Liquid Chromatographic–Electrospray Ionization–Mass Spectrometric Analysis of Cytochrome P450 Metabolites of Arachidonic Acid

Nithipatikom, Kasem; Grall, Andrew J.; Holmes, Blythe B.; Harder, David R.; Falck, John R.; Campbell, William B.

doi:10.1006/abio.2001.5395

Cited by 111 publications

(96 citation statements)

References 41 publications

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“…Eicosanoids were separated by reverse-phase HPLC using a water and acetonitrile mobile phase. 15 The presumptive HPLC peak was identified by an atmospheric pressure ionization-mass spectroscopy method using electrospray ionization and detection in the negative ion mode. 15 Deuterated EETs served as internal standards.…”

Section: Measurement Of 1112-eetmentioning

confidence: 99%

Endothelium-Derived Hyperpolarizing Factor in Human Internal Mammary Artery Is 11,12-Epoxyeicosatrienoic Acid and Causes Relaxation by Activating Smooth Muscle BK _Ca Channels

et al. 2003

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Background-Left internal mammary arteries (LIMAs) synthesize endothelium-derived hyperpolarizing factor (EDHF), a short-lived K ϩ channel activator that persists after inhibition of nitric oxide (NO) and prostaglandin synthesis. EDHF hyperpolarizes and relaxes smooth muscle cells (SMCs). The identity of EDHF in humans is unknown. We hypothesized that EDHF (1) is 11,12-epoxyeicosatrienoic acid (11,12-EET); (2) is generated by cytochrome P450-2C, CYP450-2C; and (3)

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Section: Measurement Of 1112-eetmentioning

confidence: 99%

Endothelium-Derived Hyperpolarizing Factor in Human Internal Mammary Artery Is 11,12-Epoxyeicosatrienoic Acid and Causes Relaxation by Activating Smooth Muscle BK _Ca Channels

et al. 2003

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“…The quantity of EKODE in the final extract and aliquots was determined by 3 techniques: gravimetric measurement; absorbance at 234 nm assuming a molar extinction coefficient of 16 500; 13 and isotope dilution using a trace of 14 C-labeled linoleic acid of known specific activity in the synthetic protocol.…”

Section: Ekode Synthesismentioning

confidence: 99%

“…The molecular weight of EKODE is 310, so the chromatographic eluate was monitored at masses 311 and 315 m/z, representing the mϩ1 ions of EKODE and deuterated EKODE produced by positive ion mass spectrometry. 14 Bioassays were performed using rat adrenal glomerulosa cells isolated from capsules by collagenase digestion. 15 EKODE and other lipids were dissolved in ethanol.…”

Section: Ekode Synthesismentioning

confidence: 99%

Epoxy-Keto Derivative of Linoleic Acid Stimulates Aldosterone Secretion

et al. 2004

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Abstract-Plasma levels of aldosterone are not always predictable from the activity of renin and the concentration of potassium. Among the unexplained are elevated levels of aldosterone in some obese humans. Obesity is characterized by increased plasma fatty acids and oxidative stress. We postulated that oxidized fatty acids stimulate aldosteronogenesis. The most readily oxidized fatty acids are the polyunsaturated, and the most abundant of those is linoleic acid. We tested oxidized derivatives of linoleic acid for effects on rat adrenal cells. One derivative, 12,13-epoxy-9-keto-10(trans)-octadecenoic acid (EKODE), was particularly potent. EKODE stimulated aldosteronogenesis at concentrations from 0.5 to 5 mol/L, and inhibited aldosteronogenesis at higher doses. EKODE's stimulatory effect was most prominent when angiotensin and potassium effects were submaximal. The lipid's mechanism of action was on the early pathway leading to pregnenolone; its action was inhibited by atrial natriuretic peptide. Plasma EKODE was measured by liquid chromatography/mass spectrometry. All human plasmas tested contained EKODE in concentrations ranging from 10 Ϫ9to 5ϫ10 Ϫ7 mol/L. In samples from 24 adults, levels of EKODE correlated directly with aldosterone (rϭ0.53, Pϭ0.007). In the 12 blacks in that cohort, EKODE also correlated with body mass index and systolic pressure. Those other correlations were not seen in white subjects. The results suggest that oxidized derivatives of polyunsaturated fatty acids other than arachidonic are biologically active. Compounds like EKODE, derived from linoleic acid, may affect adrenal steroid production in humans and mediate some of the deleterious effects of obesity and oxidative stress, especially in blacks. Key Words: fatty acids Ⅲ oxidative stress Ⅲ hypertension Ⅲ aldosterone Ⅲ obesity Ⅲ adrenal gland A ldosterone production by adrenal cells in vitro can be affected by more than 20 hormones, autacoids, ions, and nutrients. 1,2 It is probable, therefore, that regulation of aldosterone secretion in intact animals and humans is more complex than classical schemes that involve primarily angiotensin II and potassium. In fact, several authors have invoked unknown factors to explain levels of aldosterone that do not comply with predictions based on classical regulators of the adrenal. [3][4][5] We encountered one such inexplicable situation when we found that aldosterone levels were higher in some subjects with visceral obesity and hypertension than in lean normotensive subjects, even when we took into account the levels of renin activity and potassium. 6 Obesity and hypertension are associated with relatively high levels of fatty acids and increased oxidative stress. 7-10 These facts led us to postulate that oxidized derivatives of fatty acids might drive aldosterone secretion and possibly explain the relatively high levels of aldosterone in some subjects with visceral obesity. The most readily oxidized fatty acids are polyunsaturated, and the most prevalent polyunsaturated fatty acid in humans is l...

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“…At 0, 10, 30, 60, and 120 minutes, an aliquot (5 L) was analyzed for 5,6-EET and PTPA by liquid chromatography/mass spectrometry (LC/MS) using a modification of a method described previously. 18 The LC/MS analysis used an Agilent 1100 mass selective detection mass spectrometer with electrospray ionization (ESI). The eicosanoids were separated on a Phemomenex Prodigy C-18 (5 m; 1.0ϫ50 mm) column.…”

Section: Stability Of 56-eet and Ptpamentioning

confidence: 99%

Stable 5,6-Epoxyeicosatrienoic Acid Analog Relaxes Coronary Arteries Through Potassium Channel Activation

et al. 2005

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Abstract-5,6-Epoxyeicosatrienoic acid (5,6-EET) is a cytochrome P450 epoxygenase metabolite of arachidonic acid that causes vasorelaxation. However, investigations of its role in biological systems have been limited by its chemical instability. We developed a stable agonist of 5,6-EET, 5-(pentadeca-3(Z),6(Z),9(Z)-trienyloxy)pentanoic acid (PTPA), in which the 5,6-epoxide was replaced with a 5-ether. PTPA obviates chemical and enzymatic hydrolysis. In bovine coronary artery rings precontracted with U46619, PTPA (1 nmol/L to 10 mol/L) induced concentration-dependent relaxations, with maximal relaxation of 86Ϯ5% and EC 50 of 1 mol/L. The relaxations were inhibited by the cyclooxygenase inhibitor indomethacin (10 mol/L; max relaxation 43Ϯ9%); the ATP-sensitive K ϩ channel inhibitor glybenclamide (10 mol/L; max relaxation 49Ϯ6%); and the large conductance calcium-activated K ϩ channel inhibitor iberiotoxin (100 nmol/L; max relaxation 38Ϯ6%) and abolished by the combination of iberiotoxin with indomethacin or glybenclamide or increasing extracellular K ϩ to 20 mmol/L. Whole-cell outward K ϩ current was increased nearly 6-fold by PTPA (10 mol/L), which was also blocked by iberiotoxin. Additionally, we synthesized 5-(pentadeca-6(Z),9(Z)-dienyloxy)pentanoic acid and 5-(pentadeca-3(Z),9(Z)-dienyloxy)pentanoic acid (PDPA), PTPA analogs that lack the 8,9 or 11,12 double bonds of arachidonic acid and therefore are not substrates for cyclooxygenase. The PDPAs caused concentration-dependent relaxations (max relaxations 46Ϯ13% and 52Ϯ7%, respectively; EC 50 1mol/L), which were not altered by glybenclamide but blocked by iberiotoxin. These studies suggested that PTPA induces relaxation through 2 mechanisms: (1) cyclooxygenase-dependent metabolism to 5-ether-containing prostaglandins that activate ATP-sensitive K ϩ channels and (2) Key Words: arachidonic acids Ⅲ cyclooxygenase Ⅲ endothelium-derived factors C ytochrome P450 epoxygenases metabolize arachidonic acid to 4 regioisomeric epoxyeicosatrienoic acids (EETs): 14,15-EET, 11,12-EET, 8,9-EET, and 5,6-EET. 1 The EETs have a range of biological activities including vasodilation, 2,3 release of catecholamines 4 and hypothalamic hormones, 5 and alteration of sodium transport. 6 In some instances, the 4 EET regioisomers are equipotent in biological activity, 2 and in other instances, a particular regioisomer is active, whereas other isomers are inactive or less active. 7 EETs are metabolized by soluble epoxide hydrolase to dihydroxyeicosatrienoic acids (DHETs) 8 by ␤-oxidation to short-chain epoxides 9 or by incorporation into membrane phospholipids. 10 These pathways represent mechanisms of cellular inactivation and EET recycling. With the exception of 5,6-EET, the EETs are chemically stable in alkaline aqueous solution. 5,6-EET is hydrolyzed to 5,6-DHET and its ␦-lactone under neutral and acidic conditions. 11,12 In some studies, the methyl ester of 5,6-EET is used because of its improved chemical stability. The chemical instability has limited investigation of this EET regioiso...

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Liquid Chromatographic–Electrospray Ionization–Mass Spectrometric Analysis of Cytochrome P450 Metabolites of Arachidonic Acid

Cited by 111 publications

References 41 publications

Endothelium-Derived Hyperpolarizing Factor in Human Internal Mammary Artery Is 11,12-Epoxyeicosatrienoic Acid and Causes Relaxation by Activating Smooth Muscle BK _Ca Channels

Endothelium-Derived Hyperpolarizing Factor in Human Internal Mammary Artery Is 11,12-Epoxyeicosatrienoic Acid and Causes Relaxation by Activating Smooth Muscle BK _Ca Channels

Epoxy-Keto Derivative of Linoleic Acid Stimulates Aldosterone Secretion

Stable 5,6-Epoxyeicosatrienoic Acid Analog Relaxes Coronary Arteries Through Potassium Channel Activation

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Liquid Chromatographic–Electrospray Ionization–Mass Spectrometric Analysis of Cytochrome P450 Metabolites of Arachidonic Acid

Cited by 111 publications

References 41 publications

Endothelium-Derived Hyperpolarizing Factor in Human Internal Mammary Artery Is 11,12-Epoxyeicosatrienoic Acid and Causes Relaxation by Activating Smooth Muscle BK Ca Channels

Endothelium-Derived Hyperpolarizing Factor in Human Internal Mammary Artery Is 11,12-Epoxyeicosatrienoic Acid and Causes Relaxation by Activating Smooth Muscle BK Ca Channels

Epoxy-Keto Derivative of Linoleic Acid Stimulates Aldosterone Secretion

Stable 5,6-Epoxyeicosatrienoic Acid Analog Relaxes Coronary Arteries Through Potassium Channel Activation

Contact Info

Product

Resources

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Endothelium-Derived Hyperpolarizing Factor in Human Internal Mammary Artery Is 11,12-Epoxyeicosatrienoic Acid and Causes Relaxation by Activating Smooth Muscle BK _Ca Channels

Endothelium-Derived Hyperpolarizing Factor in Human Internal Mammary Artery Is 11,12-Epoxyeicosatrienoic Acid and Causes Relaxation by Activating Smooth Muscle BK _Ca Channels