Synthesis and vitamin D receptor affinity of 16-oxa vitamin D3 analogues

Article abstract of DOI:10.1039/c9ob02339a

Two novel 16-oxa-vitamin D₃ analogues were synthesized using a tandem Ti(ii)-mediated enyne cyclization/Cu-catalyzed allylation, Ru-catalyzed ring-closing metathesis reaction, and a low-valent titanium (LVT)-mediated stereoselective radical red

Full text of DOI:10.1039/c9ob02339a

Organic &
Biomolecular Chemistry
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Synthesis and vitamin D receptor affinity of 16-oxa
vitamin D analogues†
3
Cite this: Org. Biomol. Chem., 2019,
7, 10188
1
Kouta Ibe, Takeshi Yamada
and Sentaro Okamoto
*
Two novel 16-oxa-vitamin D
tion/Cu-catalyzed allylation, Ru-catalyzed ring-closing metathesis reaction, and a low-valent titanium
LVT)-mediated stereoselective radical reduction of 8α,14α-epoxide as the key steps for the synthesis
of the 16-oxa-C,D ring unit. The vitamin D receptor-binding affinity of the synthesized analogues,
6-oxa-1α,25-(OH) VD and 16-oxa-19-nor-1α,25-(OH) VD , was evaluated by fluorescence polarization
3
analogues were synthesized using a tandem Ti(II)-mediated enyne cycliza-
(
Received 29th October 2019,
Accepted 18th November 2019
1
2
3
2
3
DOI: 10.1039/c9ob02339a
vitamin D receptor competitor assay and time-resolved fluorescence energy transfer vitamin D receptor
rsc.li/obc
co-activator assay.
development of safer, more efficient, and more selective
vitamin D analogues remains an unmet need.
Introduction
1
α,25-Dihydroxyvitamin D3 [1α,25-(OH) VD , 1], a seco-ster-
Although a huge number of vitamin D analogues have been
2
3
oidal hormone and a potent metabolite of vitamin D (VD ), synthesized so far, most of the synthetic studies involve side
3
3
exhibits a broad spectrum of biological activities (Fig. 1). The chain and A-ring modifications such as 19-nor-1α,15-(OH)
regulation of calcium and phosphorus metabolism as well as (2),
2
VD
due to the difficulty in synthesizing a C,D-ring and its
bone remodeling are the most prominent physiological roles analogous structure.
3
1
0,11
1
,2
of 1. Moreover, 1 exhibits proliferating and differentiating
Many synthetic methods have been developed for the syn-
1
2
activities towards various types of tumor cells, while it regu- thesis of a C,D-ring. However, the C,D-ring intermediates
lates immune reactions.
3
,4
1 exerts its biological effects required for the synthesis of 1 in its natural form and its ana-
through its interaction with the vitamin D receptor (VDR), logues with a modified side chain and/or its A-ring counter-
5
which is a member of the nuclear receptor superfamily and part(s) have been synthesized by the oxidative degradation of
acts as a ligand-dependent gene transcription factor with co- natural vitamin D2 and D . Therefore, the syntheses of ana-
3
1
,6
activators.
The VDR-mediated transactivation involves a logues with modified C,D-ring skeletons have been less
1
3
ligand-binding process to the ligand-binding domain (LBD) of explored. Regarding the skeletal modifications of the C,D-
the apo-form of VDR, followed by a conformational change of ring, a heteroatom could be introduced in place of the C11,
the ligand–VDR complex into a transcriptionally active holo- C12, C15, or C16 carbon atom of the hydroindane ring, consid-
form, which binds to the co-activators and ultimately activates ering their chemical stability (steroid numbering). Most
7
the target gene.
3
The agonistic or antagonistic function of a VD –ligand
complex is controlled by the conformational change of the
ligand–VDR complex, in which helix 12, the very last
C-terminal R-helix of VDR that interacts with other proteins
8
,9
such as co-activators, plays a critical role.
Hypercalcemia
and hyperphosphatemia have been identified as serious side
effects during the therapeutic application of 1. Therefore, the
Department of Materials and Life Chemistry, Kanagawa University,
3
-27-1 Rokkakubashi, Kanagawa-ku, Yokohama 221-8686, Japan.
E-mail: okamos10@kanagawa-u.ac.jp
Electronic supplementary information (ESI) available: Copies of NMR ( H &
1
†
1
3
C) for compounds 3, 4, 6b, 7, 8, 9, 10b, 12, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23
and epi-23, 31, and 32. See DOI: 10.1039/c9ob02339a
2 3
Fig. 1 Natural 1α,25-(OH) VD and its analogues, synthesized in the
current study.
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specifically, an oxa analogue at the C16 position might be of
A novel 16-oxa type C,D-ring intermediate 6b was prepared
interest, because small variations of the C–O bond lengths, the following the procedure shown in Scheme 2. 1-Chloro-3-
D-ring size, and the bond angle between 16–17–20 atoms from methylbut-2-ene (11) and phenol were reacted through a
the corresponding values of the natural form would affect its Williamson reaction to yield the corresponding ether (80%),
biological properties by critically changing the conformation which was subjected to a SeO -catalyzed allylic oxidation with
2
of the ring system and/or the orientations of the side chain t-BuOOH (TBHP) (86%). Subsequent oxidation of the resulting
and the hydroxy moieties.
alcohol under the Parikh–Doering reaction conditions yielded
Herein, we report the synthesis of novel 16-oxa-vitamin D3 aldehyde 12 (65% through three steps). Evans’ asymmetric
analogues, wherein the synthesis of the 16-oxa-C,D-ring intermedi- aldol reaction between aldehyde 12 and oxazolidinone 13
17
2 3 3
ates was carried out using a tandem Ti(II)-mediated cyclization/ employing MgBr , Et N, and Me SiCl in ethyl acetate furn-
Cu-catalyzed allylation, Ru-catalyzed ring-closing metathesis, ished a diastereomeric mixture of adducts in a ratio of
and a low-valent titanium (LVT)-mediated stereoselective radical 95 : 4 : 1, from which the major isomer, 14, was isolated by
14
reduction of epoxide as the key steps of the synthesis. Moreover, column chromatography in 76% yield. The reduction of the
analogues 16-oxa-1α,25-(OH) VD (3) and 16-oxa-19-nor-1α,25- isolated 14 by LiBH4 gave the corresponding diol 15 (99%),
2
3
2 3
(OH) VD (4) (Fig. 1) were biologically investigated using VDR- which was converted to enyne 7 in 66% yield through a three-
binding assays and the preliminary results are described.
step process. First, a primary-selective silylation with i-Pr
3
SiCl
(
TIPSCl) and imidazole yielded 16 (86%). Propargylation of the
remaining secondary alcohol group of 16 with propargyl
bromide and NaH in the presence of a catalytic amount of
Results and discussion
n-Bu NI in THF resulted in intermediate 17 in 77% yield.
4
Synthesis of novel 16-oxa-VD analogues
3
Silylation of the terminal alkyne of 17 on treatment with
EtMgBr and then Me
3
SiCl gave 7 in 99% yield. A divalent tita-
nium alkoxide -mediated cyclization of enyne 7 was carried
out on treatment with Ti(O-i-Pr) and 2 i-PrMgCl in ether at
40 °C. The corresponding cyclized alkenyltitanium com-
The 16-oxa analogues 3 and 4 were synthesized through
Suzuki–Miyaura coupling of the A-ring unit 5 and the 16-oxa-C,
D-ring unit 6b (Scheme 1). The synthesis of A-ring units 5a
1
8
4
1
5
−
and 5b has been reported previously by our research team.
pound A was generated in situ and was further treated with
The 16-oxa-C,D-ring precursor 6b was prepared through a
Ti(II)-mediated cyclization of enyne 7, followed by an in situ
Cu-catalyzed allylation that yielded triene 8. A subsequent
ring-closing metathesis and a partial hydrogenation resulted
in compound 9.
allyl bromide in the presence of CuCN (cat.) to produce the
1
4
1
4
cyclized/allylated triene 8 as a single isomer in a high yield.
The relative stereochemistry was determined by performing
the nuclear Overhauser effect experiment on compound B,
which was generated after the protonation of the intermediate
Scheme 2 illustrates the synthetic procedures for the prepa-
ration of C,D-ring precursors 6. The natural type C,D-ring pre-
cursors, ketone 10a and alkenyl bromide 6a, were synthesized
organotitanium A with H O. Triene 8 reacted with a first-gene-
2
ration Grubbs ruthenium–carbene catalyst to undergo a ring-
1
9
1
6
closing metathesis (RCM) reaction and yielded the cyclized
cyclohex-1,4-diene derivative 18 (98%), which was selectively
hydrogenated at the less-substituted alkene position and
based on the existing literature: 10a was treated with phos-
phine ylide Ph PvCHBr to provide 6a (70%), which was then
coupled with 5a and 5b to yield 1α,25-(OH) VD (1) and 19-nor-
3
2
3
1
5
yielded 19 (99%). Protodesilylation of the SiMe
3
group under
1
α,25-(OH) VD (2), respectively.
2
3
acidic conditions (CF COOH) and subsequent removal of the
3
4
TIPS protecting group using n-Bu NF (59% in two steps)
yielded alcohol 20 (78%), which reacted with p-TsCl to produce
tosylate 9 (46% yield from 19). A side-chain was then intro-
duced to 9 through a copper-catalyzed reaction with a
Grignard reagent (87%). On treatment of 21 with
N-bromosuccinimide (NBS) in a H O/THF mixture, the corres-
2
ponding bromohydrin was obtained, which was further treated
under basic conditions to provide epoxide 22 in 55% yield.
The radical reduction of epoxide 22 was accomplished using a
method developed by our research group. In particular, 22 was
2
0
treated with a Ti(O-i-Pr)
4 3
/Me SiCl/Mg powder reagent in
1
,2-dimethoxyethane (DME)/CH Cl to produce a 3 : 1 mixture
2
2
of 23 and its epimer (epi-23) in a total 75% yield, which were
separated readily by column chromatography. The isolated
analogue 23 was oxidized on treatment with pyridinium
chlorochromate (PCC) to ketone 10b (93%), which was con-
verted to 6b (82%) following the procedures similar to
those used for the synthesis of 6a. In contrast to the prepa-
Scheme 1 General synthetic scheme for the preparation of analogues
, 3, and 4.
2
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+
−
Scheme 2 Synthesis of the 16-oxa-C,D-ring precursors 6a and 6b. Reagents: (a) Ph
3
P CH
, THF–H
, 2i-PrMgCl, ether; (k) cat. PhCHvRu(PCy
NF, THF; (o) p-TsCl, pyridine; (p) BrMgCH CH CMe OSiEt , cat. CuI–SMe
SiCl, Mg powder, THF; and (t) PCC, pyridine, CH Cl
2
Br Br , KHMDS, toluene; (b) PhOH, K
O; (g) (i-Pr) SiCl, imidazole, DMF; (h) propargyl
Cl , CH Cl ; (l) H (1 atm),
, THF; (q) NBS, THF–
2 3 3
CO , CH CN; (c)
TBHP, SeO
bromide, cat. n-Bu
5
2
, CH
2
Cl
2
; (d) SO
NI, NaH, THF; (i) EtMgCl, Me
wt% Pd/C, MeOH; (m) CF CO H, CHCl ; (n) n-Bu
O; (r) K CO , MeOH; (s) Ti(O-i-Pr) , Me
3
–py., Et
3
N, DMSO; (e) MgBr
2
, Et
3
N, Me
3
SiCl, AcOEt; (f) LiBH
4
2
3
4
3
SiCl, THF; ( j) Ti(O-i-Pr)
4
3
)
2
2
2
2
2
3
2
3
4
2
2
2
3
2
H
2
2
3
4
3
2
2
.
ration of 6a from 10a, halo-methylidenation of 16-oxa analogue trated as I and II (Scheme 3). Consequently, their reaction with
0b by a Wittig reaction resulted in a E/Z mixture in a ratio BH -THF, followed by oxidation with H , and their reaction
of 79 : 21. with m-CPBA gave the undesired alcohol 29 and β-epoxide 30,
1
3
2 2
O
Several model reactions were performed to determine the respectively. Moreover, the low-valent titanium-mediated
optimum reaction conditions (Scheme 2) to synthesize the radical reduction of 30 yielded compound 29. Meanwhile, the
desired analogues. The results are presented in Scheme 3. reaction of 26 with NBS under aqueous conditions provided
Using model compound C, which was prepared following the bromohydrins III and III′, which produced α-epoxide 27 after
synthetic procedure of 18 from the corresponding i-Pr substi- treatment with K
2 3
CO . The radical reduction of epoxide 27
2
0
tuted enyne, it was observed that the hydrogenation was with a Ti(O-i-Pr) /Me SiCl/Mg powder reagent provided a
4
3
affected by the solvent. Although the use of THF yielded a mixture of the desired alcohol 28 and its isomer epi-28. After
mixture of 24 and 25, to our delight the use of MeOH pro- screening various solvents and solvent concentrations, it was
duced partially reduced 24 in a quantitative yield.
Reaction of 3a-methylhexahydroindenes 9 and 26 with ceeded selectively to provide 28 in a satisfying yield. The rela-
electrophilic reagents such as BH -THF, m-CPBA, and NBS tive stereochemistries of 28 and epi-28 were confirmed by
proceeded on the convex face of the molecular structure, illus- the NOESY measurement of their acetates 28′ and epi-28′
found that the reaction in a DME/CH Cl mixture (v/v: 4/1) pro-
2 2
3
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Scheme 4 Synthesis of 16-oxa vitamin D analogues. Reagents: (a) cat.
PdCl
2 4
(dppf), 3 M KOH aq., THF, 60 °C; (b) n-Bu NF, THF; and (c) toluene,
110 °C.
PdCl (dppf) (8 mol%, dppf: 1,1′-bis(diphenylphosphino)ferro-
2
cene) in THF furnished the coupling products as
6 : 14 mixture of 7E/7Z isomers. The major isomer 31 was iso-
lated by column chromatography in 60% yield. 16-Oxa-
α,25-(OH) VD (3) was obtained in 92% yield after protodesi-
lylation of 31 by n-Bu NF. Similarly, 16-oxa-19-nor-1α,25-(OH) -
a
8
1
2
3
4
2
VD (4) was isolated by employing compounds 6b and 5b. The
3
obtained analogue 3 was stable in a refrigerator as well as at
ambient temperature, but it was isomerized slowly to the
corresponding pre-vitamin form (33) at 110 °C in heated
2
1
toluene (t1/2 = 12 h).
Molecular-docking study
Scheme 3 Results of model reactions.
A ligand–receptor protein docking study on novel 16-oxa ana-
logues 3, 4 and 1α,25-(OH) VD (1) and 19-nor-1α,25-(OH) VD (2)
2
3
2
3
22
was performed using the AutoDock Vina™ program, based on
(
Scheme 3). As described in a previous report, tertiary radical a reported crystal structure of VDR complexed with calcipotriol
23
IV, generated by a single electron transfer from Ti(III), predomi- (PDB: 1S19). The calculated overlapped models of 3 with 1 and
nantly absorbed a hydrogen atom in the intramolecular 4 with 2 are illustrated in Fig. 2. It was observed that the novel
pathway.
and intermolecular hydrogen absorption due to a difference in site similar to 1 and 2. However, they differed especially in the
the coordination manner to the titanium.
orientation of the side chain, presumably due to small differences
2
0b
The solvent(s) may thus affect the ratio of intra- 16-oxa-VD analogues 3 and 4 occupy the whole receptor-binding
3
With the 16-oxa-C,D-ring unit 6b in hand, we carried out between the C–O bond lengths, the size of the D-ring, and the
the synthesis of a series of 16-oxa-VD3 analogues by the bond angle between 16–17–20 atoms of the new compounds and
Suzuki–Miyaura coupling reaction with the A-ring units 5a and those of their natural form. Any interaction, such as hydrogen
1
5
5
b (Scheme 4). The reaction of 6b (a 79 : 21 mixture of 7E/7Z- bonding, between the introduced oxygen at the C16 position and
isomers) with 5a in the presence of KOH (3 M) and amino acid residue(s) in VDR was not observed.
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Fig. 2 Molecular-docking study based on a crystal structure of VDR in
complex with calcipotriol (PDB: 1S19): (a) calculated overlapped model
of 16-oxa analogues 3 (yellow) with natural 1α,25(OH)
blue) and (b) calculated overlapped model of analogue 4 (purple) with
9-nor-1α,25(OH) -VD (2) (light-blue).
2 3
-VD (1) (light-
1
2
3
VDR-binding assay
To evaluate the affinity of the synthesized 16-oxa-VD3 ana-
logues to VDR, fluorescence polarization vitamin D receptor
2
4
competitor assay
(PolarScreen™, Invitrogen) and time-
resolved fluorescence resonance energy transfer (TR-FRET)
2
4
VDR co-activator assay (LanthaScreen®, Invitrogen)‡ were
employed.
The vitamin D3 analogue bearing a fluorescence-tag exhi-
bits polarized fluorescence when it binds to VDR protein. The
potent VD3 analogues with fluorescence-tagged compounds
compete to bind with the VDR protein and decrease the polar-
ized fluorescence intensity depending on their binding
affinity. 1 μM VD3 analogues was incubated with a fluo-
rescence-tagged VD3 analogue-VDR complex and the results
are shown in Fig. 3(a). All compounds, 1, 2, 3, and 4, exhibited
competitive binding to VDR and decreased the polarized fluo-
rescence with ΔmP values of 222, 152, 119, and 101, respect-
ively. It was found that 16-oxa analogues, 3 and 4, are specific
ligands to VDR–LBD but their potency was relatively lower
than 1 and 2.
Meanwhile, the binding of VD
causes a conformational change around helix 12 in the LBD
3
analogues to the VDR–LBD
Fig. 3 (a) The results of the fluorescence polarization vitamin D recep-
tor competitor assay (PolarScreen™, Invitrogen). (b) The results of the
resulting in a higher affinity for the fluorescein-attached co- time-resolved fluorescence resonance energy transfer (TR-FRET)
VDR co-activator assay. Dose–response curve of analogues
1
activator peptide and thus allowing the fluorescence energy
transfer from the Tb complex of the receptor-tracer-antibody to
the fluorescence acceptor of the co-activator peptide. The
TR-FRET emission ratio at 520 and 495 nm (excitation:
(
red circle), 19-nor-1α,25-(OH) VD (blue square), 3 (orange triangle),
2
3
and 4 (purple triangle), as obtained using TR-FRET VDR co-activator
assay (LanthaScreen®, Invitrogen).
3
40 nm) was calculated and the EC50 values of the VD ana-
3
logues were determined by the dose–response curve. The assay
was performed in quintuplicate and, after eliminating the
highest and lowest points, the average value of the remaining
triplicate points was calculated. The dose–response curves of
the novel analogues’ VDR-binding potency are shown in
Fig. 3(b). It was demonstrated that 16-oxa-1α,25-(OH) VD (3)
with EC50 of 39.9 and 67.1 nM, respectively. However, they
were less potent than 1α,25-(OH)
2 3
VD (1) (EC50 = 16.2 nM) and
1
9-nor-1α,25-(OH) VD (2) (EC50 = 5.7 nM).
2
3
2
3
Conclusions
and 16-oxa-19-nor-1α,25-(OH) VD (4) are potent analogues
2 3
In summary, we have synthesized two novel 16-oxa-VD
analogues, 16-oxa-1α,25(OH) -VD (3) and 16-oxa-19-nor-1α,25
(OH) -VD (4). Their VDR-binding affinity was preliminarily
3
2
3
‡
See Invitrogen homepage: http://www.invitrogen.com
2
3
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evaluated using fluorescence polarization vitamin D receptor CH
2
Cl
2
(650 mL) was sequentially added an aqueous solution
competitor assay and TR-FRET VDR co-activator assay of tert-butylhydroperoxide (68 wt% in water, 72 mL, 576 mmol)
LanthaScreen®, Invitrogen). Their preparation involved and SeO (4.44 g, 40 mmol) at 0 °C. The resulting biphasic
(
2
multistep synthesis of the 16-oxa-C,D ring units through a mixture was stirred for 48 h at ambient temperature. Then,
Ti(II)-mediated cyclization of enyne 7, followed by an in situ methanol (400 mL) and NaBH4 were added until foaming
Cu-catalyzed allylation and a subsequent ring-closing meta- ceased and the reaction was checked by KI/starch paper. After
thesis. Moreover, α-epoxide 22 was selectively reduced through addition of water (1.0 L), the mixture was extracted with
a low-valent titanium-mediated radical process. It was also CHCl . The combined organic layers were washed with water
3
3 4
proven that the 16-oxa-VD analogues, 3 and 4, bind effectively and brine, dried over MgSO , filtered, and concentrated under
to the VDR, and their EC50 values, 39.9 and 67.1 nM, respect- reduced pressure. The residue was chromatographed on silica
ively, were calculated using TR-FRET VDR co-activator gel to give (E)-2-methyl-4-phenoxybut-2-en-1-ol (30.7 g) in 86%
assay. However, the binding activities of the novel analogues yield. To a mixture of (E)-2-methyl-4-phenoxybut-2-en-1-ol
were approximately 2.5–4 times less than that of natural (28.5 g, 160 mmol) and Et
3
N (67 mL, 480 mmol) in DMSO
1
α,25(OH) -VD (1). Research on further biological and (160 mL) was slowly added a pyridine–sulfur trioxide complex
2
3
pharmacological properties is in progress and the results will (51.0 g, 320 mmol) at 0 °C. After stirring for 30 min at room
be reported in the future.
temperature and then cooling to 0 °C, saturated aqueous
NaHCO3 was added. The mixture was extracted with
3
ether, washed with saturated aqueous NaHCO , aqueous 1 M
HCl, and brine, dried over MgSO , filtered, and concentrated.
The residue was purified by column chromatography on
silica gel (hexane/AcOEt) to give 12 (26.5 g) in 94% yield.
4
Experimental section
General procedures
1
NMR spectra were recorded in CDCl at 600 and 500 MHz for
3
3
H NMR (600 MHz, CDCl ) δ 9.49 (s, 1H, CH ̲O ), 7.32 (t, J =
1
13
H and 150 and 125 MHz for C on JEOL JNM-ECA600 and
ECA500 spectrometers, respectively. Chemical shifts are
reported in parts per million (ppm, δ) relative to Me Si (δ 0.00)
7
2
.6 Hz, 2H, Ar), 7.00 (t, J = 7.6 Hz, 1H, Ar), 6.92 (d, J = 7.6 Hz,
H, Ar), 6.70 (t, J = 5.5 Hz, 1H, olefinic), 4.88 (d, J = 5.5 Hz,
-
1
3
4
2H, allylic CH ), 1.84 (s, 3H, CCH
CDCl ) δ 194.1, 158.0, 147.8, 139.8, 129.7, 121.5, 114.6, 64.6,
9
̲
3); C NMR (150 MHz,
2
1
or residual CHCl
3
(δ 7.26 for H NMR) and CDCl
3
(δ 77.0 for
3
1
3
−
1
C NMR). IR spectra were recorded on a JASCO IR FT/IR 4100
.7; IR (neat) 1686, 1597, 1493, 1237, 1016, 753, 690 cm ;
+
spectrometer. High-resolution mass spectra (HR-MS) were
measured on a JEOL Accu TOF T-100 equipped with an ESI
ionization unit. All reactions sensitive to oxygen and/or moist-
ure were performed under an argon atmosphere. Dry solvents
HRMS m/z = calcd for C11
99.0750.
2
H12NaO [M + Na] : 119.0735, found
1
(R)-4-Benzyl-3-((2S,3R,E)-3-hydroxy-2,4-dimethyl-6-phenoxyhex-
[tetrahydrofuran (THF), N,N-dimethylformamide (DMF), di-
4-enoyl)oxazolidin-2-one (14)
chloromethane (CH Cl ), toluene, and diethyl ether (ether)]
2
2
were purchased from Kanto Chemicals. Ti(O-i-Pr)4 was dis-
tilled and stored under Ar. A solution of CuCN–2LiCl in THF
was prepared by mixing CuCN (10 mmol) and LiCl (20 mmol)
in THF (20 mL) and stored under Ar. 1st generation Grubbs
A mixture of oxazolidinone 13 (28.5 g, 122 mmol), MgBr2
(
26.8 g, 122 mmol), Et N (34 mL, 244 mmol), Me SiCl (23 mL,
3 3
183 mmol), and aldehyde 12 (25.8 g, 146 mmol) in AcOEt
(
300 mL) was stirred for 48 h at room temperature. The slurry
catalyst (PhCHvRuCl
chased from Sigma-Aldrich and used as received.
BrMgCH CH CMe OTES and EtMgBr were prepared from Mg
2 3 2 2
(PCy ) ) and PdCl (dppf) were pur-
was passed through a plug of silica gel with ether. The solution
was concentrated in vacuo and MeOH (600 mL) was added
2
2
2
along with CF COOH (7 mL). The solution was stirred for
3
turnings and the corresponding bromide in THF. i-PrMgCl was
prepared from i-PrCl and Mg turnings in ether and stored
under Ar.
3
0 min at room temperature and concentrated. The residue
was purified by column chromatography on silica gel (hexane/
AcOEt) to give 14 (38.0 g) in 76% yield. H NMR (600 MHz,
1
CDCl ) δ 7.33 (t, J = 7.2 Hz, 2H, Ar), 7.29–7.23 (m, 5H, Ar), 6.94
3
(
E)-2-Methyl-4-phenoxybut-2-enal (12)
(
t, J = 7.2 Hz, 1H, Ar), 6.88 (d, J = 7.8 Hz, 2H, Ar), 5.80 (t, J = 6.3
To a mixture of 1-chloro-3-methylbut-2-ene (11) (112 mL, Hz, 1H, olefinic), 4.71–4.68 (m, 1H, NCH), 4.61 (d, J = 6.0 Hz,
.0 mol) and K CO (277 g, 2.0 mol) in CH CN (500 mL) was 2H, allylic CH ), 4.25 (d, J = 8.4 Hz, 1H, CHOH), 4.22–4.15 (m,
), 3.32 (dd, J = 3.0, 13.8 Hz, 1H,
mixture was stirred for 12 h at room temperature. After benzyl), 2.88–2.73 (m, 1H, NC(O)CHCH ), 2.76 (dd, J = 9.6,
removal of the solvent under reduced pressure, aqueous 1 M 13.8 Hz, 1H, benzyl), 1.81 (s, 3H, allylic CH ), 1.12 (d, J = 7.2
̲
1
̲
2
3
3
2
2
slowly added phenol (97 mL, 1.1 mol) at 0 °C. The resulting 3H, C(O)OCH̲ and CHOH̲
̲
3
3
1
3
NaOH (200 mL) was added. The mixture was extracted with Hz, 3H, CHCH
hexane, washed with aqueous 1 M NaOH, dried over MgSO 153.7, 139.1, 135.3, 129.4, 128.9, 127.3, 124.3, 120.8, 114.6,
and filtered through a pad of Celite. The filtrate was concen- 80.3, 66.1, 64.2, 55.6, 40.3, 37.7, 14.8, 11.7; IR (KBr) 3493,
3
3
̲ ); C NMR (150 MHz, CDCl ) δ 176.4, 158.5,
4
,
−
1
trated and chromatographed on silica gel to give ((3-methyl- 2977, 1778, 1697, 1598, 1494, 1385, 1215 cm ; HR-MS m/z =
+
but-2-en-1-yl)oxy)benzene (130 g) in 80% yield. To a solution of calcd for C24
H
27NNaO
5
[M + Na] : 432.1787, found 432.1766;
2
5
(
(3-methylbut-2-en-1-yl)oxy)benzene (32.4 g, 200 mmol) in [α] −35.8 (c 0.55, CHCl3).
D
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Organic & Biomolecular Chemistry
(
2R,3R,E)-2,4-Dimethyl-6-phenoxyhex-4-ene-1,3-diol (15)
To a solution of 14 (30.6 g, 74.6 mmol) in THF–MeOH (40 : 1
v/v, 124 mL) was added in small portions a solution of LiBH
1.9 g, 90 mmol) in THF (90 mL) at 0 °C. After 5 h of stirring at
room temperature, CH Cl (100 mL) was added. The mixture
trated. The residue was purified by column chromatography
on silica gel (hexane/ether) to give 17 (19.9 g) in 77% yield. H
1
NMR (600 MHz, CDCl
t, J = 7.8 Hz, 1H, Ar), 6.91 (d, J = 8.4 Hz, 2H, Ar), 5.69 (t, J = 6.0
Hz, 1H, olefinic), 4.66 (dd, J = 6.0, 12.6 Hz, 1H, allylic CH2),
3
) δ 7.28 (dd, J = 7.8, 8.4 Hz, 2H, Ar), 6.95
4
(
(
2
2
4
1
.61 (dd, J = 5.4, 12.6 Hz, 1H, allylic CH
5.6 Hz, 1H, propargylic CH ), 3.87 (dd, J = 2.4, 15.6 Hz, 1H,
2OSi), 3.77
OSi), 3.70 (d, J = 9.6, Hz, 1H,
), 2.34 (t, J = 2.1 Hz, 1H, alkyne H), 1.85–1.80 (m, 1H,
), 1.65 (s, 3H, allylic CH ), 1.12–1.03 (m, 21H, i-Pr), 0.82
2
), 4.05 (dd, J = 2.4,
was washed with 5% aqueous citric acid and brine. The com-
bined organic layers were dried over Na SO , filtered, and con-
centrated. The residue was purified by column chromato-
graphy on silica gel (hexane/AcOEt) to give 15 (17.5 g) in 99%
yield. H NMR (600 MHz, CDCl ) δ 7.27 (t, J = 8.3 Hz, 2H, Ar),
2
2
4
propargylic CH ), 3.80 (dd, J = 5.4, 9.6 Hz, 1H, CH
̲
2
(
dd, J = 3.0, 9.6 Hz, 1H, CH
CH OCH
CH CH
̲
2
1
2
̲
3
̲
3
3
6
.95 (t, J = 7.2 Hz, 1H, Ar), 6.90 (d, J = 7.8 Hz, 2H, Ar), 5.70 (t,
1
3
(
d, J = 7.2 Hz, 3H, CH
3 3
̲ CH); C NMR (150 MHz, CDCl )
J = 5.9 Hz, 1H, olefinic), 4.61 (dd, J = 6.6, 12.6 Hz, 1H, allylic
CH ), 4.57 (dd, J = 6.0, 12.6 Hz, 1H, allylic CH ), 3.95 (d, J = 8.4
δ 158.6, 136.8, 129.4, 126.4, 120.8, 114.7, 85.4, 80.2, 73.6, 64.6,
2
2
6
1
C
4.4, 55.5, 37.9, 18.2, 13.8, 12.6, 11.2; IR (neat) 3309, 2942,
Hz, 1H, CH
CH2OH), 3.21 (brs, 1H, OH
H, CH
CH CH); C NMR (150 MHz, CDCl
̲
OH), 3.73–3.71 (m, 1H, CH
̲
2
OH), 3.66–3.62 (m, 1H,
−1
600, 1496, 1462, 1239, 1070 cm ; HR-MS m/z = calcd for
̲
̲
̲
+
26
H
42NaO
3
Si [M + Na] : 453.2801, found 453.2801.
1
̲
CH ), 1.74 (s, 3H, allylic CH ), 0.74 (d, J = 7.2 Hz, 3H,
3
3
1
3
̲
3
3
) δ 158.5, 140.5, 129.4,
(
7R,8R)-11,11-Diisopropyl-2,2,8,12-tetramethyl-7-((E)-4-
1
3
23.4, 120.8, 114.7, 83.8, 67.9, 64.3, 37.1, 13.7, 11.5; IR (KBr)
phenoxybut-2-en-2-yl)-6,10-dioxa-2,11-disilatridec-3-yne (7)
−
1
367, 2941, 1751, 1596, 1496, 1240, 1012 cm ; HR-MS m/z =
+
To a solution of 17 (20.6 g, 47.7 mmol) in THF (95 mL) was
added EtMgBr (48 mL, 1.48 M in THF, 71.5 mmol) at 0 °C.
calcd for C14
H
20NaO
3
[M + Na] : 259.1310, found 259.1287;
2
5
[α] −18.4 (c 0.80, CHCl3).
D
After being stirred for 1 h at room temperature, Me
3
SiCl
(
9.0 mL, 71.5 mmol) was added and the mixture was gradually
(
4
2R,3R,E)-2,4-Dimethyl-6-phenoxy-1-((triisopropylsilyl)oxy)hex-
-en-3-ol (16)
To a mixture of diol 15 (13.2 g, 56.0 mmol) and imidazole
warmed to room temperature over 2 h. Water (200 mL) was
added and the organic layer was separated. The aqueous layer
was extracted with ether. The combined organic layers were
washed with brine, dried over MgSO
trated. The residue was chromatographed on silica gel
(8.4 g, 123 mmol) in DMF (140 mL) was added i-Pr SiCl
3
4
, filtered, and concen-
(11.8 mL, 56.0 mmol) at room temperature. After stirring for
3
h, saturated aqueous NaHCO
3
at 0 °C was added to the
1
(
(
hexane/ether) to give 7 (23.7 g) in 99% yield. H NMR
600 MHz, CDCl ) δ 7.28 (t, J = 7.6 Hz, 2H, Ar), 6.95 (t, J = 7.6
mixture. The mixture was extracted with ether, dried over
MgSO , filtered, and concentrated. The residue was chromato-
graphed on silica gel (hexane/AcOEt) to give 16 (18.9 g) in 86%
yield. H NMR (600 MHz, CDCl ) δ 7.27 (dd, J = 7.8, 9.0 Hz, 2H,
3
4
Hz, 1H, Ar), 6.91 (d, J = 7.6 Hz, 2H, Ar), 5.66 (t, J = 6.0 Hz, 1H,
olefinic), 4.65 (dd, J = 6.0, 12.6 Hz, 1H, allylic CH ), 4.61 (dd,
2
1
3
J = 5.4, 12.6 Hz, 1H, allylic CH
pargylic CH ), 3.89 (d, J = 15.8 Hz, 1H, propargylic CH
dd, J = 3.0, 9.6 Hz, 1H, CH2OSi), 3.74 (dd, J = 6.0, 9.6 Hz, 1H,
CH OSi), 3.62 (d, J = 9.6 Hz, 1H, CH OCH), 1.87–1.81 (m, 1H,
CH CH), 1.65 (s, 3H, allylic CH ), 1.13–1.03 (m, 21H, i-Pr), 0.82
d, J = 6.6 Hz, 3H, CH CH), 0.16 (s, 9H, SiCH
2
), 4.04 (d, J = 15.8 Hz, 1H, pro-
Ar), 6.93 (t, J = 7.6 Hz, 1H, Ar), 6.90 (d, J = 7.6 Hz, 2H, Ar), 5.73
2
2
), 3.81
(t, J = 6.0 Hz, 1H, olefinic), 4.63 (dd, J = 6.0, 12.6 Hz, 1H, allylic
(
̲
CH ), 4.60 (dd, J = 6.0, 12.6 Hz, 1H, allylic CH ), 4.32 (d, J = 2.4
Hz, 1H, OH
2
2
̲
2
̲
̲
), 3.96 (dd, J = 2.4, 8.4 Hz, 1H, CH
̲O H), 3.90 (dd, J =
3
̲
3
3
.9, 10.2 Hz, 1H, CH
CH2OSi), 1.95–1.88 (m, 1H, CH
.16–1.05 (m, 21H, i-Pr), 0.76 (d, J = 7.2 Hz, 3H, CH
̲
2
OSi), 3.69 (dd, J = 7.8, 10.2 Hz, 1H,
13
(
(
1
1
̲
̲
); C NMR
3
3
̲
̲
3
3
3
150 MHz, CDCl
3
) δ 158.7, 136.9, 129.6, 126.3, 120.9, 114.8,
1
1
̲
3
CH);
C
02.3, 90.3, 86.0, 64.9, 64.5, 56.6, 38.0, 18.2, 13.9, 12.0,
NMR (150 MHz, CDCl
1
3
3
) δ 158.7, 140.3, 129.4, 123.1, 120.6,
−1
1.3, −0.16; IR (neat) 2959, 2336, 1560, 1495, 1250 cm
;
14.7, 83.4, 69.2, 64.4, 37.3, 17.9, 17.8, 13.6, 11.7; IR (neat)
+
HR-MS m/z = calcd for C29
found 525.3198.
H
50NaO
3
Si
2
[M + Na] : 525.3196,
−
1
443, 2943, 1560, 1496, 1462, 1239 cm ; HR-MS m/z = calcd
+
for C23H40NaOSi [M + Na] : 415.2644, found 415.2621.
Triisopropyl((R)-2-((2R,3S,Z)-3-methyl-4-(1-(trimethylsilyl)but-
(
((2R,3R,E)-2,4-Dimethyl-6-phenoxy-3-(prop-2-yn-1-yloxy)hex-4-
en-1-yl)oxy)triisopropylsilane (17)
To a mixture of NaH (7.5 g, 60 wt% in oil, 186 mmol) and 17.6 mmol) in ether (67 mL) was added dropwise i-PrMgCl
n-Bu NI (TBAI, 1.1 g, 3.1 mmol) in THF (150 mL) was added a (22.0 mL, 1.62 M in ether, 35.2 mmol) at −40 °C. The reaction
3-en-1-ylidene)-3-vinyltetrahydrofuran-2-yl)propoxy)silane (8)
To a mixture of 7 (6.8 g, 13.5 mmol) and Ti(O-i-Pr) (5.2 mL,
4
4
solution of 16 (23.6 g, 60 mmol) in THF (50 mL) at 0 °C. After mixture was stirred for 30 min at this temperature and then
stirring for 1 h at room temperature and then cooling to 0 °C, allowed to warm to 0 °C over 1.5 h. Allyl bromide (13.5 mL, 2.0
propargyl bromide (14.0 mL, 186 mmol) was added dropwise. M in THF, 27.0 mmol) and CuCN–2LiCl (5.4 mL, 0.5 M in
The mixture was allowed to warm to room temperature and THF, 2.7 mmol) were added at 0 °C and the mixture was
stirred for 3 h. Water (200 mL) was slowly added and the stirred for 3 h. After addition of water (0.5 mL), Celite (5 g),
mixture was extracted with ether. The organic layer was and NaF (5 g), the mixture was stirred for 30 min at room
washed with brine, dried over MgSO , filtered, and concen- temperature, and then filtered through a pad of Celite with
4
10194 | Org. Biomol. Chem., 2019, 17, 10188–10200
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Paper
ether. The filtrate was concentrated and the residue was puri- CH
fied by column chromatography on silica gel (hexane/ether) to 1.71–1.60 (m, 1H, CH
3
CH
̲
2
), 1.79 (dt, J = 12.6, 3.6 Hz, 1H, CH
2
3
̲ CCH ),
̲
CH C), 1.21 (dt, J = 5.4, 12.6 Hz, 1H,
2
2
1
give 8 (5.9 g) in 97% yield. H NMR (600 MHz, CDCl
dd, J = 10.8, 17.4 Hz, 1H, CH vCHC), 5.77–5.71 (m, 1H, (d, J = 7.2 Hz, 3H, CH
CH vCHCH ), 5.13 (d, J = 18.6 Hz, 1H, CH vCHCH ), 5.10 (150 MHz, CDCl ) δ 153.9, 125.3, 88.4, 68.4, 66.4, 42.7, 37.5,
3
) δ 5.95 CH̲ CCH ), 1.23–1.02 (m, 21H, i-Pr), 1.00 (s, 3H, CH̲ C), 0.98
2
3
3
1
3
(
2
̲
3 3 3
̲ CH), 0.06 (s, 9H, Si(CH̲ ) ); C NMR
̲
̲
2
2
2
2
3
(
1
4
CH
6
3
1
d, J = 11.7 Hz, 1H, CH
2
vCHCH
vCHC), 4.92 (dd, J = 1.8, 17.4 Hz, 1H, CH
.50 (d, J = 13.2 Hz, 1H, CH2OCH), 4.17 (d, J = 12.6 Hz, 1H, C H NaO Si [M + Na]: 447.3091, found 447.3099.
OCH), 3.81 (dd, J = 3.0, 9.0 Hz, 1H, CH
.6, 9.6 Hz, 1H, CH OSi), 3.37 (d, J = 9.6 Hz, 1H, CH
.21 (dd, J = 3.0, 16.2 Hz, 1H, allylic CH ), 2.92 (dd, J = 5.4,
2
), 4.98 (dd, J = 1.8, 10.2 Hz, 32.7, 27.1, 18.5, 18.1, 18.0, 13.1, 12.0, −1.0; IR (neat) 2942,
2
̲
vCHC), 1463, 1384, 1259, 1052, 837 cm− ; HR-MS m/z = calcd for
1
H, CH
̲
2
̲
24
48
2
2
̲
2
̲
OSi), 3.69 (dd, J =
OCH),
(
1
R)-2-((1R,7aS)-7a-Methyl-1,3,5,6,7,7a-hexahydroisobenzofuran-
-yl)propan-1-ol (20)
), 1.24 A mixture of 19 (5.2 g, 12.2 mmol) and CF
C), 1.22–1.05 (m, 21H, i-Pr), 0.90 (d, J = 6.6 Hz, 3H, 61.0 mmol) in CHCl (61 mL) was stirred for 3 h at 50 °C, and
) ); C NMR (150 MHz, CDCl ) δ then cooled to room temperature and concentrated. To a solu-
2
2
̲
2
6.2 Hz, 1H, allylic CH
2
), 1.89–1.84 (m, 1H, CH
3
CH
̲
2
3
COOH (4.7 mL,
(
s, 3H, CH
CH3CH), 0.08 (s, 9H, Si(CH3 3
58.0, 144.5, 138.4, 128.4, 115.2, 113.7, 88.5, 71.2, 66.1, 52.3, tion of the residue in THF (12 mL) was added n-Bu
7.1, 34.7, 18.0, 17.0, 14.2, 12.0, −0.06; IR (neat) 2942, 1635, (15.0 mL, 1.0 M in THF, 15.0 mmol). After stirring for 1 h at
̲
3
3
1
3
̲
̲
3
1
3
1
4
NF
−
1
463, 1249, 1071 cm ; HR-MS m/z = calcd for C H NaO Si
M + Na] : 473.3247, found 473.3239; [α] +14.8 (c 0.97, added, and then the mixture was extracted with ether. The
room temperature, saturated aqueous NH Cl (15 mL) was
2
6
50
2
2
4
+
25
D
[
CHCl
3
).
organic layer was dried over MgSO
4
, filtered, concentrated and
chromatographed on silica gel (hexane/ether) to give 20
Triisopropyl((R)-2-((1R,7aS)-7a-methyl-4-(trimethylsilyl)-
,3,5,7a-tetrahydroisobenzofuran-1-yl)propoxy)silane (18)
A mixture of 8 (10.5 g, 23.3 mmol) and 1st generation Grubbs (dq, J = 12.0, 1.8 Hz, 1H, CH
Ru-catalyst [(benzylidene)bis(tricyclohexylphosphine)ruthe- 1H, CHCH OH), 3.56 (dd, J = 3.0, 10.8 Hz, 1H, CHCH
nium(IV) dichloride, 0.575 g, 0.70 mmol] in dry CH Cl 3.23 (bs, 1H, OH), 3.17 (d, J = 9.6 Hz, 1H, CHOCH ), 2.08–1.96
120 mL) was stirred for 12 h at room temperature. The (m, 3H, allylic CH and CH CHCH ), 1.76 (dt, J = 12.6, 3.6 Hz,
1
(
1
1.41 g) in 59% yield. H NMR (600 MHz, CDCl
H, olefinic), 4.32 (dq, J = 12.0, 3.0 Hz, 1H, CH
2OCH), 3.63 (dd, J = 9.0, 10.8 Hz,
OH),
3
) δ 5.45 (brs,
1
̲
2
OCH), 4.16
̲
2
̲
2
2
2
̲
̲
2
(
̲
2
3
2
mixture was filtered through a pad of silica gel and concen- 1H, CH
trated. The residue was purified by column chromatography 12.6 Hz, 1H, CH
on silica gel (hexane/ether) to give 18 (9.7 g) in 98% yield. H 3H, CH
2
CCH
3
), 1.72–1.64 (m, 2H, CH
CCH ), 1.04 (s, 3H, CH
̲ CH); C NMR (150 MHz, CDCl ) δ 143.5, 117.5, 94.5,
2
CH
2
C), 1.24 (dt, J = 4.2,
C), 0.83 (d, J = 7.2 Hz,
2
3
3
1
3
3
3
NMR (600 MHz, CDCl ) δ 6.00 (dd, J = 3.0, 9.6 Hz, 1H, olefinic), 69.5, 68.7, 42.5, 36.2, 32.9, 24.3, 18.4, 17.7, 12.7; IR (neat)
.86 (ddd, J = 1.8, 5.4, 9.6 Hz, 1H, olefinic), 4.50 (dd, J = 3.6, 3439, 2934, 1462, 1378, 1248, 1040 cm ; HR-MS m/z = calcd
−
1
5
1
CH
6
2
2
1
+
25
3.8 Hz, 1H, CH
OCH), 3.90 (dd, J = 3.0, 9.0 Hz, 1H, CH
.6, 9.0 Hz, 1H, CH OSi), 3.31 (d, J = 10.2 Hz, 1H, CH
.72 (dd, J = 5.4, 21.0 Hz, 1H, allylic CH ), 2.62 (dquint, J =
2̲ OCH), 4.22 (dd, J = 3.0, 13.8 Hz, 1H, for C H NaO [M + Na] : 219.1361, found 219.1363; [α] +8.6
12 20 2
D
2
̲
OSi), 3.67 (dd, J = (c 1.01, CHCl
OCH),
3
).
2
2
̲
(
1
R)-2-((1R,7aS)-7a-Methyl-1,3,5,6,7,7a-hexahydroisobenzofuran-
-yl)propyl 4-methylbenzenesulfonate (9)
2
2
), 1.92–1.87 (m, 1H, CH
3 2
CH ̲C H ),
̲
3
CH), 0.99 (s, 3H, CH
̲
3
C), 0.09 To a solution of 20 (0.84 g, 4.3 mmol) in pyridine (21 mL) was
) ); C NMR (150 MHz, CDCl ) δ 154.9, 132.0, added p-TsCl (1.2 g, 6.4 mmol) at 0 °C. The mixture was stirred
26.9, 123.2, 84.3, 68.2, 66.1, 44.6, 37.7, 29.2, 19.1, 18.0, 13.4, for 24 h at room temperature and then AcOEt (30 mL) was
added. The resulting suspension was washed with aqueous
HR-MS m/z = calcd for C H NaO Si [M + Na] : 445.2934, 1 M HCl, water, and saturated aqueous NaHCO . The organic
1
(s, 9H, Si(CH
̲
3
3
3
1
1
−
1
2.0, −1.3; IR (neat) 2960, 1731, 1627, 1463, 1249, 1012 cm
;
+
2
4
46
2
2
3
2
found 445.2945; [α]D −26.3 (c 1.43, CHCl
3
).
layer was dried over MgSO
4
, filtered and concentrated. The
residue was chromatographed on silica gel (hexane/ether) to
Triisopropyl((R)-2-((1R,7aS)-7a-methyl-4-(trimethylsilyl)-
,3,5,6,7,7a-hexahydroisobenzofuran-1-yl)propoxy)silane (19)
1
give 9 (1.17 g) in 78% yield. H NMR (600 MHz, CDCl ) δ 7.79
3
1
(
d, J = 7.8 Hz, 2H, Ar), 7.33 (d, J = 7.8 Hz, 2H, Ar), 5.39 (s, 1H,
To a solution of 18 (3.8 g, 9.0 mmol) in MeOH (90 mL) was olefinic), 4.28 (dq, J = 12.6, 3.0 Hz, 1H, CH OCH), 4.20 (dd, J =
added 5% Pd/C (383 mg). Under a H atmosphere (1 atm) the 3.0, 9.0 Hz, 1H, CH2OS), 4.02 (dd, J = 6.6, 9.0 Hz, 1H, CH2OS),
suspension was stirred for 12 h at room temperature and fil- 3.70 (d, J = 12.0 Hz, CH OCH), 2.97 (d, J = 9.6 Hz, 1H,
tered through a pad of Celite. The filtrate was concentrated. CH OCH), 2.44 (s, 3H, Ar–CH ), 2.06–1.94 (m, 3H, allylic CH
The residue was purified by column chromatography on silica and CH CHCH ), 1.74–1.62 (m, 3H, CH CH CCH ), 1.21 (dt,
̲
2
̲
̲
2
̲
2
2
̲
̲
3
2
̲
̲
̲
3
2
2
2
3
1
gel (hexane/AcOEt) to give 19 (3.79 g) in 99% yield. H NMR J = 3.6, 12.0 Hz, 1H, CH
600 MHz, CDCl ) δ 4.37 (dt, J = 12.6, 3.3 Hz, 1H, CH OCH), J = 7.2 Hz, 3H, CH CH); C NMR (125 MHz, CDCl
.18 (dt, J = 12.6, 2.4 Hz, 1H, CH2OCH), 3.89 (dd, J = 3.6, 9.0 144.1, 133.1, 129.7, 128.0, 117.2, 88.0, 74.0, 68.3, 42.0, 34.8,
Hz, 1H, CH OSi), 3.64 (dd, J = 7.2, 9.6 Hz, 1H, CH OSi), 3.02 32.9, 24.2, 21.6, 18.4, 17.4, 12.9; IR (neat) 2937, 1598, 1359,
d, J = 9.6 Hz, 1H, CH OCH), 2.11 (app d, J = 17.4 Hz, 1H, 1189, 949, 847, 664 cm ; HR-MS m/z = calcd for C19
CH2CSi), 1.99–1.92 (m, 1H, CH2
2
CH3̲
2
CCH
3
), 0.96 (s, 3H, CH
3
C), 0.95 (d,
) δ 144.4,
1
(
4
3
̲
̲
3
3
̲
̲
2
̲
2
−
1
(
2
̲
H
26NaO
4
S
+
25
̲
̲ CSi), 1.90–1.83 (m, 1H, [M + Na] : 373.1450, found 373.1426; [α] +9.3 (c 1.36, CHCl3).
D
This journal is © The Royal Society of Chemistry 2019
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Organic & Biomolecular Chemistry
Triethyl(((R)-2-methyl-6-((1R,7aS)-7a-methyl-1,3,5,6,7,7a-
(150 MHz, CDCl
3
) δ 89.6, 73.5, 71.6, 68.1, 55.3, 45.3, 41.4, 34.8,
hexahydroisobenzofuran-1-yl)heptan-2-yl)oxy)silane (21)
34.4, 30.0, 29.8, 27.8, 21.1, 20.6, 15.2, 14.9, 14.8, 7.1, 6.8; IR
−
1
(
neat) 2954, 1459, 1380, 1235, 1042, 741 cm ; HR-MS m/z =
To
a solution of 4-bromo-2-methyl-2-triethylsiloxybutane
+
calcd for C23
H
44NaO
3
Si [M + Na] : 419.2957, found 419.2984;
(
(
1.4 g, 5.0 mmol) in THF (5.0 mL) was added Mg turnings
0.36 g, 15 mmol) at room temperature. After stirring for
2
2
[
α] +29.4 (c 1.00, CHCl3).
D
3
0 min at this temperature, a solution of CuI–SMe
2
(0.27 g,
(
1R,3aS,4R,7aS)-7a-Methyl-1-((R)-6-methyl-6-((triethylsilyl)oxy)
1
.3 mmol) in THF (2.0 mL) was added. To this was added a
heptan-2-yl)octahydroisobenzofuran-4-ol (23) and its epimer
epi-23
solution of 9 (0.35 g, 1.0 mmol) in THF (10 mL) at −10 °C.
After being stirred for 1 h at this temperature, aqueous NH Cl/
NH OH buffer (pH 7–8, 20 mL) was added. The mixture was
extracted with AcOEt, and the combined extracts were washed
with saturated aqueous NaHCO , brine, dried over MgSO and
concentrated. The residue was purified by column chromato-
4
To a mixture of epoxide 22 (376 mg, 0.95 mmol), Ti(O-i-Pr)
4
4
(
0.34 mL, 1.14 mmol), and Mg powder (69 mg, 2.85 mmol) in
DME–CH Cl (1 : 4 v/v, 4.8 mL) was added Me SiCl (0.14 mL,
.14 mmol) at room temperature. After being stirred for 24 h
2
2
3
3
4
1
at room temperature, saturated aqueous NaHCO was added
3
graphy on silica gel (hexane/AcOEt) to give 21 (0.33 g) in 87%
yield. H NMR (600 MHz, CDCl ) δ 5.39 (s, 1H, olefinic), 4.39
1
and the mixture was stirred for 30 min. After addition of Celite
and NaF, the mixture was stirred for 30 min and filtered
through a pad of Celite. The filtrate was concentrated and puri-
fied by column chromatography on silica gel (hexane/AcOEt)
to provide 23 (210 mg) in 56% yield and epi-23 (71 mg) in 19%
yield. Compound 23: H NMR (600 MHz, CDCl ) δ 4.02 (t, J =
3
(
dq, J = 12.0, 3.6 Hz, 1H, CH
̲
2OCH), 4.11 (dq, J = 12.0, 1.8 Hz,
OCH), 2.92 (d, J = 9.6 Hz, 1H, CH OCH), 2.06–1.95 (m,
H, allylic CH ), 1.80 (dt, J = 12.6, 3.6 Hz, 1H, CH CH2CCH3),
1
2
1
3
H, CH
2
2
̲
̲
2
2
.74–1.65 (m, 4H, CHCH CH
̲
and CH
̲
C(CH ) ), 1.50–1.33 (m,
2
2
2
3 2
1
3
H, CH
3
CH
̲
CH
2
and CH
3
CHCH
2
CH
̲
2
), 1.29–1.20 (m, 2H,
7
.5 Hz, 1H, CH
CH CHOH), 3.61 (dd, J = 7.2, 11.4 Hz, 1H, CH
J = 10.2 Hz, 1H, CH OCH), 2.07–2.03 (m, 1H, CH
CH CCH and CH COSi), 1.52–1.34
), 1.31–1.14 (m, 2H, CH CHCH and
COSi), 1.18 (s, 3H, CH COSi), 1.17 (s, 3H, CH COSi),
̲
OCH), 3.76 (dt, J = 4.8, 10.8 Hz, 1H,
OCH), 3.06 (d,
CHOH),
CH CH
̲
CCH and CH CHCH̲2), 1.18 (s, 3H, CH̲3COSi), 1.17 (s,
3
2
2
3
3
2
̲
̲
2
3
H, CH
̲
COSi), 1.13–1.05 (m, 1H, CH CHCH
̲
2), 1.02 (s, 3H,
), 0.86 (d, J = 6.6 Hz,
3
̲
̲
2
CH
H, CH̲ CH), 0.55 (q, J = 7.8 Hz, 6H, Si(CH̲ CH ) ); C NMR
3 2 3 3
̲
3
C), 0.93 (t, J = 7.8 Hz, 9H, Si(CH
2
CH
̲
3
)
3
13
1.78–1.60 (m, 7H, CH
m, 3H, CH CHCH CH
CH
̲
2
CH
̲
2
̲
2
3
2
3
(
3
̲
2
̲
2
3
̲
2
(150 MHz, CDCl ) δ 145.3, 116.6, 92.6, 73.5, 68.2, 45.4, 42.1,
3
̲
̲
̲
2
3
3
3
4.8, 34.6, 33.2, 30.0, 29.7, 24.3, 21.1, 18.5, 17.4, 14.8, 7.1, 6.8;
−
1
1.08–1.02 (m, 1H, CH
(CH CH ), 0.84 (s, 3H, CH
CH
3
CHCH
̲
2
IR (neat) 2950, 1460, 1235, 1044, 1016, 742, 723 cm ; HR-MS
m/z = calcd for C H NaO Si [M + Na] : 403.3008, found
+
2
̲
3
)
3
3
2
3
44
2
1
3
2
3
3
CH), 0.55 (q, J = 7.8 Hz, 6H, OSi(CH
150 MHz, CDCl ) δ 92.5, 73.5, 68.7, 67.0, 56.4, 45.3, 44.2, 36.1,
5.1, 35.0, 34.5, 30.0, 29.7, 21.7, 21.1, 14.5, 11.7, 7.1, 6.8; IR
2
4
3
03.3029; [α]D +20.9 (c 1.00, CHCl ).
(
3
3
Triethyl(((R)-2-methyl-6-((1aR,4aR,5R,7aR)-4a-
methylhexahydro-1aH-oxireno[2,3-d]isobenzofuran-5-yl)
heptan-2-yl)oxy)silane (22)
−1
(
neat) 3381, 2935, 2874, 1462, 1038, 1016, 742, 723 cm ;
+
HR-MS m/z = calcd for C23
H
46NaO
3
Si [M + Na] : 421.3114,
24
D
found 421.3112; [α] −9.9 (c 1.01, CHCl
3
). Compound epi-23:
1
To a solution of 21 (0.48 g, 1.25 mmol) in THF (3.8 mL) and
H NMR (600 MHz, CDCl ) δ 3.98 (quint, J = 4.8 Hz, 1H,
3
water (0.8 mL) was added NBS (0.49 g, 2.75 mmol) at 0 °C. The CH
reaction mixture was slowly warmed to 10 °C. After being Hz, 1H, CH
2
CH
̲
̲
2
OCH), 3.92 (t, J = 8.9
OCH ), 2.32 (dt,
2
̲
2
̲
2
aqueous NaHCO
AcOEt, washed with brine, dried over MgSO
3
was added. The mixture was extracted with CH
, filtered, and con- CH
2
CH
̲
2
̲
2
2
̲
2
4
3
CH
̲
2
̲
2
̲
2
J =
centrated under reduced pressure. A mixture of the residue CH CH
̲
̲
2
2
3
3
and K
2
CO
3
(0.48 g, 3.5 mmol) in MeOH (14 mL) was stirred for C(CH
̲
3
)
2
), 1.07 (s, 3H, CH
CH
organic layer was washed with brine, dried over MgSO , fil- 7.8 Hz, 6H, Si(CH
̲
3
3
h at room temperature and extracted with AcOEt. The (CH
2
̲
3
)
3
̲
3
2
1
3
̲
4
2
3 3
3
tered, and concentrated under reduced pressure. The residue 73.4, 68.9, 67.3, 49.8, 45.4, 44.7, 34.8, 34.7, 33.0, 30.1, 29.9,
was purified by column chromatography on silica gel (hexane/ 29.8, 21.8, 20.1, 19.6, 17.7, 7.1, 6.8; IR (neat) 3409, 2955, 2875,
1
−1
AcOEt) to give 22 (0.27 g) in 55% yield. H NMR (600 MHz, 1459, 1044, 742, 723 cm
;
HR-MS m/z
=
calcd for
24
+
CDCl
Hz, 1H, CH
3
) δ 4.16 (d, J = 10.2 Hz, 1H, CH
̲
2
OCH), 3.53 (d, J = 10.2
CHOC), 3.35 (d, J (c 1.03, CHCl
), 2.05–1.98 (m, 1H, OCHCH2),
), 1.76–1.69 (m, 3H, OCHCH CH
CCH and
), 1.32–1.10 (m, 2H, CH CHCH CH ), 1.19 To a suspension of PCC (16 mg, 74 μmol) and Celite (16 mg)
C
23
H
46NaO
3
Si [M + Na] : 421.3114, found 421.3140; [α]D +32.4
̲
2
OCH), 3.43 (t, J = 1.8 Hz, 1H, CH
2
̲
3
).
=
1
9.6 Hz, 1H, CH OCH
.88–1.83 (m, 1H, OCHCH
̲
̲
2
(
1R,3aS,7aS)-7a-Methyl-1-((R)-6-methyl-6-((triethylsilyl)oxy)
heptan-2-yl)hexahydroisobenzofuran-4(1H)-one (10b)
2
2
2
̲ ,
and CH
̲
2
CCH
3
), 1.60–1.36 (m, 6H, CH
̲
2
3
CH CHCH CH
̲
̲
CH
̲
̲
3
2
2
2
3
2
2
(
0
s, 3H, CH
.94 (t, J = 7.8 Hz, 9H, OSi(CH
CH CH) 0.56 (q, J = 7.8 Hz, 6H, OSi(CH
̲
3
COSi), 1.18 (s, 3H, CH
̲
3
COSi), 1.06 (s, 3H, CH
̲
3 2 2
C), in CH Cl (0.37 mL) was added pyridine (6.0 μL, 74 μmol) at
2
CH
̲ ) ), 0.84 (d, J = 6.6 Hz, 3H, room temperature. After being stirred for 30 min, the mixture
3 3
13
̲
̲
CH ) ); C NMR was cooled to 0 °C and a solution of 23 (14.6 mg, 37 μmol) in
3
2
3 3
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Paper
1
3
2 2 2 3 3 3
CH CH (0.37 mL) was added. After stirring for 3 h at room 0.56 (q, J = 7.8 Hz, 6H, Si(CH̲ CH ) ); C NMR (CDCl ,
temperature, the solid material was filtered off. The solvent 150 MHz) δ 141.2, 101.4, 92.4, 73.4, 69.0, 54.0, 45.4, 35.5, 34.6,
was removed under reduced pressure. The residue was purified 33.2, 30.0, 29.8, 27.1, 21.6, 21.2, 19.9, 17.4, 7.1, 6.8; IR (neat,
−
1
by flash column chromatography on silica gel (hexane/AcOEt) ATR) 2931, 2873, 1459, 1380, 1219, 1040, 772, 722 cm
;
1
+
to give 10b (13.6 mg) in 93% yield. H NMR (600 MHz, CDCl ) HR-MS m/z = calcd for C H BrNaO Si [M + Na] : 495.2270,
3
24 45
2
δ 3.87 (dd, J = 8.4, 10.2 Hz, 1H, CHOCH
2
), 3.82 (t, J = 8.4 Hz, found 495.2295.
1
7
2
H, CHOCH
.8, 10.8 Hz, 1H, C(O)CH
.14–2.09 (m, 1H, C(O)CH
CH CH ), 1.74–1.59 (m, 3H, CH
̲
2
), 3.24 (d, J = 10.2 Hz, 1H, CH
̲
2
), 2.93 (dd, J =
2),
), 2.06–1.93 (m, 2H, C(O) To a mixture of bromide C,D-ring (6b) (10 mg, 21 μmol),
CCH and CH COSi), aqueous 3 M KOH (7 μL) and THF (0.2 mL) was added drop-
2), 1.31–1.22 (m, 1H, wise a mixture of boronate A-ring (5a) (13 mg, 25 μmol) and
COSi), 1.18 (s, 3H, CH COSi), PdCl (dppf) (1.2 mg, 1.7 μmol) at room temperature. The
), 0.94 (t, J = 7.8 Hz, 9H, resulting mixture was stirred for 37 h at 60 °C. After addition
3)3), 0.87 (d, J = 6.6 Hz, 3H, CH3CH), 0.82 (s, 3H, of ether, the mixture was washed with aqueous 1 M HCl and
CCH), 0.56 (q, J = 7.8 Hz, 6H, Si(CH CH
CDCl , 150 MHz) δ 208.4, 91.9, 73.4, 62.8, 60.8, 49.0, 45.3, residue was purified by column chromatography on silica gel
2 3
Bis-tert-butyldimethylsilyl ether of 16-oxa-1α,25-(OH) VD (31)
̲
̲
2
2
CH̲
2
CH
2
̲
2
̲
2
̲
2
3
̲
2
1
.52–1.34 (m, 3H, CH CH
̲
̲
3
2
CH
3
CHCH
̲
2
3
2
1
.11–1.03 (m, 1H, CH
3 2
CHCH̲
Si(CH CH
CH
(
̲
̲
2
1
3
̲
3
2
3 3 4
) ); C NMR brine, dried over MgSO , and concentrated in vacuo. The
3
1
4
0.6, 35.1, 34.8, 34.6, 30.0, 29.7, 24.0, 21.0, 14.7, 12.2, 7.1, 6.8; (hexane/AcOEt) to give 31 (9.6 mg) in 60% yield. H NMR
−1
IR (neat) 2953, 2876, 1720, 1382, 1038, 742, 724 cm ; HR-MS (CDCl
3
, 600 MHz) δ 6.20 (d, J = 10.8 Hz, 1H, olefinic), 5.83 (d,
Si [M + Na] : 419.2957, found J = 10.8 Hz, 1H, olefinic), 5.17 (s, 1H, CH vC), 4.82 (s, 1H,
), 4.21–4.16 (m, 1H,
), 3.80 (t, J = 7.5 Hz, 1H, CHOCH ), 3.69 (dd, J = 7.2,
), 3.13 (d, J = 9.6 Hz, 1H, CHOCH ), 2.83
+
m/z = calcd for C23
19.2975.
H
44NaO
3
̲
2
4
CH
̲
vC), 4.37–4.35 (m, 1H, CH CH̲
2
2
CH
2
CH
̲
̲
2
Triethyl(((R)-6-((1R,3aR,7aS)-4(bromomethylene)-7a-
methyloctahydroisobenzofuran-1-yl)-2-methylheptan-2-yl)oxy)
11.4 Hz, 1H, CHOCH
̲
2
̲
2
(
(
dd, J = 4.2, 12.6 Hz, 1H, allylic), 2.47–2.42 (m, 2H, allylic), 2.22
dd, J = 7.2, 13.2 Hz, 1H, allylic), 1.89–1.60 (m, 8H,
silane (6b)
+
Br)Br− (74 mg, 0.17 mmol) and dry CH
To a mixture of Ph
3
P (CH
2
̲
2
CH
̲
2
CH
2
CCH
̲
2
O, CH
CH CH2), 1.18 (s, 3H, C(CH
), 1.08–1.03 (m, 1H, CH CHCH ), 0.94 (t, J = 7.8,
CH ), 0.873 (s, 9H, SiC(CH ), 0.870 (s, 9H, SiC
) ), 0.84 (d, J = 6.6 Hz, 3H, CH CHCH ), 0.71 (s, 3H,
̲
2
CO, and CHCH
2
CH), 1.50–1.20
toluene (0.34 mL) was added KHMDS (1.9 M in toluene, 81 μL, (m, 5H, CH CHCH
̲
̲
̲
3)3), 1.17 (s,
3
2
0
.15 mmol) at room temperature and the resulting mixture 3H, C(CH
was stirred for 1 h at this temperature. After cooling to 0 °C, to 9H, Si(CH
this was added a solution of 10b (13.5 mg, 34 μmol) and the (CH
3
)
3
3
̲
2
2
̲
3
)
3
3 3
̲ )
̲
̲
3
3
3
2
mixture was stirred for 30 min at 0 °C. The reaction mixture CH
was quenched with saturated aqueous NH Cl. The mixture was (CH
filtered through a short pad of silica gel and washed with 3H, Si(CH3 2
̲
3
CCH), 0.56 (q, J = 7.8 Hz, 6H, Si(CH
), 0.06 (s, 3H, Si(CH ), 0.05 (s, 3H, Si(CH
) ); C NMR (CDCl , 150 MHz) δ 148.3, 136.5,
̲
2
CH
3
)
3
), 0.07 (s, 3H, Si
4
3
)
2
̲
3
)
2
3 2
̲ )
), 0.04 (s,
1
3
̲
3
hexane. After concentration in vacuo, the residue was purified 133.1, 122.4, 118.9, 111.3, 107.0, 102.8, 92.8, 73.5, 71.8, 67.5,
by column chromatography on silica gel (hexane/ether) to give 65.2, 55.2, 45.9, 45.3, 45.2, 44.8, 36.6, 35.3, 35.0, 30.1, 29.7,
a mixture of (E)-6b and (Z)-6b (10 mg, 21 μmol, E : Z = 79 : 21) 28.3, 25.8, 25.7, 22.9, 21.2, 18.2, 18.1, 14.8, 11.6, 7.1, 6.8, −4.7,
1
in total 62% yield. (E)-6b: H NMR (600 MHz, CDCl
3
) δ 5.54 (s, −4.8, −5.1; IR (neat, ATR) 2954, 2875, 1471, 1380, 1256, 1085,
−
1
1
7
2
1
CH
CH
H, olefinic), 3.84 (t, J = 7.2 Hz, 1H, CHOCH
.2, 11.4 Hz, 1H, CHOCH2), 3.12 (d, J = 9.6 Hz, 1H, CH
.91–2.89 (m, 1H, allylic), 2.47 (dd, J = 7.8, 10.2 Hz, 1H, allylic),
.86–1.84 (m, 1H, CHCH O), 1.78–1.60 (m, 5H, CCH CH CH
CO), 1.50–1.33 (m, 5H, CH CHCH CH CH2), 1.18 (s, 3H, To a solution of 31 (13.7 mg, 18 μmol) in THF (0.18 mL) was
COSi), 1.09–1.02 (m, 1H, added n-Bu NF (1.0 M in THF, 63 μL, 63 μmol). After stirring
), 0.94 (t, J = 7.8 Hz, 9H, Si(CH CH ), 0.85 (d, J = for 24 h at room temperature, the solvents were removed
CH), 0.73 (s, 3H, CH CCH), 0.56 (q, J = 7.8 Hz, under reduced pressure. The residue was diluted with H O
̲
2 4 3
), 3.66 (dd, J = 835, 775 cm ; HR-MS m/z = calcd for C44H84NaO Si [M +
+
̲
̲O CH ), Na] : 783.5575, found 783.5595.
2
1
6-Oxa-1α,25-(OH) VD (3)
2
3
̲
2
̲
2
̲
2
2
,
̲
̲
̲
̲
̲
2
3
2
2
3
COSi), 1.17 (s, 3H, CH
CHCH
̲
3
4
CH
3
̲
2
2
3 3
̲ )
6
6
9
2
1
.6 Hz, 3H, CH
̲
̲
3
3
2
1
3
H, Si(CH
2.2, 73.4, 64.6, 55.0, 45.3, 44.9, 35.9, 35.1, 34.9, 30.5, 30.0, bined organic layers were dried over MgSO
9.7, 22.1, 21.1, 14.8, 11.5, 7.1, 6.8; IR (neat, ATR) 2929, 2855, purified by column chromatography on silica gel (hexane/
2
CH
3
)
3
); C NMR (150 MHz, CDCl
3
) δ 141.3, 97.9, (1 mL) and then extracted with AcOEt (4 × 1 mL). The com-
4
, concentrated and
−
1
1
3
459, 1363, 1219, 1038, 772, 743 cm ; HR-MS m/z = calcd for AcOEt) to give 3 (6.7 mg) in total 92% yield. H NMR (CDCl ,
+
C
24
H
45BrNaO
2
Si [M + Na] : 495.2270, found 495.2251. (Z)-6b: 600 MHz) δ 6.34 (d, J = 11.4 Hz, 1H, olefinic), 5.84 (d, J = 10.8
2vC), 4.96 (s, 1H, CH vC),
1
H NMR (600 MHz, CDCl ) δ 5.96 (brs, 1H, olefinic), 3.83 (t, J = Hz, 1H, olefinic), 5.32 (s, 1H, CH
̲
̲
3
2
9
(
.0 Hz, 1H, CHOCH
d, J = 6.6 Hz, 1H, CH
.05–2.00 (m, 1H, CH
CCH CH CH , CH CHCH CH
.16–1.11 (m, 1H, CH CHCH
J = 7.8 Hz, 9H, Si(CH CH3)3), 0.91 (d, J = 6.6 Hz, 3H, CH
̲
2
), 3.72 (t, J = 9.0 Hz, 1H, CHOCH
OCH ), 2.68–2.61 (m, 2H, allylic), CH
CH O), 1.74–1.22 (m, 11H, 11.4 Hz, 1H, CHOCH
̲
2
), 3.35 4.43 (dd, J = 4.2, 7.8 Hz, 1H, CH
CH), 3.79 (t, J = 7.5 Hz, 1H, CHOCH
), 3.15 (d, J = 9.6 Hz, 1H, CH̲
2
CH̲
̲
2
2
̲
̲
2
2
̲
̲
2
2
2
̲
2
̲
2
2
3
̲
̲
2
̲
2
CH
̲
2
), 1.19 (s, 6H, (CH
CCH), 0.94 (t, allylic), 2.46 (dd, J = 7.2, 10.8 Hz, 1H, allylic), 2.32 (dd, J = 6.0,
3CH), 13.2 Hz, 1H, allylic), 2.07–2.03 (m, 1H, allylic), 1.91–1.82 (m,
3
2
̲ ) COSi), (dd, J = 4.2, 12.6 Hz, 1H, allylic), 2.58 (dd, J = 3.0, 13.8 Hz, 1H,
1
3
̲
2
), 1.02 (s, 3H, CH
̲
3
̲
̲
2
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2
H, allylic, CCH
̲
CH
2
), 1.77–1.61 (m, 6H, CCH
̲
2
CH
̲
2
CH
2
,
2.51–2.45 (m, 2H, allylic), 2.23–2.15 (m, 2H, allylic), 1.99–1.94
CH CO), 1.51–1.25 (m, 5H, CH CH
̲
̲
̲
̲
2), 1.21 (s, 3H, (m, 1H), 1.86–1.70 (m, 4H), 1.67–1.40 (m, 10H), 1.30–1.25 (m,
), 1.15–1.06 (m, 1H, CH CHCH ), 1H), 1.22 (s, 3H, CH COH), 1.21 (s, 3H, CH COH), 1.16–1.09
); C NMR (m, 1H, CH CHCH ), 0.86 (d, J = 6.6 Hz, 3H, CH CHCH ), 0.72
CDCl , 150 MHz) δ 147.7, 138.4, 134.5, 124.1, 118.1, 111.6, (s, 3H, CH C); C NMR (CDCl , 150 MHz) δ 138.3, 132.6,
2
3
2
C(CH
0
(
3
)
2
), 1.20 (s, 3H, C(CH
3
)
2
3
̲
2
̲
3
̲
3
1
3
.85 (d, J = 6.6 Hz, 3H, CHCH
3
), 0.72 (s, 3H, CCH
̲
3
3
̲
3
2
̲
3
2
1
̲
3
3
3
9
3
3
7
4
2.6, 71.1, 70.4, 66.8, 65.1, 55.2, 45.2, 45.1, 44.1, 42.8, 36.4, 123.0, 116.4, 92.65, 71.1, 67.3, 67.1, 65.2, 55.2, 45.1, 44.5, 44.1,
5.0, 34.9, 29.4, 29.0, 28.5, 23.0, 21.0, 14.9, 11.7; IR (neat, ATR) 42.1, 37.2, 36.4, 35.1, 34.9, 29.5, 29.1, 28.3, 22.9, 21.1, 14.9,
366, 2927, 2854, 1746, 1648, 1457, 1376, 1065, 1026, 911, 11.7; IR (neat, ATR) 3365, 2928, 2872, 1456, 1377, 1214, 1041,
40 cm− ; HR-MS m/z = calcd for C26
1
H
42NaO
+
−1
+
4
4
Si [M + Na] : 938, 754 cm ; HR-MS m/z = calcd for C25H42NaO [M + Na] :
41.2981, found 441.2984.
429.2981, found 429.3009.
Bis-tert-butyldimethylsilyl ether of 16-oxa-19-nor-
α,25-(OH) VD (32)
Evaluation of the VDR-binding affinity of 16-oxa-VD3
analogues
1
2
3
To a mixture of bromide C,D-ring (6b) (10 mg, 21 μmol), VDR-binding affinity of 16-oxa-VD analogues were evaluated
3
aqueous 3 M KOH (7 μL) and THF (0.2 mL) was added drop- using fluorescence polarization (FP) VDR competitor assay
wise a mixture of boronate A-ring (5b) (13 mg, 25 μmol) and (PolarScreen™, Invitrogen) and time-resolved fluorescence
PdCl (dppf) (1.2 mg, 1.7 μmol) at room temperature. The resonance energy transfer (TR-FRET) VDR co-activator assay.
2
resulting mixture was stirred for 37 h at 60 °C. After addition Each assay was performed under in vitro conditions in 384-well
of ether, the mixture was washed with aqueous 1 M HCl and black polystyrene plates (Corning® 384-well Low Volume Black
brine, dried over MgSO , and concentrated in vacuo. The Round Bottom Polystyrene NBS Microplate (Corning 3676))
4
residue was purified by column chromatography on silica gel using a total volume of 40 μL per well.
1
(
hexane/AcOEt) to give 32 (8.1 mg) in 52% yield. H NMR
Fluorescence polarization (FP) VDR competitor assay
PolarScreen™, Invitrogen)
(
CDCl , 600 MHz) δ 6.14 (d, J = 11.4 Hz, 1H, olefinic), 5.63 (d,
3
(
J = 11.4 Hz, 1H, olefinic), 4.10–4.02 (m, 2H, CH
7
3
̲
.5 Hz, 1H, CHOCH
̲
2
̲
), Vitamin receptor (VDR), Fluormone™ VDR Red
D
3
̲
allylic), 2.47 (dd, J = 7.8, 11.4 Hz, 1H, allylic), 2.39–2.33 (m, 2H, purchased from Invitrogen (PolarScreen Vitamin D Receptor
allylic), 2.20 (dd, J = 1.8, 13.2 Hz, 1H, allylic), 2.10 (dd, J = 8.4, Competitor Assay). VD analogues were dissolved in dimethyl
3.2 Hz, 1H, allylic), 1.86–1.36 (m, 15H, CHCH2CH, CvCCH sulfoxide (DMSO, for molecular biology, Sigma Aldrich) and
CH CH CH CvC and CHCH CH CH ), 1.19 (s, 3H, C(CH ), diluted with VDR Red Assay Buffer with 1% DMSO to arbitrary
.18 (s, 3H, C(CH ), 1.09–1.03 (m, 1H, CH CHCH ), 0.94 (t, concentrations. A receptor–tracer complex was added to VD
J = 7.8 Hz, 9H, Si(CH CH3)3), 0.88 (s, 9H, Si(CH3)3), 0.85 (s, solutions to achieve final concentrations of 1.4 nM for VDR,
), 0.84 (d, J = 7.2 Hz, 1H, CHCH ), 0.72 (s, 3H, 2.0 nM for Fluormone and 1.0 μM for VD analogues in a total
C), 0.56 (q, J = 8.4 Hz, 6H, Si(CH CH ), 0.06 (s, 3H, volume of 20 μL per well. This mixture was incubated for 2 h at
Si(CH3)2), 0.05 (s, 3H, Si(CH ) ), 0.03 (s, 6H, Si(CH ) ); room temperature. FP was measured on an Infinite® F200 Pro
3
1
̲
̲,
̲
2
̲
2
̲
2
2
̲
2
̲
2
3 2
̲ )
1
3
)
2
3
̲
2
3
̲
̲
2
9
H, Si(CH
̲
3
)
3
3
3
CH
3
2
3 3
)
̲
̲
̲
3 2
3 2
1
3
3
C NMR (CDCl , 150 MHz) δ 136.2, 135.0, 121.1, 117.2, 92.8, microplate reader (Tecan) using a 535 nm excitation filter
7
3
1
2
3.5, 68.1, 67.9, 65.2, 55.2, 46.1, 45.3, 45.0, 43.6, 36.8, 36.6, (25 nm bandwidth) and a 590 nm emission filter (20 nm band-
5.3, 35.0, 30.1, 29.7, 28.1, 25.9, 25.8, 22.8, 21.2, 18.13, 18.07, width). Finally, the ΔmP value for VD analogues was calcu-
3
4.7, 11.7, 7.1, 6.8, −4.7, −4.8, −4.9, −4.9; IR (neat, ATR) 2954, lated to compare with the mP value of the VDR-fluorescent
−
1
856, 1462, 1252, 1085, 1028, 836 cm ; HR-MS m/z = calcd for ligand complex.
+
C H NaO Si [M + Na] : 771.5575, found 771.5546.
4
3
84
4
3
Time-resolved fluorescence resonance energy transfer
(TR-FRET) VDR co-activator assay
1
6-Oxa-19-nor-1α,25-(OH) VD (4)
2
3
To a solution of 32 (10.3 mg, 14.0 μmol) in THF (0.14 mL) was VDR ligand-binding domain tagged with GST (VDR–LBD
added n-Bu NF (1.0 M in THF, 70.0 μL, 70.0 μmol) at 0 °C. (GST)), fluorescein-TRAP220/DRIP-2 co-activator peptide (fluor-
4
After stirring for 48 h at room temperature, the solvent was escein-peptide), LanthaScreen® Tb-anti-GST (Goat) antibody
removed under reduced pressure. The residue was diluted with (Tb-anti-GST), TR-FRET co-regulator buffer G and DTT solution
H
2
O (1 mL) and then extracted with AcOEt (4 × 1 mL). The were purchased from Invitrogen (LanthaScreen® TR-FRET
combined organic layers were dried over MgSO , concentrated VDR Coactivator Assay Kit). VD analogues were dissolved in
and chromatographed on silica gel (hexane/AcOEt) to give 4 dimethyl sulfoxide (DMSO, for molecular biology, Sigma
4
3
1
(
5.4 mg) in 95% yield. H NMR (CDCl
3
, 600 MHz) δ 6.28 (d, J = Aldrich) and diluted with TR-FRET co-regulator buffer G with
1.1 Hz, 1H, olefinic), 5.67 (d, J = 11.1 Hz, 1H, olefinic), 1% DMSO to arbitrary concentrations. A receptor–tracer–anti-
.16–4.12 (m, 1H, CHOH), 4.05 (sep, J = 4.1 Hz, 1H, CHOH), body complex was added to VD solutions to achieve final con-
.86 (t, J = 7.2 Hz, 1H, CH OCH), 3.75 (dd, J = 7.8, 11.4 Hz, 1H, centrations of 1.0 nM for VDR–LBD (GST), 2.0 nM for Tb-anti-
OCH), 3.16 (d, J = 9.6 Hz, 1H, CH OCH), 2.82 (dd, J = 4.2, GST and 100 nM for fluorescein-peptide in a total volume of
2.6 Hz, 1H, allylic), 2.71 (dd, J = 3.6, 13.2 Hz, 1H, allylic), 20 μL per well. This mixture was incubated for 2 h at room
1
4
3
CH
1
̲
̲
3
2
̲
2
2
̲
10198 | Org. Biomol. Chem., 2019, 17, 10188–10200
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temperature. TR-FRET was measured on an Infinite® F200
PRO microplate reader (Tecan) using a 340 nm excitation filter
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Y. Fall, Tetrahedron Lett., 2017, 58, 3568–3570;
There are no conflicts to declare.
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This work was partly supported by the JSPS KAKENHI grant
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