INTRODUCTION

Many compounds containing a five-membered heterocyclic ring have unique chemical properties and diverse biological activity types. Thiadiazole is considered a bioisostere of pyrimidine and oxadiazole [1].

Until recently, a thiadiazole fragment was a structural component in the synthesis of antiparasitic and antimicrobial drugs, some of which are still used in clinical practice [2]. Over the last decade, studies have shown that compounds containing a thiadiazole ring in their structure possess antitumor, antibacterial, anticonvulsant, and anti-inflammatory properties [3].

The aim of this work was to determine the spatial structure parameters and the proportion of con-formers of 1-[5-(4-methoxyphenylamino)-1,2,4-thiadiazol-3-yl]-propan-2-ol, a thiadiazole derivative, in solvents of different polarities (CDCl3 and DMSO-d6). As it was shown in recent works, nuclear magnetic resonance (NMR) spectroscopy and nuclear Overhauser effect (NOE) spectroscopy in particular are convenient ways of solving such problems [4–8]. Our approach is based on determining the internuclear distances in the structure of a molecule based on cross relaxation rates obtained by analyzing the integral intensities of the signals in 1H–1H NOESY spectra.

EXPERIMENTAL

All the NMR experiments were performed on a Bruker Avance III 500 spectrometer. The operating frequencies for the 1H and 13C nuclei were 500.17 and 125.77 MHz, respectively. The temperature was controlled with a Bruker Temperature Control Unit (BVT-2000) combined with a Bruker Cooling Unit (BCU 05). The 1H NMR spectra were obtained with a pulse sequence generated by TopSpin 3.6.1 spectrometer software. The number of scans was 128. The spectral range was 7500.00 Hz, the receiver gain coefficient (RG) was 6.4, and the number of points (TD) was 16 384 [9].

Proton decoupled 13C NMR spectra were also obtained with a pulse sequence generated by TopSpin 3.6.1 spectrometer software. The number of scans was 1024. The spectral range was 34 722.22 Hz, the receiver gain was set automatically (256), and the number of points (TD) was 65536 [10].

A “noesygpphpp” pulse program was used to register the 2D NOESY NMR spectra. It generates a sequence consisting of three 90° RF pulses separated by a delay d0 between the first and second pulses, a mixing time delay d8 between the second and third pulses, and the time of free induction decay generation. The period of mixing d8 for the NOESY experiments was from 0.1 to 0.9 s with a step of 0.1 s. The number of scans (NS) was 16. The spectral range was 7500.00 Hz along both axes. The relaxation period (5T1) was chosen according to the recommendations for NMR experiments: 3 s [11–23].

Homo- and heteronuclear spectra (1H–13C HSQC, 1H–13C HMBC, and 1H–1H TOCSY) were obtained using parameters found from 1D experiments (1H and 13C) [9, 24–31].

Quantum-chemical calculations were made using the Gaussian 16 software with an APFD DFT functional that included dispersion correction, in combination with a 6-311++G (2d,2p) basis set.

RESULTS AND DISCUSSION

A molecule of the thiadiazole derivative has five dihedral angles. Changes in these angles result in the formation of a variety of conformers conformers, including those with an intramolecular O‒H···N hydrogen bond. The structures of 104 conformers were optimized. Twenty-eight conformers with intramolecular H-bonds had the lowest energies of those calculated. The remainder had no intramolecular hydrogen bonds. The mutual arrangement of two cyclic fragments that can either lie in the same plane or be displaced relative to each other also affects the formation of additional conformers of the thiadiazole derivative molecule. The vibrational spectra of the conformers were therefore calculated using quantum chemistry. None of the structures had imaginary frequencies.

Signals attributed to the conformers, in which the H2C–CH bond in the isopropanol side fragment is mobile, were observed in the NOESY spectrum. It was difficult to observe other conformers by NOESY, since the conformational distances that characterize the rotation of the remaining fragments of the molecules were beyond the method sensitivity (>5 Å).

Figure 1 shows the barriers to the intramolecular rotation around the H2C–CH (τ2) bond in the isopropanol fragment without intramolecular hydrogen bonds, which enables such structural variety. The difference between the energy barriers is due to the CH3 and OH groups. The O–H···N hydrogen bonds with the heterocycle nitrogen atoms hinder this rotation and can alter the energy barriers. Solvents capable of forming hydrogen bonds with an OH group (proton acceptors) or with N atoms (proton donors) can break the intramolecular N···H–O bond in the thiadiazole derivative molecule and change the isopropanol fragment conformation.

Fig. 1.
figure 1

Barriers to intramolecular rotation around the H2C–CH bond (τ2) in the isopropanol fragment of the thiadiazole derivative molecule.

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Nine resonance signals (Fig. 2) are observed in the 1H NMR spectrum of 1-[5-(4-methoxyphenylamino)-1,2,4-thiadiazol-3-yl]-propan-2-ol (a thiadiazole derivative) in chloroform-d1. The four resonance signals detected at 1.32 (d, H11), 2.39 (s, H12), 2.85 (dd, H9b), and 3.00 ppm (dd, H9a) are attributed to the CH3 (H11 and H12) and CH2 (H9a and H9b) groups of the molecule. The signals attributed to the CH, OH, and NH groups of the molecule are located at 4.27 (m, H10), 7.08 (dd, H2), 7.28 (dd, H3/5), 4.29 (s, ОН), and 8.54 (s, NH) ppm. The unambiguous assignment of the resonance signals was made based on the 2D NMR, published data, and the shape of the characteristic group signals.

Fig. 2.
figure 2

1H NMR spectrum (500 MHz) of the thiadiazole derivative in chloroform-d1.

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When the spatial structure of the thiadiazole derivative molecule was studied and the proportion of conformers was found, nuclear Overhauser effect spectra (1H–1H NOESY) were obtained with different mixing times. Figure 3 shows a NOESY spectrum with a mixing time of 0.9 s. It contains 14 pairs of cross peaks: H2–H11, H3/5–H11, H10–H2, H10–H3/5, H10–H9a, H10–H9b, H10–H11, H11–H9b, H11–H9a, H12–H3/5, NH–H3/5, NH–H2, NH–H10, and NH–H11 corresponding to the interactions between the hydrogen atoms. Further analysis of the integrated intensities of the cross peaks in the NOESY spectra responsible for the conformational and reference distances allowed us to calculate the populations of thiadiazole derivative conformers in the chloroform-d1 solution with high accuracy.

Fig. 3.
figure 3

1H–1H NOESY spectrum of the thiadiazole derivative in chloroform-d1.

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Table 1 shows the chemical shifts of the signals in our 1D spectra and data on the intramolecular interactions found by analyzing the two-dimensional NMR spectra.

Table 1.   1H and 13C NMR chemical shifts and cross-correlation peaks in 2D spectra of the thiadiazole derivative in chloroform-d1
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The test compound conformations in a solvent of different polarity were analyzed at the next stage. Nine resonance signals were observed in the 1H NMR spectrum of the thiadiazole derivative in DMSO-d6 (Fig. 4). The four resonance signals located at 1.17 (d, H11), 2.22 (s, H12), 2.75 (dd, H9b), and 2.89 ppm (dd, H9a) were attributed to the CH3 (H11 and H12) and CH2 (H9a and H9b) groups of the molecule. The signals at 4 to 9 ppm were attributed to the CH– (4.23 ppm, m, H10), 7.28 ppm (dd, H2), 7.21 ppm (dd, H3), 7.72 ppm (dd, H5), 4.79 ppm (s, OH), and 10.77 ppm (s, NH) groups of the molecule. The unambiguous assignment of the resonance signals was made based on the 2D NMR, published data, and the shape of the characteristic group signals.

Fig. 4.
figure 4

1H NMR spectrum (500 MHz) of the thiadiazole derivative in DMSO-d6.

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Nuclear Overhauser effect spectra (1H–1H NOESY) were obtained to determine the spatial structure of the thiadiazole derivative molecule, calculate the interproton distances and the fractions the fractions of the conformers. Figure 5 shows a NOESY spectrum with a mixing time of 0.9 s. It contains six pairs of cross-peaks (H3–H12, H10–H9b, H10–H9a, H10‒H11, H9a–H11, and H9b–H11) corresponding to the H–H interactions. Further analysis of the integrated intensities of the cross peaks in the NOESY spectra responsible for the conformational and reference distances allowed us to calculate the percentage of thiadiazole derivative conformers in the DMSO‑d6 solution with high accuracy.

Fig. 5.
figure 5

1H–1H NOESY spectrum of the thiadiazole derivative in DMSO-d6.

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Table 2 presents data on the chemical shifts in the 1D spectra and intramolecular interactions of the H–H and H–C types obtained by analyzing the 2D spectra.

Table 2.   1H and 13C NMR chemical shifts and cross-correlation peaks in 2D spectra of the thiadiazole derivative in DMSO-d6
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The H10–H11 and H9a–H10 distances were used as the references to determine the other correlations between the hydrogen atoms (H2–H11, H3/5–H11, H10–H2, H10–H3/5, H10–H9a, H10–H9b, H10–H11, H11–H9b, H11–H9a, H12–H3/5, NH–H3/5, NH–H2, NH–H10, and NH–H11) observed in the NOESY experiment. Analysis of the conformer structure indicates that the reference distances are virtually independent of the molecule conformation to a tolerance of ±0.007 Å.

Since the analysis of the barriers to the intramolecular rotation in quantum-chemical calculations shows that there are three stable conformers due to the mobile isopropanol fragment of the molecule and since the conformation-determined distances of conformers A and C are similar within the experimental error, they are considered as one conformer in this work. As a result, the H9a–H10 distances are 3.04 Å for conformer A and 2.47 Å for B. The distances between these nuclei were obtained by analyzing the structure calculated by quantum chemistry, with the reference distance of 2.60 Å.

The relationships between the average integrated intensities of the cross peaks and the period of mixing were plotted after analyzing the NOESY data, and the cross-relaxation peaks were found. They were 0.0209 ± 0.0005, 0.0207 ± 0.0005, and 0.0111 ± 0.0006 for the H9a–H10 and 0.0187 ± 0.0020 for the H10–H11 distances in the thiadiazole derivative structure in chloroform-d1 and DMSO-d6, respectively. The experimental values were found using a spin pair model: 2.51 ± 0.04, 2.60 ± 0.04, 2.84 ± 0.04, and 2.60 ± 0.04 Å for the H9a–H10, H10–H11, and H11–H9a distances in the thiadiazole derivative structure in chloroform-d1 and DMSO-d6, respectively.

The dependences of the difference between the calculated and experimental conformation-dependent distances and the proportions of the conformers (Fig. 6, red and blue lines) were obtained along with the error in determining the experimental distance (dashed line) using a two-position exchange equation. The minima of the values on the graphs correspond to the most probable proportions of conformers, and the point where the lines intersect determines the error value. The results are shown on the diagrams.

Fig. 6.
figure 6

Plot of average calculation error due to fitting the distribution of populations of conformers of thiadiazole derivative in (a) chloroform-d1 and (b) DMSO-d6 to experimental interproton distances (H9a–H10) obtained from 2D NOESY. The dashed line corresponds to the experimental distance error.

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Figure 7a shows the ratio of the most probable conformers of the thiadiazole derivative in chloroform and Fig. 7b presents the respective ratio in DMSO in DMSO. An analysis of the dependences indicates that the ratio of conformers A and B varies considerably due to the change in angle τ2. Conformer B predominates in chloroform relative to A, with 61.9% of conformer B to 38.1% for conformer A (Fig. 7a). However, the situation is reversed in DMSO (Fig. 7b), where the ratio is 79% (conformer A) to 20.2 % (conformer B).

Fig. 7.
figure 7

Distribution of the thiadiazole molecule conformers in (a) chloroform-d1 and (b) DMSO-d6, based on experimental 2D NOESY data.

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This result is similar to the one obtained for hydrazone derivatives (PLP-2FH and PLP-T2CH) and felodipine [6, 32], when the conformational inversion was observed upon the change in the concentration or chemical structure of the molecules. The intramolecular hydrogen bonds that form due to the differences in polymorphic solvates [33–35] play an important role, and the geometry of the molecules of both polymorphs differs in the terminal para-hydroxyl groups, in contrast to hydrazone and felodipine when considering changes in conformers. This effect is based on a mechanism with conformational polymorphism, referred to as solvatomorphism. The produced by the solid phase through the formation of a crystal solvate is therefore also typical of the thiadiazole derivative.

CONCLUSIONS

The proportions of conformers of a 1-[5-(4-methoxyphenylamino)-1,2,4-thiadiazol-3-yl]-propan-2-ol molecule resulting from the mobility of its isopropanol fragment in tow solvents of different polarity were determined by nuclear Overhauser effect spectroscopy and quantum-chemical calculations. The conformational inversion was probably due to the so-called solvatomorphism. Our results can be used in the synthesis of new and modification of the existing biologically active drugs based on thiadiazole derivatives.