1,3,4-Thiadiazol-2-ylphenyl-1,2,4,5-tetrazines: efficient synthesis via Pinner reaction and their luminescent properties

Due to the interest in, and diverse applications of, 1,2,4,5-tetrazines and 1,3,4-thiadiazoles, individually and, more recently, in combination, a series of novel 1,2,4,5-tetrazine derivatives conjugated directly or via a 1,4-phenylene linker with a 1,3,4-thiadiazole core, were synthesized. A six-step reaction sequence, involving the Pinner reaction and oxidation under mild conditions, was used. This approach worked well for both symmetrical and unsymmetrical arrangements. Their luminescence properties were examined and are reported. The obtained compounds may have a number of great applications potential.


Introduction
One of the most interesting heterocyclic arrangements, characterised by both the maximum content of nitrogen atoms and its ring stability, is the six-membered 1,2,4,5-tetrazine (s-tetrazine). 1 Among its most commonly described applications, one has to distinguish high-energy-density materials (HEDM), which allow their utilisation in the production of explosives and propellants. 2However, the derivatives of this compound also have great potential for applications in other fields.4][5] Optoelectronics is an intensively developing and researched field, however, new organic n-type semiconductors, which would allow improvements in the efficiency and durability of devices, are still being researched.Accordingly, nitrogen-containing compounds are of great interest.Due to the high electronegativity of nitrogen, heterocycles rich in this atom are characterised by a large electron deficit.The use of such systems allows the modification of the charge-transport properties of a given material to obtain the appropriate levels of bandgap energies.In addition to s-tetrazine derivatives, examples of systems that work perfectly in this role are compounds containing 1,3,4-thiadiazole. 6,7Its derivatives can also be used as high-performance wide-bandgap copolymer donors for efficient non-fullerene organic solar cells. 8oth s-tetrazine and 1,3,4-thiadiazole also exhibit a broad spectrum of biological activity.Consequently, their derivatives can be used in medicine [9][10][11][12][13][14][15][16][17][18][19] and 1,3,4-thiadiazole-containing systems serve as plantprotection products. 20,21In addition, an extremely interesting potential application of s-tetrazine derivatives is in medical diagnostics.][24] The combination of s-tetrazine and 1,3,4-thiadiazole seems to be very promising as well, especially for applications in optoelectronics.However, very few types of derivatives have been described in the literature so far.In our previous work, we synthesised a series of s-tetrazines directly conjugated to 1,3,4-thiadiazoles which exhibited excellent fluorescent properties. 25We were interested in how the introduction of the 1,4phenylene linker between the heterocyclic rings would affect the luminescent properties of the new compounds.
The work reported here describes a convenient and practical methodology for the synthesis of 3,6-bis(4-(1,3,4-thiadiazol-2-yl)phenyl)-1,2,4,5-tetrazine derivatives from aromatic carbonitriles, hydrazine hydrate and Lawesson's reagent.We present another series of compounds containing these two rings, this time connected via a 1,4-phenylene linker.We were also interested in the results due to the possibility of comparing the properties of the obtained compounds with analogous systems containing 1,3,4-oxadiazole. 26To the best of our knowledge, the preparation and fluorescent properties of the title compounds, containing an extended πconjugated system, have not been previously reported.

Results and Discussion
The title 1,2,4,5-tetrazine derivatives conjugated via a 1,4-phenylene linker to a 1,3,4-thiadiazole ring were obtained in a six-step reaction sequence.The first four steps were aimed at the formation of a five-membered ring without compromising the stability of the carbonitrile moiety (Scheme 1).For this purpose, commercially available 4-cyanobenzoic acid (1) was used.The specific structure of this compound allows its application as a linker between 1,2,4,5-tetrazine and 1,3,4-thiadiazole.In the first step, the carboxyl group present in the substrate was subjected to esterification using methanol and hydrochloric acid, which gave ester 2. The next step was the conversion of compound 2 by reaction with hydrazine hydrate in methanol which resulted in product 3.In order to form the necessary diacylhydrazine moiety, as well as to introduce an additional ring with substituents of various types, hydrazide 3 was reacted with freshly prepared aroyl chlorides 4a-d.A series of products, 5a-d, were obtained.The derivatives thus prepared were reacted with Lawesson's reagent in dry toluene.At this stage, the influence on the reaction yield of the type of substituents introduced by the aroyl chlorides was evident.The presence of electron-donor groups (methoxy and tert-butyl) led not only to higher yields, but also to a reduction in the heating time (5b, 5c, entries 2 and 3, Table 1).The next step involved the Pinner reaction of previously prepared 4-(5-phenyl-1,3,4-thiadiazol-2yl)benzonitrile derivatives 6a-d with hydrazine hydrate in the presence of sulphur (Scheme 2).Initially, the research involved the formation of symmetrical compounds 7a-d.In this case, there was also a clear influence of the type of substituents.Similar to the previous reaction, electron-donor groups had a very beneficial effect on the yield of the synthesized products.This impact was also visible later in the study which involved obtaining unsymmetrical derivatives 7e-j.The last step was the oxidation of 1,4-dihydro-1,2,4,5-tetrazine derivatives (7a-j) to the final products.Of the various oxidising agents described in the literature, we decided to use hydrogen peroxide since this environmentally friendly reagent worked well for similar systems containing 1,3,4-oxadiazole. 26After 24 hours of stirring at room temperature, a number of the title products 8a-j were isolated with satisfactory yields (Table 2).Traces of two symmetrical products were also detected.The impact of the type of substituents on the outcome of the reaction, and its detailed mechanism, were discussed in our previous work.

Luminescent properties
UV-Vis absorption and three-dimensional fluorescence spectra were determined for the final products of the performed reactions (Figure S29-S31, Supplementary Material).In the case of compounds 8a, 8c, 8f, and 8g, the fluorescence spectra possess one slightly deformed maximum of fluorescence.The global emission maximum of 8b, 8d, 8e, 8h, 8i, and 8j covers the second, weaker local maximum (visible as a shoulder of the symmetrical global maximum).Due to large differences in intensities of global and local maxima and a substantial overlap, the ex and em of local maxima cannot be determined unambiguously, even with the usage of deconvolution of the three-dimensional function I = f(ex,em).The described compounds emit fluorescent UV (compounds 8a, 8c, 8f, 8g) or Vis (compounds 8b, 8d, 8h, 8i, 8j) radiation upon UV irradiation (Table 3).This suggests that the presence of one terminal H substituent (R 1 or R 2 ) is sufficient for appearance of the UV fluorescence.The quantum yields () correlate with fluorescence intensities (I), an increase of  causes an increase of I (Figure S32, Supplementary Material), whereas they do not correlate with the absorption wavelength (Figure S33, Supplementary Material).This indicates that the electronic transition character is analogous in all the studied compounds, but the amount of the absorbed energy converted into internal energy differs considerably.The main source of excited states is the n→* absorption transitions.In general, the fluorescence intensity agrees with the nature of the electron-withdrawing (EWG) and electrondonating (EDG) groups of substituents.The presence of two strong EWG substituents (8d, R 1 , R 2 = NO2) decreases fluorescence due to a decrease in electron density in delocalised systems.Alternatively, the existence of EDG substituents mostly increases fluorescence.The differences between em and ex at global maxima fall in two ranges: 121-126 nm for compounds with the OCH3 substituent (8b, 8e, 8h, 8i), and 72-91 nm for compounds without the OCH3 substituent (8a, 8c, 8d, 8f, 8g, 8j).This proves that the studied compounds have diverse energy gaps between individual orbitals participating in fluorescence transitions.It should be noted that compounds with the OCH3 substituent (8b, 8e, 8h, 8i) possess uncommon fluorescence characteristics as em-ex is larger than the standard border value of 100 nm.Within each of the abovementioned groups of compounds (with or without the OCH3 substituent), ex correlates with em, i.e., a larger ex produces emission at a larger em (Figure S34, Supplementary Material).Noteworthy is the fact that irradiation of compounds 8h by ultraviolet radiation leads to the emission of violet fluorescence light visible by the naked eye.
Table 3. Luminescence properties of symmetrical (8a-d) and unsymmetrical (8e-j) derivatives of s-tetrazine.Quantum yields Φ were calculated according to Equation 1 presented in reference 27 , with the use of emission intensity registered for two standard substances: quinine sulphate (qn-SO4 2-) 28 and trans,trans-1,4-diphenyl-1,3-butadiene (dpb). 29 The studied compounds (C, Table 4) have generally lower  than their analogous compounds containing 1,3,4oxadiazole (B) 26 which is a consequence of the sulphur, the presence of which typically causes the severe quenching of fluorescence. 30On the other hand, the compounds of both mentioned groups (B and C) exhibit lower  in comparison to their directly conjugated counterparts (A). 25 It shows that the insertion of benzene rings between tetrazine and thia/oxadiazole rings leads to the disruption of the fluorescence-favourable conjugation.The absorption wavelengths of the title compounds (C) are higher than for the direct-coupled systems (A), and most often are comparable to the corresponding 1,3,4-oxadiazole arrangements (B).The above described trend of increasing Stokes shifts for the studied compounds containing a methoxy group is in line with the observations for 1,3,4-oxadiazole analogues, but does not occur in the case of directly conjugated heterocyclic rings (entries 2 and 6, Table 4).

Conclusions
In summary, we have elaborated an efficient and universal methodology for the synthesis of 1,2,4,5-tetrazine derivatives conjugated via a 1,4-phenylene linker with a 1,3,4-thiadiazole ring.The developed procedure made it possible to obtain a number of symmetrical and unsymmetrical derivatives.In addition, the methodology can be used for arrangements containing both electron-donating and electron-withdrawing groups.As a result of the research, ten new final products, not previously described in the literature, were obtained and characterised, including by their fluorescent properties.

Experimental Section
General.Melting points were measured on a Stuart SMP3 melting point apparatus.NMR spectra were recorded at 25 C on an Agilent 400-NMR spectrometer at 400 MHz for 1 H and 100 MHz for 13 C, using CDCl3 or DMSO-d6 as solvent and TMS as the internal standard.UV-Vis absorption and 3D fluorescence spectra were registered in methanol solutions (c = 5·10 -6 mol/dm 3 ) with Jasco V-660 and Jasco F-6300 spectrometers, respectively.FT-IR spectra were measured between 4000 and 650 cm -1 on an FT-IR Nicolet 6700 apparatus with a Smart iTR accessory.Elemental analyses were performed with a VarioEL analyser.High-resolution mass spectra were obtained by means of a Waters ACQUITY UPLC/Xevo G2QT instrument.Thin-layer chromatography was performed on silica gel 60 F254 (Merck) thin-layer chromatography plates using benzene/ethyl acetate (1:3 v/v) or chloroform/ethyl acetate (5:1 v/v) as the mobile phases.Materials and procedures.All chemicals, materials and solvents were purchased from Sigma-Aldrich and used as received.Compounds 1-5 were synthesized according to the literature. 26(5-Phenyl-1,3,4-thiadiazol-2-yl)benzonitrile derivatives (6a-d).N'-Benzoyl-4-cyanobenzohydrazide (5a) or derivative (5b-d) (0.02 mol) and Lawesson's reagent (0.42 g, 0.01 mol) were dissolved in toluene (40 mL) and heated in an oil bath with stirring for 7-12 hours.The resulting mixture was filtered and the residue was evaporated.The crude product was purified by column chromatography using a chloroform/ethyl acetate mixture (5:1, v/v) as the eluent.UV-VIS: λmax (MeOH) 296.5 nm (ε10 -3 = 1.97 cm -1 M -1 ), λmax (MeOH) 225.5 nm (ε10 -3 = 3.94 cm -1 M -1 ), λmax (MeOH) 204.0 nm (ε10 -3 = 5.66 cm Preparation of final products (8a-j).The mixtures of one or two of compounds 6a-d (2 mmol of each substrate), sulphur (0.08 g, 2.5 mmol) and dry toluene (60 mL) was cooled to 0 °C and hydrazine hydrate (0.5 mL, 6 mmol) was added dropwise with stirring.The slurry was allowed to reach room temperature and then heated under reflux for 2 h.The mixture was then cooled to room temperature, filtered, and the filtrate was evaporated.The crude product (7a-j) was dissolved in methanol (30 mL), and then 35% hydrogen peroxide (30 mL) was added.After stirring at room temperature for 24 h, the reaction mixture was filtered and concentrated by evaporation.The oily residue or precipitate was filtered and purified by column chromatography using a chloroform/ethyl acetate mixture (5:1, v/v) as the eluent.

Table 2 .
The resulting symmetrical and unsymmetrical derivatives of s-tetrazine conjugated via a 1,4-

Table 4 .
25,26rison of the properties of the title compounds (C) with analogous systems described in the literature (A, B)25,26