12(R)-Hydroxyeicosatrienoic acid

A vasodilator cytochrome P-450-dependent arachidonate metabolite from the bovine corneal epithelium

R. C. Murphy, J R Falck, S. Lumin, P. Yadagiri, J. A. Zirrolli, M. Balazy, J. L. Masferrer, N. G. Abraham, M. L. Schwartzman

Research output: Contribution to journalArticle

81 Citations (Scopus)

Abstract

When corneal microsomes were incubated with arachidonic acid in the presence of an NADPH-generating system, two biologically active metabolites of arachidonic acid were formed. The structure of one of the metabolites, compound C, was previously reported to be 12(R)-hydroxy-5,8,10,14-eicosatetraenoic acid and was found to be a potent inhibitor of the Na+/K+-ATPase in the cornea. The second metabolite, compound D, was found to be a potent vasodilator as well as having the property of stimulating protein influx into the aqueous humor of the eye. Following purification of compound D by thin layer chromatography and high pressure liquid chromatography, it was found to lack a UV chromophore in contrast to the previously reported cytochrome P-450-dependent metabolite. Mass spectrometric analysis using positive and negative ionization modes was carried out on derivatized compound D that had been synthesized from a mixture of labeled ([5,6,8,9,11,12,14,15-2H8]) and unlabeled arachidonic acid incubated with corneal microsomes. The novel arachidonate metabolite had abundant fragment ions consistent with compound D being a monooxygenated derivative of arachidonic acid with a hydroxyl substituent at carbon 12 of the eicosanoid backbone; only seven deuterium atoms from [2H8]arachidonate were retained in the structure. Oxidative ozonolysis yielded a product indicating that the double bonds in metabolite D resided between carbons at positions 8 and 9 and positions 14 and 15 of the 20-carbon chain. Compound D was therefore characterized as 12-hydroxy-5,8,14-eicosatrienoic acid. Model compounds were synthesized from dimethyl malate with the hydroxy at the 12 position with both the R and S absolute configuration and with all double bonds of the cis configuration. Only the 12(R) isomer was found to be a potent vasodilator and to increase aqueous humor protein concentration, suggesting that the biologically active compound D was 12(R)-hydroxy-5,8,14-(Z,Z,Z)-eicosatrienoic acid. As this compound possesses proinflammatory properties, it may play a role in the wound-healing processes of corneal injury.

Original languageEnglish (US)
Pages (from-to)17197-17202
Number of pages6
JournalJournal of Biological Chemistry
Volume263
Issue number32
StatePublished - 1988

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Corneal Epithelium
Metabolites
Vasodilator Agents
Arachidonic Acid
Cytochrome P-450 Enzyme System
Acids
Carbon
Aqueous Humor
Microsomes
12-Hydroxy-5,8,10,14-eicosatetraenoic Acid
Eicosanoids
Deuterium
Thin Layer Chromatography
NADP
High pressure liquid chromatography
Hydroxyl Radical
Wound Healing
Cornea
Arachidonic Acids
Thin layer chromatography

ASJC Scopus subject areas

  • Biochemistry

Cite this

Murphy, R. C., Falck, J. R., Lumin, S., Yadagiri, P., Zirrolli, J. A., Balazy, M., ... Schwartzman, M. L. (1988). 12(R)-Hydroxyeicosatrienoic acid: A vasodilator cytochrome P-450-dependent arachidonate metabolite from the bovine corneal epithelium. Journal of Biological Chemistry, 263(32), 17197-17202.

12(R)-Hydroxyeicosatrienoic acid : A vasodilator cytochrome P-450-dependent arachidonate metabolite from the bovine corneal epithelium. / Murphy, R. C.; Falck, J R; Lumin, S.; Yadagiri, P.; Zirrolli, J. A.; Balazy, M.; Masferrer, J. L.; Abraham, N. G.; Schwartzman, M. L.

In: Journal of Biological Chemistry, Vol. 263, No. 32, 1988, p. 17197-17202.

Research output: Contribution to journalArticle

Murphy, RC, Falck, JR, Lumin, S, Yadagiri, P, Zirrolli, JA, Balazy, M, Masferrer, JL, Abraham, NG & Schwartzman, ML 1988, '12(R)-Hydroxyeicosatrienoic acid: A vasodilator cytochrome P-450-dependent arachidonate metabolite from the bovine corneal epithelium', Journal of Biological Chemistry, vol. 263, no. 32, pp. 17197-17202.
Murphy, R. C. ; Falck, J R ; Lumin, S. ; Yadagiri, P. ; Zirrolli, J. A. ; Balazy, M. ; Masferrer, J. L. ; Abraham, N. G. ; Schwartzman, M. L. / 12(R)-Hydroxyeicosatrienoic acid : A vasodilator cytochrome P-450-dependent arachidonate metabolite from the bovine corneal epithelium. In: Journal of Biological Chemistry. 1988 ; Vol. 263, No. 32. pp. 17197-17202.
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abstract = "When corneal microsomes were incubated with arachidonic acid in the presence of an NADPH-generating system, two biologically active metabolites of arachidonic acid were formed. The structure of one of the metabolites, compound C, was previously reported to be 12(R)-hydroxy-5,8,10,14-eicosatetraenoic acid and was found to be a potent inhibitor of the Na+/K+-ATPase in the cornea. The second metabolite, compound D, was found to be a potent vasodilator as well as having the property of stimulating protein influx into the aqueous humor of the eye. Following purification of compound D by thin layer chromatography and high pressure liquid chromatography, it was found to lack a UV chromophore in contrast to the previously reported cytochrome P-450-dependent metabolite. Mass spectrometric analysis using positive and negative ionization modes was carried out on derivatized compound D that had been synthesized from a mixture of labeled ([5,6,8,9,11,12,14,15-2H8]) and unlabeled arachidonic acid incubated with corneal microsomes. The novel arachidonate metabolite had abundant fragment ions consistent with compound D being a monooxygenated derivative of arachidonic acid with a hydroxyl substituent at carbon 12 of the eicosanoid backbone; only seven deuterium atoms from [2H8]arachidonate were retained in the structure. Oxidative ozonolysis yielded a product indicating that the double bonds in metabolite D resided between carbons at positions 8 and 9 and positions 14 and 15 of the 20-carbon chain. Compound D was therefore characterized as 12-hydroxy-5,8,14-eicosatrienoic acid. Model compounds were synthesized from dimethyl malate with the hydroxy at the 12 position with both the R and S absolute configuration and with all double bonds of the cis configuration. Only the 12(R) isomer was found to be a potent vasodilator and to increase aqueous humor protein concentration, suggesting that the biologically active compound D was 12(R)-hydroxy-5,8,14-(Z,Z,Z)-eicosatrienoic acid. As this compound possesses proinflammatory properties, it may play a role in the wound-healing processes of corneal injury.",
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T2 - A vasodilator cytochrome P-450-dependent arachidonate metabolite from the bovine corneal epithelium

AU - Murphy, R. C.

AU - Falck, J R

AU - Lumin, S.

AU - Yadagiri, P.

AU - Zirrolli, J. A.

AU - Balazy, M.

AU - Masferrer, J. L.

AU - Abraham, N. G.

AU - Schwartzman, M. L.

PY - 1988

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N2 - When corneal microsomes were incubated with arachidonic acid in the presence of an NADPH-generating system, two biologically active metabolites of arachidonic acid were formed. The structure of one of the metabolites, compound C, was previously reported to be 12(R)-hydroxy-5,8,10,14-eicosatetraenoic acid and was found to be a potent inhibitor of the Na+/K+-ATPase in the cornea. The second metabolite, compound D, was found to be a potent vasodilator as well as having the property of stimulating protein influx into the aqueous humor of the eye. Following purification of compound D by thin layer chromatography and high pressure liquid chromatography, it was found to lack a UV chromophore in contrast to the previously reported cytochrome P-450-dependent metabolite. Mass spectrometric analysis using positive and negative ionization modes was carried out on derivatized compound D that had been synthesized from a mixture of labeled ([5,6,8,9,11,12,14,15-2H8]) and unlabeled arachidonic acid incubated with corneal microsomes. The novel arachidonate metabolite had abundant fragment ions consistent with compound D being a monooxygenated derivative of arachidonic acid with a hydroxyl substituent at carbon 12 of the eicosanoid backbone; only seven deuterium atoms from [2H8]arachidonate were retained in the structure. Oxidative ozonolysis yielded a product indicating that the double bonds in metabolite D resided between carbons at positions 8 and 9 and positions 14 and 15 of the 20-carbon chain. Compound D was therefore characterized as 12-hydroxy-5,8,14-eicosatrienoic acid. Model compounds were synthesized from dimethyl malate with the hydroxy at the 12 position with both the R and S absolute configuration and with all double bonds of the cis configuration. Only the 12(R) isomer was found to be a potent vasodilator and to increase aqueous humor protein concentration, suggesting that the biologically active compound D was 12(R)-hydroxy-5,8,14-(Z,Z,Z)-eicosatrienoic acid. As this compound possesses proinflammatory properties, it may play a role in the wound-healing processes of corneal injury.

AB - When corneal microsomes were incubated with arachidonic acid in the presence of an NADPH-generating system, two biologically active metabolites of arachidonic acid were formed. The structure of one of the metabolites, compound C, was previously reported to be 12(R)-hydroxy-5,8,10,14-eicosatetraenoic acid and was found to be a potent inhibitor of the Na+/K+-ATPase in the cornea. The second metabolite, compound D, was found to be a potent vasodilator as well as having the property of stimulating protein influx into the aqueous humor of the eye. Following purification of compound D by thin layer chromatography and high pressure liquid chromatography, it was found to lack a UV chromophore in contrast to the previously reported cytochrome P-450-dependent metabolite. Mass spectrometric analysis using positive and negative ionization modes was carried out on derivatized compound D that had been synthesized from a mixture of labeled ([5,6,8,9,11,12,14,15-2H8]) and unlabeled arachidonic acid incubated with corneal microsomes. The novel arachidonate metabolite had abundant fragment ions consistent with compound D being a monooxygenated derivative of arachidonic acid with a hydroxyl substituent at carbon 12 of the eicosanoid backbone; only seven deuterium atoms from [2H8]arachidonate were retained in the structure. Oxidative ozonolysis yielded a product indicating that the double bonds in metabolite D resided between carbons at positions 8 and 9 and positions 14 and 15 of the 20-carbon chain. Compound D was therefore characterized as 12-hydroxy-5,8,14-eicosatrienoic acid. Model compounds were synthesized from dimethyl malate with the hydroxy at the 12 position with both the R and S absolute configuration and with all double bonds of the cis configuration. Only the 12(R) isomer was found to be a potent vasodilator and to increase aqueous humor protein concentration, suggesting that the biologically active compound D was 12(R)-hydroxy-5,8,14-(Z,Z,Z)-eicosatrienoic acid. As this compound possesses proinflammatory properties, it may play a role in the wound-healing processes of corneal injury.

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