- Email: [email protected]
A robust and simple protocol for the synthesis of arylfluorophosphonates
Tetrahedron Letters 56 (2015) 5619–5622
Contents lists available at ScienceDirect
Tetrahedron Letters journal homepage: www.elsevier.com/locate/tetlet
A robust and simple protocol for the synthesis of arylfluorophosphonates Mario Leypold a,b, Paal W. Wallace b,c,d, Marko Kljajic a,b, Matthias Schittmayer c,d, Jakob Pletz a,b, Carina Illaszewicz-Trattner a, Georg M. Guebitz b,e, Ruth Birner-Gruenberger b,c,d, Rolf Breinbauer a,b,⇑ a
Institute of Organic Chemistry, Graz University of Technology, Stremayrgasse 9, 8010 Graz, Austria Enzymes and Polymers, Austrian Centre of Industrial Biotechnology ACIB, Petersgasse 14, 8010 Graz, Austria Institute of Pathology, Medical University of Graz, Stiftingtalstrasse 24, 8010 Graz, Austria d Omics Center Graz, BioTechMed-Graz, Stiftingtalstrasse 24, 8010 Graz, Austria e Institute of Environmental Biotechnology, University of Natural Resources and Life Sciences, Konrad-Lorenz-Strasse 20, 3430 Tulln, Austria b c
a r t i c l e
i n f o
Article history: Received 25 June 2015 Revised 19 August 2015 Accepted 22 August 2015 Available online 22 August 2015 Keywords: Arylphosphonofluoridates Activity based probes Monoesterification DAST fluorination Pyrophosphonates
a b s t r a c t Fluorophosphonates represent powerful probes for the identification and analysis of active serine hydrolases in activity based protein profiling. Although alkylphosphonofluoridates are widely used for such purposes, little is known about the synthesis and purification of arylphosphonofluoridates, which may be useful tools for screening enzyme activities toward aromatic esters. Our optimized route makes this subclass of transition state inhibitors broadly accessible for a diverse series of phosphonic acid derivatives using a combination of selective monoesterification with EDCHCl and subsequent mild fluorination with DAST. All compounds were isolated as pure materials using a simple acid–base extraction protocol in 76–93% yields over two steps. These probes can be stored under an inert atmosphere at 24 °C for several months without significant degradation. Ó 2015 Elsevier Ltd. All rights reserved.
Activated phosphonic monoester derivatives, such as phosphonofluoridates1–4 and 4-nitrophenyl phosphonates,5–7 exert their toxic ability to the broad class of serine hydrolases by mimicking the tetrahedral transition state of enzymatic hydrolysis.4,8 Acting as effective covalent inhibitors, these analogues were used as chemical warfare agents in the First World War,9,10 but have received increasing attention in recent years for their applicability as probe molecules in activity based protein profiling (ABPP).4,7,11–13 Although several strategies for the synthesis of alkylphosphonic monoester fluoridates exist, little is known about the preparation of arylphosphonic acid derivatives, which could act as active probes for the selective inhibition of enzymes specific for the hydrolysis of aromatic esters.14 Most known approaches toward alkyl-based probes are either based on the monoesterification of alkylphosphonic acids using coupling reagents (DCC,15–20 EDCHCl,21 BOP,22,23 ByBOP22,24) or on the activation via phosphonic acid dichlorides,25–29 followed by late-stage fluorination with DAST,1–3,30–32 TBAT4 or BTFFH.33 However, for the synthesis of arylphosphonic acid derivatives these protocols would need to be adapted due to the altered electronic character compared to their alkyl analogues. Herein, we report a robust and facile procedure ⇑ Corresponding author. E-mail address: [email protected] (R. Breinbauer). http://dx.doi.org/10.1016/j.tetlet.2015.08.061 0040-4039/Ó 2015 Elsevier Ltd. All rights reserved.
for the synthesis of arylphosphonic monoester fluoridates by selective monoesterification of arylphosphonic acids using N-(3-dimethylaminopropyl)-N0 -ethylcarbodiimide hydrochloride (EDCHCl) followed by fluorination with (diethylamino)sulfur trifluoride (DAST) (Scheme 1). Importantly, all intermediates as well as the final compounds were easily isolated and purified by a simple acid–base extraction protocol. Our initial attempts for the synthesis of arylphosphonic monoester fluoridates were based on the use of phosphonic acid dichlorides25,27,29 as central intermediates. While the preparation of phenylphosphonic acid dichloride using either thionyl chloride or oxalyl chloride as activating reagents occurred smoothly, the conversion with alcohol 2a and subsequent hydrolysis did not result in selective monoesterification. Instead a mixture of sideproducts was detected by NMR analysis from which the desired intermediate could not be isolated by established extractive or chromatographic methods. An alternative approach for the formation of monoester 3a by esterification of phenylphosphonic acid (1a) with alcohol 2a using catalytic amounts of phenylarsonic acid34 also failed due to the generation of a chromatographically inseparable crude mixture in moderate yields. Application of a monoesterification strategy using EDCHCl proved successful to address the issue of selectivity. Varying the reaction temperature for monoesterification of phenylphosphonic acid (1a) with alcohol 2a showed that elevated temperatures and
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M. Leypold et al. / Tetrahedron Letters 56 (2015) 5619–5622
O
O
P OH OH
R1
+
HO
R2
P R2 O OH
EDC.HCl R1
O DAST R1
P R2 O F
Scheme 1. Selective monoesterification of arylphosphonic acids with EDCHCl and subsequent mild fluorination with DAST.
Table 1 Screening results and isolated yields for the selective monoesterification of phenylphosphonic acid (1a) with alcohol 2a (Scheme 2) Entrya
Temp (°C)
Phos. acid 1a (equiv)
EDCHCl (equiv)
Conv. (%)
Prod. 3a (%)
Pyro. mono. 4 (%)
Pyro. di. 5 (%)
Yieldb (%)
1 2 3 4 5 6 7 8 9 10d
22 40 60 80 80 80 80 80 80 80
1.0 1.0 1.0 1.0 1.2 1.0 1.0 1.2 1.2 1.2
1.0 1.0 1.0 1.0 1.0 1.2 1.5 1.5 1.5 1.5
30 39 57 71 73 83 93 >99 >99 >99
27 37 54 66 68 77 88 95 70c 94
<1 <1 <1 <1 <1 <1 <1 <1 16c <1
<1 <1 <1 <1 <1 <1 <1 <1 9c <1
n.d. n.d. n.d. 55 58 66 86 93 93 93
n.d. = not determined. a Reaction control via HPLC–MS: aliquot of reaction taken, solvent was removed using a stream of N2 and the crude material frozen until measurement. b Isolated yield of 3a after conversion of pyrophosphonates by exposing to air. c HPLC–MS analysis immediately after evaporation of the solvent. d Solvent: 1,2-DCE + 10% DMF (v/v).
O
EDC.HCl 5 mol% 4-DMAP
P OH HO OH + N3 1a
2a
O P O OH
1,2-DCE, 48 h
N3
O P O O OH P O
N3
O P O O O P O
N3
3a
4
5 N3
Scheme 2. Monoesterification of phenylphosphonic acid (1a) with alcohol 2a.
rather long reaction times (80 °C, 48 h) (entries 1–4, Table 1, Scheme 2) compared to alkylphosphonic acids (typically 35 °C, 10 h with DCC)16 were essential for acceptable conversion of the starting material. Alcohol 2 was selected as the limiting component as it was expected that the unreacted excess of phosphonic acid could be easily removed by an acid–base extraction. The optimal conditions were identified as 1.2 equiv of phosphonic acid and 1.5 equiv of EDCHCl at 80 °C for 48 h (entry 8, Table 1). Importantly, in no case was the diesterification product formed in the coupling reaction. Interestingly, when the sample was directly measured in reaction control experiments via HPLC–MS, the existence of pyrophosphonate monoester 4 (HPLC–MS (ESI, MH+): m/z = 456) and diester 5 (HPLC–MS (ESI, M+Na+): m/z = 639) could be detected (compare entries 8 and 9, Table 1, Scheme 2).22,35,36 This observation was in accordance with previously published results on alkylphosphonic acids using DCC as the coupling reagent.35–37 Indeed, after solvent evaporation these two derivatives were found in the crude materials in each tested combination of arylphosphonic acids 1a–c and alcohols 2a–c (Scheme 3). Efforts to hydrolyze these products to the desired monoesterified intermediates 3a–e under either acidic (1.0 M HCl) or basic conditions (1.0 M NaOH) were not successful within acceptable reaction times at room temperature.36,38 Using observations made with HPLC–MS samples stored for extended periods of time, in which the pyrophosphonate had vanished, a very simple method to convert pyrophosphonate by-products was found. When the crude materials were exposed to air at room temperature in open vessels for 48 h, all pyrophosphonate mono- and diesters were completely
hydrolyzed to the desired monoesterified compounds 3a–e. We hypothesize that humidity in the atmosphere in combination with an unexpected concentration effect of the neat substances resulted in smooth hydrolysis in comparison to the tested aqueous acidic and basic conditions. After workup, the excess phosphonic acid, coupling reagent, base, and urea derivative could be easily separated by a simple acid–base extraction protocol (see ESI). This optimized reaction protocol was examined for different arylphosphonic acids 1a–c containing electron-donating or electron-withdrawing substituents, as well as several alcohol reaction partners 2a–c (entries 1–5, Table 2, Scheme 3). As some of these arylphosphonic acids were not soluble in 1,2-DCE, DMF was added as a co-solvent (10% (v/v)), which did not influence the outcome of the reaction with phenylphosphonic acid (1a) and alcohol 2a (entry 10, Table 1). Compounds 3a–e were isolated as pure materials in yields between 88% and 95% (Table 2). DAST was used for subsequent fluorination of the monoesterified arylphosphonic acids 3a–e as this reagent would allow use of an extractive workup as the sole purification method.2 Under optimized reaction conditions, arylphosphonic acid monoesters with both electron-donating or electron-withdrawing residues were quantitatively converted to arylphosphonofluoridates 6a–e within 1 h using 1.5 equiv of DAST (Scheme 3).1–3,30 Due to the kinetic inertness of the phosphorus–fluorine bond toward hydrolysis, products 6a–e could be purified and isolated as pure materials in yields between 86% and 98% by washing the organic layer with 0.5 M HCl (Table 2, Scheme 3, see ESI).13 All synthesized arylphosphonofluoridates 6a–e could be stored under an inert
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M. Leypold et al. / Tetrahedron Letters 56 (2015) 5619–5622
O P OH OH
O
alcohol EDC.HCl, 5 mol% 4-DMAP 1,2-DCE + 10 % DMF (v/v) 80 °C, 48 h
R R = -H: 1a -OMe: 1b -COMe: 1c
X
n
OH
N3
alcohol: n = 1, X = -CH2-: 2a n = 1, X = -O-: 2b n = 2, X = -O-: 2c
P O OH
O X n
R
N3 R= R= R= R= R=
P O F
DAST
H, n = 1, X = -CH 2-: 3a H, n = 1, X = -O-: 3b H, n = 2, X = -O-: 3c -OMe, n = 1, X = -CH2-: 3d -COMe, n = 1, X = -CH 2-: 3e
DCM, 22 °C, 1 h
R
N3 R = H, n = 1, X = -CH2-: 6a R = H, n = 1, X = -O-: 6b R = H, n = 2, X = -O-: 6c R = -OMe, n = 1, X = -CH 2-: 6d R = -COMe, n = 1, X = -CH2-: 6e
N F
X n
S F F
DAST
Scheme 3. Synthesis of arylphosphonofluoridates 6a–e via selective monoesterification with EDCHCl and fluorination with DAST.
Table 2 Monoesterification and fluorination for a selected combination of arylphosphonic acids 1a–c and alcohols 2a–c (Scheme 3)
a b
Entry
Phos. acid
Alcohol
Final product
Monoester yield (%)
Monoester puritya (%)
Fluorination yield (%)
Overall yield (%)
1 2b 3b 4b 5
1a 1a 1a 1b 1c
2a 2b 2c 2a 2a
6a 6b 6c 6d 6e
93 92 95 88 88
98 99 98 99 99
98 95 98 98 86
91 87 93 86 76
Determined via HPLC–MS analysis. Reaction time: 24 h.
atmosphere at 24 °C for several months without significant degradation, which makes these compounds highly attractive as probes for ABPP. In summary, we have identified general conditions for the twostep synthesis of a set of electronically differentiated arylphosphonofluoridates, which provide pure materials by the application of two extractive purification protocols. This methodology should prove particularly useful for targeting hydrolases with activities toward aromatic esters by ABPP. Further investigations into the activity of these probes are underway and will be reported in due course. Acknowledgments This work has been supported by the Federal Ministry of Science, Research and Economy (BMWFW), the Federal Ministry of Traffic, Innovation and Technology (bmvit), the Styrian Business Promotion Agency SFG, the Standortagentur Tirol, the Government of Lower Austria and ZIT—Technology Agency of the City of Vienna through the COMET-Funding Program managed by the Austrian Research Promotion Agency FFG (to R.B.-G., R.B. and G.G.), NAWI Graz (to R.B.), Austrian Science Fund (FWF) Project P26074 (to R. B.-G.) and the doctoral school ‘DK Metabolic and Cardiovascular Disease’ (W1226) (to R.B.-G.). Supplementary data Supplementary data (experimental procedures, compound characterization and NMR-spectra of all prepared compounds) associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.tetlet.2015.08.061. References and notes 1. Eubanks, L. M.; Stowe, G. N.; de Lamo Marin, S.; Mayorov, A. V.; Hixon, M. S.; Janda, K. D. Angew. Chem., Int. Ed. 2011, 50, 10699–10702. 2. Xu, H.; Sabit, H.; Amidon, G. L.; Showalter, H. D. H. Beilstein J. Org. Chem. 2013, 9, 89–96. 3. Raghavan, A.; Charron, G.; Flexner, J.; Hang, H. C. Bioorg. Med. Chem. Lett. 2008, 18, 5982–5986.
4. Tuin, A. W.; Mol, M. A. E.; van den Berg, R. M.; Fidder, A.; van der Marel, G. A.; Overkleeft, H. S.; Noort, D. Chem. Res. Toxicol. 2009, 102, 683–689. 5. Dijkstra, H. P.; Sprong, H.; Aerts, B. N. H.; Kruithof, C. A.; Egmond, M. R.; Klein Gebbink, R. J. M. Org. Biomol. Chem. 2008, 6, 523–531. 6. Li, N.; Rahil, J.; Wright, M. E.; Pratt, R. F. Bioorg. Med. Chem. 1997, 5, 1783–1788. 7. Birner-Gruenberger, R.; Susani-Etzerodt, H.; Kollroser, M.; Rechberger, G. N.; Hermetter, A. Proteomics 2008, 8, 3645–3656. 8. Hedstrom, L. Chem. Rev. 2002, 102, 4501–4524. 9. Yanagisawa, N.; Morita, H.; Nakajima, T. J. Neurol. Sci. 2006, 249, 76–85. 10. Norlin, R.; Lindberg, G. J. Labelled Compd. Radiopharm. 2003, 46, 599–604. 11. Rudolf, G. C.; Sieber, S. A. ChemBioChem 2013, 14, 2447–2455. 12. Simon, G. M.; Cravatt, B. F. J. Biol. Chem. 2010, 285, 11051–11055. 13. Tully, S. E.; Cravatt, B. F. J. Am. Chem. Soc. 2010, 132, 3264–3265. 14. Mironowicz, A.; Kukułczanka, K.; Siewin´ski, A. Acta Soc. Bot. Pol. 2014, 62, 21– 23. 15. North, E. J.; Osborne, D. A.; Bridson, P. K.; Baker, D. L.; Parrill, A. L. Bioorg. Med. Chem. 2009, 17, 3433–3442. 16. Tanaka, F.; Kinoshita, K.; Tanimura, R.; Fujii, I. J. Am. Chem. Soc. 1996, 118, 2332–2339. 17. Julien, O.; Beadle, J. R.; Magee, W. C.; Chatterjee, S.; Hostetler, K. Y.; Evans, D. H.; Sykes, B. D. J. Am. Chem. Soc. 2011, 133, 2264–2274. 18. Ohashi, K.; Kosai, S.; Arizuka, M.; Watanabe, T.; Fukunaga, M.; Monden, K.; Uchikoda, T.; Yamagiwa, Y.; Kamikawa, T. Tetrahedron Lett. 1988, 29, 1189– 1192. 19. Chauhan, S. S.; Varshney, A.; Verma, B.; Pennington, M. W. Tetrahedron Lett. 2007, 48, 4051–4054. 20. Gupta, A. K.; Kumar, R.; Gupta, H. K.; Tak, V.; Dubey, D. K. Tetrahedron Lett. 2008, 49, 1656–1659. 21. Cressina, E.; Lloyd, A. J.; de Pascale, G.; Mok, B. J.; Caddick, S.; Roper, D. I.; Dowson, C. G.; Bugg, T. D. H. Bioorg. Med. Chem. 2009, 17, 3443–3455. 22. Campagne, J.-M.; Coste, J.; Jouin, P. J. Org. Chem. 1995, 60, 5214–5223. 23. Pícha, J.; Budeˇšínsky´, M.; Hancˇlová, I.; Šanda, M.; Fiedler, P.; Vaneˇk, V.; Jirácˇek, J. Tetrahedron 2009, 65, 6090–6103. 24. Zakharova, V. M.; Serpi, M.; Krylov, I. S.; Peterson, L. W.; Breitenbach, J. M.; Borysko, K. Z.; Drach, J. C.; Collins, M.; Hilfinger, J. M.; Kashemirov, B. A.; McKenna, C. E. J. Med. Chem. 2011, 54, 5680–5693. 25. Liu, L.; Yuan, H.; Fu, T.; Wang, T.; Gao, X.; Zeng, Z.; Zhu, J.; Zhao, Y. J. Org. Chem. 2014, 79, 80–87. 26. McWhirter, C.; Lund, E. A.; Tanifum, E. A.; Feng, G.; Sheikh, Q. I.; Hengge, A. C.; Williams, N. H. J. Am. Chem. Soc. 2008, 130, 13673–13682. 27. Norlin, R.; Juhlin, L.; Lind, P.; Trogen, L. Synthesis 2005, 1765–1770. 28. Nowlan, C.; Li, Y.; Hermann, J. C.; Evans, T.; Carpenter, J.; Ghanem, E.; Shoichet, B. K.; Raushel, F. M. J. Am. Chem. Soc. 2006, 128, 15892–15902. 29. Stepinski, D. C.; Nelson, D. W.; Zalupski, P. R.; Herlinger, A. W. Tetrahedron 2001, 57, 8637–8645. 30. Bisogno, T.; Burston, J. J.; Rai, R.; Allarà, M.; Saha, B.; Mahadevan, A.; Razdan, R. K.; Wiley, J. L.; Di Marzo, V. ChemMedChem 2009, 4, 946–950. 31. Inoue, M.; Hiratake, J.; Sakata, K. Biosci., Biotechnol., Biochem. 1999, 63, 2248– 2251. 32. Patricelli, M. P.; Cravatt, B. F. Biochemistry 2001, 40, 6107–6115. 33. Barry, C. S.; Backus, K. M.; Barry, C. E.; Davis, B. G. J. Am. Chem. Soc. 2011, 133, 13232–13235.
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34. Crenshaw, M. D. Phosphorus, Sulfur Silicon Relat. Elem. 2004, 179, 1509–1516. 35. Hirschmann, R.; Yager, K. M.; Taylor, C. M.; Moore, W.; Sprengeler, P. A.; Witherington, J.; Phillips, B. W.; Smith, A. B., III J. Am. Chem. Soc. 1995, 117, 6370–6371.
36. Udodong, U. E.; Hansen, M. M.; Verral, D. E.; Harkness, A. R.; Grutsch, J. L.; Miller, W. D.; Astleford, B. Tetrahedron 2003, 59, 2667–2672. 37. Bartlett, P. A.; Hanson, J. E.; Giannousis, P. P. J. Org. Chem. 1990, 55, 6268–6274. 38. Diemert, K.; Kuchen, W.; Poll, W.; Sandt, F. Eur. J. Inorg. Chem. 1998, 361–366.
Contents lists available at ScienceDirect
Tetrahedron Letters journal homepage: www.elsevier.com/locate/tetlet
A robust and simple protocol for the synthesis of arylfluorophosphonates Mario Leypold a,b, Paal W. Wallace b,c,d, Marko Kljajic a,b, Matthias Schittmayer c,d, Jakob Pletz a,b, Carina Illaszewicz-Trattner a, Georg M. Guebitz b,e, Ruth Birner-Gruenberger b,c,d, Rolf Breinbauer a,b,⇑ a
Institute of Organic Chemistry, Graz University of Technology, Stremayrgasse 9, 8010 Graz, Austria Enzymes and Polymers, Austrian Centre of Industrial Biotechnology ACIB, Petersgasse 14, 8010 Graz, Austria Institute of Pathology, Medical University of Graz, Stiftingtalstrasse 24, 8010 Graz, Austria d Omics Center Graz, BioTechMed-Graz, Stiftingtalstrasse 24, 8010 Graz, Austria e Institute of Environmental Biotechnology, University of Natural Resources and Life Sciences, Konrad-Lorenz-Strasse 20, 3430 Tulln, Austria b c
a r t i c l e
i n f o
Article history: Received 25 June 2015 Revised 19 August 2015 Accepted 22 August 2015 Available online 22 August 2015 Keywords: Arylphosphonofluoridates Activity based probes Monoesterification DAST fluorination Pyrophosphonates
a b s t r a c t Fluorophosphonates represent powerful probes for the identification and analysis of active serine hydrolases in activity based protein profiling. Although alkylphosphonofluoridates are widely used for such purposes, little is known about the synthesis and purification of arylphosphonofluoridates, which may be useful tools for screening enzyme activities toward aromatic esters. Our optimized route makes this subclass of transition state inhibitors broadly accessible for a diverse series of phosphonic acid derivatives using a combination of selective monoesterification with EDCHCl and subsequent mild fluorination with DAST. All compounds were isolated as pure materials using a simple acid–base extraction protocol in 76–93% yields over two steps. These probes can be stored under an inert atmosphere at 24 °C for several months without significant degradation. Ó 2015 Elsevier Ltd. All rights reserved.
Activated phosphonic monoester derivatives, such as phosphonofluoridates1–4 and 4-nitrophenyl phosphonates,5–7 exert their toxic ability to the broad class of serine hydrolases by mimicking the tetrahedral transition state of enzymatic hydrolysis.4,8 Acting as effective covalent inhibitors, these analogues were used as chemical warfare agents in the First World War,9,10 but have received increasing attention in recent years for their applicability as probe molecules in activity based protein profiling (ABPP).4,7,11–13 Although several strategies for the synthesis of alkylphosphonic monoester fluoridates exist, little is known about the preparation of arylphosphonic acid derivatives, which could act as active probes for the selective inhibition of enzymes specific for the hydrolysis of aromatic esters.14 Most known approaches toward alkyl-based probes are either based on the monoesterification of alkylphosphonic acids using coupling reagents (DCC,15–20 EDCHCl,21 BOP,22,23 ByBOP22,24) or on the activation via phosphonic acid dichlorides,25–29 followed by late-stage fluorination with DAST,1–3,30–32 TBAT4 or BTFFH.33 However, for the synthesis of arylphosphonic acid derivatives these protocols would need to be adapted due to the altered electronic character compared to their alkyl analogues. Herein, we report a robust and facile procedure ⇑ Corresponding author. E-mail address: [email protected] (R. Breinbauer). http://dx.doi.org/10.1016/j.tetlet.2015.08.061 0040-4039/Ó 2015 Elsevier Ltd. All rights reserved.
for the synthesis of arylphosphonic monoester fluoridates by selective monoesterification of arylphosphonic acids using N-(3-dimethylaminopropyl)-N0 -ethylcarbodiimide hydrochloride (EDCHCl) followed by fluorination with (diethylamino)sulfur trifluoride (DAST) (Scheme 1). Importantly, all intermediates as well as the final compounds were easily isolated and purified by a simple acid–base extraction protocol. Our initial attempts for the synthesis of arylphosphonic monoester fluoridates were based on the use of phosphonic acid dichlorides25,27,29 as central intermediates. While the preparation of phenylphosphonic acid dichloride using either thionyl chloride or oxalyl chloride as activating reagents occurred smoothly, the conversion with alcohol 2a and subsequent hydrolysis did not result in selective monoesterification. Instead a mixture of sideproducts was detected by NMR analysis from which the desired intermediate could not be isolated by established extractive or chromatographic methods. An alternative approach for the formation of monoester 3a by esterification of phenylphosphonic acid (1a) with alcohol 2a using catalytic amounts of phenylarsonic acid34 also failed due to the generation of a chromatographically inseparable crude mixture in moderate yields. Application of a monoesterification strategy using EDCHCl proved successful to address the issue of selectivity. Varying the reaction temperature for monoesterification of phenylphosphonic acid (1a) with alcohol 2a showed that elevated temperatures and
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M. Leypold et al. / Tetrahedron Letters 56 (2015) 5619–5622
O
O
P OH OH
R1
+
HO
R2
P R2 O OH
EDC.HCl R1
O DAST R1
P R2 O F
Scheme 1. Selective monoesterification of arylphosphonic acids with EDCHCl and subsequent mild fluorination with DAST.
Table 1 Screening results and isolated yields for the selective monoesterification of phenylphosphonic acid (1a) with alcohol 2a (Scheme 2) Entrya
Temp (°C)
Phos. acid 1a (equiv)
EDCHCl (equiv)
Conv. (%)
Prod. 3a (%)
Pyro. mono. 4 (%)
Pyro. di. 5 (%)
Yieldb (%)
1 2 3 4 5 6 7 8 9 10d
22 40 60 80 80 80 80 80 80 80
1.0 1.0 1.0 1.0 1.2 1.0 1.0 1.2 1.2 1.2
1.0 1.0 1.0 1.0 1.0 1.2 1.5 1.5 1.5 1.5
30 39 57 71 73 83 93 >99 >99 >99
27 37 54 66 68 77 88 95 70c 94
<1 <1 <1 <1 <1 <1 <1 <1 16c <1
<1 <1 <1 <1 <1 <1 <1 <1 9c <1
n.d. n.d. n.d. 55 58 66 86 93 93 93
n.d. = not determined. a Reaction control via HPLC–MS: aliquot of reaction taken, solvent was removed using a stream of N2 and the crude material frozen until measurement. b Isolated yield of 3a after conversion of pyrophosphonates by exposing to air. c HPLC–MS analysis immediately after evaporation of the solvent. d Solvent: 1,2-DCE + 10% DMF (v/v).
O
EDC.HCl 5 mol% 4-DMAP
P OH HO OH + N3 1a
2a
O P O OH
1,2-DCE, 48 h
N3
O P O O OH P O
N3
O P O O O P O
N3
3a
4
5 N3
Scheme 2. Monoesterification of phenylphosphonic acid (1a) with alcohol 2a.
rather long reaction times (80 °C, 48 h) (entries 1–4, Table 1, Scheme 2) compared to alkylphosphonic acids (typically 35 °C, 10 h with DCC)16 were essential for acceptable conversion of the starting material. Alcohol 2 was selected as the limiting component as it was expected that the unreacted excess of phosphonic acid could be easily removed by an acid–base extraction. The optimal conditions were identified as 1.2 equiv of phosphonic acid and 1.5 equiv of EDCHCl at 80 °C for 48 h (entry 8, Table 1). Importantly, in no case was the diesterification product formed in the coupling reaction. Interestingly, when the sample was directly measured in reaction control experiments via HPLC–MS, the existence of pyrophosphonate monoester 4 (HPLC–MS (ESI, MH+): m/z = 456) and diester 5 (HPLC–MS (ESI, M+Na+): m/z = 639) could be detected (compare entries 8 and 9, Table 1, Scheme 2).22,35,36 This observation was in accordance with previously published results on alkylphosphonic acids using DCC as the coupling reagent.35–37 Indeed, after solvent evaporation these two derivatives were found in the crude materials in each tested combination of arylphosphonic acids 1a–c and alcohols 2a–c (Scheme 3). Efforts to hydrolyze these products to the desired monoesterified intermediates 3a–e under either acidic (1.0 M HCl) or basic conditions (1.0 M NaOH) were not successful within acceptable reaction times at room temperature.36,38 Using observations made with HPLC–MS samples stored for extended periods of time, in which the pyrophosphonate had vanished, a very simple method to convert pyrophosphonate by-products was found. When the crude materials were exposed to air at room temperature in open vessels for 48 h, all pyrophosphonate mono- and diesters were completely
hydrolyzed to the desired monoesterified compounds 3a–e. We hypothesize that humidity in the atmosphere in combination with an unexpected concentration effect of the neat substances resulted in smooth hydrolysis in comparison to the tested aqueous acidic and basic conditions. After workup, the excess phosphonic acid, coupling reagent, base, and urea derivative could be easily separated by a simple acid–base extraction protocol (see ESI). This optimized reaction protocol was examined for different arylphosphonic acids 1a–c containing electron-donating or electron-withdrawing substituents, as well as several alcohol reaction partners 2a–c (entries 1–5, Table 2, Scheme 3). As some of these arylphosphonic acids were not soluble in 1,2-DCE, DMF was added as a co-solvent (10% (v/v)), which did not influence the outcome of the reaction with phenylphosphonic acid (1a) and alcohol 2a (entry 10, Table 1). Compounds 3a–e were isolated as pure materials in yields between 88% and 95% (Table 2). DAST was used for subsequent fluorination of the monoesterified arylphosphonic acids 3a–e as this reagent would allow use of an extractive workup as the sole purification method.2 Under optimized reaction conditions, arylphosphonic acid monoesters with both electron-donating or electron-withdrawing residues were quantitatively converted to arylphosphonofluoridates 6a–e within 1 h using 1.5 equiv of DAST (Scheme 3).1–3,30 Due to the kinetic inertness of the phosphorus–fluorine bond toward hydrolysis, products 6a–e could be purified and isolated as pure materials in yields between 86% and 98% by washing the organic layer with 0.5 M HCl (Table 2, Scheme 3, see ESI).13 All synthesized arylphosphonofluoridates 6a–e could be stored under an inert
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O P OH OH
O
alcohol EDC.HCl, 5 mol% 4-DMAP 1,2-DCE + 10 % DMF (v/v) 80 °C, 48 h
R R = -H: 1a -OMe: 1b -COMe: 1c
X
n
OH
N3
alcohol: n = 1, X = -CH2-: 2a n = 1, X = -O-: 2b n = 2, X = -O-: 2c
P O OH
O X n
R
N3 R= R= R= R= R=
P O F
DAST
H, n = 1, X = -CH 2-: 3a H, n = 1, X = -O-: 3b H, n = 2, X = -O-: 3c -OMe, n = 1, X = -CH2-: 3d -COMe, n = 1, X = -CH 2-: 3e
DCM, 22 °C, 1 h
R
N3 R = H, n = 1, X = -CH2-: 6a R = H, n = 1, X = -O-: 6b R = H, n = 2, X = -O-: 6c R = -OMe, n = 1, X = -CH 2-: 6d R = -COMe, n = 1, X = -CH2-: 6e
N F
X n
S F F
DAST
Scheme 3. Synthesis of arylphosphonofluoridates 6a–e via selective monoesterification with EDCHCl and fluorination with DAST.
Table 2 Monoesterification and fluorination for a selected combination of arylphosphonic acids 1a–c and alcohols 2a–c (Scheme 3)
a b
Entry
Phos. acid
Alcohol
Final product
Monoester yield (%)
Monoester puritya (%)
Fluorination yield (%)
Overall yield (%)
1 2b 3b 4b 5
1a 1a 1a 1b 1c
2a 2b 2c 2a 2a
6a 6b 6c 6d 6e
93 92 95 88 88
98 99 98 99 99
98 95 98 98 86
91 87 93 86 76
Determined via HPLC–MS analysis. Reaction time: 24 h.
atmosphere at 24 °C for several months without significant degradation, which makes these compounds highly attractive as probes for ABPP. In summary, we have identified general conditions for the twostep synthesis of a set of electronically differentiated arylphosphonofluoridates, which provide pure materials by the application of two extractive purification protocols. This methodology should prove particularly useful for targeting hydrolases with activities toward aromatic esters by ABPP. Further investigations into the activity of these probes are underway and will be reported in due course. Acknowledgments This work has been supported by the Federal Ministry of Science, Research and Economy (BMWFW), the Federal Ministry of Traffic, Innovation and Technology (bmvit), the Styrian Business Promotion Agency SFG, the Standortagentur Tirol, the Government of Lower Austria and ZIT—Technology Agency of the City of Vienna through the COMET-Funding Program managed by the Austrian Research Promotion Agency FFG (to R.B.-G., R.B. and G.G.), NAWI Graz (to R.B.), Austrian Science Fund (FWF) Project P26074 (to R. B.-G.) and the doctoral school ‘DK Metabolic and Cardiovascular Disease’ (W1226) (to R.B.-G.). Supplementary data Supplementary data (experimental procedures, compound characterization and NMR-spectra of all prepared compounds) associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.tetlet.2015.08.061. References and notes 1. Eubanks, L. M.; Stowe, G. N.; de Lamo Marin, S.; Mayorov, A. V.; Hixon, M. S.; Janda, K. D. Angew. Chem., Int. Ed. 2011, 50, 10699–10702. 2. Xu, H.; Sabit, H.; Amidon, G. L.; Showalter, H. D. H. Beilstein J. Org. Chem. 2013, 9, 89–96. 3. Raghavan, A.; Charron, G.; Flexner, J.; Hang, H. C. Bioorg. Med. Chem. Lett. 2008, 18, 5982–5986.
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