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1. Products of metabolism of arachidonic acid, such as 20-hydroxyeicosatetraenoic acid (20-HETE), thromboxane A2 (TXA2) and prostaglandin I2 (PGI2), regulate vascular tone. Among them, 20-HETE is a potent constrictor in small arteries that also has natriuretic properties. The present study investigated changes in urinary concentrations of 20-HETE and metabolites of TXA2 and PGI2 in glucocorticoid-hypertension in rats, a sodium-independent model. 2. Male Sprague-Dawley rats were treated with saline, adrenocorticotrophic hormone (ACTH; 0.2 mg/kg) or dexamethasone (20 µg/kg) by daily s.c. injection for 12 days. Systolic blood pressure (SBP) was measured using the tail-cuff method. Metabolic cages were used for 24 h urine collection. Thymus weight and urinary concentrations of 20-HETE, TXA2 and PGI2 were determined. 3. In the present study, SBP was increased by both ACTH (from 102 ± 2 to 134 ± 7 mmHg; n = 10; P < 0.01) and dexamethasone (from 106 ± 5 to 122 ± 4 mmHg; n = 10; P < 0.01). Thymus weight, a marker for glucocorticoid activity, was significantly decreased by both ACTH and dexamethasone (56 ± 9 and 76 ± 5 mg/100 g bodyweight, respectively; n = 10; P′ < 0.01) compared with the saline control (151 ± 5 mg/100 g bodyweight; n = 20). Urinary 20-HETE excretion was increased by ACTH (501 ± 115 pmol/g creatinine; n = 10; P′ < 0.05) but not by dexamethasone (126 ± 13 pmol/g creatinine; n = 10) compared with the saline control (219 ± 54 pmol/g creatinine; n = 20). Neither ACTH nor dexamethasone affected urinary excretion of TXB2 or PGI2 compared with the saline control. 4. In conclusion, ACTH but not dexamethasone increased urinary 20-HETE excretion in male Sprague-Dawley rats. Urinary concentrations of the metabolites TXB2 and PGI2 were unchanged in both models of glucocorticoid-hypertension. The vasoconstrictor 20-HETE may play a role in the genesis of ACTH-induced hypertension. [ABSTRACT FROM AUTHOR]

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Abstract: There is considerable evidence that chronic moderate-to-high alcohol consumption increases blood pressure. The mechanisms by which this occurs are not clear. Alcohol consumption can induce oxidative stress and cytochrome P450 (CYP450) isoforms that are associated with oxidative stress and may influence vascular tone. To study the role of such mechanisms we examined whether reducing alcohol intake in moderate-to-heavy drinkers (40–110 g/day) resulted in changes in urinary excretion of 20-HETE, a CYP450 metabolite of arachidonic acid, and plasma and urinary F2-isoprostanes as markers of lipid peroxidation. After a 4-week run-in period during which healthy men maintained their usual drinking pattern they were randomized to a two-way crossover intervention study. In each of the 4-week treatment periods subjects either substituted their usual alcohol intake with a 0.9% alcohol beer or maintained their usual alcohol intake. Plasma and urinary F2-isoprostanes and urinary 20-HETE were measured by gas chromatography mass spectrometry, and serum γ-glutamyl transpeptidase (γ-GT) was measured as a biomarker of alcohol consumption, at the end of each study period. Sixteen healthy men age 51.0±2.7 years and with a BMI of 26.4±0.61 kg/m2 completed the study. The reductions in alcohol intake (72.4±5.0 vs 7.9±1.6 g/day, p <0.001) and serum γ-GT (geometric mean 24.4 U/L (95% CI 19.7, 30.2) vs 18.6 U/L (95% CI 15.5, 22.2, p <0.01) were accompanied by a significant fall in blood pressure as well as urinary 20-HETE excretion (158±23 vs 109±19 pmol/mmol creatinine, p <0.001) and plasma F2-isoprostanes (3438±158 vs 2929±145 pmol/L, p =0.01). A substantial reduction in alcohol consumption in healthy men lowered plasma F2-isoprostanes and urinary 20-HETE. Increased oxidative stress and 20-HETE production may be linked, at least in part, to the pathogenesis of alcohol-related hypertension. [Copyright &y& Elsevier]

The anti-inflammatory properties of n-3 polyunsaturated fatty acids (n-3 PUFA) have suggested a potential role of these nutrients in dietary modification for prevention of allergic disease in early life. As oxidative stress is known to modify antigen presenting cell (APC) signalling and resulting immune responses, we examined the effects of maternal n-3 PUFA supplementation in pregnancy on markers of oxidative stress and APC function in neonates at high risk of allergy. Eighty-three pregnant atopic women were randomised to receive 4 g daily of either fish oil ( n =40) or olive oil ( n =43) capsules in a controlled trial from 20 weeks gestation until delivery. Plasma (cord blood) and urinary F 2 -isoprostanes were measured as markers of lipid peroxidation. Cord erythrocyte fatty acids and markers of APC function (HLA-DR expression and cytokine responses) were measured and related to levels of plasma F 2 -isoprostanes. Maternal fish oil supplementation lowered plasma ( p <0.0001) and urinary ( p =0.06) F 2 -isoprostanes. HLA-DR expression on APC was not different between the groups. In multiple regression analysis, 28.8% of the variance in plasma F 2 -isoprostanes was explained by positive relationships with erythrocyte arachidonic acid (AA) and monocyte HLA-DR expression and a negative relationship with erythrocyte eicosapentaenoic acid (EPA). This study shows that maternal supplementation with fish oil can attenuate neonatal lipid peroxidation. Clinical follow-up of these infants will help to determine if there are sustained effects on postnatal oxidative stress and expression of allergic disease. [ABSTRACT FROM AUTHOR]

Abstract: 20-Hydroxyeicosatetraenoic acid (20-HETE) is a cytochrome P450 metabolite of arachidonic acid that regulates vascular function and sodium homeostasis. Studies showing an association between 20-HETE excretion, raised BMI, and oxidative stress suggest that 20-HETE may be important in the development of cardiovascular disease in the metabolic syndrome (MetS). We investigated whether 20-HETE and F2-isoprostanes (markers of oxidative stress) were altered in the MetS before and after weight reduction. A case-controlled comparison of 30 participants with the MetS and matched controls showed that plasma and urinary 20-HETE and F2-isoprostanes were significantly elevated in the MetS group. There was a significant gender×group interaction such that women with the MetS had higher urinary 20-HETE and F2-isoprostanes compared to controls (p <0.0001). In a randomized controlled trial, 42 participants with the MetS were assigned to 16 weeks of weight maintenance or a 12-week weight-loss program followed by 4 weeks weight stabilization. Relative to the weight-maintenance group, a 4-kg loss in weight resulted in a 2-mm Hg fall in blood pressure (BP) but did not alter urinary or plasma 20-HETE or F2-isoprostanes. 20-HETE and oxidative stress may be important mediators of cardiovascular disease risk in the MetS. Although a 4% reduction in body weight reduced BP, there were no changes in plasma or urinary 20-HETE or F2-isoprostanes. [Copyright &y& Elsevier]

Abstract: The measurement of urinary F2-isoprostanes is noninvasive and widely used to assess in vivo oxidative stress in humans. Most studies measure urinary F2-isoprostanes in the free form; however, many eicosanoids are excreted as urine glucuronide conjugates. Using gas chromatography–mass spectrometry, we examined the extent of glucuronide conjugation of F2-isoprostanes in urine collected from healthy men (n =20) and women (n =15). Incubation of urine with exogenous glucuronidase led to F2-isoprostane concentrations that were approximately 40% higher than untreated samples (P <0.001). We conclude that a significant proportion of F2-isoprostanes in urine are conjugated as glucuronides. [Copyright &y& Elsevier]

Oxidant stress has been identified as important in the pathology of many diseases. Oxidation products of polyunsaturated fatty acids collectively termed isoprostanes, neuroprostanes, and isofurans are considered the most reliable measures of in vivo lipid oxidation, and they are widely used to assess oxidant stress in various diseases. Here we describe the measurement of these lipid oxidation products using gas chromatography mass spectrometry with electron capture negative ionization.

Reperfusion injury is characterized by significant oxidative stress. F(2)-isoprostanes (F(2)-IsoP's) and isofurans (IsoF's), the latter preferentially produced during increased oxygen tension, are recognized markers of in vivo oxidative stress. We aimed to determine whether increasing oxygen tension during reperfusion modified levels of plasma total IsoF's and F(2)-IsoP's. Forty-five patients undergoing upper-limb surgery were randomized to receive inspired oxygen concentrations of 30, 50, or 80% during the last 15 min of surgery. Venous blood samples were taken before the change in inspired oxygen, after 10 min (before reperfusion), and after 15 min (5 min after reperfusion). IsoF's and F(2)-IsoP's were measured by gas chromatography-mass spectrometry. Venous oxygen tension and hemoglobin concentrations were also measured. Plasma IsoF and F(2)-IsoP levels in the 50 and 80% O(2) groups were not significantly different from those of the 30% O(2) group. In secondary analyses, using data combining all groups, levels of IsoF's, but not F(2)-IsoP's, associated with higher venous oxygen tension (P=0.038). Hemoglobin negatively modified the influence of oxygen tension on levels of IsoF's (P=0.014). This study has shown, for the first time, that plasma IsoF levels associate with higher oxygen tension in a human model of reperfusion, and this effect is significantly attenuated by hemoglobin., (Copyright © 2011 Elsevier Inc. All rights reserved.)

General and spinal anesthesia are used extensively in orthopedic surgery. However, these methods of anesthesia may result in different amounts of oxygen being delivered to the patient. Ischemia/reperfusion injury after release of the tourniquet initiates free radical-mediated oxidative stress. F₂-isoprostanes are reliable markers of in vivo lipid peroxidation. However, under conditions of high oxygen tension, isofurans are formed. We aimed to compare plasma isofurans and F₂-isoprostanes in spinal versus general anesthesia in patients undergoing knee-replacement surgery in a randomized, blinded study. Thirty-nine patients were randomized to spinal (SA; n = 19) or general anesthesia (GA; n = 20). Blood was collected before anesthesia, and a tourniquet was then applied to the limb during surgery. After release of the tourniquet, blood samples were collected at 30 min, 2 h, and 24 h for measurement of plasma F₂-isoprostanes and isofurans by gas chromatography-mass spectrometry. The two groups were comparable in age and body mass index. Plasma F₂-isoprostanes were significantly lower in the GA patients compared with the SA patients (p = 0.045). In contrast, the GA patients had significantly elevated plasma isofurans (p = 0.032). Increased isofurans during GA compared with SA are likely to reflect increased oxidative stress due to elevated oxygen concentrations during GA. Further studies are required to assess the implications of these findings on perioperative outcomes., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Analysis of F(2)-isoprostanes in urine using gas chromatography-mass spectrometry is confounded by the presence of endogenous compounds interfering with the internal standard, 15-F(2t)-IsoP-d(4) (m/z 573). Previous efforts to resolve the 15-F(2t)-IsoP-d(4) from co-eluting peaks with different solid phase extractions were unsuccessful. This study has now used a highly-deuterated, d(9)-analogue of the derivatization agent N,O-Bis(trimethyl-d(9)-silyl) trifluoroacetamide (BSTFA-d(9)) yielding trimethylsilyl ethers, but this was not successful in resolving the 15-F(2t)-IsoP-d(4) from co-eluting peaks. It was hypothesized that interfering peaks at m/z 573 could be the tetrahydro analogue of 15-F(2t)-IsoP. However, using an authentic standard showed the interfering peaks are not due to this metabolite. In subsequent experiments good resolution was shown of the 15-F(2t)-IsoP peak using 8-F(2t)-IsoP-d(4) (m/z 573) as the internal standard. These data show that care must be taken when using GC-MS for quantitation of F(2)-IsoPs to prevent interfering substances affecting the results.

Supplementation with eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) has been reported to reduce lipid peroxidation products formed from arachidonic acid (F(2)-isoprostanes) in healthy humans, as well as in those under oxidative stress. alpha-Linolenic acid (ALA) is a precursor to EPA and DHA; however, its conversion in humans is thought to be inefficient. ALA can also undergo free radical oxidation, forming compounds known as F(1)-phytoprostanes, which are found in all plants and are in high concentrations in plant pollens. In this study, we examined the effect of ALA supplementation on plasma and urine F(1)-phytoprostane and F(2)-isoprostane concentrations in men. Thirty-six nonsmoking men, aged 20-65 y, were recruited from the general population and randomly allocated to consume 9 g/d of either flaxseed oil (62% ALA, 5.4 g/d) or olive oil (placebo) for 4 wk in a parallel design. At baseline and after 4 wk of supplementation, blood samples and a 24-h urine sample were collected for measurement of plasma and urinary F(1)-phytoprostanes and F(2)-isoprostanes and plasma fatty acids. Compared with the olive oil group, plasma phospholipid ALA was greater (P < 0.0001), as were F(1)-phytoprostanes in plasma (P = 0.049) and urine (P = 0.06) in the flaxseed oil group after 4 wk supplementation. Flaxseed oil did not affect plasma or urinary F(2)-isoprostanes. The greater plasma F(1)-phytoprostane concentration in the flaxseed oil group most likely resulted from the increased plasma concentration of the ALA substrate and/or the F(1)-phytoprostane content of the flaxseed oil. Future studies are needed to determine the physiological importance of increased plasma and urine F(1)-phytoprostanes and their relevance to heart disease prevention.

Enhanced oxidative stress is implicated in the development of atherosclerosis in humans and animal models. F(2)-isoprostanes are formed in vivo via free radical peroxidation of arachidonic acid, and their quantification has allowed assessment of oxidative stress in vivo. F(2)-isoprostanes associate with lipids, although their distribution in human plasma lipoproteins is unknown. Our aim was to determine the distribution and levels of F(2)-isoprostanes in lipoproteins isolated from human plasma by ultracentrifugation and fast protein liquid chromatography (FPLC). F(2)-isoprostanes were significantly higher in HDL compared with LDL or VLDL after isolation by ultracentrifugation or FPLC. Furthermore, HDL3 particles contained elevated levels of F(2)-isoprostanes compared with HDL2. Platelet activating factor acetylhydrolase (PAF-AH), which hydrolyses esterified F(2)-isoprostanes from phospholipids, was predominantly associated with LDL. Reduced F(2)-isoprostanes in LDL may be related to higher PAF-AH activity in LDL. Paraoxonase 1 (PON-1) activity was associated with HDL2 and may be a contributing factor to the lower F(2)-isoprostanes in HDL2 compared with HDL3. Further studies are required to establish the implications of these findings on HDL function.

20-Hydroxyeicosatetraenoic acid (20-HETE) is a cytochrome P450 metabolite of arachidonic acid that regulates vascular function and sodium homeostasis. Studies showing an association between 20-HETE excretion, raised BMI, and oxidative stress suggest that 20-HETE may be important in the development of cardiovascular disease in the metabolic syndrome (MetS). We investigated whether 20-HETE and F(2)-isoprostanes (markers of oxidative stress) were altered in the MetS before and after weight reduction. A case-controlled comparison of 30 participants with the MetS and matched controls showed that plasma and urinary 20-HETE and F(2)-isoprostanes were significantly elevated in the MetS group. There was a significant gender x group interaction such that women with the MetS had higher urinary 20-HETE and F(2)-isoprostanes compared to controls (p<0.0001). In a randomized controlled trial, 42 participants with the MetS were assigned to 16 weeks of weight maintenance or a 12-week weight-loss program followed by 4 weeks weight stabilization. Relative to the weight-maintenance group, a 4-kg loss in weight resulted in a 2-mm Hg fall in blood pressure (BP) but did not alter urinary or plasma 20-HETE or F(2)-isoprostanes. 20-HETE and oxidative stress may be important mediators of cardiovascular disease risk in the MetS. Although a 4% reduction in body weight reduced BP, there were no changes in plasma or urinary 20-HETE or F(2)-isoprostanes.

1. Products of metabolism of arachidonic acid, such as 20-hydroxyeicosatetraenoic acid (20-HETE), thromboxane A(2) (TXA(2)) and prostaglandin I(2) (PGI(2)), regulate vascular tone. Among them, 20-HETE is a potent constrictor in small arteries that also has natriuretic properties. The present study investigated changes in urinary concentrations of 20-HETE and metabolites of TXA(2) and PGI(2) in glucocorticoid-hypertension in rats, a sodium-independent model. 2. Male Sprague-Dawley rats were treated with saline, adrenocorticotrophic hormone (ACTH; 0.2 mg/kg) or dexamethasone (20 microg/kg) by daily s.c. injection for 12 days. Systolic blood pressure (SBP) was measured using the tail-cuff method. Metabolic cages were used for 24 h urine collection. Thymus weight and urinary concentrations of 20-HETE, TXA(2) and PGI(2) were determined. 3. In the present study, SBP was increased by both ACTH (from 102 +/- 2 to 134 +/- 7 mmHg; n = 10; P < 0.01) and dexamethasone (from 106 +/- 5 to 122 +/- 4 mmHg; n = 10; P < 0.01). Thymus weight, a marker for glucocorticoid activity, was significantly decreased by both ACTH and dexamethasone (56 +/- 9 and 76 +/- 5 mg/100 g bodyweight, respectively; n = 10; P' < 0.01) compared with the saline control (151 +/- 5 mg/100 g bodyweight; n = 20). Urinary 20-HETE excretion was increased by ACTH (501 +/- 115 pmol/g creatinine; n = 10; P' < 0.05) but not by dexamethasone (126 +/- 13 pmol/g creatinine; n = 10) compared with the saline control (219 +/- 54 pmol/g creatinine; n = 20). Neither ACTH nor dexamethasone affected urinary excretion of TXB(2) or PGI(2) compared with the saline control. 4. In conclusion, ACTH but not dexamethasone increased urinary 20-HETE excretion in male Sprague-Dawley rats. Urinary concentrations of the metabolites TXB(2) and PGI(2) were unchanged in both models of glucocorticoid-hypertension. The vasoconstrictor 20-HETE may play a role in the genesis of ACTH-induced hypertension.

Background: Protein and fiber may be important determinants of satiety. Lupin kernel flour is a novel food ingredient that is rich in protein and fiber., Objective: The objective was to investigate the effects of lupin kernel flour-enriched bread (LB) on satiety and energy intake in humans., Design: Two randomized controlled crossover trials were performed to compare the acute effects of LB with those of white bread (WB). In study 1, the subjects (n = 16) completed 4 treatments 1 wk apart: WB breakfast (as toast) and WB lunch (as sandwiches), WB breakfast and LB lunch, LB breakfast and WB lunch, and LB breakfast and LB lunch. Energy intake at all breakfast meals was matched (1655 kJ), and ad libitum energy intake at lunch, 3 h after breakfast, was measured. In study 2, the subjects (n = 17) completed 2 treatments 1 wk apart: WB breakfast and LB breakfast (each 1655 kJ). Blood samples were taken at baseline and at regular intervals for 3 h after breakfast., Results: In study 1, the LB breakfast resulted in significantly higher self-reported satiety (P < 0.001) and lower energy intake (kJ) at lunch (-488; 95% CI: -798, -178) than did the WB breakfast. The LB lunch resulted in a significantly lower within-meal energy intake (kJ) at lunch (-1028; 95% CI: -1338, -727) than did the WB lunch. In study 2, compared with the WB breakfast, the LB breakfast significantly altered the 3-h postmeal plasma ghrelin response (P = 0.04) and resulted in significantly lower mean 3-h plasma ghrelin concentrations (P = 0.009)., Conclusion: A novel food enriched in protein and fiber derived from lupin kernel flour significantly influences energy intake acutely.

n--3 Fatty acids derived from fish oil reduce plasma triacylglycerols (triglycerides) and increase HDL-C (high-density lipoprotein cholesterol); however, the effect of n--3 fatty acid supplementation during pregnancy, a hyperlipidaemic state, remains unknown. We took the opportunity to investigate maternal lipid levels and blood pressure during and after pregnancy, and fetal lipid levels at birth, in a study that aimed primarily to examine the effect of fish oil supplementation during pregnancy on immune function in infants born to women with allergic disease. Eighty-three pregnant women who had allergic disease, but were otherwise healthy, completed the study. They were randomly allocated to receive fish oil or olive oil capsules, taken as 4 g/day, from 20 weeks of pregnancy until delivery. Compared with olive oil, fish oil supplementation did not alter triacylglycerols, total cholesterol, LDL-C (low-density lipoprotein cholesterol) or HDL-C during or after pregnancy. There was also no effect of fish oil on cord blood triacylglycerols, total cholesterol, LDL-C or HDL-C. Fish oil supplementation during pregnancy did not alter maternal blood pressure during or after pregnancy. The effects of fish oil on lipids and blood pressure in non-pregnant individuals appear to be lost when it is administered during pregnancy.