Pyrazolo[1,5-a]pyrimidines-based fluorophores: a comprehensive theoretical-experimental study

Fluorescent molecules are crucial tools for studying the dynamics of intracellular processes, chemosensors, and the progress of organic materials. In this study, a family of pyrazolo[1,5-a]pyrimidines (PPs) 4a–g has been identified as strategic compounds for optical applications due to several key characteristics such as their simpler and greener synthetic methodology (RME: 40–53%) as compared to those of BODIPYS (RME: 1.31–17.9%), and their tunable photophysical properties (going from ε = 3320 M−1 cm−1 and ϕF = 0.01 to ε = 20 593 M−1 cm−1 and ϕF = 0.97), in which electron-donating groups (EDGs) at position 7 on the fused ring improve both the absorption and emission behaviors. The PPs bearing simple aryl groups such as 4a (4-Py), 4b (2,4-Cl2Ph), 4d (Ph) and 4e (4-MeOPh), allow good solid-state emission intensities (QYSS = 0.18 to 0.63) in these compounds and thus, solid-state emitters can be designed by proper structural selection. The properties and stability found in 4a–g are comparable to commercial probes such as coumarin-153, prodan and rhodamine 6G. Ultimately, the electronic structure analysis based on DFT and TD-DFT calculations revealed that EDGs at position 7 on the fused ring favor large absorption/emission intensities as a result of the ICT to/from this ring; however, these intensities remain low with electron-withdrawing groups (EWGs), which is in line with the experimental data and allows us to understand the optical properties of this fluorophore family.


Introduction
Fluorescent organic compounds have been a major focus of research related to materials science and biological interactions over the past decades. 1 Ranging from ionic or molecular sensing 2 to bioimaging applications 3 and, from the organic light-emitting devices, 4 bio-macromolecular interactions. 5 A plethora of applications has been proposed by exploiting the benecial properties of such materials that can raise our standard of living. In this eld, the uorogenic heterocyclic compounds display advantages over hydrocarbon-based uorophores such as (i) synthetic access methodologies that allow structural diversity, (ii) heteroatoms (B, N, O or S) that make them potential chelating agents for ions, and (iii) better solubility in green solvents. Consequently, various uorescent derivatives have been applied in diverse elds, and include coumarins, 6 pyrazoles, 7 perylene bisimides, 8 boron dipyrromethene diuoride (BODIPY), 9 cyanines, 10 and rhodamines. 11 Importantly, for biological and/or optoelectronics applications, ideal uorescent probes must meet several requirements: the synthetic pathway should be straight (few steps), useful (high yields and low waste generation), and carried out by efficient heating technologies; i.e., microwave (MW) or ultrasound (US). Because a probe for bioimaging is inherently involved in complex biological interactions within the cell, the probe must be as small as possible to reduce its impacts on the biological dynamics and facilitate its diffusion inside the cell organelles. 3 Likewise, for most of the photophysical applications, the uorophore must be able to change its absorption and emission properties with small and simple chemical modications. Ful-lling the above-mentioned conditions in a single uorophore is a challenging task and many efforts need to be made in this direction. Recently, pyrazolo[1,5-a]pyrimidines (PPs) have emerged as an attractive alternative due to their small size based on [5,6]-fused N-heterocyclic systems, their efficient synthetic approaches 12 and easy functionalization methodologies, 13,14 together with their uorescence properties (high quantum yields in different solvents, and excellent photostability). 15 Usually, the PPs receive major attention in biological applications, with the cancer therapeutics eld being the most attractive area. 16 Very recently, Professor Jian-Feng Ge's group found that the combination of photophysical properties with biological activities allows the use of these compounds as lipid droplet biomarkers for HeLa cells (cancer cells) and L929 cells (normal cells), 15 demonstrating the interesting versatility of this core.
Among the synthetic procedures available for the preparation of diverse pyrazolo [1,5-a]pyrimidine (PP) derivatives, 7,12-21 the strategy involving the cyclocondensation of NH-3-aminopyrazoles with b-dicarbonyl compounds or other 1,3-bis-electrophiles (e.g., alkoxymethylene-b-dicarbonyl compounds, a,b-unsaturated systems, b-enaminones, b-ketonitriles, b-enaminonitriles, among others) has been the most frequently studied due to its excellent performance. This synthetic approach allows key structural modications at all the peripheral positions during ringconstruction and through subsequent functionalization steps. 7,[12][13][14][15][16][17][18][19][20][21] It is important to note that the theoretical calculations are an important tool for examining the electronic and reactivity properties of some interesting uorophores. [22][23][24][25] For example, the excited-state intramolecular proton transfer (ESIPT) process, 22 absorption and emission transitions, 23 aggregationcaused quenching mechanism, 24 and other crucial chemical properties of the uorescent molecules 25 have been investigated. From this perspective and in line with the aforementioned properties of the pyrazolo[1,5-a]pyrimidines, we proposed the synthesis of a family of these fused N-heterocycles (compounds 4a-g) substituted at position 7 with different electron-withdrawing (EWGs) and electron-donating groups (EDGs). The uorophores 4a-g were obtained by the interaction of the appropriate b-enaminone 2a-g with 3-methyl-1H-pyrazol-5-amine (3); their green chemistry efficiency and the cost per gram of raw materials in each case were evaluated (Scheme 1). Likewise, the photophysical properties in both solution and solid-state of 4a-g were investigated, and theoretical calculations at the DFT and TD-DFT levels were used to interpret the absorption and emission observations.

Green chemistry performance
Since uorophores 4a-g can be recognized as ne chemicals, for practical applications these compounds are required to fulll some environmental issues related to their production. Therefore, a univariate green metrics analysis based on a common criterion such as reaction mass efficiency (RME) was performed 27 (Tables 1 and S1-S7 †). The RME values found for this family of 7-substituted 2-methylpyrazolo[1,5-a]pyrimidines 4a-g were in the range of 40-53%. The high overall yield and the absence of catalysts explained the good RME performance for the synthesis of 4a-g. Remarkably, the sub-products generated during this synthesis were methanol, dimethylamine and water, useful solvents that can be recovered for use in other chemical processes. To compare the good merits found in these probes, the same calculations were performed for three BODIPY-based uorophores, a widely used compound in materials science 28 Scheme 1 Synthesis of pyrazolo[1,5-a]pyrimidine compounds 4a-g. (Tables 1 and S8-S10 †). The overall yield and the RME were better for PPs 4a-g than for BODIPY derivatives.
With the use of the MW-assisted technology in almost all procedures, a tool known for its benets such as easy handling, rapid and solvent-free synthesis, the generation of fewer subproducts, 29 and maximizing the use of raw materials indicate that the synthetic approach for these uorophores is in agreement with the main principle of the green chemistry, prevention. 30 The cost per gram of the raw materials of PPs 4a-e is just a fraction of that calculated for the uorophores BODIPY-1-BODIPY-3 (Table 1). Meanwhile, the combination of two uorophores in the hybrid systems 4f and 4g noticeably increases the cost of the product as a result of the need to employ expensive raw materials; however, the preliminary cost per gram of these compounds is better than that of BODIPY-1 and BOD-IPY-3. The pyrazolo[1,5-a]pyrimidine core can be easily modi-ed by simple aromatic substitution reactions such as nitration, halogenation and formylation. 7,12,13 As a result, suitable functional groups can be incorporated at positions 2 and 5-7 during the fused-ring construction and at position 3 via functionalization and post-functionalization strategies ( Fig. 1), demonstrating the capability of this uorophore for structural diversity. Along this direction, we recently demonstrated the improvement in the photophysical properties with the incorporation of a conjugated substituent at position 2. 31

UV-vis and emission spectra
The UV-vis and uorescence spectra of the 7-substituted 3methylpyrazolo[1,5-a]pyrimidines 4a-g were measured in different solvents ( Fig. 2 and 3 and Table 2). The absorption and emission spectra of these PPs are highly dependent on the nature of the substituent group at position 7. In general, the absorption spectra of 4a-g showed a main band between 340-440 nm that was previously assigned to an intramolecular charge transfer process. 13,36,37 Notably, with good EDGs such as 4-anisyl 4e, 7-diethylaminocoumarin-3-yl 4f and 4-diphenylaminophenyl 4g, the absorption coefficient (3) in THF was as high as 6547, 39 867 and 15 008 M À1 cm À1 , respectively. Meanwhile, the presence of EWGs in compounds 4a-c, or neutral groups (NG) such as phenyl in the derivate 4d, decreased the values of 3 as follows: 4a (3320 M À1 cm À1 ), 4b (2727 M À1 cm À1 ), 4c (7053 M À1 cm À1 ), and 4d (3827 M À1 cm À1 ) in the same solvent. In particular, coumarin derivatives 4c and 4f displayed the same differences in the absorption spectra as a result of the p-extended conjugation in the coumarin unit. For example, the absorption coefficient of the intramolecular charge transfer (ICT) band at 440 nm for 4f displayed a much higher 3 in THF when compared with that of 4d in the same solvent. A broad absorption band with different shoulders was also observed in compound 4c; this behavior can be attributed to a combination of the ICT phenomenon, as well as the p-p* and n-p* transitions of the coumarin and pyrazolo[1,5-a] pyrimidine moieties. In all cases, the absorption maximum wavelengths (l abs ) were almost unchanged, irrespective of the solvent used, while their molar absorption coefficients tended to show a subtle decrease as the solvent polarity increased.
When PPs 4a-g were excited at their l abs in an airequilibrated solution at 20 C, they exhibited uorescence bands at around 474-541 nm (Fig. 3a). For these probes, there was also an important correlation between the uorescence quantum yield (f F ) and the electronic nature of the substituent at position 7 (Fig. 3b). In a solvent of intermediate polarity (i.e., THF), substituents at position 7, known as EDGs, such as anisyl (4e), 7-diethylaminecoumarin-3-yl (4f) or 4-diphenylaminophenyl (4g), the quantum yield values were 0.23, 0.53 and 0.85, respectively. Notably, with EWGs or NG (7-Ph), the f F values decreased to 0.01 (4a), 0.07 (4b), 0.06 (4c), and 0.06 (4d) in the same solvent. The fact that the f F values in uorophores 4f and 4g are remarkably high in solvents of different polarity is a notable virtue that can be useful, for example, in monitoring macromolecules dynamics 5 or labeling pharmacological targets. 3 Interestingly, these probes displayed good emission intensities even in polar solvents such as ethanol-water 4 : 1 or THF-water mixtures (Fig. S3 †), which is a remarkable property for biological applications.

Solvatouorochromism
Compounds bearing both EDGs and EWGs as substituents, known as push-pull structures, ensure that aer light absorption, the charge is transferred from the donor group to the acceptor, which creates a highly dipolar excited state (DES). The DES interacts with the dipoles of the solvent and thus, the emission shis to longer wavelengths in more polar solvents. 1,39 Therefore, the solvatouorochromic behavior in the emission spectra is a measure of the push-pull properties of a compound. In order to evaluate the solvatochromic features of PPs 4a-g, the relationship between the solvent polarity parameter (Df) and the Stokes shi (Dn), according to the Lippert-Mataga equation, was investigated. 40 From the plots of Dn versus Df (Fig. 4), it was found that the slopes of the tting lines for 4a, 4f and 4g were high, at 10 390, 9381 and 11 015, with acceptable linearity, suggesting that the ICT in these compounds have a larger dipole moment than the ground state due to important charge redistribution. The dipole moment changes (Dm) in those compounds were calculated to be 10.3, 12.8 and 19.0 D. Interestingly, compound 4a with pyridine as an EWG at position 7 displayed a stronger solvatouorochromic effect than that observed in 4e bearing an EDG in the same position. This behavior can be explained by the p-amphoteric donor/acceptor property in the pyrazolo[1,5-a]pyrimidine core, explicitly a pexcedent-p-decient fused system. The 7-pyridyl substituent in 4a acts as an EWG and the fused-ring moiety is expected to be the EDG (i.e., a A-A-D molecular system based on pyridine, pyrimidine and pyrazole rings). The results indicated that the structural arrangement in 4a displays a better push-pull system than that observed for uorophore 4e. Meanwhile, probes 4b-e showed non-solvent polarity dependence in the emission properties because of the absence of strong EDG or EWG substituents, which reduces the charge reorganization in the excited state (Fig. 6).

Solid-state uorescence
Solid-state uorescence properties of compounds 4a-g were rst evaluated upon illumination of each solid sample with UV light of 365 nm using a hand lamp. Fluorophores 4b, 4d, 4a and 4e exhibited a strong blue light emission with quantum yields in solid-state (QY SS ) of 0.63, 0.39, 0.22 and 0.18, respectively, while other probes were less emissive (QY SS : 4g (0.13) > 4f (0.08) > 4c (0.01)) and the uorescence was red-shied into the green region of the spectrum (Table 2). Solid-state uorescence spectra of 4e-g were red-shied with respect to those in solution, indicating the presence of important intermolecular interactions in the solid-state. Likewise, the emission wavelength of probes 4b, 4d, and 4e are very close to those in solution, suggesting that the uorescence properties in the solid-state are highly dependent on different structural arrangements due to the nature of the substituent at position 7 (Fig. 5).
It is worth noting that in the donor-acceptor molecular systems, the microcrystalline arrangement allows an intermolecular uorescence quenching due to the antiparallel organization of the molecules. 31,41,42 Likewise, in N-heterocyclic compounds that lack strong electrostatic interactions due to the absence of polar functional groups, the van der Waals forces are mainly responsible for their formation in the solid-state and apparently, these forces are improved by the presence of simple aryl groups at the ring periphery. The supramolecular assembly can be promoted by the molecular chain formations having aryl groups or fused heterocyclic moieties in a strategic disposition of p/p stacking and C-H/p (or C-H/N) interactions. 31,[43][44][45][46][47] Some haloaryl substituents play a crucial role in molecular packing and the photophysical properties of their derivatives, such as the 2,4-dichlorophenyl (2,4-Cl 2 Ph) group. 44,45 In this context and as expected, probes 4c, 4f and 4g showed lower emission intensities with respect to those found in the rest of the series (intensity in 4b > 4d > 4e > 4a), since they bear bulky groups that disfavor both good packing and uorescence intensity ( Fig. 5 and 6).
Notably, the pyrazolo[1,5-a]pyrimidine core has been widely studied and various crystal arrangements were reported, 12,13,31,46 evincing a molecular packing with parallel 46 or quasi antiparallel 12,13 dispositions with respect to each other, which depend on the nature of the substituents. However, with strong donor groups at position 7 and without steric effects in other fused ring positions, an utterly antiparallel molecular organization is favored (Fig. 6). 31 Therefore, the absence of a push-pull structure in 4b, 4g and 4f, avoids the intermolecular quenching in the solid-state, 31,41,42 allowing high-emission intensities with regard to that observed in derivatives bearing a strong pushpull arrangement (4a-inside of the PP core, 4g and 4f) or bulky substituent groups (4c, 4g and 4f). Finally, the presence of the 2,4-dichlorophenyl substituent in 4b favors its emission intensity as a result of the marked dihedral angle between this group with the heterocyclic core due to the Cl atom at position 2 of this group. 44,45 This effect could reduce the electronic communication between the two rings 45 and result in aggregation-induced emission (AIE) phenomenon 47 (Fig. 5 and 6).

Probes 4a-g stability
The photobleaching characteristics and stability at extreme pH values of uorescent compounds are critical to their application as uorescent materials and bio-probes, respectively. Accordingly, the photostability of dyes 4a-g was investigated and compared with that of prodan (P), coumarin 153 (C-153) and rhodamine 6G (R6G). Aer continuous excitation at 365 nm   with a xenon lamp (4.0 mW) at different times, the normalized uorescence intensities of dyes 4a-g decreased by 89-94%, measured at their maximum wavelength, which is a very good photobleaching performance when compared with those obtained for the commercial probes (Fig. 7a). Likewise, the stability under exposure to extreme pH (pH 2 with H 2 SO 4 and pH 12 with KOH and stirring for 1 h at 50 C; aer neutralization, the emission spectra were recorded) was studied and the behavior was followed by the relative uorescence intensity (Fig. 7b). The results are similar to those found in C-153. Interestingly, acidic conditions have a greater impact on the stability of the pyrazolo[1,5-a]pyrimidine-based probes. These observations can be attributed to the high electronic density in different atoms of the fused-pyrazole and the acid-base interactions could be the beginning of the chemical decomposition.

Computational calculations
To have better insight regarding the molecular structures and electronic properties of uorophores 4a-g, geometry optimization, analytical frequencies, and excited-state energy calculations were performed at the B3LYP lever 48 with the Ahlrichs def2-TZVP basis set, [49][50][51] as implemented in the ORCA 4.2.0 package. 52,53 For further computational details, please see the attached ESI. † All geometry optimization calculations were performed on the framework of the DFT (Fig. 8). This method has been demonstrated to provide reliable results in this kind of system. 54,55 At the B3LYP level, the ground state of these compounds is a closed-shell singlet (S ¼ 0), in which the electronic structure is characterized by frontier molecular orbitals (FMOs) of antibonding p nature (Fig. 9). We performed the geometry optimizations using an implicit solvation model (as described in the ESI †) to evaluate the stability of each probe as a function of the different solvents considered in the experiments (Tables S11-S17 †). For the sake of comparison, Table 3 gives the values obtained in THF for the calculated charges on the nucleophilic carbon and nitrogen atoms (C3 and N4 in Fig. 1  and 8), dihedrals and C7-C8 bond lengths connecting the Nheterocyclic core to 7-aryl groups, the HOMO-LUMO gaps, and polarizabilities. The negative charges in C3 and N4 are comparable, ranging from À0.300 to À0.200, which explains the greater reactivity and regioselectivity of these compounds toward electrophilic species. 12,13 Importantly, N5 may be subject to alkylation reactions to generate pyrimidinium salts, a key group of compounds in organocatalysis 56 and biochemical applications. 57 While the C7-C8 distances remain virtually unchanged with the different aryl substituents (1.470 AE 0.007 A), the dihedrals are disturbed to a larger amplitude (from 34 to 64 ), causing the resonance breaking between the aryl group and the pyrazolo[1,5-a]pyrimidine core to different extents. The HOMO-LUMO gap also changes as there are variations in the substituents. The calculated gaps are situated in the energy range from 3.3 to 4.3 eV, which is in agreement with the wavelengths of the UV-vis spectra outlined above. The polarizability doubled from 25 A 3 as in 4a up to 53 A 3 as in 4g, which is in accordance with a greater dipole moment in the excited states of the latter, and the fact that the EWGs at position 7 lead to higher solvatouorochromisms.
In PPs 4a-g there was a slight decline in the dihedrals, charges, and gaps as the solvent polarity decreased (Tables S11-S17 †). Conversely, the polarizability slightly increased with the enhancement of the solvent polarity. As discussed above for the absorption and emission spectra, this behavior reveals that the polarities of the deemed solvents play a minor role in the electronic structure of the studied compounds, as well as in its geometrical arrangement. More interestingly, PPs 4f and 4g showed the opposite behavior, where the polarizability slightly diminished as the solvent polarity increased. In fact, these   This journal is © The Royal Society of Chemistry 2020 RSC Adv., 2020, 10, 39542-39552 | 39547 Paper RSC Advances systems have larger polarizabilities due to their EDGs (as well as the smaller charges, dihedrals and HOMO-LUMO gaps, Table 3), which is in line with the greater uorescence (vide infra). These results can be understood as follows: the smaller the dihedral, the more effective the p-resonance between the heterocyclic moiety and the 7-aryl group, the polarizability increased, and the HOMO-LUMO gap was reduced. Consequently, the respective probes had improved photophysical and electronic properties because of a more favored ICT process. A deeper analysis of the absorption and emission processes was achieved based on TD-DFT calculations by taking the optimized geometries as a starting point. In this sense, the energies of the ve lowest excited singlet states of uorophores 4a-g in THF were calculated. These energies allowed us to estimate the absorption wavelength of each excited singlet and their respective oscillator strengths via the transition dipole moments. As shown in Table 4 and Fig. S16a, † compounds with EDGs (4e-g) have greater oscillator strengths and higher absorbances. The highest oscillator strengths of compounds 4a-b and d-g, are associated with the rst singlet excited state, whereas for 4c, this strength matches the third singlet excited state. Their transition energies have a mean value of 3.41 AE 0.32 kcal Â mol À1 , in harmony with absorptions in the UV-vis region. The wavelengths linked to the absorption towards the cited excited singlets are well aligned with our experimental results in the same solvent (Table 2). In almost all PPs studied, absorptions are dominated by HOMO / LUMO electronic transitions (Table 4), though in 4c, it is mainly of the HOMO-1 / LUMO type. In PPs 4a-d, the HOMO mainly has the p-nature on the PP ring, while the LUMO is largely of p-character on the aryl group (Fig. 9, top). These observations are consistent with the fact that the absorption process is mainly associated with electron transfer from the PP ring to the adjoining aryl group, that is, an ICT to the EWGs and NG. Moreover, since the HOMO and LUMO in 4e-g are largely of p-nature on the aryl and PP rings, respectively, the absorption process is reversed (Fig. 9, bottom). Thus, the charge transfer goes from the EDGs to the PP ring. Ultimately, the absorption and uorescence spectra of PPs 4a-g were predicted (Fig. S16 †), including the all-vibronic transitions, based on the excited singlet of each probe. As expected, the estimated spectral behavior agrees with the experimental data (see ESI † for more details).
Importantly, compound 4f (7-diethylaminocoumarin-3-yl derivative) displayed lower emission intensity and quantum yield as compared to 4g in optical experiments, which was not reected in the computational estimations. Likewise, the simple coumarin-PP derivative 4c offered both experimental and calculated photophysical results in lower limits ( Fig. 4-6, 8 and S16 †). These ndings are possibly due to the high sensitivity of the coumarin derivatives to the microenvironments (e.g., solvent properties and solid-state aggregation), steric effects (e.g., groups near the D-A junction and irregular packing) and because 4c does not possess strong EDGs on the coumarin ring, such as the diethylamino (Et 2 N) group at 4f. 58 In fact, this group offers resonant structures for the two geometries of the 7-Et 2 N-coumarin derivatives in the excited state, from (a) a planar emissive ICT excited state, to (b) a nonuorescent twisted ICT state (TICT), shown in Fig. 10. 58,59 61 and the good photo-and acid-base exposure stability make them attractive alternatives for real applications.
Computational calculations were performed in order to describe the changes in the electronic structure associated with the absorption and emission processes in 4a-g. Geometry optimization calculations on the singlet ground state indicated that the dihedral centered on C7-C8 controls the polarizability and HOMO-LUMO gap of compounds and hence, its respective absorption and uorescence rates. The smaller the dihedral, the shorter the HOMO-LUMO gap and thus, the ICT phenomena in absorption and emission spectroscopy for the probes are favored. Moreover, the electronic transition analysis of the lowest excited singlet states in probes, revealed that the absorbance experiments at low energy, are governed by a HOMO / LUMO electron transition, while this transition is reversed for emission experiments. The electronic structure analysis based on the FMO nature indicates that EDGs favor large absorption and emission intensities as a result of the ICT process to and from the fused heterocyclic moiety, respectively. When EWGs are used, these intensities remain low, which is in line with the experimental results. Ultimately, both experimental and theoretical results of the coumarin derivatives 4c and 4f provided evidence that the electronic properties of these compounds are governed by different complex phenomena.

General procedures
General procedure for the synthesis of b-enaminones 2a-g. A 10.0 mL sealable (Teon screw cap) oven-dried tubular reaction vessel was charged with 1.0 mmol of the appropriate methyl ketone (1a, 1b, 1d, 1e, 1f, or 1g) and 1.5 mmol of DMF-DMA. The resulting mixture was irradiated with MW at 160 C (180 W) and maintained at this temperature for 15 min in a sealed tube containing a Teon-coated magnetic stir bar. The resulting reaction mixture was cooled to 55 C by airow and the excess of DMF-DMA was removed under reduced pressure, yielding the respective crude b-enaminones (2a, 2b and 2d-g) via this protocol previously reported in our lab. 12 Importantly, b-enaminone 2c was obtained under reux for 6 h from an equimolar mixture (1 mmol) of 3-acetyl-2H-chromen-2-one (1c) and DMF-DMA, according the procedure reported by El-Taweel and Elnagdi; 62 however, in this case, we used 1,4-dioxane (5.0 mL) as a solvent instead of the xylene used by those authors. Later, the 1,4-dioxane was removed under reduced pressure, yielding the crude product 2c. Finally, all the crude b-enaminones were puried by ash chromatography on silica gel (eluent: CH 2 Cl 2 ) to afford the pure products 2d-g.
General procedure for the synthesis of pyrazolo[1,5-a] pyrimidines 4a-g. A 10.0 mL sealable (Teon screw cap) ovendried tubular reaction vessel was charged with an equimolar mixture (0.5 mmol) of the respective b-enaminone (2a, 2b, 2d, 2e, or 2g) and 3-methyl-1H-pyrazol-5-amine (3, 49 mg). The resulting mixture was irradiated with MW at 180 C (200 W) and maintained at this temperature for 2 min in a sealed tube containing a Teon-coated magnetic stir bar. The resulting reaction mixture was cooled to 55 C by airow and the precipitated product formed upon the addition of cold EtOH/ H 2 O (1 : 1, 1.0 mL) was ltered off, washed and dried to give the corresponding pure product (4a, 4b, 4d, 4e, and 4g) by this protocol previously reported in our lab. 12 Meanwhile, uorophores 4c and 4f were obtained under reux in acetic acid (1.0 mL) for 3 h, starting from b-enaminone 2c and 2f, respectively. Subsequently, the resulting reaction mixture was concentrated under reduced pressure and the residue was recrystallized from ethanol.

AIE
Aggregation

Conflicts of interest
The authors declare no competing nancial interest.