Synthesis, unambiguous chemical characterization, and reactivity of 2,3,4,5-tetrahydro-1,5-benzoxazepines-3-ol

Abstract

The previous structures of several 3,4-dihydro-2H-3-hydroxymethyl-1,4-benzoxazines were erroneous (Bioorg. Med. Chem. Lett. 2006, 16, 2862; Synlett 2009, 3283 and Chinese Patent 2006-10043245). The results presented here are both detailed NMR experiments and synthetic aspects on this subject. We redress some points for the preparation of intermediates of the target molecules.

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Introduction

The 2,3-dihydro-1,4-benzodioxin ring 1a (Fig. 1) constitutes an important skeletal fragment in medicinal chemistry.¹ We have recently reviewed the chemistry of 2,3-dihydro-1,4 benzodioxins and bioisosteres as structural motifs for biologically active compounds.² We used the bioisosteres 3,4-dihydro-2H-1,5-benzoxathiepine-3-ol 2 and 2,3-dihydro-1,4-benzoxathiine-2-methanol 3 (Fig. 1) as intermediates to obtain a series of substituted-9-(2,3-dihydro-1,4-benzoxathiine-3-and -2-ylmethyl) purine derivatives with notable anti-proliferative activities.^3,4 Benzoxazepines are a well-known pharmacophore in medicinal chemistry showing promising activity against various diseases such as antipsychotic, central nervous system activity along with anti-cancer profile against breast cancer cells.^5,6

Fig. 1 Structures of benzo-fused six- and seven-membered derivatives.

Nitrogen-containing compounds are important chemical entities, many of which show unique bioactivity. 3,4-Dihydro-2H-1,4-benzoxazines have been used as a substructure of several biologically interesting agents between them.^7–12

Following a bioisosteric replacement as a useful strategy for the structural modification of compounds 1b,¹³2 and 3, we wish to report herein the synthesis and the correct chemical characterization of compounds 4a–f and 5a–c. We will discuss the misunderstanding in the scientific bibliography between the six-membered structures 4a–f and the seven-membered ones 5a–c, and the flaws encountered by us in the synthetic protocols that lead to both intermediates 4a and 5a and target molecules. Finally, we describe the Mitsunobu reaction³ of 5a with several substituted purines to obtain 4d–f (Fig. 2).

Fig. 2 Molecules discussed in this manuscript.

Results and discussion

After a search in literature we found two manuscripts dealing with the synthesis of 3,4-dihydro-2H-3-hydroxymethyl-1,4-benzoxazine (4a): (a) from 6 in the presence of Hantzsch 1,4-dihydropyridine (HEH) and Pd/C as catalyst¹⁴ (84%); (b) through a tandem reduction-oxirane opening of 1,2-epoxy-3-(o-nitrophenoxy)propane (6), in the presence of iron powder and acetic acid at reflux,¹⁵ or using Fe or Zn powder as reducing agent, and acetic acid or ammonium chloride as the acidic medium at the reflux temperature of H₂O/EtOH.¹⁶ According to them,^14–16 the nitro group was reduced, leading to the attack of the amino group on the epoxide moiety in a 6-exo-tet fashion, according to the Baldwin rules.¹⁷

The preparation of 6 was published by Xu et al.¹⁸ Although they claim that 6 was obtained as a light yellow solid in a 35% yield after recrystallization (EtOH), we did not reproduce their results on repeating their experimental conditions and 7 (82%) was obtained instead as a yellow syrup (Scheme 1). Finally, 7 was subjected to an epoxide-forming process by using sodium hydride (1 equiv.) in anhydrous THF under an inert atmosphere (Scheme 1) and a very pure compound 6 (73%) was obtained as a light yellow solid. The synthetic procedure reported¹⁹ to 6 is different than that stated previously.

Scheme 1 Reactivity of o-nitrophenol: (i) Conditions previously reported:¹⁸ epichlorohydrin (5 equiv.), NaOH (0.25 equiv.), deionized H₂O, 85 °C, 2 h; (ii) NaH (1 equiv.), anhydrous THF, rt, instantaneous reaction; (iii) epichlorohydrin (5 equiv.), NaOH (1 equiv.), deionized H₂O, 85 °C, 36 h.

As we were surprised by the fact that Xu et al.¹⁸ only used 4 fold-less equivalents of the base in relation to o-nitrophenol, we decided to equalize both equivalents (1 equiv. of both o-nitrophenol and NaOH, but keep 5 equiv. of epichlorohydrin), and to increase the reaction time to 36 h, maintaining the reaction temperature at 85 °C (Scheme 1). The expected compound 6 was obtained in a fairly good yield (64%), together with a low yield (28%) of the propylene chlorohydrin 7.

In order to speed up the process of formation of 6, we used microwave irradiation and we obtained 7 (25%) and 6 (64%). When epichlorohydrin and o-nitrophenol were reacted in equimolecular quantities under MW irradiation at 95 °C for 40 min, the product of the reaction was the commercially available 1,3-bis-(o-nitrophenoxy)-2-propanol (8, 83%).

Nevertheless, the two following facts drew our attention after the cyclization reaction from epoxide 6: (a) two compounds were obtained, whilst the exclusive formation of w4a¹⁵ was published (See Scheme 2 to understand this terminology) in 84% yield; and (b) the ¹H NMR spectrum of the mixture shows the presence of two isomers, as demonstrated by the presence of several sets of signals, each one composed of two splitting patterns.

Scheme 2 Regioselective reaction of the seven-membered ring 5a: i) according to reference 15 ii) Pd/C, EtOH, reflux, 12 h, inert atmosphere. The identification tag w4a¹⁵ denotes the wrong 4a with its bibliographic reference as superscript: it means “the incorrect structure 4a, described in the corresponding ref. 14, 15 or 16 but that is actually 5a”.

The major isomer was identified as 2,3,4,5-tetrahydro-1,5-benzoxazepine-3-ol (5a, 72%) and the minor one was 3,4-dihydro-2H-3-hydroxymethyl-1,4-benzoxazine (4a, 14%) (the ratio between 5a/4a is 5 : 1). We then decided to carry out a shorter synthesis of 4a and 5a in a one-pot process. Starting from o-aminophenol (1 equiv.) and epichlorohydrin (1 equiv.), heating under MW-assisted conditions (160 °C, 30 min) produced a mixture of 4a (34%) and 5a (55%). Our complete ¹H and ¹³C assignment is based on ¹H signal integrals, multiplicity patterns, a DEPT(135) experiment, and HSQC/HMBC correlations. As the assignment is a routine task, only those points which are crucial to the unambiguous assignment of the previously incorrect structure are emphasized.

The separation of 4a and 5a was carried out by HPLC [a mixture of H₂O/MeCN (90/10) as eluent], semi-preparative column Nova-Pak C-18, 60 Å, 6 μm, 17.8 mm × 300 mm, retention time (t_R) of 4a: 6.96 min, and t_R of 5a: 7.27 min. The ¹H and ¹³C NMR spectra (500 MHz, CDCl₃), showed an important mistake, which is one of the key problems for the incorrect structure assignment of w4a.^14–16 Zhao, Miao et al.¹⁵ literally state the following: The structure ofw4a¹⁴was confirmed by¹H NMR and¹³C NMR data, which showed the presence of two CH₂–O signals at δ 75.2 and δ 68.4 ppm, and a CH–N signal at δ 52.2 ppm (assigned to C-3), confirming the formation of only one regioisomer. The one-pot synthesis provides a novel and facile method of preparing the 2,3-dihydro-3-hydroxymethyl-1,4-benzoxazine derivatives.

In the hetero-methylene zone of the ¹³C NMR spectrum we found the three signals at δ 75.3, 68.5 and 52.3 ppm, similar to those reported.¹⁵ In order to confirm these data and according to the DEPT(135) experiment we found that the signals at δ 75.3 and δ 52.3 ppm are negative peaks, and therefore can be assigned to CH₂–O and CH₂–N signals, whereas the signal at δ 68.5 ppm is positive and can be assigned to the CH–O moiety. Accordingly, the presence of one CH₂–O, one CH₂–N, and one CH–O group is compatible with 5a. Three years later Liu and Zhou et al.¹⁴ committed the same mistake with the incorrect structure w4a¹⁴ based on the same spectroscopic data already reported.¹⁵ Following a similar reasoning for 4a, we found two negative CH₂–O signals at δ 66.1 ppm and δ 63.0 ppm, and one positive (CH–N) at δ 51.4 ppm. This structure was also confirmed with HSQC and HMBC experiments to unambiguously establish the structures 4a and 5a (Fig. 3).

Fig. 3 Representation of the HMBC interactions observed in the six- and seven-membered rings (4a and 5a).

In the HMBC experiment to three bonds, the interaction between the hydrogen atoms of the geminal H-2 and H-2′ with the quaternary C-8a of the benzene ring should be observed in 4a, whilst interactions between H-2 [δ 4.25 (ddd)] and H-2′ [δ 3.86 (dd)], and H-4 [δ 3.36 (ddd)] and H-4′ [δ 3.17 (dd)], with the quaternary C-9a (δ 150.7 ppm) and C-5a (δ 142.0 ppm) respectively, are unequivocal proofs for the structure of 5a (Fig. 3).

Both NMR data and the structure of the alcohol obtained fit in with 5a but not with the reported results published.^14–16 To reinforce our conclusions even more, we made a deep bibliographic search and found a patent, written in Japanese, in which the synthesis and ¹H NMR data of 4a were correctly reported.²⁰

Having found controversial results between our research and that published,^14–16 we decided to carry out the reduction of 6 under the conditions previously reported (Fe/HOAc, EtOH/H₂O at reflux).¹⁵ This procedure provides 5a and not 4a as stated, which we confirmed after an in-depth NMR study and the unambiguous demonstration of the 5a structure, in spite of the fact that it was published as w4a.^14–16 Compounds w4b,c were prepared according to a reported procedure,¹⁵ and we found that these compounds were actually 5b,c based on DEPT(135) data: 5b and 5c show peaks at δ 52.10 and at δ 52.13 ppm (N–CH₂), respectively and negative peaks at δ 75.12 ppm and at δ 75.18 ppm (O–CH₂) while the positive peaks at δ 68.30 ppm (O–CH) for both 5b and 5c are unambiguously assigned to the methine carbons. Table 1 shows the correct chemical shifts, multiplicity patterns and coupling constants (J) in Hz of compounds 5a–c and 4a.¹⁵

Table 1 Comparison of the chemical shifts (δ), multiplicity patterns and coupling constants (J, Hz) of the ¹H NMR spectra for compounds 4a, 5a and w4a,b,c

Compd	R	H-2	H-2′	H-3	H-4	H-4′
5a	H	4.25 (ddd)	3.86 (dd)	3.93 (m)	3.36 (ddd)	3.17 (dd)
		J = 12.3, 3.8, 1.4	J = 12.3, 2.0		J = 12.9, 4.9, 1.4	J = 12.9, 2.4
4a	H	4.18 (dd)	4.06 (dd)	3.55 (m)	3.72 (dd)	3.64 (dd)
		J = 10.8, 2.9	J = 10.8, 5.9		J = 10.7, 4.9	J = 10.7, 7.1
w4a	H	4.26 (dd)	3.88 (dd)	3.91–3.97 (m)	3.36 (dd)	3.19 (dd)
		J = 12.8, 3.8	J = 12.8, 2.0		J = 12.9, 4.8	J = 12.9, 2.4
w4b	7-Me	4.25 (dd)	3.80(dd)	3.89–3.95 (m)	3.37 (dd)	3.14 (dd)
		J = 12.3, 2.7	J = 12.3, 1.5		J = 12.8, 3.9	J = 12.8, 1.9
w4c	7-tBu	4.25 (dd)	3.84 (dd)	3.89–3.97 (m)	3.39 (dd)	3.18 (dd)
		J = 12.3, 3.7	J = 12.3, 1.9		J = 12.8, 4.7	J = 12.8, 2.2

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In spite of the fact that Liu and Zhou¹⁴ reported the incorrect structure of w4a, the formation of six- and/or seven-membered alcohols starting from di-or three-substituted epoxides is accurate. In these cases 2D NMR data correctly supported the target structures and the aromatic splitting patterns comply with our above-mentioned rules.

Conclusively and to emphasize the reactivity of the seven-membered secondary alcohol 5a, its reaction with several purines under the microwave-mediated Mitsunobu conditions was studied (Scheme 3).

Scheme 3 Reagents and conditions: substituted purines, Ph₃P, DIAD, anhydrous THF, microwave irradiation, 140 °C, 5 min.

The product of the three reactions are the six-membered compounds 4d–f (3,4-dihydro-2H-3-hydroxymethyl-1,4-benzoxazines linked to the N-9′ purine atom through a methylene linker), demonstrated by the 2D NMR data. The ring contraction can be explained through an aziridinium ion, as previously published in a similar reaction via an episulfonium intermediate.³ The anticancer activity and study of the mechanism of action of 4d–f are underway.

In summary, 2,3,4,5-tetrahydro-1,5-benzoxazepines-3-ol 5a–c have not been reported yet because they were mistaken as being the 3,4-dihydro-2H-3-hydroxymethyl-1,4-benzoxazines 4a–c.¹⁴ We have also improved the method of synthesis of 4a and 5a, in a one-step and one-pot reaction from o-aminophenol and epichlorohydrin under microwave irradiation. Compounds 5a–c may serve as synthons for the development of new biologically active prototypes. Although six-membered rings are more stable than their seven-membered counterparts, and in general favored when there are both possibilities of formation, the use of the DEPT, HSQC and HMBC techniques would have made clear the correct assignment of the target scaffold.

We wish to help people working on benzo-fused six- and seven-membered diheterocycles to facilitate their research after having made clear and unambiguous the differences between these two scaffolds. Clarification of the confusion reported in previous works will prevent the proliferation of scientific mistakes and consequently avoid a negative domino effect due to the rapid divulgation of information nowadays.

Experimental section

Melting points were taken in open capillaries on an electrothermal melting point apparatus and are uncorrected. Analytical thin layer chromatography was performed using Merck Kieselgel 60 F254 aluminum sheets, the spots being developed with UV light (λ = 254 nm). Evaporations were carried out in vacuo with a Büchi rotary evaporator and the pressure was controlled by a Vacuubrand CVCII apparatus. For flash chromatography, Merck silica gel 60 with a particle size of 0.040–0.063 mm (230–400 mesh ASTM) was used. Nuclear magnetic resonance spectra have been carried out at the Centro de Instrumentación Científica/Universidad de Granada, and recorded on a 500 MHz ¹H and 125 MHz ¹³C NMR DEPT, HMBC and HSQC NMR Varian NMR-System-TM400 (compounds 4d–f,5a) or 400 MHz 1H and 100 MHz ¹³CNMR and DEPT NMR Varian Inova-TM spectrometers (compounds 6–8) at ambient temperature. Chemical shifts (δ) are quoted in parts per million (ppm) and are referenced to the residual solvent peak. Signals are designated as follows: s, singlet; d, doublet; dd, doublet of doublets; ddd, double doublet of doublets; dt, doublet of triplets; t, triplet; m, multiplet. The HMBC spectra were measured using a pulse sequence optimized for 10 Hz (inter-pulse delay for the evolution of long-range couplings: 50 ms) and a 5/3/4 gradient combination. In this way, direct responses (J couplings) were not completely removed. High-resolution nano-assisted laser desorption/ionization (NALDI-TOF) or electrospray ionization (ESITOF) mass spectra were carried out on a Bruker Autoflex or a Waters LCT Premier Mass Spectrometer, respectively. Small scale microwave-assisted synthesis was carried out in an Initiator 2.0 single-mode microwave instrument producing controlled irradiation at 2.450 GHz (Biotage AB, Uppsala). Reaction time refers to hold time at 140 °C, not to total irradiation time. The temperature was measured with an IR sensor outside the reaction vessel. Anhydrous THF was purchased from VWR International Eurolab. Anhydrous conditions were performed under argon. 6-Chloropurine, 6-bromopurine and 2,6-dichloropurine were purchased from Aldrich. Compounds 4d–f were synthesized according to our previously reported procedure.³ An HPLC apparatus and a fluorescence detector were used for separation of 4a and 5a (semi-preparative column C-18, 60 Å, 6 μm , 17.8 mm × 300 mm).

1,2-Epoxy-(o-nitrophenoxy)propane (6) and 1-(o-nitrophenoxy)-3-chloro-2-propanol (7)

Method a): preparation of 6 and 7. The general procedure below was previously reported,¹⁸ but our experimental results were different (see Scheme 1): Xu et al.¹⁸ described the formation of 6 in a 35% yield as a yellow solid, but we obtained 7 (82%) as a yellowish syrup. ¹H NMR (400 MHz, CDCl₃) δ 7.71 (d, J = 8.1 Hz, 1H), 7.43 (t, J = 7.9, 1H), 7.01 (d, J = 8.4 Hz, 1H), 6.93 (t, J = 7.7 Hz, 1H), 4.17 (m, 3H), 3.69 (m, 2H), 3.60 (d, J = 5.6 Hz, 1H). ¹³C NMR (100 MHz, CDCl₃) δ 151.7, 139.1, 134.6, 125.6, 120.9, 114.8, 69.9, 69.3, 45.3. HR TOF MS ES+, calculated for C₉H₁₁ClNO₄ 232.0377, found C₉H₁₁ClNO₄ 232.0377. Anal. Calcd for C₉H₁₀ClNO₄: C, 46.67; H, 4.35; N, 6.05. Found: C, 46.89; H, 4.33; N, 5.89. HR TOF MS ES+.

Method b): preparation of 6 and 7. Following the general procedure,¹⁸ but increasing from 0.25 equiv. to 1 equiv. of NaOH, 6 (64%) and 7 (28%) were obtained (see Scheme 1) as a yellowish solid (mp 81.5–82 °C, lit.⁴² 79 °C) and a yellowish syrup, respectively. Compound 6: ¹H NMR (400 MHz, CDCl₃) δ 7.84 (dd, J = 8.1, 1.3 Hz, 1H), 7.57 (dt, J = 8.2, 1.2 Hz, 1H), 7.19 (d, J = 8.5 Hz, 1H), 7.09 (t, J = 7.9 Hz, 1H), 4.51 (dd, J = 11.3, 2.1 Hz, 1H), 4.11 (dd, J = 11.3, 5.6 Hz, 1H), 3.42 (m, 1H), 2.91 (m, 2H). ¹³C NMR (100 MHz, CDCl₃) δ 151.4, 139.5, 134.1, 125.1, 120.6, 114.8, 69.6, 49.5, 43.9. HR TOF MS ES+, calcd for C₉H₉NO₄Na (M + Na)⁺ 218.0429, found 218.0427. Anal. Calcd for C₉H₉NO₄: C, 55.39; H, 4.65; N, 7.18. Found: C, 55.23; H, 4.29; N, 7.00.

1,2-Epoxy-(o-nitrophenoxy)propane (6) from 1-(o-nitrophenoxy)-3-chloro-2-propanol (7)

To a solution of 7 (2.175 g, 9.3 mmol) in anhydrous THF (30 mL) under argon, 60% NaH (0.373 g, 9.3 mmol) was added dropwise at room temperature. The reaction changed instantaneously from a yellow to a cloudy orange color. 2N HCl was added at pH = 6–7, the mixture was extracted (H₂O and CH₂Cl₂), the organic layers decanted, dried (Na₂SO₄) and filtered to give 6 (1.33 g, 73%, Scheme 1).

1,2-Epoxy-(o-nitrophenoxy)propane (6), 1-(o-nitrophenoxy)-3-chloro-2-propanol (7) and 1,3-bis(o-nitrophenoxy)-2-propanol (8) under microwave irradiation

Epichlorohydrin was added to a solution of o-nitrophenol and aqueous NaOH. The reaction was irradiated in a microwave oven at 95 °C for 40 min. The mixture was cooled and poured into ice water, and the aqueous layer was extracted with CH₂Cl₂. Organic layers were combined and dried (Na₂SO₄). After filtration, the solvent was removed in vacuo and the crude was purified by gradient-elution flash column chromatography using hexane/EtOAc (9/1 → 8/2 → 0/10) mixtures.

Method a): preparation of 6 and 7. After using 1 equiv. of o-nitrophenol, 2 equiv. of epichlorohydrin, and 1.25 equiv. of NaOH in 7 mL of H₂O and following the general method, 6 (64%) and 7 (25%) were obtained.

Method b): preparation of 8

Same conditions as method a), except that 2.5 equiv. of NaOH were used and following the general method, 8 (83%) was obtained as a yellowish syrup. ¹H NMR (400 MHz, CDCl₃) δ 7.82 (d, J = 8.1 Hz, 1H), 7.50 (t, J = 7.5 Hz, 1H), 7.07 (d, J = 8.3 Hz, 1H), 7.00 (t, J = 7.7, 1H), 4.14 (m, 3H), 3.79 (m, 2H). ¹³C NMR (100 MHz, CDCl₃) δ 152.3, 139.6, 134.9, 126.1, 121.1, 115.0, 71.3, 70.0. HR TOF MS ES+, calcd for C₁₅H₁₄N₂O₇Na (M + Na)⁺ 357.0699, found 357.0694. Anal. Calcd for C₁₅H₁₄N₂O₇: C, 53.89; H, 4.22; N, 8.38. Found: C, 53.97; H, 4.12; N, 8.61.

2,3,4,5-Tetrahydro-1,5-benzoxazepine-3-ol (5a) and 3,4-dihydro-2H-3-hydroxymethyl-1,4-benzoxazine (4a)

Method a) (starting from 6). Following exactly the procedure formerly described,¹⁵4a (14%) and 5a (72%) were obtained as brownish syrups in both cases, and not w4a,¹⁵ as stated.¹⁵ Both compounds were separated by HPLC [a mixture of H₂O/MeCN (100/0 → 0/100) as eluent]. Compound 4a: ¹H NMR (500 MHz, CDCl₃) δ 6.76 (m, 2H), 6.63 (m, 2H), 4.18 (dd, J = 10.8, 2.9 Hz, 1H), 4.06 (dd, J = 10.8, 5.9 Hz, 1H), 3.72 (dd, J = 10.7, 4.9 Hz, 1H), 3.64 (dd, J = 10.7, 7.1 Hz, 1H), 3.55 (m, 1H). ¹³C NMR (125 MHz, CDCl₃) δ: 144.1, 132.9, 121.9, 119.3, 116.9, 116.1, 66.1, 63.0, 51.4. HR TOF MS ES+, calcd for C₉H₁₂NO₂ (M + H)⁺ 166.0868, found 166.0866. Anal. Calcd for C₉H₁₁NO₂: C, 65.44; H, 6.71; N, 8.48. Found: C, 65.30; H, 6.85; N, 8.49. Compound 5a: ¹H NMR (500 MHz, CDCl₃) δ 6.99 (dd, J = 7.8, 1.5 Hz, 1H), 6.89 (dt, J = 7.5, 1.5 Hz, 1H), 6.82 (dt, J = 7.6, 1.7 Hz, 1H), 6.75 (dd, 1H), 4.25 (ddd, J = 12.3, 3.8, 1.4 Hz, 1H), 3.93 (m, 1H), 3.86 (dd, J = 12.3, 2.0 Hz, 1H), 3.36 (ddd, J = 12.9, 4.9, 1.4 Hz, 1H), 3.17 (dd, J = 12.9, 2.4 Hz, 1H).¹³C NMR (125 MHz, CDCl₃) δ 150.7, 142.0, 124.1, 122.2, 122.1, 120.0, 75.4, 68.6, 52.4. HR TOF MS ES+, calcd for C₉H₁₂NO₂ (M + H)⁺ 166.0868, found 166.0868. Anal. Calcd for C₉H₁₁NO₂: C, 65.44; H, 6.71; N, 8.48. Found: C, 65.42; H, 6.54; N, 8.69.

Method b) (starting from o-aminophenol). A mixture of o-aminophenol (0.2 g, 1.83 mmol), 60% NaH (0.07 g, 1.83 mmol) and anhydrous DMF (6 mL) was stirred at room temperature for 10 min, and irradiated in a microwave oven at 160 °C for 20 min. Epichlorohydrin (0.17 g, 1.83 mmol) was added and the resulting mixture was irradiated at 160 °C for an additional 10 min. The mixture was cooled, the DMF was removed in vacuo, and the residue was extracted (H₂O/CH₂Cl₂). Organic layers were combined and dried (Na₂SO₄). After filtration, the solvent was removed in vacuo and the crude was purified by flash column chromatography using an hexane/EtOAc (8/2) mixture. Both compounds, 4a (34%) and 5a (55%) were separated by HPLC [a mixture of H₂O/MeCN (90/10) as eluent], semi-preparative column Nova-Pak C-18, 60 Å, 6 μm, 17.8 mm × 300 mm, retention time (t_R) of 4a: 6.96 min, and t_R of 5a: 7.27 min.

6-Chloro-9-(2,3-dihydro-1,4-benzoxazine-3-ylmethyl)-9H-purine (4d)

Yellowish solid (14%), (mp 77–78 °C), ¹H NMR (500 MHz, CDCl₃) δ 8.74 (s, 1H), 8.06 (s, 1H), 6.82 (m, 2H), 6.70 (td, J = 7.8, 1.5 Hz, 1H), 6.58 (dd, J = 7.8, 1.5 Hz, 1H), 4.49 (dd, J = 14.1, 5.2 Hz, 1H), 4.36 (dd, J = 14.1, 8.2 Hz, 1H), 4.13 (ddd, J = 25.6, 11.1, 2.7 Hz, 2H), 4.03 (m, 1H), 3.96 (s, 1H). ¹³C NMR (125 MHz, CDCl₃) δ 152.36, 152.20, 151.64, 146.29, 143.52, 132.08, 131.15, 122.78, 119.80, 117.55, 116.33, 65.44, 49.46, 46.53. HR TOF MS ES+, calcd for C₁₄H₁₃ClN₅O (M + H)⁺ 302.0808, found 302.0808. Anal. Calcd for C₁₄H₁₂ClN₅O: C, 55.73; H, 4.01; N, 23.21. Found: C, 55.70; H, 4.05; N, 23.50.

6-Bromo-9-(2,3-dihydro-1,4-benzoxazine-3-ylmethyl)-9H-purine (4e)

Yellowish solid (12%), (mp 72–73 °C), ¹H NMR (500 MHz, CDCl₃) δ 8.70 (s, 1H), 8.07 (s, 1H), 6.82 (m, 2H), 6.71 (td, J = 7.8, 1.5 Hz, 1H), 6.58 (dd, J = 7.8, 1.5 Hz, 1H), 4.49 (dd, J = 14.1, 5.2 Hz, 1H), 4.35 (dd, J = 14.1, 8.3 Hz, 1H), 4.13 (ddd, J = 26.6, 11.1, 2.7 Hz, 2H), 4.02 (m, 1H), 3.90 (s, 1H). ¹³C NMR (125 MHz, CDCl₃) δ 152.21, 150.84, 150.71, 146.05, 143.69, 134.60, 131.00, 122.69, 119.75, 117.48, 116.27, 65.33, 49.37, 46.46. HR TOF MS ES+, calcd for C₁₄H₁₃BrN₅O (M + H)⁺ 346.0303, found 346.0305. Anal. Calcd for C₁₄H₁₂BrN₅O: C, 48.57; H, 3.49; N, 20.23. Found: C, 48.61; H, 3.55; N, 20.00.

2,6-Dichloro-9-(2,3-dihydro-1,4-benzoxazine-3-ylmethyl)-9H-purine (4f)

White solid (15%), (mp 89–90 °C), ¹H NMR (500 MHz, CDCl₃) δ 8.03 (s, 1H), 6.82 (m, 2H), 6.71 (td, J = 7.7, 1.5 Hz, 1H), 6.59 (dd, J = 7.8, 1.5 Hz, 1H), 4.46 (dd, J = 14.1, 5.2 Hz, 1H), 4.32 (dd, J = 14.1, 8.3 Hz, 1H), 4.17 (dd, J = 11.2, 2.5 Hz, 1H), 4.08 (dd, J = 11.2, 2.6 Hz, 1H), 4.00 (s, 1H), 3.88 (s, 1H). ¹³C NMR (125 MHz, CDCl₃) δ 153.44, 153.33, 152.26, 146.91, 143.41, 131.13, 130.78, 122.77, 119.87, 117.53, 116.39, 65.20, 49.25, 46.45. HR TOF MS ES+, calcd for C₁₄H₁₂Cl₂N₅O (M + H)⁺ 336.0419, found 336.0419. Anal. Calcd for C₁₄H₁₁Cl₂N₅O: C, 50.02; H, 3.30; N, 20.83. Found: C, 49.90; H, 3.35; N, 20.91.

Acknowledgements

This study was supported by the Instituto de Salud Carlos III (FIS) through project no. PI10/00592. The M.E-G.R. FPU grant AP2007-02954 is greatly acknowledged.

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Footnote

† Electronic supplementary information (ESI) available: Copies of the ¹H, ¹³C NMR, and DEPT spectra of 4a,d–f, 5a, 6, 7, and 8, HSQC and HMBC charts for 4a,d–f and 5a are provided. See DOI: 10.1039/c2ra21706f

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