Synthesis and 1H NMR titration study of 7-deoxycholic amide or cholane ionophores containing different ion-recognizing groups at C3 and C12

Article abstract of DOI:10.1002/hc.21002

Tweezer-type ionophores containing C3-carbamoylpropanamidoacetoxy and C12-dithiocarbamoyl groups on a 7-deoxycholic amide or cholane derivative were designed and synthesized. A representative ¹H NMR titration study indicated that newly synthesi

Full text of DOI:10.1002/hc.21002

Heteroatom Chemistry
Volume 23, Number 2, 2012
1
Synthesis and H NMR Titration Study of
7-Deoxycholic Amide or Cholane Ionophores
Containing Different Ion-Recognizing Groups
at C3 and C12
Hyunju Oh, Sang Keun Han, and Byeong Hyo Kim
Department of Chemistry, Kwangwoon University, Seoul 139-701, Republic of Korea
Received 6 September 2012; revised 28 November 2012
More recently, we have found a synthetic method
ABSTRACT: Tweezer-type ionophores contain-
for introducing different functional groups in a step-
ing C3-carbamoylpropanamidoacetoxy and C12-
by-step fashion at C3 and C12 on DCAB, which made
dithiocarbamoyl groups on a 7-deoxycholic amide
possible the development of more DCAB-based neu-
or cholane derivative were designed and synthesized.
tral carriers with different ion-recognizing groups
A representative 1H NMR titration study indicated
in the same molecule [4c,5,6]. Nonsymmetrically
that newly synthesized ionophores form 1:1 complexes
substituted tweezer-type DCAB-based ionophores,
with the Ca2+ ion.
2012 Wiley Periodicals, Inc. Het-
ꢀ
C
which contain a nitrogen or sulfur atom binding site
at C3 and a sulfur atom binding site at C12, show ex-
cellent results for silver(I) ions [4c], whereas others
that contain different ion-recognizing groups at the
C3 position show outstanding results for copper and
mercury [5,6]. Accordingly, our synthetic method for
the step-by-step introduction of different functional
groups at C3 and C12 on DCAB made it possible to
be applicable to more ions.
Thus, we aimed to synthesize and study new
types of DCAB-based ion-selective ionophores,
which contain two different ion-recognizing func-
tional groups with varied lengths at C3 and C12.
Herein, we report the synthesis and 1H NMR analysis
of new nonsymmetrically substituted tweezer-type
DCAB-based ionophores for use as highly selective
metal ion hosts.
eroatom Chem 23:187–194, 2012; View this article online
at wileyonlinelibrary.com. DOI 10.1002/hc.21002
INTRODUCTION
Ion-selective electrodes are unique examples of
chemical sensors that apply the concept of molec-
ular recognition chemistry. Thus, we have devel-
oped and utilized the idea of molecular tweezer-type
neutral carriers based on a deoxycholic acid back-
bone (DCAB) to yield highly selective ionophores for
carbonate [1], chloride [2], calcium [3], silver [4],
copper [5], and mercury [6], since the DCAB-based
carriers are easy to design, easy to prepare from in-
expensive deoxycholic acid (1; Fig. 1), and applicable
to various ions by varying the types of ion recogniz-
ing groups attached to the hydroxyl linkers at C3 and
C12. The hydroxyl linkers at the C3 and C12 carbons
of the DCAB frame are approximately parallel and
RESULTS AND DISCUSSION
Synthesis of Ionophores
˚
about 6 A apart, a reasonable distance for various
ions [7].
As mentioned in the Introduction section, we re-
cently found that mono-substituted 5β-cholan-24-
amide (4) instead of di-substituted 5β-cholan-24-
amide could be prepared from deoxycholic acid
Correspondence to: Byeong Hyo Kim; e-mail: bhkim@kw.ac.kr.
c
ꢀ
2012 Wiley Periodicals, Inc.
187

188 Oh, Han, and Kim
H
Me
Me
H
Me
Me
Me
Me
R
1
O
O
CO H
2
R
R
= CON(C H ) , CH
8 17 2 3
OH
1
2
= C H , i-C H , Ph
2
5
4 9
S
O
O
OH
R
Deoxycholic acid (1)
2
S
N
S
R
2
S
FIGURE 1 Structures of deoxycholic acid (1) and representative nonsymmetrically substituted DCAB-based ion-selective
ionophores.
using a newly developed procedure [4c]. In addi-
tion to optimizing ion recognition groups at the
3α- and 12α-positions, the evaluation of lipophilic
characteristics of the 5β-side chain is also impor-
tant. Therefore, the preparation of 5β-cholan-24-
amides with long alkyl chains on the amide group
and 5β-cholane derivatives with an alkyl chain but
no amide group should likewise be tested to ob-
tain the best ionophore. Thus, 4 and 5, contain-
ing 12α-dithiocarbamoyl groups on a 7-deoxycholic
amide or cholane derivatives, were prepared from 7-
deoxycholic acid (1) using the newly developed pro-
cedure and were used as starting substrates in this
study.
Among the linking ion-recognizing moieties de-
veloped, glycolic diamide groups exhibited the best
calcium ion recognition [3]. Thus, we introduced the
suitably lengthened amide-related functional groups
at C3 of DCAB with the expectation of new cal-
cium ion-selective ionophores. For this purpose,
N, N-di(alkyl, aryl)carbamoylpropanoic acid (3) was
selected as an ion recognition linkage, was pre-
pared through the two-step synthesis described in
Scheme 1, and was introduced at the C3 positions
in the starting substrates (4 and 5) as an ion-
recognizing diamide group.
of the substrates, esterification of the 3α-hydroxyl
group was performed with α-chloroacetyl chloride
to convert the hydroxyl group into an α-chloroacetyl
ester, which can be used as a precursor for the prop-
erly extended ion-recognizing diamide moiety. Thus,
4 or 5 was treated with α-chloroacetyl chloride in
the presence of calcium hydride and tetrabutylam-
monium bromide in toluene at 90◦C for 2 h to pro-
duce 6 or 7, correspondingly (Scheme 2, (a)). Then,
the chloro group of 6 or 7 was transformed into an
amino group by a substitution reaction with sodium
azide in DMF (50◦C for 12 h), followed by the re-
duction of the azido group using nanosized zinc
powder in acetic acid at room temperature (Scheme
2, (b) and (c)). Afterward, the target ionophores
12a–12c or 13a–13c were obtained in a reason-
able yield by reacting 10a–10c or 11a–11c with
N, N-di(alkyl, aryl)carbamoylpropanoic acid (3)/1-
(3-dimethylaminopropyl)-3-ethylcarbodiimide hy-
drochloride (EDC)/1-hydroxybenzotriazole hydrate
(HOBt) in dichloromethane at room temperature for
48 h (Scheme 2, (d)). Hence, six different prospective
ionophores were successfully synthesized and their
1
structures were fully identified with H NMR, 13C
NMR, FTIR, and FAB MASS (12a–12c) or HRMS
(13a–13c). Overall, the yields of the four-step syn-
thesis from 4 to 12a–12c or from 5 to 13a–13c
were 43%–49% and 50%–65%, respectively, and were
shown to be excellent synthetic sequences.
The starting substrates (4 and 5) were prepared
from deoxycholic acid (1) using the procedure de-
scribed in our previous paper [4c]. After preparation
O
O
R
N
O
R
N
(b)
(a)
Cl
MeO
MeO
R
HO
R
O
O
O
2a; R = ethyl (49%)
2b; R = i-butyl (74%)
2c; R = phenyl (88%)
3a; R = ethyl (49%)
3b; R = i-butyl (74%)
3c; R = phenyl (88%)
SCHEME 1 (a) HNR2, TEA, CH2Cl2, rt, 3 h; (b) LiOH, H2O, THF, rt, 3 h.
Heteroatom Chemistry DOI 10.1002/hc

Synthesis and 1H NMR Titration Study of 7-Deoxycholic Amide or Cholane Ionophores 189
H
H
Me
O
Me H
O
Me
Me
Me
Me
Me
Me
Me
(a)
R1
CO2H
R1
O
OH
O
OH
1
O
O
OH
S
S
S
S
Cl
N(C2H5)2
N(C2H5)2
4; R1 = CON(C8H17 2
)
5; R1 = CH3
6; R1 = CON(C8H17)2 (96%)
7; R1 = CH3 (93%)
(b)
H
Me
H
Me
Me
O
H
Me
Me
Me
Me
Me
Me
(d)
(c)
R1
R1
R1
O
O
O
O
O
O
O
O
O
O
S
O
S
S
S
S
S
H2N
N3
N(C2H5)2
HN
N(C2H5)2
N(C2H5)2
O
12a; R1 = CON(C8H17)2, R' = C2H5 (59%)
12b; R1 = CON(C8H17)2, R' = i-C4H9 (51%)
12c; R1 = CON(C8H17)2, R' = C6H5 (55%)
13a; R1 = CH3, R' = C2H5 (77%)
8; R1 = CON(C8H17)2 (96%)
9; R1 = CH3 (97%)
10; R1 = CON(C8H17)2 (91%)
11; R1 = CH3 (94%)
O
N R'
R'
13b; R1 = CH3, R' = i-C4H9 (60%)
13c; R1 = CH3, R' = C6H5 (70%)
SCHEME 2 (a) ClCH2COCl, CaH2, Bu4NBr, toluene, 90◦C, 2 h; (b) NaN3, DMF, 50oC, 12 h; (c) nanosized Zn powder, acetic
acid, rt, 24 h; (d) 3, EDC, HOBt, CH2Cl2, DMF, rt, 48 h.
specific ion is the initial step in the development of
a new ionophore.
NMR Titration Experiments of Ionophores
with the Ca2+ Ion
1
Therefore, H NMR titration experiments were
The calcium ion, one of the most important
electrolytes in physiological systems, is known
to form 1:2 and 1:3 complexes with the non-
cyclic diamides, i.e., (–)-(R, R) − N,Nꢁ-[bis(11-
ethoxycarbonyl)undecyl]-N,Nꢁ-4,5-tetramethyl-3,6-
dioxaoctanediamide (ETH 1001) and N,N,Nꢁ,Nꢁ-
tetracyclo-3-oxapantanediamide (ETH 129), re-
spectively [8]. Our previously synthesized tweezer
-type ionophores, such as N,N-dioctyl-3α,12α-bis
(N-heptyl-N-methylcarbamoyl-methoxyacetamido-
acetoxy)-5β-cholan-24-amide ionophore [3] and
ionophores that have a bithiophene moiety on the
3α-position and diphenylaminothioxomethylth-
ioacetyloxy group on the 12α-position of cholan-
24-amide/cholane [4c], which exhibited excellent
selectivity and sensitivity to calcium and silver
ions, respectively, formed 1:1 complexes using two
ion-recognizing groups that were properly oriented
in a neutral carrier. Consequently, whether an
ionophore will form a proper complex with the
examined after the syntheses of target neutral car-
riers to verify the recognition properties of the
newly synthesized calcium ion-selective ionophores
toward the calcium ion. 1H NMR titrations were
performed with the synthesized ionophore and
Ca(SCN)2 in methanol-d4/acetone-d6 (v/v = 1/1) co-
solvent, which was determined to be the optimum
condition according to the titration experiments.
The representative best results of the NMR titrations
with 13a are shown in Fig. 2.
The 1H NMR spectrum of the free ionophore 13a
shown in Fig. 2a was altered with the addition of
the Ca2+ ion; following the addition of 1 equiv of
Ca(SCN)2 to the ionophore in methanol-d4/acetone-
d6 solution, changes in the chemical shifts of several
peaks were clearly observed (Fig. 2b). While con-
siderable downfield shifts of the Ha, Hc, Hd, and
He peaks that are adjacent to the carbonyl π-bond
or next to the nitrogen of the amide group were
observed, negligible shifts of the Hb peaks on the
Heteroatom Chemistry DOI 10.1002/hc

190 Oh, Han, and Kim
H
Me
O
Me
Me
H
Me
Me
Me
O
O
Ha
O
Ha'
S
O
O
O
S
Hc
Hb
Hb'
S
S
Hc'
O
N
HN
Ca2+
O
N(C2H5)2
O
He
He
He
He
O
HN
O
Hd
NR2'
N
Hd'
13a–Ca2+
13a
(a)
(b)
(c)
FIGURE 2 1H NMR spectra (400 MHz) of the ionophores, (a) free 13a measured in methanol-d4/acetone-d6 (v/v = 1/1), (b)
13a after addition of 1 equiv of Ca(SCN)2, and (c) 13a after addition of 2 equiv of Ca(SCN)2.
dithiocarbamoyl moiety were observed. Moreover,
interesting upfield and downfield shifts are observed
at the peaks of Ha or Hꢁa when a Ca2+ ion is added
to the free ionophore solution. While one of the di-
astereotopic protons, Ha, is shifted downfield due
to the addition of 1 equiv of Ca(SCN)2 to the free
ionophore, the other proton, Hꢁa, experiences an up-
field shift. Obviously, Ha is on the same plane as the
adjacent carbonyl π-bond, and its σ-bond donates
electrons to the carbonyl group via the hyperconju-
gation effect during Ca2+ ion complexation with the
ionophore. These 1H NMR changes can be attributed
to the complexation between the Ca2+ ion and het-
eroatoms in the dithiocarbamoyl and carbamoyl-
propanamidoacetoxy moieties in the 13a ionophore,
especially the oxygen heteroatom of the carbonyl
group.
at Hb. This occurrence is in some way expected on
the basis of the hard–soft acid base concept. In our
previous work, the ionophores that contained softer
sulfur atoms in their binding sites were well tweez-
ered for the soft ions, such as Ag+, Hg2+, and Co2+.
However, a relatively minor contribution to com-
plexation is expected between a soft sulfur atom
and a relatively harder Ca2+ ion. It is surprising that
complexation occurred almost exclusively between a
Ca2+ ion and oxygen atoms of the carbonyl groups of
the ionophores, with no noticeable participation of
the softer sulfur atom. These results strongly imply
that the oxygen atoms in the dithiocarbamoyl and
carbamoylpropanamidoacetoxy moieties are largely
responsible for the calcium ion recognition. In ad-
dition, an increase in the amount of Ca(SCN)2, up
to 2 equiv, did not notably influence the chemical
shifts of the Ha, Hc, Hd, and He peaks for 13a (Fig.
2c). This indicates that ionophore 13a forms a 1:1
complex with the Ca2+ ion, as shown in Fig. 2 (13a–
Ca2+).
Unlike the Ag+ ion ionophores [4c], sulfur het-
eroatoms on the dithiocarbamoyl group seemed to
scarcely participate in the tweezering of Ca2+ ions
as there was no observable chemical shift change
Heteroatom Chemistry DOI 10.1002/hc

Synthesis and 1H NMR Titration Study of 7-Deoxycholic Amide or Cholane Ionophores 191
CONCLUSION
were collected and evaporated in vacuo. The residue
was diluted with CH2Cl2 (200 mL) and washed with
In this article, we synthesized uniquely designed cal-
cium ion-selective ionophores based on nonsymmet-
rically substituted tweezer-type ionophores contain-
ing a carbamoylpropanamidoacetoxy moiety at the
C3-position and dithiocarbamoyl groups at the C12-
position of a 7-deoxycholic amide or cholane deriva-
tive. 1H NMR titration experiments implied that the
synthesized ionophore forms a 1:1 complex with
the Ca2+ ion using oxygen atoms from the carbonyl
groups of the ion-recognizing linkage at the C3- and
C12-positions of the neutral ionophore.
saturated NaHCO3(aq) (2 × 200 mL), washed with
water (1 × 200 mL), dried over MgSO4, and evapo-
rated in vacuo. The crude product was purified by
silica gel chromatography with ethyl acetate/hexane
(2:8) as an eluent to yield 6 and ethyl acetate–hexane
(1.5:8.5) as an eluent to yield 7 as waxy liquids.
3α-Chloroacetoxy-12α-[[[[(diethylamino)thioxo-
methyl] thio]acetyl] oxy] - N,N - dioctyl-5β-cholan-24-
amide (6). Waxy liquid; TLC (silica gel, 35% ethyl
acetate/hexane) Rf 0.57; 1H NMR (400 MHz, CDCl3)
δ 5.13 (br s, 1H, 12β-H), 4.85–4.74 (m, 1H, 3β-H),
4.39 (d, J = 16.5 Hz, 1H), 4.22 (d, J = 16.5 Hz,
1H), 4.10–4.01 (m, 4H), 3.81–3.78 (m, 2H), 3.32–
3.17 (m, 4H), 2.35–2.18 (m, 2H), 1.99–0.72 (m, 88H);
13C NMR (100 MHz, CDCl3) δ 193.3, 172.8, 167.9,
166.8, 77.4, 76.5, 50.1, 49.1, 47.9, 47.5, 46.8, 45.8,
45.2, 41.7, 41.3, 39.2, 35.6, 35.0, 34.7, 34.2, 34.0, 31.9,
31.8, 31.7, 31.4, 30.1, 29.4, 29.3, 29.2, 29.17, 29.14,
27.8, 27.4, 27.0, 26.9, 26.8, 26.4, 25.9, 25.4, 23.4, 22.9,
22.58, 22.57, 17.7, 14.0, 12.6, 12.2, 11.5; IR (CH2Cl2)
νmax 2937, 2862, 1731, 1641, 1459, 1418, 1270, 1183,
1006 cm−1; HRMS calcd. for C49H85ClN2O5S2 m/z
880.5588, found 880.5599.
3α -Chloroacetoxy-12α-[[[[(diethylamino)thioxo-
methyl]thio]acetyl]oxy]-5β-cholane (7). Waxy liquid;
TLC (silica gel, 15% ethyl acetate/hexane) Rf 0.24;
1H NMR (400 MHz, CDCl3) δ 5.13 (br s, 1H, 12β-
H), 4.83–4.75 (m, 1H, 3β-H), 4.39 (d, J = 16.4 Hz,
1H), 4.22 (d, J = 16.4 Hz, 1H), 4.10–3.97 (m, 4H),
3.83–3.78 (m, 2H), 2.00–0.72 (m, 52H); 13C NMR (100
MHz, CDCl3) δ 193.4, 167.9, 166.9, 77.5, 76.5, 50.1,
49.1, 47.6, 46.8, 45.1, 41.8, 41.3, 39.3, 38.1, 35.6, 35.0,
34.7, 34.2, 34.1, 31.9, 27.5, 26.9, 26.4, 25.9, 25.4, 23.5,
22.9, 19.2, 17.8, 14.5, 12.6, 12.2, 11.5; IR (CH2Cl2)
νmax 2958, 2869, 1753, 1732, 1490, 1418, 1271, 1186,
1009 cm−1; HRMS calcd. for C33H54ClNO4S2 m/z
627.3183, found 627.3179.
EXPERIMENTAL
General Considerations
1H and 13C NMR spectra were recorded at 400 (JEOL,
Tokyo, Japan) and 100 MHz, respectively. Chemical
shifts are reported in parts per million relative to
the residual solvent as an internal standard. High-
resolution mass was recorded mostly on a JEOL
JMS-DX303 mass spectrometer, and GC/MS spec-
tra were recorded on an Agilent 6890N GC con-
nected to an Agilent 5975 mass selective detector.
IR spectra were recorded using an MB104 FTIR
(ABB Bomem, Inc., Zurich, Switzerland). Most of
the chemical reagents were purchased from Sigma
Aldrich Co. (St. Louis, Missouri) unless noted other-
wise and were used without purification in most
cases. Solvents were purchased and dried using nor-
mal laboratory techniques. All major products were
isolated by flash column chromatography on silica
gel (230–400 mesh ATSM, purchased from Merck &
Co., Inc. (Whitehouse Station, New Jersey)) using
ethyl acetate/hexane cosolvent as the eluent.
Syntheses
General Procedure for the Preparation of 3α-
Chloroacetoxy - 12α - [[[[(diethylamino)thioxomethyl]
thio]acetyl]- oxy]-N,N-dioctyl-5β-cholan-24-amide 6
and 3α-Chloroacetoxy-12α-[[[[(diethylamino)thiox-
omethyl ] thio] - acetyl ] oxy] - 5β - cholane 7. 12α-
[[[[(diethylamino)thioxomethyl]thio]acetyl]oxy]-3α
-hydroxy-N,N-dioctyl-5β-cholan-24-amide (4) or 12
α - [[[[(diethylamino)thioxomethyl]thio]acetyl]oxy]-
3α-hydroxy-5β-cholane (5) (0.61 mmol), calcium
hydride (130 mg, 3.05 mmol), tetrabutylammonium
bromide (25 mg, 0.08 mmol), and toluene (10
mL) were mixed and stirred. Chloroacetyl chloride
(533 μL, 6.72 mmol) was slowly added and
stirred for 2 h at 90◦C. The mixture was fil-
tered through a pad of Celite (5 g) after cooling
and was then washed with dichloromethane
(200 mL), and the combined filtrate and washings
General Procedure for the Preparation of 3α-
Azidoacetoxy-12α-[[[[(diethylamino)thioxomethyl]t-
hio]acetyl]-oxy]-N,N-dioctyl-5β-cholan-24-amide
8
and 3α - Azidoacetoxy - 12α - [[[[(diethylamino) thiox-
omethyl]thio]acetyl]-oxy]-5β-cholane 9. 3α-Chloro-
acetoxy-12α-[[[[(diethylamino)thioxomethyl]thio]
acetyl]oxy]-N,N-dioctyl-5β-cholan-24-amide(6) or
3α-chloroacetoxy-12α-[[[[(diethylamino)thioxomet-
hyl]thio]acetyl]oxy]-5β-cholane(7) (0.59 mmol) and
sodium azide (60 mg, 0.88 mmol) were dissolved
in DMF (15 mL) and stirred for 12 h at 50◦C. The
reaction mixture was evaporated in vacuo, diluted
with dichloromethane (150 mL), washed with 1 N
HCl (1 × 150 mL) and then with water (1 × 150
mL), dried over MgSO4, and evaporated in vacuo.
Heteroatom Chemistry DOI 10.1002/hc

192 Oh, Han, and Kim
The crude product was purified by chromatography
on silica gel with ethyl acetate–hexane (3:7) as an
eluent to yield 8 and ethyl acetate–hexane (1:9) as
an eluent to yield 9 as waxy liquids.
N,N-Dioctyl-3α-aminoacetoxy-12α-(N,N-diethyl-
thiocarbamoylsulfanylacetoxy) - 5β - cholan - 24 -
amide (10). Waxy liquid; TLC (silica gel,
NH4OH:MeOH:CH2Cl2 = 0.1:1:15) Rf 0.43. Its
NMR spectrum could not be verified due to its
instability. The identity was confirmed after the next
step reaction.
3α - Aminoacetoxy - 12α-(N,N-diethylthiocarbam-
oylsulfanylacetoxy)-5β-cholane (11). Waxy liquid;
TLC (silica gel, NH4OH:MeOH:CH2Cl2 = 0.1:1:15)
Rf 0.33. Its NMR spectrum could not be verified due
to its instability. The identity was confirmed after
the next step reaction.
3α-Azidoacetoxy-12α-[[[[(diethylamino)thioxom-
ethyl]thio]acetyl]oxy]-N,N-dioctyl-5β-cholan-24-ami-
de (8). Waxy liquid; TLC (silica gel, 35% ethyl
acetate/hexane) Rf 0.57; 1H NMR (400 MHz, CDCl3)
δ 5.13 (br s, 1H, 12β-H), 4.85–4.78 (m, 1H, 3β-H),
4.40 (d, J = 16.6 Hz, 1H), 4.19 (d, J = 16.6 Hz, 1H),
4.11–3.95 (m, 2H), 3.91–3.78 (m, 4H), 3.35–3.14
(m, 4H), 2.36–2.14 (m, 2H), 2.00–0.72 (m, 88H);
13C NMR (100 MHz, CDCl3) δ 193.4, 172.8, 167.93,
167.86, 77.4, 76.2, 50.5, 50.1, 49.1, 48.0, 47.5,
46.8, 45.8, 45.2, 41.8, 39.2, 35.6, 35.1, 34.7, 34.2,
34.0, 32.0, 31.8, 31.7, 31.4, 30.1, 29.4, 29.3, 29.22,
29.19, 29.16, 27.8, 27.4, 27.0, 26.9, 26.8, 26.5, 25.9,
25.4, 23.5, 22.9, 22.6, 17.8, 14.1, 12.6, 12.2, 11.5;
IR (CH2Cl2) νmax 2933, 2860, 2106, 1743, 1641,
1465, 1417, 1267, 1201 cm−1; HRMS calcd. for
C49H85N5O5S2 m/z 887.5992, found 887.6002.
3α - Azidoacetoxy - 12α - [[[[(diethylamino)thioxo-
methyl]thio]acetyl]oxy]-5β-cholane (9). Waxy liquid;
TLC (silica gel, 15% ethyl acetate/hexane) Rf 0.23;
1H NMR (400 MHz, CDCl3) δ 5.13 (br s, 1H, 12β-
H), 4.86–4.78 (m, 1H, 3β-H), 4.40 (d, J = 16.4 Hz,
1H), 4.20 (d, J = 16.4 Hz, 1H), 4.11–3.96 (m, 2H),
3.91–3.77 (m, 4H), 2.01–0.72 (m, 46H); 13C NMR
(100 MHz, CDCl3) δ 193.5, 167.91, 167.87, 77.5,
76.2, 50.5, 50.1, 49.1, 47.6, 46.8, 45.1, 41.8, 39.2,
38.1, 35.6, 35.0, 34.7, 34.2, 34.1, 32.0, 27.5, 26.8,
26.5, 25.9, 25.4, 23.5, 22.9, 19.2, 17.8, 14.5, 12.6,
12.2, 11.5; IR (CH2Cl2) νmax 2953, 2869, 2107, 1745,
1492, 1418, 1355, 1267, 1204 cm−1; HRMS calcd. for
C33H54N4O4S2 m/z 634.3587, found 634.3570.
General Procedure for the Preparation of
α-[[[[4-(Di(alkyl, aryl)amino)-1,4-dioxobutyl]amino]
acetyl]oxy]-12α-[[[[(diethylamino)thioxomethyl]thio]
acetyl]-oxy]-N,N-dioctyl-5β-cholan-24-amides 12a,
12b, and 12c. N, N-Di(alkyl, aryl)carbamoylpr
opanoic
acid
(0.79
mmol)
and
1-
hydroxybenzotriazole hydrate (HOBt) (110 mg,
0.79 mmol) were dissolved in dichloromethane
(10 mL), DMF (1 mL) at room temperature,
which was stirred for 10 min. EDC (150 mg, 0.79
mmol) was added to the reaction mixture and
was stirred for 30 min. 3α-Aminoacetoxy-12α-
[[[[(diethylamino)thioxo-methyl]thio]acetyl]-oxy]-
N,N-dioctyl-5β-cholan-24-amide (10, 140 mg, 0.16
mmol) was dissolved in dichloromethane (10 mL),
slowly added to the reaction mixture at 0◦C and
stirred for 48 h at room temperature. The reaction
mixture was diluted with CH2Cl2 (100 mL) and
was washed with saturated NaHCO3(aq) (1 × 100
mL), washed with water (1 × 100 mL), dried over
MgSO4, and evaporated in vacuo. The crude product
was purified by silica gel chromatography with
ethyl acetate as an eluent to yield 12a and ethyl
acetate–hexane (6:4) as an eluent to yield 12b or
12c as waxy solid.
3α-[[[[4-(Diethylamino)-1,4-dioxobutyl]amino]
acetyl] oxy] - 12α - [[[[(diethylamino) thioxomethyl]
thio]-acetyl]oxy] - N,N - dioctyl-5β-cholan-24-amide
(12a). Waxy solid; TLC (silica gel, 20%
acetone/CH2Cl2) Rf 0.45; 1H NMR (400 MHz,
CDCl3) δ 6.67 (t, J = 5.0 Hz, 1H), 5.13 (br s, 1H,
12β-H), 4.81–4.74 (m, 1H, 3β-H), 4.38 (d, J = 16.5
Hz, 1H), 4.22 (d, J = 16.5 Hz, 1H), 4.06–3.95 (m,
4H), 3.86–3.75 (m, 2H), 3.40–3.16 (m, 8H), 2.69–2.59
(m, 4H), 2.36–0.72 (m, 84H); 13C NMR (100 MHz,
CDCl3) δ 193.4, 172.8, 172.7, 170.8, 169.4, 168.0,
77.3, 75.6, 50.2, 49.1, 48.0, 47.5, 46.9, 45.9, 45.2,
41.89, 41.84, 41.75, 40.4, 39.2, 35.7, 35.1, 34.8, 34.3,
34.1, 32.1, 31.83, 31.79, 31.5, 31.3, 30.1, 29.43, 29.38,
29.28, 29.25, 29.22, 28.6, 27.8, 27.4, 27.1, 27.0, 26.9,
26.6, 25.9, 25.5, 23.5, 23.0, 17.8, 14.1, 13.1, 12.7,
General Procedure for the Preparation of 3α-
Aminoacetoxy - 12α - [[[[(diethylamino)thioxomethyl]
thio]acetyl]-oxy]-N,N-dioctyl-5β-cholan-24-amide 10
and
3α-Aminoacetoxy-12α-[[[[(diethylamino)th-
acetyl]oxy]-5β-cholane 11. 3α-
ioxomethyl]thio]
-
Azidoacetoxy - 12α-[[[[(diethylamino)thioxomethyl]
thio]acetyl]oxy]-N,N-dioctyl - 5β - cholan - 24-amide
(8)
or
3α-azidoacetoxy-12α-[[[[(diethylamino)-
thioxomethyl]thio]acetyl]oxy]-5β-cholane (9) (0.56
mmol) and nanosized Zn powder (500 mg, 7.61
mmol) were dissolved in glacial acetic acid (20
mL) and stirred for 24 h at room temperature. The
reaction mixture was filtered through a pad of Celite
(5 g) and washed with dichloromethane (200 mL).
The crude product was purified by chromatography
on silica gel with NH4OH/MeOH/CH2Cl2 (0.1:1:15)
as an eluent to yield 10 and NH4OH/MeOH/CH2Cl2
(0.2:2:15) as an eluent to yield 11 as waxy liquids.
Heteroatom Chemistry DOI 10.1002/hc

Synthesis and 1H NMR Titration Study of 7-Deoxycholic Amide or Cholane Ionophores 193
12.3, 11.5; IR (CH2Cl2) νmax 3450, 2926, 2860, 1742,
[[[[(diethylamino)thioxo - methyl] thio]-acetyl]oxy]-
5β-cholane (11, 220 mg, 0.36 mmol) was dissolved
in dichloromethane (10 mL), slowly added to the re-
action mixture at 0◦C and stirred for 48 h at room
temperature. The reaction mixture was diluted with
CH2Cl2 (100 mL) and was washed with saturated
NaHCO3(aq) (1 × 100 mL), washed with water (1
× 100 mL), dried over MgSO4, and evaporated in
vacuo. The crude product was purified by silica gel
chromatography with ethyl acetate as an eluent to
yield 13a and ethyl acetate–hexane (6:4) as an elu-
ent to yield 13b and 13c as a waxy solid.
1724, 1636, 1463, 1418, 1271, 1203, 1143 cm−1; MS
(FAB) m/e 1018 (M+), 581 (M+-C40H71NO).
3α-[[[[4-(Diisobutylamino)-1,4-dioxobutyl]ami-
no]acetyl]oxy]-12α - [[[[(diethylamino)thioxomethyl]
thio] - acetyl]oxy] - N,N - dioctyl-5β-cholan-24-amide
(12b). Waxy solid; TLC (silica gel, 15%
acetone/CH2Cl2) Rf 0.52; 1H NMR (400 MHz,
CDCl3) δ 6.67 (t, J = 5.0 Hz, 1H), 5.13 (br s, 1H,
12β-H), 4.80–4.72 (m, 1H, 3β-H), 4.38 (d, J = 16.5
Hz, 1H), 4.22 (d, J = 16.5 Hz, 1H), 4.04–3.98 (m,
4H), 3.84–3.78 (m, 2H), 3.33–3.10 (m, 8H), 2.70 (t,J
= 6.3 Hz, 2H), 2.61 (t,J = 6.3 Hz, 2H), 2.33–0.72 (m,
92H); 13C NMR (100 MHz, CDCl3) δ 193.4, 172.9,
172.7, 172.1, 169.4, 168.0, 77.4, 75.5, 55.3, 53.3,
50.2, 49.1, 48.0, 47.5, 46.8, 45.9, 45.2, 41.8, 41.7,
39.2, 35.7, 35.1, 34.8, 34.1, 32.1, 31.81, 31.77, 31.52,
31.46, 30.1, 29.41, 29.36, 29.25, 29.23, 29.20, 28.9,
27.8, 27.7, 27.4, 27.1, 27.0, 26.9, 26.53, 26.50, 25.9,
25.5, 23.5, 23.0, 22.6, 20.2, 20.1, 17.8, 14.1, 12.7,
12.3, 11.5; IR (CH2Cl2) νmax 3323, 2958, 2927, 2865,
1745, 1729, 1636, 1466, 1419, 1272, 1202, 1144
cm−1; MS (FAB) m/e 1074 (M+), 581 (M+-C40H71NO).
3α - [[[[4- (Diphenylamino)-1,4-dioxobutyl]ami-
no] acetyl] oxy] - 12α-[[[[(diethylamino)thioxomethyl]
thio] - acetyl]oxy] - N,N - dioctyl-5β-cholan-24-amide
(12c). Waxy solid; TLC (silica gel, 10%
acetone/CH2Cl2) Rf 0.40; 1H NMR (400 MHz,
CDCl3) δ 7.31 (br s, 10H), 6.51 (t, J = 5.2 Hz, 1H),
5.13 (br s, 1H, 12β-H), 4.81–4.73 (m, 1H, 3β-H),
4.39 (d, J = 16.5 Hz, 1H), 4.22 (d, J = 16.5 Hz, 1H),
4.03–3.93 (m, 4H), 3.81–3.79 (m, 2H), 3.35–3.18 (m,
4H), 2.59 (s, 4H), 2.36–0.72 (m, 85H); 13C NMR (100
MHz, CDCl3) δ 193.3, 172.8, 172.3, 172.0, 169.3,
167.9, 142.5, 129.7, 128.8, 127.9, 126.4, 77.2, 75.5,
60.3, 50.1, 49.0, 47.9, 47.4, 46.8, 45.8, 45.1, 41.8,
39.1, 35.6, 35.0, 34.7, 34.2, 34.0, 32.0, 31.73, 31.70,
31.4, 31.2, 31.0, 30.0, 29.33, 29.28, 29.18, 29.15,
29.13, 27.7, 27.34, 27.0, 26.9, 26.8, 26.5, 25.8, 25.4,
23.4, 22.9, 22.6, 17.7, 14.1, 14.0, 12.6, 12.2, 11.5;
IR (CH2Cl2) νmax 3445, 3065, 2931, 2857, 2083,
1722, 1670, 1644, 1488, 1419, 1377, 1358, 1265,
1203, 756, 702 cm−1; MS (FAB) m/e 1114 (M+), 581
(M+-C40H71NO).
3α-[[[[4-(Diethylamino)-1,4-dioxobutyl]amino]
acetyl] oxy] - 12α - [[[[(diethylamino) thioxomethyl]
thio]-acetyl]oxy]-5β-cholane (13a). Waxy solid; TLC
1
(silica gel, 20% acetone/CH2Cl2) Rf 0.44; H NMR
(400 MHz, CDCl3) δ 6.78 (t, J = 5.2 Hz, 1H), 5.13 (br
s, 1H, 12β-H), 4.81–4.73 (m, 1H, 3β-H), 4.39 (d, J =
16.4 Hz, 1H), 4.21 (d, J = 16.4 Hz, 1H), 4.06–3.94 (m,
4H), 3.81 (m, 2H), 3.40–3.30 (m, 4H), 2.69–2.59 (m,
4H), 1.97–0.71 (m, 56H); 13C NMR (100 MHz, CDCl3)
δ 193.5, 172.7, 170.8, 169.4, 168.0, 77.4, 75.5, 50.2,
49.1, 47.6, 46.8, 45.1, 41.89, 41.86, 41.7, 40.4, 38.1,
35.7, 35.1, 34.8, 34.3, 34.1, 32.1, 31.3, 28.5, 27.5, 26.9,
26.5, 25.9, 25.5, 23.5, 23.0, 19.2, 17.8, 14.6, 14.1, 13.1,
12.7, 12.2, 11.5; IR (CH2Cl2) νmax 3379, 2955, 2936,
2872, 1741, 1673, 1626, 1543, 1487, 1420, 1270, 1203
cm−1; HRMS calcd. for C41H69N3O6S2 m/e 763.4628,
found 763.4627.
3α-[[[[4-(Diisobytylamino)-1,4-dioxobutyl]ami-
no]acetyl]oxy]-12α -[[[[(diethylamino)thioxomethyl]
thio]-acetyl]oxy]-5β-cholane (13b). Waxy solid; TLC
1
(silica gel, 15% acetone/CH2Cl2) Rf 0.53; H NMR
(400 MHz, CDCl3) δ 6.62 (t, J = 5.0 Hz, 1H), 5.13
(br s, 1H, 12β-H), 4.80–4.72 (m, 1H, 3β-H), 4.39 (d,
J = 16.4 Hz, 1H), 4.21 (d, J = 16.4 Hz, 1H), 4.07–
3.94 (m, 4H), 3.81 (m, 2H), 3.19 (d, 2H), 3.11 (d,
2H), 2.70 (t, 2H), 2.60 (t, 2H), 2.05–0.71 (m, 64H);
13C NMR (100 MHz, CDCl3) δ 193.5, 172.7, 172.0,
169.4, 168.0, 77.5, 75.6, 55.3, 53.3, 50.2, 49.1, 47.6,
46.8, 45.1, 41.9, 41.7, 39.3, 38.1, 35.7, 35.1, 34.8, 34.3,
34.1, 32.1, 31.6, 29.0, 27.7, 27.5, 26.9, 26.53, 26.51,
25.9, 25.5, 23.5, 23.0, 20.2, 20.1, 19.2, 17.8, 14.6,
12.7, 12.2, 11.5; IR (CH2Cl2) νmax 3322, 2961, 2931,
2871, 1744, 1676, 1638, 1544, 1420, 1269, 1203, 1146
cm−1; HRMS calcd. for C45H77N3O6S2 m/e 819.5254,
found 819.5253.
General Procedure for the Preparation of
3α-[[[[4-(Di(alkyl, aryl)amino)-1,4-dioxobutyl]ami-
no]acetyl]oxy]- 12α-[[[[(diethylamino)thioxomethyl]
thio]acetyl]oxy]-5β-cholanes 13a, 13b, and 13c.
N, N-Di(alkyl, aryl)carbamoylpropanoic acid (1.78
mmol) and HOBt (240 mg, 1.78 mmol) were dis-
solved in dichloromethane (10 mL), DMF (1 mL)
at room temperature and stirred for 10 min. EDC
(340 mg, 1.78 mmol) was added to the reaction mix-
ture and stirred for 30 min. 3α-Aminoacetoxy-12α-
3α -[[[[4-(Diphenylamino)-1,4-dioxobutyl]ami-
no]acetyl]oxy]-12α-[[[[(diethylamino)thioxomethyl]
thio]-acetyl]oxy]-5β-cholane (13c). Waxy solid; TLC
1
(silica gel, 10% acetone/CH2Cl2) Rf 0.45; H NMR
(400 MHz, CDCl3) δ 7.28 (br s, 10H), 6.55 (t, J = 5.1
Hz, 1H), 5.11 (br s, 1H, 12β-H), 4.78–4.71 (m, 1H, 3β-
H), 4.38 (d, J = 16.3 Hz, 1H), 4.19 (d, J = 16.3 Hz,
1H), 4.04–3.94 (m, 4H), 3.77 (m, 2H), 2.57 (s, 4H),
Heteroatom Chemistry DOI 10.1002/hc

194 Oh, Han, and Kim
1.92–0.69 (m, 49H); 13C NMR (100 MHz, CDCl3) δ
193.4, 172.3, 172.1, 169.4, 168.0, 142.5, 129.8, 128.8,
128.1, 126.4, 77.5, 75.6, 60.4, 50.2, 49.1, 47.6, 46.8,
45.1, 44.9, 41.8, 41.7, 39.2, 38.1, 35.7, 35.1, 34.8, 34.2,
34.1, 34.0, 32.1, 31.2, 31.0, 27.5, 26.9, 26.5, 25.9, 25.4,
23.5, 22.9, 21.0, 19.2, 17.8, 14.6, 14.2, 12.7, 12.2, 11.5;
IR (CH2Cl2) νmax 3341, 3061, 2955, 2936, 2871, 1951,
1890, 1871, 1739, 1667, 1493, 1269, 1205, 739, 699
cm−1; HRMS calcd. for C49H69N3O6S2 m/e 859.4628,
found 859.4614.
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2005, 66, 794–804; (c) On, J. H.; Cho, K. T.; Park, Y.;
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ACKNOWLEDGMENT
This work was supported by Kwangwoon University
in 2010.
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Heteroatom Chemistry DOI 10.1002/hc