5'-Atp → 5-Hydroxyicosa-6,8,11,14-tetraenoic acid

Participants

Reaction Steps

76 μl of 50 mM Tris-HCl buffer (pH 7.4) containing 2.56 mM of CaCl2 and 2.56 mM of ATP and 20 μl of 5-lipoxygenase sample prepared above were added to 2μl of a solution containing a prescribed amount of the diarylheptanoide derivative (1) prepared in Example 1 in DSMO. The mixture was incubated at 37° C. After 15 minutes, the reaction was terminated with the addition of 200 μl of a 145:55:0.1 mixture of acetonitrile, methanol, and acetic acid. The reaction mixture was centrifuged (1300 rpm, 30 min., 40° C.). The amount of 5-HETE in the supernatant was measured by HPLC (column: Nova-pak 5C18; eluant: a 60:40 acetonitrile:0.05 M KH2PO4 solution with 5 mM trihexyl bromide added; flow rate: 0.8 ml/min.; wavelength: 235 nm). The amount of 5-HETE produced was calculated from the peak area of the 5-HETE. This procedure was repeated twice to determine the 5-lipoxygenase inhibiting activity of the diarylheptanoide derivative (1) from the average production amount of 5-HETE. The results are shown in FIG. 1, which indicates the 5-lipoxygenase inhibiting activity of the diarylheptanoide derivative (1).

Provenance & Evidence