Synthesis of cholic acid based calixpyrroles and porphyrins

Article abstract of DOI:10.1016/j.steroids.2012.04.008

New cholic acid based calix[4]pyrroles and porphyrins were prepared and their properties were studied. It was confirmed by spectral measurements that the superassembly of 5,15-bis(3α,7α,12α-trihydroxy-5β- cholan-24-yl)-10,20-diphenylporphyrin, the best candidate for this study from the conjugates prepared, may be influenced not only by the solvent mixture composition (polar/non-polar component ratio) but by time as well.

Full text of DOI:10.1016/j.steroids.2012.04.008

Steroids 77 (2012) 858–863
Contents lists available at SciVerse ScienceDirect
Steroids
journal homepage: www.elsevier.com/locate/steroids
Synthesis of cholic acid based calixpyrroles and porphyrins
a
b
a
a
b,
⇑
ˇ
ˇ
ˇ
Thu Huong Nguyen Thi , Lenka Cardová , Michaela Dvoráková , Dagmar Rocková , Pavel Drašar
a
ˇ
ˇ
Department of Chemistry, Faculty of Natural Sciences, J.E. Purkyne University Ústí nad Labem, Ceské mládeze 8, 400 96 Ústí nad Labem, Czech Republic
b Department of Chemistry of Natural Compounds, Institute of Chemical Technology Prague, Technická 5, 166 28 Praha 6, Czech Republic
a r t i c l e i n f o
a b s t r a c t
Article history:
New cholic acid based calix[4]pyrroles and porphyrins were prepared and their properties were studied.
It was confirmed by spectral measurements that the superassembly of 5,15-bis(3 ,7 ,12 -trihydroxy-
5b-cholan-24-yl)-10,20-diphenylporphyrin, the best candidate for this study from the conjugates pre-
pared, may be influenced not only by the solvent mixture composition (polar/non-polar component ratio)
but by time as well.
Received 22 February 2012
Received in revised form 28 March 2012
Accepted 7 April 2012
a
a
a
Available online 21 April 2012
Ó 2012 Elsevier Inc. All rights reserved.
Keywords:
Oligopyrrole macrocycle
Steroidal heterocycle
Chiral receptor
Microwave synthesis
Calix[4]pyrrole
Supramolecular synthon
1. Introduction
with detection by spraying with a solution of 5 g of Ce(SO4)2(H2O)4
in 500 ml 10% H2SO4 and subsequent heating. Flash column chro-
The steroid nucleus has been quite widely adopted as a building
block for supramolecular chemistry. It is large, rigid, chiral, and
thus suitable for creating extended architectures with well-defined
conformations, capable in principle of enantiodiscrimination. Of
the available steroidal starting materials, the most versatile bile
acids are especially useful, providing inexpensive starting points
with helpful substitution patterns, arising from both region- and
stereo-chemical variation, and are thus well adapted to serve as
a scaffold for functional group arrays [1] and functional devices
and constructs [2–4]. Hence it is still very actual to search for
new types of these compounds.
matography was performed on silica gel (Merck, 100–160
with solvents distilled prior to use.
As the source of microwave (MW) irradiation a microwave
microprocessor controlled chemical reactor Plasmatronika M REOS
with electromagnetic stirring, MW generator of 2.45G and temper-
ature regulation was used.
lm)
FTIR spectrometer Nicolet iS10 (Thermo Scientific), automatic
polarimeter Autopol VI (Rudolph Research Analytical), UV/VIS Var-
ian Cary 50 UV–Vis, and Cary Eclipse fluorescence spectrophotom-
eters were used for physicochemical characterisations. IR spectra
(m .
) are given in cmꢀ1
In this work we are further extending our studies [5–12] to
pyrrole-bile acid conjugates, because calix[4]pyrroles and porphy-
rins represent a class of macrocycles, which are very important
structural elements in host guest chemistry, since calix[4]pyrroles
bind anions and neutral species and porphyrins bind cations in an
efficient way [13] and some of them may be utilized as electro-
chemical sensing devices [14].
All NMR experiments were run with Varian Gemini 300 HC,
working at 299.97 M for protons, 75.44 M for carbon-13. 1H and
13C NMR chemical shifts were referenced to the signal of internal
standard TMS, in chloroform-d solution, and are given in d ppm,
unless stated otherwise. Coupling constants J are given in Hz.
Mass spectrometric measurements were performed using Agi-
lent LC/MSD SL/ESI with hyphenated HPLC and LCQ Fleet Ion Trap
LC/MSn mass spectrometer with electrospray resp. chemical ioniza-
tion (ESI resp. APCI) or using Q TOF (Micromass) spectrometer with
direct inlet (ESI), on a ZAB-EQ (VG Analytical) instrument (FAB) with
Xe ionization, or Autospec Ultima (Micromass) by EI technique.
2. Experimental
2.1. Materials and methods
TLC was performed on either HF254 plates (Merck) with detec-
tion by UV light or on plates made from MP Biomedicals SilicaGel G
2.2. Chemical synthesis
2.2.1. Esterification of cholic acid 1 under microwave irradiation
A mixture of cholic acid (2.01 g, 4.9 mmol) and p-toluene
sulphonyl acid monohydrate (500 mg, 2.63 mmol) in methanol
⇑
Corresponding author. Tel.: +420 220 443 698.
E-mail address: Pavel.Drasar@vscht.cz (P. Drašar).
0039-128X/$ - see front matter Ó 2012 Elsevier Inc. All rights reserved.
http://dx.doi.org/10.1016/j.steroids.2012.04.008

T.H. Nguyen Thi et al. / Steroids 77 (2012) 858–863
859
(50 ml) was irradiated by MW for 6 min at 50% of maximum inten-
sity (i.e. at 375 W). After evaporation of methanol, water was
slowly added until the product starts to crystallize, and the crude
product was filtered and washed with water. Recrystallization
from methanol–water yielded 2.07 g (99%) of methyl cholate, m.p
107–109 °C. Literature [15] gives different value of m.p, 156–
157 °C.
evaporated. The crude product was purified by chromatography
(silica gel, 32–63 lM, elution by cyclohexane – ether 9:1) to yield
198 mg calix[4]pyrrole 3 (51%) as amorphous solid. ½a D20
ꢁ
–7.6
(c = 1.05 ꢂ 10ꢀ3 g/ml in CHCl3). 1H NMR: 0.73 (s, 3 H, 18-CH3),
0.84 (d, 3 H, 21-CH3), 0.86 (s, 3 H, 19-CH3), 1.49 (bs, 6 CH3
calix[4]pyrrole), 3.66 (s, 3 H, 24-COOCH3), 5.04 (bs, 1 H, 7b-H),
5.26 (bs, 1 H, 12b-H), 5.89 (m, calix[4]pyrrole), 6.86 (bs, 1 H,
NH), 6.91 (bs, 1 H, NH), 7.04 bs, 2 H, 2 NH), 8.11 (s, 1 H, 7
8.16 (s, 1 H, 12 -OOCH). IR (BaCl2) 3416, 3107 (N–H), 2952 as(CH3),
2928 as(CH2), 2872 (CH3), 1720 (C@O), 1177 (C–O). MS (ES-API):
a-OOCH),
2.2.2. Methyl-7
a
,12
a
-diformyloxy-3
a
-hydroxy-5b-cholan-24-oate
a
A solution of methyl cholate (2.07 g, 4.9 mmol) in formic acid
(5 ml) was standing at room temperature for three days. Then ice
was added to the mixture, the white crystallized product was
filtered and washed with water, yield 2.37 g (96%) of methyl
tri-O-formylcholate.
A mixture of methyl tri-O-formylcholate (2.37 g, 4.68 mmol)
and p-toluene sulphonyl acid monohydrate (890 mg, 4.68 mmol)
in propan-2-ol (30 ml) was irradiated by MW for 8 min at 50% of
maximum intensity (i.e. at 375 W). The residue after evaporation
of propan-2-ol was dissolved in ether (25 ml) and Na2CO3 was
added. Solids were filtered off, the filtrate was evaporated and
the crude product was purified by chromatography on a column
of silica gel (45 g) in cyclohexane: diethyl ether 1:1 and then
s
C52H70N4O6 (847.14), calculated monoisotopic m/z: 846.52 Da,
found: 864.2.
Calix[4]pyrrole-cholic acid conjugate 4: A mixture of 3 (0.092 g,
0.109 mmol) and NaOH (0.024 g, 0.600 mmol) in methanol
(10 ml) was stirred in room temperature over night, then NH4Cl
was added, methanol was evaporated, and the crude product was
purified by flash chromatography, elution by diethyl ether, yield
0.035 g (42%) as amorphous solid. ½a D20
ꢁ
ꢀ8.1 (c = 2.35 ꢂ 10ꢀ3 g/ml
in CHCl3). 1H NMR: 0.73 (s, 3 H, 18-CH3), 0.84 (d, 3 H, 21-CH3),
0.86 (d, 3 H, 19-CH3), 1.67 (bs, 6 CH3 calix[4]pyrrole), 3.62 (s, 3
H, 24-COOCH3), 4.05 (bs, 1 H, 7b-H), 4.22 (bs, 1 H, 12b-H), 5.15
(m, calix[4]pyrrole), 5.25 (m, calix[4]pyrrole), 6.05 (m, calix[4]pyr-
role), 8.00 (bs, 1 H, NH), 8.45 (bs, 1 H, NH), 9.39 (bd, 2 H, 2 NH). IR
(BaCl2) 3390(OH, NH), 2952 as(CH3), 2928 as(CH2), 2872 s(CH3),
1720 (C@O). MS (ES-API): C50H70N4O4 (791.12) calculated monoi-
sotopic m/z: 790.53 (808.57) Da, found: M+NH4 808.4.
diethyl ether, yield 1.46 g (65%) of methyl-7a,12a-diformyloxy-
3a-hydroxy-5b-cholan-24-oate, m.p 59–62 °C. The product was
characterized by ½a D20
ꢁ
ꢀ43.5 (c = 8.78 ꢂ 10ꢀ3 g/ml in CHCl3). 1H
NMR: 0.76 (s, 3 H, 19-CH3), 0.84 (d, J = 6.4, 3 H, 21-CH3), 0.93 (s,
3 H, 18-CH3), 3.51 (quin, J = 4.3, 1 H, 3b-H), 3.66 (s, 3 H, 24-
COOCH3), 5.07 (d, J = 2.6, 1 H, 7b-H), 5.27 (br. s, 1 H, 12b-H), 8.15
2.2.5. Methyl 3a,12a-diformyloxy-7-oxo-5b-cholan-24-oate 5
(s, 1 H, 7
a
-OOCH) 8.11 (s, 1 H, 12
a
-OOCH). IR (BaCl2) 3500 (OH),
Fresh distilled pyridine (10 ml, 124 mmol) was added to the
mixture of dry chromium (VI) oxide (2.84 g, 28.4 mmol) and
dichloromethane (30 ml) at 0 °C, and the mixture was stirred until
a suspension was formed. Then methyl cholate (1.96 g, 4.64 mmol)
dissolved in dichloromethane (15 ml) was added. The reaction
mixture was stirred 40 min at 0 °C, then the mixture was filtered
through a layer of alumina (30 g) and the filtrate was evaporated
and co-evaporated with toluene (3 ꢂ 10 ml).
2952, 2928, 2872 (CH2, CH3), 1720 (C@O), 1177 (C–O). MS (ES-
API): for C27H42O7 (478.62), calculated monoisotopic m/z: 478.29
(496.33) Da, found: M+NH4 496.2. It may be noted that biological
data of methyl-7
oate in inflammation were described elsewhere [16].
a,12a-diformyloxy-3a-hydroxy-5b-cholan-24-
2.2.3. Methyl 7a,12a-diformyloxy-3-oxo-5b-cholan-24-oate 2
Fresh distilled pyridine (4.7 ml, 59 mmol) was added to the
mixture of dry chromium (VI) oxide (1.96 g, 19.6 mmol) and dichlo-
romethane (10 ml) at 0 °C, and the mixture was stirred until a
The crude product was dissolved in 5 ml of formic acid, the
reaction mixture was standing at room temperature for three days.
Then ice was added to the mixture, organic product was extracted
to ether, organic solution was washed with concentrated solution
of Na2CO3, water and concentrated solution of NaCl, and dried with
MgSO4 over night. Next day the solution was filtered, ether was
distilled off. Crystallization from ether yielded 1.1 g (49%) of 5,
suspension was formed. Then methyl 7a,12a-diformyloxy-3-hy-
droxy-5b-cholan-24-oate (1.56 g, 3.2 mmol) dissolved in dichloro-
methane (15 ml) was added. The mixture was irradiated by MW
for 6 min at 50% of maximum intensity (i.e. at 375 W). Dichloro-
methane was distilled off and the residue was diluted by diethyl
ether (20 ml), filtered through a layer of alumina (30 g) and the
filtrate was evaporated and co-evaporated with toluene (3 ꢂ 10
ml). Recrystallization from diethyl ether yielded 1.48 g (95%) of
m.p 127–128 °C. ½a D20
ꢁ
ꢀ44.0 (c = 9.50 g/ml in CHCl3). 1H NMR:
0.75 (s, 3 H, 18-CH3), 0.84 (d, J = 6.1, 3 H, 21-CH3), 1.21 (s, 3 H,
19-CH3), 3.66 (s, 3 H, 24-COOCH3), 4.81 (tt, J = 10.1, 5.2, 1 H, 3b-
H), 5.27 (br. s, 1 H, 12b-H), 7.98 (s, 1 H, 3
a-OOCH), 8.11 (s, 1 H,
methyl 7
a
,12
a
-diformyloxy-3-oxo-5b-cholan-24-oate 2, m.p 123–
12 -OOCH). IR (BaCl2) 2951, 2928, 2875 (CH2, CH3), 1720 (C@O),
1177 (C–O). MS (ES-API): C27H40O7 (476.60) calculated monoiso-
topic m/z: 476.27 (494.31) Da, found M+NH4 494.4.
a
124.5 °C. Product 2 was characterized by ½a D20
ꢁ
ꢀ51.1 (c = 7.10
ꢂ 10ꢀ3 g/ml in CHCl3). 1H NMR: 0.78 (s, 3 H, 18-CH3), 0.84 (d,
J = 6.4, 3 H, 21-CH3), 1.03 (s, 3 H, 19-CH3), 3.65 (s, 3 H, 24-COOCH3),
5.14 (d, J = 2.6, 1 H, 7b-H), 5.30 (br. s, 1 H, 12b-H), 8.08 (s, 1 H, 7
a
-
2.2.6. Porphyrin-cholic acid conjugates
OOCH), 8.15 (s, 1 H, 12 -OOCH). IR (BaCl2) 2952, 2928, 2872 (CH2,
a
3a,7a,12a-Trihydroxy-24,24-di(pyrrol-2-yl)-5b-chol-23-ene 7: To
CH3), 1720 (C@O), 1177 (C–O). MS (ES-API): for C27H40O7 (476.60),
calculated monoisotopic m/z: 476.27 (494.31) Da, found: M+NH4
494.0.
a solution of pyrrolyl- magnesium bromide (851 mg, 5 mmol) in
dichloromethane (55 ml) was added methyl cholate (2.00 g,
4.74 mmol) and the reaction mixture was refluxed in an oil bath
in 75 °C with stirring for 60 h. After cooling, the complex was
decomposed by pouring its solution into a mixture of about
100 g of ice and 200 ml of 10% NH4Cl solution. After thorough shak-
ing, the layers are separated, and the aqueous layer was extracted
twice with ether. The combined ether solution was washed with
water, 5% Na2CO3 solution, finally with concentrated NaCl solution,
and organic solution was dried with MgSO4 over night. The solvent
was evaporated, the residue of crude product was purified by chro-
2.2.4. Calix[4]pyrrole-cholic acid conjugates
Calix[4]pyrrole-cholic acid conjugate 3: A mixture of methyl 7
-diformyloxy-3-oxo-5b-cholan-24-oate (220 mg, 0.46 mmol),
5,5-dimethyldipyrromethane (350 mg, 2,0 mmol) and trifluoroace-
tic acid (16 l, 0.2 mmol) in dichloromethane (30 ml) under argon
a,
12a
l
was strongly stirred in room temperature for 4 h. Then dried ace-
tone (20 ml) was added, and the mixture was stirred for further
1 h, and the reaction was ended by adding Na2CO3. The mixture
was filtered through a layer of silica gel, and the filtrate was
matography (silica gel, 32–63 lM, toluene and ether), yield 1.11 g
7 (46%) as amorphous solid. ½a D20
ꢁ
ꢀ12.7 (c = 2.13 ꢂ 10ꢀ3 g/ml in

860
T.H. Nguyen Thi et al. / Steroids 77 (2012) 858–863
(632.83) calculated monoisotopic m/z: 632.38 (650.41) Da, found:
M+NH4 650.
5,15-bis(3a,7a,12a-Trihydroxy-5b-cholan-24-yl)-10,20-diphenyl
porphyrin 9: A mixture of 7 (279 mg, 0.55 mmol), benzaldehyde
(140 l, 1.38 mmol) and trifluoroacetic acid (5.6 l, 0.073 mmol)
l
l
in dichloromethane (140 ml) under argon was strongly stirred in
room temperature over night. Then DDQ (140 mg, 0.62 mmol)
was added, and the mixture was stirred for further 2 hours, and
the reaction was ended by adding Na2CO3. The mixture was filtered
through a layer of silica gel, and the filtrate was evaporated. The
crude product was purified by chromatography (silica gel, 32-
63 lM, elution by toluene and toluene-acetone 1:1), yield 15 mg
Fig. 1. Spatial representation of
optimization (Cambridge Software, Chem Office, ver. 2008).
9 after molecular mechanics (MM2)geometry
of 9 (2%) as amorphous solid. ½a D20
ꢁ
ꢀ37.2 (c = 1.3 ꢂ 10ꢀ4 g/ml in
CHCl3). 1H NMR: ꢀ2.16 (bs, 2 H. NH), 0.97 (s, 6 H, 18-CH3), 1.06
(s, 6 H, 19-CH3), 1.51(d, 6 H, 21-CH3), 3.78 (m, 2 H, 3b-H), 4.15
(m, 2 H, 7b-H), 4.49 (m, 2 H, 12b-H), 5.05 (m, 4 H, 23-CH2), 7.86
(m, 6 H, phenyl), 8.35 (m, 4 H, phenyl), 9.03 (d, 4 H, porphyrin),
9.65 (d, 4 H, porphyrin). IR (BaCl2) 3390 (O-H, N-H), 2931, 2872
CHCl3). 1H NMR d ppm 0.66 (s, 3 H, 18-CH3), 0.86 (d, 3 H, 19-CH3),
0.97 (d, 3 H, 21-CH3), 3.40 (m, 1 H, 3b-H), 3.63 (bs, 1 H, 7b-H), 3.91
´
(t, 1 H, 23-H), 3.95 (bs, 1 H, 12b-H), 6.01 (bs, 2 H, 4-H-pyrrole), 6.12
e
= 1,3.103), kmin = 435 nm
´
´
(bs, 2 H, 3-H-pyrrole), 6.61 (bs, 2 H, 5-H-pyrrole), 8.15 (bs, 1 H, NH-
pyrrole), 8.24 (bs, 1 H, NH-pyrrole). IR (BaCl2) 3360 (O–H, N–H),
(CH2, CH3). UV (DMSO) kmax = 415 nm (
= 0.2.103). MS (ES-API): C78H98N4O6 (1187.64) calculated mono-
isotopic m/z: 1186.74 Da, found: (M + H) 1187.
5,15-bis(3 ,7 ,12 -Triacetoxy-5b-cholan-24-yl)-10,20-diphenylpor
phyrin 10: Preparation was the same as for compound 9 from 8
(323 mg, 0.51 mmol), benzaldehyde (130 l, 1.28 mmol) and triflu-
oroacetic acid (5.2 l, 0.067 mmol) in dichloromethane (130 ml),
and DDQ (130 mg, 0.57 mmol). The crude product was purified
by chromatography (silica gel, 32–63 M, elution by toluene and
(e
2931, 2872 (CH2, CH3). UV (CH3OH) kmax = 237 nm (e
= 3,6.103),
kmin = 333 nm (
e
= 0.88 ꢂ 103). MS (ES-API): C32H46N2O3 (506.72)
a
a
a
calculated monoisotopic m/z: 506.35 (524.38) Da, found M+NH4
524.3.
l
3a
,7a
,12a-Triacetoxy-24,24-di(pyrrol-2-yl)-5b-chol-23-ene 8:
A
l
solution of crude 3
a
,7 ,12 -trihydroxy-24,24-di(pyrrol-2-yl)-5b-
a
a
chol-23-ene 7 (2.36 g, 4.66 mmol) which was prepared similarly
above, was filtered through a layer of silica gel, the filtrate was
evaporated, and the crude product was acetylated with Ac2O
(5 ml, 50 mmol), DMAP (600 mg, 5 mmol) and pyridine (8.1 ml,
100 mmol) by microwave irradiation for 6 min at 50% of maximum
intensity (i.e. at 375 W). After cooling, the reaction mixture was
worked up normally and the product was purified by chromatogra-
l
toluene-ether 9:1), yield 15 mg of 10 (2%) as amorphous solid.
½
a 2D0
ꢁ
ꢀ40.0 (c = 2.75 ꢂ 10ꢀ3 g/ml in CHCl3). 1H NMR: ꢀ2.68 (bs, 2
H. NH), 0.87 (s, 6 H, 18-CH3), 0.96 (s, 6 H, 19-CH3), 1.48(d, 6 H,
21-CH3), 2.05 (s, 3 H, OOCCH3), 2.07 (s, 3 H, OOCCH3), 2.22 (s, 3
H, OOCCH3), 4.59 (m, 2 H, 3b-H), 4.91 (m, 2 H, 7b-H), 5.31 (m, 2
H, 12b-H), 4.74–4.84 a 5.00–5.01 (m, 4 H, 23-CH2), 7.78 (m, 6 H,
phenyl), 8.20 (m, 4 H, phenyl), 8.88 (d, 4 H, porphyrin), 9.39 (d, 4
H, porphyrin). 13C NMR (d ppm) 12.64 (C18), 18.96 (C21), 21.76,
21.87 (3,7,12-OAc), 22.85 (C19), 23.20 (C15), 25.0 (C11),
27.20(C16), 29.17 (C2), 30.5 (C23), 31.48 (C6), 32.99 (C9), 34.30
(C10), 34.61 (C4) 35.20 (C1), 36.99 (C8), 37.0 (C20), 38.01 (C22),
41.16 (C5), 43.67 (C14), 45.50 (C13), 47.81 (C17), 70.96 (C7),
74.34 (C3, C12), 111.36, 111.42, 118.5, 119.0 (porphyrin), 120.07,
126.82, 134.62, 142.73 (phenyl), 136.22 (C24), 170.67, 170.83,
170.99 (C@O-AC). IR (BaCl2) 3316 (O–H, N–H), 2931, 2872 (CH2,
phy (silica gel, 32-63 lM, toluene and ether), yield 1.88 g 8 (62%)
as amorphous solid. ½a D20
ꢁ
ꢀ35.1 (c = 2.88 ꢂ 10ꢀ3 g/ml in CHCl3).
1H NMR d ppm 0.71 (s, 3 H, 18-CH3), 0.83 (d, J = 6.4, 3 H, 21-
CH3), 0.92 (s, 3 H, 19-CH3), 2.06 (s, 3 H, OOCCH3), 2.09 (s, 3 H,
OOCCH3), 2.13 (s, 3 H, OOCCH3), 3.92 (t, J = 7.7, 1 H, 22-H), 4.52–
4.64 (m, 1 H, 3b-H), 4.90 (q, J = 3.0, 1 H, 7b-H), 5.09 (t, J = 2.9, 1
´
H, 12b-H), 6.03–6.08 (m, 2 H, 4-H-pyrrole), 6.15 (quin, J = 2.9, 2
´
´
H, 3-H-pyrrole), 6.62–6.69 (m, 2 H, 5-H-pyrrole), 7.83 (d, J = 8.4,
H, NH-pyrrole). IR (BaCl2) 3272 (N–H), 2931, 2872 (CH2,
CH3),1720 (C@O), 1250 (C–O). UV (CH3OH) kmax = 238 nm (
2
e
=
CH3), 1720 (C@O), 1249 (C–O). UV (DMSO) kmax = 414 nm (
e = 2,1.
103), kmin = 431 nm = 0.2.103). MS (ES-API): C90H110N4O12
(
e
4,1.103), kmin = 438 nm
(
e
= 0,4.103).MS (ES-API): C38H52N2O6
O
O
OH
1) MeOH, PTS, MW, 6 min
2) HCOOH, RT, 3 d
H
H
O
OH
3) i-PrOH, MW, 8 min
O
O
4) CrO3.Py/CH2Cl2, MW, 6 min
O
O
O
OH
HO
H
H
1
2
R
H
O
H
N
N
H
O
R
H
NH
NH
5)
, TFA, rt, 4 h
HN
H
N
R: OOCH
R: OH
6) acetone, TFA, rt, 1 h
3
4
Scheme 1. Synthesis of 3 and 4.

T.H. Nguyen Thi et al. / Steroids 77 (2012) 858–863
861
O
O
OH
O
1) MeOH, PTS, MW, 8 min
2) CrO3.Py/CH2Cl2, 0°C, 60 min
H
H
OH
O
3) HCOOH, RT, 3 d
O
O
O
O
OH
HO
H
H
1
5
OCHO
H
OMe
H
N
O
OHCO
NH
H
5)
, TFA, rt, 4 h
N
H
NH
6) acetone, TFA, rt, 1 h
HN
H
N
6
Scheme 2.
OH
R
1. C4H4NMgBr/Et2O
H
H
OMe
N
H
NH
O
OH
H
R
HO
R
H
R:OH
R:OOCCH
7
8
Ac2O, DMAP/Py
3
R
1. H5C6CHO, TFA, CH2Cl2
2. DDQ
H
N
H
H
N
R
R
R
R
N
H
H
N
H
R
R:OH
R:OOCCH
9
10
3
Scheme 3. Synthesis of 9 and 10.
(1439.86) calculated monoisotopic m/z: 1438.81 Da, found: M+
1438.
out with CrO3.pyridine in dicholoromethane under microwave irra-
diation again, product 2 was obtained in overall yield 59%.
Condensation of ketone 2 with dipyrromethane and then with
acetone presented two-steps-in-one-pot synthesis of mix spiroan-
elated calix[4]pyrroles. The first step was condensation of ketone 2
with dipyrromethane catalyzed by trifluoroacetic acid under argon,
quantitative consumption of ketone 2 was observed after stirring
the mixture for 4 hours at room temperature. The second step
was cyclization of the adduct from the previous step with large ex-
cess of acetone lead to better yield of the steroid spiroanelated
calix[4]pyrrole 3. Hydrolysis of 3 in methanol catalyzed by NaOH
at room temperature afforded calix[4]pyrrole 4.
3. Results and discussion
Cholic acid based calix[4]pyrroles 3 and 4 were synthesized by
acid-catalyzed condensation of keto derivative 2 with 2,2’-pro-
pane-2,2-diylbis(1H-pyrrole) (dipyrromethane) [17,18] and ace-
tone (Scheme 1). The reactivity of the hydroxy groups towards
oxidation was 7-OH > 12-OH > 3-OH and the order of acetylation
or hydrolysis was 3-OH > 7-OH > 12-OH [19], therefore it was
necessary to protect carboxyl group and 7
a
-OH, 12
a
-OH groups be-
Similar condensation of ketone 5 did not afford any calix[4]pyr-
role product (Scheme 2).
fore oxidation of 3 -OH group. Carboxylic group was protected by
a
esterification in methanol under microwave irradiation, catalyzed
by p-toluene sulphonyl acid. All OH-groups were formylated by a
treatment of formic acid at room temperature for three days. Then
the formyloxy group from position 3 was removed by selective
deformylation in propan-2-ol, again, under microwave irradiation.
Under these conditions, propan-2-ol serves both as the reactant in
excess and as the solvent for the reaction. Oxidation of methyl-
The cholic acid based porphyrin 9 was synthesized as follows.
Methyl cholate 1 was heated with pyrrolyl magnesiumbromide
[20] in ether for 60 h to give derivative 7. After chromatographic
separation, pure 7 was subjected to condensation with benzalde-
hyde in a large amount of dichloromethane. Large dilution was
used to support formation of cyclic product. After stirring the mix-
ture over night at room temperature, the reaction mixture was
then subjected to oxidation with 2,3-dichloro-5,6-dicyano-1,4-
7a,12a-diformyloxy-3a-hydroxy-5b-cholan-24-oate was carried

862
T.H. Nguyen Thi et al. / Steroids 77 (2012) 858–863
benzoquinone (DDQ). The porphyrin 9 was separated by chroma-
tography using dichloromethane/acetone mixture as eluent. Rela-
tively low yield of the porphyrin (similarily as 10) is in porphyrin
chemistry rather normal.
resonant fluorescence with narrow band are generally termed J
aggregates (after E.E. Jelley, the inventor). Aggregates with absorp-
tion bands shifted to shorter wavelength are called H-aggregates
and exhibit in most cases low or no fluorescence [22].
Dipyrrole 8 obtained from acetylation of crude 7 was subjected
to condensation with benzaldehyde, consequently to oxidation
with DDQ to yield porphyrin 10 (Scheme 3).
Molecular aggregates are investigated nowadays because of
their unique properties and possible technological applications.
Highly ordered J- and H- aggregates are interesting in a field of
study of electronic and spectroscopic properties [21], i.a. Aggre-
gates which show a narrow absorption band that is shifted to a
longer wavelength (with respect to the monomer) and a nearly
Aggregation experiments were carried out according to known
procedure [23]. Only compound 9 (Fig. 1) showed interesting
superassembly behavior under conditions used. Hence, using of
DMSO/water solution the aggregation of amphiphilic porphyrin
derivative was achieved. In parallel with our other studies [8] we
ascribe it mainly to the solvent system composition promoted
(accelerated) superassembly, having in mind that more mecha-
nisms may be involved. At first 1 lM solution of compound 9
was prepared, after that several solutions diluted by distilled water
UV A
UV B
0.35
0.35
9 9/1
9 7/3
9 5/5
9
9 9/1
9 7/3
0.3
0.3
9 3/7
9 5/5
9 3/7
9 1/9
9 1/9
0.25
0.25
0.2
0.2
0.15
0.1
0.15
0.1
0.05
0
0.05
0
380
400
420
nm
440
460
380
400
420
440
460
nm
Graph 1. UV Spectra of 9 measured without delay.
Graph 2. UV Spectra of 9 after 2 days.
Fluorescence A
Fluorescence B
700
600
500
400
300
200
100
0
700
600
500
400
300
200
100
0
9
9
9 9/1
9 7/3
9 5/5
9 3/7
9 1/9
9 9/1 b
9 7/3 b
9 5/5 b
9 3/7 b
9 1/9 b
600
650
700
nm
750
800
600
650
700
nm
750
800
Graph 3. Fluorescence Spectra of 9 measured without delay.
Graph 4. Fluorescence Spectra of 9 after 2 days.

T.H. Nguyen Thi et al. / Steroids 77 (2012) 858–863
863
ˇ
[7] Dukh M, Šaman D, Lang K, Pouzar V, Cerny´ I, Drašar P, et al. Steroid-porphyrin
conjugate for saccharide sensing in protic media. Org Biomol Chem
2003;1:3458–63.
(in ratio 9/1, 3/7, 5/5, 3/7, 1/9) were measured while preserving
M concentration of solute in the solution. This composition in-
1
l
duces efficient porphyrin self-aggregation process to form J-type
aggregates, observed by the broadening and bathochromic shift
of the Soret band (Graph 1). These samples were measured after
two days for comparison (Graph 2) and time dependence was ob-
served. During the first set of measurements (A – immediately
after addition of water) bathochromic shift was observed at sample
5/5 for the first time while after two days this shift is possible to
observe already at 7/3 solution. It means that aggregation of com-
pound 9 is promoted not only by DMSO/water ratio but by time as
well. To confirm these results fluorescence spectra were measured
under the same conditions (Graphs 3 and 4) and change in a inten-
sity is in agreement with previous findings. Excitation wavelength
was 420 nm to confirm the UV absorption of Soret band.
[8] Zelenka K, Trnka T, Tišlerová I, Monti D, Cinti S, Naitana ML, et al.
Spectroscopic, morphological, and mechanistic investigation of the solvent-
promoted aggregation of porphyrins modified in meso-positions by
glucosylated steroids. Chem Eur J 2011;17:13743–53.
ˇ
[9] Dukh M, Drašar P, Cerny´ I, Pouzar V, Shriver JA, Král V, et al. Novel deep cavity
calix[4]pyrroles derived from steroidal ketones. Supramol Chem
2002;14:237–44.
ˇ
[10] Monti D, Venanzi M, Gatto E, Mancini G, Sorrenti A, Štepánek P, et al. Study of
the supramolecular chiral assembly of meso-‘‘C-glucoside’’-porphyrin
derivatives in aqueous media. New J Chem 2008;32:2127–33.
ˇ
ˇ
[11] Štepánek P, Dukh M, Šaman D, Moravcová J, Kniezo L, Monti D, et al. Synthesis
and solvent driven self-aggregation studies of meso-‘‘C-glycoside’’-porphyrin
derivatives. Org Biomol Chem 2007;5:960–70.
[12] Zelenka K, Trnka T, Tišlerová I, Král V, Dukh M, Drašar P. Synthesis of porphyrin
receptors modified by glycosylated steroids. Coll Czech Chem Commun
2004;69:1149–60.
[13] Radha Kishan M, Radha Rani V, Murty MRVS, Sita Devi P, Kulkarni SJ, Raghavan
KV. Synthesis of calixpyrroles and porphyrins over molecular sieve catalysts. J
Mol Catal A: Chem 2004;223:263–7.
[14] Rumlerová-Lipšová A, Barek J, Drašar P, Zelenka K, Pecková K. Polarographic
Acknowledgments
and
voltammetric
determination
of
meso-tetrakis(4-
sulfonatophenyl)porphyrin tetrasodium salt at mercury electrodes. Int
Electrochem Sci 2007;2:235–47.
[15] Fieser LF, Rajagopalan S. Oxidation of steroids. III. Selective oxidations and
acylations in the bile acid series. J Am Chem Soc 1950;72:5530–6.
J
This work was supported by the Ministry of Education, Youth
and Sports of the Czech Republic in a project No. MSM6046
137305 and by the Czech Science Foundation (Grant Agency of the
CR), in projects No. 304/10/1951 and P503/11/0616.
[16] Yasukawa K, Iida T, Fujimoto Y. Relative inhibitory activity of bile acids against
12-O-tetradecanoylphorbol-13-acetate-induced
inflammation,
and
chenodeoxycholic acid inhibition of tumour promotion in mouse skin two-
stage carcinogenesis. J Pharm Pharmacol 2009;61:1051–6.
[17] Journot G, Neier R, Stoeckli-Evans H. 2,2’-(Propane-2,2-diyl)bis(1H-pyrrole).
Acta Cryst 2010;E66:o392.
References
[18] Brown WH, French WN. Synthesis of porphyrinogenlike compounds (mixed
quaterenes). Can J Chem 1958;36:371–7.
[1] Davis AP. Anion binding and transport by steroid-based receptors. Coord Chem
Rev 2006;250:2939–51.
[19] Virtanen E, Kolehmainen E. Use of bile acids in pharmacological and
supramolecular applications. Eur J Org Chem 2004:385–99.
[20] Kutscher W, Klamerth O. Histaminelike compounds of the pyrrole series. II.
Synthesis of 2-(2-aminoethyl)pyrrole. Hoppe-Seyler’s Zeitsch. Physiol Chem
1952;289:229–33.
[21] Yao H, Domoto K, Isohashi T, Kimura K. In situ detection of birefringent
mesoscopic H and J aggregates of thiacarbocyanine dye in solution. Langmuir
2005;21:1067–73.
[22] Würthner F, Kaiser TE, Saha-Möller CR. J-aggregates: from serendipitous
discovery to supramolecular engineering of functional dye materials. Angew
Chem Int Ed 2011;50:3376–410.
[23] Monti D, Venanzi M, Stefanelli M, Sorrenti A, Mancini G, Di Natale C, et al.
Chiral amplification of chiral porphyrin derivatives by templated
heteroaggregation. J Am Chem Soc 2007;129:6688–9.
[2] Girgenti E, Ricoux R, Mahy JP. Design and synthesis of a Mn(III)-porphyrin
steroid conjugate used as a new cleavable affinity label: on the road to semi-
synthetic catalytic antibodies. Tetrahedron 2004;60:10049–58.
[3] Vollmer MS, Effenberger F, Stumpfig T, Hartschuh A, Port H, Wolf HC. Steroid-
bridged anthryloligothienylporphyrins: synthesis and study on the
intramolecular energy transfer. J Org Chem 1998;63:5080–7.
[4] Weiss J. Zinc(II)-porphyrin receptors in multi-point molecular recognition:
recent progress. J Incl Phen Macrocyc Chem 2001;40:1–22.
ˇ
[5] Štepánek P, Šimák O, Nováková Z, Wimmer Z, Drašar P. Asymmetrically
substituted calix[4]pyrrole with chiral substituents. Org Biomol Chem
2011;9:682–3.
[6] Huong NTT, Klímková P, Sorrenti A, Mancini G, Drašar P. Synthesis of
spiroannulated oligopyrrole macrocycles derived from lithocholic acid.
Steroids 2009;74:715–20.