Development and validation of a high-performance liquid chromatography–electrospray mass spectrometry method for the simultaneous determination of 23 eicosanoids

Blewett, Anthony J.; Varma, Deepti; Gilles, Tiffany; Libonati, Joseph R.; Jansen, Susan A.

doi:10.1016/j.jpba.2007.11.047

Cited by 50 publications

(33 citation statements)

References 48 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Direct measurement of PGE 2 was also done on frozen mammary tissue using liquid chromatography tandem mass spectrometry (MS). Approximately 80 mg of frozen tumor tissue (n = 4 per group) was processed using a homogenization process similar to that described by Blewett et al (33). Each tissue sample was placed on ice and 400 μL of icecold methanol (containing 2.5% formic acid), and 5 ng of the d-4 form of PGE 2 (Cayman Chemical) as the internal standard were added.…”

Section: Pge 2 Level Determinationmentioning

confidence: 99%

Cyclooxygenase-2 Inhibition for the Prophylaxis and Treatment of Preinvasive Breast Cancer in a Her-2/Neu Mouse Model

Tran‐Thanh

Buttars

Wen

et al. 2010

Cancer Prevention Research

View full text Add to dashboard Cite

Ductal carcinoma in situ (DCIS) is the most common form of preinvasive breast cancer. Several molecular alterations have been identified in DCIS. Among them, cyclooxygenase 2 (COX-2) overexpression has been shown in 60% to 80% of DCIS cases. Celecoxib is a nonsteroidal anti-inflammatory drug that selectively inhibits COX-2. In this study, we evaluated whether COX-2 inhibition by celecoxib can reduce the incidence of preinvasive breast cancer and its progression to invasive breast cancer in a mouse model exhibiting a similar phenotype to human solid-pattern DCIS. We have used the mouse model mouse mammary tumor virus (MMTV)-Neu to investigate this possibility. These mice carry a rat Her-2/Neu transgene and are known to develop DCIS-like lesions. Our results showed that celecoxib (500 ppm) given as prophylaxis was neither able to prevent tumor development nor delay tumor appearance compared with untreated mice. Furthermore, when the drug was given early in tumorigenesis, it did not reduce the progression of preinvasive to invasive tumors nor prevent lung metastasis. Reduction of prostaglandin levels was, however, achieved in mammary tumors of treated mice. In addition, celecoxib treatment caused an increase in apoptosis and decreased vascular endothelial growth factor expression in treated animals. Our results contrast with some previously published studies and highlight the complexity of the relationship between COX-2 and breast cancer. Cancer Prev Res; 3(2); 202-11. ©2010 AACR.

show abstract

Section: Pge 2 Level Determinationmentioning

confidence: 99%

Cyclooxygenase-2 Inhibition for the Prophylaxis and Treatment of Preinvasive Breast Cancer in a Her-2/Neu Mouse Model

Tran‐Thanh

Buttars

Wen

et al. 2010

Cancer Prevention Research

View full text Add to dashboard Cite

show abstract

“…Furthermore, almost all of these studies were limited to human P450 enzymes (Capdevila et al, 2000;Imig, 2012). Also, previously published studies on P450-mediated AA metabolism focused mainly on EETs and 20-HETE; as a result, mHETEs and v-1→4 HETEs have been largely ignored, mainly as the result of the lack of sophisticated methods for the simultaneous measurement of mHETEs and v-1→4 HETEs along with EETs and 20-HETE (Blewett et al, 2008). The fact that the alterations of P450-AA metabolites levels are almost exclusively studied in animals, especially rats, mandates the study of animal P450 enzymes as well, keeping in mind that the determination and consequent pharmacological modulation of the involved enzymes need to be tested in animals before proceeding to clinical trials.…”

Section: Introductionmentioning

confidence: 99%

Characterization of Arachidonic Acid Metabolism by Rat Cytochrome P450 Enzymes: The Involvement of CYP1As

El-Sherbeni

2014

Drug Metab Dispos

View full text Add to dashboard Cite

Cytochrome P450 (P450) enzymes mediate arachidonic acid (AA) oxidation to several biologically active metabolites. Our aims in this study were to characterize AA metabolism by different recombinant rat P450 enzymes and to identify new targets for modulating P450-AA metabolism in vivo. A liquid chromatography-mass spectrometry method was developed and validated for the simultaneous measurements of AA and 15 of its P450 metabolites. CYP1A1, CYP1A2, CYP2B1, CYP2C6, and CYP2C11 were found to metabolize AA with high catalytic activity, and CYP2A1, CYP2C13, CYP2D1, CYP2E1, and CYP3A1 had lower activity. CYP1A1 and CYP1A2 produced v-1→4 hydroxyeicosatetraenoic acids (HETEs) as 88.7 and 62.7%, respectively, of the total metabolites formed. CYP2C11 produced epoxyeicosatrienoic acids (EETs) as 61.3%, and CYP2C6 produced midchain HETEs and EETs as 48.3 and 29.4%, respectively, of the total metabolites formed. The formation of CYP1A1, CYP1A2, CYP2C6, and CYP2C11 major metabolites followed an atypical kinetic profile of substrate inhibition. CYP1As inhibition by a-naphthoflavone or anti-CYP1As antibodies significantly reduced v-1→4 HETE formation in the lungs and liver, whereas CYP1As induction by 3-methylcholanthrene resulted in a significant increase in v-1→4 HETEs formation in the heart, lungs, kidney, and livers by 370, 646, 532, and 848%, respectively. In conclusion, our results suggest that CYP1As and CYP2Cs are major players in the metabolism of AA. The significant contribution of CYP1As to AA metabolism and their strong inducibility suggest their possible use as targets for the prevention and treatment of several diseases.

show abstract

“…We speculate that these peaks corresponded to HETEs, [19][20][21][22][23] which are also metabolites of AA, and/or impurities contained in standard solution of AA. EETs and DHETs were detected at much higher levels than the respective LLOQs in the samples incubated for 30 min with rCYP2C9 (Fig.…”

Section: Discussionmentioning

confidence: 95%

“…Blewett et al used a similar gradient mobile phase system (a mixture of acetonitrile and water containing 0.1% formic acid) for the assaying of 23 eicosanoids. 19) Their methodology was time-consuming, with DHETs, EETs, and AA eluting at approximately 29, 45, and 52 min, respectively. Our method focused on the CYP pathway of AA metabolism to investigate drug-endogenous interactions, and we sought to determine EETs and DHETs in a shorter time.…”

Section: Discussionmentioning

confidence: 99%

“…Several assay methods for eicosanoids employing LC/ MS have been reported. [19][20][21] A method using a tandem MS (LC-MS/MS) system [22][23][24][25][26] was recently developed to enhance sensitivity. However, the method described by Bylund et al 22) employed an ion trap MS system which generally shows a smaller dynamic range than a triple-quadrupole MS/MS system, and other methods use one or no internal standard for determination of eicosanoid concentrations.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Simultaneous Determination Method of Epoxyeicosatrienoic Acids and Dihydroxyeicosatrienoic Acids by LC-MS/MS System

Mukai

Toda

Takeuchi

et al. 2015

Biological & Pharmaceutical Bulletin

View full text Add to dashboard Cite

Epoxyeicosatrienoic acids (EETs) are produced primarily by CYPs from arachidonic acid (AA) and then further metabolized to the corresponding dihydroxyeicosatrienoic acids (DHETs). EETs play important roles in physiological processes such as regulating vasodilation and inflammation. Thus, the drug inhibition of CYP-mediated AA metabolism could reduce production of EETs, potentially resulting in adverse cardiovascular events. The aim of this study was to develop a simple method to simultaneously determine the concentrations of both EETs and DHETs using a conventional LC-MS/MS system to evaluate drug-endogenous substance interactions, including eicosanoids. Eight eicosanoids (5,6-EET, 8,9-EET, 11,12-EET, 14,15-EET, 5,6-DHET, 8,9-DHET, 11,12-DHET, and 14,15-DHET) were detected with their corresponding deuteriumlabeled eicosanoids as internal standards. The samples were purified by solid-phase extraction columns. Liquid chromatographic separation was achieved on a C18 column. DHETs and EETs were eluted at 4-7 and 18-26 min, respectively. The weighted (1/y 2 ) calibration curves were linear over a range of 5-2000 nmol/L for EETs and 2-2000 nmol/L for DHETs. In quality control (QC) samples, the recoveries of eicosanoids were 95.2-118%. The intra-day precisions were within 6% in all three QC samples, and the inter-day precisions were <16.7% at 50 nmol/L, <8.6% at 200 nmol/L, and <9.8% at 1000 nmol/L. We have applied this method for the determination of the eicosanoid levels in samples from incubation studies of AA by using human recombinant CYP enzyme (rCYP), and confirmed that the method has sensitivity sufficient for assessment of rCYP incubation study.Key words epoxyeicosatrienoic acid; dihydroxyeicosatrienoic acid; arachidonic acid; LC-MS/MS Arachidonic acid (AA) is metabolized to various eicosanoids via several pathways.1) These eicosanoids play important roles in physiological processes, including the regulation of inflammation.2-5) Three major metabolic pathways for conversion of AA to eicosanoids are recognized, including routes that incorporate cyclooxygenase (COX), lipoxygenase (LOX), and CYP.6) The CYP pathway, which is mediated primarily by CYP2C9, CYP2C8, and CYP2J2, produces epoxyeicosatrienoic acids (EETs) and hydroxyeicosatetraenoic acids (HETEs). 7,8) EETs exist as one of 4 regioisomers (5,6-EET, 8,9-EET, 11,12-EET, and 14,15-EET), depending on the location of the epoxidized residue. Each EET is substantially metabolized to its corresponding diol, a dihydroxyeicosatrienoic acid (DHET), by soluble epoxide hydrolase (sEH).1,9-12) The EETs have been shown to play important roles on cardiovascular homeostasis. [13][14][15] Thus, the drug inhibition of CYP-mediated AA metabolism via could reduce production of EETs, potentially resulting adverse cardiovascular events.Traditional analytical methods such as HPLC or capillary electrophoresis with UV or fluorescence detection, 16,17) and GC-MS 18) lack sufficient sensitivity to detect the low levels of AA metabolites typically found in physiological matrices. Sev...

show abstract

Development and validation of a high-performance liquid chromatography–electrospray mass spectrometry method for the simultaneous determination of 23 eicosanoids

Cited by 50 publications

References 48 publications

Cyclooxygenase-2 Inhibition for the Prophylaxis and Treatment of Preinvasive Breast Cancer in a Her-2/Neu Mouse Model

Cyclooxygenase-2 Inhibition for the Prophylaxis and Treatment of Preinvasive Breast Cancer in a Her-2/Neu Mouse Model

Characterization of Arachidonic Acid Metabolism by Rat Cytochrome P450 Enzymes: The Involvement of CYP1As

Simultaneous Determination Method of Epoxyeicosatrienoic Acids and Dihydroxyeicosatrienoic Acids by LC-MS/MS System

Contact Info

Product

Resources

About