O-Alkylation of 3-hydroxyisoxazoles predominates under Mitsunobu conditions

Article abstract of DOI:10.1016/j.tetlet.2014.01.065

Regiochemical control in the functionalization of ambident nucleophiles is of particular interest in organic chemistry. Herein, we demonstrate that O-alkylation of ambident 3-hydroxyisoxazoles, which are heterocyclic bioisosteres of carboxylic acids, predominates under Mitsunobu conditions. In several cases, excellent O-regioselectivity (≥95%) was observed. It is noteworthy that reactions were complete within 15 min at room temperature. Furthermore, the conditions are compatible with a range of alcohols that cover all of the typical protecting groups for the 3-hydroxyisoxazole motif, providing milder, simpler and less hazardous protocols to those commonly followed in the literature.

Full text of DOI:10.1016/j.tetlet.2014.01.065

Tetrahedron Letters 55 (2014) 1693–1696
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
O-Alkylation of 3-hydroxyisoxazoles predominates under Mitsunobu
conditions
Lijia Chen a, Steven Fletcher a,b,
⇑
a Department of Pharmaceutical Sciences, University of Maryland School of Pharmacy, 20 N. Pine Street, Baltimore, MD 21201, USA
b University of Maryland Greenebaum Cancer Center, 22 S. Greene Street, Baltimore, MD 21201, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Regiochemical control in the functionalization of ambident nucleophiles is of particular interest in
organic chemistry. Herein, we demonstrate that O-alkylation of ambident 3-hydroxyisoxazoles, which
are heterocyclic bioisosteres of carboxylic acids, predominates under Mitsunobu conditions. In several
cases, excellent O-regioselectivity (P95%) was observed. It is noteworthy that reactions were complete
within 15 min at room temperature. Furthermore, the conditions are compatible with a range of alcohols
that cover all of the typical protecting groups for the 3-hydroxyisoxazole motif, providing milder, simpler
and less hazardous protocols to those commonly followed in the literature.
Received 19 November 2013
Revised 13 January 2014
Accepted 15 January 2014
Available online 23 January 2014
Keywords:
Mitsunobu reaction
3-Hydroxyisoxazole,
3-isoxazolol, isoxazole-3-one
ambident (pro-)nucleophile
Regioselective
Ó 2014 Elsevier Ltd. All rights reserved.
Bioisostere
3-Hydroxyisoxazoles (3-isoxazolols or isoxazole-3-ones) feature
in naturally occurring compounds and are useful building blocks in
the construction of pharmaceuticals.1 For example, muscimol (1)
and ibotenic acid (2) are psychoactive natural products,2 and a vari-
ety of related 3-hydroxyisoxazoles have been developed that exhi-
bit excellent central nervous system (CNS) activity.3,4 Bearing a
similar pKa, the 3-hydroxyisoxazole motif is bioisosteric with the
carboxylic acid function and may evade some of the metabolic fates
suffered by carboxylic acids, further adding to its utility in drug
design.5 Moreover, whilst a carboxylic acid represents the terminus
to a drug molecule, a 3-hydroxyisoxazole motif can be incorporated
within a drug molecule (Fig. 1B), allowing for functionality to be
added either side of the acidic function analogous to the acylsulf-
onamide in the Bcl-2/Bcl-xL inhibitor ABT-737 (3).6a Indeed, as part
of our research program to develop inhibitors of the oncoprotein
Mcl-1,6b we have enlisted the 3-hydroxyisoxazole heterocycle as
an acidic scaffold designed to bind Arg263, whilst substitutions at
both the 4- and 5-positions are intended to target hydrophobic do-
mains flanking Arg263. In our hands, however, protection of the
acidic hydroxyl was non-trivial and afforded the desired products
in a variety of yields, prompting the present research.
(RHal, K2CO3, room temperature (rt)) frequently results in mixtures
of O- and N-alkylated products.7 3-Hydroxyisoxazoles have a pKa of
approximately 5 and are, therefore, suitable pronucleophiles for
the Mitsunobu reaction with the triphenylphosphine (PPh3)/di-iso-
propyl azodicarboxylate (DIAD) reagent system, wherein the pKa of
the pronucleophile must be around 12 or below for a successful
reaction to occur.8 Encouraged by our recent findings that the
regioselective N-alkylation of the benzodiazepine-2,5-dione ani-
lide function can be effectively controlled by the Mitsunobu reac-
tion,9 coupled with a similar study on pyridone,10 we wondered
if selective O/N-alkylation of 3-hydroxyisoxazoles might likewise
be mastered. To the best of our knowledge, although there is prec-
edent for Mitsunobu chemistry with 3-hydroxyisoxazoles,11 such a
study has not been reported. By analysing the pronucleophile, alco-
hol and reaction solvent, we herein demonstrate the utility of the
Mitsunobu reaction in the regioselective O-alkylation of 3-hydrox-
yisoxazoles. Mild reaction conditions, swift reaction times and
yields of up to 91% render this chemistry an attractive means
towards the protection/functionalization of this acidic heterocycle.
Employing a slight excess (1.3 equiv) of PPh3 and DIAD in THF,
which represent the most popular Mitsunobu reagents and reac-
tion solvent, we found that the coupling reaction between 5-phe-
nyl-3-hydroxyisoxazole (4 (R1 = Ph)) and isopropanol was
complete within 15 min at rt. Analysis of the 1H NMR of the crude
reaction mixture revealed a ratio of 6.7:1 for the O-isopropyl prod-
uct 5 to the regioisomeric N-isopropyl product 6 (Table 1, entry 1).
The O:N ratio was determined by comparison of the integrations of
Owing to its ambident nature, synthetic modification of the
3-hydroxyisoxazole motif under classical alkylation conditions
⇑
Corresponding author. Tel.: +1 410 706 6361.
E-mail address: sfletche@rx.umaryland.edu (S. Fletcher).
http://dx.doi.org/10.1016/j.tetlet.2014.01.065
0040-4039/Ó 2014 Elsevier Ltd. All rights reserved.

1694
L. Chen, S. Fletcher / Tetrahedron Letters 55 (2014) 1693–1696
Table 1
Mitsunobu reactions of 3-hydroxyisoxazoles with alcohols R2OHa
A
R1
R2OH
5b (%)
5:6c
B
Entry
1
82
6.7:1
2
3
4
45
ND
33
1:1
7.3:1
1.7:1
C
5
6
7
8
84
65
70
84
13:1
2.3:1
2.3:1
19:1
MeOH
9
60
13:1
3:1
Figure 1. (A) The structures of muscimol (1) and ibotenic acid (2). (B) The
3-hydroxyisoxazole motif allows the incorporation of an acidic function within a
drug molecule, in contrast to its bioisosteric carboxylic acid motif. (C) The structure
of ABT-737.
10
18d
ND, not determined.
the C4-H protons of 5 and 6 (dH (DMSO-d6) 6.73 (O), 6.43 (N)). In
addition, after purification of the reaction mixture by silica gel
flash column chromatography, the identity of the faster-running
product (Rf 0.68 (Hex/EtOAc, 1:1)) was confirmed to be O-isopro-
pyl-5 through comparison of the NMR spectral data with an
authentic sample that was prepared unambiguously.13 The more
polar N-regioisomer 6 (Rf 0.15 (Hex/EtOAc, 1:1)) was readily sepa-
rated from 5 owing to the large difference in Rf.
a
Reaction conditions:12 3-hydroxyisoxazole (1 mmol), R2OH (1.25 mmol) and
PPh3 (1.3 mmol) were dissolved in anhydrous THF (14 mL) at rt. After 2 min, DIAD
(1.3 mmol) was added dropwise. The reaction mixture was stirred at rt under an
inert atmosphere until complete by TLC (typically within 15 min).
b
Isolated yield.
c
1H NMR-determined ratio of O:N products in crude reaction mixture.
d
Reaction time = 48 h.
In contrast to the reaction with isopropanol, no selectivity was
observed with benzyl alcohol (Table 1, entry 2). Similar ratios were
observed with 3-hydroxyisoxazole (4 (R1 = H); Table 1, entries 3
and 4). On the other hand, O-alkylation predominated with methyl
3-hydroxyisoxazole-5-carboxylate (4 (R1 = CO2Me); Table 1,
entries 5–10), demonstrating the structure of the 3-hydroxyisoxaz-
ole, to some degree, dictates the regiochemistry in the Mitsunobu
reaction. In the context of methyl 3-hydroxyisoxazole-5-carboxyl-
ate, the preference for O-alkylation may result from reduced
electron density on the nitrogen atom due to the electron-
withdrawing methyl ester at C5. As shown by entry 10 in Table 1,
the coupling of methyl 3-hydroxyisoxazole-5-carboxylate with
tert-butanol generated a notable yield of O-tert-butyl-5. This
finding is especially striking since tertiary alcohols do not usually
participate in the Mitsunobu reaction due to steric hindrance,
although this transformation under Mitsunobu conditions has
been reported on occasion.14 In addition to comparing the integra-
tions of the 1H NMR chemical shifts of the C4-H protons of regioi-
somers 5 (for example, dH (DMSO-d6) 7.04–7.11 for R1 = CO2Me)
and 6 (dH (DMSO-d6) 6.58–6.72 for R1 = CO2Me), O:N ratios were
also determined by comparisons of the integrations for O- and
N-methyl, methylene or methine protons originating from the
alcohol R2OH, when not obscured by the isopropyl CH signals from
DIAD and DIAD-H2. These assignments were corroborated by
published 1H NMR spectral data where available.7,13,15 Due to large
differences in Rf (around 0.5 units), O-alkylated products 5 were
readily separated from their more polar N-regioisomers by column
chromatography.
We, and others, have shown that the reaction solvent can influ-
ence the regiochemical outcome of the Mitsunobu-mediated alkyl-
ation of ambident nucleophiles.9,10 In the present study, however,
we observed little solvent effect on the Mitsunobu reaction with
the pronucleophiles 3-hydroxyisoxazole and 5-phenyl-3-hydrox-
yisoxazole, although THF afforded a slight preference for O-alkyl-
ation relative to the chlorinated solvents and toluene (Table 2,
entries 1–6). In contrast, the coupling between methyl 3-hydrox-
yisoxazole-5-carboxylate and isopropanol gave the lowest O-selec-
tivity of 13:1, O:N in THF (Table 2, entry 7), whilst toluene
delivered the best O-selectivity of 32:1 (Table 2, entry 11) for the
panel of solvents evaluated. Surprisingly, the Mitsunobu reactions
in DMSO and CH3CN worked well, generating the O-alkylated prod-
uct 5 in high yields (Table 2, entries 10 and 12).
Standard protecting groups for 3-hydroxyisoxazoles include
methyl ether, isopropyl ether and benzyl ether. Treatment of
3-hydroxyisoxazoles with MeI often results in N-methylation or,
at best, a 1:1 mixture of the O and N regioisomers, whilst explosive
diazomethane promotes O-methylation to afford a 2:1 mixture of
O:N products.7 Regioselective O-isopropylation is accomplished
in very good yields with isopropyl halides, although heating may

L. Chen, S. Fletcher / Tetrahedron Letters 55 (2014) 1693–1696
1695
Table 2
mations typically being complete within 15 min at rt under essen-
tially neutral conditions, and, in the case of methylation, avoid the
use of hazardous diazomethane.
Solvent study of Mitsunobu reactions of 3-hydroxyisoxazoles with alcohols R2OHa
In summary, the Mitsunobu reaction of ambident 3-hydroxyis-
oxazole pronucleophiles with a panel of alcohols resulted in
O-alkylation as the predominant product in most cases with yields
of up to 91%. Notably, the structure of the 3-hydroxyisoxazole
played a role in the regiochemical control of the reaction with an
electron-withdrawing carboxylic ester at C5 particularly favouring
O-alkylation. THF proved a suitable reaction solvent to deliver high
O-selectivities (up to 13:1, O:N), whilst toluene may furnish even
greater O-selectivities with electron-withdrawing groups at posi-
tion 5 of the isoxazole ring (up to 32:1, O:N). Transformations were
typically complete within 15 min at rt, demonstrating the mildness
of the reaction conditions. Collectively, our findings provide com-
pelling reasons to adopt the Mitsunobu reaction as the preferred
chemistry to achieve the regioselective O-functionalization of
3-hydroxyisoxazoles.
Entry
1
R1
R2OH
Solvent
THF
5b (%)
5:6c
33
1.7:1
CHCl3
THF
39
45
42
28
26
1.3:1
1:1
2
3
4
5
6
CHCl3
CH2Cl2
Toulene
1:1.4
1:1.4
1:1.7
Acknowledgment
The authors wish to thank the University of Maryland School of
Pharmacy for financial support of this work.
References and notes
7
8
THF
84
91
85
79
85
82
66
72
13:1
13:1
19:1
19:1
32:1
24:1
2.3:1
3:1
1. (a) Bostrom, J.; Grant, J. A.; Fjellstrom, O.; Thelin, A.; Gustafsson, D. J. Med.
Chem. 2013, 56, 3273–3280; (b) Frydenvang, K.; Pickering, D. S.; Greenwood, J.
R.; Krosgaard-Larsen, N.; Brehn, L.; Nielsen, B.; Vogensen, S. B.; Hald, H.;
Kestrup, J. S.; Krogsgaard-Larsen, P.; Clausen, R. P. J. Med. Chem. 2010, 53, 8354–
8361; (c) Li-Fang, Y.; Zhang, H.-K.; Gunosewoyo, H.; Kozikowski, A. P. ACS Med.
Chem. Lett. 2012, 3, 1054–1058; (d) Baraldi, P. G.; Preti, D.; Zaid, A. N.;
Saponaro, G.; Tabrizi, M. A.; Baraldi, S.; Romagnoli, R.; Moorman, A. R.; Varani,
K.; Cosconati, S.; Di Maro, S.; Marinelli, L.; Novellino, E.; Borea, P. A. J. Med.
Chem. 2011, 54, 5205–5220.
2. Krogsgaard-Larsen, P.; Johnson, G. A. R.; Curtis, D. R.; Game, C. J. A.; McCulloch,
R. M. J. Neurochem. 1975, 25, 803–809.
3. Krogsgaard-Larsen, P.; Johnson, G. A. R.; Lodge, D.; Curtis, D. R. Nature 1977,
268, 53–55.
CHCl3
9
CH2Cl2
DMSO
Toulene
CH3CN
Toulene
Toulene
10
11
12
13
14
4. Frolund, B.; Jensen, L. S.; Guandalini, L.; Canillo, C.; Vestergaard, H. T.;
Kristiansen, U.; Nielsen, B.; Stensbol, T. B.; Madsen, C.; Krogsgaard-Larsen, P.;
Liljefors, T. J. Med. Chem. 2005, 48, 427–439.
5. Matzen, L.; Engesgaard, A.; Ebert, B.; Didriksen, M.; Frølund, B.; Krogsgaard-
Larsen, P.; Jaroszewski, J. W. J. Med. Chem. 1997, 40, 520–527.
MeOH
6. (a) Oltersdorf, T.; Elmore, S. W.; Shoemaker, A. R.; Armstrong, R. C.; Augeri, D. J.;
Belli, B. A.; Bruncko, M.; Deckwerth, T. L.; Dinges, J.; Hajduk, P. J.; Joseph, M. K.;
Kitada, S.; Korsmeyer, S. J.; Kunzer, A. R.; Letai, A.; Li, C.; Mitten, M. J.;
Nettesheim, D. G.; Ng, S.; Nimmer, P. M.; O’Connor, J. M.; Oleksijew, A.; Petros,
A. M.; Reed, J. C.; Shen, W.; Tahir, S. K.; Thompson, C. B.; Tomaselli, K. J.; Wang,
B.; Wendt, M. D.; Zhang, H.; Fesik, S. W.; Rosenberg, S. H. Nature 2005, 435,
677–681; (b) Friberg, A.; Vigil, D.; Zhao, B.; Daniels, R. N.; Burke, J. P.; Garcia-
Barrantes, P. M.; Camper, D.; Chauder, B. A.; Chauder, B. A.; Lee, T.; Olejniczak,
E. T.; Fesik, S. W. J. Med. Chem. 2013, 56, 15–30.
7. Schlewer, G.; Krogsgaard-Larsen, P. Acta Chem. Scand. B 1984, 38, 815–819.
8. (a) Mitsunobu, O.; Yamada, M.; Mukaiyama, T. Bull. Chem. Soc. Jpn. 1967, 40,
935; (b) Mitsunobu, O.; Eguchi, M. Bull. Chem. Soc. Jpn. 1971, 44, 3427.
9. (a) Fletcher, S.; Jung, K.-Y. Med. Chem. Commun. 2012, 3, 1160–1163; (b)
Fletcher, S. Tetrahedron Lett. 2010, 51, 2948–2950; (c) Fletcher, S.; Shahani, V.
M.; Gunning, P. T. Tetrahedron Lett. 2009, 50, 4258–4261.
Reaction conditions:12 3-hydroxyisoxazole (1 mmol), R2OH (1.25 mmol) and
PPh3 (1.3 mmol) were dissolved in anhydrous solvent (14 mL) at rt. After 2 min,
DIAD (1.3 mmol) was added dropwise. The reaction mixture was stirred at rt under
an inert atmosphere until complete by TLC (typically within 15 min).
a
b
Isolated yield.
1H NMR-determined ratio of O:N products in crude reaction mixture.
c
be required for up to 2 days.16 Meanwhile, O-benzylation is
achieved under classical alkylation conditions with benzyl bromide
and K2CO3 in yields of 50% and upwards.4 Our Mitsunobu
conditions afforded the desired O-methyl (Table 1 entry 7), O-iso-
propyl (Table 1, entry 5) and O-benzyl (Table 1, entry 6) protected
3-hydroxyisoxazoles in good to very good yields and in excellent
selectivities that are comparable with those in the literature for
similar reactions that employ alternative, non-Mitsunobu condi-
tions. For example, O-methylation of ethyl 2-(3-hydroxyisoxazol-
5-yl)acetate with diazomethane proceeded in 63% yield in a 2:1
ratio of O:N products,7 whilst treatment of methyl 4-(3-hydrox-
yisoxazol-5-yl)piperidine-1-carboxylate with isopropyl bromide
and benzyl bromide delivered the corresponding O-alkylated
products in 82% and 50% yields, respectively.16,4 Significantly,
though, our reaction conditions are more appealing, with transfor-
10. Comins, D. L.; Jianhua, G. Tetrahedron Lett. 1994, 35, 2819–2822.
11. For example: Lachance, N.; Guiral, S.; Huang, Z.; Leclerc, J.-P.; Li, C. S.; Oballa, R.
M.; Ramtohul, Y. K.; Wang, H.; Wu, J.; Zhang, L. Bioorg. Med. Chem. Lett. 2012,
22, 623–627.
12. Typical Mitsunobu procedure: 5-phenyl-3-hydroxyisoxazole (161 mg, 1.0 mmol,
1 equiv) was dissolved in anhydrous THF (14 mL), followed by isopropanol
(96
lL, 1.25 mmol, 1.25 equiv) and PPh3 (341 mg, 1.3 mmol, 1.3 equiv). The
reaction mixture was stirred for 2 min at rt. DIAD (256
lL, 1.3 mmol, 1.3 equiv)
was added dropwise. After stirring for a further 15 min at rt under an inert (N2)
atmosphere, TLC analysis confirmed the reaction was complete. The solvent
was concentrated under reduced pressure to afford the crude product, which
was purified by flash column chromatography (eluent gradient: 100% Hexane
to 9:1 Hexane/EtOAc to 1:1 Hexane/EtOAc) to give O-regioisomer (5)
3-isopropoxy-5-phenyl-isoxazole as a colourless oil (169 mg, 82%): Rf 0.68
(Hex/EtOAc, 1:1); dH (CDCl3, 400 MHz) 1.41 (d, J = 6.0 Hz, 6H, 2CH3), 4.94 (sep,
J = 6.0 Hz, 1H, CH(CH3)2), 6.08 (s, 1H, C4-H), 7.41–7.71 (m, 5H, Ph) and
N-regioisomer (6) 2-isopropoxy-5-phenyl-isoxazolone as
a colourless film
(21 mg, 10.3%): Rf 0.15 (Hex/EtOAc, 1:1); dH (CDCl3, 400 MHz) 1.41

1696
L. Chen, S. Fletcher / Tetrahedron Letters 55 (2014) 1693–1696
(d, J = 6.7 Hz, 6H, 2CH3), 4.69 (sep, J = 6.7 Hz, 1H, CH(CH3)2), 6.02 (s, 1H, C4-H),
7.44–7.67 (m, 5H, Ph).
J.; Frydenvang, K.; Kristensen, J.; Balle, T.; Jensen, A. J.; Kristiansen, U.; Frolund,
B. J. Med. Chem. 2013, 56, 993–1006.
13. El-Seedi, H. R.; Jensen, H. M.; Kure, N.; Thomsen, I.; Torssell, K. B. G. Acta Chem.
Scand. 1993, 47, 1004–1011.
14. Mukaiyama, T.; Kuroda, K.; Maruyama, Y. Heterocycles 2010, 80, 63–82.
15. (a) Kasai, S.; McGee, Jr., K. F. PCT Int. Appl. WO 2010-JP57201 2010.; (b)
Petersen, J. G.; Bergmann, R.; Moller, H. A.; Jorgensen, C. G.; Nielsen, B.; Kehler,
16. Frolund, B.; Jorgensen, A. T.; Tagmose, L.; Stensbol, T. B.; Vestergaard, H. T.;
Engblom, C.; Kristiansen, U.; Sanchez, C.; Krogsgaard-Larsen, P.; Liljefors, T. J.
Med. Chem. 2002, 45, 2454–2468.