LC–MS/MS-analysis of prostaglandin E2 and D2 in microdialysis samples of rats

An improved LC–MS/MS method for the quantification of prostaglandins E2 and D2 production in biological fluids

“…The product ion tandem mass spectra of PGE 2 Since PGD 2 and PGE 2 are geometrical isomers that show similar fragmentation patterns, complete chromatographic separation with excellent peak shape is essential for accurate quantitation. Our chromatographic method, based on the use of a phenyl-hexyl stationary phase, achieved complete and reproducible separation of 1.4 min for these isomers (see Figure 3) compared to only 0.25 min for the method of Schmidt et al [18]. Furthermore, the mobile phase was compatible with MS-MS detection using negative ion electrospray.…”

“…It should be noted that although Schmidt et al [18] reported that both PGE 2 and PGD 2 were stable at room temperature in a solution of methanol, water, and formic acid, they did not report measurement of the stability of these prostaglandins in biological fluids or under different experimental conditions that would have indicated the instability of PGD 2 . Our data in Table 3 show that whether incubated in water/methanol (1:1), 10% FBS or FBS-free medium, PGD 2 and d 4 -PGD 2 were much less stable than PGE 2 and d 4 -PGE 2 .…”

“…To overcome the limitations of GC-MS and immunoassays for prostaglandin measurements, LC-MS [15][16][17] and LC-MS-MS [18][19][20][21][22][23][24][25][26][27] based methods have evolved as powerful tools for measuring prostaglandins in biological samples because of their high sensitivity, high selectivity and simplicity of sample preparation. Although five of these LC-MS or LC-MS-MS methods have measured PGD 2 [15][16][17][18][19], none have controlled for the inherent chemical instability of PGD 2 which is critical for determining accurate levels and for comparison with the values of other, more stable eicosanoids.…”

“…Although five of these LC-MS or LC-MS-MS methods have measured PGD 2 [15][16][17][18][19], none have controlled for the inherent chemical instability of PGD 2 which is critical for determining accurate levels and for comparison with the values of other, more stable eicosanoids. Our method addresses this important issue by incorporation of both a d 4 -PGD 2 and d 4 -PGE 2 internal standards in the reaction mixture in order to accurately quantify the relative amounts of PGE 2 and PGD 2 in the same sample.…”

“…Our method addresses this important issue by incorporation of both a d 4 -PGD 2 and d 4 -PGE 2 internal standards in the reaction mixture in order to accurately quantify the relative amounts of PGE 2 and PGD 2 in the same sample. The paper by Schmidt et al [18] is the only previous publication that describes the detailed LC-MS-MS quantification of both PGD 2 and PGE 2 , but their method used d 4 -PGD 2 as the only internal standard for measuring both analytes. According to our results, this approach cannot result in accurate measurement of both PGD 2 and PGE 2 , because the degradation rates of PGD 2 and PGE 2 during sample preparation and analysis are significantly different.…”