excretion of mercapturic acids was also different between the n-6 linoleic acid and DHA-fed rats. Accordingly, we presume from these results that after dietary DHA-induced lipid peroxidation, a proportion of the lipid peroxidation-derived aldehydic degradation products is excreted into urine as mercapturic acids.

Induction of multidrug resistance-associated protein MRP3 in the liver of rats fed with docosahexaenoic acid.

To clarify the alternative mechanisms to vitamin E (VE) regulating lipid peroxide accumulation in the liver after docosahexaenoic acid (DHA) ingestion, we examined the relationship between the DHA-induced lipid peroxide formation and induction of the xenobiotic transporters, Ral-binding GTPase-activating protein (RalBP1) and multidrug resistance-associated proteins 1, 2 and 3 (MRP1-3), in the liver of rats fed with DHA. The test diets contained DHA and linoleic acid (LA) (8.7% and 2.1% of total energy, respectively) with different levels of dietary VE (normal and low: 68 and 7.7 mg of alpha-tocopherol equivalent per kg diet, respectively), and the control diet contained LA alone (11.5% of total energy). The rats were fed with these experimental diets for 14 d. The proportions of DHA in the liver, kidney and heart were higher in the DHA-fed groups than in the LA-fed group. The tissue thiobarbituric acid values as an index of lipid peroxidation were also significantly higher in the DHA-fed groups, but the value did not differ between the DHA-fed groups with different VE levels. In the liver, there were no significant differences in the glutathione S-transferase (GST) and aldehyde dehydrogenase (ALDH) activities or in the expression of GST M2, RalBP1, MRP1 and MRP2 mRNA. However, the obvious induction of expression of liver MRP3 mRNA and tendency to produce the protein were recognized after DHA ingestion. This study is the first to report the gene expression of MRP3 by DHA ingestion. There might exist, therefore, some relationship between the DHA intake and MRP3 induction in regulating lipid peroxide accumulation in the liver.

Dietary docosahexaenoic acid-induced generation of liver lipid peroxides is not suppressed further by elevated levels of glutathione in ODS rats.

OBJECTIVES: We examined the effects of ascorbic acid (AsA) and glutathione (GSH; experiment 1) and of GSH in acetaminophen-fed rats (experiment 2) on dietary docosahexaenoic acid (DHA)-induced tissue lipid peroxidation. METHODS: In experiment 1, AsA-requiring Osteogenic Disorder Shionogi/Shi-od/od (ODS) rats were fed soybean protein diets containing DHA (10.0% total energy) and AsA at 50 (low) or 300 (normal) mg/kg without (low) or with (normal) methionine at 2 g/kg for 32 d. In experiment 2, ODS rats were fed diets containing DHA (7.8% total energy) and acetaminophen (4 g/kg) with different levels of dietary methionine (low, moderate, high, and excessive at 0, 3, 6, and 9 g/kg, respectively) for 30 d. Tissue lipid peroxides and antioxidant levels were determined. RESULTS: In experiment 1, liver lipid peroxide levels in the low-AsA group were lower than those in the normal-AsA group, but kidney and testis lipid peroxide levels in the low-AsA group were higher than those in the normal-AsA group. Dietary methionine tended to decrease tissue lipid peroxide levels but did not decrease vitamin E (VE) consumption. In experiment 2, a high level of methionine (6 g/kg) decreased liver lipid peroxide levels and VE consumption. However, generation of tissue lipid peroxides and VE consumption were not decreased further by a higher dose of methionine (9 g/kg). CONCLUSIONS: Higher than normal levels of dietary methionine are not necessarily associated with decreased dietary DHA-induced generation of tissue lipid peroxides and VE consumption except that the GSH requirement is increased in a condition such as acetaminophen feeding.

Lower weight gain and higher expression and blood levels of adiponectin in rats fed medium-chain TAG compared with long-chain TAG.

Previous studies demonstrated that, compared with long-chain TAG (LCT), dietary medium-chain TAG (MCT) could improve glucose tolerance in rats and humans. It has been well established that adiponectin acts to increase insulin sensitivity. The effects of dietary MCT on adiponectin serum concentration and mRNA levels in adipose tissue were studied in rats. Male Sprague-Dawley rats were fed a diet containing 20% MCT or LCT for 8 wk. After 6 wk of dietary treatment, an oral glucose tolerance test was performed. Rats fed the MCT diet had less body fat accumulation than those fed the LCT diet (P < 0.01). The cell diameter of the perirenal adipose tissue, one of the abdominal adipose tissues, was smaller (P < 0.01) in the MCT diet group. The serum adiponectin concentration was higher (P < 0.01) in the MCT diet group than in the LCT diet group. The adiponectin content in the perirenal adipose tissue was higher (P < 0.01) in the MCT diet group. The MCT-fed group had a higher adiponectin mRNA level in their perirenal adipose tissue (P < 0.05). The increase of the plasma glucose concentration after glucose administration (area under the curve) was smaller (P < 0.01) in the MCT diet group than in the LCT diet group. These findings suggest that dietary MCT, compared with LCT, results in a higher serum adiponectin level with transcriptional activation of the adiponectin gene in rats. We speculate that improved glucose tolerance in rats fed an MCT diet may be, at least in part, ascribed to this higher serum adiponectin level.

Primary aminophospholipids in the external layer of liposomes protect their component polyunsaturated fatty acids from 2,2'-azobis(2-amidinopropane)- dihydrochloride-mediated lipid peroxidation.

We showed in our previous study that docosahexaenoic acid-rich phosphatidylethanolamine in the external layer of small-size liposomes, as a model for biomembranes, protected its docosahexaenoic acid from 2,2'-azobis(2-amidinopropane)dihydrochloride- (AAPH-) mediated lipid peroxidation in vitro. Besides phosphatidylethanolamine, both phosphatidylserine and an alkenyl-acyl analogue of phosphatidylethanolamine, phosphatidylethanolamine plasmalogen, are reported to possess characteristic antioxidant activities. However, there are few reports about the relationship between the protective activity of phosphatidylethanolamine plasmalogen and/or phosphatidylserine against lipid peroxidation and their distribution in a phospholipid bilayer. Furthermore, it is unclear whether phosphatidylethanolamine plasmalogen and/or phosphatidylserine protect their component polyunsaturated fatty acids (PUFAs) from lipid peroxidation. In the present study, we examined the relationship between the transbilayer distribution of aminophospholipids, such as phosphatidylethanolamine rich in arachidonic acid, phosphatidylethanolamine plasmalogen, and phosphatidylserine, and the oxidative stability of their component PUFAs. The transbilayer distribution of these aminophospholipids in liposomes was modulated by coexisting phosphatidylcholine bearing two types of acyl chain: dipalmitoyl or dioleoyl. The amounts of these primary aminophospholipids in the external layer became significantly higher in liposomes containing dioleoylphosphatidylcholine than in those containing dipalmitoylphosphatidylcholine. Phosphatidylethanolamine rich in arachidonic acid, phosphatidylethanolamine plasmalogen or phosphatidylserine in the external layer of liposomes, as well as external docosahexaenoic acid-rich phosphatidylethanolamine, were able to protect their component PUFAs from AAPH-mediated lipid peroxidation.

Transbilayer distribution of aminophospholipids and the oxidative stability of their component polyunsaturated fatty acids.

We examined the relationship between the transbilayer distribution of aminophospholipids, such as phosphatidylethanolamine (PE), PE plasmalogen and phosphatidylserine, and the oxidative stability of polyunsaturated fatty acids (PUFAs) in the aminophospholipids. To modulate the transbilayer distribution of aminophospholipid in liposomes, we used phosphatidylcholine (PC) with two types of acyl chain region: dipalmitoyl (PC16:0) or dioleoyl (PC18:1). In the smaller-sized liposomes, the proportions of aminophospholipid in the liposomal external layer were significantly higher in liposomes containing PC18:1 than in those containing PC16:0. Additionally, aminophospholipids in the external layer of smaller-sized liposomes were able to protect their component PUFAs from 2,2'-azobis(2-amidinopropane)dihydrochloride-mediated lipid peroxidation.

Effect of dietary DHA on DHA levels in retinal rod outer segments in young versus mature rats.

We compared the effect of direct supplementation with docosahexaenoic acid (DHA) on the fatty acid composition of the liver and the rod outer segment (ROS) membranes of the retina in young (five-week-old) and mature (one-year-old) rats. In young rats, a high content of DHA in the diet (9.7% of total energy) effectively increased the proportion of DHA in ROS membranes (41.8%), compared with the proportion observed in a linoleic acid (LA) diet group (control, 31.6%). The proportion of DHA was also significantly higher in the livers of young DHA-fed rats. These results show that direct supplementation with DHA is very effective in increasing DHA levels in the ROS membranes and livers of developing animals. In contrast, in mature rats there was no significant increase in the proportion of DHA in the ROS membranes, even after the highest dose (8.4% of total energy) of DHA, although the proportion of DHA was significantly higher in the livers of DHA-fed rats. The changes in fatty acid composition in the ROS membranes were different in young and mature rats fed high-DHA diets. Our findings indicate that mature rats maintain a constant level of DHA in the ROS membranes even after being directly supplemented with high doses of DHA.

Dietary docosahexaenoic acid-induced production of tissue lipid peroxides is not suppressed by higher intake of ascorbic acid in genetically scorbutic Osteogenic Disorder Shionogi/Shi-od/od rats.

In previous studies, we showed that docosahexaenoic acid (DHA) ingestion enhanced the susceptibility of rat liver and kidney to lipid peroxidation, but did not increase lipid peroxide formation to the level expected from the relative peroxidizability index (P-index) of the total tissue lipids. The results suggested the existence of some suppressive mechanisms against DHA-induced tissue lipid peroxide formation, as increased tissue ascorbic acid (AsA) and glutathione levels were observed. Therefore, we focused initially on the role of AsA for the suppressive mechanisms. For this purpose, we examined the influence of different levels of dietary AsA (low, moderate, high and excessive levels were 100, 300 (control), 600 and 3000 mg/kg diet respectively) on the tissue lipid peroxide and antioxidant levels in AsA-requiring Osteogenic Disorder Shionogi/Shi-od/od (ODS) rats fed DHA (6.4 % total energy) for 32 or 33 d. Diets were pair-fed to the DHA- and 100 mg AsA/kg diet-fed group. We found that the lipid peroxide concentrations of liver and kidney in the DHA-fed group receiving 100 mg AsA/kg diet were significantly higher or tended to be higher than those of the DHA-fed groups with AsA at more than the usual control level of 300 mg/kg diet. Contrary to this, the liver alpha-tocopherol concentration was significantly lower or tended to be lower in the DHA and 100 mg AsA/kg diet-fed group than those of the other DHA-fed groups. However, tissue lipid peroxide formation and alpha-tocopherol consumption were not suppressed further, even after animals received higher doses of AsA. The present results suggest that higher than normal concentrations of tissue AsA are not necessarily associated with the suppressive mechanisms against dietary DHA-induced tissue lipid peroxide formation.

Effect of docosahexaenoic acid and ascorbate on peroxidation of retinal membranes of ODS rats.

Mutant male osteogenic disorder Shionogi (ODS) rats, unable to synthesize ascorbic acid, were fed diets containing a high content of docosahexaenoic acid (DHA) and different amounts of ascorbic acid, to study the effect of DHA on peroxidative susceptibility of the retina and possible antioxidant action of ascorbic acid. ODS rats were fed from 7 weeks of age with diets containing high DHA (6.4% of total energy). A control group received a diet high in linoleic acid. The diets also contained varying amounts of ascorbic acid. Fatty acid compositions and phospholipid hydroperoxides in rod outer segment (ROS) membranes, and retinal ascorbic acid were analyzed. DHA in ROS membranes was significantly increased in rats fed high DHA, compared with the linoleic acid diet. Levels of phospholipid hydroperoxides in the DHA-fed rats were significantly higher than the linoleic acid-fed rats. Ascorbic acid supplementation did not suppress the phospholipid hydroperoxide levels after a high DHA diet, even when the supplement increased the content of retinal ascorbic acid. In conclusion, high DHA feeding induced a marked increase of phospholipid hydroperoxides in ROS membranes of ODS rats. Supplementation of ascorbic acid did not reverse this increase.