Enantioselective transport by a steroidal guanidinium receptor

Article abstract of DOI:10.1002/1521-3765(20020703)8:13<2931::AID-CHEM2931>3.0.CO;2-H

The cationic steroidal receptors 9 and 11 have been synthesized from cholic acid 3. Receptor 9 extracts N-acetyl-α-amino acids from aqueous media into chloroform with enantioselectivities (L:D) of 7-10:1. The lipophilic variant 11 has been employed for th

Full text of DOI:10.1002/1521-3765(20020703)8:13<2931::AID-CHEM2931>3.0.CO;2-H

FULL PAPER
Enantioselective Transport by a Steroidal Guanidinium Receptor
[
d]
[a]
[b]
[a]
Beatriz Baraga nƒ a, Adrian G. Blackburn, Perla Breccia, Anthony P. Davis,*
[
b]
[c]
[b]
[a]
Javier de Mendoza,* Jos e¬ M. Padr o¬ n-Carrillo, Pilar Prados, Jens Riedner, and
Johannes G. de Vries*
[
c]
Abstract: The cationic steroidal receptors 9 and 11 have been synthesized from
cholic acid 3. Receptor 9 extracts N-acetyl-a-amino acids from aqueous media into
chloroform with enantioselectivities (l:d) of 7 ±±1:±. The lipophilic variant 11 has
been employed for the enantioselective transport of N-acetylphenylalanine,
a) through dichloromethane (DCM) and dichloroethane (DCE) bulk liquid
membranes (U-tube apparatus), and b) through 2.5% (v/v) octanol/hexane via
hollow fibre membrane contactors. Significant enantioselectivities and multiple
turnovers were observed for both types of apparatus.
Keywords: chiral resolution ¥ enan-
tioselectivity ¥ membranes ¥ recep-
tors ¥ transport
incorporated[7] in polymeric membranes. Acting as shuttles,
the receptor molecules can transfer many equivalents of
substrate between reservoirs. Selectivities depend on the rates
of uptake, diffusion and delivery of the two enantiomers, but
often reflect the ratio of binding affinities.[
Chiral carboxylates are good subjects for membrane trans-
port because they are soluble in water as alkali metal salts but
readily extracted into non-polar media by lipophilic cations.
Introduction
The design of enantioselective receptors is one of the central
challenges of supramolecular chemistry.[±] Interest derives
partly from the importance of enantioselective recognition in
biology, but also from the need for enantiomerically pure
compounds in the chemical industry.[ Indeed, to quote from a
recent monograph, ™The separation of enantiomers may well
represent the economically most important application of
supramolecular chemistry∫.[ However, to fulfil this potential
it is necessary to develop practical receptor-mediated separa-
tion procedures. A key issue is that of receptor recycling. A
sophisticated receptor is only viable if it can be used many
times over, ideally in a continuous process. Membrane
5a]
2]
Enantioselective extractions have been accomplished with
3]
[5b, 8]
chiral cations,
and also with achiral cations accompanied
by electroneutral chiral receptors.[ Guanidinium cations are
especially suitable, as they are able to form well-defined salt
bridges with carboxylates through formation of two parallel
hydrogen bonds.[
9]
±1]
[4]
transport fulfils this criterion. Receptors may be located in
Steroidal guanidinium cations of general form 1 represent a
promising new class of enantioselective carboxylate receptors.
a bulk liquid membrane,[ or dissolved or covalently
5]
[6]
[
a] Prof. A. P. Davis, A. G. Blackburn, J. Riedner
School of Chemistry, University of Bristol
Cantock×s Close, Bristol BS8 ±TS (UK)
Fax : (‡44)±±7-92986±±
E-mail: anthony.davis@bristol.ac.uk
[
b] Prof. J. de Mendoza, Dr. P. Breccia, Prof. P. Prados
Departamento de QuÌmica Org a¬ nica
Universidad Aut o¬ noma de Madrid
Cantoblanco, 28149 Madrid (Spain)
Fax : (‡34)9±-3973966
E-mail: javier.demendoza@uam.es
[
c] Prof. J. G. de Vries, Dr. J. M. Padr o¬ n-Carrillo
DSM Fine Chemicals-Advanced Synthesis & Catalysis
P.O. Box ±8, 6±61 MD Geleen (The Netherlands)
Fax : (‡3±)46-4767614
Accessible from inexpensive cholic acid 3, they create a
tunable chiral environment for bound carboxylates 2. Early
examples such as 4 and 5 have proved effective, performing
extractions with enantioselectivities of 7 ±±1:± on a range of
amino acid derivatives 6.[ Herein we report that they also
E-mail: hans-jg.vries-de@dsm.com
[
d] Dr. B. Baraga nƒ a
Department of Chemistry, Trinity College
Dublin 2 (Ireland)
±±]
Chem. Eur. J. 2002, 8, No. ±3
¹ WILEY-VCH Verlag GmbH, 6945± Weinheim, Germany, 2112
1947-6539/12/18±3-293± $ 21.11+.51/1
2931

A. P. Davis, J. de Mendoza, J. G. de Vries et al.
FULL PAPER
design evolved from 5, the most successful of our previous
systems.[
±±b]
Two changes were made. Firstly, for synthetic
reasons, the six-membered ring guanidinium moiety in 5 was
replaced by a five-membered unit, as in 9. As shown in
[
±±b]
Scheme ±, 9 could be prepared in two steps from azide 7
by
employing the guanylation reagent 8.[ In contrast the six-
membered unit in 5 had required a low-yielding multistep
procedure, as conditions for direct introduction could not be
found.[ Amino acid extraction studies using 9 confirmed that
the change in ring size did not greatly affect the enantiose-
lectivity (Table ±).
±2]
±3]
Table ±. Enantioselective extraction of N-acetyl-a-amino acids from
by receptor 9.[
a]
aqueous buffer into CHCl
3
Substrate
Extraction
efficiency [%][
Enantio-
selectivity (l:d)
b]
[c]
N-Ac-dl-alanine
N-Ac-dl-valine
N-Ac-dl-phenylalanine
77
8±
87
±1:±
7:±
±1:±
[
a] For experimental details, see ref. [±±b]. [b] Concentration of substrate in
organic phase, as percentage of receptor concentration, determined by
NMR integration. [c] Determined by NMR integration of signals for
complexed substrates in extracts.
succeed as enantioselective membrane transport agents, both
in a U-tube apparatus (bulk DCE/DCM membrane) and also
in a system based on hollow fibre membranes. The latter is of
particular interest in view of a possible industrial-scale
separation method.
Secondly, extraction experiments involving methyl esters
such as 4, 5 or 9 were generally found to result in the loss of
significant amounts of receptor from the organic phase.
Moreover, 9 was found to be insoluble in the alkane/octanol
mixtures used with the hollow fibre membrane system (vide
infra). The methyl ester was therefore changed to an eicosyl
ester, solving both problems. As shown in Scheme ±, azido
ester 7 was treated with aqueous sodium hydroxide, followed
by caesium carbonate and eicosyl bromide, to give eicosyl
Results and Discussion
Receptor design and synthesis: The transport experiments
described herein required a variant on structure 1 which was
a) enantioselective, b) available in gram quantities, and
c) highly lipophilic, and soluble in non-polar solvents. The
Scheme ±. Synthesis of receptors 9 and 11: a) ±) Zn, AcOH, 2) 8, NaHCO
) HCl aq., 3) C21 4±Br, Cs CO , NaI (cat.), DMF.
3 2 2 2
, DMF, 3) CH Cl , ±m NaOH aq. then 1.5m HCl aq.; b) ±) NaOH, MeOH, H O,
2
H
2
3
2932
¹ WILEY-VCH Verlag GmbH, 6945± Weinheim, Germany, 2112
1947-6539/12/18±3-2932 $ 21.11+.51/1
Chem. Eur. J. 2002, 8, No. ±3

Enantioselective Transport by a Steroidal Guanidinium Receptor
293±±2936
ester 10. Reduction of the azide followed by treatment with 8
gave receptor 11, which was used in the transport experiments
described below.
Transport by receptor 11 through dichloromethane (DCE)/
dichloroethane (DCM) membranes (U-tube apparatus):
Transport through chlorinated solvent membranes was inves-
tigated in the U-tube apparatus depicted in Figure ±. Based on
previous extraction experiments, N-Ac-l /d -PheOH were
selected as substrates. Initial experiments were performed on
5.61 cm
1.45 cm
1
.65 cm2
S
R
Volume 3 cm3
7.09 cm
water at T = 25°C
M
Figure 2. Active transport of N-Ac-PheOH single enantiomers with
carrier 11 in U-tube apparatus (Figure ±). Triangles represent receiving
phases and circles source phases
Figure ±. Transport cell, with dimensions.
the individual enantiomers, exploring the effect of various
parameters that could affect transport rate. The source phase
consisted of N-Ac-l-PheOH or N-Ac-d-PheOH dissolved in
TRIS buffer (pH ˆ 8.±). The organic (membrane) phase
consisted of a solution of 11 in ±,2-dichloroethane, and the
receiving phase was aqueous KBr. Bromide was found to be
more effective than chloride for counter-current exchange at
the membrane-receiving phase boundary. Concentrations of
KBr were higher than those of substrate, to ensure efficient
active transport. Transport rates varied linearly with [11] at
low to moderate receptor concentrations, but reached a
Table 2. Enantioselective transport of N-Ac-PheOH in a U-tube using 11
as carrier.
Time [h]
N-Ac-Phe [%]
ee [%]
2
3
4
5
6
7
8
9
24
3
5
6
7.5
8
±2
±1
±2
27
66
7±
71
64
68
64
64
61
56
À2
plateau at concentrations of [11] of about 3.4 Â ±1 m. A
temperature increase from 258C to 418C caused no significant
changes in the transport rate. The relative transport rates
measured for the two enantiomers were consistent with the
extraction experiments, in that the l enantiomer was trans-
ported at least four times faster than the d enantiomer
(
Figure 2).
Carrier 11 was also tested for transport of racemic N-
acetylphenylalanine in the U-tube apparatus. Conditions were
similar to those above described for the single enantiomers,
except that dichloromethane was employed as the bulk
membrane phase instead of ±,2-dichloroethane. The results
are summarized in Table 2. Notable selectivity was shown for
N-Ac-l-PheOH. A consistent value of nearly 71% ee was
found at the initial stages of the experiment, the figure only
decreasing significantly after 24 h, when the source phase
become increasingly concentrated in the other enantiomer
Figure 3. Enantioselective transport of N-Ac-dl-PheOH in U-tube appa-
ratus; ee in receiving phase is plotted against total transported.
Enantioselective transport in hollow fibre membrane con-
tactors: Although useful in the laboratory, transport in
U-tubes does not scale up effectively due to the relative
decrease in contact surface area. To investigate its behaviour
in an industrially relevant setting, receptor 11 was also tested
in the separator depicted in Figure 4 (see also Table 3). The
(
Figure 3). Over the course of the experiment, about 21
equivalents of N-acetylphenylalanine was transported by
receptor 11.
Chem. Eur. J. 2002, 8, No. ±3
¹ WILEY-VCH Verlag GmbH, 6945± Weinheim, Germany, 2112
1947-6539/12/18±3-2933 $ 21.11+.51/1
2933

A. P. Davis, J. de Mendoza, J. G. de Vries et al.
FULL PAPER
was circulated an organic phase consisting of octanol/hexane
2
.5% (v/v) (±25 mL), 4 mm in host 11. The concentrations
of the enantiomers in each aqueous phase and the ee values
were monitored over 48 h (Figure 5). After this time, the
amino acid transported to the receiving phase corresponded
to 61% of the initial amount. Initially an ee of 3±% was
observed in the receiving phase, although this was eroded
over time as the system moved towards equilibrium (which
must, in principle, involve racemates in both source and
receiving phases). Up to 71 equivalents of substrate were
transported by the receptor.
Figure 4. Experimental set-up for simultaneous extraction and stripping of
N-Ac-dl-PheOH using two hollow fibre contactors.
Table 3. Characteristics of the hollow fibre modules and membrane
Microdyn LD OC 12.
Module and membrane data
diameter
2 cm
length
35 cm
number of fibres
internal diameter, dry
wall thickness, dry
active membrane area, inside
active membrane length
free flow area
±281
211 mm
9.5 mm
2
1.2 cm
24.5 cm
1.4 cm
±± mL
25 mL
2
intracapillary volume
extracapillary volume
key elements of this apparatus are contactor modules
incorporating hollow cellulose fibres. A solution of chiral
selector in a non-polar, water-immiscible solvent is pumped
through the interiors of the fibres (lumen flow), while the
aqueous source and receiving phases are in contact with the
exteriors. Due to the high surface area, efficient transfer
occurs between phases. A similar apparatus was used by
Pirkle and co-workers to separate N-(3,5-dinitrobenzoyl)leu-
cine enantiomers, with lipophilic (S)-N-(±-naphthyl)leucine
derivatives as selectors.[ However, this work used an excess
of chiral host, while the present experiments employed 11 in
just catalytic amounts. Unfortunately the material of the
hollow fibre contactor limits the choice of organic solvent for
the lumen flow; only aliphatic hydrocarbons and alcohols are
compatible. 2.5% (v/v) ±-octanol in hexane was chosen for
this work, being sufficiently polar to dissolve 11 but incapable
of supporting transport in the absence of receptor (as shown
by control experiments).
5d]
Figure 5. Enantioselective transport of N-Ac-dl-PheOH in hollow fibre
membrane apparatus. a) Depletion of substrate enantiomers in source
phase. b) Accumulation of substrate enantiomers in the receiving phase.
c) Enantiomeric excess of N-Ac-PheOH in source and receiving phases.
The separation of N-Ac-dl-PheOH in the membrane unit
was performed as follows: A source phase consisting of 5 g of
racemic N-Ac-PheOH in 511 mL of TRIS buffer was circu-
lated through the shell of one module, while a receiving phase
of 1.±m KBr in 511 mL of TRIS buffer was circulated through
the shell of the other. Through the lumen of both modules
Transport rates and enantioselectivities were found to be
very sensitive towards changes in the source and receiving
phases. With phosphate buffer instead of TRIS, the rate of
transport was very similar but no enantioselectivity was
2934
¹ WILEY-VCH Verlag GmbH, 6945± Weinheim, Germany, 2112
1947-6539/12/18±3-2934 $ 21.11+.51/1
Chem. Eur. J. 2002, 8, No. ±3

Enantioselective Transport by a Steroidal Guanidinium Receptor
293±±2936
observed. The use of unbuffered 1.± m KBr in the receiving
phase gave good enantioselectivity (33%), but transport
leveled off at ±2%. A higher concentration of KBr (±m) in the
receiving phase lowered transport rates by a factor of 3,
despite the increased thermodynamic driving force. Presum-
ably this results from lowered concentrations of the carbox-
ylate in the organic phase, due to competition by bromide for
cationic binding sites. To confirm that the enantioselective
transport was due to the chiral host and not to the cellulose in
the fibres, we ran an experiment using tetraoctylammonium
bromide (TOAB) as carrier. As expected, non-enantioselec-
tive transport was observed. The lowered initial selectivity
observed in hollow fibre membrane apparatus (ca. 31% ee)
compared to that for the U-tube (ca. 71% ee) was due to the
change in solvent. Transport of N-Ac-dl-PheOH by 11
through a bulk membrane of 2.5% (v/v) ±-octanol in hexane
also occurred with about 31% ee.[
from the atmosphere by a calcium chloride tube and stirred vigorously at
room temperature for 45 min. The zinc residues were removed by filtration
and washed with acetic acid (3 Â 5 mL). The filtrate was evaporated under
reduced pressure. Residual acetic acid was removed by repetitive addition
and evaporation under reduced pressure of distilled toluene (2 Â 5 mL) to
afford the crude alkylammonium acetate (99 mg, 95%). Guanylating
reagent 8[ (29 mg, 1.±9 mmol) was added to a well-stirred suspension of
±2]
the alkylammonium acetate (51 mg, 58 mmol) and NaHCO
3
(61 mg,
1
.72 mmol) in DMF (625 mL) at 818C. The mixture was vigorously stirred
for ± h then allowed to cool to room temperature and evaporated under
reduced pressure. The residue was purified by flash chromatography
eluting with EtOAc. To ensure chloride was the counter anion, the resulting
2 2
purified salt was taken up in CH Cl (5 mL) and washed carefully with
aqueous NaOH (±m, 2 Â 5 mL) followed by triply distilled water (5 mL).
The organic layer was then washed with aqueous HCl (1.5 m, 2 Â 5 mL)
followed by triply distilled water (2 Â 5 mL). The organic phase was then
dried by decanting and evaporated under reduced pressure. The residue
was then further dried in vacuo at 618C to afford the pure guanidinium
chloride 9 (36 mg, 72%) as a white solid; R
f
ˆ 1.69 (EtOAc); m.p. ±95 ±
), 1.86 (d,
), 3.14 (br m, ±H; 3b-
2
×s at positions 4 and 5 of the
±
±
988C; H NMR (411 MHz; CDCl
3
): d ˆ 1.76 (s, 3H; ±8-CH
), 1.94 (s, 3H; ±9-CH
), 3.73 (s, 4H; CH
3
±4]
3
J(H,H) ˆ 6.5 Hz, 3H; 2±-CH
3
3
H), 3.64 (s, 3H; CO
2
CH
3
imidazolinium ring), 4.89 (s, ±H; ±2b-H), 5.±9 (s, ±H; 7b-H), 5.6± (br s, ±H;
NH at position ± or 3 of the imidazolinium ring), 6.75 (br s, ±H; NH at
Conclusion
3
position ± or 3 of the imidazolinium ring), 7.49 (d, J(H,H) ˆ 7.9 Hz, 4H;
ArH meta), 7.94 (m, 4H; ArH ortho), 9.15 (s, ±H; NHCO), 9.59 (br s, ±H;
±
3
In conclusion, we have synthesized the lipophilic steroidal
guanidinum receptor 11, and demonstrated its effectiveness as
an enantioselective transport agent for an N-acetyl-a-amino-
carboxylate. In a conventional U-tube apparatus with chlori-
nated solvents as bulk liquid membranes, the enantioselectiv-
ities approached those measured in simple extraction experi-
ments. In a system based on hollow fibre membrane
contactors, somewhat lower selectivities were observed due
to an enforced change in membrane solvent. In both cases
substantial turnover was observed, confirming the ability of 11
to function as a catalytic agent for enantioselective separation.
Although selectivities dropped as transport proceeded, this
problem can be remedied in a number of ways: for example,
a) extraction of the source phase with two independent loops
containing hosts with opposite selectivities, b) continuous
racemisation of the source phase, and c) the use of several
membrane units in series, running the separation in a
3a-NH), 9.69 (s, ±H; NHCO); C NMR (±11 MHz; CDCl
CH ), ±6.65 (2±-CH ), 2±.31 (±9-CH ), 2±.84 (CH ), 25.17 (CH
or 4 of imidazolium ring), 26.±3 (C-3, 4 imidazolium ring), 26.±5 (CH
7.78 (CH ), 29.65 (CH), 29.96 (CH ), 31.22 (CH ), 32.77 (±1-C), 33.5±
CH ), 33.67, 33.92, 38.43 (CH), 39.99 (CH), 42.31 (CH), 44.25 (±3-C), 46.21
(CH), 51.47 (3-CH), 52.72 (CO CH ), 71.±7 (7-CH), 75.69 (±2-C), ±±7.±6
ArCH ortho), ±±7.45 (ArCH ortho), ±24.71 (ArCH para), ±24.91 (ArCH
para), ±24.94 (ArCH meta), ±24.98 (ArCH meta), ±25.12, ±25.15, ±4±.19,
3
): d ˆ ±±.51 (±8-
), 26.17 (C-3
),
3
3
3
2
2
2
2
(
2
2
2
2
2
3
(
±
8
4±.±5, ±52.2±, ±52.37 (C), ±57.51 (C), ±73.71 (COOCH
3
); FAB-MS m/z:
‡
‡
64 [M ‡ Na] ; HRMS (FAB ) calcd for C44
H N O F : 864.4±3481; found:
56 5 6 6
864.4±143±; elemental analysis calcd (%) for C H N O F Cl ¥ ±.5H O C
4
4
56
5
6
6
2
57.98, H 6.52, N 7.68; found: C 57.98, H 6.76, N 7.47.
Eicosyl 3a-azido-7a,12a-di[p-(trifluoromethyl)phenylaminocarbonyloxy]-
5b-cholan-24-oate (10): To a solution of compound 7 (5.11 g, 6.18 mmol) in
methanol/water (51 mL, 9:±) was added sodium hydroxide (±.18 g,
2
7 mmol). The mixture was stirred at room temperature overnight. The
solvent was removed under reduced pressure and the white solid was taken
up in dichloromethane. Aqueous HCl (±m) was added until pH 4 was
reached. The mixture was extracted with dichloromethane and the organic
layer was washed with water, dried and concentrated under reduced
pressure. The white solid was dissolved in methanol/water (51 mL, 9:±) and
caesium carbonate (1.99 g, 3.14 mmol) was added. The solvents were
removed and the solid was taken up in dry DMF (81 mL). Bromoeicosane
[
4]
countercurrent fashion. . We are currently investigating this
last option, as it holds promise for a versatile and practical
enantioseparation system for development-scale intermedi-
ates and products.
(
2.2 g, 6.18 mmol) and sodium iodide (91 mg, 1.6 mmol) were added and
the mixture was stirred under argon for 24 h at 478C. The solvent was
removed under reduced pressure and the product was isolated by flash
chromatography (hexane/ethyl acetate, 5:±) to yield eicosyl ester 10 (5.46 g,
8
2%). R
(film from CDCl
411 MHz, CDCl
(d, 3H; 2±-CH
f
ˆ 1.8± (ethyl acetate/hexane/chloroform, ±:±:±); m.p. ±658C; IR
˱
˱
±
Experimental Section
3
): nÄ max ˆ 2191 cm (N
3
), ±739 cm (C ˆ O); H NMR
), 1.91 ( t, 3H; esterCH ), 1.94
×s), 3.±8
), 4.97 (m, ±H; 7b-H), 5.±3 (m, ±H;
2b-H), 6.94 (br s, ±H; NH), 7.16 (br s, ±H; NH), 7.67 (s, 8H; ArH);
(
3
): d ˆ 1.75 (s, 3H; ±8-CH
3
3
General: ±H and ±3C NMR spectra were obtained from a Bruker MSL-311
3
), 1.97 (s, 3H;±9-CH
3 2
), ±.25 (m, 38H; side chain CH
(
m, ±H; 3b-H), 4.1± (t, 2H; CO CH
2
2
or a Bruker DPX-411 spectrometer. Chemical shifts are reported relative
to residual undeuterated solvent peaks. Melting points were determined on
a Gallenkamp melting-point apparatus. Infrared spectra were recorded on
a Perkin-Elmer 883 or Perkin-Elmer FT-IR Paragon ±111 spectrophotom-
eter. Reactions were monitored by thin-layer chromatography (TLC) on
Merck silica gel 61 F254 aluminium-backed plates. Spots due to steroidal
compounds were visualised by charring over a Bunsen burner. Flash
chromatography was carried out on Merck silica gel 61 (particle size 1.141 ±
±
±
3
C NMR (±11 MHz, CDCl
6.8, 27.±, 28.6, 29.±, 29.2, 29.4, 29.5, 29.6, 29.7, 31.7, 3±.2, 3±.5, 3±.9, 34.4, 34.5,
4.7, 35.1, 37.9, 4±.±, 43.7, 45.4, 47.5, 6±.2, 64.6, 72.4, ±±8.±, ±±8.2, ±26.4, ±4±.±,
52.4, ±52.7, ±74.2; HRMS (ES ) calcd for C61
3
): d ˆ ±2.3, ±4.±, ±7.6, 22.5, 22.7, 22.9, 25.8, 25.9,
2
3
±
±
‡
‡
H
87
N
5
O
6
F
6
Na [M ‡ Na] :
±±1.6458; found: ±±±1.6511.
Eicosyl 3a-(4,5-dihydro-1H-imidazol-3-ium-2-yl)-7a,12a-bis[(p-trifluoro-
methyl)phenylaminocarbonyloxy]-5b-cholan-24-oate chloride (11): To the
azide 10 (3.15 g, 2.8 mmol) in glacial acetic acid (±5 mL) was added zinc
dust (±.11 g). The mixture was stirred for 8 h at room temperature, after
which the mixture was filtered and evaporated to dryness to give the
1
.163 mm). Reagents were purchased from Aldrich, Fluka or Sigma and
were used without further purification.
Methyl 3a-(4,5-dihydro-1H-imidazol-3-ium-2-yl)-7a,12a-bis[(p-trifluoro-
methyl)phenylaminocarbonyloxy]-5b-cholan-24-oate chloride (9): Acetic
acid (6 mL) was added to a mixture of zinc dust (211 mg, 3.1 mmol) and
corresponding alkylammonium acetate as an off white solid (2.45 g, 78%).
[
±±b]
±
compound 7
(±11 mg, 1.±2 mmol). The reaction mixture was protected
H NMR (411 MHz, CDCl
3
): d ˆ 1.8± (s, 3H; ±8-CH
3
), 1.89 (m, 6H; eicosyl
Chem. Eur. J. 2002, 8, No. ±3
¹ WILEY-VCH Verlag GmbH, 6945± Weinheim, Germany, 2112
1947-6539/12/18±3-2935 $ 21.11+.51/1
2935

A. P. Davis, J. de Mendoza, J. G. de Vries et al.
FULL PAPER
CH
3
and ±9-CH
3
), 1.95 (d, 3H; 2±-CH
3
), 2.15 (s, 3H; acetate CH
3
), 2.8± (m,
solution in TRIS or phosphate buffer (as above). The system set-up for the
simultaneous extraction and stripping process using two hollow fibre
contactors is shown in Figure 4. Each hollow fibre module (Dialysis
Module type LD OC 12, Microdyn, Wuppertal, Germany) contains ±281
hydrophilic cellulose fibres. The characteristics of the module and fibres are
summarised in Table 3. In addition, the housing material is polycarbonate
and the potting material (used to secure the fibre bundle to the tube sheet)
is polyurethane. The system is driven by three peristaltic pumps (Watson &
Marlow 515U) run at 41 rpm.
±
H; 3b-H), 4.1± (t, 2H; CO
2
CH
2
), 5.13 (m, ±H; 7b-H), 5.25 (m, ±H; ±2b-
‡
H), 6.29 (br, 3H; NH
3
), 7.52 (m, 8H; ArH), 9.37 (s, ±H; 7a-NH), 9.57 (s,
±
2
4
±
2
H; ±2a-NH); ±3C NMR (±11 MHz, CDCl
5.5, 26.8, 28.2, 28.8, 29.1, 29.±, 29.3, 31.4, 3±.±, 3±.5, 33.2, 34.5, 37.2, 45.±,
7.3, 64.±, ±±7.6, ±±7.8, ±25.7, ±26.1, ±42.2, ±52.7 (NHCO), ±53.± (NHCO),
73.8 (C-24), ±78.5 (CH
.21 mmol) and sodium bicarbonate (2.18 g, 25.2 mmol) in DMF (41 mL)
3
): d ˆ ±2.3, ±3.7, ±7.3, 2±.9, 22.2,
3 2
CO ). To a solution of this material (2.4 g,
was added reagent 8 (1.94 g, 6.3 mmol). The reaction was stirred for 3 h at
18C under argon. The solvents were removed and the product was
8
Separation experiments: The source phase (511 mL) was circulated through
purified by flash chromatography (dichloromethane/methanol, 21:±). To
ensure that chloride was the counter ion, the salt was taken up in
dichloromethane and washed with sodium hydroxide (± m) followed by
distilled water. The organic layer was washed with HCl (1.5 m) followed by
distilled water. The organic phase was dried and concentrated under
the shell of one module while the receiving phase (511 mL) was circulated
˱
through the shell of the other (both at ±8 mLmin ). Through the lumen of
˱
both modules was circulated the organic phase (±25 mL, 33 mLmin ). The
concentrations of the enantiomers in each aqueous phase were monitored
for 48 h using HPLC (Chirobiotic T, Astec, USA; H O (±% ammonium
2
acetate)-MeOH, 81:21, pH 4.±). AUV detector (Spectra Physics, USA) at a
wavelength of 254 nm was used.
reduced pressure to yield 11 (±.47 g, 68%) as a white solid. R
f
ˆ 1.79
(
dichloromethane/methanol, ±5:±); m.p. 234 ±235 8C; IR (film from
˱
±
CDCl
3
): nÄ max ˆ ±729 cm (CˆO); H NMR (411 MHz, CDCl
), 1.87 ±1.91 (m, 6H; eicosyl CH and ±9-CH
), 3.14 ( m, ±H; 3b-H), 3.69 ( br s, 4H;
NH), 4.11 (t, J(H,H) ˆ 6.8 Hz, 2H; CO CH ), 4.91 ( br s, ±H; 7b-
H), 5.21 (br s, ±H; ±2b-H), 7.49 ±7.52 (m, 4H; ArH), 7.88 ±7.95 (m, 4H;
3
): d ˆ 1.77
(
s, 3H; ±8-CH
3
3
3
), 1.96 (d,
3
J(H,H) ˆ 6.5 Hz, 3H; 2±-CH
3
NH(CH
2
)
2
2
2
Acknowledgement
±
3
ArH), 9.±± (br s , ±H), 9.58 (br s, ±H), 9.68 (br s , ±H); C NMR (±11 MHz,
CDCl ), ±4.1 (±8-CH ), ±7.6 (2±-CH ), 2±.7 (±9-CH ), 22.6
), 25.3 (CH ), 25.8 (CH ), 27.1 (CH ), 27.2 (CH ), 28.±
), (CH ), 29.± (CH ), 29.4 (CH ), 29.5 (CH ),
), 31.6 (CH ), 31.9 (CH ), 33.4 (C), 34.1
), 34.7 (CH), 34.8 (CH ), 38.1 (CH), 41.6 (CH), 42.7 (CH), 44.9 (C),
7.1 (CH), 53.1 (CH), 64.5 (CH ), 7±.1 (CH), 75.5 (CH), ±±7.9 and ±±8.6
): d ˆ ±2.3 (CH
), 23.8 (CH
CH), 28.5 (CH
9.6 (CH
CH
3
3
3
3
This research was supported by the EU TMR programme (contract ERB-
FMRX-CT98 ±1233). We thank Dr. Veerle Cauwenberg and Dr. Wouter
Pronk (DSM Research, BC-PT) for stimulating discussions and help with
the membrane unit.
3
(
(
2
(
CH
2
2
2
2
2
2
2
2
2
), 29.2 (CH
), 3±.2 (CH ), 3±.8 (CH
2 2 2
2
2
2
2
2
2
2
4
2
(
±
(
C
Ar-CH), ±23.1, ±24.1 and ±25.2 (Ar-C), ±25.6 and ±25.8 (Ar-CH), ±42.6 and
42.7 (Ar-C), ±53.2 (OCONH), ±53.5 (OCONH), ±58.3 (C(NH) ), ±74.5
): d ˆ 62.3; HRMS (FAB ) calcd for
[
±] T. H. Webb, C. S. Wilcox, Chem. Soc Rev. 1993, 22, 383; X. X. Zhang,
J. S. Bradshaw, R. M. Izatt, Chem. Rev. 1997, 97, 33±3; H. Ogoshi, T.
Mizutani, Acc. Chem. Res. 1998, 31, 8±.
3
±
9
‡
CO
2
CH
2
); F NMR (376 MHz, CDCl
3
‡
63
H
94
N
5
O
6
F
6
[M ‡ H] : ±±31.7±18; found: ±±31.7±11.
[
[
[
[
2] Chirality in Industry II (Eds.: A. N. Collins, G. N. Sheldrake, J.
U-Tube transport experiments: Single enantiomer experiments: A U-tube
glass cell (Figure ±) was employed in all experiments. The temperature was
maintained at 25.1 Æ 1.±8C, and the stirring rate was adjusted to 2611 rpm,
as measured by means of a tachometer (stirring bar dimensions: h ˆ 6 mm,
Crosby), Wiley, 1997.
3] H.-J. Schneider, A. Yatsimirsky, Principles AndMetho ds in Supra-
molecular Chemistry, Wiley, Chichester, 2000, p. 287.
4] J. F. T. Keurentjens, J. F. M. Voermans in Chirality in Industry II (Eds.:
A. N. Collins, G. N. Sheldrake, J. Crosby), Wiley, 1997 pp. ±57 ±±81.
5] a) M. Newcomb, J. L. Toner, R. C. Hegelson, D. J. Cram, J. Am. Chem.
Soc. 1979, 101, 494±; b) J.-M. Lehn, Pure Appl. Chem. 1979, 51, 979;
c) J. L. Sessler, A. Andrievsky, Chem. Eur. J. 1998, 4, ±59; d) W. H.
Pirkle, W. E. Bowen, Tetrahedron-Asymmetry 1994, 5, 773.
6] W. H. Pirkle, E. M. Doherty, J. Am. Chem. Soc. 1989, 111, 4±±3.
7] K. Inoue, A. Miyahara, T. Itaya, J. Am. Chem. Soc. 1997, 119, 6±9±; A.
Maruyama, N. Adachi, T. Takatsuki, M. Torii, K. Sanui, N. Ogata,
Macromolecules 1990, 23, 2748.
À2
Æ ˆ 2 mm). N-Ac-l-PheOH or N-Ac-d-PheOH, (5.1  ±1 m) in TRIS
buffer (3 mL; pH 8.±, 3 mL) were employed as the source phase. The
organic (membrane) phase consisted of a solution of 11 in ±,2-dichloro-
À3
ethane (±1 mL; ±.7± Â ±1 m). The receiving phase was a KBr (1.5m)
solution in TRIS buffer (pH8.±, 3 mL). Amino acid uptake at the source
phase and release at the receiving phase was monitored by HPLC (Perkin
Elmer Integral 4111, Scharlau LC±8 column, UV detector at l ˆ 231 nm,
injection of 51 mL aliquots taken at one or two hours intervals) using benzyl
[
[
alcohol as an external standard. A gradient H
2
2 3
O ± CH CN (1 ±±11% in
1 minutes), followed by pure CH CN (5 min) was employed as eluent.
3
[
8] K. Konishi, K. Yahara, H. Toshishige, T. Aida, S. Inoue, J. Am. Chem.
Soc. 1994, 116, ±337; V. Andrisano, G. Gottarelli, S. Masiero, E. H.
Heijne, S. Pieraccini, G. P. Spada, Angew. Chem. 1999, 111, 2543;
Angew. Chem. Int. Ed. 1999, 38, 2386.
Retention times: N-Ac-PheOH t ˆ 9.± min; benzyl alcohol t ˆ ±4.2 min.
Transport of racemic amino acid: The same U-tube glass cell was employed.
The dimensions of the stirring bar for this experiments (h ˆ 9 mm, Æ ˆ
5
mm, egg shaped) allowed a stirring rate of only ±111 rpm. N-Ac-dl-
[
9] G. J. Pernia, J. D. Kilburn, M. Rowley, J. Chem. Soc. Chem. Commun.
À2
PheOH, (5.1 Â ±1 m) in TRIS buffer (pH 8.±, 3 mL) was employed as the
1
995, 315.
source phase, and the receiving phase consisted of a KBr (1.5 m) solution in
[
[
±1] C. L. Hannon, E. V. Anslyn, Bioorg. Chem. Frontiers 1993, 3, ±93; A.
Gal a¬ n, D. Andreu, A. M. Echavarren, P. Prados, J. de Mendoza, J. Am.
Chem. Soc. 1992, 114, ±5±±; E. Fan, S. A. V. Arman, S. Kincaid, A. D.
Hamilton, J. Am. Chem. Soc. 1993, 115, 369; A. Metzger, K. Gloe, H.
Stephan, F. P. Schmidtchen, J. Org. Chem. 1996, 61, 215±.
±±] a) A. P. Davis, L. J. Lawless, Chem. Commun. 1999, 9; b) L. J. Lawless,
A. G. Blackburn, A. J. Ayling, M. N. P e¬ rez-Pay a¬ n, A. P. Davis, J.
Chem. Soc. Perkin Trans. 1 2001, ±329.
±2] M. S. Muche, M. W. Gˆbel, Angew. Chem. 1996, 108, 2263; Angew.
Chem. Int. Ed. Engl. 1996, 35, 2±26.
±3] The six-membered ring analogue of 8 appears to be significantly less
reactive, presumably due to steric hindrance and/or ring strain effects.
TRIS buffer (pH8.±, 3 mL), as above. A solution of 11 in dichloromethane
À4
(
8.6 Â ±1 m, ±1 mL) was employed as the bulk membrane. The concen-
trations of the amino acid enantiomers in the receiving phase were
monitored by HPLC (Waters 611 Controller, Waters 2487 UV/Vis detector
at l ˆ 254 nm), employing a chiral column (Chirobiotic T, Astec, USA)
2
under reverse phase conditions (H O (±% TEAA)-MeOH, 81:21).
Retention times: N-Ac-l-PheOH t ˆ 4.3 min; N-Ac-d-PheOH t ˆ 6.5 min.
Aliquots of ±1 mL from the receiving phase were injected every hour.
[
[
˱
˱
Calibration was performed injecting ±.± Â ±1 mg mL of a standard
solution of N-Ac-dl-PheOH.
Transport through hollow fibre membranes: Materials andapparatus : The
À2
source phase consisted of N-Ac-dl-PheOH (4.8 Â ±1 m) in TRIS buffer
[±4] This experiment was performed in a ™W-tube∫, the organic solvent
being less dense than water.
À2
(
4.8 Â ±1 m, pH 8.1 Æ 1.±) or phosphate buffer (1.± m, pH 7.4). The organic
phase consisted of a solution of 11 (3 ±8 m m) in octanol/hexane 2.5% (v/v)
Merck, Germany). The receiving phase was a 1.± ±± m potassium bromide
(
Received: January 25, 2112 [F3822]
2936
¹ WILEY-VCH Verlag GmbH, 6945± Weinheim, Germany, 2112
1947-6539/12/18±3-2936 $ 21.11+.51/1
Chem. Eur. J. 2002, 8, No. ±3