A method for simultaneous determination of plasma and erythrocyte antioxidant status. Evaluation of the antioxidant activity of vitamin E in healthy volunteers

1. Since oxygen free radicals are directly involved in a variety of pathologies such as atherosclerosis, diabetes mellitus, inflammation and/or when a deficit of defences of the organism against radicals occurs, we developed a suitable and simple method to determine both the erythrocyte sensitivity to an oxidative stress and plasma antioxidant protective capacity. 2. This test is based on the introduction at 37 degrees C of a radical initiator, 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH), within an erythrocyte suspension leading to a membrane alteration and ultimately to haemolysis. The latter can be quantified by determining the lacticodeshydrogenase activity released in the medium. The erythrocyte sensitivity to haemolysis and the volume of plasma inhibiting 50% of the haemolysis were determined. 3. Intra-assay CVs were 1.9% for erythrocyte sensitivity to oxidative stress and 3.4% for inhibitory 50% plasma volume. Inter-assay CVs for both erythrocyte sensitivity and inhibitory 50% plasma volume were 4%. 4. The reliability of this method was assessed and applied to test the protective effect of vitamin E, a well known antioxidant agent, in six healthy volunteers. Two weeks after daily administration of 500 mg of vitamin E, the mean plasma vitamin E concentration increased by 41% from 10.7 +/- 2.0 mg l-1 before treatment (P < 0.05). As the vitamin E concentration increased, the mean inhibitory 50% plasma volume and the percentage of haemolysed erythrocytes decreased respectively by 29% from 3.35 +/- 0.5 microliter (P < 0.05) and 18% from 71.5 +/- 3.8% (P < 0.05). No significative variation of these parameters was observed in six adult men without vitamin E supplementation. 5. Thus, this global and simple test permits an antioxidant status evaluation of a patient. It can be applied to various pathologies and allows the potency of new antioxidant molecules to be evaluated.     

Effects of folic acid and combinations of folic acid and vitamin B-12 on plasma homocysteine concentrations in healthy, young women

BACKGROUND: Elevated plasma homocysteine concentrations are considered to be a risk factor for vascular disease and fetal malformations such as neural tube defects. Recent studies have shown that plasma homocysteine can be lowered by folic acid in amounts corresponding to 1-2 times the recommended dietary allowance. Preliminary evidence indicates that vitamin B-12 may be beneficial when included in supplements or in a food-fortification regimen together with folic acid. OBJECTIVE: We aimed to compare the homocysteine-lowering potential of a folic acid supplement with that of 2 supplements containing different doses of vitamin B-12 in addition to folic acid. DESIGN: Female volunteers of childbearing age (n = 150) received a placebo for 4 wk followed by a 4-wk treatment with either 400 microg folic acid, 400 microg folic acid + 6 microg vitamin B-12, or 400 microg folic acid + 400 microg vitamin B-12. RESULTS: Significant reductions (P < 0.001) in plasma homocysteine were observed in all groups receiving vitamin treatment. The effect observed with the combination of folic acid + 400 microg vitamin B-12 (total homocysteine, -18%) was significantly larger than that with a supplement containing folic acid alone (total homocysteine, -11%) (P < 0.05). Folic acid in combination with a low vitamin B-12 dose (6 microg) affected homocysteine as well (-15%). CONCLUSIONS: These results suggest that the addition of vitamin B-12 to folic acid supplements or enriched foods maximizes the reduction of homocysteine and may thus increase the benefits of the proposed measures in the prevention of vascular disease and neural tube defects.     

Fish oil supplementation prevents diabetes-induced nerve conduction velocity and neuroanatomical changes in rats

Diabetic neuropathy has been associated with a decrease in nerve conduction velocity, Na,K-ATPase activity and characteristic histological damage of the sciatic nerve. The aim of this study was to evaluate the potential effect of a dietary supplementation with fish oil [(n-3) fatty acids] on the sciatic nerve of diabetic rats. Diabetes was induced by intravenous streptozotocin injection. Diabetic animals (n = 20) were fed a nonpurified diet supplemented with either olive oil (DO) or fish oil (DM), and control animals (n = 10) were fed a nonpurified diet supplemented with olive oil at a daily dose of 0.5 g/kg by gavage for 8 wk. Nerves were characterized by their conduction velocity, morphometric analysis and membrane Na, K-ATPase activity. Nerve conduction velocity, as well as Na,K-ATPase activity, was improved by fish oil treatment. A correlation was found between these two variables (R = 0.999, P < 0.05). Moreover, a preventive effect of fish oil was observed on nerve histological damage [endoneurial edema, axonal degeneration (by 10-15%) with demyelination]. Moreover, the normal bimodal distribution of the internal diameter of myelinated fibers was absent in the DO group and was restored in the DM group. These data suggest that fish oil therapy may be effective in the prevention of diabetic neuropathy.     

alpha-Tocopherol concentrations in plasma but not in lipoproteins fluctuate during the menstrual cycle in healthy premenopausal women

Because premenopausal women experience cyclic fluctuations of plasma carotenoids and their lipoprotein carriers, it was hypothesized that plasma alpha-tocopherol (A-T) fluctuates by phase of the menstrual cycle. Twelve free-living women, with a confirmed ovulatory cycle, were given a controlled diet for two consecutive menstrual cycles. Blood was drawn during the menses, early follicular, late follicular and luteal phases to simultaneously measure serum hormones, plasma lipoproteins and A-T concentrations, and A-T distribution in the lipoprotein fractions. Plasma A-T concentrations were significantly lower during menses than during the luteal phase by approximately 12% in each controlled diet cycle (P < 0.001). Adjustment for serum cholesterol and triglyceride concentrations did not alter these findings. The distributions of A-T in lipoprotein cholesterol fractions were not significantly different by menstrual phase. From 61 to 62% of A-T was concentrated in the LDL fraction, with another 9-14% in HDL2, 17-22% in HDL3 and the remaining 6-8% in VLDL+ IDL. There were no significant differences in lipoprotein cholesterol fractions by menstrual phase, except for a significant increase (P = 0.03) in HDL2 cholesterol from the early follicular to the late follicular phase. Spearman rank correlations from data during the second controlled diet month showed A-T in HDL2 in the late follicular phase was positively correlated with HDL cholesterol in the early follicular (r = 0.88), late follicular (r = 0.86) and luteal phases (r = 0.86) and with luteal apolipoprotein (ApoA-1) level (r = 0.90), and luteal HDL2 cholesterol (r = 0.83). A-T in HDL3 in the early follicular phase was negatively correlated with HDL2 cholesterol (r = -0.96) and ApoA-1 (r = -0.85), whereas luteal A-T in HDL3 was correlated with luteal HDL3 cholesterol (r = -0.79). Late follicular A-T in VLDL was positively correlated with early follicular HDL3 cholesterol and late follicular HDL3 cholesterol (r = 0.83). Fluctuations of A-T concentrations by phase of the menstrual cycle should be taken into consideration in future research concerning premenopausal women and the risk of chronic disease.     

Change in tissue concentrations of lipid hydroperoxides, vitamin C and vitamin E in rats with streptozotocin-induced diabetes

The purpose of these studies was to determine the distribution of a lipophilic antimalarial agent, halofantrine hydrochloride (Hf), in fasted plasma from hypo-, normo-, and hyperlipidemic patients that displayed differences in lipoprotein concentration and lipid transfer protein I (LTP I) activity. To assess the influence of modified lipoprotein concentrations and LTP I activity on the plasma distribution of Hf, Hf at a concentration of 1000 ng/mL was incubated in either hypo-, normo-, or hyperlipidemic human plasma for 1 h at 37 degreesC. Following incubation, the plasma samples were separated into their lipoprotein and lipoprotein-deficient plasma (LPDP) fractions by density gradient ultracentrifugation and assayed for Hf by high-pressure liquid chromatography. The activity of LTP I in the dyslipidemic plasma samples was determined in terms of its ability to transfer cholesteryl ester from low-density lipoproteins (LDL) to high-density lipoproteins (HDL). Total plasma and lipoprotein cholesterol (esterified and unesterified), triglyceride, and protein levels in the dyslipidemic plasma samples were determined by enzymatic assays. When Hf was incubated in normolipidemic plasma for 1 h at 37 degreesC, the majority of drug was found in the LPDP fraction. When Hf was incubated in human plasma of varying total lipid, lipoprotein lipid, and protein concentrations and LTP I activity, the following relationships were observed. As the triglyceride-rich lipoprotein (TRL) lipid and protein concentration increased from hypolipidemia through to hyperlipidemia, the proportion of Hf associated with TRL increased (r > 0.90). As the HDL lipid and protein concentration increased, the proportion of Hf associated with HDL decreased (r > 0.70). As the total and lipoprotein lipid levels increased, the LTP I activity of the plasma also proportionally increased (r > 0.85). Furthermore, with the increase in LTP I activity, the proportion of Hf associated with the TRL fraction increased (r > 0.70) and the proportion of Hf associated with the HDL fraction decreased (r > 0.80). In addition, a positive correlation between the proportion of apolar lipid and Hf recovered within each lipoprotein fraction was observed within hypo- (r > 0.80), normo- (r = 0.70), and hyperlipidemic (r > 0.90) plasmas. These findings suggest that changes in the HDL and TRL lipid and protein concentrations, LTP I activity, and the proportion of apolar lipid within each lipoprotein fraction may influence the plasma lipoprotein distribution of Hf in dyslipidemia.     

Fatty acid composition in maternal milk and plasma during supplementation with cod liver oil

OBJECTIVE: We investigated how cod liver oil influences the amount of essential fatty acids in mothers' breast milk. DESIGN AND INTERVENTION: Lactating mothers (n =22) were randomized into four groups 3-8 weeks after parturition. They were supplemented for 14 days with 0, 2.5, 5 and 10 ml cod liver oil (7.7 g eicosapentaenoic acid (EPA, 20:5n-3), 10.2 g docosahexaenoic acid (DHA, 22:6n-3) and 22.9 g n-3 fatty acids in total per 100 ml). RESULTS: In maternal plasma phospholipids there was an increase in the content of EPA and DHA in the group supplemented with 10 ml cod liver oil daily (P < or = 0.05). DHA concentrations in breast milk pre-supplementation ranged from 0.15 to 1.56 wt% and increased in all supplemented groups (P< or =0.05). The concentration of EPA in breast milk increased in the groups supplemented with 5 or 10 ml cod liver oil (P< or =0.05), whereas the concentration of arachidonic acid (AA, 20:4n-6) did not change in any of the supplemented groups. Total intake of DHA adjusted to body mass index (BMI), correlated to DHA concentrations in plasma (r = 0.49, P = 0.02) and breast milk (r = 0.45, P = 0.04). The concentration of tocopherol did not change during the supplementation period, neither in plasma nor in breast milk. CONCLUSION: Dietary intake of DHA is reflected in the concentration of DHA in breast milk, without affecting the concentration of AA or tocopherol.     

Factors affecting serum selenium and vitamin E concentrations in dairy cows

Supplementation of selenium and vitamin E to enhance disease resistance in dairy cattle has become common, particularly to prevent periparturient reproductive disorders and mastitis. To establish reference values for serum vitamin E and selenium concentrations in postparturient dairy cattle and to determine whether serum concentrations of these micronutrients varied with season and stage of lactation, cows from a stratified random sample of 50 herds were studied for 1 year. Blood samples were collected from each of the 50 study herds twice, from the 10 most recently parturient cows or from 10% of the herd, whichever was greatest. Mean concentration of vitamin E and selenium was 2.55 micrograms/ml and 78.12 ng/ml, respectively. Vitamin E concentrations were significantly (P < 0.05) higher during the summer and fall than during the winter and spring. Selenium concentrations were significantly (P < 0.05) lower during the summer and fall than during the winter and spring. Herd, season of blood sample collection, and time since parturition were significant (P < 0.02) in explaining variation in vitamin E and selenium concentrations.     

Influence of beta carotene, vitamin E, and vitamin C on endogenous antioxidant defenses in erythrocytes

OBJECTIVE: To evaluate the in vivo radical scavenger activity of vitamin E, vitamin C, and beta carotene on erythrocyte membranes. DESIGN: A prospective, open trial without placebo. SETTING: Department of Clinical Pharmacy. PATIENTS: Ten healthy volunteers being supplemented with beta carotene, vitamin E, and vitamin C. MEASUREMENTS: Erythrocytes were incubated in water bath with 2,2' azobis (2 amidinopropane) hydrochloride (AAPH). AAPH decomposes spontaneously at 37 degrees C to generate free radicals inducing membrane cellular damage and hemolysis. The absorbance was measured at 405 nm at 0, 30, and 60 min, and then every 20 minutes for four hours. The time for 50 percent of maximal hemolysis (T50%), which expresses the radical scavenger activity of erythrocytes, was determined. RESULTS: The physiologic T50% value determined in 52 healthy volunteers is 117 +/- 12 min. Patients receiving these supplements have a higher value of T50% (143.2 +/- 11.6 min at 30 d and 145.7 +/- 10.5 min at 60 d) than the physiologic value (p < 0.001). CONCLUSIONS: These data suggest that vitamin C, vitamin E, and beta carotene stimulate the radical scavenger activity of erythrocyte membranes after 30 days.     

Whole blood production of thromboxane, prostacyclin and leukotriene B4 after dietary fish oil supplementation in man: effect of vitamin E

Antigenic peptides derived from several differentiation antigens of the melanocyte lineage were recently identified in human melanomas as targets for HLA-A2.1-restricted cytotoxic T lymphocytes (CTLs). To examine their potential role in tumour-directed immune responses in vivo, we determined CTL reactivity against seven antigenic peptides derived from the Melan A/MART-1, tyrosinase and gp100/Pmel17 antigens in the peripheral blood of 10 HLA-A2+ healthy controls and 26 HLA-A2+ melanoma patients. The influenza matrix peptide (GILGFVFTL) presented by HLA-A2.1 was used as a control peptide. CTL reactivity was assessed in a mixed lymphocyte 'peptide' culture assay. Reactivity against Melan A/MART-1-derived peptide antigens was readily detectable in both melanoma patients and controls. Reactivity directed against tyrosinase-derived peptide antigens was also detected in both melanoma patients and healthy individuals, but less frequently. A measurable response against gp100/Pmel17-derived antigens was found in 1/10 controls and in 1/26 of the melanoma patients. Reactivity against the influenza matrix peptide was common in both melanoma patients and controls. Our findings show that precursor CTLs against melanocyte differentiation antigens can be detected in peripheral blood of melanoma patients and healthy individuals. The pattern of CTL reactivity directed against melanoma-associated antigens does not seem to be altered in melanoma patients. Despite antigen-specific CTL reactivity, tumour growth was not prevented in melanoma patients and autoimmune phenomena were not detected in healthy individuals. It remains to be determined whether precursor CTLs recognizing melanocyte differentiation antigens can be activated by immunization and lead to effective tumour rejection in vivo.     

Tissue manganese concentrations and antioxidant enzyme activities in rats given total parenteral nutrition with and without supplemental manganese

BACKGROUND: Manganese is an essential but potentially toxic mineral. Parenteral administration of manganese via total parenteral nutrition (TPN) bypasses homeostatic mechanisms (intestinal absorption and presystemic hepatic elimination). Our objective in this study was to determine the effect of supplemental manganese in TPN solutions on manganese status in a rat model. METHODS: Male Sprague-Dawley rats underwent jugular catheterization and were given 61.0 +/- 0.4 g/d TPN solution providing 0.5 +/- 0.2 nmol manganese/g (Mn-; n = 6) or 16 +/- 3 nmol manganese/g (Mn+; n = 7) for 7 days. Reference rats (RF; n = 8) were fed a purified diet containing 1.3 mmol manganese/g. RESULTS: Liver manganese decreased in both TPN groups, but tibia, spleen, and pancreas manganese concentrations were greater in Mn+ rats than in Mn- or RF rats. Although no treatment differences were seen in heart or liver manganese superoxide dismutase activity, heart copper-zinc superoxide dismutase activity was lower in the Mn+ rats than in Mn- or RF rats (p < .05). Glutathione peroxidase activity was depressed in livers of both Mn- and Mn+ rats relative to RF rats (p < .0001), which was not due to selenium deficiency. CONCLUSIONS: Supplemental parenteral manganese is taken up to a greater extent by peripheral tissues than the liver. In this first report of antioxidant enzyme activities in animals maintained with TPN, we found that TPN as well as supplemental manganese can influence antioxidant enzyme activities. We conclude that it is generally unnecessary and potentially toxic to supplement TPN solutions with manganese during short-term usage.     

Acute inflammation induces hyporetinemia and modifies the plasma and tissue response to vitamin A supplementation in marginally vitamin A-deficient rats

Plasma retinol is reduced during numerous infections, and inflammation alters the hepatic synthesis of retinol-binding protein (RBP). In this study, we have investigated the effects of endotoxin-induced inflammation on vitamin A (VA) supplementation in a rat model of marginal VA deficiency. Marginally VA-deficient rats received an intraperitoneal dose of lipopolysaccharide (LPS, n = 14) or saline (n = 10); 6 h later, six LPS + VA and six saline + VA rats received 7.1 micromol VA orally. Twenty-four hours after endotoxin administration, rats with inflammation (LPS) had lower plasma retinol, RBP, and hepatic RBP than saline rats (37, 31 and 44%, respectively, P < 0.05). Inflammation did not affect VA concentrations in liver and perirenal adipose tissue, although kidney VA was reduced relative to saline rats. However, urinary VA was not detected. Eighteen hours after VA supplementation, inflammation reduced the plasma unesterified retinol response (P < 0. 05) in LPS + VA relative to saline + VA rats, although total VA increased as a result of the presence of retinyl esters in LPS + VA rats. Hepatic esterified retinol concentration was reduced (P < 0. 01) in LPS + VA compared with saline + VA rats; however, hepatic unesterified retinol did not differ. Renal total retinol increased in VA-supplemented rats, but urinary retinol excretion, when observed, was low, independently of inflammation. These findings indicate that inflammation-induced hyporetinemia does not necessarily imply a loss of VA, but rather represents a redistribution of tissue VA brought about by a reduced hepatic synthesis of RBP. Practical implications from these collective results are to recommend the determination of both unesterified and esterified retinol to fully assess the plasma response to VA supplementation and to caution the use of VA assessment methodologies that depend on the hepatic synthesis of RBP during acute inflammation.     

Raised plasma endothelin-1 concentrations in patients with primary hypercholesterolemia without evidence of atherosclerosis

This study investigated the Psychosocial adjustment in 40 patients who received orthotopic liver transplantation (OLT) for several endstage liver diseases. Twenty patients were grafted because they suffered from liver Cancer as well as cirrhosis. Particular attention was paid to evaluating whether cancer could affect recipients' coping with transplant. Each patient underwent a semi-structured interview to obtain information on their psychosocial life, relationship with the donor, organ acceptance and life expectancy. Interview was performed I year after transplantation. A psychodiagnostic evaluation was also performed using a Minnesota Multiphasic Personality Inventory (MMPI) and a Human Figure Test. Psychosocial adaptation in everyday life following liver transplantation seemed good in most of the patients, whatever the indication for transplantation might be. It can he seen that by replacing the diseased organ a high percentage of oncological patients overcame their fear of cancer.     

Reversal of defective nerve conduction with vitamin E supplementation in type 2 diabetes: a preliminary study

OBJECTIVE: The present study has examined the effect of vitamin E, the principal modulator of free radical activity, on electrophysiological parameters in patients with diabetic peripheral sensorimotor polyneuropathy, matched for duration of disease and metabolic control. RESEARCH DESIGN AND METHODS: A total of 21 subjects with type 2 diabetes were enrolled in this double-blind randomized placebo-controlled study (vitamin E, 11 patients; placebo, 10 patients). Patients were randomly assigned to receive either 900 mg vitamin E or placebo for 6 months. The average dietary vitamin E consumption of the subjects was similar during the study. The main outcome measure was the electrophysiological tests assessing nerve conduction. Fasting plasma glucose, HbA1, postprandial plasma glucose, and electrophysiological parameters in the basal state and after 6 months of treatment were studied. RESULTS: Glycemic indexes did not show any significant changes during the study, whereas nerve conduction improved significantly in 2 of the 12 studied electrophysiological parameters after 6 months in patients on vitamin E supplementation. The changes in the electrophysiological parameters were obvious in the median motor nerve fibers and tibial motor nerve fibers. Nerve conduction velocity in the median motor nerve fibers (P = 0.0019) and tibial motor nerve distal latency (P = 0.0284) improved significantly after 6 months of vitamin E supplementation. CONCLUSIONS: This study shows that defective nerve conduction in diabetic subjects with mild-to-moderate peripheral neuropathy may be improved by pharmacological doses of vitamin E supplementation. Further studies with a larger number of patients for longer periods of time are needed.     

The effect of vitamin E on antioxidant tissue activity in pregnant rats with streptozocin-induced diabetes]

When home enteral nutrition is needed, there is still, despite the undisputed increase in the quality of life that can be achieved with a Percutaneous Endoscopic Gastrostomy (PEG), a reservation in its use because this technique and its maintenance is considered to be very costly. We aim to assess the true cost of home enteral nutrition using the oral route, a nasogastric tube, and PEG. PATIENTS AND METHODS: The data of 65 patients who required home enteral nutrition during 1996, were analyzed retrospectively. The access route was a nasogastric tube in 20 cases, 18 patients had PEG, and 27 candidates used an oral route. The average age was 56 years. 50% were men and 50% were women. The most common diagnoses that led to the indication were oropharyngeal-maxillofacial neoplasms and neurological disorders. In all cases the material and formula used was assessed, as were the associated complications and the cost of the at home enteral nutrition. RESULTS: The average duration of the treatment was 175 +/- 128 days, and this was similar in all three groups. The average formula/day cost was slightly higher in the patients using the oral access route. The average total day cost and the average material/day cost was slightly higher in patients with a PEG. Patients with a PEG presented fewer complications than those with a nasogastric tube. The cost derived from possible complications must be higher in the nasogastric tube group, especially considering the repeated tube changes due to obstruction or loss. CONCLUSIONS: The cost of home enteral nutrition is slightly lower if one uses a nasogastric tube. The greater incidence of complications that were mild but required a tube change, in this case a nasogastric tube, suggests higher indirect costs. The oral route is associated with the need for special formulae that are more expensive.     

Effects of vitamin E and C supplementation on oxidative stress and viral load in HIV-infected subjects

OBJECTIVES: The HIV-infected population is known to be oxidatively stressed and deficient in antioxidant micronutrients. Since in vitro replication of HIV is increased with oxidative stress, this study assessed the effect of antioxidant vitamin supplementation on lipid peroxidation, a measure of oxidative stress, and viral load in humans. DESIGN: A randomized placebo-controlled, double-blind study. METHODS: Forty-nine HIV-positive patients were randomized to receive supplements of both DL-alpha-tocopherol acetate (800 IU daily) and vitamin C (1000 mg daily), or matched placebo, for 3 months. Plasma antioxidant micronutrient status, breath pentane output, plasma lipid peroxides, malondialdehyde and viral load were measured at baseline and at 3 months. New or recurrent infections for the 6-month period after study entry were also recorded. RESULTS: The vitamin group (n = 26) had an increase in plasma concentrations of alpha-tocopherol (P < 0.0005) and vitamin C (P < 0.005) and a reduction in lipid peroxidation measured by breath pentane (P < 0.025), plasma lipid peroxides (P < 0.01) and malondialdehyde (P < 0.0005) when compared with controls (n = 23). There was also a trend towards a reduction in viral load (mean +/- SD changes over 3 months, -0.45 +/- 0.39 versus +0.50 +/- 0.40 log10 copies/ml; P = 0.1; 95% confidence interval, -0.21 to -2.14). The number of infections reported was nine in the vitamin group and seven in the placebo group. CONCLUSION: Supplements of vitamin E and C reduce oxidative stress in HIV and produce a trend towards a reduction in viral load. This is worthy of larger clinical trials, especially in HIV-infected persons who cannot afford new combination therapies.     

Variability of plasma HDL subclass concentrations in men and women over time

1. Nitric oxide (NO) has been suggested as a gastrointestinal neurotransmitter, mediating the gastric receptive relaxation and the relaxation in the peristaltic reflex. The aim of the present study was to measure nerve-induced NO formation in vivo in the gastrointestinal tract. 2. Formation of the nitric oxide oxidation products nitrite and nitrate during vagal nerve stimulation were measured in the anaesthetized rabbit. Microdialysis probes were inserted into the wall of the stomach and proximal colon, and nitrite and nitrate in dialysate measured by capillary electrophoresis. 3. During bilateral vagal nerve stimulation there was an increase in nitrite and nitrate formation at the level of the stomach and in nitrite formation at the level of the colon. This increase was inhibited by intravenous administration of the NO synthase inhibitor N omega-nitro-L-arginine methyl ester (L-NAME 30 mg kg-1). Furthermore, L-NAME significantly increased nerve-induced gastric and colonic contractions, as well as spontaneous colonic contractions. 4. In summary, we present a new methodological procedure for quantification of small changes in nitric oxide formation in vivo. This study provides evidence that nitric oxide is released in the stomach and colonic wall during vagal nerve activity, at concentrations able to cause inhibition of smooth muscle contractions in vivo.     

Incorporation of n-3 fatty acids into plasma lipid fractions, and erythrocyte membranes and platelets during dietary supplementation with fish, fish oil, and docosahexaenoic acid-rich oil among healthy young men

The effects of n-3 fatty acid supplementation in the form of fresh fish, fish oil, and docosahexaenoic acid (DHA) oil on the fatty acid composition of plasma lipid fractions, and platelets and erythrocyte membranes of young healthy male students were examined. Altogether 59 subjects (aged 19-32 yr, body mass index 16.8-31.3 kg/m2) were randomized into the following diet groups: (i) control group; (ii) fish diet group eating fish meals five times per week [0.38 +/- 0.04 g elcosapentaenoic acid (EPA) and 0.67 +/- 0.09 g DHA per day]; (iii) DHA oil group taking algae-derived DHA oil capsules (1.68 g/d DHA in triglyceride form); and (iv) fish oil group (1.33 g EPA and 0.95 g DHA/d as free fatty acids) for 14 wk. The fatty acid composition of plasma lipids, platelets, and erythrocyte membranes was analyzed by gas chromatography. The subjects kept 4-d food records four times during the study to estimate the intake of nutrients. In the fish diet, in DHA oil, and in fish oil groups, the amounts of n-3 fatty acids increased and those of n-6 fatty acids decreased significantly in plasma lipid fractions and in platelets and erythrocyte membranes. A positive relationship was shown between the total n-3 polyunsaturated fatty acids (PUFA) and EPA and DHA intake and the increase in total n-3 PUFA and EPA and DHA in all lipid fractions analyzed. DHA was preferentially incorporated into phospholipid (PL) and triglyceride (TG) and there was very little uptake in cholesterol ester (CE), while EPA was preferentially incorporated into PL. and CE. The proportion of EPA in plasma lipids and platelets and erythrocyte membranes increased also by DHA supplementation, and the proportion of linoleic acid increased in platelets and erythrocyte membranes in the DHA oil group as well. These results suggest retroconversion of DHA to EPA and that DHA also interferes with linoleic acid metabolism.