Polymer Journal
Supplementary Information for
Synthesis and evaluation of resins bearing substrate-like
inhibitor functions for capturing copper amine oxidases
Marco Pocci,* Silvana Alfei, Sara Castellaro,
Francesco Lucchesini, Marco Milanese and Vincenzo Bertini
Dipartimento di Farmacia, Università di Genova
Via Brigata Salerno, 13 I-16147 Genova, Italy.
*Corresponding author: Marco Pocci
Tel.: +39 010 3532685; Fax: +39 010 3532684
E-mail: pocci@dictfa.unige.it
Contents
1. Synthesis of 2,6-dimethoxy-4-hydroxybenzaldehyde (4)
2. Synthesis of 2,6-dimethoxy-4-hydroxybenzaldoxime (7)
3. Synthesis of 2,6-diethoxy-4-hydroxybenzaldoxime (8)
4. Synthesis of 2,6-dimethoxy-4-hydroxybenzonitrile (9)
5. Synthesis of 2,6-diethoxy-4-hydroxybenzonitrile (10)
6. Synthesis of (E)-2,6-dimethoxy-4-[(2-tetrahydropyranyloxy)-2-butenyloxy]benzonitrile (12)
7. Synthesis of (E)-2,6-diethoxy-4-[(2-tetrahydropyranyloxy)-2-butenyloxy]benzonitrile (13)
8. References
1.
Synthesis of 2,6-dimethoxy-4-hydroxybenzaldehyde (4)
OEt
HO
OEt
DMF/POCl3
rt HO
OEt -10 °C
OEt
OEt
CH=O
OEt
4 (50%)
+
O=HC
+
HO
OEt
CH=O
5 (26%)
HO
OEt
CH=O
6 (11%)
A mixture of 3,5-diethoxyphenol [1] (3.52 g, 19.3 mmol) and POCl3 (5.67 g, 37.0 mmol, 3.4 mL) was cooled to -10 °C,
treated dropwise with DMF (2.14 g, 29.3 mmol, 2.3 mL), stirred under N2 at rt for 24 h 40 min then poured in iced
water and treated with solid NaOH (7.01 g, 175.3 mmol) up to pH = 9. The solution was acidified with 12% aq. HCl
1
(pH = 6-7) to give a solid precipitate which was filtered, dried over P 2O5, washed with CHCl3 (2x10 mL) and
crystallized from MeOH to afford 2,6-diethoxy-4-hydroxybenzaldehyde (4) (2.02 g, 50%). Purity 99% by HPLC. Mp:
178-180 °C. FTIR (KBr, cm-1): 3218 (OH); 1659 (CH=O). 1H-NMR (DMSO-d6, δ ppm): 1.33 (t, 6H, J = 7.0 Hz); 4.00
(q, 4H, J = 7.0 Hz); 6.06 (s, 1H); 10.20 (s, 1H); 10.50 (s, broad OH). 13C-NMR: 14.33, 63.82, 92.44, 107.04, 162.84,
164.77, 185.31.
The CHCl3 washings after removal of the solvent left 2,4-diethoxy-6-hydroxybenzaldehyde (5) (1.06 g, 26%). For
characterization purposes a sample was sublimed at reduced pressure in the form of a white waxy solid. Purity 98% by
HPLC. Mp: 70-71°C. FTIR (KBr, cm-1): 1643 (CH=O). 1H-NMR (CDCl3, δ ppm): 1.41 (t, 3H, J = 7.0 Hz); 1.43 (t, 3H,
J = 7.0 Hz); 4.048 (q, 2 H, J = 7.0 Hz); 4.052 (q, 2 H, J = 7.0 Hz); 5.88 (d, 1H, J = 2.0 Hz); 5.97 (d, 1H, J = 2.0 Hz);
10.10 (d, 1H, J = 0.4 Hz); 12.50 (s, 1H). 13C-NMR: 14.42, 14.48, 64.04, 64.17, 91.44, 93.17, 105.94, 162.91, 166.26,
167.47, 191.87. 2- Hydroxy-4,6-diethoxy-1,3-benzenedicarboxaldehyde (6) was obtained by slow spontaneous
precipitation from the aqueous phase of the hydrolytic step (0.536 g, 11%). Purity 99% by HPLC. Mp: 168-171 °C.
FTIR (KBr, cm-1): 3427 (OH); 1690 (CH=O). 1H-NMR (CDCl3, δ ppm): 1.51 (t, 6H, J = 7.0 Hz); 4.20 (q, 4H, J = 7.0
Hz); 5.91 (s, 1H); 10.22 (s, 1H); 13.47 (s, OH). 13C-NMR: 14.35, 65.15, 86.94, 106.07, 167.76, 168.32, 189.44.
2. Synthesis of 2,6-dimethoxy-4-hydroxybenzaldoxime (7)
OMe
CH=O
HO
OMe
3
OMe
CH=NOH
NH 2OH . HCl
abs EtOH
reflux
Pyridine
HO
OMe
7 (90%)
A mixture of 3 [2,3] (2.98 g, 16.4 mmol), EtOH 100% (35 mL) and pyridine (3.5 mL) was treated under stirring at rt
with NH2OH·HCl (1.37 g, 19.7 mmol) to afford an orange suspension which was refluxed for 3 h and left at rt
overnight. After removal of the solvent at reduced pressure the solid residue was crystallized from EtOH 95% to give 7
as an orange solid (2.91 g, 90%). Purity 97% by HPLC. Mp: 210-218 °C (dec.). FTIR (KBr, cm-1): 3316, 3234 (OH),
1597 (C=N). 1H-NMR (DMSO-d6, δ ppm): 3.72 (s, 6H); 6.12 (s, 2H); 8.13 (s,1H); 9.93 (s, broad, OH); 10.67 (s, very
broad, OH). 13C-NMR: 55.45, 92.03, 100.74, 142.56, 159.24, 159.98.
3. Synthesis of 2,6-diethoxy-4-hydroxybenzaldoxime (8)
OEt
CH=O
HO
OEt
4
OEt
NH 2OH . HCl
abs EtOH
reflux
Pyridine
CH=NOH
HO
OEt
8 (78%)
A solution of 4 (2.44 g, 11.6 mmol), EtOH 100% (28.5 mL) and pyridine (2.9 mL) was treated as described for 7 with
NH2OH·HCl (0.967 g, 13.9 mmol). After 2 h reflux the solvent was removed at reduced pressure to afford 8 which was
crystallized from EtOH 90% (2.02 g, 78%). Purity 97% by HPLC. Mp: 150-152 °C. FTIR (KBr, cm-1): 3411 (OH),
1611 (C=N). 1H-NMR (DMSO-d6, δ ppm): 1.30 (t, 6H, J = 7.0 Hz); 3.96 (q, 4H, J = 7.0 Hz); 6.07 (s, 2H); 8.12 (s, 1H);
9.76 (s, broad, OH), 10.59 (s, broad, OH). 13C-NMR: 14.56, 63.61, 92.98, 101.42, 142.91, 158.55, 159.71.
2
4. Synthesis of 2,6-dimethoxy-4-hydroxybenzonitrile (9)
OMe
CH=NOH
HO
OMe
7
OMe
C N
Cu(OAc)2
CH 3CN
reflux
HO
OMe
9 (77%)
A mixture of 7 (3.52 g, 17.9 mmol), CH3CN (19 mL) and a solution of Cu(OAc)2 (0.376 g, 2.1 mmol) in CH3CN (19
mL) was heated under N2 with stirring for 2 h 45 min until disappearance of 7 (HPLC). The brown suspension was
hydrolyzed with 5% H2SO4 (100 mL), extracted first with Et 2O (3x30 mL) then with EtOAc (3x30 mL). The extracts
were combined, washed with H2O then dried (Na2SO4).The removal of the solvent at reduced pressure afforded 9 (2.45
g, 77%). Purity 97% by HPLC. Mp: 261-262 °C (CH3CN); FTIR (KBr, cm-1): 3222 (OH), 2228 (CN); 1H-NMR
(DMSO-d6, δ ppm): 3.81 (s, 6H); 6.14 (s, 2H); 10.69 (s, OH). 13C-NMR (δ, ppm): 55.97, 80.70, 92.08, 114.67, 163.32,
164.02.
5. Synthesis of 2,6-diethoxy-4-hydroxybenzonitrile (10)
OEt
HO
OEt
CH=NOH
Cu(OAc)2
OEt
CH 3CN
reflux
8
C N
HO
OEt
10 (70%)
A mixture of 8 (1.03 g, 4.6 mmol), CH3CN (4.5 mL) and a solution of Cu(OAc)2 (0.092 g, 0.51 mmol) in CH3CN (44
mL) was heated under N2 with stirring for 1 h 30 min until disappearance of 8 (TLC, petroleum ether/EtOAc = 1/2 as
eluent). The blue solution was hydrolyzed with 5% H2SO4 (30 mL), extracted first with Et 2O (30 mL) then with EtOAc
(2x30 mL). The extracts were combined, washed with H 2O then dried (Na2SO4). The removal of the solvent at reduced
pressure afforded crude 10 which was crystallized from toluene as a yellow solid (0.658 g, 70%). Purity 99% by HPLC.
Mp: 173-175 °C. FTIR (KBr, cm-1): 3247 (OH), 2222 (CN). 1H-NMR (DMSO-d6, δ ppm): 1.34 (t, 6H, J = 7.0 Hz); 3.34
(s, OH); 4.06 (q, 4H, J = 7.0 Hz); 6.11 (s, 2H). 13C-NMR: 14.25, 64.15, 81.26, 92.62, 114.64, 162.55, 163.83.
6. Synthesis of (E)-2,6-dimethoxy-4-[(2-tetrahydropyranyloxy)-2-butenyloxy]benzonitrile (12)
OMe
C N
HO
OMe
9
OMe
C N
1) NaH/DMF
2) THPO
THP O
Cl
O
OMe
12 (63%)
A solution of benzonitrile 9 (3.70 g, 20.7 mmol) in dry DMF (38.5 mL) was added under N2 to a suspension of NaH
[0.933 g (23.3 mmol) as 60% mineral oil dispersion, washed with pentane (3x2 mL)] in dry DMF (8 mL). The
suspension was left under stirring at rt for 1 h 30 min, cooled to 0 °C, treated with (E)-1-chloro-4-(2tetrahydropyranyloxy)-2-butene (11) [4] (5.79 g, 30.4 mmol, 5.5 mL), refluxed for 3 h, hydrolyzed with H 2O (150 mL)
and extracted with Et2O (3x50 mL). The extracts were combined, washed with 10% NaOH and dried (Na2SO4). The
removal of the solvent at reduced pressure left a residue (7.69 g) which was subjected to column chromatography using
3
a mixture EtOAc/petroleum ether = 40/60 as eluent to give 12 as an oil (4.38 g, 63%). Purity 99% by HPLC. FTIR
(film, cm-1): 2219 (CN), 970 (CH=CH, trans). 1H-NMR (CDCl3, δ ppm): 1.52-1.95 (m, 6H); 3.47-3.58 (m, 1H); 3.833.90 (m, 1H); 3.85 (s, 6H); 4.00-4.07 (s, 1H); 4.27-4.34 (m, 1H); 4.57-4.60 (m, 2H); 4.63-4.68 (m, 1H); 5.94-6.03 (m,
2H); 6.09 (s, 2H). 13C-NMR: 19.45, 25.41, 30.57, 56.12, 62.30, 66.66, 68.45, 84.26, 91.09, 98.20, 114.55, 125.98,
131.29, 163.77, 164.29.
The last fractions of the column eluted with 100% EtOAc allowed the isolation of solid (E)-2,6-dimethoxy-4-[4hydroxy-(2-butenyloxy)]benzonitrile (0.46 g, 9%). Purity 98% by HPLC. Mp: 201-202 °C. FTIR (KBr, cm-1): 3412
(OH), 2216 (CN). 1H-NMR (Acetone-d6, δ ppm): 2.89 (s, OH); 3.91 (s, 6H); 4.10- 4.17 (m, 2H); 4.66-4.72 (m, 2H);
5.85-6.10 (m, 2H); 6.33 (s, 2H). 13C-NMR: 56.71, 62.33, 69.45, 84.44, 92.32, 114.64, 124.55, 135.85, 164.59, 165.53.
7. Synthesis of (E)-2,6-diethoxy-4-[(2-tetrahydropyranyloxy)-2-butenyloxy]benzonitrile (13)
OEt
HO
10
C N
1) NaH/DMF
OEt
2) THPO
OEt
C N
Cl
THP O
O
OEt
13 (82%)
A solution of benzonitrile 10 (2.00 g, 9.7 mmol) in dry DMF (13 mL) was added under N2 to a suspension of NaH [0.44
g (11.0 mmol) as 60 % mineral oil dispersion, washed with pentane (3x2 mL)] in dry DMF (3.7 mL). The suspension
was left under stirring at rt for 1 h 30 min, cooled to 0 °C, treated with 11 [4] (2.71 g, 14.2 mmol, 2.6 mL), heated at 70
°C for 4 h 30 min, hydrolyzed with H 2O (75 mL) and extracted with Et2O (3x25 mL). The extracts were combined,
washed with 10% NaOH and dried (Na2SO4). The removal of the solvent at reduced pressure left a residue (4.19 g)
which was subjected to column chromatography using a mixture EtOAc/petroleum ether = 40/60 as eluent to give 13 as
oil (2.89 g, 82%). Purity 99% by HPLC. FTIR (film, cm-1): 2220 (CN), 970 (CH=CH, trans). 1H-NMR (Acetone-d6, δ
ppm): 1.40 (t, 6H, J = 7.0 Hz); 1.45-1.85 (m, 6H); 3.42-3.48 (m, 1H); 3.74-3.83 (m, 1H); 3.94-4.02 (m, 1H); 4.15 (q,
4H, J = 7.0 Hz); 4.17-4.27 (m, 1H); 4.61-4.64 (m, 1H); 4.65-4.69 (m, 2H); 5.88-6.07 (m, 2H); 6.27 (s, 2H). 13C-NMR:
14.74, 19.95, 26.17, 31.21, 62.15, 65.36, 66.98, 69.09, 84.87, 92.73, 98.41, 114.62, 126.77, 131.86, 163.72, 165.13.
The last fractions of the column eluted with 100% EtOAc allowed the isolation of solid (E)-2,6-diethoxy-4-[4-hydroxy(2-butenyloxy)]benzonitrile (0.0944 g, 4%). Purity 99% by HPLC. Mp: 98-100 °C. FTIR (KBr, cm-1): 3476 (OH), 2230
(CN). 1H-NMR (Acetone-d6, δ ppm): 1.40 (t, 6H, J = 7.0 Hz); 3.85-4.12 (m, 2H); 4.17 (q, 4H, J = 7.0 Hz); 4.65-4.68
(m, 2H); 5.88-6.06 (m, 2H); 6.30 (s, 2H). 13C-NMR: 14.79, 62.33, 65.48, 69.39, 85.09, 92.96, 114.65, 124.59, 135.81,
163.88, 165.40.
8. References
1. Maillard J, Vincent M, Delaunay P, Fumeron B (1996) Derivés du phloroglucinol. 1. Synthèse de mono, di et
triéthers d’acides trihydroxy-2,4,6-benzoiques mono et dichlorés, et leur acétylation. Bull. Soc. Chim. Fr. (8): 25202524
2. Albericio F, Kneib-Cordonier N, Biancalana S, Gera L, Masada RI, Hudson D, Barany G (1990) Preparation and
application of the 5-(4-(9-fluorenylmethyloxycarbonyl)aminomethyl-3,5-dimethoxyphenoxy)valeric acid (PAL) handle
for the solid-phase synthesis of C-terminal peptide amides under mild conditions. J. Org. Chem. 55(12): 3730-3743
4
3. Jin J, Graybill TL, Wang MA, Davis LD, Moore ML (2001) Convenient preparation of 4-formyl-3,5dimethoxyphenol and its incorporation into linkers and resins for solid-phase synthesis. J. Comb. Chem. 3(1): 97-101
4. Deslongschamp P, Lamothe S, Ho-Shen-Lin (1987) A simple and direct method of cyclization for the synthesis of
10-membered rings. Can. J. Chem. 65(6): 1298-1307
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