The first chemoenzymatic total synthesis of the phytotoxic nonenolide putaminoxin and its (5S,6E,9S)-diastereomer

Article abstract of DOI:10.1016/j.tet.2018.12.027

The limitations of an alcohol dehydrogenase regarding the oxidative kinetic resolution of homoallylic alcohol containing alkyl chains were investigated, leading to a valuable building block for the total synthesis of phytotoxic nonenolide putaminoxin. The enzymatic approach towards the enantioenriched homoallylic alcohol was compared to classical, nonenzymatic approaches using asymmetric reagent controlled allyl additions and the obtained building block was used for the total synthesis of putaminoxin and its (5S,6E,9S)-diastereomer. After the spectroscopic analysis of the synthesized compounds, discrepancies were observed to already published data of isolated and synthesized putaminoxin. Therefore, a systematic comparison of NMR data was carried out. The result underlines the necessity of total synthesis for the absolute assignment of configuration.

Full text of DOI:10.1016/j.tet.2018.12.027

Accepted Manuscript
The first chemoenzymatic total synthesis of the phytotoxic nonenolide putaminoxin
and its (5S,6E,9S)-diastereomer
David Dickmann, Martin Diekmann, Claudia Holec, Jörg Pietruszka
PII:
S0040-4020(18)31505-9
https://doi.org/10.1016/j.tet.2018.12.027
TET 30013
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 31 August 2018
Revised Date: 7 December 2018
Accepted Date: 13 December 2018
Please cite this article as: Dickmann D, Diekmann M, Holec C, Pietruszka Jö, The first chemoenzymatic
total synthesis of the phytotoxic nonenolide putaminoxin and its (5S,6E,9S)-diastereomer, Tetrahedron
(2019), doi: https://doi.org/10.1016/j.tet.2018.12.027.
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The first chemoenzymatic total synthesis of
the phytotoxic nonenolide putaminoxin and
its (5S,6E,9S)-diastereomer
Leave this area blank for abstract info.
a,b
b
b
David Dickmann , Martin Diekmann , Claudia Holec ,
a,b
and Jörg Pietruszka
a
b
Forschungszentrum Jülich, IBG-1: Biotechnology, 52425 Jülich, Germany; Heinrich-Heine-Universität
Düsseldorf, Institut für Bioorganische Chemie im Forschungszentrum Jülich, Gebäude: 15.13, 52426 Jülich,
Germany

1
ACCEPTED MANUSCRIPT
Tetrahedron
journal homepage: www.elsevier.com
The first chemoenzymatic total synthesis of the phytotoxic nonenolide putaminoxin
and its (5S,6E,9S)-diastereomer
a,b
b
b
a,b
David Dickmann , Martin Diekmann , Claudia Holec , and Jörg Pietruszka
a
Forschungszentrum Jülich, IBG-1: Biotechnology, 52425 Jülich, Germany
Heinrich-Heine-Universität Düsseldorf, Institut für Bioorganische Chemie im Forschungszentrum Jülich, Gebäude: 15.13, 52426 Jülich, Germany
b
ARTICLE INFO
ABSTRACT
Article history:
Received
Received in revised form
Accepted
The limitations of an alcohol dehydrogenase regarding the oxidative kinetic resolution of
homoallylic alcohol containing alkyl chains were investigated, leading to a valuable building
block for the total synthesis of phytotoxic nonenolide putaminoxin. The enzymatic approach
towards the enantioenriched homoallylic alcohol was compared to classical, nonenzymatic
approaches using asymmetric reagent controlled allyl additions and the obtained building block
was used for the total synthesis of putaminoxin and its (5S,6E,9S)-diastereomer. After the
spectroscopic analysis of the synthesized compounds, discrepancies were observed to already
published data of isolated and synthesized putaminoxin. Therefore, a systematic comparison of
NMR data was carried out. The result underlines the necessity of total synthesis for the absolute
assignment of configuration.
Available online
Keywords:
Chemoenzymatic synthesis
Oxidative kinetic resolution
Alcohol dehydrogenase
Nonenolides
2
009 Elsevier Ltd. All rights reserved.
Putaminoxin
1
. Introduction
Recently the chemoenzymatic total synthesis of the proposed
structures of putaminoxins B and D (2) were achieved by
employing alcohol dehydrogenases for the selective generation of
Putaminoxin (1) is a phytotoxic compound from Phoma
putaminum, the causal agent of leaf necrosis of a common weed
14
the two stereogenic centers. Both enantiomers of the employed
1-2
of field, Erigeron annuus. It is a 10-membered macrolactone
with a hydroxyl group in position 5, a trans-configured double
bond at position 6 and a propyl-sidechain at position 9. After
isolation from liquid culture of Phoma putaminum the structure
of putaminoxin was investigated and the stereogenic center at
position 5 was determined as being (S)-configured. The
stereogenic center at position 9 could not be assigned initially,
but was found to be (R)-configured after total synthesis and
comparison of the spectroscopic data to the isolated sample. Due
non-1-en-4-ol (5) building block were obtained in good yield and
up to excellent ee-values in either oxidative kinetic resolution of
racemic non-1-en-4-ol (rac-5), or asymmetric reduction of the
corresponding ketone 6 (vide infra), by utilization of an alcohol
dehydrogenase from Thermoanaerobacter brockii. (Tb-ADH). In
the present study the limitations of this enzyme regarding the
length of the alkyl-chain were investigated, giving potential
access to more analogs of these interesting macrolactones via the
identical retrosynthetic approach thus also providing an answer
of questions raised concerning their configuration (Scheme 1).
1
3
to its biological activity, putaminoxin (1) and its analogs
putaminoxin B and D (2), hypocreolide A (3), and aspinolide A
3
-14
(4), were the basis of several synthetic studies (Figure 1).
Figure 1: Structure of putaminoxin and related nonelides.
Scheme 1: Retrosynthesis of nonenolide 1 (PG: protection group).

2
Tetrahedron
2
. Results and Discussion:
and much slower, but continuous oxidation of the remaining (S)-
ACCEPTED MANUSCRIPT
hept-1-en-4-ol [(S)-14] (Figure 2).
2
.1. Oxidative kinetic resolution of aliphatic homoallylic alcohols
Key to the successful synthesis of the proposed structures of
putaminoxin B and D was a convenient enantioselective
synthesis of the required homoallylic alcohol via an oxidative
kinetic resolution utilizing an alcohol dehydrogenase. Here, the
alcohol dehydrogenase from Thermoanaerobacter brockii. was
shown to be able to sterically distinguish between the two alkyl
14-15
moieties of non-1-en-4-ol (5).
To analyze the scope of the Tb-
ADH in the oxidative kinetic resolution of aliphatic homoallylic
alcohols, homoallylic alcohols with increased and decreased
chain length were employed. The oxidation of the longer chain
homoallylic alcohols was slow and their limited solubility
impaired the reliable measurement of conversion, because the
samples were no longer homogenous under the tested reaction
conditions. This effect could unfortunately not be circumvented
by reducing to analytical transformations and working up the
entire reaction since we had to work with stock solutions.
Nevertheless, it could be observed that dec-1-en-4-ol (10) was
oxidized and enantiomerically pure after 8 hours, whereas undec-
Figure 2: Oxidative kinetic resolution of hept-1-en-4-ol (14) ■ and oct-1-
en-4-ol (13) ○ with the alcohol dehydrogenase from Thermoanaerobacter
1
-en-4-ol (11) and dodec-1-en-4-ol (12) were nearly left
brockii. Reaction conditions: Homoallylic alcohol (10 mM), acetone (5 Vol.-
untouched, even after prolonged reaction times (see
supplementary material). These observations are in line with
those reported for the asymmetric reduction of aliphatic ketones
+
%
), Tb-ADH (0.5 U/mL), NADP (300 µM), KP
i
-buffer (50 mM, pH 7.0, 1
2
mM MgCl ), 30 °C, 130 rpm. Samples were taken after 0 h, 1 h, 3 h, 4 h, 5 h,
6
h, 8 h and 24 h.
16
of comparable chainlength. Reducing the concentration of the
longer aliphatic homoallylic alcohols (10-12) from 10 mM to
7
.5 mM led to a decreased reaction time and enantiopure or
For the total synthesis of putaminoxin (1) the reaction was
enantioenriched alcohols after 4 to 24 hours (Table 1). On the
other hand, (S)-oct-1-en-4-ol [(S)-13] and (S)-non-1-en-4-ol [(S)-
upscaled from 50 to 250 mL and (S)-hept-1-en-4-ol [(S)-14]
could be isolated in 25% yield with an ee of 94%. The
stereogenic center was found to be (S)-configured by comparison
with products from the Brown asymmetric reagent controlled
5
] could be obtained with an ee >99% at 50% conversion after
just 3 hours. Surprisingly the Tb-ADH was also able to
differentiate between the alkyl moieties of the nearly symmetrical
hept-1-en-4-ol (14). After 58% conversion an ee of 94% could be
achieved (Table 1).
17
allyl addition to butanal (20) (Scheme 2). This selectivity of the
Tb-ADH is in line with previously reported results and is retained
1
5-16,
even in case of the smaller, and nearly symmetrical substrate.
18
The low yield of this kinetic resolution can probable be
Table 1: Oxidative kinetic resolution of homoallylic alcohols with Tb-
attributed to the volatility of the product, something that was
observed in the Brown asymmetric reagent controlled allyl
addition as well. The selective allyl addition with commercial
Brown reagent led to 23% yield with a maximum (S)-ee of 95%
and was highly dependent on the quality of every single charge
of the reagent. Selective allyl additions with commercially
ADH. Reaction conditions: Homoallylic alcohol (10 mM), acetone (5 Vol.-
+
%
), Tb-ADH (0.5 U/mL), NADP (300 µM), KP
i
-buffer (50 mM, pH 7.0, 1
2
mM MgCl ), 30 °C, 130 rpm.
19
available Leighton-reagent were unsuccessful.
2
.2. Total synthesis of putaminoxin (1) and its (5S,6E,9S)-
diastereomer [(5S,6E,9S)-1]
Substrate
Product
Conversion
%]
ee
[%]
94
>99
>99
Time
[h]
3
3
The enantiomerically enriched (S)-hept-1-en-4-ol [(S)-14] was
further used for the synthesis of putaminoxin (1) and its
[
1
4 (n = 1) (S)-14 (n = 1)
58
50
50
(
5S,6E,9S)-diastereomer [(5S,6E,9S)-1]. The synthetic route was
planned based on previous publications, with a cross-metathesis
of the two major building blocks 6 8 14 and and
Therefore the
1
5
3 (n = 2)
(n = 3)
(S)-13 (n = 2)
(S)-5 (n = 3)
(S)-10 (n = 4)
(S)-11 (n = 5)
(S)-12 (n = 6)
3
a
a
9
a
1
1
1
0 (n = 4)
1 (n = 5)
2 (n = 6)
>99
4
8
,
a
a
macrolactonization as the key steps.
99
90
a
a
homoallylic alcohol (S)-14 was protected with a benzoylic
protection group, because the acetate protected hept-1-en-4-ol
proved to be even more volatile than the unprotected alcohol. For
the synthesis of putaminoxin (1) the protection of the homoallylic
alcohol was performed via Mitsunobu-esterification to invert the
24
a
Substrate concentration was reduced to 7.5 mM.
In previous experiments, Keinan et al. could not observe
product formation in the asymmetric reduction of symmetrical
heptan-4-on, in presence of Tb-ADH, presumably because the n-
pentyl chain does not fit into the small pocket of the substrate
20
configuration. Both protected alcohols (S)-8 and (R)-8, were
obtained in 89% and 83% yield respectively (Scheme 2).
16
binding site of the alcohol dehydrogenase. The oxidative kinetic
resolution of racemic hept-1-en-4-ol (rac-14) revealed that the n-
pentyl chain is indeed not readily accepted in contrast to the
slightly smaller n-pentenyl chain. The discrimination is not as
accurate as for the n-butyl chain of oct-1-en-4-ol (13) though,
leading to a decreased enantiomeric excess after 50% conversion

3
ACCEPTED MANUSCRIPT
Scheme 2: THP = tetrahydropyranyl. Bz = benzoyl. Reaction conditions
+
for A: (a) Tb-ADH, acetone, NADP , KP
MgCl ), 30 °C, 120 rpm, 3.5 h; (b) (+)-B-Allyldiisopinocampheylborane,
Et O, -78 °C, 1 h; (c) benzoyl chloride, pyridine, CH Cl , 24 h; (d) benzoic
acid, triphenyl phosphine, diisopropyl azodicarboxylate, THF, 4.5 h; (e)
i
-buffer (50 mM, pH 7.0, 1 mM
Scheme 3: THP = tetrahydropyranyl. Bz = benzoyl. Reaction conditions:
nd
(a) Grubbs 2 generation catalyst, CH
2
Cl
2
, 40 °C, 3 d; (b) LiOH,
2
2
THF:MeOH:H O (2:1:1), 60 °C, 48 h; (c) 1. 2,4,6-trichlorobenzoyl chloride,
2
2
2
Et N, THF, rt, 2 h; 2. 4-(N,N-dimethylamino)pyridine (DMAP), toluene,
3
nd
Grubbs 2 generation catalyst, CH
2
Cl
2
, 40 °C, 24 h; (f) Lb-ADH, 2-propanol,
); (g) pyridinium para-
, rt, 14 h.
2
reflux, 3 h; (d) PPTS, p-TsOH·H O, EtOH, 40 °C, 16 h.
+
NADP , KP
i
-buffer (50 mM, pH 7.0, 1 mM MgCl
Cl
2
toluenesulfonate, 3,4-dihydro-2H-pyran, CH
2
2
13
2
.3. Comparison of C NMR data of nonenolides
Since a previous publication revealed inconsistencies in the
The second building block (S)-24 could be provided in a
three-step procedure by employing an ADH from Lactobacillus
brevis for the asymmetric reduction of ketone 23, which was
obtained in two steps, starting from ethyl 4-bromobutanoate (22)
13
C NMR data of a variety of putaminoxin analogs that just differ
14
in the length of the alkyl chain at position 9, a comparison of
the synthesized molecules with already published data was
drawn. Even though the MS and IR data are quite similar to those
21
in accordance to the previously described protocol. Reduction
of the ketone yielded the alcohol in 92% yield with an excellent
ee of >99%. The allylic alcohol was protected with a THP-group
in 94% yield before the following cross-metathesis reaction. The
metathesis reaction of protected homoallylic alcohol 8 and
protected allylic alcohol 9 with Grubbs second generation
catalyst afforded the diastereomer 25 in 24% yield. The primary
product was in both cases the homodimerized homoallylic
alcohol 21. Therefore, it was attempted to dimerize the
homoallylic alcohol 8 first, before employing it in the cross-
metathesis, which was beneficial in previous attempts with
1
reported by Evidente et al. for the isolated sample, and Sabitha
3
13
et al. for the synthesized compound, the differences in the
C
14
NMR data are identical to those reported recently. (Figure 3, A)
The newly acquired
13
C NMR data of the synthesized
(
5S,6E,9R)-putaminoxin (1) do not fit to those reported by
1
3
Evidente et al. and Sabitha et al. The data fits very well to the
13
C NMR data of the synthesis of the proposed structures of
14
6
putaminoxin B and D (2) and those of hypocreolide A (3)
though, which are likewise (5S,6E,9R) configurated (Table 2,
Figure 3, A).
6,
14
13
acetate protected non-1-en-4-ol.
Even though the first
On the other hand the C NMR data of the newly synthesized
metathesis led to good yields ranging from 83-86% yield, the
second step did not exceed yields of 41%. However, unreacted
allylic and homodimerized homoallylic alcohol could be
reisolated and reused in a subsequent metathesis reaction giving
nearly identical yields of the heterodimer 25 of 40%. The last
step of the synthesis was a three-step procedure of saponification,
followed by Yamaguchi macrolactonization, and subsequent
deprotection of the THP-protection group. In accordance to a
(5S,6E,9S)-diastereomer [(5S,6E,9S)-1] are in full agreement
13
1
with the C NMR data provided by Evidente et al. and Sabitha
3
et al. for the (5S,6E,9R)-putaminoxin (1) (Figure 3, B).
13
Additionally, the C NMR data of the (5S,6E,9S)-diastereomer
[(5S,6E,9S)-1] fit very well to the data of (5S,6E,9S)-2 by
14
Bisterfeld and Holec et al. and the (5R,6E,5R) configurated.
22
aspinolide A (4), isolated by Fuchser and Zeeck. Significant
differences in the data of the mentioned examples are just in the
signals of the alkyl chains at position 9, which is to be expected
because of their different length. A structural proposition that
explains these correlations would therefore lead to a structure that
is (5R,6E,9R) configurated, rather than (5S,6E,9R) (Figure 3).
6
procedure of Götz et al., the intermediates were not isolated. The
synthesis of putaminoxin (1) and its (5S,6E,9S)-diastereomer
[
(5S,6E,9S)-1] were finalized with moderate yields of 57-66%
over the last three steps (Scheme 3).

4
Tetrahedron
ACCEPTED MANUSCRIPT
1
3
Table 2: Comparison of C NMR signals of the newly synthesized compounds (5S,6E,9R)-1 and (5S,6E,9S)-1 with previously reported data for the originally
1
3
provided structures. Relevant C NMR signals were highlighted by color (green = similar to (5S,6E,9R)-1, red = similar to (5S,6E,9S)-1).
Struc-
tures
1
6
Ref.
Atom
C-1
C-2
C-3
C-4
C-5
C-6
C-7
This
paper
176.8
35.9
18.0
36.8
68.6
136.8
126.4
40.9
76.6
36.5
19.3
14.0
Sabitha
et al.
Evidente et al.
This paper
Bisterfeld and
Holec et al.
Götz et al.
Fuchser and
Zeeck
3
14
22
175.6
35.7
22.3
38.7
74.1
137.1
131.7
40.4
75.3
36.4
19.1
13.9
175.8
35.6
22.2
38.7
74.0
137.2
131.5
40.3
75.3
36.3
19.1
13.8
175.9
35.8
22.5
38.9
74.3
137.3
131.8
40.5
75.5
36.6
19.3
14.0
176.7
35.7
17.8
36.6
68.4
136.6
126.3
40.8
76.8
34.2
25.6
31.6
22.7
14.1
176.6
35.9
18.0
36.8
68.6
136.8
126.5
40.9
76.9
34.4
26.1
29.5
29.3
31.9
22.8
14.2
175.5
35.6
22.3
38.7
74.1
137.1
131.8
42.1
71.6
19.8
C-8
C-9
C-10
C-11
C-12
C-13
C-14
C-15
C-16

5
ACCEPTED MAin
N
cre
U
as
S
es
C
an
R
d
I
t
P
he
T
substrate concentration needs to be lowered,
probably owing to their poor solubility in aqueous media
Table 1). The newly acquired C NMR-data of the final
13
(
products 1 and [(5S,6E,9S)-1] showed a discrepancy with already
13
published data. A comparison with the C NMR data of closely
related analogs, which just differ in the length of the sidechain at
position 9, indicate that the configuration of the stereogenic
centers of putaminoxin (1) should be reconsidered (Table 2,
Figure 3). Additionally, the rotatory powers of the isolated and
synthesized nonenolides 1-4 demonstrate, that the stereogenic
center at position 9 dictates the direction of the optical rotation,
since all (9R)-configurated analogs of putaminoxin (1) showed a
negative optical rotatory power, whereas (9S)-configurated
analogs showed a positive optical rotatory power (Table 3).
Table 3: Optical rotatory powers of different nonenolides.
Author
Molecule
Optical rotatory power
1
ꢁꢂ
Evidente et al.
(5S,6E,9?)-1
[α]ꢀ = -23.1,
(c = 1.6), CHCl3
3
ꢁꢂ
Sabitha et al.
(5S,6E,9R)-1
(5S,6E,9R)-1
[α]ꢀ = -25.2,
(
c = 1), CHCl3
ꢁꢂ
Dickmann et al.
Dickmann et al.
Bisterfeld
[α] = -13.4,
ꢀ
(
c = 0.7), CHCl3
ꢁꢂ
(5S,6E,9S)-1
[α] = +22.3,
ꢀ
(
c = 0.7), CHCl3
ꢁꢃ
and (5S,6E,9R)-2
and (5S,6E,9S)-2
(5S,6E,9R)-3
[α] = -25.6,
ꢀ
1
4
Holec et al.
(
c = 1), CHCl3
ꢁꢃ
Bisterfeld
Holec et al.
[α] = +16.1,
ꢀ
1
4
(
c = 1), CHCl3
6
ꢁꢂ
Götz et al.
[α]ꢀ = -29.0,
(c = 0.35), CHCl3
ꢁ
ꢄ
Fuchser
Zeeck
and (5R,6E,9R)-4
[α] = -43.8,
ꢀ
2
2
1
3
Figure 3: Comparison of the C NMR shifts of nonenolides with the
(
c = 0.3), MeOH
1
3
reported C NMR shifts of the isolated compound (5S,6E,9?)-1 by Evidente
1
et al. and a structural proposition on the basis of observed correlations. A)
1
3
13
Comparison of C NMR shifts of (5S,6E,9?)-1 with the C NMR shifts of
Therefore, it is likely that the originally isolated compound by
3, 6, 14
13
synthesized (5S,6E,9R) configurated nonenolides.
B) Comparison of
C
Evidente et al. is indeed (9R) configurated as well. By
1
3
NMR shifts of (5S,6E,9?)-1 with the C NMR shifts of synthesized
13
comparison of the C NMR data of our synthesized compounds
14, 22
(
5S,6E,9S) and isolated (5R,6E,9R) configurated nonenolides.
and those of putaminoxin B/D (2), hypocreolide A (3), and
aspinolide A (4) one can assume though, that putaminoxin (1)
may not be (5S)-, but (5R)-configurated instead. Taking these
observations into consideration, we come to the conclusion, that
the originally isolated putaminoxin from Evidente et al. might be
(5R,6E,9R)-configurated, instead of (5S,6E,9R). The specific
rotation of the newly synthesized enantiomer (5S,6E,9S)-1 of
+22.3 supports this argument, as it is the opposite of the reported
3
. Conclusion:
In the present work we were able to realize the
chemoenzymatic total synthesis of the phytotoxic compound
putaminoxin (1) and its (5S,6E,9S)-diastereomer [(5S,6E,9S)-1]
by employing alcohol dehydrogenases for the generation of both
stereogenic centers. The oxidative kinetic resolution of aliphatic
homoallylic alcohols revealed that the alcohol dehydrogenase
from Thermoanaerobacter brockii is capable of distinguishing
between the n-pentyl and n-pentenyl residue of the challenging
substrate hept-1-en-4-ol (14), giving access to the valuable
building block (S)-14 with a cheap and easy reaction setup and
under mild reaction conditions. The Tb-ADH is most suited for
the conversion of oct-1-en-4-ol (13) and non-1-en-4-ol (5)
though, combining short reaction times and excellent
selectivities. Longer aliphatic homoallylic alcohols can still be
converted with excellent selectivities, but the reaction time
13
-23.1 by Evidente et al. (Table 3) and the C NMR data of these
enantiomers are consistent as well (Table 2, Figure 3, B). A more
13
concise comparison of C NMR data of closely related
13
nonenolides 2-4 revealed even more inconsistencies in the
C
NMR data of these intriguing compounds (see supplementary
material), emphasizing the importance of total synthesis of these
biologically active lactones.

6
Tetrahedron +
4
. Experimental Section:
NADP (300 µM) in KP -buffer (50 mM, pH 7.0, 1 mM MgCl )
i 2
ACCEPTED MANUSCRIPT
to a final volume of 50 mL. The reaction was shaken at 30 °C
and 130 rpm and after 0 h, 1 h, 3 h, 4 h, 5 h, 6 h, 8 h and 24 h a
4
.1. General:
2
00 µL sample was taken. The sample was extracted by addition
Optical rotations were measured in CHCl3 using
PerkinElmer 341 and Krüpps Optotronic polarimeter at the
sodium ᴅ-line using a cell with 100 mm path length. Absorbance
a
of 500 µL MTBE, containing 2.5 mM of 1-Hexanol as internal
standard, and measured via gas chromatography over chiral
stationary phase (individual programs are as stated in the
supplementary material).
measurements
were
conducted
using
an
UV-160
spectrophotometer. IR data were recorded on a PerkinElmer
SpectrumOne and SpectrumTwo instrument as a thin film, and
Analytical Scale:
−1
1
13
absorbance frequencies are reported in cm . H- and C NMR
spectra were recorded on an Avance/DRX 600 NMR
A 5 mL glass vial was charged with homoallylic alcohol (0.02
+
mmol), acetone (5 Vol.-%), Tb-ADH (1 U), and NADP (300
µM) in KP -buffer (50 mM, pH 7.0, 1 mM MgCl ) to a final
volume of 2 mL. The reaction mixture was briefly vortexed and
distributed into 200 µL aliquots. The aliquots were shaken at 30
spectrometer (Bruker) at ambient temperature in CDCl at 600
3
and 151 MHz, respectively. The chemical shifts are given in ppm
i
2
1
relative to the solvent signal [ H: δ (CHCl ) = 7.26 ppm] and
3
13
[
C: δ (CDCl ) = 77.16 ppm, for the centerline of the CDCl3
3
°
C and 130 rpm and after 0 h, 1 h, 3 h, 4 h, 5 h, 6 h, 8 h and 24 h
triplet]. NMR signals were assigned by means of COSY and
HSQC experiments. GC/MS measurements were carried out
using a Hewlett-Packard HP 6890 Series GC System/Hewlett-
Packard 5973 mass selective detector or Thermo Scientific ISQ
QD/Thermo Scientific Trace 1310 GC System. HRMS (ESI,
positive ion) was performed by the analytical service of the
Forschungszentrum Jülich (ZEA-3). GC analysis was performed
on a Trace GC Ultra gas chromatograph (Thermo Finnigan,
Thermo Scientific) or a GC-17A gas chromatograph (Shimadzu)
with a flame ionization detector (FID). The Trace GC Ultra gas
chromatograph was equipped with a FS-Lipodex G (25 m × 0.25
mm, Macherey Nagel)., FS-Hydrodex-βTBDAc (25 m × 0.25
mm, Macherey Nagel) or FS-Hydrodex-β3p column (25 m × 0.25
mm, Macherey Nagel). The injector and detector were operated
a 200 µL sample was extracted by addition of 500 µL MTBE,
containing 2.5 mM of 1-hexanol as internal standard, and
measured via gas chromatography over chiral stationary phase
(individual programs are as stated in the supplementary material).
Ethyl (5S)-5-(tetrahydropyran-2´´-yloxy)-hept-6-enoate
[(S)-9]: To a solution of allylic alcohol (S)-24 (400 mg, 2.00
mmol.) in dry CH Cl2 (38 mL) were added pyridinium p-
2
toluenesulfonate (60.0 mg, 240 ꢀmol, PPTS) and 3,4-dihydro-
2
1
H-pyran (550 ꢀL, 6 mmol). The mixture was stirred at rt for
4 h and then quenched by the addition of saturated aqueous
NaHCO -solution (60 mL) and extracted with Et O (3 × 30 mL).
3
2
The combined organic layer was dried over MgSO , filtered over
4
a pad of Celite, and concentrated under reduced pressure.
Chromatography of the crude product on silica gel (petroleum
ether/EtOAc, 90:10) afforded the product (S)-7 (533 mg, 1.88
mmol, 94%) as a colorless oil. The analytical data were in
at 250 °C, and H was used as the carrier gas at a flow rate of 30
2
−1
mL·min . The GC-17A gas chromatograph was equipped with a
CP-Chirasil-Dex CB column (25 m × 0.25 mm, Varian), and He
−1
21
was used as the carrier gas at a flow rate of 1.3 mL·min .
Substances were dissolved in MTBE and analyzed according to
the given temperature protocols. Chiral stationary-phase HPLC
measurements were performed on a Dionex system equipped
with a pump with a gradient mixer and devolatilizer, including a
WPS-3000TSL autosampler and a DAD-3000 UV-detector. A
Lux Amylose-1 column (250 mm × 4.6 mm, Phenomenex) and a
mixture of n-heptane/2-propanol (99.8:0.2) as solvent were used,
accordance with the literature. Compound (S)-7 was due to the
THP-protecting group obtained as a diastereomeric mixture
indicated as A and B; dr (A:B) = 1:1.2. R 0.28 (petroleum
f
ꢁꢃ
ether/EtOAc, 9:1); [α]ꢀ = -13.8 (c 1.0, CHCl ); FT-IR ṽ 2942,
3
max
2
867, 1735, 1445, 1379, 1240, 1175, 1129, 1076, 1021, 967, 923,
-1 1
8
69, 813 cm ; H NMR (600 MHz, CDCl ) δ[ppm]= 1.25 (t,
3
AB AB AB
3
J
= 7.1 Hz, 6H, 2-H ´), 1.45-1.90 (m, 20H, 3-H , 4-H , 3-
2
´,1´
AB
AB AB AB
H ´´, 4-H ´´, 5-H ´´), 2.29-2.35 (m, 4H, 2-H ), 3.43-3.54
−1
AB
AB
applying a flow rate of 0.5 mL·min at room temperature. Flash
column chromatography was performed on silica gel 60, particle
size 40−63 ꢀm (230−240 mesh).
(m, 2H, 6-Hb ´´), 3.85-3.91 (m, 2H, 6-H ´´), 4.05-4.13 (m,
a
AB
AB
3
B
6
H, 5-H , 1-H ´), 4.65 (t, J
= 3.6 Hz, 1H, 2-H ´´), 4.69
2
B´´,3B´´
3
A
3
(
t, J
= 3.8 Hz, 1H, 2-H ´´), 5.11 (dt, J
= 10.5 Hz,
7aA,6A
B
2
A´´,3A´´
2
A
J7aA,7bA = 1.4 Hz), 1H, 7-H ), 5.19 (m, 2H, 7-H ), 5.23 (dt,
J7bA,6A = 17.3 Hz, J
J6B,7bB = 17.6 Hz, J
4
.2. Chemicals:
a
3
3
2
A
= 1.4 Hz, 1H, 7-H ), 5.62 (ddd,
7bA,7aA
b
3
3
B
All reagents were used as purchased from commercial
= 10.0 Hz, J
= 17.3 Hz, J
= 7.7 Hz, 1H, 6-H ),
= 10.5 Hz, J
6B,7aB
6B,5B
3
3
3
suppliers without further purification. The nicotinamide cofactor
5.86 (ddd, J
= 6.8 Hz,
6
A,7bA
6A,7aA
6A,5A
+
A
13
NADP was a generous gift from Codexis. Petroleum ether,
1H, 6-H ); C NMR (151 MHz, CDCl ) δ[ppm]= 14.40 (C-2´),
3
diethyl ether, n-pentane, and EtOAc for column chromatography
were distilled before usage. The synthesis of ethyl 5-oxo-6-
heptenoate (23) and ethyl (S)-5-hydroxyhept-6-enoate [(S)-24]
were carried out according to a published procedure by Fischer
19.71/19.82/20.67/21.19/25.60/25.72/30.88/31.03/34.40/35.09
(C-3, C-4, C-3´´, C-4´´, C-5´´), 34.04/34.35 (C-2), 60.36/60.40
(C-1´), 62.44/62.74 (C-6´´), 76.17/77.77 (C-5), 95.17/97.92 (C-
2´´), 115.20/117.5 (C-7), 138.36/139.50 (C-6), 173.72 (C-1);
EIMS m/z 55, 70, 85, 98, 155.
21
and Pietruszka.
4
.3. Enzyme Production:
(S)-Hept-1-en-4-ol (Brown allylation) [(S)-14]: A solution of
5
mL (+)-B-Allyldiisopinocampheylborane (5 mmol, 1 M in n-
For heterologous protein expression, the Escherichia coli
pentane) in 5 mL dry Et O was stirred and cooled to -78 °C and a
solution of butanal (20) (403 µL, 4.5 mmol) in 500 µL dry Et O
was slowly added. The mixture was stirred for 1 h after which the
reaction was warmed to rt. 1.6 mL of a 3 M NaOH solution and
6
refluxed for 1 h. The organic phase was separated, washed with
H O and brine, dried over MgSO , filtered and concentrated
under reduced pressure. Chromatography of the crude product
over silica gel (n-pentane/Et O, 87:13) afforded product (S)-14 as
colorless oil (118 mg, 1,03 mmol, 23%, ee 95%). The analytical
2
strain BL21 (DE3) was used. Expression, purification, and
quantification of production and activity measurements of Lb-
2
23
ADH and Tb-ADH, were performed as described earlier.
.4. Oxidative kinetic resolution of homoallylic alcohols:
Preparative Scale:
A 250 mL Erlenmeyer flask was charged with homoallylic
alcohol (0.5 mmol), acetone (5 Vol.-%), Tb-ADH (25 U), and
50 µL of H O (30% (w/w)) were added and the reaction was
4
2 2
2
4
2

7
2
4
data were in accordance with those reported in
A
th
C
e l
C
ite
E
ra
P
tu
T
re.ED
R
MA
w
N
ere
U
di
S
ss
C
ol
R
ve
I
d
P
in 5 mL dry CH Cl and the reaction was heated
2 2
T
f
ꢁꢃ
0
.21 (n-pentane/Et O, 87:13); [α] = -12.3 (c 1.0, CHCl ); FT-
IR ṽmax 3355, 2964, 2929, 2874, 1643, 995, 912 cm ; H NMR
600 MHz, CDCl ) δ[ppm]= 0.93 (t, J = 7.2 Hz, 3H, 7-H), 1.32
to 40 °C and stirred for 24 h. The reaction mixture was filtrated
through a pad of celite and concentrated under reduced pressure.
Chromatography over silica gel (n-pentane/Et O, 94:6) afforded
2
ꢀ
3
-
1
1
3
(
3
7,6
2
–
1
1.58 (m, 5H, 4-OH, 5-H, 6-H), 2.14 (m, 1H, 3-H ), 2.30 (m,
the product (4S,6E,9S)-21 (116 mg, 0.28 mmol, 83%) as a
a
ꢁꢃ
H, 3-H ), 3.66 (m, 1H, 4-H), 5.09-5.16 (m, 2H, 1-H), 5.79-5.87
colorless oil. R 0.23 (n-pentane/Et O, 94:6); [α] = -19.5 (c 0.7,
b
f
2
ꢀ
13
(m, 1H, 2-H); C NMR (151 MHz, CDCl ) δ[ppm]= 14.23 (C-7),
CHCl ); FT-IR ṽ
cm ; H NMR (600 MHz, CDCl ) δ[ppm]= 0.82-0.93 (m, 6H , 1-
2958, 2873, 1714, 1451, 1270, 1111, 710
3
3
max
-
1
1
1
1
9.01 (C-6), 39.14 (C-5), 42.11 (C-3), 70.55 (C-4), 118.21 (C-1),
35.06 (C-2). EIMS m/z 55, 71, 73.
3
H, 12-H), 1.23-1.45 (m, 4H, 2-H, 11-H), 1.51-1.70 (m, 4H, 3-H,
1
0-H), 2.33-2.53 (m, 4H,5-H, 8-H), 5.12 (m, 2H, 4-H, 9-H), 5.53
(
S)-Hept-1-en-4-ol (oxidative kinetic resolution) [(S)-14]: A
(m, 2H, 6-H, 7-H), 7.40-7.45 (m, 4H, 4-H ´,4-H ´, 4-H ´´, 4-
a
b
a
1
3
(
L Erlenmeyer flask was charged with substrate 14 (500 µL+,
H
´´), 7.52-7.56 (m, 2H, 5-H´,5-H´´), 8.00-8.04 (m, 4H, 3-H ´, 3-
a
13
b
,68 mmol), acetone (5 Vol.-%), Tb-ADH (119 U) and NADP
H ´, 3-H ´´,3-H ´´); C NMR (151 MHz, CDCl ) δ[ppm]= 14.07
b
a
b
3
300 µM) in KP -buffer (50 mM, pH 7.0, 1 mM MgCl ) to a final
i
2
(C-1, C-12), 18.73 (C-2, C-11), 35.76 (C-3, C-10), 37.56 (C-5, C-
volume of 250 mL. The reaction was stirred at 30 °C and 130
rpm for 3.5 h after it was stopped and extracted by addition of
CH Cl . Combined organic layers were dried over MgSO ,
8
1
1
), 74.19 (C-4, C-8), 128.43 (C-4´,C-4´´), 128.60 (C-6, C-7),
29.66 (C-3´,C-3´´), 130.88 (C-2´,C-2´´), 132.86 (C-5´,C-5´´)
2
2
4
66.31 (C-1´,C-1´´); EIMS m/z 77, 105; HRMS m/z 409.2374
filtered
and
concentrated
under
reduced
pressure.
+
(calcd for C H O 409.2373).
26
33
4
Chromatography of the crude product on silica gel (n-
pentane/Et O, 87:13) afforded product (S)-14 as colorless oil
(4R,6E,9R)-Dodec-6-ene-4,9-diyl dibenzoate [(4R,6E,9R)-
2
(
102.7 mg, 0,90 mmol, 24%, ee 94%) The analytical data were in
21]: 151 mg (0.69 mmol) of the benzoyl-protected (R)-Hept-1-
en-4-ol (R)-8 and 30 mg (0.03 mmol) Grubbs 2 generation
nd
accordance to those stated for compound (S)-14 (see above).
catalyst were dissolved in 5 mL dry CH Cl and the reaction was
heated to 40 °C and stirred for 24 h. The reaction mixture was
filtrated through a pad of celite and concentrated under reduced
2
2
(
S)-Hept-1-en-4-yl benzoate [(S)-8]: To a stirring solution of
(
S)-14 (150 mg, 1.31 mmol) in 5 mL dry CH Cl , benzoyl
chloride (198 µL, 1.71 mmol) and pyridine (179 µL, 2.23 mmol)
were slowly added, and the reaction was kept stirring at rt for
2
2
pressure. Chromatography over silica gel (n-pentane/Et O, 94:6)
2
afforded the product (4R,6E,9R)-21 (116 mg, 0.30 mmol, 86%)
2
4 h. The reaction was stopped by addition of saturated
ꢁꢃ
as a colorless oil. [α]ꢀ = +28.3 (c 1, CHCl ); The analytical data
3
Na HCO -solution and the aqueous phase was extracted with
2
3
are in accordance to those of (4S,6E,9S)-21 (see above).
CH Cl . Combined organic layers were washed with saturated
2
2
copper(II) sulfate-solution and brine, afterwards dried over
Ethyl (5S,6E,9R)-9-benzoyl-5-(tetrahydropyran-2´´-yloxy)-
dodec-6-enoate [(5S,6E,9R)-25]: To a solution of 100 mg (0.24
mmol) of (4R,6E,9R)-21 and 121 mg (0.47 mmol) (S)-7 in 5 mL
MgSO and filtrated. The crude solution was concentrated under
4
reduced pressure before chromatography over silica gel (n-
2nd
pentane/Et O, 98:2) yielded the product (S)-8 (254 mg, 1.16
dry CH Cl , 22 mg (0.02 mmol) of Grubbs
generation catalyst
2
2
2
mmol, 89 %) as a colorless oil. R 0.23 (n-pentane/Et O, 98:2);
were added, and the reaction stirred at 40 °C for 3 d. The reaction
was filtrated through a pad of celite and concentrated under
f
2
ꢁꢃ
[
2
α] = +15.7° (c 1.0, CHCl , 95% ee); FT-IR ṽ 2959, 2929,
ꢀ
3
max
-
1 1
866, 1713, 1269, 1111, 710 cm ; H NMR (600 MHz, CDCl )
reduced
pressure.
Chromatography
over
silica
gel
3
3
δ[ppm]= 0.93 (t, J1 = 7.4 Hz, 3H, 7-H), 1.34-1.49 (m, 2H, 6-H),
(n-pentane/Et O, 93:7→83:17→75:25) yielded the product
,2
2
3
3
1
2
5
.59-1.75 (m, 2H, 5-H), 2.45 (dd, J = 7.2 Hz, J = 5.9 Hz,
(5S,6E,9R)-25 as a colorless oil (90 mg, 0.20 mmol, 41%).
Unreacted substrates (4R,6E,9R)-21 (44 mg, 0.11 mmol, 44%)
and (S)-7 (34 mg, 0.13 mmol, 28%) could be reisolated. Due to
the THP-protecting group compound (5S,6E,9R)-25 was obtained
3
,2
3,4
2
3
H, 3-H), 5.06 (dt, J
= 1.6 Hz, J = 10.1Hz, 2H, 1-Ha),
= 17.2Hz, 2H, 1-H ), 5.78-5.87
1b,2 b
3
1a,1b
1a,2
2
3
.11 (dt J
= 1.6 Hz, J
1b,1a
3
3
(ddt, J2 = 17.2Hz, J = 10.1Hz, J = 7.2Hz, 1H, 2-H), 7.40-
,1b 2,1a 2,3
1
7
.45 (m, 2H, 4-H ´,4-H ´), 7.52-7.56 (m, 1H, 5-H´), 8.00-8.04
as a diastereomeric mixture indicated as A and B; dr = 1:1.2 ( H
a
b
1
3
ꢁꢃ
(m, 2H, 3-H ´, 3-H ´); C NMR (151 MHz, CDCl ) δ[ppm]=
NMR). R 0.33 (n-pentane/Et O, 75:25); [α] = -9.5 (c 1,
a
b
3
f
2
ꢀ
-
1
1
4.11 (C-7), 18.77 (C-6), 36.01 (C-5), 38,89 (C-5), 73.98 (C-4),
17.87 (C-1), 128.45 (C-4´), 129.69 (C-3´) 130.94 (C-2´), 132.87
CHCl ); FT-IR ṽ 2938, 2873, 1716, 1272, 1112, 1022, 712 cm
3 max
1 1 3
; H NMR (600 MHz, CDCl ) δ[ppm]= 0.93 (t, J
= 7.4 Hz,
3
12,11
AB
3
AB
(C-5´), 133.89 (C-2), 166.38 (C-1´); EIMS m/z 77, 105.
6H, 12-H ), 1.24 (t, J = 7.1 Hz, 6H, 2-H ´), 1.28-1.76 (m,
2
´,1´
AB
AB
AB
AB
AB
AB
AB´´
2
2
8H, 3-H , 4-H , 10-H , 11-H , 3-H 3´´,4-H ´´, 5-H ),
(R)-Hept-1-en-4-yl benzoate [(R)-8]: 116 mg (0.95 mmol)
AB 3
.25 (m, 4H, 2-H ), 2.43 (q, J = 5.8 Hz, J = 5.6 Hz, 4H, 8-
8
,9
8,7
benzoic acid were dissolved in 5 mL dry THF and 251 mg (0.96
mmol) triphenyl phosphine and 100 mg (0.88 mmol) of (S)-hept-
AB
2
3
H ), 3.31-3.36 (dt, J
= 10.5 Hz, J
= 4.5 Hz,
6bB´´,5aB´´
2
6
bB´´,6aB´´
3
B
J6bB´´,5bB´´3= 4.5 Hz, 1H, 6-H ´´), 3.36-3.41 (dt, 1H, J6bA´´,6aA´´=
b
1
0
-en-4-ol (S)-14 were added before the reaction was cooled to
°C. 188 µL (0.95 mmol) of DIAD were slowly added to the
3
A
b
1
3
0.5 Hz, J
.78 (ddd, J6
= 4.8 Hz, J
=4.8 Hz, 6-H ´´) 3.72-
6
bA´´,5aA´´
6bA´´,5bA´´
2
3
3
= 11.5 Hz, J
aB´´,6bB´´
=8.5 Hz, J
= 11.5 Hz, J
=3.3
=
6aB´´,5bB´´
6aB´´,5aB´´
stirring solution which was then left to warm to rt. After 4.5 h the
B
2
3
Hz, 1H, 6-H ´´), 3.78-3.83 ( dt, J
5
5
Hz, 1H, 2-H ´´), 4.64 (t, J
2
6
a
6aA´´,6bA´´
6aA´´,5bA´´
reaction was stopped by addition of saturated NaHCO -solution
3
A
A
3
.6 Hz, J
=5.6 Hz, 1H, 6-H ´´), 4.00-4.04 (m, 2H, 5-H ,
aA´´,5aA´´ a
AB 3
6
and the organic phase was washed with brine and water. The
crude product was concentrated under reduced pressure and then
B
3
-H ), 4.11 (q, J = 7.1 Hz, 4H, 1-H ´), 4.47 (t, J = 3.6
1´,2´
2B´´,3B´´
B
3
A
= 3.7 Hz, 1H, 2-H ´´), 5.18 (m,
2
A´´,3A´´
purified via chromatography over silica gel (n-pentane/Et O,
A
B
3
3
2
H, 9-H , 9-H ), 5.32 (dd, J
= 15.5 Hz, J
= 8.3 Hz, 1H,
6B,7B
6B, 5B
9
8
8:2) to yield the final product (R)-8 as colorless oil (159 mg,
3%, 87 % ee). The configurational change of the stereogenic
B
A
AB
-H ), 5.54-5.70 (m, 3H, 6-H , 7-H ), 7.40-7.45 (m, 4H, 4-
Ha ´´´, 4-Hb ´´´), 7.52-7.57 (m, 2H, 5-H ´´´), 8.01-8.05 (m,
H, 3-Ha ´´´, 3-Hb ´´´). C NMR (151 MHz, CDCl ) δ[ppm]=
4.11 (C-12), 14.40 (C-2´), 18.75/18.76 (C-11), 19.66/19.81 (C-
´´), 20.78/21.27 (C-3), 25.60/25.67, 30.83/30.94, 35.31 (C-10,
AB
AB
AB
center was confirmed via HPLC over a chiral stationary phase
AB
AB
13
4
1
4
3
(
[
see supplementary material). R 0.23 (n-pentane/Et O, 98:2);
α] = -17.4 (c 1.0, CHCl , 94% ee); The analytical data are in
ꢀ
3
f
2
ꢁꢃ
accordance to those reported for compound (S)-6 (see above).
C-3´´,C-5´´) , 34.36/34.38 (C-2), 36.02/36.20 (C-4), 37.27/37.36
(C-8), 60.31/60.34 (C-1´) 62.48/62.60 (C-6´´) 73.87/74.17 (C-9),
(4S,6E,9S)-Dodec-6-ene-4,9-diyl dibenzoate [(4S,6E,9S)-
B
A
2
4
1]: 151 mg (0.69 mmol) of the benzoyl-protected (S)-Hept-1-en-
-ol (S)-8 and 30 mg (0.03 mmol) Grubbs 2 generation catalyst
75.39/76.76 (C-5), 95.04 (C-2 ´´), 97.52 (C-2 ´´) 126.64 (C-7),
128.43/128.47 (C-4´´´), 129.36/129.70 (C-3´´´), 130.83/130.94
nd

8
Tetrahedron
(
C-2´´´), 132.86/132.93 (C-5´´´), 133.6
A
4/
C
13
C
4.6
E
8
PT(C
E
-
D
+
6), MAun
N
de
U
r r
S
ed
C
uc
R
edIPpr
T
essure. Chromatography of the crude product (n-
1
1
4
66.22/166.28 (C-1´´´), 173.62/173.70 (C-1); EIMS m/z 77, 105,
64, 341; HRMS m/z 469.2560 (calcd for C H O Na
69.2561).
pentane/Et O, 65:35) yielded 12 mg (0.06 mmol, 57%) of the
2
ꢁꢂ
final product (1). R 0.16 (n-pentane/Et O, 65:35); [α] = -13.4
26
38
6
f
2
ꢀ
(c 0.7, CHCl ); FT-IR ṽ 3441, 2963, 2931, 2874, 1727, 1442,
3 max
-1 1
1
226, 1158 cm ; H NMR (600 MHz, CDCl ) δ[ppm]= 0.93 (t,
3
Ethyl (5S,6E,9S)-9-benzoyl-5-(tetrahydropyran-2´´-yloxy)-
3
J12,11 = 7.4 Hz, 3H, 12-H), 1.32-1.60 (m, 4H, 4-H , 10-H , 11-H),
a
a
docec-6-enoate [(5S,6E,9S)-25]: To a solution of 94 mg (0.22
mmol) of (4S,6E,9S)-21 and 124 mg (0.48 mmol) (S)-7 in 5 mL
dry CH Cl , 22 mg (0.02 mmol) of Grubbs
Catalyst were added, and the reaction stirred at 40 °C for 3 d. The
reaction was filtrated through a pad of celite and concentrated
under reduced pressure. Chromatography over silica gel (n-
1
.62-1.70 (m, 2H, 3-H , 10-H ), 1.75-1.91 (br s, 1H, 5-OH), 1.93-
a
b
2nd
2.16 (m, 4H, 2-H
H ), 4.44 (br s, 1H, 5-H), 5.04 (ddd, J = 10.5 Hz, 8.3 Hz, 4.7 Hz,
b
a
, 3-H , 4-H , 8-H ), 2.40-2.50 (m, 2H, 2-H , 8-
b b a b
Generation
2
2
1
1
H, 9-H),5.44 (dd, J = 15.5 Hz, 1.7 Hz, 1H, 6-H),5.55 (dddd, J =
5,5 Hz, 10.5 Hz, 4.9 Hz, 2.4 Hz, 1H, 7-H); C NMR (151 MHz,
13
CDCl ) δ[ppm]= 14.01 (C-12), 18.01 (C-3), 19.31 (C-11), 35.91
3
pentane/Et O, 92:8→8:2→7:3) yielded the product (5S,6E,9S)-25
as a colorless oil (47 mg, 0.11 mmol, 23%). Unreacted substrates
2
(C-2), 36,45 (C-10), 36.84 (C-4), 40,91 (C-8), 68.60 (C-5), 76.60
(C-9), 126.42 (C-7), 136.82 (C-6), 176.79 (C-1); HRMS m/z
(4S,6E,9S)-21 (59 mg, 0.14 mmol, 63%) and (S)-7 (60 mg, 0.23
+
2
13.1486 (calcd for C H O 213.1485).
12
21
3
mmol, 48%) could be reisolated. Due to the THP-protecting
group compound (5S,6E,9S)-25 was obtained as a diastereomeric
(5S,6E,9S)-5-Hydroxy-9-propyl-6-nonen-9-olide
[(5S,6E,9S)-1]: 9 mg LiOH (0.38 mmol) were added to a
solution of 35 mg (0.08 mmol) (5S,6E,9S)-25 in 8.1 mL of
1
mixture indicated as A and B; dr = 1:1.2 ( H NMR). R 0.37
f
ꢁ
ꢃ
(
2
n-pentane/Et O, 7:3); [α] = -17.0 (c 1, CHCl ); FT-IR ṽ
937, 2873, 1716, 1272, 1112, 1022, 713 cm ; H NMR (600
= 7.4 Hz, 6H, 12-H ), 1.24
t, J = 7.1 Hz, 6H, 2-H ´), 1.28-1.76 (m, 28H, 3-H , 4-H ,
0-H , 11-H , 3-H ´´,4-H ´´, 5-H ´´), 2.19 (m, 4H, 2-H ),
.44 (m, 4H, 8-H ), 3.37-3.45 (m, 2H 6-H ´´, 6-H ´´) 3.79-
2
ꢀ
3
max
-
1
1
THF:MeOH:H O (2:1:1) and heated to 60 °C for 24 h. The
2
3
AB
MHz, CDCl ) δ[ppm]= 0.93 (t, J
reaction was diluted with 25 mL of Et O washed with saturated
3
12,11
2
3
AB
AB
AB
(
KH PO -solution and brine. The organic phase was dried of
2
´,1´
2
4
AB
AB
AB
AB
AB
AB
1
2
3
6
5
MgSO and concentrated under reduced pressure. 8 mL dry THF
4
AB
A
B
was added to the crude product together with 71 µL (0.44 mmol)
2,4,6-trichlorobenzoyl chloride and 67 µL (0.48 mmol) of
triethylamine. The reaction mixture was stirred for 120 min at rt,
diluted with 20 mL dry toluene and then filtrated through a pad
of celite. 20 mL of dry toluene were added and the mixture was
slowly added to a refluxing solution of 67 mg (0.55 mmol)
DMAP in 57 mL dry toluene over the course of 2.5 h. The
reaction was stirred for another 30 min before it was allowed to
cool to rt. The mixture was quenched with 1M aqueous HCl and
b
b
A
B
3
3
.85 (m, 2H 6-H ´´, 6-H ´´), 3.98 (q, J
= 6.2 Hz, J
=
a
a
5A, 4aA
3
5A,6A
A
3
.2 Hz, 1H, 5-H ), 4.03 (q J
= 6.5 Hz, J
= 6.5 Hz, 1H,
5B,4aB
3
5
B,6B
B
3
AB
-H ) 4.10 (q, J = 7.1 Hz, 4H, 1-H ´), 4.57 (t, J = 3.7
1
´,2´
2B´´,3B´´
B
3
A
Hz, 1H, 2-H ´´), 4.64 (t, J
2
6
= 3.7 Hz, 1H, 2-H ´´), 5.18 (m,
2
A´´,3A´´
A
B
3
3
H, 9-H , 9-H ), 5.31 (dd, J
= 15.5 Hz, J
= 8.3 Hz, 1H,
6
B,7B
6B, 5B
B
3
3
-H ), 5.53-5.59 (dd, J
= 15.5 Hz, J
=7.1 Hz, 1H, 6-
6A,5A
AB
6
A,7A
A
AB
H ), 5.64 (m, 2H, 7-H ), 7.40-7.45 (m, 4H, 4-H ´´´, 4-
a
AB
AB
Hb ´´´), 7.52-7.57 (m, 2H, 5-H ´´´), 8.01-8.05 (m, 4H, 3-
Ha ´´´, 3-Hb ´´´). C NMR (151 MHz, CDCl ) δ[ppm]= 14.12
AB
AB
13
washed with saturated NaHCO -solution and brine. The
3
3
(
2
C-12), 14.39 (C-2´), 18.76 (C-11), 19.77/19.82 (C-4´´),
0.70/21.20 (C-3), 25.59/25.70, 30.86/30.98, 35.24 (C-10, C-3´´,
C-5´´), 34.28/34.31 (C-2), 36.01/36.04 (C-4), 37.26/37.50 (C-8),
0.31/60.34 (C-1´), 62.46/62.68 (C-6´´), 73.78/74.00 (C-9),
5.36/77.23 (C-5), 94.92/97.74 (C-2´´), 126.76/129.22 (C-7),
28.42/128.46 (C-4´´´), 129.67 (C-3´´´), 130.76/130.85 (C-2´´),
32.88/132.94 (C-5´´´), 133.59/134.74 (C-6), 166.24/166.28 (C-
combined organic layers were dried over MgSO4 and
concentrated under reduced pressure. The residue was dissolved
in 7 mL EtOH and treated with 27 mg PPTS (0.09 mmol) and 4.5
mg (0.02 mmol) p-TsOH monohydrate. The reaction mixture was
heated to 40 °C and stirred for 16 h before being quenched with
6
7
1
1
1
20 mL of ice water and saturated NaHCO -solution. The mixture
3
was extracted with EtOAc (3× 20 mL), dried over MgSO4,
filtered over a pad of celite and concentrated under reduced
´´´), 173.61/173.68 (C-1); EIMS m/z 77, 105, 164, 341; HRMS
+
m/z 469.2560 (calcd for C H O Na 469.2561).
pressure. Chromatography of the crude product (n-pentane/Et O,
26
38
6
2
6
0:40->50:50) yielded 11 mg (0.05 mmol, 66%) of the final
(
5S,6E,9R)-5-Hydroxy-9-propyl-6-nonen-9-olide [1]: 11.3
ꢁꢂ
product (5S,6E,9S)-1. R 0.19 (n-pentane/Et O, 50:50); [α] =
+
1
0
OH, 10-H , 11-H), 1.62-1.70 (m, 1H, 10-H ), 1.86-1.97 (m, 3H,
3
2
f
2
ꢀ
mg LiOH (0.47 mmol) were added to a solution of 44 mg (0.10
mmol) (5S,6E,9R)-25 in 10.2 mL of THF:MeOH:H O (2:1:1) and
heated to 60 °C for 48 h. The reaction was diluted with 30 mL of
Et O washed with saturated KH PO -solution and brine. The
organic phase was dried over MgSO and concentrated under
reduced pressure. 12 mL dry THF was added to the crude product
together with 86 µL (0.55 mmol) 2,4,6-trichlorobenzoyl chloride
and 84 µL (0.60 mmol) of triethylamine. The reaction mixture
was stirred for 60 min at rt, diluted with 25 mL dry toluene and
then filtrated through a pad of celite. 40 mL of dry toluene were
added and the mixture was slowly added to a refluxing solution
of 86 mg (0.69 mmol) DMAP in 50 mL dry toluene over the
course of 2.5 h. The reaction was stirred for another 30 min
before it was allowed to cool to rt. The mixture was quenched
with 1 M aqueous HCl and washed with saturated NaHCO3-
solution and brine. The combined organic layers were dried over
22.3 (c 0.7, CHCl ); FT-IR ṽ 3426, 2956, 2926, 2866, 1728,
3
max
2
-1
1
442, 1181, 1002 cm ; H NMR (600 MHz, CDCl ) δ[ppm]=
3
3
.92 (t, J1 = 7.2 Hz, 3H, 12-H), 1.32-1.59 (m, 5H, 4-H , 5-
2,11
a
2
2
4
a
b
4
-H, 8-H ), 1.98-2.06 (m, 2H, 2-H , 4-H ), 2.35 (m, 1H, 8-H ),
a a b b
.44 (m, 1H, 2-H ), 4.01 (m, 1H, 5-H), 5.03 (m, 1H, 9-H), 5.32
b
13
(
(
(
(
dt, J = 15.0 Hz, 6.5 Hz, 1H, 6-H), 5.54 (m, 1H, 7-H); C NMR
151 MHz, CDCl ) δ[ppm]= 14.01 (C-12), 19.28 (C-11), 22.45
C-3), 35.82 (C-2), 36.58 (C-10), 38.88 (C-4), 40.51 (C-8), 74.28
C-5), 75.48 (C-9), 131.83 (C-7), 137.30 (C-6), 175.93 (C-1);
3
+
HRMS m/z 213.1485 (calcd for C H O 213.1485).
12
21
3
5
. Acknowledgments:
The authors thank the Fonds of the Chemical Industry
(scholarship to C.H.), the Ministry of Innovation, Science, and
Research of the German federal state of North Rhine-Westphalia,
the Heinrich Heine University Düsseldorf, and the
Forschungszentrum Jülich GmbH for their generous support of
our project. We gratefully acknowledge Birgit Henßen for
analytical support and Andreas Klein for his help with graphics.
MgSO and concentrated under reduced pressure. The residue
was dissolved in 12 mL EtOH and treated with 27 mg PPTS
4
(0.11 mmol) and 5.6 mg (0.03 mmol) p-TsOH monohydrate. The
reaction mixture was heated to 40 °C and stirred for 16 h before
being quenched with 40 mL of ice water and saturated NaHCO3-
solution. The mixture was extracted with EtOAc (3× 20 mL),
dried over MgSO , filtered over a pad of celite and concentrated
4

9
6
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