Table 2

Structural identification of E₂ metabolites

E₂ Standards and Metabolites	Retention Times on HPLC		Retention Times on GC		Characteristic Mass Fragments ( m/z ) of the Trimethylsilylated E₂Metabolites^2-b	Quality of Match
E₂ Standards and Metabolites	Standard	Metabolite	Standard	Metabolite		Quality of Match
	min (RRT^2-a )		min			%
E₁	50.27 (1.038)	50.30 (1.039)	25.66	25.55	342, 257, 218, 73	99
2-OH-E₁	40.50 (0.833)	40.80 (0.842)	30.51	30.45	430, 73	98
4-OH-E₁	41.90 (0.862)	42.20 (0.871)	32.16	32.11	430, 73	98
6α-OH-E₁	22.00 (0.453)	N.D.	29.69	N.D.	430, 340, 73
6β-OH-E₁	23.87 (0.491)	24.63 (0.509)	28.06	28.18	430, 415, 340, 280, 73	97
6-keto-E₁	32.40 (0.664)	N.D.	33.50	N.D.	356, 341, 299, 73
7α-OH-E₁	20.80 (0.428)	N.D.	26.78	N.D.	430, 340, 312, 283, 73
7β-OH-E₁	19.80 (0.407)	N.D.	30.44	N.D.	430, 414, 340, 312, 73
11α-OH-E₁	22.80 (0.469)	N.D.	28.71	N.D.	430, 415, 340, 242, 218, 73
11β-OH-E₁	29.98 (0.614)	N.D.	28.18	N.D.	430, 415, 340, 313, 242, 73
11-keto-E₁	31.83 (0.651)	N.D.	26.69	N.D.	356, 340, 312, 218, 73
12β-OH-E₁	29.33 (0.603)	N.D.	30.14	N.D.	430, 415, 323, 218, 73
14-OH-E₁	21.33 (0.439)	N.D.	32.83	N.D.	340, 325, 312, 297, 284, 73
15α-OH-E₁	20.63 (0.424)	N.D.	31.93	N.D.	430, 415, 286, 217, 73
15β-OH-E₁	26.53 (0.546)	N.D.	30.88	N.D.	430, 286, 218, 73
16α-OH-E₁	25.37 (0.522)	24.63 (0.509)	31.81	31.49	430, 415, 286, 73	96
16β-OH-E₁	25.10 (0.516)	24.63 (0.509)	32.55	32.84	430, 415, 286, 73	99
16-keto-E₁	35.70 (0.731)	N.D.	32.17	N.D.	356, 341, 286, 232, 73
E₂	48.60 (1.00)	48.43 (1.00)	26.71	26.66	416, 401, 326, 298, 285, 73	99
2-OH-E₂	37.17 (0.765)	36.90 (0.762)	31.88	32.09	504, 73	99
4-OH-E₂	34.80 (0.716)	34.67 (0.716)	33.92	33.87	504, 73	99
6α-OH-E₂	11.23 (0.232)	10.50 (0.217)	31.39	31.46	504, 414, 324, 309, 283, 73	83
6β-OH-E₂	19.57 (0.403)	19.30 (0.398)	30.09	29.81	504, 414, 324, 283, 229, 73	99
6-keto-E₂	24.80 (0.510)	24.63 (0.509)	35.84	35.78	430, 415, 340, 299, 129, 73	96
7α-OH-E₂	10.47 (0.215)	10.50 (0.217)	26.81	26.89	504, 414, 324, 309, 283, 73	99
7β-OH-E₂	11.67 (0.241)	N.D.	31.66	N.D.	504, 414, 324, 309, 283, 73
11α-OH-E₂	20.70 (0.426)	N.D.	28.91	N.D.	504, 489, 414, 324, 217, 73
11β-OH-E₂	29.22 (0.597)	N.D.	27.63	N.D.	504, 414, 324, 297, 244, 73
12β-OH-E₂	30.30 (0.623)	31.17 (0.640)	35.40	35.45	504, 399, 324, 283, 73	96
12-keto-E₂	28.37 (0.584)	N.D.	32.25	N.D.	430, 415, 340, 323, 73
14-OH-E₂	12.57 (0.259)	N.D.	32.53	N.D.	414, 324, 309, 283, 73
15α-OH-E₂	9.47 (0.195)	9.33 (0.193)	31.85	31.96	504, 324, 298, 245, 217, 73	96
15β-OH-E₂	17.57 (0.361)	17.17 (0.355)	30.72	30.81	504, 325, 298, 283, 217, 73	99
16α-OH-E₂ (E₃)	15.83 (0.321)	15.67 (0.323)	35.20	35.09	504, 386, 345, 324, 311, 73	99
16β-OH-E₂	27.87 (0.575)	27.80 (0.574)	36.74	36.52	504, 489, 386, 345, 311, 73	99
16-keto-E₂	23.78 (0.487)	24.63 (0.509)	33.38	33.22	430, 415, 312, 285, 258, 73	89
16α-OH-17α-E₂	30.55 (0.625)	N.D.	34.20	N.D.	504, 489, 414, 386, 345, 73
16β-OH-17α-E₂	15.87 (0.325)	N.D.	32.60	N.D.	504, 489, 386, 345, 311, 73
2-OH-E₃	7.15 (0.146)	N.D.	41.70	N.D.	592, 147, 129, 73
6-keto-E₃	7.40 (0.152)	N.D.	49.44	N.D.	518, 325, 245, 73
15α-OH-E₃	7.27 (0.149)	N.D.	46.94	N.D.	592, 502, 191, 73
2-MeO-E₂	50.80 (1.045)	N.D.	30.80	N.D.	446, 431, 416, 315, 73

Human liver microsomes were incubated with E₂ under conditions as described under Materials and Methods. Each peak was collected from the HPLC, derivatized with BSTFA containing 1% TMCS, and then analyzed by GC/MS as described under Materials and Methods.

N.D., Not detected.
↵2-a Relative retention time in relation to E₂.
↵2-b The major fragment (base peak) for each metabolite is underlined.