Synthesis of ursodeoxycholic acid from plant-source (20S)-21-hydroxy-20-methylpregn-4-en-3-one

Article abstract of DOI:10.1016/j.steroids.2020.108600

A novel synthetic route of producing ursodeoxycholic acid (UDCA) was developed through multiple reactions from cheap and commercially available bisnoralcohol (BA). The key reaction conditions, including solvents, bases and reaction temperatures of the route were investigated and optimized. In the straightforward route for preparation of UDCA, most of the reaction steps have high conversions with average yields of 91%, and overall yield up to 59% (6 steps) from the plant-source BA. Especially in the last step of reduction and hydrolysis, there are five functional groups converted with calcd 97% per conversion in one-pot reaction. This promising route offers economical and efficient strategies for potential large-scale production of UDCA.

Full text of DOI:10.1016/j.steroids.2020.108600

Steroids157(2020)108600
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Steroids
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Synthesis of ursodeoxycholic acid from plant-source (20S)-21-hydroxy-20-
methylpregn-4-en-3-one
⁎
Jie Wanga, Xiang-Zhong Gub, Li-Ming Hea, Chen-Chen Lia, Wen-Wei Qiua,
a Shanghai Engineering Research Center of Molecular Therapeutics and New Drug Development, School of Chemistry and Chemical Engineering, East China Normal
University, Shanghai 200241, China
b Department of Research and Development, Jiangsu Jiaerke Pharmaceuticals Group Co., Ltd., Zhenglu Town, Changzhou 213111, China
A R T I C L E I N F O
A B S T R A C T
Keywords:
A novel synthetic route of producing ursodeoxycholic acid (UDCA) was developed through multiple reactions
from cheap and commercially available bisnoralcohol (BA). The key reaction conditions, including solvents,
bases and reaction temperatures of the route were investigated and optimized. In the straightforward route for
preparation of UDCA, most of the reaction steps have high conversions with average yields of 91%, and overall
yield up to 59% (6 steps) from the plant-source BA. Especially in the last step of reduction and hydrolysis, there
are five functional groups converted with calcd 97% per conversion in one-pot reaction. This promising route
offers economical and efficient strategies for potential large-scale production of UDCA.
Ursodeoxycholic acid
Bisnoralcohol
Synthesis
1. Introduction
(Fig. 1) by regioselective oxidation of 7α-OH to 7-oxo and then reduction
to 7β-OH [14–16]. Hyodeoxycholic acid (HDCA) is a relatively less ex-
Ursodeoxycholic acid (UDCA) (Fig. 1) is the major bile acid of the bile
of black bears which has been used in Chinese traditional medicine as a
remedy for liver diseases [1]. It is an active pharmaceutical ingredient
(API) widely used in clinics and approved by the US Food and Drug Ad-
ministration (FDA) for the treatment of primary biliary cholangitis (PBC)
since the year 1988 [2,3]. PBC, previously termed primary biliary cir-
rhosis, is a rare autoimmune inflammatory liver disease, which produces
bile duct injury, fibrosis, cholestasis and eventual cirrhosis [4]. UDCA also
has a variety of biological activities, such as non-surgical treatment of
cholesterol gallstones [5], anti-apoptotic effects [6], anti-inflammatory
actions [7], improvement of insulin sensitivity [8], antitumor activity [9]
and treatment of Alzheimer's disease [10]. Accordingly, exploitation of
economical and efficient synthetic methods of UDCA will not only benefit
PBC patients, but also promote its further development and applications.
Cholic acid (CA) can be used as a commercial starting material for
preparation of UDCA. For examples, Ferrari et al. provided a low overall
yield method by a crucial high-temperatured Wolff-Kishner reduction
[11]; Dangate et al. exploited an optimized protection-free route with o-
iodoxybenzoic acid as key regioselective oxidant [12]; previously, we
also developed a high-yield route (7 steps) by a key elimination reaction
of C12-mesylate ester group of CA, in which most of the reaction steps
have high conversions with average yields of 94% [13]. Currently, UDCA
is usually obtained by epimerization of chenodeoxycholic acid (CDCA)
pensive commercially available bile acid compared with CDCA. Zhou
et al. prepared of UDCA from the HDCA (Fig. 1) with a 15% poor total
yield [17]. More recently, Dou et al. developed a route based on the
HDCA with an improved 26% overall yield via a crucial Shapiro reaction
[18]. However, no matter the starting materials of CA, CDCA and HDCA
are extracted from the bile acid and belonging to starting materials of
animal origin. To our knowledge, there is no synthetic route to UDCA
based on a plant-source commercial starting material. The (20S)-21-hy-
droxy-20-methylpregn-4-en-3-one, also called bisnoralcohol (BA)
(Fig. 1), which is obtained from side-chain degradation of the phytos-
terols by an industrial strain mycobacterium neoaurum [19,20]. Pre-
sently, the BA is abundant and commercially available, which price is
much cheaper than CDCA. Thus, it is rather valuable to develop an im-
proved synthetic route for large scale production of UDCA using the
readily-available starting material BA.
Herein, we report a high-efficiency and economical synthetic route
of UDCA from cost-effective and plant-source BA.
2. Experimental
2.1. General procedures
All reagents and chemicals were purchased from commercial
⁎ Corresponding author.
E-mail address: wwqiu@chem.ecnu.edu.cn (W.-W. Qiu).
https://doi.org/10.1016/j.steroids.2020.108600
Received 19 December 2019; Received in revised form 21 January 2020; Accepted 27 January 2020
Availableonline14February2020
0039-128X/©2020ElsevierInc.Allrightsreserved.

J. Wang, et al.
Steroids157(2020)108600
Fig. 1. Structures of bile acids and bisnoralcohol.
suppliers and used without further purification unless otherwise stated.
When needed, the reactions were carried out in oven-dried glassware
under a positive pressure of dry N2. Column chromatography was
performed on silica gel (QinDao, 200–300 mesh) using the indicated
eluents. Thin-layer chromatography was carried out on silica gel plates
(QinDao) with a layer thickness of 0.25 mm. Melting points were de-
termined using the MEL-TEMP 3.0 apparatus and uncorrected. 1H
(400 MHz or 500 MHz) and 13C (100 MHz or 125 MHz) NMR spectra
were recorded on Bruker 400 MHz or 500 MHz spectrometer with
CDCl3 or DMSO‑d6 as solvent and tetramethylsilane (TMS) as the in-
ternal standard. All chemical shift values were reported in units of δ
(ppm). The following abbreviations were used to indicate the peak
multiplicity: s = singlet; d = doublet; t = triplet; m = multiplet;
TBAB (870 mg, 2.70 mmol) in H2O (40 mL) was added. The reaction
mixture was stirred at 0 °C for an additional 20 min, and then NCS
(4.1 g, 31.00 mmol) was added. After 5 h, the reaction mixture was
allowed to warm to room temperature, treated with sodium thiosulfate
pentahydrate (1.3 g in 25 mL H2O). The reaction mixture was stirred at
0 °C for 20 min. The organic phase was separated, and the aqueous
phase was extracted with CH2Cl2 (50 mL × 2). The organic layer was
washed with sodium hydroxide solution, brine, dried with anhydrous
Na2SO4 and concentrated to give 3 (9.6 g, 95%) as a light yellow solid;
mp: 168–171 °C. 1H NMR (500 MHz, CDCl3) δ 9.56 (d, J = 3.3 Hz, 1H),
5.36–5.31 (m, 1H), 3.97–3.90 (m, 4H), 2.58–2.53 (m, 1H), 2.39–2.31
(m, 1H), 2.11 (dd, J = 14.2, 2.9 Hz, 1H), 2.00–1.93 (m, 2H), 1.91–1.82
(m, 1H), 1.81–1.73 (m, 2H), 1.68–1.62 (m, 3H), 1.59–1.53 (m, 1H),
1.52–1.44 (m, 3H), 1.40–1.29 (m, 2H), 1.28–1.15 (m, 2H), 1.12 (d,
J = 6.8 Hz, 3H), 1.11–1.03 (m, 2H), 1.02 (s, 3H), 0.72 (s, 3H). 13C NMR
(125 MHz, CDCl3) δ 205.20, 140.26, 122.10, 109.54, 64.56, 64.35,
56.11, 51.08, 49.79, 49.61, 43.09, 41.90, 39.58, 36.74, 36.45, 32.01,
31.80, 31.19, 27.16, 24.78, 21.10, 19.00, 13.59, 12.37. HRMS (ESI):
calcd for C24H36NaO3 [M + Na]+, 395.2557, found 395.2542.
br
= broad. High-resolution mass data were obtained on a
BrukermicroOTOF-Q II spectrometer.
2.2. Chemical synthesis
2.2.1. 3-dioxolane-23,24-bisnorchola-5-en-22-ol (2)
To a solution of bisnoralcohol (10.0 g, 30.26 mmol), ethylene glycol
(16.8 mL, 302.60 mmol) and pTSA (57 mg, 0.30 mmol) in benzene
(300 mL) was refluxed with Dean-Stark column for 24 h. The reaction
mixture was treated with saturated NaHCO3 solution (20 mL) and ex-
tracted with AcOEt (60 mL × 3). The organic layer was washed with
brine, dried over anhydrous Na2SO4 and then concentrated. The residue
was purified by silica gel chromatography (petroleum ether/AcOEt, 3/
1, v/v) to give 2 (10.0 g, 88%) as a white solid; mp: 175–177 °C [lit.
[21] 173–174 °C]. 1H NMR (500 MHz, CDCl3) δ 5.36–5.32 (m, 1H),
3.97–3.90 (m, 4H), 3.63 (dd, J = 10.5, 3.2 Hz, 1H), 3.35 (dd, J = 10.5,
6.9 Hz, 1H), 2.58–2.53 (m, 1H), 2.11 (dd, J = 14.2, 2.9 Hz,1H),
2.03–1.91 (m, 2H), 1.85–1.72 (m, 3H), 1.69–1.58 (m, 3H), 1.57–1.49
(m, 2H), 1.49–1.39 (m, 2H), 1.36–1.27 (m, 3H), 1.22–1.15 (m, 2H),
1.12–1.07 (m, 1H), 1.04 (d, J = 6.7 Hz, 3H), 1.02 (s, 3H), 1.00–0.97
(m, 1H), 0.70 (s, 3H). 13C NMR (125 MHz, CDCl3) δ 140.26, 122.25,
109.60, 68.12, 64.55, 64.34, 56.59, 52.52, 49.79, 42.55, 41.91, 39.74,
38.90, 36.73, 36.45, 32.04, 31.84, 31.19, 27.85, 24.51, 21.16, 19.00,
2.2.3. (20S)-3-oxopregn-4-ene-20-carboxaldehyde (4)
By a similar procedure described for 3, 4 was obtained as a light
yellow solid (yield 95%), mp: 155–157 °C. 1H NMR (400 MHz, CDCl3) δ
9.55 (s, 1H), 5.71 (s, 1H), 2.45–2.23 (m, 5H), 1.99 (t, J = 13.7 Hz, 2H),
1.91–1.78 (m, 2H), 1.68 (t, J = 10.2 Hz, 2H), 1.43 (m, 5H), 1.30–1.19
(m, 2H), 1.17 (s, 3H), 1.11 (d, J = 5.5 Hz, 3H), 1.06–0.89 (m, 3H), 0.75
(s, 3H). 13C NMR (100 MHz, CDCl3) δ 205.00, 199.65, 171.31, 123.99,
55.25, 53.84, 51.04, 49.54, 43.10, 39.39, 38.68, 35.80, 35.68, 34.06,
32.93, 32.05, 27.11, 24.64, 21.06, 17.48, 13.53, 12.44. HRMS(ESI):
calcd for C22H32NaO2 [M + Na]+, 351.2295, found 351.2292.
2.2.4. (22E)-3-oxo-4,22-choladien-24-oic acid ethyl ester (5) [22]
To a solution of 4 (9.5 g, 28.92 mmol) and Ph3P = CHCOOC2H5
(20.2 g, 57.84 mmol) in toluene (150 mL) was refluxed for 4 h. After
cooling, the reaction mixture was concentrated. The residue was pur-
ified by silica gel chromatography (petroleum ether/AcOEt, 3/1, v/v)
to give 5 (11.3 g, 98%) as a white solid; mp: 160–162 °C. [lit. [23]
155–157 °C]. 1H NMR (400 MHz, CDCl3) δ 6.81 (dd, J = 15.3, 9.0 Hz,
1H), 5.71 (d, J = 13.4 Hz, 2H), 4.24–4.09 (m, 2H), 2.45–2.21 (m, 5H),
2.00 (d, J = 12.6 Hz, 2H), 1.80 (m, 1H), 1.76–1.33 (m, 7H), 1.26 (m,
6H), 1.17 (s, 3H), 1.08 (d, J = 6.2 Hz, 3H), 1.05–0.86 (m, 3H), 0.73 (s,
3H).13C NMR (100 MHz, CDCl3) δ 199.70, 171.51, 167.16, 154.56,
123.94, 119.21, 60.27, 55.78, 54.98, 53.84, 42.82, 39.80, 39.54, 38.69,
16.89, 12.07. HRMS (ESI): calcd for
397.2713, found 397.2704.
C
24H38NaO3 [M
+
Na]+
,
2.2.2. 3-dioxolane-23,24-bisnorchola-5-en-22-al (3)
To
a solution of 2 (10.1 g, 26.96 mmol), TEMPO (42 mg,
0.27 mmol) in CH2Cl2 (100 mL) was stirred for 20 min under nitrogen
atmosphere at 0 °C. Then a solution of NaHCO3 (3.1 g, 36.40 mmol),
2

J. Wang, et al.
Steroids157(2020)108600
35.80, 35.70, 34.08, 32.98, 32.06, 28.19, 24.28, 21.10, 19.31, 17.49,
14.40, 12.32. HRMS(ESI): calcd for C26H38NaO3 [M + Na]+, 421.2713,
found 421.2708.
filtrate was evaporated under reduced pressure. The residue was pur-
ified by silica gel chromatography (petroleum ether/AcOEt, 3/1, v/v)
to give 7 (4.4 g, 85%) as a white solid; mp: 139–141 °C. 1H NMR
(500 MHz, CDCl3) δ 6.82 (dd, J = 15.6, 9.0 Hz, 1H, H-22), 5.72 (d,
J = 15.6 Hz, 1H, H-23), 5.65 (d, J = 1.7 Hz, 1H, H-6), 4.16 (q,
J = 7.1 Hz, 2H, –OCH2CH3), 3.98–3.90 (m, 4H, –OCH2CH2O–), 2.66
(dd, J = 14.7, 1.8 Hz, 1H), 2.44–2.36 (m, 1H), 2.34–2.18 (m, 3H),
2.02–1.95 (m, 1H), 1.89–1.83 (m, 2H), 1.78–1.71 (m, 2H), 1.64–1.52
(m, 3H), 1.52–1.43 (m, 1H), 1.27 (m, 8H), 1.19 (s, 3H, H-19), 1.08 (d,
J = 6.6 Hz, 3H, H-21), 0.70 (s, 3H, H-18). 13C NMR (125 MHz, CDCl3) δ
201.52 (C-7), 167.11 (C-24), 164.66 (C-5), 154.53 (C-22), 126.73 (C-6),
119.23 (C-23), 108.98 (C-3), 64.70 (–OCH2CH2O–, 64.62
(–OCH2CH2O–), 60.24 (–OCH2CH3), 53.82, 50.01, 49.65, 45.41, 43.58,
41.84, 39.59, 38.65, 38.35, 35.73, 31.16, 28.34, 26.44, 21.25, 19.58 (C-
21), 17.08 (C-19), 14.40 (–OCH2CH3), 12.40 (C-18). HRMS(ESI): calcd
for C28H40NaO5 [M + Na]+, 479.2768, found 479.2770.
2.2.5. (E)-3,3-ethylenedioxy-5,22-choladienoic acid ethyl ester (6) [22]
To a solution of 3 (9.6 g, 25.77 mmol) and Ph3P = CHCOOC2H5
(18.0 g, 51.54 mmol) in toluene (150 mL) was refluxed for 4 h. After
cooling, the reaction mixture was concentrated. The residue was pur-
ified by silica gel chromatography (petroleum ether/AcOEt, 3/1, v/v)
to give 6 (11.2 g, 98%) as a white solid; mp: 122–124 °C.
To a solution of 5 (5.0 g, 12.54 mmol), ethylene glycol (7.0 mL,
125.40 mmol) and pTSA (25 mg, 0.13 mmol) in toluene (150 mL) was
refluxed with Dean-Stark column for 24 h. The reaction mixture was
treated with saturated NaHCO3 solution (20 mL) and extracted with
AcOEt (50 mL × 3). The organic layer was washed with brine, dried
over anhydrous Na2SO4 and then concentrated. The residue was pur-
ified by silica gel chromatography (petroleum ether/AcOEt, 3/1, v/v)
to give 6 (4.9 g, 88%) as a white solid; mp: 122–124 °C. 1H NMR
(500 MHz, CDCl3) δ 6.82 (dd, J = 15.6, 8.9 Hz, 1H, H-22), 5.72 (d,
J = 15.6 Hz, 1H, H-23), 5.39–5.28 (m, 1H, H-6), 4.16 (q, J = 7.1 Hz,
2H, –OCH2CH3), 3.97–3.90 (m, 4H, –OCH2CH2O–), 2.58–2.53 (m, 1H),
2.26 (d, J = 6.7 Hz, 1H), 2.11 (dd, J = 14.2, 2.9 Hz, 1H), 2.00–1.92
(m, 2H), 1.81–1.73 (m, 2H), 1.72–1.61 (m, 3H), 1.60–1.52 (m, 2H),
1.51–1.41 (m, 2H), 1.37–1.30 (m, 1H), 1.27 (t, J = 7.1 Hz, 3H,
–OCH2CH3), 1.25–1.18 (m, 3H), 1.08 (d, J = 6.7 Hz, 3H, H-21),
1.04–1.06 (m, 2H), 1.02 (s, 3H, H-19), 1.00–0.97 (m, 1H), 0.71 (s, 3H,
H-18). 13C NMR (125 MHz, CDCl3) δ 167.20 (C-24), 154.84 (C-22),
140.24 (C-5), 122.20 (C-6), 119.07 (C-23), 109.57 (C-3), 64.56
(OCH2CH2O), 64.35 (OCH2CH2O), 60.22 (OCH2CH3), 56.63 (C-14),
55.01 (C-17), 49.78 (C-9), 42.79 (C-13), 41.91 (C-4), 39.85 (C-20),
39.72 (C-12), 36.75 (C-10), 36.45 (C-1), 32.02 (C-8), 31.80 (C-7), 31.20
(C-2), 28.25 (C-16), 24.42 (C-15), 21.14 (C-11), 19.38 (C-21), 19.00 (C-
19), 14.42 (OCH2CH3), 12.24 (C-18). HRMS(ESI): calcd for C28H42NaO4
[M + Na]+, 465.2975, found 465.2990.
2.2.7. (E)-7-hydroxy-3-oxo-4,6,22-choladienoic acid ethyl ester (8)
To a solution of 7 (4.4 g, 9.64 mmol) in a mixed solvent (50 mL) of
THF-H2O (9:1, v/v) was added H2SO4 (2 mL) at 0 °C. Then the reaction
mixture was stirred for 4 h at room temperature. The reaction mixture
was treated with saturated NaHCO3 solution (80 mL) and extracted
with AcOEt (30 mL × 3). The organic layer was washed with brine,
dried over anhydrous Na2SO4 and then concentrated. The residue was
purified by silica gel chromatography (petroleum ether/AcOEt, 3/1, v/
v) to give 8 (3.9 g, 98%) as a white solid; mp: 167–169 °C. 1H NMR
(400 MHz, DMSO‑d6) δ 10.22 (s, 1H), 6.74 (dd, J = 15.4, 9.0 Hz, 1H),
5.78 (d, J = 15.5 Hz, 1H), 5.29 (s, 1H), 5.27 (s, 1H), 4.09 (dd, J = 13.2,
6.4 Hz, 2H), 2.37–2.13 (m, 5H), 1.94–1.85 (m, 2H), 1.58–1.42 (m, 4H),
1.34–1.25 (m, 3H), 1.20 (m, 6H), 1.05 (d, J = 7.0 Hz, 6H), 0.69 (s, 3H).
13C NMR (100 MHz, DMSO‑d6) δ 198.89, 165.98, 165.30, 164.15,
154.50, 118.79, 117.65, 99.83, 59.69, 53.03, 50.42, 48.87, 44.90,
43.47, 38.80, 38.27, 35.45, 32.49, 27.95, 26.30, 25.43, 21.04, 19.19,
16.91, 14.17, 12.09. HRMS(ESI): calcd for C26H36NaO4 [M + Na]+
435.2506, found 435.2501.
,
2.2.6. (E)-3,3-ethylenedioxy-7-oxo-5,22-choladienoic acid ethyl ester (7)
To a solution of 6 (5.0 g, 11.30 mmol) in a mixed solvent (100 mL)
of acetone-H2O (9:1, v/v) was added NHPI (2.0 g, 12.43 mmol), PDC
(4.7 g, 12.43 mmol) at room temperature. The reaction mixture was
stirred for 20 h and the solid residue was removed by filtration, then the
2.2.8. Ursodeoxycholic acid (UDCA)
To a solution of 8 (2.0 g, 4.85 mmol) in 2-methyl tetrahydrofuran
(20 mL) in autoclave was hydrogenated in the presence of Raney-Ni
(2.0 g) under H2 (4.0 MPa) for 24 h at 90 °C. Then i-PrOH (20 mL), t-
Scheme 1. Synthesis of ursodeoxycholic acid from bisnoralcohol. Reagents and conditions: (a) ethylene glycol, pTSA, benzene, reflux, 88%; (b) TEMPO, NaHCO3,
TBAB, NCS, CH2Cl2, H2O, 0 °C, 95%; (c) Ph3 = CHCOOC2H5, toluene, reflux, 98%; (d) TEMPO, NaHCO3, TBAB, NCS, CH2Cl2, H2O, 0 °C, 95%; (e)
Ph3 = CHCOOC2H5, toluene, reflux, 98%; (f) ethylene glycol, pTSA, benzene, reflux, 88%; (g) PDC, NHPI, acetone, H2O, rt, 85%; (h) H2SO4, THF, rt, 98%; (i) Raney
Ni, H2, t-BuONa, 2-methyl tetrahydrofuran, i-PrOH, 90 °C, 87%.
3

J. Wang, et al.
Steroids157(2020)108600
BuONa (932 mg, 9.70 mmol) was added and flushed with H2 (4.0 MPa).
The reaction mixture was stirred for 24 h at 90 °C. After cooling, the
reaction mixture was acidified to pH 5 with AcOH and the catalyst was
removed by filtration through celite, then the filtrate was evaporated
under reduced pressure. The residue was poured into H2O (100 mL) and
extracted with AcOEt (30 mL × 3). The organic layer was washed with
brine, dried with anhydrous Na2SO4 and concentrated. The residue was
purified by silica gel chromatography (CH2Cl2/MeOH, 20/1, v/v) to
give UDCA (1.65 g, 87%) as a white solid; mp: 200–202 °C [lit. [13]
198–200 °C]. [α]25 D + 60.9 (c 1.0, CH3CH2OH). [lit. [13] [α]25
D + 59.7 (c 1.0, CH3CH2OH)].1H NMR (400 MHz, DMSO‑d6) δ 11.94
(s, 1H, –COOH), 4.46 (s, 1H, H-3), 3.88 (d, J = 6.7 Hz, 1H, H-7),
3.35–3.24 (m, 2H), 2.26–2.19 (m, 1H), 2.13–2.05 (m, 1H), 1.95–1.81
(m, 2H), 1.78–1.63 (m, 4H), 1.51–1.42 (m, 3H), 1.41–1.28 (m, 7H),
1.23–1.08 (m, 6H), 1.05–0.91 (m, 2H), 0.88 (d, J = 6.5 Hz, 6H, H-19,
H-21), 0.61 (s, 3H, H-18). 13C NMR (100 MHz, DMSO‑d6) δ 174.93 (C-
24), 69.75 (C-3), 69.46 (C-7), 55.87 (C-17), 54.70 (C-14), 43.11 (C-13),
43.02 (C-8), 42.20 (C-5), 39.94 (C-12), 39.84 (C-9), 39.73 (C-4), 37.75
(C-6),37.28 (C-20), 34.88 (C-1), 33.78 (C-10), 30.78 (C-22, C-23),
30.26 (C-2), 28.21 (C-16), 26.75 (C-15), 23.34 (C-19), 20.89 (C-11),
3. Results and discussion
The synthesis of UDCA is shown in Scheme 1. Compound 2 was
obtained by glycol protection of the C-3 carbonyl group in 88% yield.
Oxidation of side chain hydroxyl groups to aldehydes in the presence of
TEMPO and NCS gave Compound 3 and 4 in 95% yield respectively.
Compound 5 was prepared by witting reaction in 98% yield. The key
intermediate compound 6 was afforded by witting reaction from com-
pound 3 in 98% yield or glycol protection of the C-3 carbonyl group
from compound 5 in 88% yield. The witting reaction with a resonance-
stabilized is known to predominantly deliver trans stereoisomer, and
the two isomers (cis- and trans- isomers) show different chemical shifts
on 1H NMR and 13C NMR spectra. Herein, the compounds 5 and 6
obtained by witting reaction showed a single trans structure on 1H NMR
and 13C NMR spectra respectively. The yield of oxidation at the C-7
position to afford compound 7 was unsatisfactory. Therefore, a model
system was employed to find an ideal solvent and optimal reaction
conditions (Table 1). After screening various oxidant, we found that the
oxidant of PDC/NHPI was the optimal oxidant for this reaction, and the
yield was up to 72% (Table 1, entry 1–12). Then we examined several
different solvents, and the results showed that acetone-H2O (9:1, v/v) is
the optimal solvent (Table 1, entry 4, 10 and 13–24). We also
18.32 (C-21), 12.06 (C-18). HRMS(ESI): calcd for
[M + Na]+, 415.2819, found 415.2834.
C24H40NaO4
Table 1
Oxidation of compound 6.a
Entry
Solvent
Oxidant
RTb(°C)
Yieldc(%)
1
2
Toluene
Toluene
Na2Cr2O7/NHPI
Na2Cr2O7/NHPI/Acetic
acid
25
25
Nd
54
3
Toluene
K2Cr2O7/NHPI/Acetic
acid
25
52
4
5
6
7
8
Toluene
Toluene
Toluene
Acetone
Acetone
PDC/NHPI
25
25
25
25
25
66
43
42
Nd
62
PDC/TBHP
BPO/NHPI
Na2Cr2O7/NHPI
Na2Cr2O7/NHPI/Acetic
acid
9
Acetone
K2Cr2O7/NHPI/Acetic
acid
25
61
10
11
12
13
14
15
16
17
18
19
20
21
22
Acetone
PDC/NHPI
25
25
25
25
25
25
25
25
25
25
25
25
25
72
59
51
38
Nd
Nd
70
Nd
68
77
79
79
80
Acetone
PDC/TBHP
BPO/NHPI
Acetone
DMF
PDC/NHPI
t-BuOH
PDC/NHPI
Ethyl acetate
PDC/NHPI
MeCN
PDC/NHPI
NMP
PDC/NHPI
DCM
PDC/NHPI
MeCN/H2O (4:1, v/v)
PDC/NHPI
MeCN/H2O (9:1, v/v)
PDC/NHPI
MeCN/H2O (14:1, v/v)
PDC/NHPI
Acetone/H2O (4:1, v/
PDC/NHPI
v)
23
24
25
26
Acetone/H2O (9:1, v/
PDC/NHPI
PDC/NHPI
PDC/NHPI
PDC/NHPI
25
25
0
85
82
65
73
v)
Acetone/H2O (14:1, v/
v)
Acetone/H2O (9:1, v/
v)
Acetone/H2O (9:1, v/
v)
50
a
b
c
All the reactions were performed for 20 h, and the ratio of oxidant/compound 6 was 1.1:1 (mol: mol).
Reaction temperature.
Isolated yield.
d
No reaction.
4

J. Wang, et al.
Steroids157(2020)108600
Table 2
Reduction and hydrolysis of compound 8.a
Entry
Solvent
Base
RTb(°C)
Yieldc(%)
1
i-PrOH
t-BuOK
60
60
60
60
40
90
90
90
90
90
90
90
38
35
36
Nd
33
61
44
43
72
81
70
87
2
n-BuOH
t-BuOK
3
t-BuOH
t-BuOK
4
THF
t-BuOK
5
i-PrOH
t-BuOK
6
i-PrOH
t-BuOK
7
i-PrOH
CH3ONa
C2H5ONa
t-BuONa
t-BuONa
t-BuONa
t-BuONa
8
i-PrOH
9
i-PrOH
10
11
12
THF/i-PrOH (1/1, v/v)
1, 4-dioxane/i-PrOH (1/1, v/v)
2-methyl tetrahydrofuran/i-PrOH (1/1,
v/v)
13e
14e
15e
i-PrOH
t-BuONa
t-BuONa
t-BuONa
90
90
90
60
65
68
THF/i-PrOH (1/1, v/v)
2-methyl tetrahydrofuran/i-PrOH (1/1,
v/v)
a
b
c
d
e
All the reactions were performed for 48 h, and the ratio of Raney-Ni/compound 8 was 1/1 (m/m).
Reaction temperature.
Isolated yield.
No UDCA.
Solvent and base were added together (The rest were added separately).
development of novel UDCA derivatives for PBC disease.
Acknowledgment
This work was supported by Shanghai Science and Technology
Council (Grant 18ZR1411200) and Jiangsu Jiaerke Pharmaceuticals
Group Co., Ltd.
Fig. 2. Single-crystal X-ray structure of ursodeoxycholic acid.
investigated the reaction temperature, and discovered that 25 °C was
the most suitable temperature, and the yield was up to 85% for the
oxidation reaction (entry 23, 25 and 26). Compound 8 was furnished by
hydrolysis of the ethylene ketal of compound 7 in the presence of
H2SO4 and THF in 98% yield. Finally, UDCA was afforded by reduction
and hydrolysis of compound 8 under H2 (4 MPa), Raney-Ni and base in
autoclave. After screening various solvents, reaction temperature and
bases (Table 2) of the reaction, we found that a mixed solvent of 2-
methyl tetrahydrofuran-i-PrOH, 90 °C and t-BuONa were optimum
conditions and the yield reach 87%. In this step, five functional groups
were converted with calcd 97% per conversion in one-pot reaction. The
relative stereo-configuration of UDCA was determined by X-ray analysis
[24] (Fig. 2). In this straightforward methodology for preparation of
UDCA, most of the conversions are very efficient with an average yield
of 91% in 6 steps and overall yield up to 59%.
Appendix A. Supplementary data
Supplementary data to this article can be found online at https://
doi.org/10.1016/j.steroids.2020.108600.
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