Organic & Biomolecular Chemistry
Paper
of epiandrosterone and epiandrosterone 3-glucuronide with a
90% conversion as determined by 600 MHz 1H NMR inte-
gration of the H3 protons. A copy of the H NMR conversion
Pregnenolone
SPE
Testosterone
Trenazone
UGT
3β-Hydroxypregn-5-en-20-one
Solid-phase extraction
17β-Hydroxyandrost-4-en-3-one
17β-Hydroxyestra-4,9-dien-3-one
Uridine 5′-diphosphoglucuronosyl-
transferase
1
spectrum is reproduced in the ESI.†
Method B (synthesis). The reaction was carried out as for
method A on a larger scale with epiandrosterone (5.03 mg,
1.73 μmol). The reaction was then subjected to solid-phase
extraction. An Oasis WAX SPE cartridge (500 mg, 6 mL) was
pre-conditioned with methanol (5 mL) and milliQ water
(15 mL). The crude reaction was loaded onto the cartridge and
washed with aqueous formic acid (15 mL, 2% v/v), milliQ
water (15 mL), methanol (15 mL) to elute free steroid, and
finally with saturated aqueous ammonium hydroxide in
methanol (30 mL, 5% v/v) to elute the steroid glucuronide. The
appropriate fractions were combined and the solvent removed
under reduced pressure at 30 °C to afford epiandrosterone
3-glucuronide 13 as a white solid (6.8 mg, 84%). Copies of the
1H and 13C NMR spectra are reproduced in the ESI.† Rf 0.44
(7 : 2 : 1 EtOAc–MeOH–H2O); δH (600 MHz, CD3OD) 4.41 (1 H,
d, JH20–H21 8.0, H20), 3.78 (1 H, m, H3), 3.55 (1 H, d, JH24–H23
9.5, H24), 3.43 (1 H, t, JH23–H24 ≈ JH23–H22 9.2, H23), 3.39 (1 H,
t, JH22–H23 ≈ JH22–H21 8.9, H22), 3.18 (1 H, t, JH21–H22 ≈ JH21–H20
8.4, H21), 2.43 (1 H, dd, J 19.3, 8.7), 2.06 (1 H, dt, J 19.3, 9.4)
1.97–1.91 (2 H, m), 1.83 (1 H, m), 1.75–1.49 (7 H, m), 1.40–1.28
(5 H, m), 1.23 (1 H, m), 1.15 (1 H, m), 1.06–1.01 (2 H, m), 0.88
(3 H, s, CH3), 0.87 (3 H, s, CH3), 0.75 (1 H, m); δC (150 MHz,
CD3OD) 224.0 (C17), 176.7 (C25), 101.8 (C20), 78.5 (C3),
77.8 (C22), 75.4 (C24), 74.9 (C21), 73.7 (C23), 55.8, 52.6, 45.9,
38.1, 36.8, 36.6, 36.3, 35.1, 32.7, 32.0, 30.2, 29.7, 22.6, 21.5,
14.1 (CH3), 12.6 (CH3), one carbon overlapping or obscured;
LRMS (–ESI) m/z: 465 ([M − H]−); HRMS (–ESI) m/z: Calcd for
C25H37O8 ([M − H]−) 465.2488, found 465.2488.
Acknowledgements
We thank the Australian Research Council (DP110101235) for
financial support. We thank Ms Tracy Murray for assistance
with protein expression and purification and Mr Chris Blake
for assistance with NMR spectroscopy.
Notes and references
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Abbreviations
Androstanolone
Androsterone
Boldenone
17β-Hydroxy-5α-androstan-3-one
3α-Hydroxy-5α-androstan-17-one
17β-Hydroxyandrost-1,4-dien-3-one
5β-Cholestan-3β-ol
Coprostanol
Dehydroepiandrosterone 3β-Hydroxyandrost-5-en-17-one
Epiandrosterone
Epitestosterone
Estradiol
3β-Hydroxy-5α-androstan-17-one
17α-Hydroxyandrost-4-en-3-one
Estra-1,3,5(10)-triene-3,17β-diol
3-Hydroxyestra-1,3,5(10)-trien-
17-one
3α-Hydroxy-5β-androstan-17-one
2,2,3,4,4-d5-3α-Hydroxy-5β-andro-
stan-17-one
17β-Hydroxy-1α-methyl-5α-andro-
stan-3-one
17α-Methylandrost-5-ene-
3β,17β-diol
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Chem., 2011, 22, 752–758.
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and R. Kostiainen, Steroids, 2008, 73, 257–265.
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R. Kostiainen, Bioconjugate Chem., 2002, 13, 194–199.
15 T. Kuuranne, M. Kurkela, M. Thevis, W. Schänzer, M. Finel
and R. Kostiainen, Drug Metab. Dispos., 2003, 31, 1117–
1124.
Estrone
Etiocholanolone
d5-Etiocholanolone
Mesterolone
Methandriol
Methyltestosterone
Nandrolone
17β-Hydroxy-17α-methylandrost-
4-en-3-one
17β-Hydroxyestr-4-en-3-one
16 R. H. Tukey and C. P. Strassburg, Annu. Rev. Pharmacol.
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