Alkoxyallene-based syntheses of preussin and its analogs and their cytotoxicity

Article abstract of DOI:10.1039/c8ob02645a

Short syntheses of oxa-preussin, racemic preussin and (-)-preussin are reported. Starting from a racemic 3-nonyl-substituted methoxyallene derivative, its lithiation and addition to phenylethanal provided the corresponding allenyl alcohol that was converted into two diastereomeric dihydrofuran derivatives by silver nitrate-catalyzed 5-endo-trig cyclization. The acid hydrolysis of the enol ether moiety gave heterocyclic ketones and subsequent highly stereoselective reductions with l-selectride furnished 2-benzyl-5-nonylfuran-3-ol derivatives in good overall yield. The major all-cis-diastereomer has the skeleton and relative configuration of preussin and is hence called oxa-preussin. An analogous sequence with the same allene, but an N-sulfonyl imine as the electrophile, finally led to racemic preussin. The stereoselectivities of the individual steps are discussed in detail. With an enantiopure 2-benzyl-5-nonylpyrrolidin-3-one intermediate the preparation of (-)-preussin with an enantiomeric ratio of >95:5 could be accomplished in a few steps. The sign of the optical rotation of this product finally proved the absolute configurations of its precursors and demonstrated that our chiral auxiliary-based route led to the antipode of the natural product. The cytotoxicity of several of the prepared heterocycles against MCF-7 tumor cells was investigated and five compounds, including racemic and enantiopure (-)-preussin, were identified as highly cytotoxic with IC₅₀ values in the range of 3-6 μM.

Full text of DOI:10.1039/c8ob02645a

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Page 1 of 20
Organic & Biomolecular Chemistry
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DOI: 10.1039/C8OB02645A
Alkoxyallene-Based Syntheses of Preussin and Analogs and Their Cytotoxicity
Arndt Hausherr,a Gerhard Siemeisterb and Hans-Ulrich Reissig*a
a Institut für Chemie und Biochemie, Freie Universität Berlin, Takustr. 3, 14195 Berlin (Germany)
E-mail: hans.reissig@chemie.fu-berlin.de
b Bayer AG, Research & Development, Pharmaceuticals Müllerstraße 178, 13353 Berlin (Germany)
Electronic supplementary information (ESI) available: Experimental details and spectra of all products.
Abstract: Short syntheses of oxa-preussin, racemic preussin and (-)-preussin are reported. Starting
from a racemic 3-nonyl-substituted methoxyallene derivative, its lithiation and addition to phenylethanal
provided the corresponding allenyl alcohol that was converted into two diastereomeric dihydrofuran
derivatives by silver nitrate-catalyzed 5-endo-trig cyclization. The acidic hydrolysis of the enol ether
moiety gave heterocyclic ketones and subsequent highly stereoselective reductions with L-Selectride
furnished 2-benzyl-5-nonylfuran-3-ol derivatives in good overall yield. The major all-cis-diastereomer
has the skeleton and relative configuration of preussin and is hence called oxa-preussin. An analogous
sequence with the same allene, but an N-sulfonyl imine as electrophile, finally led to racemic preussin.
The stereoselectivities of the individual steps are discussed in detail. With an enantiopure 2-benzyl-5-
nonylpyrrolidin-3-one intermediate the preparation of (-)-preussin with an enantiomeric ratio of >95:5
could be accomplished in a few steps. The sign of the optical rotation of this product finally proved the
absolute configurations of its precursors and demonstrated that our chiral auxiliary-based route led to
the antipode of the natural product. The cytotoxicity of several of the prepared heterocycles against
MCF-7 tumor cells was investigated and five compounds, including racemic and enantiopure (-)-
preussin, were identified as highly cytotoxic with IC50 values in the range of 3–6 μM.
Introduction
Compounds with hydroxylated pyrrolidine, pyrrolizidine or indolizidine skeletons are frequently occurring
natural products with many interesting biological activities.1 Applying suitably substituted alkoxyallenes2
and imines we developed short and stereoselective syntheses of the amino acid (-)-detoxinine3 and the
biologically active alkaloids (-)-anisomycin4 and (+)-codonopsinine.5 Employing an arabinose-derived
nitrone and the requisite alkoxyallenes the preparation of (+)-australine and (+)-casuarine6 or (-)-
hyacinthacine B4 and hyacinthacine C5 epimers7 could be achieved in a close collaboration with the Goti
research group (Fig. 1). As a related interesting target compound we planned to prepare the pyrrolidin-
3-ol natural product preussin and analogs such as its oxygen congener oxa-preussin.

Organic & Biomolecular Chemistry
Page 2 of 20
OH
OH
Me
OAc
View Article Online
DOI: 10.1039/C8OB02645A
HO
An
HO
CO2H
An
N
N
N
H
OH
H
Me
(-)-detoxinine
(-)-anisomycin
(+)-codonopsinine
An = 4-MeOC6H4
HO
OH
HO
OH
HO
OH
OH
OH
OH HO
N
N
N
Me
OH
(+)-casuarine
OH
OH
(-)-hyacinthacine B4
OH
(+)-australine
OH
OH
Non
Non
O
Non
N
N
Ph
Ph
Ph
Me
Me
(+)-preussin
(-)-preussin
oxa-preussin
Fig. 1 Hydroxylated pyrrolidine and pyrrolizidine natural products.
The naturally occurring (+)-preussin was isolated independently by two research groups either from the
fermentation broth of Aspergillus Ochraceus (ATCC 22947)8 or of a Preussia species.9 This pyrrolidin-
3-ol alkaloid has a broad activity spectrum against bacteria and filamentous fungi8,9 and 1997 Yoshida
et al. also described the cytotoxic activity against rat fibroblast cells 3Y1.10 Later Müller et al. reported
that the compound directly interferes in the cell cycle and induces apoptosis of human tumor cells.11 It
could be shown with different human cell lines that (+)-preussin inhibits in vitro the cyclin E kinase
(CDK2-cyclin E)12 with an IC50 value of ca. 0.5 μM by blocking the cell cycle progression into S phase.
It was also found that (+)-preussin inhibits the -1 programmed ribosomal frameshifting and virus
propargation.13 More recently, the closely related (+)-preussin B (a compound with a 5-heptyl instead of
a 5-nonyl substituent) was isolated from Simplicillium Ianosoniveum and its biosynthesis was
elucidated.14 In 2018 Kijjoa et al. isolated together with 14 other compounds a second close relative of
(+)-preussin from the marine sponge-associated fungus Aspergillus Candidus KUFA0062: preussin C
is the N-demethylated version of (+)-preussin.15 These authors found that preussin C has cytotoxic
effects against different tumor cell lines, but that the N-methyl group of (+)-preussin seems to be crucial
for antibiotic and high cytotoxic activity.
As consequence of these interesting biological activities and the moderate structural complexity
of this natural product, it became a popular target for the proof of new synthetic concepts or methods.
A large number of syntheses of (+)-preussin, a few of its antipode (-)-preussin and of the racemic
compound have been reported.16 Since most of these syntheses are based on chiral pool compounds,
there is still room for improved selectivity and flexibility with respect to an access to analogs. For the
envisioned preparation of compounds with the preussin skeleton we planned stereoselective reductions
of ketones of type A that should be available from the corresponding heterocyclic compounds B bearing
an enol ether moiety (Scheme 1). These 2,5-dihydrofuran or 2,5-dihydropyrrole derivatives B should be
prepared from the corresponding allenyl alcohols[17] or amines C, [18] respectively, by suitable 5-endo-
trig cyclizations.[19] Hence this analysis leads to starting materials such as lithiated 3-nonyl-substituted
alkoxyallenes E as crucial nucleophilic component and phenylethanal or its imine congener D as
electrophiles.

Page 3 of 20
Organic & Biomolecular Chemistry
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OH
O
OR
DOI: 10.1039/C8OB02645A
Non
Non
Non
X
X
X
Ph
Ph
Ph
A
B
preussin skeleton
Ph
OR
OR
Li
•
•
+
Non
Non
X
Ph
HX
D
C
E
R = Me or chiral auxiliary
X = O or N-Tos
Scheme 1 Retrosynthetic analysis of compounds with preussin skeleton leading via heterocyclic ketones A, enol ethers B, and
allenyl alcohols or allenyl amines C to nucleophilic building blocks E and electrophiles D.
The synthesis and lithiation of 3-alkyl-substituted alkoxyallenes and the addition to electrophiles had
been reported by our group in preceding publications.20 These model reactions revealed that only low
diastereoselectivities in additions of the axially chiral allenes to prochiral electrophiles are to be
expected.21 In addition, we also investigated the feasibility of this (3+2) route to five-membered
heterocycles using allenes with carbohydrate-derived auxiliaries at the oxygen, that led to highly enantio-
enriched pyrrolidine building blocks.22
Results and Discussion
First, the short synthesis of racemic oxa-preussin (X = O) is described. The axially chiral 3-nonyl-
substituted methoxyallene 120 was used in racemic form, lithiated with n-butyllithium under standard
conditions and treated with phenylethanal (2) at -80 °C. After warm-up and aqueous quench crude
allenyl alcohol 3 was quantitatively obtained (Scheme 2). The low diastereoselectivity in the addition
step was expected since in model reaction of lithiated 1 or related allenes with various electrophiles
similarly unselective additions were observed.21 The slim nonyl group of 1 seems to be too far away
from the C-C bond forming event to have strong influence on this step. Since allenyl alcohols such as 3
rapidly undergo decomposition, crude 3 was treated with 0.2 equivalents of silver nitrate in the presence
of potassium carbonate17d to furnish the expected 2,5-dihydrofuran derivative 4. The diastereomeric
ratio was slightly shifted in favor of the cis-compound (assignments after the next step). This is likely
due to a partial equilibration at the allenyl alcohol stage by silver(I) catalysis and a faster cyclization of
the pro-cis-allenyl alcohol 3 to furnish cis-4 in slight excess. This behavior has also been found and
explained in allenyl amine cyclizations.22 Intermediate 4 was not purified but directly hydrolyzed to furan-
3-one derivatives 5 obtained in 66% overall yield after column chromatography. This good yield
demonstrates the efficacy of each step of this approach to specifically substituted furan-3-ones. The
separation of the two isomers by HPLC provided cis-5 and trans-5 in 42% and 17% overall yield,
respectively.

Organic & Biomolecular Chemistry
Page 4 of 20
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DOI: 10.1039/C8OB02645A
1. nBuLi, THF
OMe
H
OMe
-40 °C, 30 min
•
•
Non
2.
Ph
Non
Ph
HO
O
2
3
1
-80 °C
to -30 °C
2 h
(d.r. 55:45)
O
0.2 equiv. AgNO3 acetonitrile
r.t., 12 h
2 equiv. K2CO3
Non
Non
O
Ph
Ph
OMe
5
cis-
2 M H2SO4/H2O
+
THF,
3.5 h
Non
O
O
Ph
4
66%
(cis:trans 70:30)
O
(d.r. 70:30)
5
trans-
overall yield after separation
cis- : 42%, trans- : 17%
5
5
Scheme 2 Synthesis of cis- and trans-furan-3-one 5 by lithiation of alkoxyallene 1 and addition to phenylethanal (2) followed by
silver nitrate-promoted cyclization and acidic hydrolysis of 4. (The shown formulas refer to the relative configurations; the
compounds are racemic mixtures).
In order to obtain oxa-preussin, the stereoselective reduction of ketones cis-5 and trans-5 were studied
(Scheme 3). Treatment of cis-5 with sodium borohydride in ethanol afforded a 3:1 mixture of the desired
all-cis-6 and r-2,t-3,c-5-6, that were isolated after purification in 77% and 18% yield, respectively.
Gratifyingly, the use of L-Selectride in tetrahydrofuran at -78 °C exclusively gave all-cis-6 in essentially
quantitative yield. A NOESY experiment confirmed the configurational assignment of all-cis-6. Starting
from alkoxyallene 1, the synthesis of oxa-preussin (all-cis-6) was accomplished in four steps with an
overall yield of 42%.
The reduction of trans-5 with L-Selectride was less selective furnishing a mixture of the two possible
diastereomers in 62% and 8% yield (Scheme 3). The depicted configurations are based on our
observations with related pyrrolidin-3-ones showing that in trans-compounds the substituent at C-5 has
a stronger influence on the reduction than the C-3 substituent.21 The r-2,t-3,t-5 configuration of the major
diastereomer of 6 is thus very likely.
OH
O
OH
NaBH4, EtOH
r.t., 14 h
+
Non
Non
r-2,t-3,c-5-
18%
Non
77%
O
O
O
Ph
6
Ph
Ph
5
6
cis-
all-cis-
OH
O
L-Selectride, THF
-78 °C, 3 h
Non
Non
O
O
Ph
Ph
6
all-cis-
5
cis-
99%
oxa-preussin
OH
O
OH
L-Selectride, THF
-78 °C, 3 h
+
Non
8%
Non
Non
O
O
O
Ph
6
Ph
Ph
5
trans-
6
r-2,c-3,t-5-
r-2,t-3,t-5-
62%
Scheme 3 Synthesis of oxa-preussin (all-cis-6) by stereoselective reduction of cis-5 and reduction of trans-5. (The shown
formulas refer to the relative configurations; the compounds are racemic mixtures).

Page 5 of 20
Organic & Biomolecular Chemistry
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For the synthesis of preussin we planned the use of N-tosyl imine 8 since this electrophile should be
DOI: 10.1039/C8OB02645A
highly reactive. The N-tosyl group also leads to fairly stable products and on the other hand, it can easily
be removed under mild conditions at later stages. However, compound 8 is not particularly stable due
to its fast isomerization into the corresponding enamine tautomer.23 Therefore we examined the C,N-
ditosyl amine 9 as precursor, since under basic conditions this compound undergoes smooth elimination
of sulfinate to provide in situ imine 8 that can be trapped by nucleophiles.24 In most cases the employed
base and nucleophile are identical. In a first model reaction, two equivalents of methoxyallene (7) were
lithiated and imine precursor 9 was added at -80 °C giving the expected allenyl amine 10 quantitatively
(Scheme 4). This intermediate was cyclized with silver nitrate to furnish the expected 2,5-dihydropyrrole
derivative 11 in 60% overall yield. This experiment taught us that 9 should be a suitable building block
for the preparation of the target compound preussin.
1. 2 equiv. nBuLi
THF, -40 °C, 30 min
OMe
OMe
H
•
•
Ph
Ph
7
HN
-80 °C
to -30 °C
2 h
2.
Tos
Tos
(2 equiv.)
N
quant.
10
8
0.2 equiv. AgNO3
2 equiv. K2CO3
Ph
acetonitrile
r.t., 12 h
Tos
Tos
NH
OMe
9
N
Ph
Tos
11
60%
Scheme 4 Synthesis of model compound 11 by lithiation and addition of methoxyallene (7) to in situ generated imine 8 and
subsequent cyclization of allenyl amine 10.
With the knowledge gained during the preparation of oxa-preussin and the model reaction above we
combined racemic 3-nonyl-substituted methoxyallene 1 with imine precursor 9 (Scheme 5). Two
equivalent of lithiated 1 and 9 afforded the expected diastereomers pro-cis-12 and pro-trans-12 in good
yield, but with no selectivity. The two isomers were separated by conventional column chromatography
and both were isolated in good quantities in 36% yield. The excess of 1 required in this experiment could
be re-isolated and used again.
OMe
•
Non
Ph
HN
Tos
12
36% pro-cis-
1. 2 equiv. nBuLi
THF, -40 °C, 30 min
OMe
H
•
+
OMe
Non
2.
Ph
•
-80 °C
to -30 °C
3 h
1
Non
36%
Ph
(2 equiv.)
HN
N
Tos
Tos
8
12
pro-trans-
Ph
Tos
Tos
NH
9

Organic & Biomolecular Chemistry
Page 6 of 20
Scheme 5 Synthesis of diastereomeric allenyl amines pro-cis-12 and pro-trans-12 by lithiation of 3-nonyl-substitutedView Article Online
DOI: 10.1039/C8OB02645A
methoxyallene 1 and addition to in situ generated imine 8. (The shown formulas refer to the relative configurations; the
compounds are racemic mixtures).
Next, two methods were used for the cyclization step in order to receive the required cis-13 from both
diastereomeric precursors.21 The cyclization of pro-trans-12 with silver nitrate under buffered conditions
led to a mixture of cis-13 and trans-13 in 87% yield (Scheme 6). The two isomers were separated by
HPLC to furnish the two crystalline isomers in 42% and 30% yield, respectively. Whereas this method
is not stereospecific due to the already mentioned equilibration at the allenyl amine stage, the alternative
cyclization under strongly basic conditions with potassium tert-butoxide in dimethyl sulfoxide proceeds
stereospecifically.21 Starting with pro-cis-12 after 12 h the desired 2,5-disubstituted dihydropyrrole
1
derivative cis-13 was isolated exclusively in good yield. The H NMR data allow an unambiguous
assignment of the relative configurations of the compounds. Altogether, preussin precursor cis-13 was
isolated in 43% overall yield when the results of Schemes 5 and 6 are combined.
OMe
Non
N
Ph
Tos
0.2 equiv. AgNO3
2 equiv. K2CO3
42%
Non
13
cis-
OMe
•
+
OMe
acetonitrile
r.t., 12 h
Non
Ph
HN
Tos
N
Ph
12
pro-trans-
Tos
13
30% trans-
OMe
0.25 equiv.
KOtBu
OMe
•
Non
N
Non
DMSO
50 °C, 12 h
Ph
Ph
HN
Tos
Tos
13
cis-
78%
12
pro-cis-
Scheme 6 Cyclizations of diastereomeric allenyl amines 12 to cis- and trans-configured dihydropyrrole derivatives 13 under
silver nitrate or potassium tert-butoxide promotion. (The shown formulas refer to the relative configurations; the compounds are
racemic mixtures).
The hydrolysis of cis-13 and trans-13 could be routinely achieved with 20% aqueous sulfuric acid,
furnishing the required 2,5-disubstituted pyrrolidin-3-ones cis-14 and trans-14 quantitatively (Scheme 7,
equations 1 and 2). A cis/trans-equilibration via the corresponding enol form might be possible due to
the CH acidic position at C-2, however, even under the relatively harsh hydrolysis conditions no cross-
over between cis-14 and trans-14 was found. Since only cis-14 has the required relative configuration
to approach preussin, we also examined the equilibration under basic conditions. Whereas no reaction
was observed with weak bases (K2CO3 or NEt3), 1,8-diazabicyclo-[5.4.0]undec-7-ene (DBU) converted
trans-14 within 2 h into the thermodynamically more stable cis-14. Unfortunately, a second compound
was formed in similar quantities (Scheme 7, equation 3) whose NMR data reveal the constitution of
product 15. Apparently, a base-promoted sulfinate elimination competes with the desired trans/cis-
equilibration. The conversion into 15 could be completed by applying DBU for longer reaction times
(Scheme 7, equation 4). Without the disturbing NMR signals of compound cis-14 we could detect that
compound 15 is in equilibrium (ratio ca. 3:1) with its tautomer 16 (E/Z ca. 1:1).

Page 7 of 20
Organic & Biomolecular Chemistry
O
OMe
View Article Online
DOI: 10.1039/C8OB02645A
20% H2SO4/H2O
(1)
(2)
Non
Non
N
N
THF, , 4 h
quant.
Ph
Ph
Ph
Tos
Tos
14
cis-
13
cis-
O
OMe
20% H2SO4/H2O
Non
Non
N
N
THF, ,4 h
quant.
Ph
Tos
Tos
13
trans-
14
trans-
O
DBU, CH2Cl2
r.t., 2 h
(3)
14
cis-
14
trans-
+
Non
N
Ph
(ca. 1:1)
O
15
O
DBU, CH2Cl2
(4)
Ph
14
cis-
Non
Non
N
H
N
r.t., 5.5 h
Ph
15
16
(ca. 3:1)
Scheme 7 Hydrolysis of the enol ether moiety of 2,5-dihydropyrroles cis-13 and trans-13 to pyrrolidin-3-ones cis-14 and trans-
14 and equilibration experiments finally leading to 15 and 16. (The shown formulas refer to the relative configurations; the
compounds are racemic mixtures).
For the completion of the synthesis of rac-preussin the N-tosyl group of cis-13 has to be replaced by an
N-methyl group. Among the several possibilities to reductively remove N-sulfonyl substituents we
examined sodium naphthalenide as reagent.25 At -78 °C compound cis-13 was smoothly converted into
cis-17 when the mixture was quenched with water (Scheme 8). The analogous reaction, similarly
executed but quenched with an excess of methyl iodide, furnished cis-18 in 53% yield. The moderate
efficacy of this step is probably due to the formation of the quarternary ammonium salt. Compound cis-
18 was transformed by acidic hydrolysis into 2,5-disubstituted pyrrolidin-3-one derivative cis-19 in 60%
yield. The final reduction with sodium borohydride provided an 80:20 mixture of the two possible
diastereomers, but with L-Selectride it proceeded with high diastereoselectivity and exclusively gave all-
cis-20 in good yield. The NMR data of the sample were in full agreement with those published for
preussin in the literature.26 In conclusion, we could accomplish a synthesis of rac-preussin in five steps
(from alkoxyallene 1 and imine precursor 9). The moderate overall yield of 11% is mainly due to the
unselective formation of pro-cis-12 from alkoxyallene 1 and imine 8. On the other hand, the isolation of
pro-trans-12 in similar quantities should allow the preparation of diastereomeric analog of rac-preussin
with trans orientation of the benzyl and nonyl substituents at C-2 and C-5. Starting with differently 3-
alkyl-substituted alkoxyallenes the preparation of other preussin analoga, e.g. the new relative preussin
B, should be easily possible.

Organic & Biomolecular Chemistry
Page 8 of 20
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DOI: 10.1039/C8OB02645A
1. 3 equiv. Na-Naph
OMe
DME
OMe
-78 °C, 1 h
Non
Non
N
N
2. H2O
75%
H
Ph
Ph
Tos
17
cis-
13
cis-
1. 3 equiv. Na-Naph
DME
OMe
OMe
-78 °C, 1.5 h
Non
Non
N
N
2. 10 equiv. MeI
-78 °C to r.t.
1 h
Ph
Ph
Tos
Me
13
cis-
18
cis-
53%
20% H2SO4/H2O
THF,
4 h
60%
O
OH
L-Selectride, THF
Non
Non
N
N
-78 °C, 3 h
Ph
Ph
Me
Me
80%
d.r.>99:1
19
cis-
20
all-cis-
rac-preussin
Scheme 8 Synthesis of rac-preussin (all-cis-20) by reductive removal of the N-tosyl group of cis-13, N-methylation to cis-18,
conversion into pyrrolidin-3-one cis-19 followed by stereoselective reduction. (The shown formulas refer to the relative
configurations; the compounds are racemic mixtures).
As a result of our systematic studies of 3-alkyl-substituted alkoxyallenes bearing carbohydrate-derived
auxiliaries, the pyrrolidin-3-one cis-14 was also available in enantiopure form.22 Enantiomers 2R,5S-cis-
14 and 2S,5R-cis-14 had been prepared in a three-step sequence employing the diacetone fructose-
derived alkoxyallene congener of 1 and imine 8 as crucial precursors. Without knowing the absolute
configuration by certainty,22 we converted the obtained major isomer cis-14 into enantiopure preussin
whose optical rotation showed that we have prepared the unnatural (-)-enantiomer (Scheme 9). Hence
the configuration of the used cis-14 could finally be confirmed to be 2R,5S. For the preparation of (-)-
preussin we slightly modified the sequence of steps, starting with the reduction of the ketone moiety and
finalizing it by a reductive amination to introduce the N-methyl group. Whereas the reduction of 2R,5S-
cis-14 was unselective with sodium borohydride giving an 80:20 mixture of the two diastereomers, the
use of L-Selectride exclusively afforded 2R,3R,5S-all-cis-21 in good yield. The subsequent reductive
removal of the N-tosyl group furnished gave 2R,3R,5S-all-cis-22 that is the optical antipode of the
recently isolated preussin C. Since we had learned during the synthesis of racemic preussin that the
direct N-methylation of the intermediate amide anion with methyl iodide proceeded only with moderate
efficacy (Scheme 8), we employed a literature known method27 for a reductive methylation with aqueous
formaldehyde and sodium cyanoborohydride under acidic conditions. The obtained 2R,3R,5S-all-cis-20
was isolated in 80% yield and its NMR data agree very well with those of the literature.26

Page 9 of 20
Organic & Biomolecular Chemistry
O
View Article Online
DOI: 10.1039/C8OB02645A
OH
L-Selectride
Non
Non
N
N
THF, -78 °C, 9 h
73%
Ph
14
Ph
Tos
Tos
2R,3R,5S-all-cis-
2R,5S-cis-
21
5 equiv. Na-Naph
DME, -78 °C, 1.5 h
87%
NaCNBH3
CH2O, H2O
MeCO2H
OH
OH
Non
Non
N
N
H
THF
Ph
Ph
Me
acetonitrile
r.t., 1 h
22
2R,3R,5S-all-cis-
20
2R,3R,5S-all-cis-
80%
(-)-preussin
OH
O
Non
Non
N
N
Ph
Ph
Me
Tos
14
2S,5R-cis-
(+)-preussin
Scheme 9 Synthesis of enantiopure (-)-preussin by stereoselective reduction of 2R,5S-cis-15 followed by transformations into
all-cis-22 and all-cis-20.
The negative sign of the optical rotation of the final product revealed that we have obtained the unnatural
enantiomer and the absolute value of -25.8 (in chloroform) indicated the high enantiomeric purity.28 The
optical purity was further evidenced by converting the sample of all-cis-20 into its Mosher ester. NMR
and HPLC analyses showed that the obtained product has a ratio of diastereomers of > 95:5 indicating
an ee of at least 90%.
Summing up, our route to (-) preussin involved six steps (starting from the diacetone fructose-derived
allene and imine precursor 9), but the overall yield of only 16% is due to the formation of diastereomers
during the route to the required allenyl amine. It should also be mentioned that the 2S,5R-cis-14 isomer,
also available in lower quantities by this route,22 will allow the reparation of the natural (+) preussin in
analogous fashion.
Twenty-one racemic or enantiopure precursors and analogs of preussin (including oxa-preussin)
obtained in this study and the preceding report were investigated in vitro for their cytotoxicity against
MCF-7 tumor cells during 96 hours of incubation. Only five compounds were found to be cytotoxic with
IC50 values in the range of 3 to 6 μM (Fig. 2). Racemic preussin showed an IC50 value of 6 μM whereas
for the (-)-preussin 3.5 μM were determined. In the literature an IC50 value of 4.1 μM is reported for the
naturally occurring (+)-preussin.11 These results indicate that the absolute configuration of the preussin
enantiomers are not decisive for its cytotoxicity. This is also in accordance with an investigation of the
eight stereoisomers with preussin constitution that revealed that the relative and absolute configuration
of the compounds has no crucial effect on their inhibitory activity against the cell growth of fission
yeast.28b

Organic & Biomolecular Chemistry
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OMe
OH
OMe
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DOI: 10.1039/C8OB02645A
Non
Non
N
N
N
H
Ph
Ph
20
Ph
Tos
11
Me
(rac) cis-
rac preussin
17
(rac) cis-
3 M
3 M
6 M
O
OH
Non
Non
N
N
Ph
14
Ph
Tos
Me
20
2R,5S-cis-
3.8 M
2R,3R,5S-cis-
(-)-preussin
3.5 M
Fig. 2 IC50 values of cytotoxic compounds of this studies against the human tumor cell line MCF-7.
As mentioned in the introduction, it was reported that (+)-preussin directly intervenes in the cell cycle
and inhibits the entry to the S phase.11 In order to confirm this finding, the five cytotoxic compounds
depicted in Figure 2 were additionally investigated in a cyclinE/CDK2-assay. However, none of the
compounds inhibited this kinase. The cellular active compounds 11, cis-17 and cis-20 [(-)-preussin] were
tested as inhibitors of other kinases, but none showed activity. A final conclusion about the mode of
action of this type of compounds is therefore not possible.
Conclusions
The routes to racemic preussin and its oxa-analog oxa-preussin based on 3-nonyl-substituted
methoxyallene 1 as key precursor are short and flexible. The diastereoselectivity of the addition of the
lithiated allene to the electrophiles is low, but this stereodivergent step opens the way for the synthesis
of diastereomeric target compounds. A synthesis of the antipode of the natural product – (-)-preussin –
proceeds efficiently by starting from highly enantio-enriched 2R,5S-cis-14 and proves the absolute
configuration of this compound and its precursors. On the way to (-)-preussin its demethylated relative
(-)-preussin C was passed by that is the antipode of a recently isolated natural product. Our approach
to compounds with preussin skeleton is highly flexible since we do not follow a chiral pool but an
auxiliary-based strategy. As a consequence, the 3-alkyl group of the starting allene and the C-
substituent of the imine can easily be varied and a broad range of target compounds with differing
substituents at C-2 and C-5 should easily be accessible, e.g. preussin B or related compounds. Racemic
preussin and (-)-preussin showed IC50 values against MCF-7 tumors cells in the range of 3.5-6 μM,
similar to a literature reported value for (+)-preussin. Unexpectedly, the absolute configuration of the
compounds does not play no essential role for its cytotoxicity.
Experimental
For general information and details of remaining experiments, see ESI.
Starting materials: 1-methoxydodeca-1,2-diene (1)20b, imine precursor 924.
General procedure for the addition of lithiated alkoxyallenes to aldehydes or imines (GP1)

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Organic & Biomolecular Chemistry
For generation of the lithiated alkoxyallene, the corresponding alkoxyallene was dissolved in THF and n-butyllithium wasVaiedwdAerdticle Online
DOI: 10.1039/C8OB02645A
at -40 °C. After 30 min, the solution was cooled to -80 °C and the corresponding electrophile was added. The mixture was allowed
to warm up to -30 °C and stirred for the time given in the individual experiments. After quenching with saturated aqueous NaHCO3
solution (10 mL/), the organic phase was separated and the aqueous phase was extracted with diethyl ether (3 x 15 mL/mmol of
alkoxyallene). The combined organic phases were dried (Na2SO4), filtered and evaporated in vacuo to provide the crude product
that was used directly or purified as given in the individual experiment. The yields refer to the amount of electrophile used.
3-Methoxy-1-phenyltetradeca-3,4-diene-2-ol (3)
According to GP1, 1-methoxydodeca-1,2-diene (1) (0.620 g, 3.16 mmol), n-butyllithium (1.30 mL, 3.16 mmol of 2.43 M solution in
hexanes) and phenylethanal (2) (0.380 g, 3.16 mmol) in THF (40 mL) provided after 2 h crude 3 (1.13 g, d.r. 55:45) as light yellow
oil that was cyclized without purification.
1H NMR (CDCl3, 270 MHz), major diastereomer:  = 0.89 (t, J = 6.6 Hz, 3 H, Me), 1.15–1.40, 1.85–2.00 (2 m, 14 H, 2 H, CH2),
2.01 (d, J = 5.9 Hz, 1 H, OH), 2.88, 2.98 (AB part of ABX system, JAB = 13.6 Hz, JAX = 7.4 Hz, JBX = 2.6 Hz, 1 H each, 1-H), 3.41
(s, 3 H, OMe), 4.28–4.44 (m, 1 H, 2-H), 5.84 (d, J = 7.0 Hz, 1 H, 5-H), 7.10–7.40 ppm ( m, 5 H, Ph); the following signals of the
minor diastereomer are distinguishable from those of the major isomer:  = 2.15 (d, J = 5.9 Hz, 1 H, OH), 2.88, 3.00 (AB part of
ABX system, JAB = 13.6 Hz, JAX = 7.4 Hz, JBX = 3.3 Hz, 1 H each, 1-H), 5.89 ppm (td, J = 6.8 Hz, J = 1.7 Hz, 1 H, 5-H). 13C NMR
(CDCl3, 67.9 MHz), major diastereomer:  = 14.0 (q, Me), 22.6, 28.5, 29.1, 29.2, 29.4, 29.5, 31.4, 31.8 (8 t, CH2), 40.9 (t, C-1),
55.9 (q, OMe), 71.2 (d, C-2), 109.8 (d, C-5), 126.2, 128.1, 129.4 (3 d, Ph), 134.1 (s, C-3), 138.0 (s, Ph),188.8 ppm (s, C-4); the
following signals of the minor diastereomer are distinguishable from those of the major isomer:  = 31.5 (t, CH2), 41.1 (t, C-1),
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72.5 (d, C-2), 108.8 (d, C-5), 129.5 (d, Ph), 135.0 (s, C-3), 137.8 (s, Ph), 188.5 ppm (s, C-4). IR (film): = 3460 (OH), 3030–2855
(C-H), 1950 cm-1 (C=C=C). MS (EI, 80 eV): m/z (%) = 316 (1) [M+], 301 (1) [M+ - Me], 239 (21) [M+ - Ph], 227 (65), 91 (100) [Bn+].
General procedure for silver nitrate-promoted cyclization (GP2)
To a solution of the corresponding allenyl alcohol or amine in acetonitrile (5 mL/mmol) were added under a stream of argon via a
funnel potassium carbonate and silver nitrate. The resulting mixture was stirred at room temperature under light exclusion for 12
h, then filtered and evaporated. The residue was dissolved in a small amount ethyl acetate and filtered through a pad of Celite
(elution with ethyl acetate). After removal of the solvents in vacuo, the crude product was purified as indicated in the individual
experiments.
2-Benzyl-3-methoxy-5-nonyl-2,5-dihydrofuran (4)
According to GP2, crude allenyl alcohol 3 (1.13 g, d.r. 55:45) in acetonitrile (40 mL) was treated with AgNO3 (0.107 g, 0.63 mmol)
and K2CO3 (0.873 g, 6.30 mmol) for 12 h to provide crude 4 (1.11 g, d.r. 70:30) as brown oil, that was directly hydrolyzed in the
next step. The cis/trans assignments are based on the assignments of cis-5 and trans-5.
General procedure for acid-promoted hydrolysis of 2,5-dihydrofurans and 2,5-dihydropyrroles (GP3)
A solution of the corresponding substrate in THF was heated under reflux with aqueous sulfuric acid (2 M or 20%) for the time
given in the individual experiments (progress of conversion was followed by TLC). After cooling to room temperature the mixture
was cautiously neutralized with saturated aqueous NaHCO3 solution and the organic phase was separated. The aqueous phase
was extracted with diethyl ether (2 x 15 mL). The combined organic phases were dried (Na2SO4), filtered and evaporated in vacuo.
The purification is indicated in the individual experiments.
2-Benzyl-5-nonyltetrahydrofuran-3-one (5)
According to GP3, crude 4 (1.11 g, d.r. 70:30) in THF (20 mL) and H2SO4 (14 mL of 2 M aqueous solution) provided after 3.5 h
crude 5 (1.06 g, d.r. 70:30) as yellow oil. Purification by column chromatography (silica gel, hexanes/ethyl acetate 10:1) furnished
5 (0.630 g, 66%, d.r. 70:30) as colorless oil and subsequent separation of the diastereomers by HPLC (hexanes/ethyl acetate
97:3) afforded trans-5 (0.165 g, 17%, containing minor impurities) and cis-5 (0.402 g, 42%) as colorless oils. Overall yield for three
steps: 59%.
Data of cis-5: 1H NMR (CDCl3, 270 MHz):  = 0.88 (t, J = 6.9 Hz, 3 H, Me), 1.20–1.40, 1.47–1.60 1.62–1.70, (3 m, 14 H, 1 H each,
CH2), 1.83 (dd, J = 17.9 Hz, J = 10.6 Hz, 1 H, 4-H), 2.45 (dd, J = 17.9 Hz, J = 5.6 Hz, 1 H, 4-H), 2.87, 3.08, 4.00 (ABX system,
JAB = 14.4 Hz, JAX = 6.9 Hz, JBX = 3.9 Hz, 1 H each, PhCH2, 2-H), 4.06 (ddt, J = 10.6 Hz, J = 5.9 Hz, J = 5.6 Hz, 1 H, 5-H), 7.20–
7.32 ppm (m, 5 H, Ph). 13C NMR (CDCl3, 67.9 MHz):  = 14.1 (q, Me), 22.7, 25.2, 29.3, 29.4, 29.5*, 31.9, 35.5, 37.2, 41.2 (9 t,
CH2, C-4, PhCH2,), 75.9, 81.9 (2 d, C-2, C-5), 126.5, 128.1, 129.7, 137.2 (3 d, s, Ph), 216.0 ppm (s, C-3); * signal with higher

Organic & Biomolecular Chemistry
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-1
+
+
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DOI: 10.1039/C8OB02645A
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intensity. IR (film): = 3090–2855 (C-H), 1740 cm (C=O). MS (EI, 80 eV): m/z (%) = 302 (20) [M ], 181 (16), 91 (100) [Bn ].
C20H30O2 (302.5): calcd. C 79.42, H 10.00; found C 79.34, H 9.75.
Data of trans-5: 1H NMR (CDCl3, 270 MHz):  = 0.88 (t, J = 6.7 Hz, 3 H, Me), 1.20–1.40, 1.45–1.60, 1.60–1.75 (3 m, 14 H, 1 H
each, CH2), 2.15, 2.34 (AB part of ABX system, JAB = 18.0 Hz, JAX = 7.3 Hz, JBX = 6.8 Hz, 1 H each, 4-H), 2.90, 2.98, 4.23 (ABX
system, JAB = 14.2 Hz, JAX = 6.8 Hz, JBX = 4.6 Hz, 1 H each, PhCH2, 2-H), 4.03–4.16 (m, 1 H, 5-H), 7.16–7.31 ppm (m, 5 H, Ph).
13C NMR (CDCl3, 67.9 MHz):  = 14.1 (q, Me), 22.6, 25.4, 29.2, 29.4, 29.5*, 31.8, 35.6, 37.0, 42.6 (9 t, CH2, C-4, PhCH2), 75.5,
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80.0 (2 d, C-2, C-5), 126.5, 128.3, 129.5, 137.1 (3 d, s, Ph), 216.4 ppm (s, C-3); * signal with higher intensity. IR (film): = 3065–
2855 (C-H), 1755 cm-1 (C=O). MS (EI, 80 eV): m/z (%) = 302 (6) [M+], 136 (16), 120 (18), 91 (100) [Bn+].
General procedure for sodium borohydride-promoted reductions of cyclic ketones (GP4)
The corresponding substrate was dissolved in dry ethanol and at 0 °C sodium borohydride was added under a stream of argon
via a funnel. After stirring at room temperature for the time indicated, the mixture was treated with 2 N HCl until the precipitate
dissolved and was extracted with diethyl ether (3 x 10 mL). The combined organic phases were washed with saturated aqueous
NaHCO3 solution (until the extract is neutral), dried (Na2SO4), filtered and concentrated in vacuo. The crude product was purified
by chromatography.
(all-cis)- and (r-2,t-3,c-5)-2-Benzyl-5-nonyltetrahydrofuran-3-ol (6)
According to GP4, a solution of cis-5 (0.027 g, 0.09 mmol), NaBH4 (0.007 g, 0.17 mmol) in EtOH (3 mL) provided after 14 h crude
6 (0.032 g, d.r. 75:25) as a colorless oil. Column chromatography (silica gel, hexanes/ethyl acetate 5:1) furnished all-cis-6 (0.021
g, 77%) as colorless crystals (m.p. 49–50 °C) and r-2,t-3,c-5-6 (0.005 g, 18%) as colorless crystals (m.p. 42–43 °C).
Data of all-cis-6: 1H NMR (CDCl3, 500 MHz):  = 0.88 (t, J = 6.5 Hz, 3 H, Me), 1.20–1.40 (m, 14 H, CH2), 1.54 (ddd, J = 13.9 Hz,
J = 6.5 Hz, J = 1.8 Hz, 1 H, 4-H), 1.51–1.58 (m, 1 H, CH2), 1.68 (d, J = 7.5 Hz, 1 H, OH), 1.66–1.78 (m, 1 H, CH2), 2.34 (ddd, J =
13.9 Hz, J = 8.2 Hz, J = 6.3 Hz, 1 H, 4-H), 3.01 (d, J = 7.1 Hz, 2 H, PhCH2), 3.75 (td, J = 7.1 Hz, J = 3.3 Hz, 1 H, 2-H), 3.79 (dq,
J = 8.2 Hz, J = 6.5 Hz, 1 H, 5-H), 4.05–4.13 (m, 1 H, 3-H), 7.15–7.33 ppm (m, 5 H, Ph). 13C NMR (CDCl3, 125.8 MHz):  = 14.1
(q, Me), 22.7, 26.2, 29.3, 29.5, 29.6*, 31.9, 36.8, (7 t, CH2), 35.1 (t, PhCH2), 41.6 (t, C-4), 72.3 (d, C-3), 77.8 (d, C-5), 83.8 (d, C-
2), 126.2, 128,4, 129.2, 138.6 ppm (3 d, s, Ph); * signal with higher intensity. IR (KBr): = 3410 (O-H), 3030–2850 cm-1 (C-H). MS
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(EI, 80 eV): m/z (%) = 304 (10) [M+], 213 (100) [M+ - Bn], 177 (34) [M+ - C9H19]. C20H30O2 (304.5): calcd. C 78.90, H 10.59; found
C 78.73, H 10.57.
Data of r-2,t-3,c-5-6: 1H NMR (CDCl3, 270 MHz):  = 0.88 (t, J = 6.6 Hz, 3 H, Me), 1.20–1.50 (m, 16 H, CH2), 1.62 (ddd, J = 13.1
Hz, J = 9.6 Hz, J = 6.6 Hz, 1 H, 4-H), 1.55–1.70 (m, 1 H, OH), 1.87 (ddd, J = 13.1 Hz, J = 5.7 Hz, J = 2.4 Hz, 1 H, 4-H), 2.71, 2.96,
3.94 (ABX system, JAB = 13.7 Hz, JAX = 7.4 Hz, JBX = 6.0 Hz, 1 H each, PhCH2, 2-H), 4.07 (dtd, J = 9.6 Hz, J = 6.0 Hz, J = 5.7 Hz,
1 H, 5-H), 4.08–4.16 (m, 1 H, 3-H), 7.20–7.35 ppm (m, 5 H, Ph). 13C NMR (CDCl3, 67.9 MHz):  = 14.1 (q, Me), 22.7, 26.0, 29.3,
29.6*, 29.7, 31.9, 35.1, 35.8, 40.6 (9 t, CH2, C-4, PhCH2), 75.7 (d, C-3), 78.2 (d, C-5), 87.1 (d, C-2), 126.4, 128.4, 129.4, 137.9
-1
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ppm (3 d, s, Ph); * signal with higher intensity. IR (KBr): = 3410 (O-H), 3085, 3020, 2955, 2920, 2820 cm (C-H). MS (EI, 80
eV): m/z (%) = 304 (13) [M+], 213 (100) [M+ - Bn], 177 (37) [M+ - C9H19]. HRMS (EI, 80 eV): calcd. for C20H32O2: 304.2402; found
304.2444.
General procedure for the reduction of cyclic ketones with L-Selectride (GP5)
The corresponding substrate was dissolved in THF and at –78 °C an excess of L-Selectride (1 M in THF) was added dropwise.
The mixture was stirred at this temperature for the time indicated and then quenched by addition of water (10 mL). The aqueous
phase was separated and the organic phase was extracted with ethyl acetate (3 x 10 mL). The combined organic phases were
dried (Na2SO4), filtered and the solvents were removed in vacuum. The crude product was purified by chromatography.
(all-cis)-2-Benzyl-5-nonyltetrahydrofuran-3-ol (6)
According to GP5, a solution of cis-5 (0.036 g, 0.12 mmol), L-Selectride (0.18 mL of 1 M solution in THF, 0.18 mmol) in THF (3
mL) provided after 3 h crude 6 (0.061 g). Purification by column chromatography (silica gel, hexanes/ethyl acetate 5:1) afforded
all-cis-6 (0.036 g, 99%) as colorless crystals (m.p. 49–50 °C). The spectroscopic data agree with those of the sample obtained
before.
(r-2,t-3,t-5)- and (r-2,c-3,t-5)-2-Benzyl-5-nonyltetrahydrofuran-3-ol (6)

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Organic & Biomolecular Chemistry
According to GP5, a solution of trans-5 (0.147 g, 0.49 mmol), L-Selectride (0.73 mL of 1 M solution in THF, 0.73 mmol) iVniewTHArFticle Online
DOI: 10.1039/C8OB02645A
(20 mL) provided after 3 h the crude product (0.250 g, d.r. 90:10). The ratio of diastereomers was determined by HPLC. Purification
by column chromatography (silica gel, hexanes/ethyl acetate 5:1) and subsequent separation by HPLC (hexanes/ethyl acetate
5:1) afforded r-2,t-3,t-5-6 (0.091 g, 62%) as colorless crystals (m.p. 54–55 °C) and r-2,c-3,t-5-6 (0.010 g, 8%) as colorless oil.
Data of r-2,t-3,t-5-6: H NMR (CDCl3, 270 MHz):  = 0.88 (t, J = 6.5 Hz, 3 H, Me), 1.20–1.45 (m, 15 H, CH2), 1.55–1.73 (m, 2 H,
OH, CH2), 1.70 (ddd, J = 13.4 Hz, J = 9.5 Hz, J = 4.4 Hz, 1 H, 4-H), 2.08 (dd, J = 13.4 Hz, J = 6.1 Hz, 1 H, 4-H), 2.96, 3.01, 4.05
(ABX system, JAB = 13.5 Hz, JAX = 7.7 Hz, JBX = 6.7 Hz, 1 H each, PhCH2, 2-H), 4.09–4.17 (m, 1 H, 3-H), 4.28 (dq, J = 9.5 Hz, J
= 6.1 Hz, 1 H, 5-H), 7.17–7.33 ppm (m, 5 H, Ph). 13C NMR (CDCl3, 67.9 MHz):  = 14.1 (q, Me), 22.7, 26.0, 29.3, 29.5, 29.6*,
31.9, 35.3, 36.3, 41.6 (9 t, CH2, C-4, PhCH2), 72.7 (d, C-3), 77.5 (d, C-5), 83.6 (d, C-2), 126.2, 128.4, 129.2, 138.5 ppm (3 d, s,
Ph); * signal with higher intensity. IR (KBr): = 3420 (O-H), 3035–2850 cm-1 (C-H). MS (EI, 80 eV): m/z (%) = 304 (18) [M+], 213
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(100) [M+ - Bn], 91 (81) [Bn+]. HRMS (EI, 80 eV): calcd. for C20H32O2: 304.2402; found 304.2434.
Data of r-2,c-3,t-5-6: 1H NMR (CDCl3, 270 MHz):  = 0.87 (t, J = 6.9 Hz, 3 H, Me), 1.15–1.73 (m, 17 H, OH, CH2), 1.57 (ddd, J =
13.2 Hz, J = 7.3 Hz, J = 5.6 Hz, 1 H, 4-H), 2.33 (dd, J = 13.2 Hz, J = 6.3 Hz, 1 H, 4-H), 2.71, 2.92 (AB part of ABX system, JAB
=
13.6 Hz, JAX = 7.2 Hz, JBX = 6.3 Hz, 1 H each, PhCH2), 3.95–4.05, 4.08–4.16 (2 m, 3 H, 2-H, 3-H, 5-H), 7.17–7.35 ppm (m, 5 H,
Ph). 13C NMR (CDCl3, 67.9 MHz):  = 14.1 (q, Me), 22.7, 26.0, 29.3, 29.5, 29.6, 29.7, 31.9, 36.6, 39.4, 40.3 (10 t, CH2, C-4,
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PhCH2), 75.9, 77.4, 85.0 (3 d, C-2, C-3, C-5), 126.4, 128.5, 129.3, 137.8 ppm (3 d, s, Ph). IR (film): = 3410 (O-H), 3085–2855
cm-1 (C-H). MS (EI, 80 eV): m/z (%) = 304 (11) [M+], 213 (100) [M+ - Bn], 91 (96) [Bn+]. HRMS (EI, 80 eV): calcd. for C20H32O2:
304.2402; found 304.2439.
N-(3-Methoxy-1-phenyltetradeca-3,4-dien-2-yl)-p-toluenesulfonamide (12)
According to GP1, 1-methoxydodeca-1,2-diene (1) (1.32 g, 6.71 mmol), n-butyllithium (2.80 mL, 6.71 mmol of 2.40 M solution in
hexanes) and 9 (1.44 g, 3.35 mmol) in THF (50 mL) provided after 3 h crude 12 (2.34 g, d.r. 50:50) as light yellow oil. The crude
product was purified by column chromatography (alumina III, hexanes/ethyl acetate 5:1) to give 12 (0.778 g), pro-cis-12 (0.569 g,
36%) as light yellow solid (melting range 36–40 °C) and pro-trans-12 (0.566 g, 36%) as light yellow crystals (m.p. 48 °C).
Data of pro-cis-12: 1H NMR (CDCl3, 270 MHz):  = 0.89 (t, J = 6.3 Hz, 3 H, Me), 1.00–1.40, 1.60–1.70 (2 m, 14 H, 2 H, CH2), 2.40
(s, 3 H, Tos-Me), 2.92, 2.99 (AB part of ABX system, JAB = 13.8 Hz, JAX = 7.0 Hz, JBX = 5.9 Hz, 1 H each, 1-H), 3.20 (s, 3 H, OMe),
4.05–4.18 (m, 1 H, 2-H), 4.56 (d, J = 9.6 Hz, 1 H, NH), 5.45 (t, J = 5.9 Hz, 1 H, 5-H), 7.09 (d, J = 7.4 Hz, 2 H, Ph), 7.15–7.30 (m,
5 H, Ph, Tos), 7.66 ppm (d, J = 8.1 Hz, 2 H, Tos). 13C NMR (CDCl3, 67.9 MHz):  = 14.1 (q, Me), 21.5 (q, Tos-Me), 22.7, 28.7,
29.2, 29.3, 29.4, 29.6, 31.2, 31.9 (8 t, CH2), 40.1 (t, C-1), 58.8, 55.4 (q, d, OMe, C-2), 109.7 (d, C-5), 126.5, 127.1, 128.1, 129.3,
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129.8 (5 d, Ph, Ts), 131.2 (s, C-3), 136.5*, 142.9 (2 s, Ph, Ts), 189.0 ppm (s, C-4); * signal with higher intensity. IR (KBr):
=
3255 (N-H), 3065, 2925, 2850 (C-H), 1965 (C=C), 1325, 1165 cm-1 (Tos-N). MS (pos. FAB): m/z (%) = 470 (9) [M+ + H], 456 (11)
[M+ + H - Me], 378 (5) [M+ - Bn], 274 (34), 154 (73), 136 (66), 91 (100) [Bn+].
Data of pro-trans-12: 1H NMR (CDCl3, 500 MHz):  = 0.89 (t, J = 7.0 Hz, 3 H, Me), 1.00–1.45, 1.50–1.60 (2 m, 14 H, 2 H, CH2),
2.40 (s, 3 H, Tos-Me), 2.89, 2.97 (AB part of ABX system, JAB = 13.2 Hz, JAX = 8.8 Hz, JBX = 5.7 Hz, 1 H each, 1-H), 3.06 (s, 3 H,
OMe), 4.05–4.13 (m, 1 H, 2-H), 4.86 (d, J = 9.4 Hz, 1 H, NH), 5.42 (t, J = 6.7 Hz, 1 H, 5-H), 7.08 (d, J = 7.1 Hz, 2 H, Ph), 7.10–
7.30 (m, 5 H, Ph, Ts), 7.67 ppm (d, J = 8.4 Hz, 2 H, Ts). 13C NMR (CDCl3, 125.8 MHz):  = 14.2 (q, Me), 21.5 (q, Tos-Me), 22.7,
28.4, 29.1, 29.3, 29.4, 29.5, 30.9, 31.9 (8 t, CH2), 40.1 (t, C-1), 55.5, 57.4 (q, d, OMe, C-2), 108.0 (d, C-5), 126.4, 128.6, 128.9,
129.1, 130.0 (5 d, Ph, Ts), 136.9, 137.8, 142.9 (3 s, Ph, Ts)*, 189.3 ppm (s, C-4) ppm; * signal of C-3 is hidden by the aryl signals.
IR (KBr): = 3270 (N-H), 3065, 2925, 2855 (C-H), 1965 (C=C), 1335, 1160 cm-1 (Tos-N). MS (pos. FAB): m/z (%) = 470 (21) [M+
휈
+ H], 438 (7) [M+ - OMe], 378 (12) [M+ - Bn], 314 (22), 299 (44), 91 (100) [Bn+].
cis- and trans-2-Benzyl-3-methoxy-5-nonyl-1-tosyl-2,5-dihydro-pyrrole (13)
According to GP2, allenyl amine pro-trans-12 (0.200 g, 0.43 mmol) in acetonitrile (8 mL) was treated with AgNO3 (0.015 g, 0.09
mmol) and K2CO3 (0.118 g, 0.86 mmol) for 12 h to provide crude 13 (0.220 g, d.r. 55:45) as brown oil that was purified by column
chromatography (silica gel, hexanes/ethyl acetate 9:1) to give the pure diastereomers of 13 (0.174 g, 87%). Separation HPLC
(hexanes/ethyl acetate 93:7) furnished trans-13 (0.060 g, 30%) as colorless crystals (m.p. 66 °C) and cis-13 (0.083 g, 42%) as
colorless crystals (m.p. 57–58 °C).
Data of cis-13: 1H NMR (CDCl3, 500 MHz):  = 0.89 (t, J = 6.6 Hz, 3 H, Me), 0.95–1.41 (m, 16 H, CH2), 2.42 (s, 3 H, Tos-Me),
3.06, 3.17 (AB part of ABX system, JAB = 13.6 Hz, JAX = 2.6 Hz, JBX = 5.5 Hz, 1 H each, PhCH2), 3.54 (s, 3 H, OMe), 3.94–4.02
(m, 1 H, 5-H), 4.27 (s, 1 H, 4-H), 4.37–4.43 (m, 1 H, 2-H), 7.15–7.27 (m, 5 H, Ph), 7.30, 7.71 ppm (2 d, J = 8.1 Hz, 2 H each, Tos).
13C NMR (CDCl3, 125.8 MHz):  = 14.0 (q, Me), 21.5 (q, Tos-Me), 22.7, 28.7, 25.6, 28.7, 29.3, 29.4, 29.5, 31.9 (8 t, CH2), 38.8 (t,

Organic & Biomolecular Chemistry
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PhCH2), 56.6 (q, OMe), 65.0, 65.2 (2 d, C-2, C-5), 94.4 (d, C-4), 126.3, 127.5, 127.7, 129.8, 131.0 (5 d, Ph, Tos), 134.3,V1ie3w6A.4rt,icle Online
DOI: 10.1039/C8OB02645A
143.3 (3 s, Ph, Tos), 154.0 ppm (s, C-3). IR (KBr): = 3060, 3030, 2925, 2855 (C-H), 1670 (C=C), 1345, 1165 cm-1 (Tos-N). MS
휈
(EI, 80 eV): m/z (%) = 469 (0.3) [M+], 378 (100) [M+ - Bn], 342 (20) [M+ - C9H19], 91 (100) [Bn+]. C28H39NO3S (469.3): calcd. C 71.61,
H 8.37, N 2.98; found C 71.62, H 8.22, N 2.91.
Data of trans-13: 1H NMR (CDCl3, 500 MHz):  = 0.88 (t, J = 7.1 Hz, 3 H, Me), 0.90–1.31, 1.80–1.88 (2 m, 15 H, 1 H, CH2), 2.42
(s, 3 H, Tos-Me), 3.01 (dd, J = 13.7 Hz, J = 2.2 Hz, 1 H, PhCH2), 3.56 (s, 3 H, OMe), 3.74 (dd, J = 13.7 Hz, J = 4.5 Hz, 1 H,
PhCH2), 3.97–4.02 (m, 1 H, 5-H), 4.23 (s, 1 H, 4-H), 4.71 (sbr, 1 H, 2-H), 7.19–7.28 (m, 3 H, Ph), 7.27 (d, J = 8.1 Hz, 2 H, Tos),
7.35 (d, J = 6.9 Hz, 2 H, Ph), 7.77 ppm (d, J = 8.1 Hz, 2 H, Tos). 13C NMR (CDCl3, 125.8 MHz):  = 14.1 (q, Me), 21.4 (q, Tos-
Me), 22.6, 25.1, 29.2, 29.4*, 29.5, 31.8, 33.9, (8 t, CH2), 38.4 (t, PhCH2), 56.3 (q, OMe), 65.7 (2 d, C-2, C-5), 94.6 (d, C-4), 126.1,
126.6, 127.6, 129.3, 130.5 (5 d, Ph, Tos), 136.5, 140.1, 142.6 (3 s, Ph, Tos),153.8 ppm (s, C-3); signal with higher intensity. IR
(KBr): = 3030, 2925, 2855 (C-H), 1675 (C=C), 1335, 1160 cm-1 (Tos-N). MS (EI, 80 eV): m/z (%) = 469 (0.5) [M+], 378 (100) [M+
휈
- Bn], 342 (12) [M+ - C9H19], 91 (50) [Bn+]. HRMS (EI, 80 eV): calcd. for C28H39NO2S: 469.2651; found 469.2636. C28H39NO3S
(469.3): calcd. C 71.61, H 8.37, N 2.98; found C 71.65, H 8.33, N 2.95.
2-Benzyl-3-methoxy-5-nonyl-1-tosyl-2,5-dihydropyrrole (cis-13)
A solution of pro-cis-12 (0.520 g, 1.11 mmol) in 15 mL of dry DMSO was heated to 50 °C. In a stream of argon, freshly sublimed
KOtBu (0.031 g, 0.28 mmol) was added via a funnel and the mixture was stirred at this temperature for 12 h. After cooling to room
temperature, saturated aqueous NaHCO3 solution (10 mL) was added. The organic phase was separated and the aqueous phase
was extracted with ethyl acetate (2 x 20 mL). The combined organic phases were washed with saturated aqueous NaHCO3
solution (2 x 20 mL), dried (Na2SO4), filtered and evaporated in vacuo to provide crude cis-13 (0.534 g) as yellow oil. The crude
product was purified by column chromatography (silica gel, hexanes/ethyl acetate) to furnish cis-13 (0.405 g, 78%) as colorless
crystals (m,p. 57–58 °C). For the analytical data see above.
2-Benzyl-5-nonyl-1-tosylpyrrolidin-3-one (cis-14)
According to GP3, cis-13 (0.120 g, 0.26 mmol) in THF (6 mL) and 20% aqueous H2SO4 (4 mL) provided after 4 h crude cis-14
(0.189 g) as light yellow oil. Purification by column chromatography (silica gel, hexanes/ethyl acetate 7:1) furnished cis-14 (0.116
g, quant.) as colorless crystals (m.p. 63–65 °C).
1H NMR (CDCl3, 270 MHz):  = 0.90 (t, J = 6.7 Hz, 3 H, Me), 0.90–1.40 (m, 16 H, CH2), 1.76, 2.14 (AB part of ABX system, JAB
18.2 Hz, JAX = 3.0 Hz, JBX = 9.3 Hz, JB,CH = 1.3 Hz, 1 H each, 4-H), 2.43 (s, 3 H, Tos-Me), 3.22, 3.27, 3.93 (ABX system, JAB
=
=
13.5 Hz, JAX = 5.7 Hz, JBX = 4.0 Hz, 1 H each, PhCH2, 2-H), 3.77–3.88 (m, 1 H, 5-H), 7.20–7.30 (m, 5 H, Ph), 7.33, 7.73 ppm (2
d, J = 8.3 Hz, 2 H each, Tos). 13C NMR (CDCl3, 67.9 MHz):  = 14.1 (q, Me), 21.5 (q, Tos-Me), 22.6, 25.8, 29.0, 29.2, 29.3, 29.4,
31.8, 37.0, 37.9, 42.2 (10 t, CH2, PhCH2, C-4,), 56.8, 65.4 (2 d, C-2, C-5), 126.9, 127.5, 128.2, 130.0, 130.9 (5 d, Ph, Tos), 134.1,
136.2, 144.1 (3 s, Ph, Tos), 211.3 ppm (s, C-3). IR (KBr): = 3060–2855 (CH), 1760 (C=O), 1355, 1155 cm-1 (Tos-N). MS (EI, 80
휈
eV): m/z (%) = 455 (7) [M+], 364 (100) [M+ - Bn], 155 (88) [Tos+], 91 (82) [Bn+].
2-Benzyl-5-nonyl-1-tosylpyrrolidin-3-one (trans-14)
According to GP3, trans-13 (0.060 g, 0.13 mmol) in THF (4 mL) and 20% aqueous H2SO4 (2 mL) provided after 4 h crude trans-
14 (0.058 g, quant.) as light yellow solid that was not purified but directly used in the epimerization experiments.
1H NMR (CDCl3, 270 MHz):  = 0.87 (t, J = 6.8 Hz, 3 H, Me), 0.90–1.35, 1.60–1.70 (2 m, 15 H, 1 H, CH2), 1.65, 1.99 (AB part of
ABX system, JAB = 17.2 Hz, JAX = 1.0 Hz, JBX=9.0 Hz, 1 H each, 4-H), 2.44 (s, 3 H, Tos-Me), 3.12 (dd, J = 13.7 Hz, J = 3.2 Hz, 1
H, PhCH2), 3.64 (dd, J = 13.7 Hz, J = 5.1 Hz, 1 H, PhCH2), 3.94 (dd, J = 5.0 Hz, J = 3.2 Hz, 1 H, 2-H), 4.00–4.10 (m, 1 H, 5-H),
7.20–7.36 (m, 7 H, Ph, Tos), 7.77 ppm (d, J = 8.3 Hz, 2 H, Tos). 13C NMR (CDCl3, 67.9 MHz):  = 14.1 (q, Me), 21.5 (q, Tos-Me),
22.6, 24.6, 27.4, 29.3,* 29.4, 31.8, 33.2, 37.9, 42.8 (9 t, CH2, PhCH2, C-4), 57.6, 64.8 (2 d, C-2, C-5), 127.0, 127.2, 128.3, 129.7,
130.6 (5 d, Ph, Tos), 135.3, 137.8, 143.6 (3 s, Ph, Tos), 210.9 ppm (s, C-3); * signal with higher intensity.
2-Benzyl-3-methoxy-1-methyl-5-nonyl-2,5-dihydropyrrole (cis-18)
A solution of sodium naphthalenide (0.98 mL of a 1 M solution in DME, prepared according to ref. 25) was added to a solution of
cis-13 (0.153 g, 0.33 mmol) in DME (17 mL) at -78 °C. After 1.5 h the mixture was quenched at this temperature with methyl iodide
(0.468 g, 3.30 mmol), then stirred at room temperature for 1 h and concentrated in vacuo. The residue was taken up in saturated
aqueous NaHCO3 solution (20 mL) and extracted with ethyl acetate (3 x 10 mL). The combined organic phases were dried

Page 15 of 20
Organic & Biomolecular Chemistry
(Na2SO4), filtered and evaporated in vacuo to obtain the crude product (0.250 g) as yellow solid. Purification by cVoielwumArnticle Online
DOI: 10.1039/C8OB02645A
chromatography (alumina III, hexanes/ethyl acetate 25:1 + 1% of NEt3) gave cis-18 (0.057 g, 53%) as yellow oil.
1H NMR (CDCl3, 270 MHz):  = 0.88 (t, J = 6.6 Hz, 3 H, Me), 1.05–1.40 (m, 16 H, CH2), 2.19 (s, 3 H, Me), 2.67, 2.95 (AB part of
ABX system, JAB = 13.2 Hz, JAX = 5.9 Hz, JBX = 4.4 Hz, 1 H each, PhCH2), 3.08–3.17, 3.46–3.53 (2 m, 1 H each, 2-H, 5-H), 3.57
(s, 3 H, OMe), 4.39 (s, 1 H, 4-H), 7.10–7.30 ppm (m, 5 H, Ph). 13C NMR (CDCl3, 67.9 MHz):  = 14.1 (q, Me), 22.7, 25.7, 29.4,
29.6, 29.7, 30.0, 31.9, 36.8 (8 t, CH2), 40.1, 40.9 (t, q, PhCH2, NMe), 56.5 (q, OMe), 69.5, 71.7 (2 d, C-2, C-5), 95.0 (d, C-4),
125.5, 127.6, 130.1, 139.5 (3 d, s, Ph), 157.7 ppm (s, C-3). IR (film): = 3085–2775 (C-H), 1660 cm-1 (C=C). MS (EI, 80 eV): m/z
휈
(%) = 329 (0.7) [M+], 328 (3) [M+ - H], 238 (100) [M+ - Bn], 202 (48) [M+ - C9H19], 91 (25) [Bn+].
2-Benzyl-1-methyl-5-nonylpyrrolidin-3-one (cis-19)
According to GP3, cis-18 (0.054 g, 0.16 mmol) in THF (4 mL) and 20% aqueous H2SO4 (4 mL) provided after 4 h crude cis-19
(0.053 g) as yellow oil. Purification by column chromatography (alumina III, hexanes/ethyl acetate 20:1 + 1% of NEt3) furnished
cis-19 (0.031 g, 60%) as light yellow oil.
1H NMR (CDCl3, 270 MHz):  = 0.88 (t, J = 6.7 Hz, 3 H, Me), 1.10–1.38, 1.68–1.84 (2 m, 15 H, 1 H, CH2), 1.76 (dd, J = 17.4 Hz,
J = 10.3 Hz, 1 H, 4-H), 2.31 (s, 3 H, NMe), 2.38 (dd, J = 17.4 Hz, J = 5.9 Hz, 1 H, 4-H), 2.37–2.53 (m, 1 H, 5-H), 2.75 (X part of
ABX system, JAX = 5.1 Hz, JBX = 4.7 Hz, 1 H, 2-H), 2.84, 3.05 (AB part of ABX system, JAB = 14.2 Hz, JAX = 5.1 Hz, JBX = 4.7 Hz,
1 H each, PhCH2), 7.14–7.29 ppm (m, 5 H, Ph). 13C NMR (CDCl3, 67.9 MHz):  = 14.1 (q, Me), 22.7, 25.6, 29.3, 29.5, 29.6, 29.8,
31.9, 32.9, 35.9, 42.8 (10 t, CH2, PhCH2, C-4), 39.2 (q, NMe), 62.5, 74.3 (2 d, C-2, C-5), 126.1, 128.0, 129.7, 138.5 (3 d, s, Ph),
214.8 ppm (s, C-3). IR (film): = 3060–2855 (C-H), 1755 cm-1 (C=O). MS (EI, 80 eV): m/z (%) = 315 (1.5) [M+], 224 (100) [M+ -
휈
Bn], 91 (21) [Bn+]. HRMS (EI, 80 eV): calcd. for C21H33NO [M+]: 315.2562; found 315.2539; calcd. for C14H26NO [M+ - Bn] 224.2014;
found 224.2042.
2-Benzyl-1-methyl-5-nonylpyrrolidin-3-ol (all-cis-20), rac-preussin
According to GP5, a solution of cis-19 (0.010 g, 0.04 mmol), L-Selectride (0.06 mL of 1 M solution in THF, 0.73 mmol) in THF (4
mL) provided after 3 h the crude product (0.038 g) as yellow oil. Purification by column chromatography (silica gel,
dichloromethane/methanol 9:1) provided pure all-cis-20 (0.008 g, 80%, d.r. >99:1 according to HPLC) as light yellow oil.
1H NMR (CDCl3, 270 MHz):  = 0.88 (t, J = 6.6 Hz, 3 H, Me), 1.15–1.36 (m, 16 H, CH2), 1.42 (ddd, J = 13.0 Hz, J = 5.5 Hz, J = 1.4
Hz, 1 H, 4-H), 1.65–1.78 (m, 1 H, OH), 2.05–2.15 (m, 1 H, 5-H), 2.18 (dd, J = 13.0 Hz, J = 6.4 Hz, 1 H, 4-H), 2.27, 2.84, 2.89 (X
part and AB part of ABX system, JAB = 13.4 Hz, JAX = 5.3 Hz, JBX = 9.1 Hz, 1 H each, 2-H, PhCH2), 2.34 (s, 3 H, NMe), 3.76–3.84
(m, 1 H, 3-H), 7.16–7.32 ppm (m, 5 H, Ph). 13C NMR (CDCl3, 67.9 MHz):  = 22.7, 26.3, 29.3, 29.5, 29.6, 29.9, 31.9, 33.7, 34.9,
39.2 (10 t, CH2, PhCH2, C-4), 38.6 (q, NMe), 65.8, 70.4, 73.6 (3 d, C-2, C-3, C-5), 126.0, 128.4, 129.3, 139.4 ppm (3 d, s, Ph).
The spectroscopic data of the sample agree with those reported in literature.26
(2R,3R,5S)-2-Benzyl-5-nonyl-1-tosylpyrrolidin-3-ol (all-cis-21)
According to GP5, a solution of 2R,5S-cis-15 (0.055 g, 0.12 mmol), L-Selectride (0.24 mL of 1 M solution in THF, 0.24 mmol) in
THF (5 mL) provided after 6 h the crude product that still contained starting material. It was treated again with L-Selectride (0.96
mL of 1 M solution in THF, 0.96 mmol) in THF (5 mL) for 3 h and finally furnished crude 2R,3R,5S-all-cis-21 (0.080 g) as yellow
oil., d.r. 90:10). Pre-purification by column chromatography (silica gel, hexanes/ethyl acetate 4:1) and purification by HPLC
(hexanes/ethyl acetate 4:1) afforded diastereomerically pure 2R,3R,5R-all-cis-21 (0.040 g,73%) as colorless oil.
[]20
Optical rotation:
= -20.4 (c = 0.8, CHCl3). 1H NMR (CDCl3, 270 MHz):  = 0.89 (t, J = 6.4 Hz, 3 H, Me), 1.20–1.40, 1.50–1.80,
D
1.90–2.10 (3 m, 14 H, 4 H, 1 H, CH2, 4-H, OH), 2.42 (s, 3 H, Tos-Me), 3.08, 3.24 (AB part of ABX system, JAB = 13.9 Hz, JAX = 9.0
Hz, JBX = 3.9 Hz, 1 H each, PhCH2), 3.55–3.70 (m, 1 H, 5-H), 3.80–3.90 (m, 2 H, 2-H, 3-H), 7.17–7.30 (m, 7 H, Ph, Tos), 7.71
ppm (d, J = 8.0 Hz, 2 H, Tos). 13C NMR (CDCl3, 67.9 MHz):  = 14.1 (q, Me), 21.5 (q, Tos-Me), 22.7, 26.5, 29.3, 29.4, 29.5, 29.6,
31.9, 37.1, 37.3, 37.8 (10 t, CH2, C-4, PhCH2), 59.9, 65.6, 71.6 (3 d, C-2, C-3, C-5), 126.3, 127.6, 128.5, 129.4, 129.7 (5 d, Ph,
Tos), 135.0, 128.9, 143.4 ppm (3 s, Ph, Tos). IR (film): = 3520 (OH), 3085–2855 (C-H), 1600 (Ph), 1340, 1160 cm-1 (Tos-N). MS
휈
(EI, 80 eV): m/z (%) = 457 (0.1) [M+], 456 (0.2) [M+ - H], 366 (100) [M+ - Bn], 155 (20) [Tos+], 91 (52) [Bn+]. HRMS (EI, 80 eV):
calcd. for C27H39NO3S [M+]: 457.2651; found 457.2677; calcd. for C27H38NO3S [M+-H]: 456.2572; found 456.2547.
(2R,3R,5S)-2-Benzyl-5-nonylpyrrolidin-3-ol (all-cis-22)
A solution of sodium naphthalenide (0.44 mL of a 1 M solution in DME, 0.44 mmol, prepared according to ref. 25) was added to a
solution of all-cis-21 (0.040 g, 0.09 mmol) in DME (7 mL) at -78 °C. After 1.5 h the mixture was quenched with water (5 mL) and

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concentrated in vacuo. The residue was taken up in saturated aqueous NaHCO3 solution (20 mL) and extracted with ethyl aVcieewtaAtreticle Online
DOI: 10.1039/C8OB02645A
(3 x 10 mL). The combined organic phases were dried (Na2SO4), filtered and evaporated in vacuo to obtain the crude product
(0.090 g) as yellow solid. Purification by column chromatography (silica gel, hexanes/ethyl acetate 1:3, + 1% of NEt3) gave all-cis-
22 (0.021 g, 87%) as light yellow crystals (melting range 96-100 °C; ref. 29: m.p. of the enantiomer: 101-102 °C). The
spectroscopic data agree with those of the enantiomer.29
[]20
[]20
Optical rotation:
= +14.1 (c = 0.4, CHCl3); for the enantiomer
= -15.6 (c = 1.0, MeOH).29 1H NMR (CDCl3, 270 MHz):  =
D
D
0.86 (t, J = 6.6 Hz, 3 H, Me), 1.20–1.38 (m, 14 H, CH2), 1.35 (ddd, J = 14.2 Hz, J = 6.7 Hz, J = 1.5 Hz, 1 H, 4-H), 1.45–1.60 (m, 2
H, CH2), 2.08–2.25 (m, 2 H, OH, NH), 2.26 (ddd, J = 14.2 Hz, J = 8.6 Hz, J = 6.2 Hz, 1 H, 4-H), 2.85, 2.94 (AB part of ABX system,
JAB = 13.0 Hz, JAX = 7.4 Hz, JBX = 6.6 Hz, 2 H, PhCH2), 2.89–3.07 (m, 2 H, 2-H, 5-H), 3.99 (ddd, J = 6.2 Hz, J = 3.4 Hz, J = 1.5
Hz, 1 H, 3-H), 7.16–7.36 ppm (m, 5 H, Ph). 13C NMR (CDCl3, 67.9 MHz):  = 14.0 (q, Me), 22.7, 27.2, 29.3, 29.5, 29.6, 29.7, 31.9,
35.6, 37.5, 42.0 (10 t, CH2, PhCH2, C-4), 57.0, 65.7, 72.2 (3 d, C-2, C-3, C-5), 126.1, 128.5, 128.9, 139.9 ppm (3 d, s, Ph). IR
(KBr): = 3420 (N-H), 3065, 3030, 2925, 2855 cm-1 (C-H). MS (EI, 80 eV): m/z (%) = 303 (0.5) [M+], 302 (2) [M+ - H], 212 (100)
휈
[M+ - Bn], 176 (22) [C9H19+], 91 (16) [Bn+]. HRMS (EI, 80 eV): calcd. for C18H20NO [M+]: 303.2562; found 303.2575; calcd. for
C18H19NO [M+ - H]: 302.2484; found 302.2442.
(2R,3R,5S)-2-Benzyl-1-methyl-5-nonylpyrrolidin-3-ol (all-cis-20), (-)-Preussin
2R,3R,5S-all-cis-22 (0.019 g, 0.06 mmol) was dissolved in a 1:1 mixture of THF/acetonitrile (4 mL) and aqueous formaldehyde
(0.03 mL, 35%, 0.38 mmol) was added dropwise. After 5 min sodium cyanoborohydride (0.008 g, 0.13 mmol) and after 1 h stirring
at room temperature acetic acid (0.5 mL) were added. After 30 min solid NaHCO3 was added, with aqueous 2 N NaOH the solution
adjusted to pH 10 and the organic phase was separated. The aqueous phase was extracted with dichloromethane (3 x 10 mL)
and the combined organic phases were dried (Na2SO4). After filtration and evaporation of solvents in vacuo the residue was
purified by column chromatography (silica gel, dichloro-methane/methanol 9:1) to afford pure 2R,3R,5S-all-cis-20 (0.016 g, 80 %)
as yellow oil.
[]2D0
Optical rotation:
(see above).
= -25.8 (c = 0.8, CHCl3); for literature values see ref. 28. The NMR data agree with those of rac-preussin
MCF-7 cell proliferation assay
MCF-7 cells (ATCC HTB-22, USA) were seeded in RPMI 1640 medium (Biochrome, Germany) supplemented with fetal calf serum
(10%; PAA, Austria), l-glutamine (2 mM), estradiol (0.1 nM), and insulin (1 U/mL) at 5000 cells per well in 96-well plates. Cells
were allowed to adhere for 24 h, and then fresh growth medium plus compound was added. The final concentration of DMSO was
0.5%. After four days of continuous incubation, the cells were fixed with glutaraldehyde and stained with crystal violet, and the
absorbance was recorded at 595 nm using Tecan Sunrise equipment. All measurements were performed in quadruplicate. The
values were normalized to the absorbance of solvent-treated cells (100 %), and the absorbance of a reference plate which was
fixed at the time point of compound application (0 %). Half-maximal growth inhibition (IC50) was determined as the compound
concentration required to achieve 50% inhibition of cell growth.
Conflicts of interest
There are no conflicts of interest to declare.
Acknowledgements
Generous support of this work by the Fonds der Chemischen Industrie (Kekulé fellowship for Arndt Hausherr), the Deutsche
Forschungsgemeinschaft and Schering AG is most gratefully acknowledged. We thank Winfried Münch and Christiane Groneberg
for their assistance in analytical work and Dr. Reinhold Zimmer for his great help during preparation of the manuscript.
Notes and references
1 (a) D. O’Hagan, Nat. Prod. Rep. 2000, 17, 435–446; (b) N. Asano, R. J. Nash, R. J. Molyneux and G. W. Fleet,
Tetrahedron: Asymmetry 2000, 11, 1645−1680; (c) J. R. Liddell, Nat. Prod. Rep. 2002, 19, 773–781; (d) H. Yoda,
Curr. Org. Chem. 2002, 6, 223–243; (e) P. Compain and O. R. Martin, Iminosugars: from Synthesis to Therapeutic

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Applications, Wiley-VCH: Weinheim, New York, 2007; (f) J. P. Michael, Nat. Prod. Rep. 2007, 24, 191–222; (g)
DOI: 10.1039/C8OB02645A
V. Desvergnes and Y. Landais, Stud. Nat. Prod. Chem. 2014, 42, 373−419.
2 Reviews on the use of alkoxyallenes in organic synthesis: (a) R. Zimmer, Synthesis 1993, 165–178; (b) M.
Brasholz, H.-U. Reissig and R. Zimmer, Acc. Chem. Res. 2009, 42, 45–56; (c) F. Pfrengle and H.-U. Reissig,
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2374 (AC).
17 Typical examples: a) S. Hoff, L. Brandsma and J. F. Arens, Recl. Trav. Chim. Pays-Bas 1969, 88, 609–619; (b)
D. Gange and P. Magnus, J. Am. Chem. Soc. 1978, 100, 7746–7747; (c) S. Hormuth and H.-U. Reissig, J. Org.
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18 Typical examples: a) M. G. Okala Amombo, A. Hausherr and H.-U. Reissig, Synlett 1999, 1871–1874; (b) O.
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19 Recent computational study: F. Cumine, A. Young, H.-U. Reissig, T. Tuttle and J. A. Murphy, Eur. J. Org. Chem.
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20 (a) A. Hausherr, B. Orschel, S. Scherer and H.-U. Reissig, Synthesis 2001, 1377–1385; (b) A. Hausherr and H.-
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21 A. Hausherr and H.-U. Reissig, Eur. J. Org. Chem. 2018, 4071–4080.
22 A. Hausherr, R. Zimmer and H.-U. Reissig, Synthesis 2018, 50, doi.org/10.1055/s-0037-1609942.
23 M. Haddad, F. Dahan, J.-P. Legros, L. Lopez, M.-T. Boisdon and J. Barrans, J. Chem. Soc., Perkin Trans. 2
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24 F. Chemla, V. Hebbe and J.-F. Normant, Synthesis 2000, 75-77.
25 A. B. Smith, T. A. Rano, N. Chida, A. G. Sulikowski and J. L. Wood, J. Am. Chem. Soc. 1992, 114, 8008–8022.
26 See for instance: a) T. Bach, H. Brummerhop and K. Harms, Chem. Eur. J. 2000, 6, 3838–3848.
27 (a) H. M. Davies, J. J. Matasi, L. M. Hodges, N. J. S. Huby, C. Thornley, N. Kong and J. H. Houser, J. Org.
Chem. 1997, 62, 1095-1105; (b) R. Lin, J. Castells and H. Rapoport, J. Org. Chem. 1998, 63, 4069-4078.
28 Optical rotations reported for (-)-preussin: (a) -21.6 (c 1.0, CHCl3): W. Deng and L. E. Overman, J. Am. Chem.
Soc. 1994, 116, 11241–11250; b) -28.8 (c 1.01 CHCl3) M. Okue, H. Watanabe, K. Kasahara, M. Yoshida, S.

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16s.
29 A. Kanazawa, S. Gillet, P. Delair and A. E. Greene, J. Org. Chem. 1998, 63, 4660–4663.

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DOI: 10.1039/C8OB02645A
Alkoxyallene-Based Syntheses of Preussin and Analogs and Their Cytotoxicity
Arndt Hausherr, Gerhard Siemeister and Hans-Ulrich Reissig
OH
OH
OR
H
or
•
+
Non
Non
Ph
O
N
Non
Ph
Ph
X
Me
oxa-preussin
(-)-preussin
X = O or N-Tos
Preussin made in Prussia! Axially chiral alkoxyallenes provided oxa-preussin, racemic and
enantiopure (-)-preussin (cytotoxicities: IC50 = 3-6 μM).