Synthesis of Homodrimane Sesquiterpenoids Bearing 1,3-Benzothiazole Unit and Their Antimicrobial Activity Evaluation

Based on some homodrimane carboxylic acids and their acyl chlorides, a series of fourteen 2-homodrimenyl-1,3-benzothiazoles, N-homodrimenoyl-2-amino-1,3-benzothiazoles, 4′-methyl-homodrimenoyl anilides and 4′-methyl-homodrimenthioyl anilides were synthesized and their biological activities were evaluated on five species of fungi (Aspergillus niger, Fusarium solani, Penicillium chrysogenum, P. frequentans, and Alternaria alternata) and two strains of bacteria (Bacillus sp. and Pseudomonas aeruginosa). The synthesis involved the decarboxylative cyclization, condensation and thionation of the said acids, anhydrides or their derivatives with 2-aminothiophenol, 2-aminobenzothiazole, p-toluidine and Lawesson’s reagent. As a result, together with the desired compounds, some unexpected products 8, 25, and 27 were obtained, and the structures and mechanisms for their formation have been proposed. Compounds 4, 9, and 25 showed higher antifungal and antibacterial activity compared to the standards caspofungin (MIC = 1.5 μg/mL) and kanamycin (MIC = 3.0 μg/mL), while compound 8 had comparable activities. In addition, compounds 6, 17, and 27 showed selective antifungal activity at MIC = 2.0, 0.25, and 1.0 μg/mL, respectively.


Introduction
Natural labdane-type diterpenes isolated from terrestrial plants and marine sources are still interesting objects of study due to a wide range of their biological activities [1]. Some of them are obtained from sources in sufficient amounts to be used as precursors for the synthesis of natural analogs, special purpose compounds, or pharmaceutical agents with remarkable properties [2].
The chemistry of 1,3-benzothiazole and its 2-substituted derivatives has become a separate area of research due to a high degree of structural diversity, which generates a wide variety of their applications or pharmacological activities [3][4][5].
A wide variety of reagents and methods which lead to 2-substituted 1,3-benzothiazoles are known. One of the most requested methods of their synthesis involves the cyclocondensation of aromatic aldehydes or others carbonyl compounds such as carboxylic acids, esters, acyl halides, etc., with o-aminophenol [6,7] or its disulfides [8]. Frequently, for the conversion of the resulting amides into the corresponding thioamides, Lawesson's reagent is used, but the course of reactions and their yields strongly depend on the structure of substrates [9].
The synthesis of terpeno-heterocyclic hybrid compounds with a cumulative biological potential is a new direction of organic chemistry that has emerged in the last decade. such as carboxylic acids, esters, acyl halides, etc., with o-aminophenol [6,7] or its disulfides [8]. Frequently, for the conversion of the resulting amides into the corresponding thioamides, Lawesson's reagent is used, but the course of reactions and their yields strongly depend on the structure of substrates [9].
In continuation of our work aimed at the preparation of hybrid terpeno-heterocyclic compounds, herein, we report the result of the synthesis of novel homodrimane sesquiterpenoids bearing 2-substituted 1,3-benzothiazole, N-substituted 2-amino-1,3-benzothiazole and N-substituted p-toluidine units and their antimicrobial properties evaluation.

Synthesis and Characterization
According to the synthesis strategy of the desired compounds, at first, the intermediate carboxylic acids were obtained from commercial (+)-sclareolide (1). It was converted to methoxyester 2 in two steps, with an overall yield of 25%, applying the known procedure [19], followed by the saponification into acid 3 in 89% yield. Starting from sclareolide (1) carboxylic acids 5 and 7 were obtained in five and six steps, with overall yields of 81% and 62%, respectively [13,20] (Scheme 1).
The structures of intermediary compounds as well as final products were confirmed by 1 H, 13 C, 15 N, and 2D NMR spectroscopy and HRMS analysis. The formation of the desired hybrid compounds 4, 6, 8, and 9 was proved, first of all, with the presence of signals attributed to aromatic protons from a common 2-substituted-1,3-benzothiazole unit in a range of 7.32-7.96 ppm. In addition, some individual signals such as a singlet corresponding to protons of C 7 -bonded methoxy group at 3.39 ppm, or doublet of doublets of C 6 -and C 7 -bonded protons at 5.88 and 5.95 ppm confirmed the presence of a terpene unit. Those structures were fully confirmed by the carbon spectral data.
It should be noted that, in the case of acid 7, surprisingly, in addition to the desired compound 9, obtained with a yield of only 5%, the compound 8 with an unexpected structure was afforded as a major reaction product, in 27% total yield. The rearrangement of the carbon skeleton of compound 8 was confirmed by a shift of some signals in the 1 H NMR spectrum compared to the starting acid 7, e.g., by singlet signals of the C 8 -and C 9 -bonded methyl groups at 0.92 and 1.05 ppm and the appearance of new multiplet signals of the C 8 -bonded proton at 1.54-1.56 ppm. The 13 C NMR spectra confirmed this by signals of the C 8 (34.5 ppm) and C 9 (42.1 ppm), those at 130.8 and 139.4 ppm being attributed to C 10 and C 5 , respectively.
The NMR data of compound 8 have been assigned on the basis of their 1D ( 1 H, 13 C, DEPT-135 • ) and 2D homo-( 1 H/ 13 C HSQC, 1 H/ 13 C HMBC and 1 H/ 1 H COSY-45 • ) correlation spectra. An analysis of the 1 H, 13 C, 1 H/ 1 H COSY and 1 H/ 13 C HSQC NMR spectra suggested the presence of two isolated spin systems: CH 2 CH 2 CH 2 (C 1 to C 3 ) and CH 2 CH 2 CH (C 6 to C 8 ) (Figure 1). The rearranged carbon framework of compound 8 was indisputable according to a detailed analysis of its 1 H/ 13 C HMBC spectrum. Thus, the observed correlations of H2-C 2 with two sp 2 hybridized carbons (C 5 , δ C 139.4 and C 10 , δ C 130.8) were indicative of the ∆ 5,10 double bond localization, which was also supported by the correlations of H3-C 18 /C 5 , H3-C 9 /C 10 and H2-C 11 /C 10 . The migration of H3-C 20 methyl from the C 10 to C 9 position was ascertained by the H3-C 20 /C 10 , H3-C 20 /C 8 and H3-C 20 /C 9 and the H3-C 20 /C 11 cross-peaks in the HMBC spectrum.
unit in a range of 7.32-7.96 ppm. In addition, some individual signals such as a singlet corresponding to protons of C7-bonded methoxy group at 3.39 ppm, or doublet of doublets of C6and C7-bonded protons at 5.88 and 5.95 ppm confirmed the presence of a terpene unit. Those structures were fully confirmed by the carbon spectral data.
It should be noted that, in the case of acid 7, surprisingly, in addition to the desired compound 9, obtained with a yield of only 5%, the compound 8 with an unexpected structure was afforded as a major reaction product, in 27% total yield. The rearrangement of the carbon skeleton of compound 8 was confirmed by a shift of some signals in the 1 H NMR spectrum compared to the starting acid 7, e.g., by singlet signals of the C8-and C9-bonded methyl groups at 0.92 and 1.05 ppm and the appearance of new multiplet signals of the C8-bonded proton at 1.54-1.56 ppm. The 13 C NMR spectra confirmed this by signals of the C8 (34.5 ppm) and C9 (42.1 ppm), those at 130.8 and 139.4 ppm being attributed to C10 and C5, respectively.
The NMR data of compound 8 have been assigned on the basis of their 1D ( 1 H, 13 C, DEPT-135°) and 2D homo-( 1 H/ 13 C HSQC, 1 H/ 13 C HMBC and 1 H/ 1 H COSY-45°) correlation spectra. An analysis of the 1 H, 13 C, 1 H/ 1 H COSY and 1 H/ 13 C HSQC NMR spectra suggested the presence of two isolated spin systems: CH2CH2CH2 (C1 to C3) and CH2CH2CH (C6 to C8) ( Figure 1). The rearranged carbon framework of compound 8 was indisputable according to a detailed analysis of its 1 H/ 13 C HMBC spectrum. Thus, the observed correlations of H2-C2 with two sp 2 hybridized carbons (C5, δC 139.4 and C10, δC 130.8) were indicative of the Δ 5,10 double bond localization, which was also supported by the correlations of H3-C18/C5, H3-C9/C10 and H2-C11/C10. The migration of H3-C20 methyl from the C10 to C9 position was ascertained by the H3-C20/C10, H3-C20/C8 and H3-C20/C9 and the H3-C20/C11 cross-peaks in the HMBC spectrum. The rearrangement of 2-homodrimenyl-1,3-benzothiazole 8 carbon skeleton can be explained by the following reaction pathway (Scheme 2). The reaction started with formation of triphenylphosphonium chloride as product of Ph3P and CCl4 interaction, followed by its condensation with carboxylic acid which led to acylphosphonium intermediate 10. Next, the nucleophilic attack of the amino group to the carbonyl gave the intermediate amide 11 and triphenylphosphine oxide. Then, the nucleophilic attack of the deprotonated sulfur atom led to the unstable cyclic intermediate 12. Next, the formation of compound 8 was a result of the elimination reaction, which led to the desired 2-homodrimenyl-1,3-benzothiazole 9 that by protonation gave carbocation 13. The latter suffered a rearrangement of the carbon skeleton as a result of the C10-bonded methyl group migration to C9, followed by both C5 deprotonation and C5-C10 double bond formation. The rearrangement of 2-homodrimenyl-1,3-benzothiazole 8 carbon skeleton can be explained by the following reaction pathway (Scheme 2). The reaction started with formation of triphenylphosphonium chloride as product of Ph 3 P and CCl 4 interaction, followed by its condensation with carboxylic acid which led to acylphosphonium intermediate 10. Next, the nucleophilic attack of the amino group to the carbonyl gave the intermediate amide 11 and triphenylphosphine oxide. Then, the nucleophilic attack of the deprotonated sulfur atom led to the unstable cyclic intermediate 12. Next, the formation of compound 8 was a result of the elimination reaction, which led to the desired 2-homodrimenyl-1,3-benzothiazole 9 that by protonation gave carbocation 13. The latter suffered a rearrangement of the carbon skeleton as a result of the C 10 -bonded methyl group migration to C 9 , followed by both C 5 deprotonation and C 5 -C 10 double bond formation. Next, a series of new N-homodrimenoyl-2-amino-1,3-benzothiazoles were prepared, starting from the intermediate carboxylic acids 3, 5, 7, and 20, via their acyl chlorides 14, 16, 18, and 21, generated in situ in 25-65% yields. It should be mentioned that the acid 20 was obtained from the commercially available (+)-sclareolide (1) in 6 steps, with an overall yield of 60%, according to the known method [11]. The desired N-substituted 2-amino-1,3-benzothiazoles 15, 17, 19, and 22 were obtained with yields between 40-84% by acylation of 2-amino-1,3-benzothiazole with the mentioned sesquiterpene acyl chlorides under the mentioned conditions (Scheme 3).
According to the NMR spectra, the hybrids involved both heterocyclic and terpene units, and their accurate masses were confirmed by a high-resolution mass spectrometry (HRMS) analysis. All proton spectra of compounds 15, 17, 19, and 22, include the signals of aromatic protons in a range of 7.30-7.82 ppm, together with the signals specific for terpene unit such as singlets of C 8 -bonded methyl groups at 1.69-1.82 ppm and C 7 -bonded methoxy group at 3.40 ppm, C 6 -and C 7 -bonded protons at 3.50, 5.93, and 5.94 ppm, singlets of C 17exomethylene group at 4.39 and 4.73 ppm, and broad singlets of amidic protons in a range of 9.73-11.26 ppm. The structures of the reported N-substituted 2-amino-1,3-benzothiazoles were additionally confirmed by the 13 C NMR spectra.

Scheme 2.
A plausible reaction pathway of formation of compound 8.
According to the NMR spectra, the hybrids involved both heterocyclic and terpene units, and their accurate masses were confirmed by a high-resolution mass spectrometry (HRMS) analysis. All proton spectra of compounds 15, 17, 19, and 22, include the signals of aromatic protons in a range of 7.30-7.82 ppm, together with the signals specific for terpene unit such as singlets of C8-bonded methyl groups at 1.69-1.82 ppm and C7-bonded methoxy group at 3.40 ppm, C6-and C7-bonded protons at 3.50, 5.93, and 5.94 ppm, singlets of C17-exomethylene group at 4.39 and 4.73 ppm, and broad singlets of amidic protons in a range of 9.73-11.26 ppm. The structures of the reported N-substituted 2-amino-1,3-benzothiazoles were additionally confirmed by the 13 C NMR spectra.
An attempt to perform the direct amidation of acids 3 and 5 with p-toluidine in the presence of N,N -dicyclohexylcarbodiimide (DCC) and 4-dimethylaminopyridine (4-DMAP) gave better results, because the yields of amides 26 and 28 increased up to 76% and 94%, respectively (Scheme 4). Together with the signals of protons from terpene units, the proton spectra of amides 23, 26, and 28 contained the singlets of C 4 -bonded methyl in a range of 2.30-2.32 ppm and doublets of aromatic protons from 7.11 ppm to 7.37 ppm, and a broad singlet of the amidic proton at 7.53-7.70 ppm. The structures of the mentioned compounds were fully confirmed by the 13 C NMR spectra. presence of N,N′-dicyclohexylcarbodiimide (DCC) and 4-dimethylaminopyridine (4-DMAP) gave better results, because the yields of amides 26 and 28 increased up to 76% and 94%, respectively (Scheme 4). Together with the signals of protons from terpene units, the proton spectra of amides 23, 26, and 28 contained the singlets of C4′-bonded methyl in a range of 2.30-2.32 ppm and doublets of aromatic protons from 7.11 ppm to 7.37 ppm, and a broad singlet of the amidic proton at 7.53-7.70 ppm. The structures of the mentioned compounds were fully confirmed by the 13 C NMR spectra. After that, amides 23, 26, and 28 were submitted to the thionation reaction using Lawesson's reagent (LR) in toluene [22]. In the case of amide 23, a reaction occurred, and thioamide 24 was obtained in 50% yield (Scheme 4). Its structure was confirmed by the NMR and HRMS analyses. In the 1 H and 13 C NMR spectra, down field shifts of the amidic proton to 9.03 ppm and of C12 to 200.9 ppm compared to the initial amide 23 were observed.
Heterocyclization of thioamide 24 performed in the presence of potassium ferricyanide under basic conditions (30% NaOH) [22] did not lead to the desired 2-substituted 6-methyl-benzothiazole and gave an unexpected compound 25 in 41% yield After that, amides 23, 26, and 28 were submitted to the thionation reaction using Lawesson's reagent (LR) in toluene [22]. In the case of amide 23, a reaction occurred, and thioamide 24 was obtained in 50% yield (Scheme 4). Its structure was confirmed by the NMR and HRMS analyses. In the 1 H and 13 C NMR spectra, down field shifts of the amidic proton to 9.03 ppm and of C 12 to 200.9 ppm compared to the initial amide 23 were observed.
Heterocyclization of thioamide 24 performed in the presence of potassium ferricyanide under basic conditions (30% NaOH) [22] did not lead to the desired 2-substituted 6-methylbenzothiazole and gave an unexpected compound 25 in 41% yield (Scheme 4). Its structure was elucidated based on the NMR and HRMS spectra. Comparing with the initial thioamide 24, its 1 H NMR spectra did not contain the signals of an amidic proton and of one of C 11bonded proton, but the singlets of a C 8 -bonded methyl group and one of C 11 -H were shifted to 1.72 and 6.19 ppm, respectively. The same is true of the carbon spectrum, where some signals are strongly shifted, e.g., C 8 (62.9 ppm), C 11 (123.6 ppm), C 9 (170.4 ppm), and C 12 (176.6 ppm).
The analysis of the structure of compound 25 by 1 H, 13 C, 1 H/ 1 H COSY and 1 H/ 13 C HSQC NMR spectra suggested the presence of two isolated spin systems: CH 2 CH 2 CH 2 (C 1 to C 3 ) and CHCH 2 CH 2 (C 5 to C 7 ) (Figure 2). In the 1 H/ 13 C HMBC spectrum correlations of H-C 11 with quaternary carbons (C 9 , δ C 170.4 and C 12 , δ C 176.6) were observed, which was also supported by the correlations of H-C 11 /C 8 . In addition, the correlation of H3-C 17 /C 12 confirms the formation of the 5-membered heterocycle.
In the NOESY spectrum of compound 25, there is no NOE correlation between C 11 -H and C 2 -H from the aromatic ring ( Figure 2) that clearly indicates the E-configuration for C 12 = N-double bond. HSQC NMR spectra suggested the presence of two isolated spin syst (C1 to C3) and CHCH2CH2 (C5 to C7) (Figure 2). In the 1 H/ 13 C HMBC spe of H-C11 with quaternary carbons (C9, δC 170.4 and C12, δC 176.6) were was also supported by the correlations of H-C11/C8. In addition, H3-C17/C12 confirms the formation of the 5-membered heterocycle. In the NOESY spectrum of compound 25, there is no NOE correlat and C2′-H from the aromatic ring ( Figure 2) that clearly indicates the E C12 = N-double bond.
The thionation of amide 26 under the same conditions occur unexpected cyclic thioamide 27 in 52% yield. The formation of th confirmed by the absence of an amidic proton signal, a shift of the C8-CH3 to 1.56 ppm and by the appearance of the C9-H doublet at 1.88 chemical shift of C8 and C9 atoms to 59.2 and 58.2 ppm and a downfiel C12 (175.9 ppm) in the 13 C NMR spectra also confirmed the structure of Structural analysis of compound 27 by 1 H, 13 C, 1 H/ 1 H COSY and 1 H spectra suggested the presence of two isolated spin systems: CH2CH2C CH2CHCH (C5 to C7) ( Figure 3). The rearranged carbon framework of indisputable by a detailed analysis of its 1 H/ 13 C HMBC spectrum. T correlations of H-C5 with two sp 2 hybridized carbons (C6, δC 126.8 and indicative of the Δ 6,7 double bond localization, which was also s correlations of H3-C17/C7. The relative configuration at C17 was deduced of H3-C17/H3-C10 NOESY correlation. The position of N has been confir HMBC spectra and was supported by the correlations of H2-C1 cross-peaks ( Figure 3).  The thionation of amide 26 under the same conditions occurred and gave an unexpected cyclic thioamide 27 in 52% yield. The formation of the N-C 8 bond was confirmed by the absence of an amidic proton signal, a shift of the singlet signal of C 8 -CH 3 to 1.56 ppm and by the appearance of the C 9 -H doublet at 1.88 ppm. The upfield chemical shift of C 8 and C 9 atoms to 59.2 and 58.2 ppm and a downfield shifted signal of C 12 (175.9 ppm) in the 13 C NMR spectra also confirmed the structure of compound 27.
Structural analysis of compound 27 by 1 H, 13 C, 1 H/ 1 H COSY and 1 H/ 13 C HSQC NMR spectra suggested the presence of two isolated spin systems: CH 2 CH 2 CH 2 (C 1 to C 3 ) and CH 2 CHCH (C 5 to C 7 ) (Figure 3). The rearranged carbon framework of compound 27 was indisputable by a detailed analysis of its 1 H/ 13 C HMBC spectrum. Thus, the observed correlations of H-C 5 with two sp 2 hybridized carbons (C 6 , δ C 126.8 and C 7 , δ C 130.6) were indicative of the ∆ 6,7 double bond localization, which was also supported by the correlations of H3-C 17 /C 7 . The relative configuration at C 17 was deduced from the absence of H3-C 17 /H3-C 10 NOESY correlation. The position of N has been confirmed by the 1   In the NOESY spectrum of compound 25, there is no NOE correla and C2′-H from the aromatic ring ( Figure 2) that clearly indicates the C12 = N-double bond.
The thionation of amide 26 under the same conditions occu unexpected cyclic thioamide 27 in 52% yield. The formation of th confirmed by the absence of an amidic proton signal, a shift of th C8-CH3 to 1.56 ppm and by the appearance of the C9-H doublet at 1.8 chemical shift of C8 and C9 atoms to 59.2 and 58.2 ppm and a downfie C12 (175.9 ppm) in the 13 C NMR spectra also confirmed the structure o Structural analysis of compound 27 by 1 H, 13 C, 1 H/ 1 H COSY and spectra suggested the presence of two isolated spin systems: CH2CH CH2CHCH (C5 to C7) ( Figure 3). The rearranged carbon framework of indisputable by a detailed analysis of its 1 H/ 13 C HMBC spectrum. T correlations of H-C5 with two sp 2 hybridized carbons (C6, δC 126.8 and indicative of the Δ 6,7 double bond localization, which was also correlations of H3-C17/C7. The relative configuration at C17 was deduce of H3-C17/H3-C10 NOESY correlation. The position of N has been confi HMBC spectra and was supported by the correlations of H2-C cross-peaks ( Figure 3).  However, in the case of amide 28, from the thionation reaction mixture, the same amides 26 and 27 were isolated in 37% and 21% yields, respectively (Scheme 4). Their spectral data were in accordance with those obtained earlier.
Returning to the compound 25, it can be said that its formation in place of the desired benzothiazole is due to the reaction conditions and the molecular structure of thioamide 24 which permits the existence of a tautomeric thioketo-enothiol 24<->29 system. In the basic medium, the tautomer 29 easily generated enothiolate which due to the activation by hexacyanofferate ions attacked the C 8 -C 9 double bond and generated the intermediate carbocation 30 (Scheme 5). In such a way, the formation the new C 8 -S bond occurred simultaneously with the one C 11 -proton elimination giving compound 25.
Returning to the compound 25, it can be said that its formation in place of the desired benzothiazole is due to the reaction conditions and the molecular structure of thioamide 24 which permits the existence of a tautomeric thioketo-enothiol 24<->29 system. In the basic medium, the tautomer 29 easily generated enothiolate which due to the activation by hexacyanofferate ions attacked the C8-C9 double bond and generated the intermediate carbocation 30 (Scheme 5). In such a way, the formation the new C8-S bond occurred simultaneously with the one C11-proton elimination giving compound 25. Scheme 5. Plausible reaction pathway for formation of compound 25.
The formation of compound 27 can be explained by a sequence of transformations depicted in Scheme 6. In this case, the Lawesson's reagent played a double role. The first role is to interact with axial the C7-bonded methoxy group of the amide 28, stimulating its elimination and generating the carbocation 32 which by deprotonation offered amide 26 (see Schemes 4 and 6). On the other hand, the thionation with Lawesson's reagent gave an intermediate carbocation 33 which suffered a cyclization followed by deprotonation into cyclic thioamide 27. Note that there are several resonance structures, but from our point of view, intermediates 32 and 33 are more stable. Scheme 6. Plausible reaction pathway for formation of compound 27.

Antimicrobial Activity
All synthesized compounds were subjected to preliminary screening for their in vitro antifungal and antibacterial activities [23] against pure cultures of fungal species Aspergillus niger, Fusarium solani, Penicillium chrysogenum, Penicillium frequentans, and Alternaria alternata and both Gram-positive Bacillus sp. and Gram-negative Pseudomonas aeruginosa bacteria strains. The obtained minimum inhibitory concentration (MIC) values revealed that compounds 4 and 17 possessed a high nonselective antifungal (MIC 0.094 and 0.25 μg/mL, respectively) activity (Table 1, entries 1 and 6) in comparison with caspofungin. Moreover, compounds 6, 8, 9, 25, and 27 possessed a promising antifungal activity (Table 1,  The formation of compound 27 can be explained by a sequence of transformations depicted in Scheme 6. In this case, the Lawesson's reagent played a double role. The first role is to interact with axial the C 7 -bonded methoxy group of the amide 28, stimulating its elimination and generating the carbocation 32 which by deprotonation offered amide 26 (see Schemes 4 and 6). On the other hand, the thionation with Lawesson's reagent gave an intermediate carbocation 33 which suffered a cyclization followed by deprotonation into cyclic thioamide 27. Note that there are several resonance structures, but from our point of view, intermediates 32 and 33 are more stable.
Returning to the compound 25, it can be said that its formation in place of the desired benzothiazole is due to the reaction conditions and the molecular structure of thioamide 24 which permits the existence of a tautomeric thioketo-enothiol 24<->29 system. In the basic medium, the tautomer 29 easily generated enothiolate which due to the activation by hexacyanofferate ions attacked the C8-C9 double bond and generated the intermediate carbocation 30 (Scheme 5). In such a way, the formation the new C8-S bond occurred simultaneously with the one C11-proton elimination giving compound 25. Scheme 5. Plausible reaction pathway for formation of compound 25.
The formation of compound 27 can be explained by a sequence of transformations depicted in Scheme 6. In this case, the Lawesson's reagent played a double role. The first role is to interact with axial the C7-bonded methoxy group of the amide 28, stimulating its elimination and generating the carbocation 32 which by deprotonation offered amide 26 (see Schemes 4 and 6). On the other hand, the thionation with Lawesson's reagent gave an intermediate carbocation 33 which suffered a cyclization followed by deprotonation into cyclic thioamide 27. Note that there are several resonance structures, but from our point of view, intermediates 32 and 33 are more stable. Scheme 6. Plausible reaction pathway for formation of compound 27.

Antimicrobial Activity
All synthesized compounds were subjected to preliminary screening for their in vitro antifungal and antibacterial activities [23] against pure cultures of fungal species Aspergillus niger, Fusarium solani, Penicillium chrysogenum, Penicillium frequentans, and Alternaria alternata and both Gram-positive Bacillus sp. and Gram-negative Pseudomonas aeruginosa bacteria strains. The obtained minimum inhibitory concentration (MIC) values revealed that compounds 4 and 17 possessed a high nonselective antifungal (MIC 0.094 and 0.25 μg/mL, respectively) activity (Table 1, entries 1 and 6) in comparison with caspofungin. Moreover, compounds 6, 8, 9, 25, and 27 possessed a promising antifungal activity (Table 1,

Synthesis and Characterization
The IR spectra were recorded on a Spectrum 100 FT-IR spectrometer (Perkin-Elmer, Shelton, CT, USA) using an ATR technique. The 1 H, 13 C, and 15 N NMR (400, 100, and 40 MHz, respectively) and COSY, 1 H-13 C HSQC, 1 H-13 C HMBC, DEPT, and 1 H-15 N HSQC, 1 H-15 N HMBC spectra were acquired on a Bruker Avance DRX 400 spectrometer (Bruker BioSpin, Rheinstetten, Germany) in CDCl 3 (NMR spectra for all the compounds are available online, see the Supplementary Materials). The 1 H NMR chemical shifts were reported relative to the residual solvent protons as internal standards (7.26 ppm). The solvent carbon atoms served as internal standard for the 13 C NMR spectra (77.0 ppm). The 15 N NMR spectra were obtained using MeNO 2 (380.5 ppm) and urea (73.4 ppm) as internal standards. Optical rotations measurements were performed on a Jasco DIP-370 polarimeter (Rudolph Research Analytical, Hackettstown, NJ, USA) with a 10 cm microcell. Melting points were determined on a Boetius (VEB Analytik, DDR) hot stage apparatus and were not uncorrected. The progress of reactions and purity of products were examined by TLC on Merck silica gel 60 plates, eluent CH 2 Cl 2 or a mixture of CH 2 Cl 2 -MeOH, 99:1; 49:1. Visualization was achieved by the treatment with conc. H 2 SO 4 and heating at 80 • C or using an UV lamp (254 or 365 nm). All solvents were purified and dried by standard techniques prior to use.