Preparation and purification of lipid hydroperoxides from arachidonic and γ‐linolenic acids

Funk, Max O.; Isaac, Ramdas; Porter, Ned A.

doi:10.1007/bf02532660

Cited by 224 publications

(77 citation statements)

References 15 publications

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“…As an in vitro PGI2 synthetase inhibitor (12), this was synthesized enzymatically with soybean lipoxygenase and purified as previously described (13). PG standards.…”

Section: Methodsmentioning

confidence: 99%

Platelet and Blood Vessel Arachidonate Metabolism and Interactions

Needleman¹,

Wyche²,

Raz³

1979

J. Clin. Invest.

179

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A B S T R A C T Exogenous arachidonate addition to intact platelets, in the absence or the presence of blood vessel microsomes, results in the production of thromboxane B2 (the stable degradation product of thromboxane A2) only. Prostaglandin (PG) endoperoxides are released from intact platelets only when thromboxane synthetase is inhibited. Thus, addition of exogenous arachidonate to imidazole-pretreated platelets in the presence of bovine aorta microsomes (source of prostacyclin synthetase) results predominantly in the synthesis of 6-keto-PGF,a (the stable degradation product of prostacyclin). Strips of intact aorta were removed from aspirin-treated rabbits, thus the isolated blood vessels were unable to convert endogenous or exogenous arachidonate to prostacyclin. Human platelets, with ['4C]arachidonate-labeled phospholipids, adhered to the blood vessel segments and released some thromboxane B2. The subsequent addition of thrombin facilitated the release of endogenous arachidonate and thromboxane, but no labeled 6-keto-PGF,, was detectable. There is therefore no direct chemical evidence of PG-endoperoxide release from human platelets during either aggregation or adhesion, which therefore precludes the possibility that blood vessels use platelet PG-endoperoxide for prostacyclin synthesis. Imidazole inhibited the thromboxane synthetase in the labeled platelets, and thereafter thrombin stimulation resulted in the release of platelet-derived, labeled PG-endoperoxides that were converted to labeled prostacyclin by the vascular prostacyclin synthetase. The latter result suggests a potential antithrombotic therapeutic benefit might be achieved using an effective thromboxane synthetase inhibitor.

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“…As an in vitro PGI2 synthetase inhibitor (12), this was synthesized enzymatically with soybean lipoxygenase and purified as previously described (13). PG standards.…”

Section: Methodsmentioning

confidence: 99%

Platelet and Blood Vessel Arachidonate Metabolism and Interactions

Needleman¹,

Wyche²,

Raz³

1979

J. Clin. Invest.

179

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“…The hydroperoxy eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids, the two major fatty acids of fish oil (19), were analyzed by HPLC as described below. Separation of different lipid fractions in lymph was achieved by TLC (20).…”

Section: Methodsmentioning

confidence: 99%

“…Oxidized triglycerides were digested with pancreatic lipase (21) and purified by TLC (20 concentration that is found in rat bile ( 15 ), and measured luminal hydroperoxide recovery. The results show that GSH supplementation significantly attenuated peroxide accumulation in all segments of the intestine ( Fig.…”

Section: Methodsmentioning

confidence: 99%

Biliary glutathione promotes the mucosal metabolism of luminal peroxidized lipids by rat small intestine in vivo.

Aw¹

1994

J. Clin. Invest.

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We previously found that exogenous GSH enhances mucosal GSH and promotes lipid hydroperoxide metabolism by rat small intestine (Aw, T. Y., and M. W. Williams. 1992. Am. J. PhysioL 263:G665-G672). In this study, we have developed an in vivo bile and lymph fistula rat model to test the hypothesis that biliary GSH is an important luminal source of GSH. Peroxidized fish oil was infused into the proximal intestine, and hydroperoxide accumulation in lumen, mucosa, and lymph was determined. Diversion of bile decreased mucosal GSH and increased hydroperoxide accumulation in all fractions. Supplementation with GSH, but not with GSSG, increased tissue GSH and attenuated hydroperoxide accumulation (50-60%), consistent with enhancement of hydroperoxide removal by exogenous GSH. Addition of native bile deficient in GSH, but not cysteine, cystine, or GSSG, decreased luminal and lymph hydroperoxide levels by 20-30%. Amino acid supplementation concurrently attenuated hydroperoxide recoveries in these fractions by 30-40% and increased mucosal GSH by 40%, indicating a role for biliary amino acids in hydroperoxide elimination. The effect of amino acids was abolished by buthionine sulfoximine, confirming their role in GSH biosynthesis. Collectively, the results demonstrate that bile is a rich source of reductant for maintaining mucosal GSH to promote intestinal metabolism of luminal peroxidized lipids. (J. Cmn. Invest. 1994Invest. .94:1218Invest. -1225.) Key words: omega-3 fatty acid hydroperoxides * en-

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“…Currently, researchers prepare their own in-house reference LOOH [i.e., lipids (e.g., phospholipids, cholesterols, triacylglycerols, and fatty acids)] are subjected to photo- (12), free radical (13), or enzymatic oxidation (14,15). The resultant crude or partly purified LOOH is generally used as a calibrator.…”

mentioning

confidence: 99%

Preparation of pure lipid hydroperoxides

Ibusuki

Nakagawa

Asai

et al. 2008

Journal of Lipid Research

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Increasing evidence of lipid peroxidation in food deterioration and pathophysiology of diseases have revealed the need for a pure lipid hydroperoxide (LOOH) reference as an authentic standard for quantification and as a compound for biological studies in this field. Generally, LOOH is prepared from photo-or enzymatically oxidized lipids; however, separating LOOH from other oxidation products and preparing pure LOOH is difficult. Early studies showed the usability of reaction between hydroperoxide and vinyl ether for preparation of pure LOOH. Because the reactivity of vinyl ether with LOOHs other than fatty acid hydroperoxides has never been reported, here, we employed the reaction for preparation of a wide variety of pure LOOHs. Phospholipid, cholesteryl ester, triacylglycerol, or fatty acid was photo-or enzymatically oxidized; the resultant crude sample containing hydroperoxide was allowed to react with a vinyl ether [2-methoxypropene (MxP)]. Liquid chromatography (LC) and mass spectrometry confirmed that MxP selectively reacts with LOOH, yielding a stable MxP adduct (perketal). The lipophilic perketal was eluted at a position away from that of intact LOOH and identified and isolated by LC. Upon treatment with acid, perketal released the original LOOH, which was finally purified by LC. Using our optimized purification procedures, for instance, we produced 75 mg of pure phosphatidylcholine hydroperoxide (.99%) from 100 mg of phosphatidylcholine. Our developed method expands the concept of the perketal method, which provides pure LOOH references. The LOOHs prepared by the perketal method would be used as "gold standards" in LOOH methodology. Because lipid peroxidation is involved in food deterioration (1) and pathophysiology of human diseases (2, 3), there has been a great interest in the accurate measurement of lipid hydroperoxide (LOOH) concentration. This can be performed by several quantitative methods (4-11), and the most sensitive and reliable one is chemiluminescence detection-liquid chromatography (CL-LC) (9-11). Since there is no approved LOOH calibrator ("gold standard") available, it is impossible to compare the LOOH levels from various laboratories around the world.Currently, researchers prepare their own in-house reference LOOH [i.e., lipids (e.g., phospholipids, cholesterols, triacylglycerols, and fatty acids)] are subjected to photo-(12), free radical (13), or enzymatic oxidation (14, 15). The resultant crude or partly purified LOOH is generally used as a calibrator. However, these references are neither officially approved nor do they correspond to each other, particularly with regard to their purity. As frequently mentioned by LOOH researchers, this problem is mainly caused by the difficulty in distinguishing and isolating LOOH from other oxidation products such as hydroxides (13). Therefore, efficient purification of a wide variety of LOOHs is the key to the development of the gold standard not only for the accurate quantification of LOOH but also for the evaluation of its biological fu...

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Preparation and purification of lipid hydroperoxides from arachidonic and γ‐linolenic acids

Cited by 224 publications

References 15 publications

Platelet and Blood Vessel Arachidonate Metabolism and Interactions

Platelet and Blood Vessel Arachidonate Metabolism and Interactions

Biliary glutathione promotes the mucosal metabolism of luminal peroxidized lipids by rat small intestine in vivo.

Preparation of pure lipid hydroperoxides

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