Next Article in Journal
Effect of Extraction Methods on Polyphenols, Flavonoids, Mineral Elements, and Biological Activities of Essential Oil and Extracts of Mentha pulegium L.
Previous Article in Journal
Stabilizing the Exotic Carbonic Acid by Bisulfate Ion
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Molecules
Volume 27
Issue 1
10.3390/molecules27010012
molecules-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Color-Tunable Indolizine-Based Fluorophores and Fluorescent pH Sensor

by
Taegwan Kim
1 and
Jonghoon Kim
1,2,*
1
Department of Chemistry, Soongsil University, Seoul 06978, Korea
2
Integrative Institute of Basic Science, Soongsil University, Seoul 06978, Korea
*
Author to whom correspondence should be addressed.
Molecules 2022, 27(1), 12; https://doi.org/10.3390/molecules27010012
Submission received: 3 December 2021 / Revised: 17 December 2021 / Accepted: 20 December 2021 / Published: 21 December 2021
(This article belongs to the Section Chemical Biology)
Browse Figures
Review Reports Versions Notes

Abstract

:
A new fluorescent indolizine-based scaffold was developed using a straightforward synthetic scheme starting from a pyrrole ring. In this fluorescent system, an N,N-dimethylamino group in the aryl ring at the C-3 position of indolizine acted as an electron donor and played a crucial role in inducing a red shift in the emission wavelength based on the ICT process. Moreover, various electron-withdrawing groups, such as acetyl and aldehyde, were introduced at the C-7 position of indolizine, to tune and promote the red shift of the emission wavelength, resulting in a color range from blue to orange (462–580 nm). Furthermore, the ICT effect in indolizine fluorophores allowed the design and development of new fluorescent pH sensors of great potential in the field of fluorescence bioimaging and sensors.

Graphical Abstract

1. Introduction

Optical fluorescence imaging for the non-invasive study of living systems has contributed to the organic fluorophore and fluorescent probe exploitation in the fields of life sciences, medicine, chemical biology, and biotechnology [1,2,3,4,5,6,7,8]. Due to a high demand for the development of novel fluorescent core skeletons, considerable progress has been made over the past few decades, which has resulted in the synthesis of various new fluorophores characterized by specific photophysical properties allowing the development of novel fluorescent probes [9,10,11,12,13]. As an example, indolizine-based organic molecules were successfully exploited in new fluorophores and fluorescent probes [14,15,16,17,18,19,20]. Synthetic methods for various types of indolizine scaffolds have led to the development of well-defined fluorophore scaffolds with an in-depth studied structure-fluorescence relationship. Figure 1a reports an emission-tunable and predictable indolizine-based fluorescent core skeleton, 9-aryl-1,2-dihydropyrrolo[3,4-b]indolizin-3-one, named Seoul-Fluor, and various fluorescent probes developed by Park et al. Perturbation of the electron density of the substituents at the C-7 and C-9 positions in Seoul-Fluor dramatically changed the emission wavelength, allowing a systematic tuning of the emission in the range 420–613 nm [21,22,23]. Moreover, computational studies provided guidelines for tuning the emission wavelength of Seoul-Fluor and developing Seoul-Fluor analogs with improved molar absorptivity [24]. More recently, Kim et al. prepared a new colorful fluorescent scaffold, named Kaleidolizine (KIz), with unique aggregation-induced emission (AIE) properties, on which the resulting fluorogenic probes were based [25,26]. However, previous studies typically focused on the synthesis of indolizine via the cycloaddition reaction of pyridinium ylide with alkenes or alkynes, with results critically influenced by the substrates [27,28,29]. Thus, most of these reactions are limited by incorporation of various functional groups at specific indolizine positions [30]. To address this, substantial efforts have been devoted to the development of straightforward synthetic methods for the incorporation of functional groups into indolizine. An indolizine-based full-color-tunable (405–616 nm) fluorophore, named C3-Indo-Fluoro, was developed by Lan et al. through the metal catalytic C–H activation of indolizine [31]. Despite the notable advances in indolizine-based fluorophores, the development of a straightforward synthetic strategy for the versatile functionalization of the indolizine core, providing convenient access to new indolizine fluorophores with unique photophysical properties, is challenging. To this end, we conceived a new synthetic route to prepare 3,7-disubstitued indolizine from the pyrrole ring (Figure 1b). The methodology reported in this study revealed a systematic tunability of the emission wavelength (462–580 nm) through the perturbation of the substituent electron density based on the intramolecular charge transfer (ICT) process [32]. Finally, the introduction of a pH-sensitive functional group into this new indolizine-based fluorescent scaffold allowed the successful development of a fluorescent pH sensor.

2. Results and Discussion

Based on the retrosynthetic analysis, the synthetic pathway employed to provide 3,7-disubstitued indolizine compounds is illustrated in Scheme 1. First, the SN2 reaction of 1H-pyrrole-2-carboxaldehyde (1) with ethyl 4-bromobutyrate afforded compound 2, followed by aldol condensation using sodium hydride (NaH), resulting in the cyclic compound 3. Regioselective bromination of compound 3 with N-bromosuccinimide (NBS) generated compound 4. Afterwards, the dehydrogenative oxidation with manganese dioxide (MnO2) generated the indolizine compound 5 with bromo and ester functional groups at the C-3 and C-7 positions, respectively, which allow facile conversion into other functional groups. Finally, to introduce an aryl moiety into the C-3 position for the new π-conjugated indolizine, we adopted the Suzuki cross-coupling reaction. The optimized cross-coupling conditions were conventional heating in water in the presence of palladium(II) acetate (Pd(OAc)2) and tetrabutylammonium bromide (TBAB) without any organic co-solvents [33]. Under the optimized reaction conditions, we successfully prepared a series of 7-aryl substituted indolizine-based fluorescent compounds with various R1 substituents to change the electronic characteristics of the phenyl ring.
Our systematic evaluation of the structure–photophysical property relationship included the maximum absorption and emission wavelengths, Stokes shifts, and quantum yields measurements (Table 1). Normalized absorption and emission spectra of each compound are shown in Figure S1 in Supplementary Materials. Interestingly, for all the functional groups introduced at the R1 position (from an electron-donating group, EDG, such as the methoxy group, to an electron-withdrawing group, EWG, such as the cyano group), the maximum absorption and emission wavelength change was negligible. However, compound 9 bearing an N,N-dimethylamino group, which is the most electron-donating group of the series, exhibited a red shift in the emission spectra (533 nm), compared to other compounds (462–492 nm). We hypothesized that this feature originated from the ICT process between the N,N-dimethylamino group at the R1 position and the ester group at the C-7 position of indolizine. Contrarily to other donating groups, with the ester group acting as an electron acceptor, in this electron-push-pull π-conjugated indolizine-based system, the N,N-dimethylamino group acted as a powerful electron donor, triggering an ICT process. In the electronic push-pull system in fluorescent scaffolds, the effects of the substituent electronic characteristics on the energy gap between the highest occupied molecular orbital (HOMO) and the lowest unoccupied molecular orbital (LUMO) resulted in a red or blue shift in emission [34]. In the case of compound 9, the N,N-dimethylamino group on the phenyl ring might increase the HOMO energy level, with a minimal effect on the LUMO energy level, causing a red shift of the emission wavelength. Moreover, ICT causes a large change in the excited state dipole moment, promoting a solvent-dependent emission shift [35]. To determine the effect of ICT, we further studied the solvatochromic fluorescence emission change of compound 9. As shown in Figure 2, the solvent polarity had limited effects on the absorption wavelength. Conversely, an increase in solvent polarity induced a bathochromic shift of the emission wavelength from 485 nm in non-polar diethyl ether to 533 nm in polar methanol. This positive emission solvatochromism suggested that the ICT process was the dominant mechanism for the red-shift emission of compound 9.
Based on this observation, we designed and synthesized indolizine-based fluorescent compounds to tune the emission wavelength, which can cover a wide range of colors. As the N,N-dimethylamino group at the R1 position could successfully reduce the HOMO-LUMO energy gap by increasing the HOMO energy level, we hypothesized that the EWG group at the C-7 position influences the stabilization of the LUMO state, allowing a red-shift emission. As shown in Scheme 2, to introduce a stronger electron-withdrawing group than the ester group at the C-7 position, compound 9 was subjected to a hydrolysis reaction in the presence of sodium hydroxide (NaOH), to afford compound 14. Compound 15 was generated through an amide coupling reaction with N,O-dimethylhydroxylamine after the activation of carboxylic acids using carbonyldiimidazole (CDI). The N-methoxy-N-methyl amide group (Weinreb amide) was converted into aldehyde or ketone after treatment with lithium aluminum hydride (LiAlH4) or methylmagnesium bromide (CH3MgBr), to produce compounds 16 and 17, respectively. Compounds 16 and 17 bearing aldehyde and acetyl groups, respectively, which are stronger electron-withdrawing groups than ester, showed a 47 nm and 45 nm red shift of emission wavelength, respectively, compared to compound 9 (Table 1). Nevertheless, the carboxylic acid group at the C-7 position induced a 28 nm blue shift in emission (Table 1). This was reasonably due to the reduction of the electron-withdrawing strength upon the introduction of the carboxylic acid group at the C-7 position. Figure 3 clearly shows a positive solvatochromism (from 499 to 574 nm) and a reduction in fluorescence intensity under polar conditions in compound 17. These more considerable bathochromic shifts and changes in fluorescence intensity compared to compound 9 can be ascribed to the strengthening the ICT process through the introduction of more electron-withdrawing acetyl groups than the ester group at the C-7 position of the indolizine fluorophores. Overall, the synthesized new indolizine fluorophore compounds covered blue to orange-red color emission ranges (462–580 nm). The normalized emission spectra of representative compounds 12 (blue), 14 (green), 9 (yellow), and 16 (red-orange) are shown in Figure 4a. The colors, brought about irradiation at 365 nm, were visible to the naked eye, as shown in the fluorescence image (Figure 4b).
Because ICT processes are extensively used in the design of fluorescent probe [36,37,38,39], we hypothesized that the unique features of this indolizine-based fluorescent scaffold could lead to new applications for environmental changes as a fluorescent pH sensor. Therefore, compound 14 was selected as a potential pH sensor probe, owing to its pH-responsive functional group, namely the N,N-dimethylamino group, at the R1 position and carboxylic acid group at the C-7 position as well as considering high solubility in water solution. The protonation and deprotonation processes of these pH-responsive groups can induce changes in their electronic characteristics according to their pKa values, generating subsequent dramatic shifts in the emission wavelength. As an example, under acidic conditions, protonation of the dialkylamino group at the R1 position can promote the blue shift of the emission wavelength through the reduction of electron-donating characteristics. Nevertheless, under neutral conditions, the deprotonation of carboxylic acid can reduce the electron-donating character, causing a red-shifted emission. To demonstrate this, the emission response of compound 14 at different pH values (3.0–7.0) was examined. We observed a blue shift of the emission wavelength from 529 nm to 473 nm and an improvement of fluorescence intensity with the pH decreasing from 7.0 to 3.0 (Figure 5a,b). This result suggested that the protonation and deprotonation of the functional groups on indolizine fluorophores perturbed the electronic states, which could control the ICT process. In neutral conditions, the dimethylamino group could strengthen the ICT process compared to protonated dimethylamino group in acid conditions, leading to a red shift of the emission wavelength and reduction of emission intensity. As shown in Figure 5c, the emission intensity at 473 nm gradually decreased at higher pH values, revealing the potential of compound 14 as a fluorescent pH sensor. The inflection point of the sigmoid curve for compound 14 was pH 4.07. Besides compound 14, the emission response of compounds 9, 12, 16, and 17 at different pH values (3.0, 5.0, and 7.0) was examined. As expected, compound 9, containing a pH-responsive element such as a dimethylamino group, clearly shows a blue shift of the emission wavelength from 522 nm to 476 nm and improvement of fluorescence intensity with the pH decreasing from 7.0 to 3.0, similar to compound 14. In addition, compounds 16 and 17 exhibit a more dramatic reduction of emission intensity with the pH increasing from 3.0 to 7.0 compared to compound 9, which is caused by strengthening the ICT process through the introduction of a more electron-withdrawing group (aldehyde and acetyl group) at the C-7 position, consistent with the findings in a previous solvatochromic study (Figure 6c,d). On the other hand, in the case of compound 12 containing cyano group at the R1 position, the emission wavelength and intensity cannot be obviously changed depending on the pH value, which can be ascribed to less intense ICT through the introduction of an electron-withdrawing group into the R1 position (Figure 6b). Thus, we clearly demonstrated that the ICT phenomenon in the indolizine fluorophore allowed the successful development of fluorescent pH sensors by introducing pH-responsive functional groups such as N,N-dimethylamino, revealing the potential of this new indolizine fluorophore for versatile fluorescent probes to explore biological systems and environmental changes.

3. Materials and Methods

All commercially available reagents were used without further purification unless noted otherwise. Dried solvents were passed through a solvent purification system equipped with activated alumina columns (glass contours). NMR analyses were carried out using a JEOL ECZ500/S1 spectrometer (500 MHz, Jeol, Tokyo, Japan). The chemical shifts of the proton (1H) and carbon (13C) NMR spectra were reported in parts per million (δ), with respect to the internal standard tetramethylsilane (TMS) or to the residual solvent peak (CDCl3, 1H: 7.26, 13C: 77.00; DMSO-d6, 1H: 2.49, 13C: 39.52; CD2Cl2, 1H: 5.32, 13C: 53.8). The multiplicities of the 1H NMR peaks are reported as follows: s (singlet), d (doublet), t (triplet), q (quartet), quintet, m (multiplet), dd (doublet of doublets), dt (doublet of triplets), td (triplet of doublets), and br s (broad singlet). Signals marked with an asterisk (*) correspond to the peaks assigned to the minor rotamer conformation. The coupling constants (J) were reported in Hz. High-resolution mass spectrometry (HRMS) spectra were obtained on a compact QToF spectrometer (Bruker, Billerica, MA, USA) using the electrospray ionization (ESI) method. Low-resolution mass spectrometry (LRMS) spectra were obtained on a compact mass spectrometer (Advion, Ithaca, NY, USA) using the atmospheric pressure chemical ionization (APCI) method. To monitor the progress of the reactions, analytical thin-layer chromatography (TLC) analysis was performed using glass plates pre-coated with silica gel (60 F254, Merck, Darmstadt, Germany), and the components were observed under UV light (254 and 365 nm). Flash column chromatography was performed using silica gel 60 (230–400 mesh, Merck, Darmstadt, Germany). Absorption spectra were recorded using a UV-3101PC spectrophotometer (Shimadzu, Kyoto, Japan) or V-770 spectrophotometer (Jasco, Easton, MD, USA). Fluorescence emission spectra were obtained with a RF-5301PC Fluorescence spectrophotometer (Shimadzu, Kyoto, Japan), and the absolute quantum yield was measured by QE-2000 (Otsuka Electronics, Osaka, Japan).
Ethyl 4-(2-formyl-1H-pyrrol-1-yl)butanoate (2). To a solution of 1H-pyrrole-2-carbaldehyde (4.33 g, 45.5 mmol) in acetonitrile (ACN; 230 mL), cesium carbonate (Cs2CO3; 29.6 g, 91.0 mmol) was added. The reaction mixture was stirred at room temperature (r.t.) for 30 min. Ethyl 4-bromobutyrate (7.8 mL, 54.6 mmol) was slowly added to the resultant solution at 0 °C, and the reaction mixture was stirred at r.t. for 18 h. The crude reaction mixture was partitioned into water and ethyl acetate (EtOAc). The aqueous layer was extracted twice with ethyl acetate (EtOAc). The combined organic layers were dried over anhydrous magnesium sulfate (MgSO4), filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded ethyl 4-(2-formyl-1H-pyrrol-1-yl)butanoate (2) (9.66 g, 44.6 mmol, 98%, Rf = 0.70 (EtOAc/hexane = 1:5) as a pale yellow oil. 1H NMR (500 MHz, CDCl3) δ (ppm): 9.52 (d, J = 1.0 Hz, 1H), 6.94–6.92 (m, 1H), 6.22 (dd, J = 4.0, 3.0 Hz, 1H), 4.37 (t, J = 7.0 Hz, 2H), 4.12 (q, J = 7.0 Hz, 2H), 2.27 (t, J = 7.5 Hz, 2H), 2.08 (quintet, J = 7.5 Hz, 2H), 1.25 (t, J = 7.0 Hz, 3H). 13C NMR (125 MHz, CDCl3) δ (ppm): 179.3, 172.8, 131.5, 131.3, 125.0, 109.7, 60.5, 47.9, 30.8, 26.4, 14.2. LRMS (APCI +): calcd for C11H16NO3+ [M + H]+ 210.1, found 210.3.
Ethyl 5,6-dihydroindolizine-7-carboxylate (3). To a solution of compound 2 (1.96 g, 9.35 mmol) in ethanol (EtOH; 500 mL), sodium hydride (NaH; 0.41 g of 60 % dispersion in paraffin liquid, 10.3 mmol) was added portion-wise at 0 °C. The reaction mixture was stirred at 70 °C for 18 h and then cooled to r.t. The resultant mixture was quenched with saturated ammonium chloride (NH4Cl, aq). EtOH was evaporated under reduced pressure, and the crude mixture was partitioned into water and EtOAc. The aqueous layer was extracted twice with EtOAc. The combined layer was dried over anhydrous MgSO4, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded ethyl 5,6-dihydroindolizine-7-carboxylate (3) (1.39 g, 7.27 mmol, 78%, Rf = 0.48 (EtOAc/hexane = 1:5)) as a pale white oil. 1H NMR (500 MHz, CDCl3) δ (ppm): 7.47 (s, 1H), 6.73 (t, J = 1.5 Hz, 1H), 6.35 (dd, J = 3.5, 1.5 Hz, 1H), 6.21 (dd, J = 4.0, 3.0 Hz, 1H), 4.25 (q, J = 7.0 Hz, 2H), 4.01(t, J = 7.0 Hz, 2H), 2.80 (td, J = 7.5, 1.5 Hz, 2H), 1.33 (t, J = 8.5 Hz, 3H). 13C NMR (125 MHz, CDCl3) δ (ppm): 167.1, 128.2, 128.0, 123.9, 119.3, 111.8, 109.7, 60.4, 43.9, 23.6, 14.3. LRMS (APCI +): calcd for C11H14NO2+ [M + H]+ 192.1, found 192.3.
Ethyl 3-bromo-5,6-dihydroindolizine-7-carboxylate (4). To a solution of compound 3 (0.50 g, 2.63 mmol) in tetrahydrofuran (THF; 2.6 mL), N-bromosuccinimide (NBS; 0.47 g, 2.63 mmol) was added portion-wise at r.t. After the reaction was complete, as indicated by TLC, the reactant was extracted with EtOAc twice, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded ethyl 3-bromo-5,6-dihydroindolizine-7-carboxylate (4) (0.64 g, 2.37 mmol, 90%, Rf = 0.43 (EtOAc/hexane = 1:5)) as a pale white oil. 1H NMR (500 MHz, CDCl3) δ (ppm): 7.37 (t, J = 1.0 Hz, 1H), 6.33 (d, J = 4.0 Hz, 1H), 6.22 (d, J = 3.5 Hz, 1H), 4.25 (q, J = 7.5 Hz, 2H), 3.97 (t, J = 7.5 Hz, 2H), 2.80 (td, J = 7.5, 1.5 Hz, 2H), 1.33 (t, J = 7.0 Hz, 3H). 13C NMR (125 MHz, CDCl3) δ (ppm): 166.9, 129.6, 127.2, 119.5, 112.6, 112.2, 106.7, 60.5, 42.5, 23.2, 14.4. LRMS (APCI +): calcd for C11H13BrNO2+ [M + H]+ 270.0, found 270.2.
Ethyl 3-bromoindolizine-7-carboxylate (5). To a solution of compound 4 (0.71 g, 3.0 mmol) in dichloromethane (DCM; 150 mL) manganese dioxide (MnO2; 13.0 g, 150 mmol) was added. The reaction mixture was stirred under reflux for 18 h and then cooled to r.t. The mixture was filtered through Celite and solvent was evaporated. Subsequent silica-gel flash column chromatography afforded ethyl 3-bromoindolizine-7-carboxylate (5) (0.59 g, 2.20 mmol, 73%, Rf = 0.46 (EtOAc/hexane = 1:6)) as a pale-yellow solid. 1H NMR (500 MHz, CDCl3) δ (ppm): 8.17 (s, 1H), 7.94 (d, J = 7.5 Hz, 1H), 7.21 (dd, J = 7.5, 2.0 Hz, 1H), 6.90 (d, J = 4.5 Hz, 1H), 6.77 (d, J = 4.0 Hz, 1H), 4.38 (q, J = 7.5 Hz, 2H), 1.41 (t, J = 7.5 Hz, 3H). 13C NMR (125 MHz, CDCl3) δ (ppm): 165.8, 132.4, 122.7, 122.1, 118.7, 117.5, 110.4, 105.3, 96.1, 61.0, 14.4. LRMS (APCI +): calcd for C11H11BrNO2+ [M + H]+ 268.0, found 268.2.
Ethyl 3-phenylindolizine-7-carboxylate (6). To a solution of compound 5 (0.27 g, 1.0 mmol) in water (3 mL), phenyl boronic acid (0.13 g, 1.1 mmol), potassium carbonate (K2CO3; 0.35 g, 2.5 mmol), tetrabutylammonium bromide (TBAB; 0.32 g, 1.0 mmol), and palladium(II) diacetate (Pd(OAc)2; 4 mg, 0.02 mmol) were added. The reaction mixture was stirred at 70 °C for 2 h. The crude mixture was extracted twice with EtOAc. The combined organic layers were dried over anhydrous MgSO4, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded ethyl 3-phenylindolizine-7-carboxylate (6) (0.22 g, 0.82 mmol, 82%, Rf = 0.43 (EtOAc/hexane = 1:6)) as a yellow oil. 1H NMR (500 MHz, CDCl3) δ (ppm): 8.24 (s, 1H), 8.23 (dt, J = 7.5, 0.5 Hz, 1H), 7.58 (dd, J = 8.5, 1.5 Hz, 2H), 7.50 (t, J = 7.5 Hz, 2H), 7.39 (tt, J = 7.0, 1.5 Hz, 1H), 7.07 (dd, J = 7.5, 1.5 Hz, 1H), 6.96 (d, J = 4.0 Hz, 1H), 6.83 (d, J = 4.5 Hz, 1H), 4.38 (q, J = 7.5 Hz, 2H), 1.41 (t, J = 7.0 Hz, 3H). 13C NMR (125 MHz, CD2Cl2) δ (ppm): 166.1, 133.0, 132.1, 129.4, 128.5, 128.4, 128.1, 123.5, 121.8, 118.9, 116.2, 110.0, 105.6, 61.1, 14.5. HRMS (ESI +): calcd for C17H15NO2Na+ [M + Na]+ 288.1000, found 288.0995.
Ethyl 3-(p-tolyl)indolizine-7-carboxylate (7). To a solution of compound 5 (0.27 g, 1.0 mmol) in water (3 mL), p-tolylboronic acid (0.15 g, 1.1 mmol), K2CO3 (0.35 g, 2.5 mmol), TBAB (0.32 g, 1.0 mmol), and Pd(OAc)2 (4 mg, 0.02 mmol) were added. The reaction mixture was stirred at 70 °C for 2 h. The crude mixture was extracted twice with EtOAc. The combined organic layers were dried over anhydrous MgSO4, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded ethyl 3-(p-tolyl)indolizine-7-carboxylate (7) (0.27g, 0.95 mmol, 95%, Rf = 0.43 (EtOAc/hexane = 1:6)) as a yellow oil. 1H NMR (500 MHz, CDCl3) δ (ppm): 8.24 (s, 1H), 8.19 (dt, J = 7.5, 1.0 Hz, 1H), 7.47 (d, J = 8.0 Hz, 2H), 7.31 (d, J = 8.0 Hz, 2H), 7.05 (dd, J = 7.5, 2.0 Hz, 1H), 6.93 (d, J = 4.0 Hz, 1H), 6.81 (d, J = 3.5 Hz, 1H), 4.38 (q, J = 7.0 Hz, 2H), 2.43 (s, 3H), 1.41 (t, J = 7.0 Hz, 3H). 13C NMR (125 MHz, CDCl3) δ (ppm): 166.1, 137.7, 132.3, 129.7, 128.8, 128.2, 128.1, 123.5, 121.4, 118.2, 115.7, 109.7, 105.2, 60.8, 21.3, 14.4. HRMS (ESI +): calcd for C18H17NO2Na+ [M + Na]+ 302.1157, found 302.1151.
Ethyl 3-(4-methoxyphenyl)indolizine-7-carboxylate (8). To a solution of compound 5 (0.27 g, 1.0 mmol) in water (3 mL), (4-methoxyphenyl)boronic acid (0.17 g, 1.1 mmol), K2CO3 (0.35 g, 2.5 mmol), TBAB (0.32 g, 1.0 mmol), and Pd(OAc)2 (4 mg, 0.02 mmol) were added. The reaction mixture was stirred at 70 °C for 2 h. The crude mixture was extracted twice with EtOAc. The combined organic layers were dried over anhydrous MgSO4, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded ethyl 3-(4-methoxyphenyl)indolizine-7-carboxylate (8) (0.25 g, 0.83 mmol, 83%, Rf = 0.25 (EtOAc/hexane = 1:6)) as a yellow solid. 1H NMR (500 MHz, CDCl3) δ (ppm): 8.23 (s, 1H), 8.14 (d, J = 7.5 Hz, 1H), 7.49 (d, J = 8.5 Hz, 2H), 7.06–7.03 (m, 3H), 6.89 (d, J = 4.5 Hz, 1H), 6.81 (d, J = 4.0 Hz, 1H), 4.37 (q, J = 7.0 Hz, 2H), 3.88 (s, 3H), 1.41 (t, J = 7.5 Hz, 3H). 13C NMR (125 MHz, CDCl3) δ (ppm): 166.1, 159.2, 132.1, 129.6, 128.0, 124.1, 123.4, 121.2, 118.0, 115.5, 114.5, 109.6, 105.1, 60.8, 55.4, 14.4. HRMS (ESI +): calcd for C18H17NO3Na+ [M + Na]+ 318.1106, found 318.1101.
Ethyl 3-(4-(dimethylamino)phenyl)indolizine-7-carboxylate (9). To a solution of compound 5 (0.27 g, 1.0 mmol) in water (3 mL), (4-(dimethylamino)phenyl)boronic acid (0.18 g, 1.1 mmol), K2CO3 (0.35 g, 2.5 mmol), TBAB (0.32 g, 1.0 mmol), and Pd(OAc)2 (4 mg, 0.02 mmol) were added. The reaction mixture was stirred at 70 °C for 2 h. The crude mixture was extracted twice with EtOAc. The combined organic layers were dried over anhydrous MgSO4, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded ethyl 3-(4-(dimethylamino)phenyl)indolizine-7-carboxylate (9) (0.24 g, 0.77 mmol, 77%, Rf = 0.25 (EtOAc/hexane = 1:6)) as a yellow solid. 1H NMR (500 MHz, CDCl3) δ (ppm): 8.22 (s, 1H), 8.17 (d, J = 7.5 Hz, 1H), 7.44 (d, J = 9.0 Hz, 2H), 7.02 (dd, J = 7.5, 2.0 Hz, 1H), 6.87 (d, J = 4.0 Hz, 1H), 6.84 (d, J = 9.0 Hz, 2H), 6.80 (d, J = 4.0 Hz, 1H), 4.37 (q, J = 7.5 Hz, 2H), 3.03 (s, 6H), 1.41 (t, J = 7.5 Hz, 3H). 13C NMR (125 MHz, CDCl3) δ (ppm): 166.2, 149.9, 131.8, 129.1, 128.8, 123.4, 121.3, 119.3, 117.4, 115.1, 112.5, 109.3, 105.1, 60.7, 40.4, 14.4. HRMS (ESI +): calcd for C19H21N2O2+ [M + H]+ 309.1603, found 309.1598.
Ethyl 3-(4-fluorophenyl)indolizine-7-carboxylate (10). To a solution of compound 5 (0.27 g, 1.0 mmol) in water (3 mL), (4-fluorophenyl)boronic acid (0.15 g, 1.1 mmol), K2CO3 (0.35 g, 2.5 mmol), TBAB (0.32 g, 1.0 mmol), and Pd(OAc)2 (4 mg, 0.02 mmol) were added. The reaction mixture was stirred at 70 °C for 2 h. The crude mixture was extracted twice with EtOAc. The combined organic layers were dried over anhydrous MgSO4, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded ethyl 3-(4-fluorophenyl)indolizine-7-carboxylate (10) (0.24 g, 0.84 mmol, 84%, Rf = 0.43 (EtOAc/hexane = 1:6)) as a yellow solid. 1H NMR (500 MHz, CDCl3) δ (ppm): 8.24 (s, 1H), 8.12 (d, J = 7.5 Hz, 1H), 7.55–7.51 (m, 2H), 7.20 (t, J = 8.5 Hz, 2H), 7.07 (dd, J = 7.5, 2.0 Hz, 1H), 6.91 (d, J = 4.0 Hz, 1H), 6.81 (d, J = 4.0 Hz, 1H), 4.38 (q, J = 7.0 Hz, 2H), 1.41 (t, J = 7.0 Hz, 3H). 13C NMR (125 MHz, CDCl3) δ (ppm): 165.9, 163.2, 161.2, 132.4, 130.0, 129.9, 127.8, 127.0, 123.5, 121.1, 118.5, 116.2, 116.0, 115.9, 109.9, 105.2, 60.8, 14.4. LRMS(ESI+): calcd for C17H15FNO2+ [M+H]+ 284.1, found 284.1. HRMS (ESI +): calcd for C17H14FNO2Na+ [M + Na]+ 306.0906, found 306.0901.
Ethyl 3-(4-formylphenyl)indolizine-7-carboxylate (11). To a solution of compound 5 (0.27 g, 1.0 mmol) in water (3 mL), (4-formylphenyl)boronic acid (0.16 g, 1.1 mmol), K2CO3 (0.35 g, 2.5 mmol), TBAB (0.32 g, 1.0 mmol), and Pd(OAc)2 (4 mg, 0.02 mmol) were added. The reaction mixture was stirred at 70 °C for 90 min. The crude mixture was extracted twice with EtOAc. The combined organic layers were dried over anhydrous MgSO4, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded ethyl 3-(4-formylphenyl)indolizine-7-carboxylate (11) (0.19 g, 0.63 mmol, 63%, Rf = 0.11 (EtOAc/hexane = 1:6)) as a yellow solid. 1H NMR (500 MHz, CDCl3) δ (ppm): 10.05 (s, 1H), 8.31 (d, J = 7.5 Hz, 1H), 8.26 (s, 1H), 8.00 (d, J = 6.5 Hz, 2H), 7.76 (d, J = 8.0 Hz, 2H), 7.15 (dd, J = 7.5, 1.5 Hz, 1H), 7.07 (d, J = 4.0 Hz, 1H), 6.85 (d, J = 4.0 Hz, 1H), 4.39 (q, J = 7.5 Hz, 2H), 1.41 (t, J = 6.5 Hz, 3H). 13C NMR (125 MHz, CDCl3) δ (ppm): 191.4, 165.7, 137.6, 134.9, 133.8, 130.6, 127.7, 126.7, 123.5, 121.5, 119.6, 117.2, 110.6, 106.1, 61.0, 14.4. HRMS (ESI +): calcd for C18H15NO3Na+ [M + Na]+ 316.0950, found 316.0944.
Ethyl 3-(4-cyanophenyl)indolizine-7-carboxylate (12). To a solution of compound 5 (0.27 g, 1.0 mmol) in water (3 mL), (4-cyanophenyl)boronic acid (0.16 g, 1.1 mmol), K2CO3 (0.35 g, 2.5 mmol), TBAB (0.32 g, 1.0 mmol), and Pd(OAc)2 (4 mg, 0.02 mmol) were added. The reaction mixture was stirred at 70 °C for 2 h. The crude mixture was extracted twice with EtOAc. The combined organic layers were dried over anhydrous MgSO4, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded ethyl 3-(4-cyanophenyl)indolizine-7-carboxylate (12) (0.19 g, 0.66 mmol, 66%, Rf = 0.14 (EtOAc/hexane = 1:6)) as a yellow solid. 1H NMR (500 MHz, CDCl3) δ (ppm): 8.26–8.25 (m, 2H), 7.77 (d, J = 8.5 Hz, 2H), 7.70 (d, J = 8.5 Hz, 2H), 7.15 (dd, J = 7.5, 1.5 Hz, 1H), 7.03 (d, J = 4.5 Hz, 1H), 6.85 (d, J = 4.5 Hz, 1H), 4.39 (q, J = 7.0 Hz, 2H), 1.41 (t, J = 7.0 Hz, 3H). 13C NMR (125 MHz, CDCl3) δ (ppm): 165.6, 136.2, 133.8, 132.9, 127.8, 126.0, 123.5, 121.3, 119.8, 118.7, 117.1, 110.7, 110.6, 106.1, 61.0, 14.3. HRMS (ESI +): calcd for C18H14N2O2Na+ [M + Na]+ 313.0953, found 313.0947.
Ethyl 3-(3-cyanophenyl)indolizine-7-carboxylate (13). To a solution of compound 5 (0.27 g, 1.0 mmol) in water (3 mL), (3-cyanophenyl)boronic acid (0.16 g, 1.1 mmol), K2CO3 (0.35 g, 2.5 mmol), TBAB (0.32 g, 1.0 mmol), and Pd(OAc)2 (4 mg, 0.02 mmol) were added. The reaction mixture was stirred at 70 °C for 2 h. The crude mixture was extracted twice with EtOAc. The combined organic layers were dried over anhydrous MgSO4, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded ethyl 3-(3-cyanophenyl)indolizine-7-carboxylate (13) (0.16 g, 0.54 mmol, 54%, Rf = 0.14 (EtOAc/hexane = 1:6)) as a yellow solid. 1H NMR (500 MHz, CDCl3) δ (ppm): 8.26 (s, 1H), 8.17 (d, J = 7.0 Hz, 1H), 7.86 (t, J = 1.5 Hz, 1H), 7.82 (dt, J = 7.5, 1.5 Hz, 1H), 7.65 (dt, J = 7.5, 1.5 Hz, 1H), 7.61 (t, J = 8.0 Hz, 1H), 7.14 (dd, J = 7.5, 1.5 Hz, 1H), 6.99 (d, J = 4.0 Hz, 1H), 6.84 (d, J = 4.0 Hz, 1H), 4.39 (q, J = 7.0 Hz, 2H), 1.41 (t, J = 7.0 Hz, 3H). 13C NMR (125 MHz, CDCl3) δ (ppm): 165.7, 133.3, 133.1, 131.9, 131.3, 130.8, 130.0, 125.4, 123.5, 120.9, 119.4, 118.4, 116.6, 113.5, 110.6, 105.7, 61.0, 14.4. HRMS (ESI +): calcd for C18H14N2O2Na+ [M + Na]+ 313.0953, found 313.0947.
3-(4-(Dimethylamino)phenyl)indolizine-7-carboxylic acid (14). To a solution of compound 9 (0.37 g, 1.2 mmol) in EtOH (4 mL) and water (2 mL), sodium hydroxide (NaOH; 0.10 g, 2.4 mmol) was added. The reaction mixture was stirred at 60 °C. After the reaction was complete, as indicated by TLC, the mixture was acidified with 1 N hydrogen chloride (HCl; aq) to give a yellow precipitate. The resulting precipitate was filtered and dried to afford desired product, 3-(4-dimethylamino)phenyl)indolizine-7-carboxylic acid (14) (0.29 g, 1.0 mmol, 84%, Rf = 0.20 (EtOAc/hexane = 2:1)) as a yellow solid. 1H NMR (500 Mz, DMSO-d6) δ (ppm): 12.67 (br s, 1H), 8.25 (d, J = 7.5 Hz, 1H), 8.15 (s, 1H), 7.43 (d, J = 9.0 Hz, 2H), 6.96 (dd, J = 7.5, 1.5 Hz, 1H), 6.92 (d, J = 4.0 Hz, 1H), 6.86–6.84 (m, 3H), 2.96 (s, 6H). 13C NMR (125 MHz, CDCl3)) δ (ppm): 171.5, 150.1, 131.8, 129.6, 129.2, 124.9, 121.4, 119.2, 116.1, 115.5, 112.6, 109.4, 106.2, 40.4. HRMS (ESI +): calcd for C17H17N2O2+ [M + H]+ 281.1290, found 281.1285.
3-(4-(Dimethylamino)phenyl)-N-methoxy-N-methylindolizine-7-carboxamide (15). To a solution of compound 14 (0.36 g, 1.3 mmol) in THF (13 mL), 1,1′-carbonyldiimidazole (CDI; 0.25 g, 1.6 mmol) was added at 0 °C. The reaction mixture was stirred at r.t for 30 min. To the resultant mixture, N,O-dimethylhydroxylamine hydrochloride (0.15 g, 1.5 mmol) was added at r.t and stirred for 18 h. The reaction mixture was partitioned into 1 N sodium hydroxide (NaOH; aq) solution and EtOAc. The aqueous solution was extracted twice with EtOAc. The combined organic solutions were washed with 1N NaOH, dried anhydrous MgSO4, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded 3-(4-(dimethylamino)phenyl)-N-methoxy-N-methylindolizine-7-carboxamide (15) (0.24 g, 0.75 mmol, 59%, Rf = 0.22 (EtOAc/hexane = 1:1)) as an orange oil. 1H NMR (500 MHz, CDCl3) δ (ppm): 8.16 (d, J = 7.5 Hz, 1H), 8.02 (s, 1H), 7.44 (d, J = 9.0 Hz, 2H), 6.89 (dd, J = 7.5, 1.5 Hz, 1H), 6.86–6.83 (m, 3H), 6.72 (d, J = 4.0, 1H), 3.64/3.62* (s, 3H), 3.39/3.38* (s, 3H), 3.05*/3.03 (s, 6H). LRMS (APCI +): calcd for C19H22N3O2+ [M + H]+ 324.2, found 324.2.
3-(4-(Dimethylamino)phenyl)indolizine-7-carbaldehyde (16). To a solution of compound 15 (0.11 g, 0.35 mmol) in dry THF (3.5 mL), under an argon atmosphere, a solution of lithium aluminium hydride in THF (LiAlH4; 0.2 mL, 2.0 M) was added dropwise at 0 °C. The reaction mixture was stirred at r.t for 2 h. After the reaction was complete, as indicated by TLC, the mixture was quenched with saturated NH4Cl (aq), and then extracted twice with EtOAc. The combined organic layers were dried over anhydrous MgSO4, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded 3-(4-(dimethylamino)phenyl)indolizine-7-carbaldehyde (16) (47 mg, 0.18 mmol, 51%, Rf = 0.32 (EtOAc/hexane = 1:5)) as a yellow solid. 1H NMR (500 MHz, CDCl3) δ (ppm): 9.76 (s, 1H), 8.18 (d, J = 7.5 Hz, 1H), 7.90 (s, 1H), 7.44 (d, J = 9.0 Hz, 2H), 6.98 (dd, J = 7.0, 1.5 Hz, 1H), 6.94 (d, J = 4.0 Hz, 1H), 6.91 (d, J = 4.5 Hz, 1H), 6.84 (d, J = 9.0 Hz, 2H), 3.04 (s, 6H). 13C NMR (125 MHz, CDCl3) δ (ppm): 189.4, 150.2, 131.6, 130.8, 129.2, 128.7, 125.5, 122.3, 118.7, 115.8, 112.3, 107.7, 106.3, 40.3. HRMS (ESI +): calcd for C17H17N2O+ [M + H]+ 265.1341, found 265.1335.
1-(3-(4-(Dimethylamino)phenyl)indolizin-7-yl)ethan-1-one (17). To a solution of compound 15 (0.11 g, 0.35 mmol) in dry THF (3.5 mL), under an argon atmosphere, a solution of methylmagnesium bromide in diethyl ether (CH3MgBr; 0.35 mL, 3.0 M) was added dropwise at 0 °C. The reaction mixture was stirred at 0 °C for 2 h. After the reaction was complete, as indicated by TLC, the mixture was quenched with saturated NH4Cl (aq), and then extracted twice with EtOAc. The combined organic layers were dried over anhydrous MgSO4, filtered, and evaporated. Subsequent silica-gel flash column chromatography afforded 1-(3-(4-(dimethylamino)phenyl)indolizin-7-yl)ethan-1-one (17) (78 mg, 0.28 mmol, 81%, Rf = 0.26 (EtOAc/hexane = 1:3)) as a yellow solid. 1H NMR (500 MHz, CDCl3) δ (ppm): 8.16 (d, J = 8.0 Hz, 1H), 8.08 (d, J = 1.0 Hz, 1H), 7.44 (d, J = 8.5 Hz, 2H), 7.07 (dd, J = 7.0, 1.5 Hz, 1H), 6.88 (d, J = 4.0 Hz, 1H), 6.85 (d, J = 5.5 Hz, 2H), 6.83 (s, 1H), 3.03 (s, 6H), 2.58 (s, 3H). 13C NMR (125 MHz, CDCl3) δ (ppm): 195.6, 150.1, 131.4, 129.6, 129.2, 125.2, 123.4, 121.6, 119.1, 115.4, 112.5, 108.1, 106.4, 40.4, 25.6. HRMS (ESI +): calcd for C18H19N2O+ [M + H]+ 279.1497, found 279.1492.

4. Conclusions

In summary, a novel 3,7-disubstituted indolizine-based fluorophore, whose emission wavelengths were tunable to cover a wide color range from blue to red-orange (462–580 nm), was designed and synthesized through a straightforward synthetic scheme starting from the pyrrole ring. Most of the functional groups introduced into the aryl ring at the C-3 position of indolizine did not induce changes in the photophysical properties, such as the absorption and emission wavelength. However, compound 9 bearing an N,N-dimethylamino group at the aryl ring exhibited a pronounced red-shift emission spectra (533 nm), compared to the other compounds (462–492 nm). This result was caused by the ICT process between the N,N-dimethylamino group and the ester group at the C-7 position in the indolizine scaffold. Furthermore, indolizine-based fluorophores showed more evident red-shift emission spectra (580 nm) through strengthening the ICT process by the introduction of more electron-withdrawing groups than the ester group, finally successfully leading to the potential fluorescent pH probe. Unfortunately, these newly developed indolizine fluorophores show broad emission spectra, which can cause fluorescent cross-talk problems [40,41,42]. In addition, they have absorption wavelengths in the UV and blue light range (~400 nm). UV and blue light range possess higher energy than visible light (> 400 nm), can be more damaging to the live cell, and usually cause high scattering more than other color light, which occasionally leads to a lower bioavailability [43]. Despite these drawbacks, many other advantages, such as easy access for absorption and emission tunability and predictability guided by computational studies, have rendered indolizine-based fluorophores versatile tools to generate fluorescent and fluorogenic probes. The past decade has witnessed remarkable advances in the development of new indolizine-based fluorophores. Thus, we are sure that our indolizine-based fluorophores could act as the new molecular template for the development of fluorophores with better photophysical properties and fluorescent probes, which leads to providing convenience for live-cell imaging. The following study will be reported in due course.

Supplementary Materials

The following are available online, Figure S1. Absorption and Emission Spectra of Compounds, Figure S2. 1H and 13C NMR Spectra.

Author Contributions

T.K. and J.K. conceived and designed the project. T.K. performed all experiments. T.K. and J.K. analyzed the data. T.K. and J.K. wrote the manuscript. All authors have read and agreed to the published version of the manuscript.

Funding

This work was supported by the Soongsil University Research Fund (New Professor Support Research) of 2019.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The data presented in this study are available in supplementary materials.

Conflicts of Interest

The authors declare no conflicts of interest.

Sample Availability

Not available.

References

  1. Fernández-Suárez, M.; Ting, A.Y. Fluorescent probes for super-resolution imaging in living cells. Nat. Rev. Mol. Cell Biol. 2008, 9, 929–943. [Google Scholar] [CrossRef] [PubMed]
  2. Dean, K.M.; Palmer, A.E. Advances in fluorescence labeling strategies for dynamic cellular imaging. Nat. Chem. Biol. 2014, 10, 512–523. [Google Scholar] [CrossRef]
  3. Lavis, L.D.; Raines, R.T. Bright ideas for chemical biology. ACS Chem. Biol. 2008, 3, 142–155. [Google Scholar] [CrossRef] [Green Version]
  4. Lukinavicius, G.; Umezawa, K.; Olivier, N.; Honigmann, A.; Yang, G.Y.; Plass, T.; Mueller, V.; Reymond, L.; Correa, I.R.; Luo, Z.G.; et al. A near-infrared fluorophore for live-cell super-resolution microscopy of cellular proteins. Nat. Chem. 2013, 5, 132–139. [Google Scholar] [CrossRef] [PubMed]
  5. Uno, S.N.; Kamiya, M.; Yoshihara, T.; Sugawara, K.; Okabe, K.; Tarhan, M.C.; Fujita, H.; Funatsu, T.; Okada, Y.; Tobita, S.; et al. A spontaneously blinking fluorophore based on intramolecular spirocyclization for live-cell super-resolution imaging. Nat. Chem. 2014, 6, 681–689. [Google Scholar] [CrossRef] [PubMed]
  6. Kobayashi, H.; Ogawa, M.; Alford, R.; Choyke, P.L.; Urano, Y. New strategies for fluorescent probe design in medical diagnostic imaging. Chem. Rev. 2010, 110, 2620–2640. [Google Scholar] [CrossRef] [Green Version]
  7. Yun, S.W.; Kang, N.Y.; Park, S.J.; Ha, H.H.; Kim, Y.K.; Lee, J.S.; Chang, Y.T. Diversity oriented fluorescence library approach (DOFLA) for live cell imaging probe development. Acc. Chem. Res. 2014, 47, 1277–1286. [Google Scholar] [CrossRef]
  8. Kwon, H.Y.; Liu, X.; Choi, E.G.; Lee, J.Y.; Choi, S.Y.; Kim, J.Y.; Wang, L.; Park, S.J.; Kim, B.; Lee, Y.A.; et al. Development of a universal fluorescent probe for gram-positive bacteria. Angew. Chem. Int. Ed. 2019, 58, 8426–8431. [Google Scholar] [CrossRef]
  9. Vendrell, M.; Zhai, D.; Er, J.C.; Chang, Y.T. Combinatorial strategies in fluorescent probe development. Chem. Rev. 2012, 112, 4391–4420. [Google Scholar] [CrossRef]
  10. Schiedel, M.S.; Briehn, C.A.; Bauerle, P. Single-compound libraries of organic materials: Parallel synthesis and screening of fluorescent dyes. Angew. Chem. Int. Ed. 2001, 40, 4677–4680. [Google Scholar] [CrossRef]
  11. Loudet, A.; Burgess, K. BODIPY dyes and their derivatives: Syntheses and spectroscopic properties. Chem. Rev. 2007, 107, 4891–4932. [Google Scholar] [CrossRef]
  12. Boens, N.; Verbelen, B.; Dehaen, W. Postfunctionalization of the BODIPY core: Synthesis and spectroscopy. Eur. J. Org. Chem. 2015, 2015, 6577–6595. [Google Scholar] [CrossRef] [Green Version]
  13. Choi, S.K.; Kim, J.; Kim, E. Overview of syntheses and molecular-design strategies for tetrazine-based fluorogenic probes. Molecules 2021, 26, 1868–1883. [Google Scholar] [CrossRef] [PubMed]
  14. Cheema, H.; Baumann, A.; Loya, E.K.; Brogdon, P.; McNamara, L.E.; Carpenter, C.A.; Hammer, N.I.; Mathew, S.; Risko, C.; Delcamp, J.H. Near-infrared-absorbing indolizine-porphyrin push-pull dye for dye-sensitized solar cells. ACS Appl. Mater. Interfaces 2019, 11, 16474–16489. [Google Scholar] [CrossRef]
  15. Gayton, J.; Autry, S.A.; Meador, W.; Parkin, S.R.; Hill, G.A.; Hammer, N.I.; Delcamp, J.H. Indolizine-cyanine dyes: Near infrared emissive cyanine dyes with increased stokes shifts. J. Org. Chem. 2019, 84, 687–697. [Google Scholar] [CrossRef]
  16. Ji, R.X.; Liu, A.K.; Shen, S.L.; Cao, X.Q.; Li, F.; Ge, Y.Q. An indolizine-rhodamine based FRET fluorescence sensor for highly sensitive and selective detection of Hg2+ in living cells. RSC Adv. 2017, 7, 40829–40833. [Google Scholar] [CrossRef] [Green Version]
  17. Park, J.; Lim, B.; Lee, N.K.; Lee, J.H.; Jang, K.; Kang, S.W.; Kim, S.; Kim, I.; Hwang, H.; Lee, J. Dual-functioning IQ-LVs as lysosomal viscosity probes with red-shifted emission and inhibitors of autophagic flux. Sens. Actuators B Chem. 2020, 309, 127764. [Google Scholar] [CrossRef]
  18. Park, S.; Kwon, D.I.; Lee, J.; Kim, I. When indolizine meets quinoline: Diversity-oriented synthesis of new polyheterocycles and their optical properties. ACS Comb. Sci. 2015, 17, 459–469. [Google Scholar] [CrossRef]
  19. Albaladejo, M.J.; Gonzalez-Soria, M.J.; Alonso, F. Metal-free remote-site C-H alkenylation: Regio and diastereoselective synthesis of solvatochromic dyes. Green Chem. 2018, 20, 701–712. [Google Scholar] [CrossRef] [Green Version]
  20. Sokolova, E.A.; Festa, A.A.; Subramani, K.; Rybakov, V.B.; Varlamov, A.V.; Voskressensky, L.G.; Van der Eycken, E.V. Microwave-assisted synthesis of fluorescent pyrido[2,3-b]indolizines from alkylpyridinium salts and enaminones. Molecules 2020, 25, 4059–4070. [Google Scholar] [CrossRef]
  21. Kim, E.; Koh, M.; Lim, B.J.; Park, S.B. Emission wavelength prediction of a full-color-tunable fluorescent core skeleton, 9-aryl-1,2-dihydropyrrolo[3,4-b]indolizin-3-one. J. Am. Chem. Soc. 2011, 133, 6642–6649. [Google Scholar] [CrossRef] [PubMed]
  22. Choi, E.J.; Kim, E.; Lee, Y.; Jo, A.; Park, S.B. Rational perturbation of the fluorescence quantum yield in emission-tunable and predictable fluorophores (Seoul-Fluors) by a facile synthetic method involving C-H activation. Angew. Chem. Int. Ed. 2014, 53, 1346–1350. [Google Scholar] [CrossRef] [Green Version]
  23. Kim, E.; Lee, Y.; Lee, S.; Park, S.B. Discovery, understanding, and bioapplication of organic fluorophore: A case study with an indolizine-based novel fluorophore, Seoul-Fluor. Acc. Chem. Res. 2015, 48, 538–547. [Google Scholar] [CrossRef] [PubMed]
  24. Lee, Y.; Jo, A.; Park, S.B. Rational improvement of molar absorptivity guided by oscillator strength: A case study with furoindolizine-based core skeleton. Angew. Chem. Int. Ed. 2015, 54, 15689–15693. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  25. Choi, S.K.; Rho, J.; Yoon, S.E.; Seok, J.H.; Kim, H.; Min, J.; Yoon, W.; Lee, S.; Yun, H.; Kwon, O.P.; et al. Full color tunable aggregation-induced emission luminogen for bioimaging based on an indolizine molecular framework. Bioconjug. Chem. 2020, 31, 2522–2532. [Google Scholar] [CrossRef] [PubMed]
  26. Choi, S.K.; Lee, Y.; Yoon, S.E.; Choi, H.; Kim, J.; Kim, J.H.; Lee, S.; Kim, W.; Kim, E. A tetrazine-fused aggregation induced emission luminogen for bioorthogonal fluorogenic bioprobe. Sens. Actuators B Chem. 2021, 340, 129966. [Google Scholar] [CrossRef]
  27. Kucukdisli, M.; Opatz, T. A modular synthesis of polysubstituted indolizines. Eur. J. Org. Chem. 2012, 2012, 4555–4564. [Google Scholar] [CrossRef]
  28. Bonte, S.; Ghinea, I.O.; Dinica, R.; Baussanne, I.; Demeunynck, M. Investigation of the pyridinium ylide-alkyne cycloaddition as a fluorogenic coupling reaction. Molecules 2016, 21, 332–346. [Google Scholar] [CrossRef] [Green Version]
  29. Douglas, T.; Pordea, A.; Dowden, J. Iron-catalyzed indolizine synthesis from pyridines, diazo compounds, and alkynes. Org. Lett. 2017, 19, 6396–6399. [Google Scholar] [CrossRef]
  30. Sadowski, B.; Klajn, J.; Gryko, D.T. Recent advances in the synthesis of indolizines and their π-expanded analogues. Org. Biomol. Chem. 2016, 14, 7804–7828. [Google Scholar] [CrossRef] [Green Version]
  31. Liu, B.; Wang, Z.; Wu, N.J.; Li, M.L.; You, J.S.; Lan, J.B. Discovery of a full-color-tunable fluorescent core framework through direct C-H (Hetero)arylation of N-Heterocycles. Chemistry 2012, 18, 1599–1603. [Google Scholar] [CrossRef]
  32. de Silva, A.P.; Gunaratne, H.Q.; Gunnlaugsson, T.; Huxley, A.J.; McCoy, C.P.; Rademacher, J.T.; Rice, T.E. Signaling recognition events with fluorescent sensors and switches. Chem. Rev. 1997, 97, 1515–1566. [Google Scholar] [CrossRef]
  33. Badone, D.; Baroni, M.; Cardamone, R.; Ielmini, A.; Guzzi, U. Highly efficient palladium-catalyzed boronic acid coupling reactions in water: Scope and limitations. J. Org. Chem. 1997, 62, 7170–7173. [Google Scholar] [CrossRef] [PubMed]
  34. Grabowski, Z.R.; Rotkiewicz, K.; Rettig, W. Structural changes accompanying intramolecular electron transfer: Focus on twisted intramolecular charge-transfer states and structures. Chem. Rev. 2003, 103, 3899–4032. [Google Scholar] [CrossRef]
  35. Reichardt, C. Solvatochromic dyes as solvent polarity indicators. Chem. Rev. 1994, 94, 2319–2358. [Google Scholar] [CrossRef]
  36. He, L.; Dong, B.; Liu, Y.; Lin, W. Fluorescent chemosensors manipulated by dual/triple interplaying sensing mechanisms. Chem. Soc. Rev. 2016, 45, 6449–6461. [Google Scholar] [CrossRef]
  37. Tang, Y.; Kong, X.; Xu, A.; Dong, B.; Lin, W. Development of a two-photon fluorescent probe for imaging of endogenous formaldehyde in living tissues. Angew. Chem. Int. Ed. 2016, 55, 3356–3359. [Google Scholar] [CrossRef] [PubMed]
  38. Hossain, S.M.; Singh, K.; Lakma, A.; Pradhan, R.N.; Singh, A.K. A schiff base ligand of coumarin derivative as an ICT-Based fluorescence chemosensor for Al3+. Sens. Actuators B Chem. 2017, 239, 1109–1117. [Google Scholar] [CrossRef]
  39. Jiang, C.; Huang, H.; Kang, X.; Yang, L.; Xi, Z.; Sun, H.; Pluth, M.D.; Yi, L. NBD-based synthetic probes for sensing small molecules and proteins: Design, sensing mechanisms and biological applications. Chem. Soc. Rev. 2021, 50, 7436–7495. [Google Scholar] [CrossRef]
  40. Wu, X.; Sun, X.; Guo, Z.; Tang, J.; Shen, Y.; James, T.D.; Tian, H.; Zhu, W. In vivo and in situ tracking cancer chemotherapy by highly photostable NIR fluorescent theranostic prodrug. J. Am. Chem. Soc. 2014, 136, 3579–3588. [Google Scholar] [CrossRef]
  41. Araneda, J.F.; Piers, W.E.; Heyne, B.; Parvez, M.; McDonald, R. High Stokes shift anilido-pyridine boron difluoride dyes. Angew. Chem. Int. Ed. 2011, 50, 12214–12217. [Google Scholar] [CrossRef] [PubMed]
  42. Shcherbakova, D.M.; Hink, M.A.; Joosen, L.; Gadella, T.W.; Verkhusha, V.V. An orange fluorescent protein with a large Stokes shift for single-excitation multicolor FCCS and FRET imaging. J. Am. Chem. Soc. 2012, 134, 7913–7923. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  43. Oliveira, E.; Bertolo, E.; Nunez, C.; Pilla, V.; Santos, H.M.; Fernandez-Lodeiro, J.; Fernandez-Lodeiro, A.; Djafari, J.; Capelo, J.L.; Lodeiro, C. Green and Red Fluorescent Dyes for Translational Applications in Imaging and Sensing Analytes: A Dual-Color Flag. ChemistryOpen 2018, 7, 9–52. [Google Scholar] [CrossRef] [PubMed]
Molecules 27 00012 g001 550
Figure 1. (a) Indolizine-based fluorescent core skeletons. (b) Retrosynthetic analysis for the 3,7-disubstituted new indolizine-based fluorescent skeletons under investigation.
Figure 1. (a) Indolizine-based fluorescent core skeletons. (b) Retrosynthetic analysis for the 3,7-disubstituted new indolizine-based fluorescent skeletons under investigation.
Molecules 27 00012 g001
Molecules 27 00012 sch001 550
Scheme 1. (a) Synthetic scheme of 3,7-disubstitued indolizine scaffolds and (b) list of synthesized indolizine-based new fluorescent compounds 6–13.
Scheme 1. (a) Synthetic scheme of 3,7-disubstitued indolizine scaffolds and (b) list of synthesized indolizine-based new fluorescent compounds 6–13.
Molecules 27 00012 sch001
Molecules 27 00012 g002 550
Figure 2. Solvatochromism of compound 9. (a) Absorbance and (b) emission spectra of compound 9 in solvents with different polarity (diethyl ether, DCM, ACN, or MeOH). (c) Fluorescence photo image of compound 9 in different solvents taken under 365 nm UV light irradiation.
Figure 2. Solvatochromism of compound 9. (a) Absorbance and (b) emission spectra of compound 9 in solvents with different polarity (diethyl ether, DCM, ACN, or MeOH). (c) Fluorescence photo image of compound 9 in different solvents taken under 365 nm UV light irradiation.
Molecules 27 00012 g002
Molecules 27 00012 g003 550
Figure 3. Solvatochromism of compound 17. (a) Absorbance and (b) emission spectra of compound 17 in solvents with different polarity (diethyl ether, DCM, ACN, or MeOH). (c) Fluorescence photo image of compound 17 in different solvents taken under 365 nm UV light irradiation.
Figure 3. Solvatochromism of compound 17. (a) Absorbance and (b) emission spectra of compound 17 in solvents with different polarity (diethyl ether, DCM, ACN, or MeOH). (c) Fluorescence photo image of compound 17 in different solvents taken under 365 nm UV light irradiation.
Molecules 27 00012 g003
Molecules 27 00012 g004 550
Figure 4. (a) Normalized emission spectra of representative compounds 12 (blue), 14 (green), 9 (yellow), and 16 (red-orange). (b) Fluorescence photo image of each compound in MeOH taken under 365 nm UV light irradiation.
Figure 4. (a) Normalized emission spectra of representative compounds 12 (blue), 14 (green), 9 (yellow), and 16 (red-orange). (b) Fluorescence photo image of each compound in MeOH taken under 365 nm UV light irradiation.
Molecules 27 00012 g004
Molecules 27 00012 sch002 550
Scheme 2. Synthetic scheme for the modification of ester functional group of compound 9 at the C-7 position.
Scheme 2. Synthetic scheme for the modification of ester functional group of compound 9 at the C-7 position.
Molecules 27 00012 sch002
Molecules 27 00012 g005 550
Figure 5. pH response of compound 14. (a) Photophysical properties and visible color change of compound 14 in MeOH, pH 7.0, 5.0, and 3.0 buffer solution. The fluorescence photo images of compound 14 were taken under 365 nm UV light irradiation. (b) Emission spectra of compound 14 at different pH buffer solution measured with excitation wavelength at 388 nm. (c) Scatter plots of the normalized fluorescence emission intensity of compound 14 over the pH range 3–7 with an excitation/emission wavelength at 388/473 nm.
Figure 5. pH response of compound 14. (a) Photophysical properties and visible color change of compound 14 in MeOH, pH 7.0, 5.0, and 3.0 buffer solution. The fluorescence photo images of compound 14 were taken under 365 nm UV light irradiation. (b) Emission spectra of compound 14 at different pH buffer solution measured with excitation wavelength at 388 nm. (c) Scatter plots of the normalized fluorescence emission intensity of compound 14 over the pH range 3–7 with an excitation/emission wavelength at 388/473 nm.
Molecules 27 00012 g005
Molecules 27 00012 g006 550
Figure 6. pH response of compound 9 (a), compound 12 (b), compound 16 (c) and compound 17 (d). Emission spectra of compounds at different pH buffer solution (3.0, 5.0, and 7.0).
Figure 6. pH response of compound 9 (a), compound 12 (b), compound 16 (c) and compound 17 (d). Emission spectra of compounds at different pH buffer solution (3.0, 5.0, and 7.0).
Molecules 27 00012 g006
Table
Table 1. Photophysical properties of 3,7-disubstituted indolizine compounds.
Table 1. Photophysical properties of 3,7-disubstituted indolizine compounds.
Molecules 27 00012 i001
CpdR1R2λabs[nm] [a]ε[M−1cm−1] [b]Λem[nm] [c]△v[cm−1] [d]ΦF[e]
64-HCO2Et38911,00046843990.24
74-CH3CO2Et390955047445440.26
84-OMeCO2Et39011,10048349370.32
94-N(Me)2CO2Et40410,80053359910.46
104-FCO2Et38610,50046744930.23
114-CHOCO2Et42018,30049237510.02
124-CNCO2Et39417,70046639210.15
133-CNCO2Et38212,20046245330.19
144-N(Me)2CO2H390870050558390.33
164-N(Me)2CHO42315,35058063990.04
174-N(Me)2COCH342011,50057463880.15
All the experimental data were obtained using methanol. [a] Only the largest absorption maxima are shown. [b] Molar absorption coefficient at maximum absorption wavelength. [c] Excited at the maximum absorption wavelength. [d] Stokes Shift. [e] Absolute fluorescence quantum yield.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Share and Cite

MDPI and ACS Style

Kim, T.; Kim, J. Color-Tunable Indolizine-Based Fluorophores and Fluorescent pH Sensor. Molecules 2022, 27, 12. https://doi.org/10.3390/molecules27010012

AMA Style

Kim T, Kim J. Color-Tunable Indolizine-Based Fluorophores and Fluorescent pH Sensor. Molecules. 2022; 27(1):12. https://doi.org/10.3390/molecules27010012

Chicago/Turabian Style

Kim, Taegwan, and Jonghoon Kim. 2022. "Color-Tunable Indolizine-Based Fluorophores and Fluorescent pH Sensor" Molecules 27, no. 1: 12. https://doi.org/10.3390/molecules27010012

Article Metrics

Back to TopTop