Preparation of (S)-γ-cyclogeraniol by lipase-catalyzed transesterification and synthesis of (+)-trixagol and (+)-luffarin-P

Article abstract of DOI:10.1016/j.molcatb.2015.11.019

Lipase-catalyzed kinetic resolution of γ-cyclogeraniol by Candida antarctica lipase B yielded 23% of enantiomerically pure (S)-γ-cyclogeraniol. (+)-trixagol and (+)-luffarin-P were synthesized from the obtained (S)-γ-cyclogeraniol, and the absolute config

Full text of DOI:10.1016/j.molcatb.2015.11.019

Journal of Molecular Catalysis B: Enzymatic 123 (2016) 160–166
Contents lists available at ScienceDirect
Journal of Molecular Catalysis B: Enzymatic
journal homepage: www.elsevier.com/locate/molcatb
Preparation of (S)-␥-cyclogeraniol by lipase-catalyzed
transesterification and synthesis of (+)-trixagol and (+)-luffarin-P
Mikio Fujiia,b,∗, Yosuke Morimotoa, Machiko Onob, Hiroyuki Akitaa
a Faculty of Pharmaceutical Sciences, Toho University, 2-2-1 Miyama, Funabashi, Chiba 247-8510, Japan
b School of Pharmacy, International University of Health and Welfare, 2600-1 Kitakanemaru, Ohtawara, Tochigi 324-8501, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Lipase-catalyzed kinetic resolution of ␥-cyclogeraniol by Candida antarctica lipase B yielded 23% of enan-
tiomerically pure (S)-␥-cyclogeraniol. (+)-trixagol and (+)-luffarin-P were synthesized from the obtained
(S)-␥-cyclogeraniol, and the absolute configuration of natural (+)-luffarin-P was determined to have an
S configuration by our first synthesis of (S)-luffarin-P for the first time.
Received 24 August 2015
Received in revised form
17 November 2015
Accepted 17 November 2015
Available online 22 November 2015
© 2015 Elsevier B.V. All rights reserved.
Keywords:
Lipase CAL-B
␥-Cyclogeraniol
Lufferin-P
Lactone
Trixagol
1. Introduction
the direct enantiomeric resolution of 4 has several advantages
including avoiding racemization during the derivation of enan-
Several natural products possess ␥-cyclogeranyl unit, includ-
ing (+)-trixagol 1, [1] (+)-luffarin-P 2 [2] and (S)-(+)-luffarin-W
3 [2] (Fig. 1). Compounds 1 and 2 are precursors of antibacte-
rial terpenes such as agelasines, manoalide, and lufferins, which
has prompted us to study their synthesis [2–4]. Enantiomatically
enriched ␥-cyclogeraniol (4) is considered to be an impor-
tant chiral building block for the synthesis of these products.
Thus facile preparation methods of enantiomerically active 4 are
desired [5–8]. Fehr and Galindo reported two methods to pre-
pare enantiomatically enriched 4 (Scheme 1) [5]: enantiomeric
resolution by sequential recrystallization of ␥-cyclogeranic acid
and asymmetric protonation of ␣-cyclogeranic acid phenylth-
ioester using N-isopropylephedrin. Previously, lipase-catalyzed
enantiomeric resolution of rac-4 was tested; however, the results
were unsatisfactory [6]. As an alternative method, lipase-catalyzed
transesterification of ␤-hydroxyester was conducted (Scheme 1).
The enantiomatically enriched ␤-hydroxyester and its acetate were
converted to the corresponding enantiomerically active 4 [6]. While
preparation methods of enantiomerically active 4 were developed,
tiomerically active 4 from the chiral compounds and reducing the
conversion steps necessary to produce enantiomerically active 4. In
this study, we report the lipase-catalyzed enantiomeric resolution
of rac-4 and the syntheses of (S)-1 and (S)-2 from (S)-4.
2. Experimental
2.1. General
The NMR spectra were recorded on JEOL AL-400 spectrome-
ter. Mass spectra were measured by JEOL JMS-AM II 50. IR spectra
were carried out on JASCO FT/IR 4100. Specific rotation was mea-
sured on JASCO P-2200. HPLC analysis was performed on SSC-3210
pump equipped with chiral column, SSC-5200 UV detector and
SIR Chromatocorder 21. Chemicals were purchased from Tokyo
Chemical Industry Co. Ltd., Wako Pure Chemicals Industry, Ltd. or
Sigma–Aldrich Inc. LLC., otherwise indicated. rac-␥-Cyclogeraniol
(rac-4) was synthesized by previously reported procedure [6].
Geraniol, geranylacetone, lipase CAL-B (polymer supported Can-
dida antarctica lipase B) and PPL (Porcine pancreas lipase) were
purchased from Sigma–Aldrich Inc. LLC. Amano PS-C (Burkholderia
cepacia lipase on ceramic) and Amano AK (Pseudomonas fluorescens
lipase) were gifted from Amano Enzymes. Inc. Lipase QL (Alcaligenes
∗
Corresponding author at: School of Pharmacy, International University of Health
and Welfare, 2600-1 Kitakanemaru, Ohtawara, Tochigi 324-8501, Japan.
E-mail address: mfujii@iuhw.ac.jp (M. Fujii).
http://dx.doi.org/10.1016/j.molcatb.2015.11.019
1381-1177/© 2015 Elsevier B.V. All rights reserved.

M. Fujii et al. / Journal of Molecular Catalysis B: Enzymatic 123 (2016) 160–166
161
Table 1
Screening of enzymes for the enantiomeric resolution of rac-4.
Enzyme
Time (h)
23
143
180
6
6
180
48
Ee of 4 (%)a
Ee of ester (%)b
Conv. (%)c
Ed
Amano PS–C
PPL
13.6 (S)
2.3 (S)
14.0 (S)
15.5 (S)
33.1 (R)
3.0 (R)
34.6 (R)
9.0 (R)
28.2
20.4
31.8
36.5
57.2
8.6
2.3
1.2
2.1
2.0
2.2
2.0
7.1
Amano AK
Lipase QL
Lipase OF
Lipase MY
CAL-B
30.0 (R)
27.0 (R)
24.8 (S)
31.9 (S)
70.8 (R)
20.2 (S)
22.2
a
Determined by Chiral HPLC analysis (CHIRALPAK OD-H, Daicel Corp.) after ben-
zoylation.
b
Calculated based on the ee of 4 and the conversion [12].
Determined by 1H NMR spectra of the reaction mixture.
Calculated based on ee of recovered 4 and the conversion [12].
c
d
(1 mL) to convert unreacted 4 to the corresponding benzoyl ester
and the ee of 4 was determined by HPLC analysis equipped with
chiralpak OD-H. E value of the reaction was calculated based on the
ee of recovered 4 and the conversion. HPLC analysis: n-hexane/i-
PrOH = 100/1, Flow rate; 1.0 mL/min, Detection; 220 nm. Benzoate
of (S)-4; tR = 11.8 min, benzoate of (R)-4: tR = 13.2 min. The results
are summarized in Tables 1 and 2. The signals of 1H NMR that were
used to calculate the conversion are listed below.
4: 1H NMR (CDCl3) ␦ 3.60 (1H, dd, J = 4.8, 11.2 Hz), 3.69 (1H, dd,
J = 10.6, 11.2 Hz). Acetate of 4:1H NMR (CDCl3) ␦ 4.20 (1H, dd, J = 5.2,
10.8 Hz), 4.24 (1H, dd, J = 9.2, 10.8 Hz). Propanoate of 4: 1H NMR
(CDCl3) ␦ 4.21 (1H, dd, J = 5.2, 10.8 Hz), 4.26 (1H, dd, J = 9.2, 10.8 Hz).
Butanoate of 4; 1H NMR: ␦ 4.21 (1H, dd, J = 5.2, 10.8 Hz), 4.25 (1H,
dd, J = 9.2, 10.8 Hz), 4.58 (1H, s), 4.80 (1H, s). EI-MS m/z: 224 (Calced
for C14H24O2: 224). Hexanoate of 4; 1H NMR: ␦ 4.22 (1H, dd, J = 5.2,
11.2 Hz), 4.25 (1H, dd, J = 9.2, 11.2 Hz), 4.58 (1H, s), 4.80 (1H, s). EI-
MS m/z: 252 (Calced for C16H28O2: 252). Octanoate of 4; 1H NMR:
␦ 4.22 (1H, dd, J = 4.8, 11.0 Hz), 4.25 (1H, dd, J = 8.8, 11.0 Hz), 4.59
(1H, s), 4.81 (1H, s). EI-MS m/z: 280 (Calced for C18H32O2: 280).
Decanoate of 4; 1H NMR: ␦ 4.23(1H, dd, J = 4.8, 11.0 Hz), 4.28 (1H,
dd, J = 8.8, 11.0 Hz), 4.59 (1H, s), 4.81 (1H, s). EI-MS m/z: 308 (Calced
for C20H36O2: 308). Laurate of 4; 1H NMR: ␦ 4.22 (1H, dd, J = 4.8,
11.0 Hz), 4.25 (1H, dd, J = 8.8, 11.2 Hz), 4.58 (1H, s), 4.80 (1H, s). EI-
MS m/z: 336 (Calced for C22H40O2: 336). Myristate of 4; 1H NMR:
␦ 4.22 (1H, dd, J = 4.8, 11.0 Hz), 4.26 (1H, dd, J = 8.8, 11.2 Hz), 4.59
(1H, s), 4.81 (1H, s). EI-MS m/z: 364 (Calced for C24H44O2: 364).
Crotonate of 4; 1H NMR: ␦ 4.22 (1H, dd, J = 4.8, 11.0 Hz), 4.26 (1H,
dd, J = 8.8, 11.2 Hz), 4.62 (1H, s), 4.83 (1H, s). EI-MS m/z: 222 (Calced
for C14H22O2: 222). Benzoate of 4; 1H NMR: ␦ 4.44 (1H, dd, J = 10.0,
11.2 Hz), 4.53 (1H, dd, J = 5.4, 11.2 Hz).
Scheme 1. Reported methods for synthesis of enantiomerically active 4.
sp. lipase), lipase OF (Candida rugosa lipase) and lipase MY (C. rugosa
lipase) were purchased from Meito Sangyo Co. Ltd.
2.2. General procedure for enzymatic kinetic resolution of
ꢀ-cyclogeraniol.
To a solution of rac-4 (100 mg, 0.45 mmol) and acyl donor (10
eq.) in diisopropyl ether (1 mL) was added lipase (100 mg) and the
resulting mixture was stirred at 25 ◦C. The conversion was deter-
mined by the measurement of 1H NMR of the reaction mixture. The
reaction was stopped by filtration through a pad of Celite 545. The
residue was washed with ether, then the filtrate and ether solution
were combined and evaporated under reduced pressure to afford
the mixture of the substrate and the corresponding product. The
residue was treated with benzoyl chloride (300 mg) in pyridine
2.3. Preparation of (S)-ꢀ-cyclogeraniol
To a solution of rac-4 (3.00 g, 19.5 mmol) and vinyl propanoate
(10 mL) in diisopropyl ether (100 mL) was added lipase CAL-B
(1.0 g) and the resulting mixture was stirred at room temperature
for 7 days. The reaction mixture was filtrated through a pad of
Celite 545. The residue was washed with ether, then the filtrate and
ether solution were combined, and evaporated under reduced pres-
sure. The residue was treated with the same conditions described
above {vinyl propanoate (10 mL), diisopropyl ether (100 mL) and
lipase CAL-B (1.0 g)}, and then the residue was purified by gel col-
umn chromatography [silica gel (80 g), hexane/ethyl acetate (15/1)]
to afford (S)-4 (702 mg, 4.55 mmol, 23%, >99% ee) and the propyl
ester of 4 (2.95 g, 14.0 mmol, 72%, 32% ee). After small amount of
Fig. 1. Structures of natural terpenoids possessing ␥-cyclogeranyl unit.

162
M. Fujii et al. / Journal of Molecular Catalysis B: Enzymatic 123 (2016) 160–166
2.4.2. 4-((1ꢀS)-2ꢀ,2ꢀ-Dimethyl-6-methylenecyclohexyl)
butan-2-ol (7)
Table 2
Screening of acyl donor on the lipase-catalyzed enantiomeric resolution of rac-4.
To a solution of (S)-6 (1.26 g, 4.53 mmol) in dry THF (30 mL)
were added n-butyl lithium in hexane (2.6 M, 1.5 mL, 3.9 mmol)
at 0 ◦C and then propylene oxide (1.0 mL, 16 mmol) were added
to the mixture. The mixture was stirred at rt for 2 h, then poured
into water and extracted with ethyl acetate. The organic layer was
dried over Na2SO4 and evaporated under reduced pressure. The
residue was chromatographed by silica gel [silica gel (50 g), hex-
ane/AcOEt (10/1)] to afford crude product (1.5 g). To the mixture of
the crude product, Na2HPO3 (1.50 g) and EtOH (2 mL) in THF (50 mL)
was added Na (1.0 g) at 0 ◦C and the mixture was stirred at the same
temperature for 12 h. The mixture was evaporated under reduced
pressure and the residue was purified by column chromatography
[silica gel (30 g), hexane/AcOEt (10/1)] to afford a diastereomeric
mixture of (1ꢀS)-7 (639 mg, 3.26 mmol, 84%).
Acyl donor
Time (h) Ee of 4(%)a Ee of ester(%)b Conv. (%)c Ed
Vinyl acetate
48
50
20.2
24.3
25.7
57.0
53.0
37.5
38.7
34.0
41.5
1.9
70.8
68.4
66.4
50.1
46.6
27.0
39.2
48.7
48.8e
4.6
22.2
26.2
27.9
53.2
53.2
58.1
49.7
41.1
46.0
29.4
7.1
6.7
6.3
5.2
4.5
2.4
3.2
4.0
4.3f
1.1
Vinyl propanoate
Vinyl butanoate
Vinyl hexanoate
Vinyl octanoate
Vinyl decanoate
Vinyl laurylate
Vinyl myristate
Vinyl benzoate
Vinyl crotonate
24
11
11
24
15
15
157
282
2.4.3. 5-[4-((1S)-2,2-Dimethyl-6-methylenecyclohexyl)
butan-2-ylsulfanyl]-1-phenyltetrazole (8a)
a
Determined by Chiral HPLC analysis equipped with CHIRALPAK OD-H after ben-
zoylation.
b
c
d
e
f
To a solution of 7 (596 mg, 3.04 mmol), 1-phenyl-1H-tetrazole-
5-thiol (PTSH) (587 mg, 3.30 mmol) and triphenylphosphine
(0.988 g, 3.77 mmol) in THF (10 mL) were added DEAD (2.2 M
toluene solution, 1.5 mL, 3.3 mmol) in toluene at 0 ◦C. The mixture
was stirred at 0 ◦C for 2 h, then poured into water and extracted
with ethyl acetate. The organic layer was dried over Na2SO4 and
evaporated under reduced pressure. The residue was purified by
column chromatography [silica gel (15 g), hexane/AcOEt (10/1)] to
afford 8a (565 mg, 2.91 mmol, 96%) [8].
Calculated based on the ee of 4 and the conversion.
Determined by 1H NMR spectra of the reaction mixture.
Calculated based on ee of recovered (S)-4 and the conversion.
Determined by Chiral HPLC analysis.
Calculated by the ee of ester and the conversion.
(S)-4 was converted to the corresponding benzoyl ester by ben-
zoyl chloride and pyridine, the ee of 4 was determined as >99% ee
by HPLC analysis equipped with chiralpak OD-H. [␣]D21 + 23.7 (c
1.00, CHCl3). Lit [␣]D21 + 23.7 (c 0.31, CHCl3)[7]. Propanoate of 4:
2.4.4. 5-[4-((1S)-2,2-Dimethyl-6-methylenecyclohexyl)
butan-2-ylsulfonyl]-1-phenyltetrazole (9a)
IR (neat) ␯ 1734 cm−1 1H NMR (CDCl3) ␦ 0.87 (3H, s), 0.98 (3H, s),
.
1.11 (1H, t, J = 7.6 Hz), 1.14–1.28 (4H, m), 2.01–2.12 (2H, m), 2.29
(2H, q, J = 7.6 Hz), 4.21 (1H, dd, J = 5.2, 10.8 Hz), 4.26 (1H, dd, J = 9.2,
10.8 Hz), 4.59 (1H, s), 4.81 (1H, s). 13C NMR (CDCl3, 100 MHz) ␦ 9.2,
23.4, 25.2, 27.7, 28.7, 33.3, 34.3, 37.6, 52.2, 62.6, 109.7, 147.1, 174.6.
EI-HRMS 210.1622 (Calced for C13H22O2: 210.1620).
To
a
mixture of 8a (212 mg, 0.862 mmol) and
(NH4)6Mo7O24·H2O (12 mg, 0.01 mmol) in EtOH (5 mL) was
added aq. 30% hydrogen peroxide (30%, 1.5 mL) at 0 ◦C, the mixture
was stirred at rt for 8 h and added ice-cold aq. sodium sulfite.
The mixture was extracted with ethyl acetate, the organic layer
was dried over Na2SO4 and evaporated under reduced pressure.
The residue was purified by silica gel column chromatography
[silica gel (10 g), hexane/ethyl acetate (10/1)] to afford 9a (239 mg,
0.86 mmol, 99% yield). Spectral date of 9a were identical with
those of reported value [8].
2.4. Synthesis of (S)-trixago1
2.4.1. (S)-1-[1ꢀ-Benzenesulfonyl]methyl]-2,2-dimehtyl-6-
methylenecyclohexane
(6)
2.4.5. 2-[4-((1S)-2,2-Dimethyl-6-methylenecyclohexyl)
butan-2-ylsulfonyl]benzothiazole (9b)
To a solution of (S)-4 (1.40 g, 9.09 mmol) in pyridine (5 mL) was
added mesyl chloride (1.0 mL, 12 mmol) at 0 ◦C. The mixture was
stirred at rt for 2 h, poured into water and extracted with a mix-
ture of hexane and ethyl acetate (3/1). The organic layer was dried
over sodium sulfate and evaporated under reduced pressure. The
residue was added to a mixture of sodium hydride (40% in min-
eral oil, 1.2 g) and thiophenol (2.2 g, 20 mmol) in DMF (100 mL) and
stirred at 90 ◦C for 6 h. The mixture was cooled to rt, poured into
water and extracted with a mixture of hexane and ethyl acetate
(3/1). The organic layer was dried over sodium sulfate and evap-
orated under reduced pressure. The residue was purified by silica
gel column chromatography [silica gel (50 g), hexane/ethyl acetate
(50/1)] to afford (S)-5 (2.12 g, 8.62 mmol, 89% yield). To a mix-
ture of (S)-5 (2.12 g, 8.62 mmol) and (NH4)6Mo7O24·H2O (120 mg,
0.1 mmol) in ethanol (30 mL) was added aq. 30% hydrogen peroxide
(30%, 15 mL) at 0 ◦C, the mixture was stirred at rt for 8 h and added
ice-cold aq. sodium sulfite. The mixture was extracted with ethyl
acetate, the organic layer was dried over sodium sulfate and evap-
orated under reduced pressure. The residue was purified by silica
gel column chromatography [silica gel (30 g), hexane/ethyl acetate
(10/1)] to afford (S)-6 (2.39 g, 8.60 mmol, 99%). [␣]D + 13.7 (c 1.0,
CHCl3). Lit[8]. [␣]D + 10.8 (c 2.1, CHCl3). 1H and 13C NMR spectra
were identical with those of reported value [8].
9b (441 mg, 1.17 mmol, 87% 2 steps) was prepared from 7
(290 mg, 1.48 mmol) by the same procedure described above using
2-benzothiazolethiol (BTSH) instead of PTSH. Spectral date of (S)-9b
were identical with those of reported value [8].
2.4.6. (6E)-8-(2,4-Tetrahydropyranyloxy)-2,6-dimethyloct-6-
ene-2,3-diol
To
a solution of potassium carbonate (8.0 g, 62.5 mmol),
K3[Fe(III)(CN)6] (14.5 g, 0.044 mol), potassium osmate(IV) (15.4 mg,
41.8 mmol) and methanesulfonamide (2.0 g, 20 mmol) in water
(100 mL) was added a solution of geraniol THP ether (5.0 g,
21 mmol) in t-BuOH (100 mL) at 0 ◦C, and the mixture was stirred at
room temperature for 20 h. A saturated aqueous solution of sodium
hydrogen sulfate (100 mL) was added to the reaction mixture and
the resulting mixture was extracted by CH2Cl2 four times. The
organic layers were collected, dried over Na2SO4 and evaporated
under reduced pressure. The residue was purified by silica gel col-
umn chromatography [silica gel (150 g), hexane/ethyl acetate (2/1)]
to afford the diastereomeric mixture of 6,7-diol of THP-geraniol
(4.14 g, 14.9 mmol, 71% yield). IR (neat) ␯ 3433, 1021 cm−1 1H NMR
.
(CDCl3) ␦ 1.10 (3H, s), 1.15 (3H, s), 1.30–1.70 (7H, m) 1.64 (3H,
s), 1.70–1.82 (1H, m), 1.99–2.09 (1H, m), 2.20–2.31 (1H, m), 2.60

M. Fujii et al. / Journal of Molecular Catalysis B: Enzymatic 123 (2016) 160–166
163
Table 3
Reaction conditions for one-pot Julia coupling for the synthesis of 11.
Entry
sulfone
Base
Temp
Yield(%)
E/Z
1
2
3
4
5
9a
9a
9a
9a
9b
LiHMDS
NaHMDS
KHMDS
KHMDS
KHMDS
78
78
78
-90
-78
47
43
30
46
23
1.2/1
1.1/1
3.3/1
5/1
1/1.1
Conditions: 9, 200 mg, THF, 5 mL, Base, 1.1 eq.
(2H, br) 3.29 (1H, d, J = 10.8 Hz), 3.41-3.51 (1H, m), 3.78-3.90 (1H,
m), 3.91–3.42 (1H, m), 4.13–4.24 (1H, m), 4.57 (1H, t, J = 2.0 Hz),
5.30–5.54 (1H, m). 13C NMR (CDCl3) ␦ 16.3 (16.3 for the diastere-
omer), 19.5, 23.2, 25.4, 26.3, 29.4,30.6, 36.7, 62.3, 63.6 (63.7 for the
diastereomer), 73.0, 78.0 (78.1 for the diastereomer), 98.0 (98.1
for the diastereomer), 121.0 (121.0 for the diastereomer), 139.9.
EI HR-MS m/z: 272.1992 (Calced for C15H28O4: 272.1988).
5.20–5.32 (1H, m). EI HR-MS m/z: 374.3179 (Calced for C25H42O2:
374.3185).
2.4.9. (S)-Trixagol tetrahydropyranyl ether 11
0.5 M KHMDS in toluene (1.1 mL) was added to a solution of (S)-
9a (198 mg, 0.529 mmol) in dry THF (5 mL) was added at −30 ◦C
under Ar atmosphere and stirred for 1 h. The mixture was cooled
at −90 ◦C and 10 (126 mg, 0.594 mmol) was added to the mixture
and stirred for 3 h at the same temperature. The reaction mixture
was warmed up to room temperature and was concentrated under
reduced pressure. The residue was purified by silica gel column
chromatography [silica gel (10 g), hexane/AcOEt (20/1)] afford (S)-
11 (88.1 mg, 0234 mmol, 46%) as stereoisomeric mixture.
2.4.7. (4E)-6-(2,4-Tetrahydropyranyloxy)-4-methylhex-4-enal
(10)
To a solution of sodium periodate (0.863 g, 4.05 mmol) in water
(30 mL) was added a solution of the diol (1.00 g, 3.67 mmol) pre-
pared in the Section 2.4.6. in EtOH (30 mL) at 0 ◦C, and the mixture
was stirred at the same temperature for 2 h. A saturated aque-
ous solution of sodium hydrogen sulfate (100 mL) was added to
the reaction mixture and the resulting mixture was extracted
by CH2Cl2 three times. The organic layers were collected, dried
over Na2SO4 and evaporated under reduced pressure to afford 10
(763 mg, 3.67 mmol, 100%), which was then used immediately for
the next coupling reaction without further purification. 1H NMR
(CDCl3) ␦ 1.70 (3H, s), 1.45–1.88 (6H, m), 2.37 (2H, t, J = 7.2 Hz), 2.58
(2H, t, J = 7.2 Hz), 3.48–3.57 (1H, m), 3.90–3.97 (1H, m), 4.02 (1H, dd,
J = 7.2, 12.0 Hz), 4.23 (1H, dd, J = 6.0, 12.0 Hz), 4.61 (1H, s), 5.39 (1H,
t, J = 2.0 Hz), 9.78 (1H, s). 13C NMR (CDCl3, 100 MHz) ␦ 19.8, 22.8,
28.7, 33.9, 34.8, 45.1, 65.6, 66.8, 101.3, 124.9, 141.1, 205.3.
2.4.10. (S)-Trixagol (S)-1
To a solution of (S)-11 (E/Z = 5/1) (63.4 mg, 0.169 mmol) in EtOH
(2 mL) was added PPTS (2.57 mg, 0.015 mmol) and the mixture was
stirred for 2 h under Ar atmosphere. The mixture was poured into
sat. NaHCO3 and extracted with AcOEt twice. The organic layer was
dried over Na2SO4 and evaporated under reduced pressure. The
residue was purified by silica gel column chromatography [silica gel
(60N) (5 g), hexane/AcOEt (15/1)] afford the stereoisomeric mixture
at C6 position of (S)-1 (E/Z = 5/1) (46.7 mg, 0.161 mmol, 95%). The
mixture was further purified by silica gel column chromatography
[silica gel (60N) (50 g), benzene/ether (10/1)] afford (S)-1 (21.0 mg,
45%) and stereoisomeric mixture (25.7 mg, 55%) of (6E)-1 and (6Z)-
1. [␣]D21 = +14.1 (c 0.5, CHCl3), lit [3]. [␣]D = +14 (c 2.6, CHCl3). The
spectral data was identical with those of reported value [1,3]. 1H
NMR (CDCl3): ␦ 0.81 (3H, s), 0.88 (3H, s), 1.14–1.75 (10H, m), 1.57
(3H, s), 1.67 (3H, s), 1.85–2.13 (6H, m), 4.13 (2H, d, J = 6.8 Hz), 4.51
(1H, s), 4.73 (1H, s), 5.07 (1H, t, J = 6.4 Hz), 5.41 (1H, t, J = 6.4 Hz).
2.4.8. General procedure of one pot Julia coupling of 9 and 10
To a solution of (1S)-9a (200 mg, 0.52 mmol) or (1S)-9b in dried
THF (5 mL) was added base (1.1 eq.) indicated in Table 3 at −78 ◦C.
The mixture was stirred for 1 h at the same temperature, then 10
was added to the mixture. The mixture was stirred for 3 h at the
same temperature, and was concentrated under reduced pressure.
The residue was purified by silica gel column chromatography [sil-
ica gel (10 g), hexane/AcOEt (20/1)] afford 11 as stereoisomeric
mixture. The results were listed in Table 3. The stereoisomeric ratio
were determined by signals at 4.5 ppm on 1H NMR. 11: 1H NMR: ␦
[0.75, 0.76] (3H, s each), [0.83, 0.84] (3H, s each), [1.51, 1.61] (3H, s
each), 1.54 (3H, s), 3.36–3.48 (1H, m), 3.75–3.86 (1H, m), 3.90–3.99
(1H, m), 4.10–4.19 (1H, m) [4.46(E), 4.50 (Z)] (1H, s each), 4.56 (1H,
t, J = 4.4 Hz), [4.67(E), 4.70 (Z)] (1H, s each), 5.01 (1H, t, J = 6.8 Hz),
2.5. Synthesis of Luffarin-P
2.5.1. 1-Geranyl-3-hydroxyacetone.
To a solution of geranylacetone (15.7 g, 79 mmol) in THF
(100 mL) was dropwise added LiHMDS (80 mL, 1 M solution in THF,
80 mmol) at −78 ◦C, the mixture was stirred for 30 min at −78 ◦C,
then TMSCl (12 mL, 94 mmol) was added to the mixture at the
same temperature and the resulting mixture was stirred for 30 min.
The mixture was concentrated under reduced pressure, the residue
was dissolved in hexane and the resulting participate was filtrated.
The filtrate was concentrated under reduced pressure to afford the
crude TMS-enolether of geranylacetone. The residue was added
to a suspension of NBS (13.5 g, 75.8 mmol) in CH2Cl2 (300 mL) at
−78 ◦C was stirred for 1 h at the same temperature. The mixture
was concentrated under reduced pressure, the residue was dis-
solved in hexane and the resulting participate was removed by
filtration. The filtrate was concentrated under reduced pressure
Scheme 2. Preparation of (S)-4.

164
M. Fujii et al. / Journal of Molecular Catalysis B: Enzymatic 123 (2016) 160–166
Bakkestein
et al.
SPh
8
OR
TsO
+
8
6
OH
9
1
1
Our approach
Scheme 4. Synthesis of sulfone 9.
H
SO2R
6
OR
O
115.5, 122.4, 137.2, 170.4, 174.3. EI HR-MS m/z: 268.1669 (Calced
for C15H24O4: 268.1675).
+
2.5.4. (3E)-3-(7-Formyl-4-mehtylhex-3-enyl) but-2-en-4-olide
(12)
Scheme 3. Synthetic routes for synthesis of 1.
To a solution of sodium periodate (0.231 g, 1.00 mmol) in water
(30 mL) was added a solution of diol (0.232 g, 0.865 mmol) in EtOH
(30 mL) at 0 ◦C, and the mixture was stirred at the same tem-
perature for 2 h. A saturated aqueous solution of NaHSO3 (10 mL)
was added to the reaction mixture and the resulting mixture was
extracted by CH2Cl2 four times. The organic layers were collected,
dried over Na2SO4 and evaporated under reduced pressure to afford
12 (180 mg, 0.865 mmol, 100%), which was then used immediately
for the next coupling reaction without further purification. 1H NMR
(CDCl3) ␦ 1.64 (3H, s), 2.27–2.38 (2H, m), 2.45 (2H, t, J = 7.2 Hz), 2.54
(2H, td, J = 6.6, 1.6 Hz), 4.73 (2H, s), 5.40 (1H, t, J = 5.6 Hz), 5.84 (1H,
s), 9.75 (1H, t, J = 1.6 Hz). 13C NMR (CDCl3) ␦ 16.1, 25.5, 28.4, 31.5,
41.9, 73.0, 115.5, 122.9, 135.5, 169.9, 173.9, 201.9.
to afford the crude product of 1-bromo-3-geranylacetone. To the
crude product of 1-bromo-3-geranylacetone in DMF (100 mL) was
added KOAc (10 g, 0.10 mol) and stirred for 6 h at 50 ◦C. The mix-
ture was poured into ice-cooled water (1 L) and was extracted with
hexane/AcOEt (20/1), and the organic layer was concentrated in
reduced pressure. The residue (13 and 13ꢀ) was added to a sus-
pension of lipase PS-C (1.0 g) in 10% ethanol in hexane (300 mL),
and was stirred for 6 h. The reaction mixture was filtrated by Celite
545, evaporated under reduced pressure, and purified by silica gel
column chromatography [silica gel (100 g), hexane/AcOEt (12/1)]
to afford 1-geranyl-3-hydroxyacetone 14 (9.53 g, 45.3 mmol, 60%)
[9].
2.5.5. Lufferin-P ((S)-2)
2.5.2. (3E,7E)-3-(4,8-Dimethylnon-3,7-dienyl) but-2-en-4-olide
(15)
To a solution of (S)-9a (122 mg, 0.314 mmol) in dry THF (3 mL)
was added 0.5 M KHMDS in toluene (0.6 mL, 0.3 mmol) at −30 ◦C,
the mixture was cooled at −90 ◦C after stirring for 1 h at −30 ◦C,
and 18 was added to the mixture. The mixture was concentrated
under reduced pressure. The residue was purified by silica gel
column chromatography [silica gel (10 g), hexane/AcOEt (50/1)]
afford the stereoisomeric mixture at C3ꢀ position of (S)-2 (82.2 mg,
0.245 mmol, 74%). The mixture was further purified by silica gel
column chromatography [silica gel (60N) (50 g), benzene/ether
(10/1)] afford (S)-2 (9.8 mg, 12%) and the stereoisomeric mixture
To a solution of 12 (333 mg, 0.169 mmol) in benzene (8 mL) was
added (triphenylphosphoranylidene) ketene (485 mg, 0161 mmol)
and the mixture was stirred for 5 h under Ar atmosphere at 60 ◦C.
The mixture was concentrated under reduced pressure. The residue
was purified by silica gel column chromatography [silica gel (10 g),
hexane/AcOEt (20/1)] to afford 15 (312 mg, 0.133 mmol, 83%). Spec-
tral date of 15 were identical with those of reported value [10].
20
(72.4 mg, 88%) of 2. [␣]D +3.7 (c 0.13, CHCl3). Lit [2]. [␣]D +5
2.5.3. (3E)-3-(7,8-Dihydroxy-4,8-dimehtylnon-3-enyl)
(c 0.1, CHCl3) Spectral data were identical with those of reported
value [2]. 1H NMR (CDCl3) ␦ 0.81 (3H, s), 0.91 (3H, s), 1.17–1.27
(1H, m), 1.38-1.61 (4H, m), 1.59 (3H, s), 1.62 (3H, s) 1.65–2.11 (10H,
m) 2.29 (2H, t, J = 7.2 Hz), 2.45 (2H, t, J = 6.6 Hz), 4.53 (1H, dd, J = 2.4,
15.2 Hz), 4.72 (2H, s), 4.77 (1H, d, J = 11.2 Hz), 5.02-5.12 (2H, m), 5.84
(1H, t, J = 1.6 Hz). EI HR-MS m/z: 370.2884 (Calced for C25H38O2:
370.2872).
but-2-en-4-olide
To a solution of K2CO3 (800 mg, 6.25 mmol), K3[Fe(III)(CN)6]
(1.45 g, 4.4 mmol), potassium osmate(IV) (3.08 mg, 8.36 ␮mol) and
methanesulfonamide (20 mg, 0.2 mmol) in water (10 mL) was
added a solution of 15 (521 mg, 2.22 mmol) in t-BuOH (10 mL), and
the mixture was stirred at room temperature for 20 h. A saturated
aqueous solution of NaHSO3 (100 mL) was added to the reaction
mixture and the resulting mixture was extracted by CH2Cl2 four
times. The organic layers were collected, dried over Na2SO4 and
evaporated under reduced pressure. The residue was purified by
silica gel column chromatography [silica gel (10 g), hexane/ethyl
acetate (2/1)] to afford (3E)-3-(7,8-dihydroxy-4,8-dimehtylnon-3-
enyl) but-2-en-4-olide (435 mg, 1.60 mmol, 72% yield). IR (neat) ␯
3. Results and discussion
3.1. Enantiomeric resolution of rac-4
Because the chiral center of the primary alcohol 4 is distant
from the reaction center, enzymatic resolution of rac-4 is diffi-
cult [11]. In addition, as 4 does not contain a heteroatom, except
for the hydroxyl group, the enzyme must recognize the small dif-
ferences between the methylene and dimethyl groups, which are
3420, 1780, 1742 cm−1 1H NMR (CDCl3) ␦ 1.11 (3H, s), 1.15 (3H,
.
s), 1.29-1.60 (2H, m) 1.58 (3H, s), 1.98–2.48 (8H, m), 3.24 (1H, dd,
J = 6.0, 8.8 Hz), 4.70 (2H, s), 5.11 (1H, t, J = 6.4 Hz), 5.80 (1H, s). 13C
NMR (CDCl3): ␦ 16.0, 23.0, 25.6, 26.4, 28.5, 29.5, 36.5, 73.0, 73.2 77.8,

M. Fujii et al. / Journal of Molecular Catalysis B: Enzymatic 123 (2016) 160–166
165
separated from the hydroxyl group by three carbon atoms. Thus,
OAc
enzymatic resolution of rac-4 is challenging. rac-4 was prepared
from 6-methylhept-5-en-2-one in four steps to achieve 74% over-
all yield [6]. Several lipases, including pig porcine lipase (PPL), C.
antarctica lipase B (CAL-B), Amano PS-C, Amano AK, Lipase QL,
Lipase OF and Lipase MY were tested for their stereoselectivity of
rac-4 using a our conventional assay method [12]. The results are
shown in Table 1. The quality of the reaction was determined by
the E value [13] calculated on the basis of the conversion and the
enantiomeric excess (ee) of recovered 4. The reactions catalyzed by
Amano PS, PPL, Amano AK, Lipase QL and CAL-B afforded (R)-ester
whereas Lipase OF and Lipase MY afforded (S)-ester. Among them,
CAL-B showed the best stereoselectivity as indicated by the E value
(E = 7.1). It is known that changing the acyl donor affects the stere-
oselectivity and reactivity of lipase [12,14–16]. Several acyl donors
in the transesterification of rac-4 were tested, but their E values
were lower than that of vinyl acetate (Table 2). However, elonga-
tion of the carbon chain of the acyl donor increased the reaction
rate. Thus, to the reaction period for obtaining enantiomerically
pure (S)-4, vinyl propionate was employed over vinyl acetate in the
large-scale experiment. To achieve this, 3.0 g of rac-4 was treated
with CAL-B in vinyl propionate and i-Pr2O for 14 days, propionate
of (R)-4 with a 32% ee was obtained in 72% yield, and enantiomer-
ically pure (S)-4 with >99% ee was recovered in 23% yield (700 mg)
(Scheme 2). As CAL-B was deactivated in a week by acetaldehyde
generated from vinyl propionate, fresh CAL-B, vinyl propionate, and
diisopropyl ether were added after the evaporation of the reaction
mixture to complete the reaction.
O
a
13
+
OAc
O
geranylacetone
O
13'
b
c
OH
60%
2 steps
O
83%
14
d
OHC
O
O
O
71%
O
15
12
O
e
f
O
2
(S)-
74%
12%
2
(S)- (E : Z = 7 : 2)
Conditions: a) 1. LiHMDS, TMSCl, 2. NBS, 3. AcOK, b) Lipase PS-C, EtOH,
c) (triphenylphosphoranylidene)ketene. d) 1) K2OsO4, K3Fe(CN6),
K2CO3, CH3SO2NH2, 2) KIO4, e) , KHMDS, DMA, -90 oC,
9a
f) silica gel chromatography
Scheme 6. Synthesis of Lufferin-P.
a diastereomer mixture of 7 in 84% yield in two steps. Treating
7 with PTSH under Mitsunobu reaction conditions gave 8a in 96%
yield, followed by oxidation by hydrogen peroxide and the addition
of a catalytic amount of molybdenum complex afforded sulfone 9a
in 99% yield. Treating 7 with BTSH instead of PTSH using the same
procedure afforded 8b (88%) and then 9b (97%)
3.2. Synthesis of (S)-trixagol (1)
(S)-(+)-Trixagol
1 was isolated from Bellardia trixago [1].
Recently, several groups have achieved their total synthesis
[3,17,18]. Bakkestein et al. reported the chiral synthesis of the (S)-
1 skeleton at the C8–C9 bond (Scheme 3) [3,19]. We attempted
the skeleton construction of 1 at the C6–C7 double bond by one-
pot Julia olefination [20] because it seemed difficult to apply their
methods to the synthesis of 2, which possesses the reactive ␣,␤-
unsaturated lactone moiety.
Mesylation of (S)-4, followed by treatment with sodium
phenylthiolate at 80 ◦C, yielded 89% of sulfide 5 (Scheme 4). Oxida-
tion by hydrogen peroxide in the presence of molybdenum complex
afforded sulfone 6 in 99% yield. An anion generated from 6 and n-
butyl lithium was reacted with racemic propylene oxide and then
desulfonated by sodium in a mixture of THF and ethanol to give
Aldehyde 10 was prepared by selective oxidation from
tetrahydropyranyl-protected geraniol (THP-geraniol) in 71% yield
(Scheme 5). Several bases were tested for one-pot Julia coupling
of 9a or 9b and aldehyde 10; the results are listed in Table 3.
The use of NaHMDS on the reaction with 9a gave 11 in compar-
atively higher yield (43%) but the stereoselectivity of the reaction
was low (E/Z = 1.1/1). One-pot Julia coupling of 9a and aldehyde
10 using KHMDS yielded 30% of 11 with the best stereoselectivity.
Based on the stereoselectivity of (E)-11 and (Z)-11, the reaction of
9a with KHMDS was chosen for the preparation of 11. At −90 ◦C,
the reaction yielded 46% of 11 with a stereoselectivity of 5/1. After
removal of the THP group on 1 in 95% yield, silica gel column chro-
matography was applied to isolate 45% of (S)-1 from the mixture
of geometrical isomers of 1. The spectral signature of synthesized
1, including specific rotation, was superimposable with those of
natural 1 [1,3].
a
OTHP
OHC
OTHP
(71%)
THP-geraniol
b
10
OTHP
3.3. Synthesis of (S)-lufferin-P (2)
c
(+)-Luffarin-P (2) was isolated from the Australian marine
sponge, Luffariella geometrica [2]. While S was assumed to be
the absolute configuration of (+)-2 by comparison of its [M]D
sign with that of (+)-luffarin W (3), this has not yet been deter-
mined [2,21]. (S)-2 was synthesized from (1S)-9a and aldehyde
12 to determine the absolute configuration of (+)-2 (Scheme 6).
Aldehyde 12 was synthesized from geranylacetone. Geranylace-
tone was converted to TMS-enolether at −78 ◦C, and then treated
with NBS. Following treatment with potassium acetate, crude
1-acetoxy-3-geranylacetone (13) contaminated with 1-acetoxy-
1-geranylacetone (13ꢀ) (ca. 15%) was produced. On treating the
mixture of 13 and 13ꢀ with lipase Amano PS-C in hexane contain-
ing 10% ethanol, the lipase selectively reacted with 13 to afford
11
(S)- (46%, E/Z = 5:1)
OH
d
1
(S)-
45%
1
(S)- (95%)
Conditions: a) 1) K2OsO4, K3Fe(CN6), K2CO3, CH3SO2NH2, 2) KIO4,
b) , KHMDS, DMA, -90 oC, c) aq. HCl, d) silica-gel chromatography
9a
Scheme 5. Synthesis of (S)-1.

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M. Fujii et al. / Journal of Molecular Catalysis B: Enzymatic 123 (2016) 160–166
14 in 60% yield from geranylacetone. Treatment of 14 with Best-
man’s ketene [22] gave lactone 15 in 76% yield. The regioselective
lipase-catalyzed reaction was necessary to obtain pure 15, since
13ꢀ and its derivatives such as hydroxyketone and lactone were
difficult to remove from 13 to 15. Regioselective dihydroxylation
of 15, followed by oxidative cleavage of the diol by sodium peri-
odate afforded 12 in 71% yield (two steps). One-pot Julia coupling
between (S)-9a and 12 gave a mixture of geometrical isomers of
(S)-2 (E/Z = 7/2) in 74% yield. After separation by silica gel chro-
matography 12% of pure (S)-2 could be isolated and 88% of the
mixture of geometrical isomers could be recovered. The spectral
signature of (S)-2 was superimposable with those of natural (+)-2.
Comparison of the specific rotation sign of synthetic (S)-2 {+3.7 (c
0.13, CHCl3)} with that of natural 2 {+5 (c 0.1, CHCl3)} revealed that
the absolute configuration of the natural (+)-2 was S [2].
Acknowledgements
The authors thank Ms. Yurie Kurimoto and Ms. Sumie Yasuhara
for their supports.
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4. Conclusion
In this study, we attempted the lipase-catalyzed enantiomeric
resolution of rac-4. Several reaserch groups have attempted the
enzymatic enantiomeric resolution of rac-4, but with less than
satisfactory results [8]. The lipase screening results showed rac-4
undergoes a CAL-B catalyzed selective acylation. In addition, car-
bon chain elongation of the acyl donor accelerated the reaction
rate to enhance the enantiomeric purity of the starting material.
In case of the use of vinyl acetate, the reaction gave (S)-4 with 96%
ee at 3 weeks. Applying vinyl propionate to enhance enzymatic
resolution results in >99% ee of (S)-4 at 2 weeks. While the stereos-
electivity and reactivity of the enzymatic enantiomeric resolution
were not satisfied, enantiomerically pure (S)-4 (23% isolated yield)
was obtained by a simple procedure. Synthesized (S)-4 was then
applied for the first synthesis of (+)-lufferin-P. Thus, the absolute
configuration of (+)-lufferin-P possesses an S-configuration.
[15] K. Nakamura, K. Takenaka, A. Ohno, Tetrahedron: Asymmetry 11 (2000)
4429–4439.
[16] M. Kawasaki, M. Goto, S. Kawabata, T. Kometani, Tetrahedron: Asymmetry 12
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[21] The absolute configuration of lufferin-P was assumed as R-configuration [2],
but the structure of lufferin-P was written as S-configuration. We assume the
authors of reference 2 mistake the numbering of the stereochemical
determination, since they mention that (+)-trixagol possessed
R-configuration.
[22] H.J. Bestmann, D. Sandmeier, Angew. Chem. 87 (1975) 630.