Abstract
Four phthalazine derivatives have been prepared from substituted 2-bromobenzaldehyde acetals by a sequence involving: (1) lithiation and formylation; (2) deprotection; and (3) condensative cyclization with hydrazine. Two additional phthalazines were prepared by a similar sequence following direct lithiation of benzaldehyde acetals substituted by anion-stabilizing groups at C3. These syntheses can be conveniently carried out to give phthalazines in overall yields of 40–70%.
Introduction
Structural modifications to optimize the activity of antibiotic 1 are currently under investigation in our laboratory. These drugs act against inhalation anthrax and multidrug-resistant staph by inhibiting dihydrofolate reductase (DHFR), a key enzyme required for bacterial growth. An important feature of these compounds is that they selectively target bacterial DHFR, while not harming human DHFR (Bourne et al., 2009).
In an earlier study (Bourne et al., 2009), X-ray analysis of the DHFR-(S)-1 complex indicated interactions between DHFR and the dihydrophthalazine portion of the drug. This work revealed that space was available in the active site for a small substituent at C5, C6 or C7 on this ring. Based upon this finding and with the goal of maximizing the antibacterial activity of 1, we directed our efforts toward preparing several phthalazines substituted by small groups that could be accommodated in the DHFR active site.
Results and discussion
Our approach to phthalazine derivatives is outlined in Scheme 1. The starting bromoacetal derivatives 2a–d were either known or readily prepared using standard methodology (Remy et al., 1985; Moody and Warrellow, 1990; Balczewski et al., 2006). Lithium-bromide exchange was carried out by adding 1.2 equiv. of n-butyllithium to a solution of each bromide 2 in THF at -78°C and warming to -40°C for 30 min. The mixture was then recooled to -78°C, and 1.2 equiv. of anhydrous N,N-dimethylformamide was added. Stirring was continued for 30 min, and the reaction was worked up to give aldehydes 3a–d in 56–84% yields. Deprotection of 3a–d was accomplished by stirring with wet Amberlyst® 15 in acetone (Rohm and Haas Co., 1978; Kalesse, 1995). This cleanly converted the acetals back to the aldehydes to give phthalaldehydes 4a–d in 73–93% yields. Finally, o-dialdehydes 4a–d were each reacted with 1.1 equiv. of anhydrous hydrazine in absolute ethanol at 0°C–23°C for 3 h (Hirsch and Orphanos, 1965; Bhattacharjee and Popp, 1980) to give phthalazines 5a–d in 78–98% yields.
Synthesis of 5a–d; (a) i. n-BuLi, THF, -78°C, ii. warm to -40°C, iii. cool to -78°C, iv. DMF, -78°C–23°C; (b) wet Amberlyst® 15, acetone, 23°C; (c) anhyd NH2NH2, EtOH, 0°C–23°C.
Our route is similar to one reported earlier (Tsoungas and Searcey, 2001) for the preparation of the 6-methoxyphthalazine (5a). In the current study, however, additional examples of this transformation are described and more procedural details are given. Finally, the methodology has generally been streamlined to minimize extensive purification of intermediates.
During our study, it was found that two cases did not require the presence of bromine on the aromatic ring for lithiation to proceed, although the reaction regioselectivity was altered (see Scheme 2). Direct lithiation of piperonal and 3-fluorobenzaldehyde acetals 6a (Charlton et al., 1996) and 6b (Dellaria, 2001) yielded preferential metalation of the aromatic site flanked by two heteroatom groups, that is, at C2 rather than at C6 (Gschwend and Rodriguez, 1979). Thus, for these two substrates, the resulting aldehydes 7a (89%) and 7b (66%) were substituted at C3 and C4 rather than at C4 and C5. Similar precursors having a methyl or a methoxy group at C3 were insufficiently activated or too hindered to allow direct lithiation at C2 and gave no products under our conditions. Finally, while 7a smoothly underwent deprotection to 8a (98%) and conversion to phthalazine 9a (86%), deprotection of 7b was difficult to monitor by thin layer chromatography and phthalaldehyde 8b was sensitive toward purification. To circumvent this problem, crude 8b was reacted directly with hydrazine in anhydrous ethanol to give phthalazine 9b in a two-step yield of 61%.
Synthesis of 9a–b; (a) i. n-BuLi, THF, -78°C, ii. warm to -40°C, iii. cool to -78°C, iv. DMF, -78°C–23°C; (b) wet Amberlyst® 15, acetone, 23°C; (c) anhyd NH2NH2, EtOH, 0°C–23°C.
Conclusion
We have developed a convenient synthesis of a series of substituted phthalazines from readily available benzaldehyde acetals. The procedure allows the preparation of phthalazines substituted by groups stable to metalation conditions with n-butyllithium. The lithiation process is facilitated by the acetal group positioned ortho to the bromine, but direct C2 metalation is also possible when anion-stabilizing groups are present at C3. These two processes allow access to phthalazines with different substitution patterns in yields ranging from 40% to 70%.
Experimental
All reactions were run in oven-dried glassware. Reactions were monitored by TLC using silica gel GF plates. Flash chromatography was performed in quartz columns using silica gel (Davisil®, grade 62, 60–200 mesh). Band elution for all chromatographic separations was monitored using a hand-held UV lamp. 1H and 13C NMR spectra were measured in CDCl3 at 300 MHz and 75 MHz, respectively, and were referenced to internal tetramethylsilane. Low-resolution mass spectra (EI) were recorded at 30 eV.
General procedure for lithium-bromine exchange and formylation
2-(1,3-Dioxolan-2-yl)-4-methoxybenzaldehyde (3a)
To a solution of 5.18 g (20.0 mmol) of 2a (Remy et al., 1985) in 50 mL of dry THF at -78°C was added dropwise over 1 h, 10.9 mL (24.0 mmol, 1.2 equiv.) of 2.2 M n-butyllithium in hexanes. The reaction mixture was stirred for an additional 15 min at -78°C, and then warmed to -40°C and maintained at this temperature for 30 min. The mixture was again cooled to -78°C, and 1.75 g (1.86 mL, 24.0 mmol, 1.2 equiv.) of anhydrous DMF was added dropwise over 30 min with stirring at -78°C for an additional 30 min. The crude reaction mixture was added to aqueous NH4Cl and extracted with ether (2 × 100 mL). The combined organic layers were washed with aqueous NaCl, dried (MgSO4), and concentrated under reduced pressure to give a yellow oil. This material was purified by flash chromatography on a 50-cm × 2-cm column eluting with 2–10% ethyl acetate in hexanes to give 3.49 g (84%) of 3a as a light yellow oil. IR: 2845, 1689 cm-1; 1H NMR: δ 10.23 (s, 1H), 7.90 (d, J = 8.5 Hz, 1H), 7.26 (d, J = 2.7 Hz, 1H), 6.98 (dd, J = 8.5, 2.7 Hz, 1H), 6.45 (s, 1H), 4.15 (m, 2H), 4.09 (m, 2H), 3.90 (s, 3H); 13C NMR: δ 190.2, 163.8, 141.6, 133.3, 127.6, 114.2, 112.1, 110.3, 65.3, 55.7; MS: m/z 208 (M+).
2-(1,3-Dioxolan-2-yl)-4,5-dimethoxybenzaldehyde (3b)
Scale: 20.0 mmol of 2b (Moody and Warrellow, 1990); yield 68% of 3b as a white solid, mp 96–98°C (lit mp 98–99°C; Moody and Warrellow, 1990); IR: 2835, 1682 cm-1; 1H NMR: δ 10.34 (s, 1H), 7.48 (s, 1H), 7.22 (s, 1H), 6.36 (s, 1H), 4.18 (m, 2H), 4.12 (m, 2H), 3.99 (s, 3H), 3.95 (s, 3H); 13C NMR: δ 189.6, 153.4, 149.5, 134.0, 127.7, 110.5, 108.9, 100.5, 65.3, 56.2, 56.1; MS: m/z 238 (M+).
2-(1,3-Dioxolan-2-yl)-4,5-(methylenedioxy)benzaldehyde (3c)
Scale: 20.0 mmol of 2c (Balczewski et al., 2006); yield 56% of 3c as a colorless oil that solidified at 0°C following flash chromatography as above, mp 66–68°C (lit mp 69–71°C; Moody and Warrellow, 1990); IR: 2896, 2788, 1680, 1610 cm-1; 1H NMR: δ 10.25 (s, 1H), 7.36 (s, 1H), 7.16 (s, 1H), 6.32 (s, 1H), 6.05 (s, 2H), 4.14 (m, 2H), 4.06 (m, 2H); 13C NMR: δ 188.9, 152.0, 148.4, 136.5, 129.3, 107.8, 106.6, 102.1, 100.0, 65.2; MS: m/z 222 (M+).
2-(1,3-Dioxolan-2-yl)-4-methylbenzaldehyde (3d)
Scale: 20.0 mmol of 2d; yield 70% of 3d as a colorless oil following flash chromatography as above; IR: 1692, 1389 cm-1; 1H NMR: δ 10.33 (s, 1H), 7.83 (d, J = 7.9 Hz, 1H), 7.55 (s, 1H), 7.32 (d, J = 7.9 Hz, 1H), 6.40 (s, 1H), 4.16 (m, 2H), 4.09 (m, 2H), 2.44 (s, 3H); 13C NMR: δ 191.4, 144.7, 138.8, 132.0, 130.7, 130.0, 127.5, 100.8, 65.3, 21.8; MS: m/z 192 (M+).
General procedure for aldehyde deprotection
4-Methoxyphthalaldehyde (4a)
A solution of 3.22 g (15.5 mmol) of 3a in 50 mL of acetone was treated with 0.50 g of wet Amberlyst®15 and stirred vigorously for 1 h, during which time a white solid formed. At this point, 30 mL of dichloromethane was added to dissolve the product, the mixture was filtered through Celite®, and the solution was concentrated under reduced pressure. The resulting oil was purified by flash chromatography using increasing concentrations (5–20%) of ether in hexane to give 1.86 g (73%) of 4a as a white solid, mp 39–41°C (lit mp 41–42°C; Pappas et al., 1968). IR: 2751, 2848, 1692 cm-1; 1H NMR: δ 10.66 (s, 1H), 10.33 (s, 1H), 7.94 (d, J = 8.2 Hz, 1H), 7.46 (d, J = 2.7 Hz, 1H), 7.23 (dd, J = 8.2, 2.7 Hz, 1H), 3.96 (s, 3H); 13C NMR: δ 191.9, 191.0, 163.8, 138.6, 134.6, 129.4, 118.7, 114.7, 55.9; MS: m/z 164 (M+).
4,5-Dimethoxyphthalaldehyde (4b)
Scale: 2.20 mmol of 3b; yield 93% of 4b as a white solid, mp 165–167°C (lit mp 168–169°C; Bhattacharjee and Popp, 1980); IR: 2849, 2772, 1677 cm-1; 1H NMR: δ 10.59 (s, 2H), 7.48 (s, 2H), 4.04 (s, 6H); 13C NMR: δ 190.0, 153.1, 130.9, 111.5, 56.4; MS: m/z 194 (M+).
4,5-(Methylenedioxy)phthalaldehyde (4c)
Scale: 2.20 mmol of 3c; yield 91% of 4c, mp 143–145°C (lit mp 143.5°C; Kessar et al., 1991); IR: 2862, 2754, 1691, 1675 cm-1; 1H NMR: δ 10.50 (s, 2H), 7.42 (s, 2H), 6.19 (s, 2H); 13C NMR: δ 189.7, 152.1, 133.4, 109.5, 103.0; MS: m/z 178 (M+).
4-Methylphthalaldehyde (4d)
Scale: 15.5 mmol of 3d; yield 85% of 4d as a colorless oil (lit mp 37–38°C; Pappas et al., 1968), which was used without further purification; IR: 2861, 2745, 1695 cm-1; 1H NMR: δ 10.55 (s, 1H), 10.46 (s, 1H), 7.88 (d, J = 7.7 Hz, 1H), 7.77 (s, 1H), 7.57 (d, J = 7.7 Hz, 1H), 2.51 (s, 3H); 13C NMR: δ 192.4, 191.9, 144.8, 136.1, 134.0, 133.7, 131.4, 131.2, 21.3; MS: m/z 132 (M+).
General procedure for condensative cyclization using hydrazine
6-Methoxyphthalazine (5a)
To a stirred solution of 1.64 g (10.0 mmol) of 4a in 30 mL of absolute ethanol at 0°C was added dropwise 0.35 g (0.34 mL, 11.0 mmol, 1.1 equiv.) of anhydrous hydrazine. Stirring was continued with gradual warming to 23°C until TLC indicated the reaction was complete (3 h). The solvent was removed under vacuum, and the resulting product was crystallized from benzene-pentane to give 1.31 g (82%) of 5a as a tan solid; mp 117–119°C; IR: 2854, 1616 cm-1; 1H NMR: δ 9.47 (s, 1H), 9.40 (s, 1H), 7.87 (d, J = 8.8 Hz, 1H), 7.51 (dd, J = 8.8, 2.2 Hz, 1H), 7.20 (d, J = 2.2 Hz, 1H), 4.01 (s, 3H); 13C NMR: δ 162.4, 150.6, 150.0, 128.5, 128.0, 124.9, 122.0, 103.9, 55.8; MS: m/z 160 (M+).
6,7-Dimethoxyphthalazine (5b)
Scale: 1.20 mmol of 4b; yield 82% of 5b as a tan solid, mp 196–198°C (lit mp 198–200°C; Bhattacharjee and Popp, 1980); IR: 2840, 1612 cm-1; 1H NMR: δ 9.38 (s, 2H), 7.18 (s, 2H), 4.09 (s, 6H); 13C NMR: δ 154.1, 149.4, 123.2, 104.2, 56.4; MS: m/z 190 (M+).
6,7-(Methylenedioxy)phthalazine (5c)
Scale: 1.20 mmol of 4c; yield 98% of 5c as a tan solid, mp 255–257°C (lit mp 255°C; Dallacker et al., 1961); IR: 1606 cm-1; 1H NMR: δ 9.33 (s, 2H), 7.19 (s, 2H), 6.21 (s, 2H); 13C NMR: δ 152.0, 149.8, 124.9, 102.43, 102.37; MS: m/z 174 (M+).
6-Methylphthalazine (5d)
Scale: 10.0 mmol of 4d; yield 78% of 5d as a tan solid, mp 69–71°C (lit mp 72°C; Robev, 1981); IR: 1620, 1374 cm-1; 1H NMR: δ 9.47 (s, 2H), 7.87 (d, J = 8.5 Hz, 1H), 7.76 (d, J = 8.5 Hz, 1H), 7.73 (s, 1H), 2.63 (s, 3H); 13C NMR: δ 150.7, 150.6, 143.5, 134.6, 126.6, 125.9, 125.1, 124.7, 22.1; MS: m/z 144 (M+).
4.4. General procedure for direct metalation and formylation
6-(1,3-Dioxolan-2-yl)-2,3-(methylenedioxy)benzaldehyde (7a)
Using the lithium-bromide exchange conditions above, 2.50 g (12.9 mmol) of 6a (Charlton et al., 1996) was directly metalated and treated with anhydrous DMF to give a yellow solid. Trituration of this product in ether gave 2.55 g (89%) of 7a as a white solid; mp 70–72°C; IR: 1690 cm-1; 1H NMR: δ 10.41 (s, 1H), 7.18 (d, J = 8.0 Hz, 1H), 6.96 (d, J = 8.0 Hz, 1H), 6.23 (s, 1H), 6.15 (s, 2H), 4.11 (m, 2H), 4.07 (m, 2H); 13C NMR: δ 188.5, 149.9, 149.5, 130.9, 129.2, 120.7, 117.5, 111.7, 102.8, 101.5, 65.0; MS: m/z 222 (M+).
2-(1,3-Dioxolan-2-yl)-6-fluorobenzaldehyde (7b)
Scale: 10.0 mmol of 6b (Dellaria, 2001); yield 66% of 7b as a colorless oil following flash chromatography as above; IR: 1700 cm-1; 1H NMR: δ 10.51 (s, 1H), 7.61–7.55 (complex m, 2H), 7.19 (ddd, J = 10.4, 9.9, 3.8 Hz, 1H), 6.50 (s, 1H), 4.11–4.05 (complex m, 4H); 13C NMR: δ 188.4 (d, J = 9.1 Hz), 164.8 (d, J = 258.5 Hz), 140.7, 135.1 (d, J = 10.0 Hz), 122.9 (d, J = 6.8 Hz), 122.4 (d, J = 3.4 Hz), 117.1 (d, J = 21.8 Hz), 99.7 (d, J = 2.9 Hz), 65.3; MS: m/z 196 (M+).
3,4-(Methylenedioxy)phthalaldehyde (8a)
Using wet Amberlyst® 15 as described above, 0.22 g (1.00 mmol) of 7a was reacted to give 0.18 g (98%) of 8a as a white solid; mp 145–148°C. IR: 1682 cm-1; 1H NMR: δ 10.65 (s, 1H), 10.21 (s, 1H), 7.54 (d, J = 8.0 Hz, 1H), 7.10 (d, J = 8.0 Hz, 1H), 6.26 (s, 2H); 13C NMR: δ 190.9, 189.1, 153.5, 150.0, 130.3, 129.3, 118.4, 111.4, 103.7; MS: m/z 178 (M+).
5,6-(Methylenedioxy)phthalazine (9a)
Scale: 4.70 mmol of 8a and 5.17 mmol of anhydrous hydrazine; yield: 0.71 g (86%) of 9a as a tan solid; mp 167–169°C; IR: 1639 cm-1; 1H NMR: δ 9.52 (s, 1H), 9.35 (s, 1H), 7.55 (d, J = 8.0 Hz, 1H), 7.51 (d, J = 8.0 Hz, 1H), 6.34 (s, 2H); 13C NMR: δ 150.5, 149.5, 144.6, 141.0, 121.5, 121.2, 115.8, 112.0, 103.4; MS: m/z 174 (M+). Anal. Calcd for C9H6N2O2: C, 62.07; H, 3.45; N, 16.09. Found: C, 62.21; H, 3.49; N, 15.97.
5-Fluorophthalazine (9b)
To a solution of 1.08 g (5.50 mmol) of 7b dissolved in 30 mL of acetone was added 50 mg of wet Amberlyst®15, and the mixture was stirred vigorously for 2.5 h. The crude reaction mixture was filtered through Celite® and concentrated under reduced pressure. The resulting oil (1.01 g of crude 8b) was dissolved in 20 mL of absolute ethanol, cooled to 0°C, and treated with 0.20 g (0.20 mL, 6.25 mmol) of anhydrous hydrazine. The reaction mixture was stirred for 2.5 h with gradual warming to 23°C and then was concentrated to give a product that was triturated in ether to yield 0.51 g (62% for two steps) of 9b as a tan solid; mp 110–112°C (lit mp 109–110°C; Omata et al., 1989); IR: 1619 cm-1; 1H NMR: δ 9.80 (s, 1H), 9.59 (s, 1H), 7.92 (td, J = 8.2, 5.5 Hz, 1H), 7.79 (d, J = 8.2 Hz, 1H), 7.60 (t, J = 8.2 Hz, 1H); 13C NMR: δ 157.4 (d, J = 258.8 Hz), 150.1 (d, J = 2.6 Hz), 144.5 (d, J = 2.9 Hz), 133.4 (d, J = 7.7 Hz), 127.1 (d, J = 3.4 Hz), 122.1 (d, J = 4.9 Hz), 116.9 (d, J = 18.6 Hz), 116.8 (d, J = 15.8 Hz); MS: m/z 148 (M+).
T.H. gratefully acknowledges the Department of Chemistry at Oklahoma State University (OSU) for a teaching assistantship. Funding for the 300-MHz NMR spectrometers of the Oklahoma Statewide Shared NMR Facility was provided by NSF (BIR-9512269), the Oklahoma State Regents for Higher Education, the W.M. Keck Foundation, and Conoco, Inc. The authors also wish to thank the OSU College of Arts and Sciences for funds to upgrade our departmental FT-IR and GC-MS instruments.
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