A direct stereoselective preparation of a fish pheromone and application of the zinc porphyrin tweezer chiroptical protocol in its stereochemical assignment

Article abstract of DOI:10.1002/chir.22186

A two-step stereoselective preparation of a goldfish pheromone, 17α,20β-dihydroxy-4-pregnen-3-one, is reported from the readily available cortexolone in 64% overall yield. The (20S)-epimer was also synthesized in three steps from cortexolone with an overall yield of 47%. A microscale chiroptical technique based on a host/guest complexation mechanism between the substrate and a dimeric metalloporphyrin host (tweezer) was used to confirm the stereochemical assignment, while Density Functional Theory (DFT) calculations were employed to explain the high stereoselectivity induced by the 17α-hydroxyl and C18-methyl groups. Chirality 00:000-000:, 2013. 2013 Wiley Periodicals, Inc.

Full text of DOI:10.1002/chir.22186

CHIRALITY 25:575–581 (2013)
A Direct Stereoselective Preparation of a Fish Pheromone and
Application of the Zinc Porphyrin Tweezer Chiroptical Protocol in Its
Stereochemical Assignment
1
1
1
2
2
YANNICK P. OUEDRAOGO, LONGCHUAN HUANG, MARIANA P. TORRENTE, GLORIA PRONI, EKATERINA CHADWICK,
1
1
RUDOLF J. WEHMSCHULTE, AND NASRI NESNAS *
1
Department of Chemistry, Florida Institute of Technology, Melbourne, Florida
Science Department, John Jay College of Criminal Justice, New York, New York
2
ABSTRACT
A two-step stereoselective preparation of a goldfish pheromone, 17a,20b-
dihydroxy-4-pregnen-3-one, is reported from the readily available cortexolone in 64% overall
yield. The (20S)-epimer was also synthesized in three steps from cortexolone with an overall
yield of 47%. A microscale chiroptical technique based on a host/guest complexation mechanism
between the substrate and a dimeric metalloporphyrin host (tweezer) was used to confirm the
stereochemical assignment, while Density Functional Theory (DFT) calculations were employed
to explain the high stereoselectivity induced by the 17a-hydroxyl and C18-methyl groups. Chirality
25:575–581, 2013. © 2013 Wiley Periodicals, Inc.
KEY WORDS: goldfish pheromone; stereoselective reduction; cortexolone; circular dichroism;
DFT; computational modeling; zinc porphyrin tweezer
1
13
INTRODUCTION
chromatography were performed on silica gel 60 F254. H and
nuclear magnetic resonance (NMR) spectra were recorded on 400 or
60 MHz spectrometers. CDCl and DMSO-d were used as NMR sol-
C
We herein report a direct enantioselective preparation
of 17a,20b-dihydroxy-4-pregnen-3-one, (20R)-1, a goldfish
Carassius auratus) pheromone, from cortexolone
Scheme 1). The pheromone is at least a thousand times
more expensive than the readily available corticosteroid,
cortexolone. The female goldfish releases the pheromone
3
3
6
vents and their signals were used as internal standards. Chemical shifts
were expressed in ppm, followed by multiplicity (s, singlet; d, doublet; t,
triplet; q, quartet; quin, quintet; m, multiplet; br, broad), and the number
of corresponding protons. High-resolution mass spectrometry (HRMS) of
positive ions was obtained on an AccuTOF mass spectrometer with a
DART ion source. The circular dichroism (CD) spectra were measured
(
(
1
–4
causing an increased sperm production in males.
-
1
-1
5
–9
in millidegrees and normalized into emax [Lmol cm ]/l [nm] units.
An extensive body of research conducted on the phero-
mone’s biological activity emphasizes its importance and its
potential to be a key element for the aquaculture of other
2
1-Chloro-17a-hydroxy-4-pregnen-3,20-dione (2). Cortexolone
(500 mg, 1.44 mmol) and mesyl chloride (330 mg, 2.88 mmol)
were dissolved in CH Cl (20 ml), and a catalytic amount
2 mg) of DMAP was slowly added and the mixture was
refluxed for 4 h. The crude was diluted with 10 ml of CH Cl2
and washed with brine (10 ml) three times. The organic layer
was then dried with Na SO and the solvent was evaporated.
organisms.1
0–15
Outstanding work by Sorensen and coworkers
2
2
on sea lampreys (Petromyzon marinus) strongly suggests
that other fish pheromones have a related steroidal skele-
(
7
,8,16,17
Previous syntheses include the classical18 and
2
ton.
enzymatic reduction of 17-hydroprogesterone precursor.
1
9
2
4
The synthetic approach employed in this article owes its
strength to both its stereoselective nature and the ease
of its extension to a wide variety of other potentially
biologically active pheromones.
The initial synthetic approach was based on a one-pot
hydride reduction of the readily generated chlorocortexolone
Purification by column chromatography and recrystallization
ꢀ
in EtOAc afforded 2 (443 mg, 85%), mp = 227-230 C; IR n/
-
1
cm = 3495 (br, 17-OH), 1724 (C = O, at C20), 1637
1
(
C = C-C = O); H NMR (360 MHz, DMSO-d ) d = 5.63 (s, 1H,
H-C = C), 5.54 (s, 1H, 17-OH), 4.77 (d, J = 16.8 Hz, 1H of CH Cl),
.47 (d, J = 16.9 Hz, 1H of CH Cl), 2.55 (t, J =13.2Hz, 1H, 16b-H),
.40 (dt, J =14.9, 3.4 Hz, 2H: overlap of 6b-Hax and 2b-Hax),
.24 (m, 1H, 6a-H ), 2.15 (m, 1H, 2a-H ), 1.96 (dq, J = 13.3,
.3 Hz, 1H, 1b-H ), 1.79 (m, 1H, 7b-H ), 1.66-1.75 (m, 2H,
overlap of 12b-Heq and 14-H), 1.51-1.66 (m, 4H, overlap of
5a-H, 1a-H , 8-H, and 11a-H ), 1.30-1.50 (m, 3H, overlap
6
2
4
2
2
2
2
2
(Scheme 2). Reduction of 2 generates chloroalcohol 3,
which, under reductive conditions in the presence of the
alkoxide, may close up to form epoxide 4. The epoxide, in
the presence of an additional equivalent of hydride may open
up from the less hindered side to afford target pheromone 1
after workup. Epoxide 4 itself can be used as a key intermediate
in the generation of multiple variants of the pheromone in an
attempt to target other aquatic species. For instance, nucleophilic
opening of the epoxide using alkyl aluminates may afford the
general target structure 5.
eq
eq
eq
eq
1
ax
eq
of 16a-H, 11b-Hax, and 12a-H ), 1.23 (m, 1H, 15b-H), 1.14
ax
(
s, 3H, 19-CH ), 1.01 (dq, J = 11.8, 2.9 Hz, 1H, 7a-H ), 0.91
3
a
x
Additional Supporting Information may be found in the online version of this
article.
Correspondence to: N. Nesnas, Department of Chemistry, Florida Insti-
*
tute of Technology, 150 West University Boulevard, Melbourne, Florida
2901. E-mail: nesnas@fit.edu
Received for publication 01 March 2013; Revised 04 April 2013; Accepted
EXPERIMENTAL
3
General
04 April 2013
Anhydrous solvents were dried and distilled (THF from potassium,
DOI: 10.1002/chir.22186
Published online 26 June 2013 in Wiley Online Library
(wileyonlinelibrary.com).
CH
chromatography (HPLC) grade or reagent grade. Column and flash
2013 Wiley Periodicals, Inc.
2 2 2
Cl from CaH ). Solvents used were either high-performance liquid
©

5
76
OUEDRAOGO ET AL.
1
7a, 20b-Dihyroxy-4-pregnen-3-one [(20R)-1]. Pheromone (20R)-1
was obtained in a one-pot reductive procedure after the dechlo-
rination of 2 with Ti to generate 6 in situ. To the latter formed
after 10 h of reaction time, NaBH (23 mg, 0.61 mmol) was
3+
4
added. The reduction was complete in 30 min, after which the
mixture was extracted with EtOAc. The organic layers were
combined, washed with brine, dried over Na SO , rotavaped,
2
4
and then purified on a silica gel column using 25% EtOAc/
hexanes. Product (20R)-1 (125 mg, 0.376 mmol) was
obtained with an overall yield of 64% from cortexolone,
Scheme 1. Fish pheromone in two steps from cortexolone.
and showed identical NMR spectra to the authentic phero-
ꢀ
mone. R = 0.43 (50% EtOAc/hexanes), mp = 199-203 C; IR
f
-
1
n/cm = 3444 (br, OH), 2938 (aliphatic C-H), 1659 (C = C-
C = O); 1H NMR (400MHz, DMSO-d ): d = 5.62 (s, 1H, H-
6
C = C), 4.04 (d, J =7.1 Hz, 1H, 20-OH), 3.75 (app quin,
J = 6.6 Hz, 1H, 20-H), 3.43 (s, 1H, 17-OH), 2.39 (m, 2H: overlap
of 6b-Hax and 2b-H ), 2.15-2.24 (m, 2H, 6a-H overlap with
ax
eq
2
a-H ), 1.95 (m, 1H, 1b-H ), 1.80 (m, 1H, 7b-H ), 1.64-1.75
eq eq eq
(
m, 2H, overlap of 12b-Heq and 14-H), 1.42-1.63 (m, 5H,
overlap of 16b, 15a-H, 1a-H , 8-H, and 11a-H ), 1.20-1.41
ax
eq
(
m, 3H, overlap of 16a-H, 11b-H , and 12a-H ), 1.19 (m,
ax ax
1
2
0
H, 15b-H), 1.18 (s, 3H, 19-CH ), 1.00 (d, J = 6.3 Hz, 3H,
3
1-CH3 overlap with m, 1H, 7a-H ), 0.82 (m, 1H, 9-H),
ax
.76 (s, 3H, 18-CH ); HRMS m/z calcd. for C H O
M + H] 333.2430, found 333.2392.
3
21 33
3
+
[
21-Chloro-17a,20a-dihydroxy-4-pregnene-3-one [(20S)-3]. Inter-
mediate 2 (200 mg, 0.55 mmol) was dissolved in 20 ml of
anhydrous THF, and 2.5 ml of 0.5 M HAlO in anhydrous
THF were added while stirring. The mixture was stirred at
room temperature for 3 h, then extracted with EtOAc
Scheme 2. General concept of a one-pot basic reduction. The stereochemistry
of C20 was intentionally not shown here since it is dependent on the reductive
method employed as explained below.
(
20 ml) three times. The combined organic layers were
washed with brine (15 ml) three times, dried over Na SO ,
2
4
and concentrated in vacuo and purified on silica gel to
afford 198 mg (98% yield) of chloroalcohol (20S)-3. R = 0.49
f
(
dt, J = 11.9, 3.1 Hz, 9-H), 0.56 (s, 3H, 18-CH ); HRMS m/z
3
ꢀ
-1
+
(50% EtOAc/hexanes), mp = 194-197 C; IR n/cm = 3430
calcd. for C H ClO [M + H] 365.1883, found 365.1813
2
1
30
3
1
(
(
(
(
17-OH, 20-OH), 2940 (CH), 1656 (C = C-C = O); H NMR
(
displaying typical isotopic chlorine pattern shown in the
360 MHz, DMSO-d ): d = 5.62 (s, 1H, H-C = C), 4.93
d, J = 7.7 Hz, 1H, 20-OH), 3.86 (s, 1H, 17-OH), 3.69
m, 2H, 21-CH Cl), 3.51 (m, 1H, 20-H), 2.37 (dt, J =14.9,
6
Supporting Information).
2
1
0
7a-Hydroxyprogesterone (6). Intermediate
2
(182 mg,
.5 mmol) was dissolved in 30% NH3 in MeOH (20 ml).
A solution of 45% Ti (SO ) in dilute sulfuric acid
3.4 Hz, 2H: overlap of 6b-Hax and 2b-H ), 2.24 (dt, J = 13.3,
ax
2.2 Hz, 1H, 6a-H ), 2.15 (dt, J = 13.3, 2.2 Hz, 1H, 2a-H ), 1.95
eq
eq
(dq, J = 13.3, 2.3 Hz, 1H, 1b-H ), 1.66-1.82 (m, 3H, 7b-H ,
2
4 3
eq
eq
(
4 mmol, 2.35 ml) was added at once. The reaction
12b-Heq and 14-H), 1.42-1.62 (m, 6H, overlap of 16b-H,
16a-H, 15a-H, 1a-H , 8-H, and 11a-H ), 1.20-1.40 (m, 2H, over-
mixture was stirred for 10 h, resulting in the formation
ax
eq
of a suspension of white powder (TiO ) in a dark solution.
The mixture was concentrated, extracted with EtOAc
lap of 11b-Hax, and 12a-H ), 1.15 (m, 1H, 15b-H overlap with s,
2
ax
3H, 19-CH ), 0.99 (m, 1H, 7a-H ), 0.88 (m, 1H, 9-H), 0.78
3
ax
+
(
20 ml) three times; the organic layers were combined,
(s, 3H, 18-CH ); HRMS m/z calcd. for C H ClO [M + H]
3 21 32 3
dried with Na SO , and concentrated under reduced
pressure. Silica gel column chromatography afforded a
367.2040, found 367.2085 (displaying typical isotopic chlorine
pattern shown in the Supporting Information).
2
4
major fraction of methylketone 6 (137mg, 0.414 mmol) in 83%
ꢀ
1
yield, R = 0.55 (50% EtOAc in hexanes), mp = 280-285 C; H
17-Hydroxy-20,21-epoxy-4-pregnene-3-one [(20S)-4]. Interme-
diate (20S)-3 (125 mg, 0.345 mmol) was dissolved in MeOH
(15 ml). A solution of NaOH (40 mg, 0.68 mmol) dissolved in
5 ml of water was added to the solution, and the mixture
was stirred at room temperature for 2 h. The crude was then
extracted with EtOAc and the organic layers were combined
and washed with brine and dried over Na SO . It was then
f
NMR (360 MHz, DMSO-d ): d = 5.63 (s, 1H, H-C= C), 5.25
6
(
(
2
1
1
s, 1H, 17-OH), 2.53 (dt, J = 13.2, 3.1 Hz, 1H, 16b-H), 2.39
dt, J =14.9, 3.4 Hz, 2H: overlap of 6b-Hax and 2b-Hax),
.24 (dt, J = 13.3, 2.2 Hz, 1H, 6a-H ), 2.15 (dt, J = 13.3, 2.2 Hz,
eq
H, 2a-H ), 2.08 (s, 3H, 21-CH ), 1.96 (dq, J = 13.3, 2.3 Hz, 1H,
eq
3
b-H ), 1.79 (m, 1H, 7b-H ), 1.66-1.75 (m, 2H, overlap
eq
eq
2
4
of 12b-Heq and 14-H), 1.51-1.66 (m, 4H, overlap of 15a-H,
concentrated under reduced pressure to afford a nearly
1
1
a-H , 8-H, and 11a-H ), 1.30-1.50 (m, 3H, overlap of
quantitative yield of epoxide (20S)-4 (110 mg, 96%). R = 0.47
ax
eq
f
ꢀ
1
6a-H, 11b-Hax, and 12a-H ), 1.23 (m, 1H, 15b-H), 1.12
(50% EtOAc/hexanes), mp = 235-239 C; H NMR (400 MHz,
ax
(
s, 3H, 19-CH ), 0.99 (dq, J = 11.8, 2.9 Hz, 1H, 7a-H ),
DMSO-d ) d = 5.63 (s, 1H, H-C = C), 4.01 (s, 1H, 17-OH), 3.01
3
ax
6
0
.88 (dt, J = 11.9, 3.1 Hz, 9-H), 0.53 (s, 3H, 18-CH ).
(dd, J = 4.0, 2.8 Hz, 1H, 20-H), 2.67 (dd, J = 5.2, 2.8 Hz,
3
Chirality DOI 10.1002/chir

STEREOSELECTIVE PREPARATION OF A FISH PHEROMONE
577
1
=
H, 21-H ), 2.58 (dd, J = 5.2, 4.0 Hz, 1H, 21-H ), 2.40 (dt, J
(m, 1H, 20-H), 4.12 (br s, 1H, 17-OH) overlap with 4.11 (s, 2H,
(C = O)CH NBoc), 3.88 (m, 2H, -NBocCH CH CH NHBoc),
S
R
14.9, 3.4 Hz, 2H: overlap of 6b-Hax and 2b-H ), 2.24
ax
2
2
2
2
(
2
dt, J = 13.3, 2.2 Hz, 1H, 6a-H ), 2.15 (dt, J = 13.3, 2.2 Hz, 1H,
2.90 (m, 2H, CH CH NHBoc), 2.39 (m, 2H: overlap of 6b-H
and 2b-H ), 2.24 (m, 1H, 6a-H ), 2.15 (m, 1H, 2a-H ), 1.97
ax eq eq
eq
2 2 ax
a-H ), 1.96 (dq, J = 13.3, 2.3 Hz, 1H, 1b-H ), 1.75-1.87
eq
eq
(
m, 2H, overlap of 7b-Heq and 14-H), 1.49-1.68 (m, 6H, overlap
(m, 2H, CH CH CH ) overlap with 1.96 (m, 1H, 1b-H ),
2 2 2 eq
of 16a-H, 16b-H, 1a-H , 12b-H , 8-H, and 11a-H ), 1.30-1.46
1.45-1.81 (m, 8H, 7b-H ,12b-H 14-H, 16b-H, 15a-H, 1a-H ,
ax
eq
eq
eq
eq,
ax
(
m, 3H, overlap of 11b-Hax, 12a-H , and 15a-H), 1.22 (m, 1H,
8-H, and 11a-H ), 1.37 (s, 9H, NHBoc), 1.34 (s, 9H, NBoc),
ax
eq
1
5b-H), 1.16 (s, 3H, 19-CH ), 0.99 (m, 1H, 7a-H ), 0.85
1.30-1.45 (m, 3H, overlap of 16a-H, 11b-Hax, and 12a-Hax),
1.22 (m, 1H, 15b-H) overlap with 1.21 (br s, 3H, 21-CH3),
1.13 (s, 3H, 19-CH ), 0.96 (m, 1H, 7a-H ), 0.84 (m, 1H, 9-H),
3
ax
(
m, 1H, 9-H), 0.81 (s, 3H, 18-CH ); HRMS m/z calcd. for
3
+
C H O [M + H] 331.2273, found 331.2292.
2
1
31
3
3
ax
0
.67 (s, 3H, 18-CH ); HRMS m/z calcd. for C H N O
3 31 51 2 6
+
1
7a,20a-Dihyroxy-4-pregnen-3-one [(20S)-1]. Intermediate (20S)-3
[M-Boc + H] 547.3747, found 547.3779.
(
(
323 mg, 0.88 mmol) was dissolved in 30% NH in MeOH
20 ml). A solution of 45% Ti (SO ) (1.76 mmol, 1.0 ml) was
3
Host-guest complex between 9 and the Zn tetraphenylporphyrin
tweezer
2
4 3
2
0,21
.
The purified 8 (1.0 mg) was dissolved in
added at once. The reaction mixture was stirred for 10 h,
resulting in the formation of a suspension of white powder
0
.5 ml CH Cl , and 0.1 ml TFA was added to the solution to
2
2
form the TFA salt of 9. The mixture was stirred under argon
for 2 h. The solvent was evaporated and the residue was
further dried on the vacuum line. The residue was suspended
in MeOH and washed with Na CO solution to yield 9. The
MeOH was then evaporated and the compound was
redissolved in anhydrous CH Cl (0.3 ml). A 10-mL aliquot of
(
TiO ) in a dark solution. The mixture was concentrated,
2
extracted with EtOAc (20 ml) three times; the organic layers
were combined, dried with Na SO , and concentrated under
reduced pressure. Silica gel column chromatography
afforded (163 mg, 0.49 mmol, 56%) in 47% overall yield from
cortexolone. Alternatively, intermediate (20S)-4 (100 mg,
2
4
2
3
2
2
the solution was added to the solution of Zn porphyrin tweezer
commercially available) in CH Cl (1mM). The CD spectra
0
.303 mmol) was dissolved in 10 ml of anhydrous THF. An
(
2
2
amount of 2.0 ml of 0.5 M HAlO in anhydrous THF was added
into the solution while stirring. The reaction mixture was
20,21
were then recorded according to the published procedure.
ꢀ
heated in an oil bath at 65 C overnight, then extracted with
EtOAc. The organic layers were combined, washed with
brine twice, dried over Na SO , then concentrated under
RESULTS AND DISCUSSION
Chlorination of cortexolone was performed using mesyl
chloride and DMAP to afford chlorocortexolone 2 in 85%
yield (not shown).22 Reduction of 2 using sodium borohy-
2
4
reduced pressure. Silica gel column chromatography using
2
6
5% EtOAc/hexanes afforded (20S)-1 (70 mg, 0.21 mmol,
dride (NaBH ) generated chloroalcohol (20R)-3. Although
9%) in 55% overall yield from cortexolone. R = 0.40
4
f
ꢀ
(20R)-3 was expected to be close to the epoxide under basic
conditions, the isolation and characterization of that interme-
diate preceded further basic treatment. Chloroalcohol (20R)-
(
50% EtOAc/hexanes), mp = 203-206 C. Mixed melting
points of 1:1 (wt%) of (20R)-1 and (20S)-1 was measured
as 188–195 C; and the melting point of a 1:1 (wt%) mixture
of a sample of the commercially available pheromone
ꢀ
2
3
3
was isolated in ca. 98% yield. Only the (20R)-3 (i.e., 20b)
ꢀ
1
was obtained and there was no evidence for a mixture of
diastereomers by either thin-layer chromatography (TLC) or
NMR. The reduction is stereocontrolled by the presence of
the 17-a-hydroxy and the C18-methyl groups according to
the discussion below in conjunction with computational
molecular modeling.
(
20R)-1 with (20S)-1 was measured as 190–195 C;
H
NMR (400 MHz, CDCl ) d = 5.73 (s, 1H, H-C = C), 3.86
3
(
2
quin, J = 6.3 Hz, 1H, 20-H), 3.49 (d, J = 5.53 Hz, 1H, 20-OH),
.31-2.48 (m, 3H, overlap of 2b-H , 2a-H , 6b-H ), 2.24-2.31
ax
eq
ax
(
1
m, 1H, 6a-H ), 1.98-2.10 (m, 2H, 16b-H and 1b-H ),
.82-1.90 (m, 1H, 7b-H ), 1.81 (s, 1H, 17-OH), 1.66-1.81
eq
eq eq
Treatment of (20R)-3 with KOH resulted in the formation
of epoxide (20R)-4 in about 96% yield. Reduction of the
epoxide with NaBH was unsuccessful under the mild conditions
needed to avoid excessive reduction of the ketone or alkene
function of ring A. An alternative approach using Ti3+ proved
successful as a more direct pathway to obtaining target phero-
(
1
1
m, 5H, overlap of 16a-H, 15a-H, 1a-H , 14-H, and 12b-H ),
.51-1.66 (m, 3H, overlap of, 11a-H , 8-H, and 12a-H ),
eq ax
.35-1.48 (m, 1H, 11b-H ), 1.23 (m, 1H, 15b-H), 1.21
ax eq
4
ax
(
(
d, J = 6.5 Hz, 3H, 21-CH ), 1.19 (s, 3H, 19-CH ), 1.10
m, 1H, 7a-H ), 0.98 (m, 1H, 9-H), 0.78 (s, 3H, 18-CH ); HRMS
3 3
ax
3
+
m/z calcd. for C H O [M + H] 333.2430, found 333.2916.
21 33 3
24
mone (20R)-1. Reductive dechlorination of chloroketone 2
with Ti3+ generated hydroxyprogesterone 6, which was
General procedure for conjugate ester 8. Conjugation of bis-Boc
18
carrier acid2
0,21
7 with the 2 chiral alcohol of the pheromones
ꢀ
followed by NaBH reduction leading to (20R)-1 in an overall
4
7
6% yield from 2 (Scheme 3). The latter reduction is both time-
(
commercially available pheromone, (20R)-1, and (20S)-1
sensitive and dependent on the equivalency of NaBH . As
respectively): a solution of (20R)-1 (5.3 mg, 0.16 mmol, 1 eq),
4
bis-Boc carrier 7 (20.0 mg, 0.60 mmol, 4 eq), EDC (32.1 mg,
0
.167 mmol, 10 eq) and DMAP (2.4mg, 0.019 mmol, 1.2 eq) in
anhydrous CH Cl (8ml) was stirred in a 50-ml round-bottom
2
2
flask overnight under nitrogen. The mixture was diluted with
CH Cl (10 ml), washed with a saturated NaHCO solution,
2
2
3
and then with brine. The solution was dried with Na SO and
2
4
then the solvent was rotavaped. The residue was quickly
purified by pipette column chromatography (CH Cl /MeOH,
2
2
4
0:2, R = 0.42) to yield the bis-Boc derivative of conjugate ester
f
1
(
20R)-8 as an oily film (4.0 mg, 39% yield); H NMR (400 MHz,
Scheme 3. Synthesis of the (20R)-epimer (the pheromone). These two
steps were combined in a one-pot transformation.
DMSO-d ) d = 6.75 (br s, 1H, N-H), d = 5.62 (s, 1H, H-C = C), 4.92
6
Chirality DOI 10.1002/chir

5
78
OUEDRAOGO ET AL.
previously reported,18 we also observed the reduction of both
the C3 enone and the C20 ketone with increased reaction times
determination of configuration with microscale quantities. Li
6,27
et al.2
elegantly described how the tweezer methodology
(
>30 min) and NaBH equivalencies (>1.1 eq). Careful control
could be applied to erythro and threo diols; however, none of
the substrates studied contained a stereogenic tertiary alcohol.
Pheromone (20R)-1 was coupled to a bidentate carrier 7
(Scheme 5) to afford conjugate (20R)-8. The derivatization
was carried out on the secondary alcohol with no interference
from the vicinal tertiary alcohol, as previously observed for the
4
of reaction conditions favored the C20 over the C3 reduction as
discussed below with the aid of computational calculations. We
were also able to combine the dechlorination and reduction
steps in one pot with an overall yield of 64% for the conversion
of cortexolone to (20R)-1 in just two steps. The target structure
showed identical spectra to those of the commercially available
sample of the pheromone.
20
ginkgolide derivative reported by Kurtan et al. Deprotection
of (20R)-8 with trifluoroacetic acid (TFA) resulted in (20R)-9
which was complexed with a bis-chromophoric zinc porphyrin
We aimed to accomplish an equaly stereoselective prepara-
tion of the pheromone C20 epimer, and provide a ratio-
nale for the stereoselectivity. Although it was evident
that aluminumoxyhydride (HAlO) was capable of
exhibiting selectivity towards reducing ketones, the pres-
ence of the 17-OH guided such reduction to the C20 over
the C3 ketone which was reduced preferentially in the case of
20,21
tweezer (Fig. 1).
The well-established chiroptical method
takes advantage of the presence of a large moiety (L) and a
medium moiety (M) on the secondary alcohol carbon, in which
the small hydrogen (H) will align syn to the ester carbonyl.
25
O
progesterone. The HAlO reduction of chlorocortexolone,
shown in Scheme 4, generated the diastereomer of the
M
H3C
N
NH2
H
O
chloroalcohol that was obtained by NaBH reduction. Further
H
OH
L
O
4
M
L H
3
+
reductive dechlorination was achieved using Ti on (20S)-3
to afford the C20 epimer (i.e., 20a) of the pheromone, (20S)-1
H
H
N
NH2
H
H
O
(
three steps from cortexolone, overall 47% yield). Alternatively,
O
basic treatment of (20S)-3 led to epoxide (20S)-4 which upon
selective reduction with HAlO afforded (20S)-1 in an overall
N
N
(
20R)-9
N
N
Zn
Zn
5
5% yield (over four steps) from cortexolone.
N
N
N
N
We proceeded to characterize the stereochemistry of the
secondary alcohol of both C20 epimers using a previously
established microscale chiroptical method developed by
2
0,21
Nakanishi, Berova, and colleagues.
The advantages of
this chiroptical method, over traditional derivatization
methods combined with 2D NMR analysis, is the unambiguous
O
O
O
O
Zn Porphyrin Tweezer
+
-
L
M
L
>
>>
M
favored
conformation
less favored
conformation
Fig. 1. The diamino conjugate of (20R)-9 was complexed with Zn porphy-
rin tweezer leading to two populations of conformers, a favored one and a less
favored one. The predominantly favored conformer(s) lead to a negative exci-
ton couplet, as expected. The purple rectangular shapes, highlighting the
twist of the complex, represent the transition moments of the two porphyrins
in the tweezer; L and M represent the large and medium groups, respectively.
The (20S)-9 was also complexed leading to the opposite twist for its predomi-
nant conformer, resulting in an opposite couplet (not shown in this figure).
Scheme 4. Synthesis of the (20S)-epimer of the pheromone, (20S)-1.
Scheme 5. Coupling the secondary alcohol to a bidentate carrier 7. This was carried out for both authentic and synthesized pheromone (20R)-1 as well as the
20S)-1, forming (20R)-9 and (20S)-9 (not shown), respectively.
(
Chirality DOI 10.1002/chir

STEREOSELECTIVE PREPARATION OF A FISH PHEROMONE
579
In this case, M is the methyl group and the L group is an
unambiguously much larger group; namely, the steroid moiety.
Therefore, it is expected that there will be a clear overpopulation
of the favored conformation of the diamino conjugate with
the Zn porphyrin tweezer, in which the L group will prefer-
entially reside away from the crevice of the complex. This
induces a negative twist for the transition moments of the
Zn porphyrin twist, which results in a negative exciton
couplet. The resultant complex was subjected to CD
studies in methylcyclohexane (Fig. 2). Similarily, the C20
epimer, (20S)-1, was subjected to conjugation with 7
(
Scheme 5), followed by deprotection and complexation
with the tweezer prior to CD studies. As predicted on the basis
of the steric size of the substituents flanking the stereogenic
20,21,28–30
center,
CD spectrum of the tweezer complex formed from (20R)-1
compound (20R)-9) had a negative exciton couplet, reflective
as described above, the exciton coupled
(
of the negative twist of the transition moments of the porphyrin
tweezer and indicative of an (R) absolute configuration, whereas
the complex formed from (20S)-1(compound (20S)-9) had a pos-
itive couplet, predicting an (S) absolute configuration.
Molecular modeling Density Functional Theory (DFT)
calculations at the (R)B3LYP/6-31G(d) level were computed
to elucidate the pathway of the reduction of the carbonyl
at C20 to yield either the pheromone, (20R)-1, or its
epimer (20S)-1. The ground state equilibrium geometries,
i.e., lowest energy conformers, of all synthesized molecules
Fig. 2. (a) CD spectra of the complexes of (20R)-9 (dashed) and of (20S)-9
solid) with Zn porphyrin tweezer. (b) Respective UV spectra of these
(
complexes: (20R)-9 (dashed) and of (20S)-9 (solid).
-
and suggested stable intermediates ([2-HAlO], [6] and
[
6-BH ]) were optimized in the gas phase at 298 K. All
3
ground states were true minima with no imaginary vibra-
tional frequencies (see Electronic Supporting Information
for energy differences between conformers of the opti-
mized structures).3
1–35
The equilibrium geometry of methylketone 6 has an H-bond
between the OH at C17 and the oxygen of the carbonyl at C20.
This exposes the re face of the carbonyl toward nucleophilic
attack, as the opposite si face is hindered by the C18-methyl
group (Fig. 3).36
Fig. 3. Hydride attack on LUMO + 1 is hindered from the si face due to the
C18 methyl (highlighted in yellow for clarity).
-
Fig. 4. (a) NaBH
4
and (b) HAlO proposed mechanisms with optimized structures. Structures [6] , [2-HAlO] and [6-BH
3
] are the only suggested stable inter-
mediates that were not isolated and characterized.
Chirality DOI 10.1002/chir

5
80
OUEDRAOGO ET AL.
5
. Sorensen PW, Stacey NE. Brief review of fish pheromones and discussion
of their possible uses in the control of non-indigenous teleost fishes. N Z J
Mar Freshwater Res 2004;38:399–417.
In the case of the reduction of 6 with NaBH in MeOH
4
the (20R)-epimer is formed exclusively; i.e., the (20S) is
not formed. The sodium cation is not as oxophilic as
aluminum in HAlO and will therefore not chelate the two
oxygens at C17 and C20 (as shown in HAlO reduction,
below). Thus, the methoxide ion of the basic methanolic
solution abstracts the proton from the 17-OH, resulting in
a near 180-degree rotation of the C17-C20 bond to minimize
6
. Appelt C, Sorensen PW. Female goldfish signal spawning readiness by
altering when and where they release a urinary pheromone. Anim Behav
2007;74:1329–1338.
7. Fine JM, Sorensen PW. Isolation and biological activity of the multi-component
sea lamprey migratory pheromone. J Chem Ecol 2008;34:1259–1267.
8. Stacey NE, Sorensen PW. Hormonally derived sex pheromones in fish.
Fish Reprod 2008; Chap. 6:201–244.
-
repulsion between the two oxygen atoms (6 ! [6] ). This
9
. Scott AP, Sumpter JP, Stacey N. The role of the maturation-inducing
steroid, 17,20beta-dihydroxypregn-4-en-3-one, in male fishes: a review. J
Fish Biol 2010;76:183–224.
exposes the si face of the carbonyl, contrary to the case
-
presented in Fig. 3. Intermediate [6] is then reduced from
the accessible si face of the carbonyl, leading to the formation
of the (20R)-epimer, i.e., pheromone (20R)-1 as shown in
Fig. 4a. A similar stereochemical outcome would result in
1
0. Milla S, Jalabert B, Rime H, Prunet P, Bobe J. Hydration of rainbow trout
oocyte during meiotic maturation and in vitro regulation by 17,20beta-
dihydroxy-4-pregnen-3-one and cortisol. J Exp Biol 2006;209:1147–1156.
a site delivery of the hydride from a [6-BH ] complex at
11. Inbaraj RM, Haider S, Baqri SSR. Dynamics of 17alpha,20beta-dihydroxy-
-pregnen-3-one and 17alpha,20beta,21-trihydroxy-4-pregnen-3-one in
3
4
the C17 alkoxy. Previous work by Evans and co-workers
also report similar high stereoselectivities in related
plasma and oocyte incubation media of catfish (Clarias batrachus) in
response to salmon gonadotropin. Curr Sci 2001;80:455–458.
3
7,38
b-hydroxyketone reductions using borohydrides.
1
2. King HR, Young G. Milt production by non-spermiating male Atlantic
salmon (Salmo salar) after injection of a commercial gonadotropin
releasing hormone analog preparation, 17alpha-hydroxyprogesterone
or 17alpha,20beta-dihydroxy-4-pregnen-3-one, alone or in combination.
Aquaculture 2001;193:179–195.
In the case of HAlO, the more oxophilic aluminum coordi-
nates the two oxygen atoms on C17 and C20 while the
hydride attacks the carbonyl from the nonhindered re
face leading to an exclusive stereoselection for the
3
9
13. Todo T, Ikeuchi T, Kobayashi T, Kajiura-Kobayashi H, Suzuki K,
Yoshikuni M, Yamauchi K, Nagahama Y. Characterization of a testicular
(
20S)-epimer (Fig. 4b).
1
7alpha,20beta-dihydroxy-4-pregnen-3-one (a spermiation-inducing steroid
CONCLUSION
in fish) receptor from a teleost, Japanese eel (Anguilla japonica). FEBS
Lett 2000;465:12–17.
4. Barry TP, Riebe JD, Parrish JJ, Malison JA. Effects of 17alpha,20beta-
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We developed a two-step procedure for the construction
1
1
1
of a goldfish pheromone in an overall 64% yield with exclu-
sive enantioselectivity from a rather inexpensive starting
material. A second procedure, using a fairly newly devel-
oped aluminum reducing agent (HAlO), was designed for
a three-step synthesis of the C20 epimer of the pheromone
in an overall 47% yield. Future studies with collaborators
will be focused on elucidating the difference in the biological
activities of the two C20 epimers, as well as novel constructs,
based on the general structure 5, resulting from the opening
of the intermediate epoxide 4. Moreover, transition state
computations should provide more insight to the proposed
reaction mechanisms.
1
1
7. Sorensen PW, Hara TJ, Stacey NE. Sex pheromones selectively stimulate
the medial olfactory tracts of male goldfish. Brain Res 1991;558:343–347.
8. Kovganko NV, Kashkan ZN, Shkumatov VM. Simple synthesis of
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19. Nagahama Y. 17alpha,20beta-Dihydroxy-4-pregnen-3-one:
a
teleost
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ACKNOWLEDGMENTS
20. Kurtan T, Nesnas N, Li Y-Q, Huang X, Nakanishi K, Berova N. Chiral recog-
nition by CD-sensitive dimeric zinc porphyrin host. 1. Chiroptical protocol
for absolute configurational assignments of monoalcohols and primary
monoamines. J Am Chem Soc 2001;123:5962–5973.
21. Kurtan T, Nesnas N, Koehn FE, Li Y-Q, Nakanishi K, Berova N. Chiral
recognition by CD-sensitive dimeric zinc porphyrin host. 2. Structural
studies of host-guest complexes with chiral alcohol and monoamine conju-
gates. J Am Chem Soc 2001;123:5974–5982.
The authors thank the Materials, Science, and Nanotechnology
Institute at Florida Institute of Technology for purchasing the
AccuTOF-DART. G. Proni thanks the John Jay College
Research Assistance Fund and PRISM (Program for Research
Initiatives for Science Majors) for financial support. E. Chadwick
thanks PRISM for financial support. We also thank K. Nakanishi
and N. Berova for enabling the CD studies and providing insight,
J. C. Baum and R. Terryn for advice on computational studies,
and P. J. Cohen for help with mass spectrometry.
2
2. Draper RW, Hu B, McPhail AT, Puar MS, Vater EJ, Weber L. Unusual
hydroxy-gamma-sultone byproducts of steroid 21-methanesulfonylation.
An efficient synthesis of mometasone 17-furoate (Sch 32088). Tetrahe-
dron 1999;55:3355–3364.
2
3. Although using (20R) and (20S) is less ambiguous to denote the stereo-
chemistry at acyclic stereocenters such as C20, we used the traditional
notations of 20b and 20a, respectively, to refer to the names of the
pheromone and its epimer, as in previous reports. The use of 20a/
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Chirality DOI 10.1002/chir