Synthesis of diastereomers of 1,3-cis-25-dihydroxy-19-norvitamin D3

Article abstract of DOI:10.1248/cpb.c16-00311

1β,3β,25-Dihydroxy-19-norvitamin D₃ (4a) and 1α,3α,25-dihydroxy-19-norvitamin D₃ (4b) were synthesized by employing a new A-ring synthon, (1R,3S)-3-((tert-butyldimethylsilyl)oxy)-5-oxocyclohexyl benzoate (19), which was derived from D-(-)-quinic acid in 12 steps. The A-ring was coupled with the circular dichroism (CD) ring by means of Julia-Kocienski olefination to construct the diene unit. The structures of the products were confirmed by ¹H-NMR and nuclear Overhauser effect (NOE) experiments.

Full text of DOI:10.1248/cpb.c16-00311

1190
Chem. Pharm. Bull. 64, 1190–1195 (2016)
Vol. 64, No. 8
Regular Article
Synthesis of Diastereomers of 1,3-cis-25-Dihydroxy-19-norvitamin D3
Kosuke Usuda,a,# Tanima Biswas,b,# Takuya Yamaguchi,a Yusuke Akagi,a Koji Yasui,a
,b
,a
Motonari Uesugi,c,d Isao Shimizu,b Seijiro Hosokawa,* and Kazuo Nagasawa*
a Graduate School of Technology, Tokyo University of Agriculture and Technology; 2–14–16 Naka-cho, Koganei,
b
Tokyo 184–8588, Japan: Faculty of Science and Technology, Waseda University; 3–4–1 Okubo, Shinjuku-ku,
c
Tokyo 169–8555, Japan: Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University; Yoshida-
d
Honmachi, Sakyo-ku, Kyoto 606–8115, Japan: and Institute for Chemical Research, Kyoto University; Gokasho, Uji,
Kyoto 611–0011, Japan.
Received April 4, 2016; accepted May 12, 2016
1β,3β,25-Dihydroxy-19-norvitamin D3 (4a) and 1α,3α,25-dihydroxy-19-norvitamin D3 (4b) were synthe-
sized by employing a new A-ring synthon, (1R,3S)-3-((tert-butyldimethylsilyl)oxy)-5-oxocyclohexyl benzoate
(19), which was derived from D-(ꢀ)-quinic acid in 12 steps. The A-ring was coupled with the circular dichroism
(CD) ring by means of Julia–Kocienski olefination to construct the diene unit. The structures of the products
1
were confirmed by H-NMR and nuclear Overhauser effect (NOE) experiments.
Key words 19-norvitamin D3; cis-A ring synthon; Julia–Kocienski coupling
1α,25-Dihydroxyvitamin D3 (2, Fig. 1; also known as calcitri-
Synthesis of ketone 6 is illustrated in Chart 2. 1,3,5-
ol), which is the hormonally active form of vitamin D3 (1) in Cyclohexanetriol (7) was reacted with the phenylboronic acid
mammalians, regulates calcium and phosphorus homeostasis,1–3) in toluene under reflux to give the boronate, whose free hy-
and also has antiproliferative, cell differentiation-inducing and droxy group was then protected as benzoate ester by reaction
immune-regulatory activities.1,4–6) It is used to treat hyperprolif- with benzoyl chloride in the presence of triethylamine, afford-
erative disorders, but its clinical utility is limited by side effects ing 8 in 76% yield (2 steps).19,20) The boronate ester in 8 was
such as hypercalcemia and hypercalciuria, which are induced at hydrolyzed using pinacol in the presence of BF3·Et2O, and the
clinically effective doses. Hence, there is a need for structural free hydroxy groups were protected as tert-butyldimethylsilyl
development to separate these biological activities.7–11)
(TBS) ether using TBS chloride in the presence of triethyl-
19-Norvitamin D3 (3) was first reported by DeLuca and col- amine and tetrabutylammonium bromide (TBAB), affording
leagues in 1990,12) and has a characteristic biological activity 9 in 52% yield. After hydrolysis of benzoyl group in 9 with
profile, exhibiting high differentiation potency towards vari- sodium hydroxide in methanol, the resulting hydroxy group
ous malignant tissues with a substantially reduced calcemic was oxidized with pyridinium chlorochromate (PCC) to give
effect.13) Since then, there have been many synthetic studies ketone 6 in 56% yield.
of 3, mainly focusing on the A-ring synthon.14,15) We recently
With ketone 6 in hand, the CD ring synthon 521–23) and
reported a new synthetic strategy.16) We are particularly inter- ketone 6 were coupled using lithium bis(trimethylsilyl)amide
ested in structural derivatization involving the C1 and C3 hy-
droxy groups, whose geometries are believed to be crucial for
antiproliferative activity.17,18) Herein, we described syntheses
of 1β,3β,25-dihydroxy-19-norvitamin D3 (4a) and 1α,3α,25-
dihydroxy-19-norvitamin D3 (4b), using a newly developed
methodology to obtain the 1,3-cis-type A-ring synthon 19.
Results and Discussion
Synthetic methodologies for 1,3-trans-19-nor type vitamin
D derivatives are well developed, but to our knowledge, there
is no report of the synthesis of 1,3-cis type 19-nor analogues
or their A-ring synthons. As we were interested in synthesiz-
ing 1,3-cis type analogues, we decided to investigate Julia–
Kocienski olefination as a means to obtain two diastereomers
at the same time, namely 1β,3β,25-dihydroxy-19-norvitamin
D3 (4a) and 1α,3α,25-dihydroxy-19-norvitamin D3 (4b) (Chart
1). However, because of the structural similarity and similar
polarity of these compounds, separation, and structural identi-
fication could be challenging.
# These authors contributed equally to this work.
Fig. 1. Structures of Vitamin D3 (1), 1α,25-Dihydroxyvitamin D3 (2),
Isao Shimizu (Deceased on 30th October 2014.)
and 1,3-trans-(3) and 1,3-cis-25-Dihydroxy-19-norvitamin D3 (4a, b)
*To whom correspondence should be addressed. e-mail: seijiro@waseda.jp; knaga@cc.tuat.ac.jp
© 2016 The Pharmaceutical Society of Japan

Vol. 64, No. 8 (2016)
Chem. Pharm. Bull.
1191
Chart 1. Retrosynthetic Strategy for 1,3-cis-25-Dihydroxy-19-norvita-
min D3 (4a, b) Employing Julia–Kocienski Olefination
Chart 2. Synthesis of A-Ring Synthon 6
Chart 3. Julia–Kocienski Coupling of Compounds 5 and 6
(LiHMDS) in tetrahydrofuran (THF)24,25) to give 10 in 98%
yield as a 1:1 mixture of diastereomers 10a and b (Chart 3).
However, separation of these two diastereomers was difficult.
1
Moreover, the chemical shifts of H at C1 and C3 of these two
isomers 10a and b overlapped (δ=3.6–3.9ppm), and structural
identification of these isomers was problematic. Therefore, we
decided to modify the hydroxy protecting groups at the A-
ring in order to obtain products with different polarity, which
would be more easily separable.
Specifically, we planned to change one silyl ether to benzo-
ate ester in ketone 6 to obtain 19. The strongly electron-with-
drawing nature of the benzoyl group would make the adjacent
Chart 4. Attempt to Synthesize 19 from 6
C1–H more deshielded than its C3 counterpart, which should
1
enable an easy identification of both H’s in H-NMR. This TBAF in THF (1M in THF, 1eq) at 0°C to give a mixture of
group was also expected to induce some polarity difference four stereoisomers 16 in 70% yield. This mixture of isomers
between the two target compounds, enabling us to separate was oxidized with tetra-n-propylammonium perruthenate-N-
them easily on a preparative scale. Thus, we attempted selec- methylmorpholine N-oxide (TPAP-NMO) to give ketone 17 (a
tive deprotection of one TBS group in 6 using tetrabutylam- mixture of four isomers). Various conditions were investigated
monium fluoride (TBAF) (1eq) or hydrogen fluoride·pyridine for the stereoselective reduction of ketone 17 to 1,3-cis alcohol
(HF·Py) (1eq). However, β-elimination and subsequent aro- 18. Among them, diisobutylaluminium hydride (DIBAL)-H
26)
matization reaction took place to generate 12 and 13, respec- in the presence of ZnCl2 was effective, and 1,3-cis alcohol
tively (Chart 4). These results led us to develop a new route to 18 (more polar) was obtained in 67% yield, together with
synthesize an A-ring synthon with different hydroxy protect- 1,3-trans alcohol 16 (less polar, 29%) after silica gel column
ing groups at the C1 and C3 positions (19).
separation.
We started our synthesis of 19 from 14, which can be ob-
After protection of the hydroxy group in 18 as benzoate by
tained from D-(−)-quinic acid in five steps12) (Chart 5). The reaction with benzoyl chloride and triethylamine, deprotec-
diol in 14 was protected with benzaldehyde dimethylacetal to tion of benzylidene acetal under hydrogenation conditions in
give acetal 15 in 82% yield. We found that selective deprotec- the presence of palladium hydroxide followed by oxidative
tion of one TBS group in 15 proceeded upon slow addition of cleavage of the resulting diol with NaIO4 aqueous solution in

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Chem. Pharm. Bull.
Vol. 64, No. 8 (2016)
Chart 5. Synthesis of 1,3-cis Alcohol 18
Experimental
All reactions have been carried out in dry solvents, and
under inert atmosphere unless otherwise mentioned. Reagents
were purchased from Sigma-Aldrich (St. Louis, MO, U.S.A.),
TCI and Wako Pure Chemical Industries, Ltd. (Osaka,
Japan). Flash chromatography was performed on silica gel 60
(spherical, particle size 40–100mm; Kanto (Tokyo, Japan)).
1H-NMR spectra have been recorded in deuteriochloroform
at 300 and 400MHz using JMTC 300 and JNM-ECX 400
spectrometers, whereas 13C-NMR spectra have been recorded
at 75 and 100MHz using same spectrometers. The spectra
are referenced internally according to residual solvent signal
of CDCl3 (1H-NMR: δ=7.26ppm; 13C-NMR: δ=77.0ppm).
1
Data for H-NMR are recorded as follows: chemical shift (δ
ppm), multiplicity (s=singlet; d=doublet; t=triplet; q=quartet;
m=multiplet; br=broad), coupling constant (Hz), and integra-
tion. Data for 13C-NMR are reported in terms of chemical
shift (δ ppm). For mixture of diastereomers NMR of single
pure diastereomer or the peaks for major isomers is reported,
in other cases diastereomeric protons have been reported in
italics. Mass spectra were recorded on a JMS-T100X (JEOL)
spectrometer in electrospray ionization (ESI)-MS mode using
methanol as solvent.
Chart 6. Synthesis of 4a and b
methanol afforded ketone 19. Coupling reaction of CD-ring
(1R,5S)-3-Phenyl-2,4-dioxa-3-borabicyclo[3.3.1]nonan-
synthon 5 with ketone 19 was performed to give 20a and b. 7-yl Benzoate (8) 1,3,5-Cyclohexanetriol (3.0g, 22.69mmol)
These two diastereomers were separable by silica gel column was dissolved in toluene (250mL) under argon, and the
chromatography, affording 20a and b in 36 and 35% yield, mixture was refluxed for 6h, when no left over starting ma-
respectively. The structures of these isomers were confirmed terial was found at TLC, the reaction was cooled to room
1
by H-NMR and nuclear Overhauser effect correlated spec- temperature and the solvent was evaporated under reduced
troscopy (NOESY)27) (Chart 6).
pressure, the white solid left behind was the boronate ester
These isomers were converted to 1,3-cis-19-nor vitamin D3 (4.95g, quant.). The boronate ester (1g, 4.58mmol) was then
4a and b in 56 and 59% yield, respectively, by reaction with dissolved in pyridine (15mL) at room temperature, then ben-
potassium carbonate in methanol–THF followed by deprotec- zoyl chloride (0.51mL, 0.62g, 4.41mmol) was added, and
tion of silyl ethers with HF·Py.
the mixture was allowed to stir for 12h at room temperature.
After the reaction was finished, pyridine was evaporated off
under reduced pressure, the residue was dissolved in benzene
Conclusion
In conclusion, synthesis of 1,3-cis-19-nor type A-ring syn- and was filtered. The filtrates were concentrated in vacuo and
thon from 1,3,5-cyclohexanetriol was problematic due to the dried under high vacuum for 12h. Then, it was dissolved in
difficulty of separation and identification of the 1,3-cis-19-nor- minimum amount of benzene at 80°C and was allowed to cool
vitamin D3 isomers 4a and b. A simple strategic modification down gradually overnight, the solids were filtered out and
to protect one hydroxy group with benzoate afforded A-ring washed with hexane to give 8 (1.12g, 76%).
synthon 19. Finally, 4a and b were synthesized by Julia–Ko-
(3R,5S)-3,5-Bis[(tert-butyldimethylsilyl)oxy]cyclohexyl
cienski olefination of this A-ring synthon and a circular dichro- Benzoate (9) Compound 8 (500mg, 1.55mmol) was first
ism (CD) ring synthon, thus opening up a new route that should dissolved in CH2Cl2 (15mL), at room temperature, under
be available to build a library of 1,3 modified 1,3-cis-19-nor argon, then pinacol (365mg, 3.0mmol) was added followed
type vitamin D derivatives in a convergent manner.
by BF3·OEt2 (32mL, 0.77mmol). This mixture was stirred

Vol. 64, No. 8 (2016)
Chem. Pharm. Bull.
1193
for 4h at room temperature, then the solvent was evaporated Combined organic layer was washed with water and brine,
off under reduced pressure, the left over gum was dissolved dried on MgSO4, filtered and evaporated in vacuo. The residue
in methanol and evaporated under reduced pressure, this was purified on silica gel column (2% ethyl acetate–hexanes)
process was repeated for five times, and then the left over to give 15 (2.54g, 82%) as a mixture of two diastereomers.
sticky substance was treated as the crude for the next step.
The crude product was dissolved in N,N-dimethylformamide
Spectral Data for 15
1H-NMR (300MHz, CDCl3) δ: 7.50–7.46 (2H, m), 7.41–7.34
(DMF) (15.5mL) at 0°C. To the mixture was added tetrabutyl (3H, m), 5.92 (5.81) (1H, s), 4.26–4.25 (1H, m), 4.08–3.98
ammonium bromide (48mg, 0.15mmol), N,N-dimethyl-4-ami- (2H, m), 3.92–3.68 (1H, m), 2.28 (2.17) (1H, d, J=11.9Hz),
nopyridine (DMAP) (189mg, 1.55mmol), and TBSCl (2.34g, 1.97–1.84 (3H, m), 1.67–1.59 (1H, m), 1.48–1.34 (1H, m), 0.90
15.5mmol), then triethylamine (0.59mL, 434mg, 4.27mmol) (18H, d, J=5.9Hz), 0.1–0.05 (12H, q, J=6.4Hz); 13C-NMR
was added dropwise, after the addition was over the reaction (75MHz, CDCl3) δ: 138.71 (138.11), 129.29 (129.15), 128.43,
was raised at room temperature, and allowed to stand at room 126.79 (126.58), 102.57 (101.49), 81.52 (80.99), 75.29, 67.38
temperature for 15h. After the reaction was finished, water (67.16), 66.09 (65.83), 46.34 (44.42), 42.73 (42.68), 41.89
(75mL) was added to the reaction, and then the aqueous layer (41.80), 25.99, 25.89, 18.27, 18.05, −4.54, −4.83. HR-MS ESI:
was extracted with CH2Cl2 for three times. The combined m/z: Calcd for C26H46Na1O4Si2: 501.2832 [M+Na]+. Found:
organic layer was washed with water for three times, and then 501.2805.
was washed with brine, dried over MgSO4, filtered and evapo-
(7S,9S)-9-[(tert-Butyldimethylsilyl)oxy]-2-phenyl-1,3-di-
rated in vacuo. The residue was purified by silica gel column oxaspiro[4.5]decan-7-ol (16) To a solution of 15 (1.15 g,
(5% ethyl acetate–hexanes) to give 9 (372mg, 2 steps 52%).
Spectral Data for 9
2.4mmol) in THF (48mL) was added TBAF (1M in THF,
2.4mL, 2.4mmol) at 0°C dropwise over 1h. The reaction was
1H-NMR (300MHz, CDCl3) δ: 7.60–7.33 (5H, m), 5.06–4.89 monitored by TLC carefully. At the point of polar bis-TBS de-
(1H, m), 3.74–3.63 (2H, m), 2.27–2.04 (3H, m), 1.66–1.39 (3H, protected spot appearance on TLC, the reaction was quenched
m), 0.87 (18H, s), 0.06 (12H, s); 13C-NMR (75MHz, CDCl3) with saturated NH4Cl. The aqueous layer was extracted with
δ: 165.98, 134.83, 133.06, 131.3, 130.64, 129.68, 70.75, 68.62, ethyl acetate for three times, and the combined organic layer
66.29, 45.85, 40.89, 25.90, 18.30, −4.54. High resolution (HR)- was washed with water, followed by brine, dried over MgSO4,
MS ESI: m/z: Calcd for C25H44Na1O4Si2: 487.2676 [M+Na]+. filtered and evaporated in vacuo. The residue was chromato-
Found: 487.2658.
graphed on silica gel column (10–15% ethyl acetate–hexanes)
(3R,5S)-3,5-Bis[(tert-butyldimethylsilyl)oxy]cyclohexa- to give 16 as four diastereomers mixtures (399mg, 70%, brsm).
none (6) To a solution of 9 (500mg, 1.08mmol) in CH2Cl2
(22mL) was added a solution of NaOH (260mg, 6.5mmol) in
Spectral Data for 16
1H-NMR (300MHz, CDCl3) δ: 7.50–7.46 (2H, m), 7.40–7.36
MeOH (44mL), dropwise at 0°C. The resulting mixture was (3H, m), 5.85 (1H, s), 4.34–4.26 (1H, m), 4.17–4.08 (1H, m),
allowed to stand overnight, the reaction was quenched with 4.02 (1H, d, J=8.6Hz), 3.80 (1H, d, J=8.9Hz), 2.07 (1H, dd,
1M HCl solution, the aqueous layer was washed with CH2Cl2 J=3.4, 13.4, Hz), 2.00–1.66 (5H, m), 0.89 (9H, s), 0.09 (6H, s);
twice, the combined organic layer was washed with water, 13C-NMR (75MHz, CDCl3) δ: 137.63, 129.56, 128.52, 126.73,
followed by brine, dried over MgSO4, filtered and evapo- 103.22, 82.46, 67.20, 65.11, 44.29, 42.31, 40.06, 25.43, 18.19,
rated under reduced pressure. The residue was dissolved in −4.69, −4.73. HR-MS ESI: m/z: Calcd for C20H32Na1O4Si1:
CH2Cl2 (11mL), and to the solution was added NaOAc (44mg, 387.1976 [M+Na]+. Found: 387.1994.
0.54mmol) and PCC (395mg 1.87mmol) at 0°C and the result-
(R)-9-[(tert-Butyldimethylsilyl)oxy]-2-phenyl-1,3-dioxas-
ing mixture was stirred for 15min. The reaction mixture was piro[4.5]decan-7-one (17) To a solution of alcohol 16
filtered through a pad of Celite, the filtrates were washed with (410mg, 1.1mmol) in CH2Cl2 (11mL) was added activated
saturated Na2S2O3 solution, the aqueous layer was extracted molecular sieves (553mg) and N-methylmorpholene-N-oxide
with CH2Cl2. The combined organic layer was washed with (322mg, 2.75mmol) at 0°C under argon. After stirring for
brine, dried over MgSO4, filtered and evaporated in vacuo. The 30min, TPAP (39mg, 0.11mmol) was added to the mixture,
residue was purified on silica gel column (2% ethyl acetate– and the resulting mixture was stirred at 0°C for another 3h.
hexanes) to give ketone 6 (217mg, 2 steps 56%).
Spectral Data for 6
The reaction mixture was filtered through a short silica gel col-
umn (6–8% ethyl acetate–hexanes) to give 17 (407mg, 99%).
Spectral Data for 17
1H-NMR (300MHz, CDCl3) δ: 3.83–3.73 (2H, m), 2.56–2.50
(1H, dd, J=5.2, 14.1Hz), 2.38–2.30 (2H, t, J=11.0Hz),
1H-NMR (300MHz, CDCl3) δ: 7.46–7.42 (2H, m),
2.29–2.22 (1H, m), 1.82–1.71 (1H, dd, J=11.0, 12.7Hz), 7.39–7.35 (3H, m), 5.88 (1H, s), 4.39–4.31 (1H, m), 4.00 (1H,
0.86 (18H, s), 0.06 (12H, s); 13C-NMR (75MHz, CDCl3) δ: d, J=8.6Hz), 3.78 (1H, d, J=8.6Hz), 2.69–2.60 (3H, m),
207.67, 66.85, 51.00, 45.11, 25.81, 18.13, −4.68. HR-MS ESI: 2.43–2.31 (2H, m), 2.03–1.96 (1H, m), 0.87 (9H, s), 0.05 (6H,
m/z: Calcd for C18H38Na1O3Si1: 381.2257 [M+Na]+. Found: s); 13C-NMR (75MHz, CDCl3) δ: 206.06, 137.78, 129.46,
381.2298.
128.50, 126.55, 103.25, 81.18, 75.25, 66.86, 50.16, 49.75,
{[(7S,9S)-2-Phenyl-1,3-dioxaspiro[4.5]decane-7,9-diyl]- 43.54, 25.83, 18.10, −4.78, −4.82. HR-MS ESI: m/z: Calcd for
bis(oxy)}bis(tert-butyldimethylsilane) (15) To a solution of C20H30Na1O4Si1: 385.1811 [M+Na]+. Found: 385.1851.
14 (2.53g, 6.47mmol) in CH2Cl2 (64.7mL) was added activat-
(7R,9S)-9-[(tert-Butyldimethylsilyl)oxy]-2-phenyl-1,3-
ed 4Α molecular sieves (3.23g), benzaldehyde dimethyl acetal dioxaspiro[4.5]decan-7-ol (18) To a mixture of ketone 17
(1.24mL, 1.27g, 8.41mmol), and the mixture was stirred for (400mg, 1.1mmol) and ZnCl2 (300mg, 2.2mmol) in CH2Cl2
12h at room temperature. The reaction mixture was filtered (11mL) was added DIBAL-H (4.4mL, 1M in toluene) at −78°C
through a pad of Celite, the filtrate was washed with water, dropwise, and the resulting mixture was stirred at −78°C for
aqueous layer was extracted with CH2Cl2 for three times. 12h. To the reaction mixture was added MeOH (3mL) at

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Chem. Pharm. Bull.
Vol. 64, No. 8 (2016)
−78°C, and the resultant mixture was warmed to room tem- To a mixture of CD-ring synthon 5 (36mg, 59µmol) and ke-
perature. Then, saturated Rochelle salt solution (5.2mL) was tone 19 (14mg, 39µmol) in THF (0.6mL) was added LiHMDS
added, and allowed to stir for 30min. The aqueous layer was (79µL, 78µmol) was slowly at −78°C under argon, and the
extracted with CH2Cl2 for three times. The combined organic mixture was stirred for 2.5h, then the mixture was raised
layer was washed with saturated Rochelle salt and brine, dried at room temperature and allowed to stand for 10min. To the
over MgSO4, filtered, and concentrated in vacuo. The residue reaction mixture was added water, and extracted with ether
was chromatographed on silica gel column (6–8, 9–10% ethyl for three times. The combined extracts were washed with
acetate–hexanes) to give trans alcohols 16 (less polar, 115mg, brine, dried over MgSO4, filtered and concentrated in vacuo.
29%) and cis alcohols 18 (more polar, 226mg, 67%).
Spectral Data for 18
The residue was passed through a small silica gel column to
give 20 as two diastereomer’s mixture (35mg, 72%). These
1H-NMR (300MHz, CDCl3) δ: 7.48–7.44 (2H, m), 7.41–7.32 diastereomers were separated on a silica gel column (0.3%
(3H, m), 5.85 (1H, s), 4.13–4.02 (3H, m), 3.85 (1H, d, ether–hexane) to give 20a (18mg, 36%) and 20b (17mg, 35%).
J=8.3Hz), 2.13–2.00 (3H, m), 1.64–1.43 (3H, m), 0.89 (9H,
Spectral Data for 20a
s), 0.07 (6H, s); 13C-NMR (75MHz, CDCl3) δ: 137.91, 129.47,
1H-NMR (400MHz, CDCl3) δ: 8.05 (2H, d, J=6.9Hz),
128.49, 126.79, 103.35, 80.40, 67.35, 66.71, 44.36, 42.91, 7.55 (1H, t, J=7.3Hz), 7.44 (2H, t, J=7.8Hz), 6.26 (1H, d,
42.34, 25.94, 18.19, −4.69, −4.76. HR-MS ESI: m/z: Calcd for J=10.9Hz), 5.80 (1H, d, J=11.5Hz), 4.91–4.90 (1H, m),
C20H32Na1O4Si1: 387.1966 [M+Na]+. Found: 387.1994.
3.71–3.67 (1H, m), 3.13 (1H, dd, J=4.6, 12.8Hz), 2.80 (1H,
(1R,3S)-3-[(tert-Butyldimethylsilyl)oxy]-5-oxocyclohexyl d, J=11.5Hz), 2.45 (1H, dd, J=4.6, 13.1Hz), 2.36 (1H, m),
Benzoate (19) To a solution of cis alcohols 18 (219mg, 2.14 (1H, t, J=11.9Hz), 2.02–1.86 (3H, m), 1.67–1.60 (4H, m),
0.6mmol) in CH2Cl2 (5mL) was added benzoyl chloride 1.41–1.24 (13H, m), 1.17 (6H, s), 0.96–0.82 (22H, m), 0.58–0.52
(0.15mL, 186mg, 1.32mmol), triethylamine (0.24mL, 182mg, (9H, m), 0.07 (6H, d, J=6.9Hz); 13C-NMR (100MHz, CDCl3)
1.8mmol), and DMAP (102mg, 0.84mmol) at room tempera- δ: 165.98, 143.35, 132.99, 130.65, 130.23, 129.72, 128.42,
ture under argon, and the resultant mixture was stirred for 4h. 123.05, 115.57, 73.59, 70.49, 68.76, 56.73, 56.46, 46.45, 45.94,
To the reaction mixture was added saturated NH4Cl, and the 45.62, 41.85, 40.62, 36.55, 36.22, 33.74, 30.11, 29.93, 28.97,
aqueous layer was extracted with CH2Cl2 for three times. 27.73, 25.95, 23.59, 22.39, 20.94, 18.92, 18.21, 12.26, 7.24, 6.91,
Combined extracts were washed with water and brine, dried −4.48, −4.54. HR-MS ESI: m/z: Calcd for C45H76Na1O4Si2:
over MgSO4, filtered and concentrated in vacuo. The residue 759.5180 [M+Na]+. Found: 759.5205.
was chromatographed on the silica gel column (5% ethyl ace-
tate–hexanes) to give benzoate-protected cis alcohols (264mg,
Spectral Data for 20b
1H-NMR (400MHz, CDCl3) δ: 8.04 (2H, d, J=7.3Hz),
94%). The mixture of benzoate-protected cis alcohols (264mg, 7.55 (1H, t, J=7.8Hz), 7.44 (2H, t, J=7.8Hz), 6.29 (1H, d,
0.564mmol) was dissolved in THF (6mL) under argon, and J=10.9Hz), 5.84 (1H, d, J=11.5Hz), 4.91–4.88 (1H, m),
10% Pd(OH)2 (26mg) was added, then argon was purged with 3.62–3.58 (1H, m), 2.99 (1H, dd, J=4.6, 10.9Hz), 2.79 (1H,
hydrogen gas, and the reaction mixture was allowed to stir d, J=12.4Hz), 2.66 (1H, dd, J=4.6, 12.4Hz), 2.37–2.34 (1H,
under the hydrogen atmosphere for 8h. The reaction mixture br), 2.25–2.20 (1H, t, J=11.5Hz), 2.02–1.66 (9H, m), 1.43–1.30
was filtered through a pad of Celite, and filtrates were concen- (7H, m), 1.19 (6H, s), 0.96–0.83 (23H, m), 0.59–0.53 (9H, m),
trated in vacuo. The residue was dissolved in MeOH (28mL), 0.09–0.07 (6H, d, J=8.2Hz); 13C-NMR (100MHz, CDCl3) δ:
and a saturated aqueous solution of NaIO4 (542mg, 2.53mmol, 165.96, 143.38, 132.96, 130.62, 130.38, 129.68, 128.39, 123.05,
in 0.28mL H2O) was added, the resulting mixture was stirred 115.54, 73.56, 71.04, 68.60, 56.73, 56.41, 45.84, 45.61, 42.07,
for 3h. To the reaction mixture was added water, and aqueous 41.76, 40.56, 38.32, 36.53, 36.23, 30.09, 29.92, 29.79, 28.99,
layer was extracted with ethyl acetate until no product was 27.78, 25.96, 23.63, 22.23, 20.93, 18.91, 18.29, 12.15, 7.22, 6.89,
found in the aqueous layer, on TLC. The extracts were washed −4.59, −4.67. HR-MS ESI: m/z: Calcd for C45H76Na1O4Si2:
with brine, dried over MgSO4, filtered and concentrated in 759.5180 [M+Na]+. Found: 759.5170.
vacuo. The residue was chromatographed on silica gel column
(2% ethyl acetate–hexanes) to give ketone 19 (158mg, 64%).
Spectral Data for 19
(1R,3S,E)-5-[(E)-2-[(1R,7aR)-1-[(R)-6-Hydroxy-6-
methylheptan-2-yl]-7a-methylhexahydro-1H-inden-4(2H)-
ylidene]ethylidene]cyclohexane-1,3-diol (4a) To a solution
1H-NMR (400MHz, CDCl3) δ: 8.01 (2H, d, J=8.2Hz), of 20a (18mg, 24µmol) in MeOH (0.6mL) and THF (2.4mL)
7.54 (1H, t, J=7.3Hz), 7.42 (2H, t, J=7.8Hz), 5.29–5.22 (1H, was added K2CO3 (5mg, 36µmol) at room temperature under
m), 4.12–4.05 (1H, m), 2.81 (1H, dd, J=5.5, 13.5Hz), 2.64 argon, and the mixture was stirred overnight. To the reaction
(1H, dd, J=5.0, 15.2Hz), 2.59–2.53 (1H, m), 2.48–2.42 (2H, mixture was added saturated NH4Cl, and aqueous layer was
m), 2.08–2.00 (1H, m), 0.83 (9H, s), 0.03 (6H, s); 13C-NMR extracted with ethyl acetate for three times. The combined
(100MHz, CDCl3) δ: 205.67, 165.66, 133.27, 129.69, 129.78, organic layer was washed with water and brine, dried over
128.45, 68.15, 66.34, 50.61, 46.30, 39.50, 25.77, 18.07, −4.77, MgSO4, filtered and concentrated in vacuo. The residue was
−4.71. HR-MS ESI: m/z: Calcd for C19H28Na1O4Si: 371.1655 purified by silica gel column (10% ethyl acetate–hexanes) to
[M+Na]+. Found: 371.1655.
give alcohol, which was then dissolved in THF (1mL), and
(1S,3R,E)-3-[(tert-Butyldimethylsilyl)oxy]-5-[(E)-2- HF·Py (181mg, 0.16mL, 6.3mmol) was added, and the result-
[(1R,7aR)-7a-methyl-1-[(R)-6-methyl-6-[(triethylsilyl)oxy]- ing mixture was stirred for 24h at room temperature. To the
heptan-2-yl]hexahydro-1H-inden-4(2H)-ylidene]ethylidene]- reaction mixture was added saturated Na2S2O3, and aqueous
cyclohexyl Benzoate (20a) and (1R,3S,E)-3-[(tert-Butyl- layer was extracted with ethyl acetate for three times. The ex-
dimethylsilyl)oxy]-5-[(E)-2-[(1R,7aR)-7a-methyl-1-[(R)-6- tracts were washed with water and brine, dried over MgSO4,
methyl-6-[(triethylsilyl)oxy]heptan-2-yl]hexahydro-1H- filtered and concentrated in vacuo. The residue was first eluted
inden-4(2H)-ylidene]ethylidene]cyclohexyl Benzoate (20b) using 3% MeOH–CH2Cl2 on a silica gel column, and then

Vol. 64, No. 8 (2016)
Chem. Pharm. Bull.
1195
(2005).
again purified on a preparative TLC plate using 5% MeOH–
CH2Cl2 as the mobile phase to give 4a (5mg, 56%).
Spectral Data for 4a
7) Deluca H. F., BoneKEy Reports, 3, 514 (2014).
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hedron Lett., 31, 1823–1824 (1990).
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Smith C. M., DeLuca H. F., J. Med. Chem., 45, 3366–3380 (2002).
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H. F., Tetrahedron Lett., 32, 7663–7666 (1991).
1H-NMR (400MHz, CDCl3) δ: 6.23 (1H, d, J=10.9Hz), 5.86
(1H, d, J=11.5Hz), 3.91 (2H, m), 2.82 (1H, dd, J=10.3, 4.1Hz),
2.59 (1H, dd, J=11.5, 3.7Hz), 2.49 (1H, dd, J=13.3, 3.7Hz),
2.43–2.38 (1H, m), 2.28–2.23 (1H, m), 2.13–1.24 (27H, m),
1.22 (6H, s), 0.94 (3H, d, J=6.4Hz), 0.55 (3H, s); 13C-NMR
(100MHz, CDCl3) δ: 142.89, 130.21, 123.82, 115.48, 71.22,
68.92, 68.65, 56.61, 56.41, 45.87, 45.23, 44.50, 41.33, 40.59,
36.95, 36.48, 36.18, 29.46, 29.28, 28.99, 27.75, 23.57, 22.37,
20.89, 18.89, 12.16. HR-MS ESI: m/z: Calcd for C26H44Na1O3:
427.3188 [M+Na]+. Found: 427.3172.
(1R,3S, Z)-5-[(E)-2-[(1R,7aR)-1-[(R)-6-Hydroxy-6-
methylheptan-2-yl]-7a-methylhexahydro-1H-inden-4(2H)-
ylidene]ethylidene]cyclohexane-1,3-diol (4b) Similar to
20a, compound 20b (17mg, 24µmol) in MeOH (0.6mL) and
THF (2.4mL) was added K2CO3 (4.9mg, 36µmol) at room
temperature under argon, and the mixture was stirred over-
night. To the reaction mixture was added saturated NH4Cl,
and aqueous layer was extracted with ethyl acetate for three
times. The combined organic layer was washed with water
and brine, dried over MgSO4, filtered and concentrated in
vacuo. The residue was purified by silica gel column (10%
ethyl acetate–hexanes) to give alcohol, which was then dis-
solved in THF (1mL), and HF·Py (0.16mL) was added, and
the resulting mixture was stirred for 24h at room temperature.
To the reaction mixture was added saturated Na2S2O3, and
aqueous layer was extracted with ethyl acetate. The extracts
were washed with water and brine, dried over MgSO4, filtered
and concentrated in vacuo. The residue was first eluted using
3% MeOH–CH2Cl2 on a silica gel column, and then again pu-
rified on a preparative TLC plate using 5% MeOH–CH2Cl2 as
the mobile phase to give 4b (5.5mg, 59%).
15) Hanazawa T., Wada T., Masuda T., Okamoto S., Sato F., Org. Lett.,
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K., Ida K., Nagasawa K., Asian Journal of Organic Chemistry, 3,
994–999 (2014).
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J. W., Glorieux F. H., Elsevier Academic Press, New York, 2005,
pp. 1405–1422.
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Takayama H., Bioorg. Med. Chem., 8, 123–134 (2000).
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54, 4505–4508 (2013).
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3730–3735 (1994).
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Kishimoto S., Sugiura T., Waku K., Takayama H., Kittaka A., J.
Org. Chem., 68, 7407–7415 (2003).
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M., DeLuca H. F., J. Med. Chem., 49, 2909–2920 (2006).
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1175–1178 (1991).
Spectral Data for 4b
26) Cavallo A. S., Suffert J., Adib A., Solladié G., Tetrahedron Lett.,
31, 6649–6652 (1990).
1H-NMR (400MHz, CDCl3) δ: 6.34 (1H, d, J=10.9 Hz),
5.87 (1H, d, J=11.5Hz), 4.01 (2H, br), 2.83 (1H, dd, J=4.6,
10.3Hz), 2.58 (1H, dd, J=6.4, 13.5Hz), 2.32–2.24 (3H, m),
2.04–1.97 (3H, m), 1.91–1.84 (2H, m), 1.72–1.21 (23H, m), 0.94
(3H, d, J=6.4Hz), 0.55 (3H, s); 13C-NMR (100MHz, CDCl3)
δ: 142.89, 130.05, 124.14, 115.44, 71.22, 68.96, 68.77, 56.58,
56.39, 45.85, 45.14, 44.50, 40.56, 40.36, 36.76, 36.47, 36.20,
29.44, 29.31, 29.02, 27.78, 23.60, 22.37, 20.90, 18,89, 12.17.
HR-MS ESI: m/z: Calcd for C26H44Na1O3: 427.3188 [M+Na]+.
Found: 427.3196.
27) H-4β and H-10β (20a) shows large coupling constant, which
confirms the axial disposition of 1 and 3 protons to avoid 1,3-di-
axial interaction, similar situation can be found in case of H-4α and
H-10α (20b). Both compounds show NOE correlation between H-4α
and H-6, and H6-and H-9β, revealing the fact that each time H-3 is
placed at the same side to that of H-9β of CD ring. In this manner
stereochemistry of two isomers was deduced.
Conflict of Interest The authors declare no conflict of
interest.
References and Notes
1) “Vitamin D,” Chapter 86, ed. by Kudobera N., Feldman D., Pike J.
W., Glorieux F. H., Elsevier Academic Press, New York, 2005, pp.
152–154.
2) Fukumoto S., BoneKEy Reports, 3, 497 (2014).
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Physiol., 289, F8–F28 (2005).
4) Samuel S., Sitrin M. D., Nutr. Rev., 66 (Suppl. 2), S116–S124
(2008).
5) Philip J. L., Martin H., John S. A., Holick M. F., “Vitamin D: Physi-
ology Molecular Biology and Clinical Application,” Part II, Chapter
13, Humana Press, New York, 2010, pp. 297–310.
6) Nagpal S., Na S., Rathnachalam R., Endocr. Rev., 26, 662–687