The Solid-Phase Zincke Reaction: Preparation of ω-Hydroxy Pyridinium Salts in the Search for CFTR Activation

Article abstract of DOI:10.1021/jo0001636

A study of structural modifications of MPB-07 was undertaken as part of a synthetic program aimed at discovering small molecules with CFTR activation potential. Solid-phase synthesis techniques were used to prepare derivatives of MPB-07 employing the Zincke reaction for the construction of aromatic, quaternary ammonium salts such as those found in 2 or 3. In this transformation, primary amines react with highly electrophilic N-2,4-dinitrophenylpyridinium (DNP) salt 4 to afford pyridinium salt 8 with release of 2,4-dinitroaniline 6. Thus, the reaction of 1-(2,4-dinitrophenyl)pyridinium salts with various polymer-bound amino ethers, followed by cleavage from the resin, delivers the desired salts in good yield and high purity.

Full text of DOI:10.1021/jo0001636

J . Org. Chem. 2000, 65, 5131-5135
5131
Th e Solid -P h a se Zin ck e Rea ction : P r ep a r a tion of ω-Hyd r oxy
P yr id in iu m Sa lts in th e Sea r ch for CF TR Activa tion
†
,‡
‡
Masahiro Eda, Mark J . Kurth,* and Michael H. Nantz
Department of Chemistry, University of California, One Shields Avenue, Davis, California 95616-5295,
and Welfide Corporation, 2-25-1, Shodai-Ohtani, Hirakata, Osaka, J apan 573-1153
E-mail: mjkurth@ucdavis.edu
Received February 7, 2000
A study of structural modifications of MPB-07 was undertaken as part of a synthetic program
aimed at discovering small molecules with CFTR activation potential. Solid-phase synthesis
techniques were used to prepare derivatives of MPB-07 employing the Zincke reaction for the
construction of aromatic, quaternary ammonium salts such as those found in 2 or 3. In this
transformation, primary amines react with highly electrophilic N-2,4-dinitrophenylpyridinium
(DNP) salt 4 to afford pyridinium salt 8 with release of 2,4-dinitroaniline 6. Thus, the reaction of
1
-(2,4-dinitrophenyl)pyridinium salts with various polymer-bound amino ethers, followed by cleavage
from the resin, delivers the desired salts in good yield and high purity.
In tr od u ction
Cystic fibrosis (CF) is the most common fatal genetic
disease of Caucasians with an incidence of 1 in 2500
newborns.1 In 1989, the product of the gene whose
mutation results in CF was identified as CF transmem-
2
brane conductance regulator (CFTR) which is a mem-
brane protein that functions as a chloride-selective ion
3
channel. Patients with CF ultimately suffer pulmonary
failure and death caused by reduced CFTR function.
F igu r e 1.
Recently, Becq reported that the benzo[c]quinolizinium
The Zincke reaction affords excellent methodology for
the construction of aromatic, quaternary ammonium salts
such as those found in 2 or 3. In this transformation,
primary amines react with highly electrophilic N-2,4-
dinitrophenylpyridinium (DNP) salt 4 to afford pyri-
derivative MPB-07 1 (Figure 1) activates wild-type CFTR
4
in a variety of cell systems. Although the mechanism of
CFTR activation by MPB-07 has not been fully eluci-
dated, this compound class represents a promising new
structural lead for CF treatment.
dinium salt 8 with release of 2,4-dinitroaniline 6 (Scheme
As part of a synthetic program aimed at discovering
small molecules with CFTR activation potential, one of
our approaches was to modify the rigid conformational
aspects of MPB-07 to deliver more flexible structural
types; for example, simple pyridinium alkyl alcohol
derivatives 2 or slightly more complex 1-phenyl pyri-
dinium derivatives 3. Like MPB-07, these compounds
contain a quaternary nitrogen, a lipophilic component,
and a hydroxyl moiety. We report here the use of solid-
phase synthesis techniques to prepare these and other
derivatives of MPB-07.
5
1
). Recent interest in this old reaction, first reported by
Sch em e 1
†
Welfide Corp.
University of California.
‡
(
1) Collins, F. S. Science 1992, 256, 774.
(2) (a) Riordan, J . R.; Rommens, J . M.; Kerem, B. S.; Alon, N.;
Rozmahel, R.; Grzelczak, Z.; Zeilenski, J .; Lok, S.; Plavsic, N.; Chou,
J .; et al. Science 1989, 245, 1066. (b) Kerem, B.; Rommens, J . M.;
Buchanan, J . A.; Markiewicz, D.; Cox, T. K.; Chakravarti, A.; Buch-
wald, M.; Tsui, L. C. Science 1989, 245, 1073.
(
3) Li, M.; MacCann, J . D.; Anderson, M. P.; Clancy, J . P.; Liedtke,
C. M.; Nairn, A. C.; Greengard, P.; Welssch, M. J . Science 1989, 244,
353.
4) Becq, F.; Mettey, Y.; Gray, M. A.; Galietta, L. J .; Dormer, R. L.;
Merten, M.; Metaye, T.; Chappe, V.; Marvingt-Mounir, C.; Zegarra-
Moran, O.; Tarran, R.; Bulteau, L.; D e´ rand, R.; Pereira, M. M. C.;
McPherson, M. A.; Rogier, C.; J offre, M.; Argetn, B. E.; Sarrouilhe,
D.; Kammouni, W.; Figarella, C.; Verrier, B.; Gola, M.; Verfond, J .-M.
J . Biol. Chem. 1999, 39, 27415.
1
6
(
Zincke in 1903, has been focused mainly on mechanistic
7
considerations. In contrast, relatively less attention has
(5) Kost, A. N.; Gromov, S. P.; Sagitullin, R. S. Tetrahedron 1981,
37, 3423.
1
0.1021/jo0001636 CCC: $19.00 © 2000 American Chemical Society
Published on Web 07/22/2000

5
132 J . Org. Chem., Vol. 65, No. 17, 2000
Eda et al.
Sch em e 3
F igu r e 2.
been paid to the synthetic potential of this transformation
because purification of the desired salt 8 from dinitro-
aniline 6 and other reaction intermediates is tedious due
8
to their highly polar character. Furthermore, salt 4, a
convenient preparation of which has been reported by
9
Vompe, is only marginally soluble in organic solvents
and the chloride counteranion is nucleophilic enough to
result in decomposition of this salt under the conditions
of the Zincke reaction.10 Recently, the Marazano group
Sch em e 4
1
0,11
reported a useful approach to solving these problems.
Specifically, they reported the use of a lipophilic dodecyl
sulfate counteranion (cf. 9), instead of chloride, which
minimizes nucleophilic attack on the pyridinium salt and
has improved solubility in organic solvents. With this
modification, Zincke product 10 was prepared in good
yield (Scheme 2). This background information led us to
adapt the Zincke reaction for application in solid-phase
synthesis as outlined in Figure 2 and we report here a
solid-phase route to a small library of pyridinium salts
2
and 3.
Sch em e 2
â-alaninol, valinol, phenylglycinol in the presence of a
catalytic amount of boron trifluoride in the mixed solvent
THF and dichloromethane (1:2) gave 13, whereas reac-
tion with o-nitrobenzyl alcohol gave 14. Removal of the
Fmoc group from 13 gave resin 15, whereas reduction of
the nitro group with tin chloride13 gave 16.
Resu lts a n d Discu ssion
Dodecyl sulfate salts 9a -d were easily prepared from
To optimize the Zincke reaction for solid-phase, we first
carried out the reaction of 9d with 15a (loading ) 0.55
mmol/g based on a 0.60 mmol/g loading for 11a ) in
various solvents (Table 1), including dichloromethane,
tetrahydrofuran, n-butanol, and toluene. A convenient
procedure for the systematic investigation of this reaction
was to estimate the ratio of the desired isoquinolizinium
9
the chloride salts according to the method of Vompe and
Barbier10 (Scheme 3). Thus, treatment of pyridine with
1
4
equiv of 1-chloro-2,4-dinitrobenzene led to chloride salt
. Subsequent reaction of 4 with 1.1 equiv of sodium
dodecyl sulfate in refluxing dichloromethane, followed by
filtration over Celite diatomaceous earth, gave dodecyl
sulfate salt 9 in good yield after recrystallization.
The various polymer-bound ethers 13 and 14 were pre-
pared by modification of a method reported by Hanessian
1
salt 10 and phenylglycinol salt 18 by H NMR analysis
3 3
(300 MHz; CD OD/CDCl , 1:5) of the crude reaction
mixture obtained after cleavage from the resin (TFA
treatment). This NMR analysis was accomplished by
comparing integrations for the hydroxyl-substituted
methylene protons in 10 [4.51 ppm (1H, dd) and 4.37
ppm (1H, dd)] versus the same protons in phenylglycinol
(
Scheme 4).12 Thus, reaction of Wang resin 11a or
NovaSyn TGA resin 11b (polymer-bound poly(ethylene
glycol) hydroxymethylphenyl resin) with trichloroaceto-
nitrile in the presence of DBU gave trichloroacetimidate
resin 12. Reaction of this activated resin with N-(9-
fluorenylmethoxycarbonyl)-protected (Fmoc-protected)
[3.78 ppm (2H, d)]. Likewise, the benzylic methine proton
in 10 [6.12 ppm (1H, dd)] could be compared with the
benzylic methine proton in phenylglycinol [4.24 ppm
(
6) (a) Zincke, Th.; Heuser, G.; Moller, W. J ustus Liebigs Ann. Chem.
(1H, t)]. We discovered that this reaction required a high
1
3
904, 333, 296. (b) Zincke, Th. J ustus Liebigs Ann. Chem. 1903, 330,
61.
reaction temperature (>80 °C) and a solvent capable of
eliciting efficient resin swelling. Among the solvents
investigated, good results were obtained by heating the
reaction mixture in toluene at 80-100 °C. In contrast,
n-butanol was less effective due to its inability to swell
the resin (Table 1, entry 1 vs 2).
(7) (a) Kunugi, S.; Okubo, T.; Ise, N. J . Am. Chem. Soc. 1976, 98,
2
9
5
282. (b) Marvell, E. N.; Caple, G.; Shahidi, I. J . Am. Chem. Soc. 1970,
2, 5641. (c) Marvell, E. N.; Shahidi, I. J . Am. Chem. Soc. 1970, 92,
646.
(
8) Genisson, Y.; Marazano, C.; Mehmandoust, M.; Gnecco, D.; Das,
B. C. Synlett 1992, 431.
9) Vompe, A. F.; Turitsyna, N. F. Dokl. Akad. Nauk SSSR 1949,
4, 341 (Chem. Abstr. 1949, 43, 4671a).
10) Barbier, D.; Marazano, C.; Das, B. C.; Potier, P. J . Org. Chem.
996, 61, 9596.
11) Genisson, Y.; Marazano, C.; Das, B. C. J . Org. Chem. 1993, 58,
052.
12) Hanessian, S.; Xie, F. Tetrahedron Lett. 1998, 39, 733.
(
6
1
2
Varying the number of equivalents of 9d employed in
the reaction with 15a in toluene did not improve the
(
(
(13) Mayer, J . P.; Zhang, J .; Bjergarde, K.; Lenz, D. M.; Gaudino,
J . J . Tetrahedron Lett. 1996, 37, 8081.
(

Zincke Reaction Salt Preparation for CFTR Activation
J . Org. Chem., Vol. 65, No. 17, 2000 5133
Ta ble 1. Op tim iza tion of th e Solid -P h a se Zin ck e
Rea ction
counteranion exchange was carried out by column chro-
matography of the crude dodecyl sulfate salt with reversed-
phase silica gel using MeOH/H
2
O/1N HCl (50:50:1) as the
eluent. Under these conditions, the dodecyl sulfate and
side product contaminants were found to strongly absorb
onto the silica gel, and the desired chloride salt was easily
isolated. However, we could not separate 2e and 2h from
the valinol. Our purified yields, based on initial resin
loading and purity analyzed by reversed-phase HPLC,
of the chloride salts are shown in Table 2.
The Zincke reaction is adversely affected by electron-
donor substituents on the pyridine ring which prevent
5
nucleophilic attack by the primary amine. Vompe re-
ported that heating salt 4b with aniline in ethanol gives
dinitrodiphenylamine which was derived by elimination
of 3-methoxypyridine from 4b by ipso substitution with
aniline.15 Interestingly, under the conditions in our
experiments, salts 3b, 2b, 2f, and 2j were obtained in
very good yield.
Because 1-arylpyridinium salts such as 8 are shown
to be obtained via the intermediacy of glutaconaldehyde
dianil 7 (which requires that the 1°-amine moiety adds
twice), one can heat an alcohol solution of 4 with more
resin
entry (mmol/g)
temp time equiv 10:18
a
b
solvent
(°C)
(h)
9d
ratio
1
2
3
4
5
6
7
8
9
0
1
15a (0.55) n-BuOH
15a (0.55) toluene
15a (0.55) toluene
15a (0.55) toluene
15a (0.55) toluene
15a (1.09) toluene
15b (0.24) toluene
15b (0.24) toluene
100
100
92
90
90
85
73
73
85
85
78
40
40
72
40
40
40
40
40
40
40
40
1
1
1
2
3
3
3
4
2
3
3
56:44
64:36
65:35
65:35
69:31
68:32
32:68
25:75
84:16
>95:<5
65:35
than 2 equiv of arylamine to obtain 8 in one synthetic
step.6
,15
In the case of aliphatic amines, which are
15a (0.55) toluene + Et3N
15a (0.55) toluene + Et3N
15b (0.24) toluene + Et3N
1
1
more reactive than arylamines, only a slight excess of
16
amine is required. The amine moiety in our solid-
a
phase Zincke reaction is attached to the resin, and
Resin loading is based on the initial loading of resin 11a or
1b. b Bath temperature.
17
1
dianil formation thus requires site-site interaction
of two solid-phase 1°-amines. Entries 1-8 in Table 1
all give product 10 via this solid-phase site-site
process. The nearly identical outcome in entries 5 and 6
(0.55 vs 1.09 mmol/g loading; Table 1) suggests that
there is an upper limit to the effectiveness of this site-
site process. Lower resin loading (0.24 mmol/g NovaSyn
TGA resin) diminishes site-site effectiveness and results
in a decreased product-to-starting material ratio (entries
7 and 8; Table 1). Addition of triethylamine (entries 9-11;
Table 1), dramatically improves the product-to-starting
material ratio by mediating the 5 f 8 (Scheme 1)
conversion by a pathway which does not require site-
site interaction. Similar triethylamine benefits were
observed in the reaction of all solid-phase Zincke reac-
tions investigated.
product-to-starting material ratio (10:18; entries 2-5 in
Table 1). Switching from medium-loading Wang resin
(15a , 0.55 mmol/g) to higher-loading Wang resin (1.09
mmol/g) or lower-loading NovaSyn TGA (15b, 0.24 mmol/
g) gave disappointing results (entries 6-8). However, a
significant improvement was noted upon addition of 1
equiv triethylamine to the solid-phase Zincke reaction
(entry 9). Finally, the reaction could be driven to comple-
tion by use of 3 equiv of 9d in the presence of 1 equiv of
triethylamine; under these conditions, NMR analysis of
the crude product after cleavage from the resin showed
no phenylglycinol remained (entry 10). The material
obtained from this reaction protocol was further purified
by flash chromatography on silica gel under conditions
reported by Barbier.10 Isoquinolizinium salt 10 was
obtained in 74% overall isolated yield from 11a .
Con clu sion
We applied this method to other combinations of DNP
salts 9a -d and resin-bound amines 15 or 16 to prepare
a small library of pyridinium derivatives as shown in
Table 2. From the NMR analysis of these crude products,
most of these compounds showed high purity of the
desired salt without starting material contamination.
However, the combination of valinol and pyridine 2e or
isoquinoline 2h resulted in product with 10 and 30%
valinol contamination, respectively, and the combination
of o-aminobenzyl alcohol and isoquinoline 3d could not
be isolated due to low yield.
Thus, while crude product purity was generally excel-
lent and chromatographic purification was not required,
structure-activity relationship studies with MPB-07
indicate that the counteranion affects CFTR activity. A
chloride counteranion shows best potency.14 Therefore,
We have established a convenient method for the
preparation of pyridinium alkyl and aryl alcohol struc-
tural derivatives of MPB-07 using a solid-phase Zincke
reaction. The production of a larger library of these
compounds, as well as their biological evaluation, are
currently under investigation.
Exp er im en ta l Section
Gen er a l P r oced u r es. All chemicals were obtained from
commercial suppliers and used without further purification.
Wang resin (substitution 0.60 and 1.28 mmol/g) and NovaSyn
TG HMP resin (substitution 0.25 mmol/g) were purchased
from Novabiochem. Reaction solvents were distilled from an
appropriate drying agent before use. Analytical TLC was
(15) Grigor'eva, N. E.; Yavlinskii, M. D. Chem. Abstr. 1954, 48,
1
1411a.
(14) Becq, F., Laboratoire de Neurobiologie, UPR-9024 CNRS, 31
(16) Zincke, Th.; Wurker, W. J ustus Liebigs Ann. Chem. 1905, 341,
ch. J . Aiguier, F-13402 Marseille cedex 20, France. Personal com-
munication, 2000.
365.
(17) Hodge, P. Chem. Soc. Rev. 1997, 26, 417.

5
134 J . Org. Chem., Vol. 65, No. 17, 2000
Ta ble 2. A Zin ck e Rea ction Dem on str a tion Libr a r ya
Eda et al.
H
H
Ph
H
OMe
H
2a : 51 (97)
2b: 61 (96)
2c: 43 (96)
2d : 52 (93)
2e: 76c
2f: 80 (97)
2g: 74 (95)
2h : 70d
2i: 61 (98)
2j: 81 (96)
2k : 63 (99)
10: 74e
3a : 89 (99)
3b: 89 (96)
3c: 75 (97)
b
-
CHdCH)2-
3d : -
a
b
Yields of isolated product from initial resin loading with the reverse-phase HPLC purity given in parentheses. Yield was less than
d
e
1
0%. c Contains 10% valinol. Contains 30% valinol. Dodecyl sulfate salt.
carried out on precoated plates (Merck silica gel 60 for normal
phase and C18 silica gel 60 for reversed phase) and visualized
with UV light. Flash column chromatography was performed
with silica (Merck, 70-230 mesh) for normal phase and
Chromatorex ODS (Fuji Silysia Chemical Ltd., 100-200 mesh)
2.7, 8.7 Hz), 9.22 (d, 2 H, J ) 6.9 Hz), 9.27 (d, 1 H, J ) 2.7
Hz). 13C NMR (100 MHz, CD
OD): δ 14.5, 23.7, 26.9, 30.4,
30.5 (2C), 30.7 (2C), 30.8 (2C), 33.1, 69.1, 123.2, 125.8 (2C),
129.8 (2C), 131.1, 131.2 (2C), 132.8, 134.5, 134.8, 140.1, 144.7,
146.9 (2C), 151.1, 160.7.
P olym er -Bou n d 2-Am in o-2-p h en yleth yl Eth er (15a ). To
a suspension of the Wang resin (0.60 mmol/g, 20 g, 12 mmol)
2 2
in dry CH Cl (140 mL) was added trichloroacetonitrile (22.5
mL), and the mixture was cooled to 0 °C. To this suspension
was added DBU (1.5 mL) dropwise over a period of 5 min and
stirred at 0 °C for 40 min. The resin was collected on a glass
2 2 2 2
filter, washed with CH Cl , DMSO, THF, and CH Cl (30 mL
each), and dried under reduced pressure to give a trichloro-
3
1
13
for reversed phase. H and C NMR spectra were recorded in
1
13
CD
NMR), and chemical shifts were reported in parts per million
δ) relative to the solvent peaks (δ 3.30 and 49.0 ppm,
3
OD (300 MHz for H NMR and 75 or 100 MHz for
C
(
respectively). The purity of selected final compounds was
determined by HPLC using Nova-Pak C18 (3.9 × 150 mm,
Waters); linear gradient elution of 20-100% MeOH/water
containing 0.05% TFA for 10 min; flow rate 1 mL/min;
detection, 254 nm.
acetimidate resin 12. The obtained resin 12 (3.25 g, 1.79 mmol)
1
-(2,4-Din itr op h en yl)-3-m eth oxyp yr id in iu m Ch lor id e
was washed well with anhydrous THF (60 mL x 3) under N
atmosphere and suspended in the mixed solvent of CH Cl (40
2
(
4b). A mixture of finely powdered 1-chloro-2,4-dinitrobenzene
2
2
(
95%, 3.00 g, 14.1 mmol) and 3-methoxypyridine (97%, 1.46
mL) and THF (20 mL). To this suspension was added Fmoc-
phenylglycinol (1.29 g, 3.59 mmol), and the suspension was
mL, 14.1 mmol) was heated at 80 °C for 0.25 h. Then, 1 mL of
acetone was added, and the resulting mixture was refluxed
for 2 h. The resulting precipitate was collected, washed with
acetone, and recrystallized from MeOH/AcOEt/hexane to give
stirred for 5 min, followed by addition of BF
3
2
‚OEt (0.16 mL,
1.26 mmol). The resulting mixture was stirred at room
temperature for 1h. The resin was collected on the glass filter,
a product (3.73 g, 85%) as a slightly yellow crystal; mp 174-
2 2 2 2 2 2
washed with THF, CH Cl , THF, CH Cl , THF, and CH Cl
1
1
1
75 °C. H NMR (300 MHz, CD
3
OD): δ 4.15 (s, 3 H), 8.29 (dd,
(40 mL each), and dried under reduced pressure to give a resin
13. To this resin 13 was added 20% piperidine in DMF solution
(30 mL); the mixture was stirred for 20 min and filtered, and
20% piperidine in DMF solution (30 mL) was added. After the
suspension was stirred for an additional 20 min, the resin was
H, J ) 6.0, 9.0 Hz), 8.34 (d, 1 H, J ) 8.7 Hz), 8.54 (ddd, 1 H,
J ) 0.6, 2.4, 9.0 Hz), 8.91 (dd, 1 H, J ) 2.4, 8.7 Hz), 8.94 (brd,
1
2
1
H, J ) 6.0), 9.21 (dd, 1 H, J ) 1.2, 2.4 Hz), 9.25 (d, 1 H, J )
13
.4 Hz). C NMR (100 MHz, CD
31.1, 132.6, 134.6, 135.0, 139.7, 140.1, 144.5, 151.1, 160.4.
-(2,4-Din itr op h en yl)-3-m eth oxyp yr id in iu m Dod ecyl
Su lfa te (9b). A mixture of 4b (2.00 g, 6.42 mmol) and sodium
dodecyl sulfate (2.04 g, 7.06 mmol) in CH Cl (35 mL) was
3
OD): δ 58.6, 123.1, 129.9,
collected on the glass filter, washed with DMF, CH
2 2
Cl , DMF,
1
CH Cl , DMF, and CH Cl (30 mL each), and dried under
2
2
2
2
reduced pressure to give a product.
2
2
P olym er -Bou n d o-Am in oben zyl Eth er (16). The resin
14 was prepared using a method similar to that described for
15a . To this resin (2.25 g, 1.24 mmol) was added a solution of
refluxed for 3 h. The reaction solution was filtered through
Celite diatomaceous earth, and the filtrate was evaporated in
vacuo. The residue was recrystallized from AcOEt/hexane to
2 M SnCl
at room temperature for 5 h. The resin was collected on a glass
filter, washed with DMF, water, DMF, water, DMF, and CH
Cl (30 mL each), and dried under reduced pressure to give a
product.
2
in DMF (23 mL), and the suspension was stirred
give a product (3.03 g, 98%) as a slightly orange crystal; mp
1
9
6
3-94 °C. H NMR (300 MHz, CD
3
OD): δ 0.92 (t, 3 H, J )
2
-
.0 Hz), 1.29 (br, 18 H), 1.64 (m, 2 H), 3.90 (t, 2 H, J ) 6.0
2
Hz), 4.12 (s, 3 H), 8.27 (dd, 1 H, J ) 6.0, 9.0 Hz), 8.28 (d, 1 H,
J ) 8.7 Hz), 8.52 (ddd, 1 H, J ) 0.9, 2.4, 9.0 Hz), 8.87 (m, 1
H), 8.91 (dd, 1 H, J ) 2.4, 8.7 Hz), 9.13 (dd, 1 H, J ) 1.2, 2.4
1-(2-Hyd r oxym eth ylp h en yl)p yr id in iu m Ch lor id e (3a ).
To a suspension of resin 16 (200 mg, 0.11 mmol) in toluene (2
mL) were added 9a (158 mg, 0.34 mmol) and triethylamine
(0.016 mL), and the resulting mixture was stirred at 85 °C for
40 h under nitrogen atmosphere. The resin was collected on a
glass filter and washed with MeOH, DMF, MeOH, DMF,
1
3
Hz), 9.26 (d, 1 H J ) 2.4 Hz). C NMR (100 MHz, CD
δ 14.5, 23.7, 26.8, 30.3-30.7 (7C), 33.0, 58.6, 68.8, 123.1, 129.8,
31.1, 132.6, 134.3, 135.1, 139.7, 140.1, 144.3, 150.9, 160.2.
-(2,4-Din itr oph en yl)-4-ph en ylpyr idin iu m Dodecyl Su l-
fa te (9c) was prepared as a white crystal using a method
3
OD):
1
1
MeOH, CH
was added 10% trifluoroacetic acid in CH
mixture was stirred for 30 min and filtered, and 10% trifluo-
roacetic acid in CH Cl (2 mL) was added again. After the
mixture was stirred an additional 30 min, the suspension was
filtered and the resin was washed with MeOH, CH Cl , MeOH,
2
Cl
2
, MeOH, and CH
2
Cl
2
(4 mL each). To this resin
1
similar to that described for 9b (75%); mp 142-148 °C.
NMR (300 MHz, CD OD): δ 0.89 (t, 3 H, J ) 6.9 Hz), 1.27
br, 18 H), 1.59 (q, 2 H, J ) 6.6 Hz), 3.90 (t, 2 H, J ) 6.6 Hz),
.65-7.75 (m, 3 H), 8.17 (dd, 2 H, J ) 2.1, 8.4 Hz), 8.32 (d, 1
H, J ) 8.7 Hz), 8.68 (d, 2 H, J ) 6.9 Hz), 8.91 (dd, 1 H, J )
H
2
2
Cl (2 mL), the
3
(
7
2
2
2
2

Zincke Reaction Salt Preparation for CFTR Activation
CH Cl , MeOH, and CH Cl (2 mL each). Filtrate and washings
were combined and evaporated in vacuo. The residue was
purified by reversed phase column chromatography (MeOH/
J . Org. Chem., Vol. 65, No. 17, 2000 5135
2
2
2
2
(dd, 1 H, J ) 6.9, 8.1 Hz), 8.31 (d, 1 H, J ) 8.4 Hz), 8.48 (m,
1
3
2 H), 8.68 (d, 1 H, J ) 6.6 Hz), 9.94 (s, 1H). C NMR (100
MHz, CD OD): δ 34.4, 58.9, 60.4, 127.4, 128.4, 129.1, 131.4,
3
H
2
O/1N HCl, 50:50:1) to give a product as a yellow solid (22
132.5, 135.8, 138.2, 138.9, 151.3.
mg, 89%); mp 194-196 °C; 98.5% purity based on HPLC (t
R
)
1-(1-Hyd r oxy-3-m eth ylbu ta n -2-yl)-3-m eth oxyp yr id in -
iu m Ch lor id e (2f) was prepared as a yellow viscous oil using
1
2
7
.78 min). H NMR (300 MHz, CD
3
OD): δ 4.48 (s, 2 H), 7.60-
.72 (m, 4 H), 8.25 (t, 2 H, J ) 6.6 Hz), 8.78 (t, 1 H, J ) 7.8
a method similar to that described for 3a (80%); 96.8% purity
1
3
1
Hz), 9.08 (d, 2 H, J ) 5.7 Hz). C NMR (75 MHz, CD
3
OD): δ
based on HPLC (t
R
) 2.30 min). H NMR (300 MHz, CD
3
OD):
1
1
27.4, 129.1, 130.7, 131.7 (2C), 133.0, 137.1, 147.5 (2C), 148.3,
52.9.
-(2-Hydr oxym eth ylph en yl)-3-m eth oxypyr idin iu m Ch lo-
δ 0.76 (d, 3 H, J ) 6.6 Hz), 1.17 (d, 3 H, J ) 6.6 Hz), 2.46 (m,
1 H), 4.06 (br, 5 H), 4.40 (m, 1 H), 8.02 (dd, 1 H, J ) 6.3, 8.7
Hz), 8.22 (d, 1 H, J ) 9.0 Hz), 8.66 (d, 1 H, J ) 5.7 Hz), 8.78
1
(s, 1 H). 13C NMR (100 MHz, CD
OD): δ 19.4, 19.7, 30.8, 58.1,
r id e (3b) was prepared as a yellow viscous oil using a method
similar to that described for 3a (89%); 95.7% purity based on
3
62.5, 82.4, 129.6, 131.5, 133.2, 137.1, 160.3.
1
HPLC (t
(
5
8
1
1
R
) 2.89 min). H NMR (300 MHz, CD
3
OD): δ 4.11
1-(1-H yd r oxy-3-m et h ylb u t a n -2-yl)-4-p h en ylp yr id in i-
u m Ch lor id e (2g) was prepared as a yellow viscous oil using
a method similar to that described for 3a (74%); 95.0% purity
s, 3 H), 4.52 (s, 2 H), 7.63-7.72 (m, 4 H), 8.16 (dd, 1 H, J )
.7, 8.7 Hz), 8.40 (d, 1H, J ) 8.7 Hz), 8.70 (d, 1 H, J ) 5.4 Hz),
1
3
1
.90 (s, 1 H). C NMR (100 MHz, CD
3
OD): δ 58.4, 61.3, 127.3,
based on HPLC (t
R
) 5.18 min). H NMR (300 MHz, CD
3
OD):
29.4, 130.6, 131.7, 133.0, 133.2, 135.0, 137.0, 139.9, 143.4,
60.1.
δ 0.82 (d, 3 H, J ) 6.6 Hz), 1.20 (d, 3 H, J ) 6.6 Hz), 2.49 (m,
1 H), 4.10 (d, 2 H, J ) 5.4 Hz), 4.43 (m, 1 H), 7.64 (m, 3 H),
8
(
3
.02 (dd, 2 H, J ) 2.1, 7.5 Hz), 8.42 (d, J ) 6.9 Hz, 2 H), 9.03
1
-(2-Hydr oxym eth ylph en yl)-4-ph en ylpyr idin iu m Ch lo-
d, 2 H, J ) 6.9 Hz). 13C NMR (75 MHz, CD
OD): δ 19.5, 19.7,
0.7, 62.5, 81.2, 126.0 (2C), 129.2 (2C), 131.0 (2C), 133.5, 135.2,
r id e (3c) was prepared as a yellow viscous oil using a method
3
similar to that described for 3a (75%); 96.8% purity based on
1
145.3 (2C), 158.4.
-(2-Hyd r oxy-1-p h en yleth yl)p yr id in iu m Ch lor id e (2i)
HPLC (t
(
8
R
) 4.48 min). H NMR (300 MHz, CD
3
OD): δ 4.54
1
s, 2H), 7.60-7.80 (m, 7 H), 8.08 (dd, 2 H, J ) 2.7, 4.8 Hz),
.52 (d, 2 H, J ) 6.3 Hz), 9.02 (d, 2 H, J ) 6.3 Hz). 1 C NMR
3
was prepared as a yellow viscous oil using a method similar
to that described for 3a (61%). The spectral data were identical
(100 MHz, CD
3
OD): δ 60.4, 124.5, 126.4, 128.3 (2C), 129.6,
to those of the literature;19 97.8% purity based on HPLC (t
)
1
29.9 (2C), 130.7 (2C), 131.8, 132.7, 134.0, 136.1, 141.8, 146.0
R
3
.29 min).
-(2-H yd r oxy-1-p h e n yle t h yl)-3-m e t h oxyp yr id in iu m
(2C), 157.8.
1
1
-(3-Hyd r oxyp r op yl)p yr id in iu m Ch lor id e (2a ) was pre-
Ch lor id e (2j) was prepared as a yellow viscous oil using a
method similar to that described for 3a (81%); 95.5% purity
pared as a yellow viscous oil using a method similar to that
described for 3a (51%). The spectral data were identical to
1
1
8
based on HPLC (t
R
) 3.67 min). H NMR (300 MHz, CD
3
OD):
those of the literature; 96.6% purity based on HPLC (t
.79 min).
-(3-Hyd r oxyp r op yl)-3-m eth oxyp yr id in iu m Ch lor id e
2b) was prepared as a yellow viscous oil using a method
similar to that described for 3a (61%); 96.3% purity based on
R
)
δ 4.05 (s, 3 H), 4.34 (dd, 1 H, J ) 3.9, 12.3 Hz), 4.55 (dd, 1 H,
1
J ) 8.7, 12.3 Hz), 6.04 (dd, 1 H, J ) 3.9, 9.0 Hz), 7.43-7.56
1
(
m, 5 H), 8.02 (dd, 1 H, J ) 6.0, 8.4 Hz), 8.20 (d, 1 H, J ) 8.7
(
13
Hz), 8.66 (d, 1 H, J ) 6.0 Hz), 8.84 (s, 1H). C NMR (75 MHz,
CD OD): δ 58.1, 63.3, 77.9, 129.2 (2C), 129.8, 130.6 (2C), 131.2,
31.7, 133.6, 135.2, 137.1, 160.3.
-(2-Hydr oxy-1-ph en yleth yl)-4-ph en ylpyr idin iu m Ch lo-
1
3
HPLC (t
quint, 2 H, J ) 6.3 Hz), 3.63 (t, 2 H, J ) 5.7 Hz), 4.07 (s, 3
H), 4.74 (t, 2 H, J ) 7.2 Hz), 8.00 (dd, 1 H, J ) 5.7, 8.7 Hz),
.18 (d, 1 H, J ) 9.0 Hz), 8.62 (d, 1 H, J ) 5.7 Hz), 8.82 (s, 1
R
) 2.03 min). H NMR (300 MHz, CD
3
OD): δ 2.22
1
(
1
r id e (2k ) was prepared as a viscous oil using a method similar
to that described for 3a (63%). The spectral data were identical
8
1
3
H). C NMR (100 MHz, CD
29.5, 131.5, 133.7, 138.3, 160.2.
-(3-Hydr oxypr opyl)-4-ph en ylpyr idin iu m Ch lor ide (2c)
3
OD): δ 34.6, 58.1, 58.8, 60.8,
to those of the literature;19 98.7% purity based on HPLC (t
)
R
1
5
.66 min).
1
was prepared as a yellow viscous oil using a method similar
to that described for 3a (43%); 96.3% purity based on HPLC
Ack n ow led gm en t. We are grateful to the National
Science Foundation and the Cystic Fibrosis Foundation
for financial support of this research. The 400 and 300
MHz NMR spectrometers used in this study were
funded in part by a grant from NSF (CHE-9808183).
1
(
3
t
R
) 4.18 min). H NMR (300 MHz, CD
3
OD): δ 2.24 (m, 2 H),
.66 (t, 2 H, J ) 5.4 Hz), 4.75 (t, 2 H, J ) 6.9 Hz), 7.64 (br, 3
H), 8.01 (dd, 2 H, J ) 3.9, 4.8 Hz), 8.40 (d, 2 H, J ) 5.7 Hz),
.98 (d, 2 H, J ) 6.0 Hz). 1 C NMR (75 MHz, CD
3
8
5
1
3
OD): δ 34.4,
8.9, 59.7, 126.1 (2C), 129.2 (2C), 130.9 (2C), 133.4, 135.3,
46.2 (2C), 157.9.
Su p p or tin g In for m a tion Ava ila ble: 1H NMR spectra for
1
crude resin cleavage products 2i and 3a , H NMR spectra for
2
-(3-Hyd r oxyp r op yl)isoqu in olin iu m Ch lor id e (2d ) was
prepared as a yellow viscous oil using a method similar to that
described for 3a (52%); 92.9% purity based on HPLC (t ) 3.07
OD): δ 2.30 (m, 2 H), 3.66 (t, 2
H, J ) 5.1 Hz), 4.90 (br, 2 H), 8.06 (t, 1 H, J ) 7.2 Hz), 8.24
13
compounds 2a -k , 3a -c, and 10, and C NMR spectra for
compounds 2a -d , f, g, and i-k , and 3a -c. This material is
R
available free of charge via the Internet at http://pubs.acs.org.
1
min). H NMR (300 MHz, CD
3
J O0001636
(
18) Glarner, F.; Thornton, S. R.; Scharer, D.; Bernardinelli, G.;
(19) Genisson, Y.; Mehmandoust, M.; Marazano, C.; Das, B. C.
Heterocycles 1994, 39, 811.
Burger, U. Helv. Chim. Acta 1997, 80, 121.