Organic & Biomolecular Chemistry
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Cite this: DOI: 10.1039/c5ob01223f Received 16th June 2015, Accepted 29th June 2015
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De novo synthesis of phenolic dihydroxanthene near-infrared emitting fluorophores† Jean-Alexandre Richard*
DOI: 10.1039/c5ob01223f www.rsc.org/obc
We report a flexible de novo synthesis of phenolic dihydroxanthene fluorophores. The synthesis relies on a one-pot formation of an aldehyde intermediate which can be diversified in 60–70% overall yield, providing an efficient access to this family of near-infrared emitting fluorophores.
Near-infrared (NIR) imaging is becoming a serious candidate to complement the range of imaging techniques currently available in the clinic. The deep photon penetration combined with a good safety profile due to the non-invasiveness of NIR radiations have made optical in vivo imaging a powerful technique for medical diagnostics1 and image-guided surgery.2 Fluorescent tracers emitting above 700 nm are necessary in order to perform in vivo imaging because autofluorescence and absorption by biomolecules are minimized compared to the visible range.3 Scattering of light in tissues also decreases but the absorption of light by water above 900 nm precludes the use of fluorophores emitting at longer wavelengths. A good signal to background ratio thus requires the development of imaging tools in the optimum 700–900 nm window of the electromagnetic spectrum.4 Despite this relatively well-accepted paradigm the chemical tools available to probe biological functions in the near-infrared still require improvement.5 In particular, fluorophores whose light emission can be switched on depending on the biological environment provide an excellent platform to obtain sensitive fluorogenic probes.6 One simple and powerful strategy in this regard is the use of phenol fluorophores where their substitution as aryl ether or esters switches off their emission while the fluorescence can be restored upon release of the phenol moiety. This attractive strategy has so far mostly been applied to visible-emitting dyes and only a few examples of fluo-
Organic Chemistry, Institute of Chemical and Engineering Sciences (ICES), Agency for Science, Technology and Research (A*STAR), 11 Biopolis Way, The Helios Block, #03-08, Singapore 138667, Singapore. E-mail: [email protected] † Electronic supplementary information (ESI) available: Detailed experimental procedures and copies of 1H and 13C NMR spectroscopy. See DOI: 10.1039/ c5ob01223f
This journal is © The Royal Society of Chemistry 2015
resceins7 and 7-hydroxycoumarins8 pro-fluorophores emitting in the near-infrared have been reported so far. In this context, the serendipitous discovery of the phenolic dihydroxanthene skeleton by Lin and co-workers has attracted our attention because this skeleton would fill the current gap existing for near-infrared phenolic pro-fluorophores (Scheme 1A).9 Since the disclosure of this scaffold and its use as sensor for hydrogen peroxide (H2O2), applications have
Scheme 1 A. Fluorogenic probes based on the phenolic dihydroxanthene scaffold; B. Degradation of cyanine dyes for the synthesis of phenolic dihydroxanthenes; C. De novo synthesis of the phenolic dihydroxanthene scaffold.
Org. Biomol. Chem.
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been reported for the detection or monitoring of nitroxyl (HNO),10 heavy metals,11 β-lactamase,12 nitroreductase,13 hydrazine,14 liposomal pH15 and thiols.16 These early studies showed great promise for the development of near-infrared emitting fluorogenic probes but so far the access to phenolic dihydroxanthenes requires the destruction of valuable cyanine9–15 or fluorescein dyes16a in order to forge the skeleton of the fluorophore (Scheme 1B). This degradative approach lacks flexibility and limits the chemical space accessible to obtain structural analogues. A de novo synthesis would give more room for diversification but so far the only route reported by Lin and co-workers provided the skeleton of dye 7 in 20% yield (Scheme 1C). This low yield combined to the effort needed to prepare analogues of Fisher aldehyde 6 for diversification limit the efficiency and scope of the structures potentially accessible. The development of a reliable, high yielding and flexible route to this scaffold thus appears necessary to fully reveal its potential and foster its use as fluorogenic probe. Inspired by this early investigations we aimed at the design of a synthetic route which would provide a quick access to the dihydroxanthene skeleton and offer a stable precursor for derivatization. We targeted aldehyde 10 as key intermediate which allowed us to choose Fisher’s bases 11 as condensation partner instead of the less readily accessible Fischer aldehydes (Scheme 1C).17 The strategy to access aldehyde 10 would rely on a one-pot cascade sequence involving commercially available 2-hydroxy-4-methoxybenzaldehyde 8 and β-haloenal 9. The working hypothesis for achieving this transformation
Organic & Biomolecular Chemistry
would involve an oxa-Michael addition of a phenolate moiety to the electrophilic β-haloenal 9. A subsequent retro-Michael reaction would then generate aryl ether 13 which in the presence of a base would form enolate 14. A vinylogous aldol reaction followed by a final dehydration would lead to the desired aldehyde 10 (Scheme 2A). An alternative mechanism could also start with the vinylogous aldol reaction, forming first the C–C bond at the meso position of the pseudo xanthene skeleton. A sequence oxa-Michael addition/retro-Michael/ dehydration would lead to the same key intermediate 10 featuring the aldehyde function necessary for further derivatization (Scheme 2B). Reaction with indolium species 11 would then provide methylated dyes 18 and phenolic dihydroxanthene NIR fluorophores 19 after final deprotection (Scheme 2C). An overview of the reaction conditions screened in order to achieve the cascade reaction is presented in Table 1. We observed that the nature of the β-haloenal 9 used did not significantly affect the yields obtained so that eventually 9a was preferred over 9b because it was more stable and easier to handle.18 Organic bases were tried first and showed a moderate level of success. A catalytic amount of DABCO in CH2Cl2 did not form any desired product (entry 1) but traces of aldehyde 10 could be isolated using DBU (entry 2). Larger quantities of DABCO (2 eq.) in DMF slightly improved the yield (entry 3) and pyrrolidine could increase it up to 17% (entry 4). This encouraging result prompted us to try aminocatalysis conditions in the presence of L-proline or Jørgensen–Hayashi’s diphenylprolinol trimethylsilyl ether catalyst19 to promote enamine and iminium activation. Unfortunately, these con-
Scheme 2 A. Working hypothesis relying on a cascade oxa-Michael/retro-Michael/vinylogous aldol/dehydration to forge the phenolic dihydroxanthene scaffold; B. Alternative mechanism cascade vinylogous aldol/oxa-Michael/retro-Michael/dehydration; C. Approach for a divergent, de novo synthesis of phenolic dihydroxanthene near-infrared emitting dyes.
Org. Biomol. Chem.
This journal is © The Royal Society of Chemistry 2015
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Organic & Biomolecular Chemistry
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Published on 06 July 2015. Downloaded by UNIVERSITY OF OTAGO on 06/07/2015 16:10:33.
Table 1 Optimization of the oxa-Michael/retro-Michael/vinylogous aldol/dehydration cascade sequencea
Entry
β-haloenal
Base (eq.)
Solvent
Yieldb (%)
1 2 3 4 5 6
9a 9a 9a 9a 9b 9b
CH2Cl2 CH2Cl2 DMF CH2Cl2 DMSO CHCl3
0
Published on 06 July 2015. Downloaded by UNIVERSITY OF OTAGO on 06/07/2015 16:10:33.
COMMUNICATION
Cite this: DOI: 10.1039/c5ob01223f Received 16th June 2015, Accepted 29th June 2015
View Article Online View Journal
De novo synthesis of phenolic dihydroxanthene near-infrared emitting fluorophores† Jean-Alexandre Richard*
DOI: 10.1039/c5ob01223f www.rsc.org/obc
We report a flexible de novo synthesis of phenolic dihydroxanthene fluorophores. The synthesis relies on a one-pot formation of an aldehyde intermediate which can be diversified in 60–70% overall yield, providing an efficient access to this family of near-infrared emitting fluorophores.
Near-infrared (NIR) imaging is becoming a serious candidate to complement the range of imaging techniques currently available in the clinic. The deep photon penetration combined with a good safety profile due to the non-invasiveness of NIR radiations have made optical in vivo imaging a powerful technique for medical diagnostics1 and image-guided surgery.2 Fluorescent tracers emitting above 700 nm are necessary in order to perform in vivo imaging because autofluorescence and absorption by biomolecules are minimized compared to the visible range.3 Scattering of light in tissues also decreases but the absorption of light by water above 900 nm precludes the use of fluorophores emitting at longer wavelengths. A good signal to background ratio thus requires the development of imaging tools in the optimum 700–900 nm window of the electromagnetic spectrum.4 Despite this relatively well-accepted paradigm the chemical tools available to probe biological functions in the near-infrared still require improvement.5 In particular, fluorophores whose light emission can be switched on depending on the biological environment provide an excellent platform to obtain sensitive fluorogenic probes.6 One simple and powerful strategy in this regard is the use of phenol fluorophores where their substitution as aryl ether or esters switches off their emission while the fluorescence can be restored upon release of the phenol moiety. This attractive strategy has so far mostly been applied to visible-emitting dyes and only a few examples of fluo-
Organic Chemistry, Institute of Chemical and Engineering Sciences (ICES), Agency for Science, Technology and Research (A*STAR), 11 Biopolis Way, The Helios Block, #03-08, Singapore 138667, Singapore. E-mail: [email protected] † Electronic supplementary information (ESI) available: Detailed experimental procedures and copies of 1H and 13C NMR spectroscopy. See DOI: 10.1039/ c5ob01223f
This journal is © The Royal Society of Chemistry 2015
resceins7 and 7-hydroxycoumarins8 pro-fluorophores emitting in the near-infrared have been reported so far. In this context, the serendipitous discovery of the phenolic dihydroxanthene skeleton by Lin and co-workers has attracted our attention because this skeleton would fill the current gap existing for near-infrared phenolic pro-fluorophores (Scheme 1A).9 Since the disclosure of this scaffold and its use as sensor for hydrogen peroxide (H2O2), applications have
Scheme 1 A. Fluorogenic probes based on the phenolic dihydroxanthene scaffold; B. Degradation of cyanine dyes for the synthesis of phenolic dihydroxanthenes; C. De novo synthesis of the phenolic dihydroxanthene scaffold.
Org. Biomol. Chem.
View Article Online
Published on 06 July 2015. Downloaded by UNIVERSITY OF OTAGO on 06/07/2015 16:10:33.
Communication
been reported for the detection or monitoring of nitroxyl (HNO),10 heavy metals,11 β-lactamase,12 nitroreductase,13 hydrazine,14 liposomal pH15 and thiols.16 These early studies showed great promise for the development of near-infrared emitting fluorogenic probes but so far the access to phenolic dihydroxanthenes requires the destruction of valuable cyanine9–15 or fluorescein dyes16a in order to forge the skeleton of the fluorophore (Scheme 1B). This degradative approach lacks flexibility and limits the chemical space accessible to obtain structural analogues. A de novo synthesis would give more room for diversification but so far the only route reported by Lin and co-workers provided the skeleton of dye 7 in 20% yield (Scheme 1C). This low yield combined to the effort needed to prepare analogues of Fisher aldehyde 6 for diversification limit the efficiency and scope of the structures potentially accessible. The development of a reliable, high yielding and flexible route to this scaffold thus appears necessary to fully reveal its potential and foster its use as fluorogenic probe. Inspired by this early investigations we aimed at the design of a synthetic route which would provide a quick access to the dihydroxanthene skeleton and offer a stable precursor for derivatization. We targeted aldehyde 10 as key intermediate which allowed us to choose Fisher’s bases 11 as condensation partner instead of the less readily accessible Fischer aldehydes (Scheme 1C).17 The strategy to access aldehyde 10 would rely on a one-pot cascade sequence involving commercially available 2-hydroxy-4-methoxybenzaldehyde 8 and β-haloenal 9. The working hypothesis for achieving this transformation
Organic & Biomolecular Chemistry
would involve an oxa-Michael addition of a phenolate moiety to the electrophilic β-haloenal 9. A subsequent retro-Michael reaction would then generate aryl ether 13 which in the presence of a base would form enolate 14. A vinylogous aldol reaction followed by a final dehydration would lead to the desired aldehyde 10 (Scheme 2A). An alternative mechanism could also start with the vinylogous aldol reaction, forming first the C–C bond at the meso position of the pseudo xanthene skeleton. A sequence oxa-Michael addition/retro-Michael/ dehydration would lead to the same key intermediate 10 featuring the aldehyde function necessary for further derivatization (Scheme 2B). Reaction with indolium species 11 would then provide methylated dyes 18 and phenolic dihydroxanthene NIR fluorophores 19 after final deprotection (Scheme 2C). An overview of the reaction conditions screened in order to achieve the cascade reaction is presented in Table 1. We observed that the nature of the β-haloenal 9 used did not significantly affect the yields obtained so that eventually 9a was preferred over 9b because it was more stable and easier to handle.18 Organic bases were tried first and showed a moderate level of success. A catalytic amount of DABCO in CH2Cl2 did not form any desired product (entry 1) but traces of aldehyde 10 could be isolated using DBU (entry 2). Larger quantities of DABCO (2 eq.) in DMF slightly improved the yield (entry 3) and pyrrolidine could increase it up to 17% (entry 4). This encouraging result prompted us to try aminocatalysis conditions in the presence of L-proline or Jørgensen–Hayashi’s diphenylprolinol trimethylsilyl ether catalyst19 to promote enamine and iminium activation. Unfortunately, these con-
Scheme 2 A. Working hypothesis relying on a cascade oxa-Michael/retro-Michael/vinylogous aldol/dehydration to forge the phenolic dihydroxanthene scaffold; B. Alternative mechanism cascade vinylogous aldol/oxa-Michael/retro-Michael/dehydration; C. Approach for a divergent, de novo synthesis of phenolic dihydroxanthene near-infrared emitting dyes.
Org. Biomol. Chem.
This journal is © The Royal Society of Chemistry 2015
View Article Online
Organic & Biomolecular Chemistry
Communication
Published on 06 July 2015. Downloaded by UNIVERSITY OF OTAGO on 06/07/2015 16:10:33.
Table 1 Optimization of the oxa-Michael/retro-Michael/vinylogous aldol/dehydration cascade sequencea
Entry
β-haloenal
Base (eq.)
Solvent
Yieldb (%)
1 2 3 4 5 6
9a 9a 9a 9a 9b 9b
CH2Cl2 CH2Cl2 DMF CH2Cl2 DMSO CHCl3
0
