Summary of evidence that the peak with a retention time of 20.75 min in the HPLC separation of products derived from incubation of 5,8,11,14,17-eicosapentaenoic acid with huPGHS-2 is 14-HEPE
| Data | Result | Conclusion |
|---|---|---|
| UV, MS, and MS/MS data for peak at 20.75 min | ||
| [M-H]− = m/z 317 | Acidic functional group | Mr = 318 |
| Fragment ion at m/z 299 | Loss of H2O | Monohydroxy |
| Fragment ion at m/z 273 | Loss of CO2 | Carboxylic acid |
| UV absorbance at 230 nm | UV peak at 20.75 min | Conjugated diene |
| Intense fragment ion at m/z 207 | Probable α-hydroxy-β-ene rearrangement | 14-Hydroxyl group |
| MS/MS data after reduction of peak at 20.75 min with Adam's catalyst | ||
| [M-H]− = m/z 327 | Increase of 10 mass units | Five double bonds |
| Retention time increase of >10 min | More hydrophobic | Saturated chain |
| Fragment ion at m/z 309 | Loss of H2O | Monohydroxy acid |
| Fragment ion at m/z 283 | Loss of CO2 | Carboxylic acid |
| Fragment ion at m/z 325 | Loss of H2 | Formation of double bond |
| Fragment ion at m/z 211 | Probable α-hydroxy-β-ene rearrangement | 14-Hydroxy; C11–C12 double bond |