Table 5. Study in Vitro Target Occupancy (k_off) of Selected sEH Inhibitors against Human sEH.

entry	inhibitor	koff (× 10–4 s–1)a	t1/2 (min)b	entry	inhibitor	koff (× 10–4 s–1)a	t1/2 (min)b
1	APAU	19.2 ± 0.70	6.03 ± 0.2	11	17	3.51 ± 0.20	33.0 ± 1.9
2	18 (TPPU)	10.5 ± 0.20	11.0 ± 0.2	12	19	6.14 ± 0.18	18.8 ± 0.6
3	4	6.57 ± 0.30	17.6 ± 0.8	13	21	5.05 ± 0.02	22.9 ± 0.1
4	5	7.91 ± 0.31	14.6 ± 0.6	14	24	4.39 ± 0.43	26.5 ± 2.6
5	6	5.76 ± 0.26	20.1 ± 0.9	15	25	23.1 ± 1.1	5.02 ± 0.3
6	7	5.19 ± 0.09	22.3 ± 0.4	16	31	10.3 ± 0.1	11.0 ± 0.1
7	9	4.75 ± 0.11	24.3 ± 0.6	17	32	8.90 ± 0.35	13.0 ± 0.5
8	11	5.79 ± 0.43	20.0 ± 1.5	18	33 (TPAU)	26.4 ± 2.30	4.40 ± 0.4
9	13	3.13 ± 0.06	37.0 ± 0.7	19	TUPS	20.0 ± 0.64	5.79 ± 0.2
10	14	5.39 ± 0.39	21.5 ± 1.6	20	t-TUCB	7.19 ± 0.36	16.1 ± 0.8

^ak_off was determined by FRET-based displacement assay described by Lee et al.³⁸ and described in detail in Supporting Information. Brieftly, a preincubated human sEH–inhibitor complex (8 μM) was diluted by ×40 times by fluorescent reporter–APCU (2 μM, 0.1 M sodium phosphate, pH 7.4). The fluorescent enhancement (λ_excit = 280 nm, λ_emit = 450) was measured over time (5100 s). The results are the average of triplicates with ± SD

^bt_1/2 = ln(2)/k_off, which describes the half-life of enzyme–inhibitor complex.