Bis(2-cyanoacetohydrazide) as precursors for synthesis of novel azoles/azines and their biological evaluation

Mohamed Ahmed Elian Sophy; Mohamed Ahmed Mahmoud Abdel Reheim

doi:10.1515/hc-2022-0167

Article Open Access

Bis(2-cyanoacetohydrazide) as precursors for synthesis of novel azoles/azines and their biological evaluation

Mohamed Ahmed Elian Sophy and Mohamed Ahmed Mahmoud Abdel Reheim

Published/Copyright: December 12, 2023

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From the journal Heterocyclic Communications Volume 29 Issue 1

Abstract

The present work deals with the synthesis of 2-cyano-N′-(3-(2-(2-cyanoacetyl) hydrazinyl) cyclohex-2-en-1-ylidene) acetohydrazide 3 that used as a key precursor in the manufacture of new heterocyclic derivatives, such as pyrazole, pyrane, pyridine, and pyrole incorporating cyclohexene moiety via its reaction with a variety of nucleophilic and electrophilic reagents. The newly synthesized compounds were characterized by their elemental analyses (IR, ¹H-NMR, ¹³C-NMR, and Mass spectra) and assessed for their anti-microbial activity.

Graphic abstract

Keywords: hydrazide–hydrazone; pyrazole; pyrane; pyridine; pyrole; spectral data; anti-microbial activity

1 Introduction

1,3-Cyclohexanedione and their related heterocyclic attracted our attention due to their derivatives' pronounced biological activity. Their various applications include antimalarials [1], plant growth promoters [2–4], herbicides [5–7], and other applications [8,9]. The present investigation aimed to synthesize and characterize newer pyrazole, pyrane, pyridine, and pyrole incorporating cyclohexene moiety that has a specific biological activity. The literature survey reveals that pyrazole derivatives exist in various pharmaceutical formulations with anti-inflammatory effects [10] and other applications [11–15]. Pyrane derivatives are used as antimicrobial [16], anticancer [17], and antifungal agents [18]. Additionally, pyridine derivatives have been found to possess diverse pharmacological activities such as anticancer [19], antimicrobial [20,21], and anti-diabetic [22]. Moreover, the pyrole derivatives have received a great deal of attention due to their pharmacological importance. Pyroles were reported to possess antimicrobial [23], anticancer [24], and antiinflammatory effects [25].

2 Results and discussion

The study reported in this article, involved cyanoethanoic acid hydrazide and its derivatives, which are considered versatile reagents for the synthesis of a wide variety of heterocyclic compounds, due to their active methylene moiety and the β-functional nitrile moiety of the molecule is a favorable unit for addition reactions followed by cyclization or via cycloaddition with some electrophilic and nucleophilic reagents. Thus, the reaction of 1,3-cyclohexanedione 1 with cyanoacetic acid hydrazide in glacial acetic acid solution at room temperature, due to the activity of 1,3-dione and the methylene moiety at C₂ one mole of cyanoacetic acid hydrazide condensed with carbonyl group while another mole reacted with enol form gave 2-cyano-N′-(3-(2-(2-cyanoacetyl)hydrazinyl)cyclohex-2-en-1-ylidene)acetohydrazide 3 [26,27]. The structure of compound 3 was established and confirmed as the reaction product based on its elemental analyses and spectral data. The IR spectrum revealed the presence of NH groups stretching at 3,447–3,224 cm⁻¹, the CN groups stretching at 2,216 cm⁻¹, and revealed the carbonyl group at 1,678 cm⁻¹. Also, the ¹H-NMR spectrum showed characteristic singlet signals at δ = 8.52, 9.96, and 10.38 ppm for 3NH protons, a multiplet signals in the region of δ = 1.72–1.91 ppm due to the methylene group in the cyclohexene moiety, a multiplet signals at δ = 2.23–2.32 ppm corresponding to the hydrogens of the two methylene groups in the cyclohexene moiety and shows characteristic singlet signals at δ = 3.36, 3.95, and 5.54 ppm for two methylene groups and CH-cyclohexene, respectively. Moreover, the ¹³C-NMR spectrum of compound 3 shows the following signals at δ = 21.86, 24.70, 26.52, 26.63, 31.45, 86.93, 116.66, 116.88, 156.79, 160.20, 162.12, and 164.67. The structure assigned for compound 3 was fully supported by its mass spectrum, which showed a molecular formula C₁₂H₁₄N₆O₂ (m/z% = 275, M⁺ + 1, 1.01).

The proclivity of bis (2-cyanoacetohydrazide) derivative 3 toward some electrophilic reagents was investigated. Thus, the condensation of compound 3 with dimethylformamide–dimethylacetal (DMF–DMA) in dioxane produced a crylohydrazide derivative 4 as outlined in (Scheme 1). The structure of compound 4 was confirmed from their elemental analyses and spectral data. The structure of compound 4 was mainly elucidated by the ¹H-NMR spectra which showed a singlet signal at δ = 3.10 ppm assigned to the N (CH₃)₂, at δ = 3.13 ppm assigned to the N(CH₃)₂, at δ = 7.51 ppm assigned to CH-oleffinic and at δ = 7.72 ppm assigned to CH-oleffinic. The mass spectrum of compound 4 gave a strong peak at m/z = 384 (M⁺) corresponding to its molecular weight.

Scheme 1

Synthesis of hydrazide derivatives (3–7).

A crylohydrazide derivative 4 showed interesting reactivity toward binucleophilic reagent. So, when compound 4 was reacted with hydrazine hydrate afforded the 3-amino-N′-(3-(2-(3-amino-1H-pyrazole-4-carbonyl) hydrazinyl) cyclohex-2-en-1-ylidene)-1H-pyrazole-4-carbohydrazide 5. The structure of compound 5 was established and confirmed as the reaction product based on their elemental analyses and spectral data. The IR spectrum showed an absorption band in the region 3,400, 3,163 cm⁻¹ assignable for NH₂/NH, in addition to the disappearance of the cyano function group. Its ¹H-NMR spectrum revealed a new signal at δ = 7.12 ppm for CH-pyrazoles and a singlet signal at δ = 6.99 ppm assignable to the NH₂ groups. Its mass spectrum showed a molecular ion peak at m/z = 358 (M⁺) corresponding to a molecular formula C₁₄H₁₈N₁₀O₂.

Cyclocondensation of hydrazide–hydrazone 3 with thioglycolic acid in boiling DMF, anhydrous ZnCl₂ was added as a catalyst furnished the bis(2-(4-hydroxythiazol-2-yl) acetohydrazide) derivative 7 [26]. We first assumed that the reaction product is formed via the initial nucleophilic addition of the mercapto function to the nitrile group, followed by dehydration and subsequent tautomerization. The IR, ¹H-NMR, ¹³C-NMR, and Mass spectra are in agreement in all respects with the proposed structure.

The reactivity of hydrazide–hydrazone derivative 3 toward aryl aldehyde was studied. Thus, benzaldehyde condensed with compound 3 afforded the corresponding arylidene derivative 8. The structure of compound 8 was based on its analytical and spectral data (experimental section). Further evidence for structure 8 was obtained by studying their chemical reactivity towards some chemical reagents. So, when compound 8 was reacted with hydrazine hydrate afforded the corresponding bis(3-amino-5-phenyl-1H-pyrazole-4-carbohydrazide) derivative 9. The structure of compound 9 was mainly elucidated by the ¹H-NMR spectra which showed a new singlet signal at δ = 5.90 ppm for NH₂ groups [28,29].

Furthermore, the behavior of hydrazide-hydrazone derivative 3 towards binucleophilic reagents was also investigated. Thus, the reaction of compound 3 with salicyladehyde in ethanol in the presence of a catalytic amount of piperidine afforded acyclic structure 10. The reaction probably takes place through condensation of the aldehydic group with the active methylene group. The presence of the nitrile groups in the IR spectra at 2,222 cm⁻¹ revealed no cyclization process. Trials to cyclize compound 10 by nucleophilic attack of the hydroxyl group on the nitrile group succeeded under mild conditions such as refluxing of 10 in dioxane in a basic medium to afford 11. The absence of the nitrile function in the IR spectra confirms the cyclization process. Also, the ¹H-NMR spectra of compound 11 reveal the CH-pyrane as a singlet signal at δ = 8.03 ppm [30,31] (Scheme 2).

Scheme 2

Synthesis of carbohydrazidederivatives (8–11).

The behavior of hydrazide-hydrazone derivative 3 toward active methylene reagents was also investigated. Thus, a considerable approach for the synthesis of 4,6-diamino-1-((3-((4,6-diamino-3-cyano-2-oxopyridin-1(2H)-yl)amino)cyclohex-2-en-1-ylidene)amino)-2-oxo-1,2-dihydropyridine-3-carbonitrile 12, 1-((3-((3-cyano-4,6-dimethyl-2-oxopyridin-1(2H)-yl)amino)cyclohex-2-en-1-ylidene) amino)-4,6-dimethyl-2-oxo-1,2-dihydropyridine-3-carbonitrile 13 were fulfilled through interaction of hydrazide-hydrazone derivative 3 with malononitrile and acetylacetone, respectively. Assignment of the structure 12 for the reaction product was based on its correct analyses and compatible spectroscopic data, the ¹H-NMR of the structure revealed a singlet signal at δ = 5.77, 6.20 ppm corresponding to NH₂ groups. Also, a nice singlet signal at δ = 6.88 ppm is due to CH-pyridine. Also, the mass spectrum of compound 12 agrees with the proposed structure, it showed a molecular ion peak at m/z = 406 (M⁺) in agreement with its molecular formula C₁₈H₁₈N₁₀O₂. However, the structure confirmation of 13 was assisted by their analytical and spectral data. The IR spectrum displayed characteristics of an absorption band at 3,448 cm⁻¹ related to the NH group, an absorption band at 2,220 cm⁻¹ related to the CN as well as the carbonyl group absorption band at 1,627 cm⁻¹. In the ¹H-NMR spectrum two characteristic singlet signals at δ = 2.23, 2.31 ppm attributable to the four methyl groups, a singlet signal at δ = 6.20 ppm due to CH-pyridine moiety, the mass spectrum showed a molecular ion peak in accordance with the proposed structure [32,33,29]. Also, we converted the target compound 3 into the corresponding 1-((3-((5-cyano-4-methyl-2,6-dioxo-3,6-dihydropyridin-1(2H)-yl)amino)cyclohex-2-en-1-ylidene) amino)-4-methyl-2,6-dioxo-1,2,5,6-tetrahydropyridine-3-carbonitrile 14. This was accomplished by the fusion of equimolar amounts of compound 3 and ethyl acetoacetate without solvent for about 12 h. The structure of compound 14 was established and confirmed as the reaction product based on their elemental analyses and spectral data. The IR spectrum of 14 showed the presence of an NH stretching at 3,426 cm⁻¹, one cyano group stretching at 2,223 cm⁻¹ and one carbonyl group stretching at 1,727 cm⁻¹. Also, the ¹H-NMR and mass spectrum of the same product are in accordance with the proposed structure (Scheme 3).

Scheme 3

Synthesis of 1,2-dihydropyridine-3-carbonitrile derivatives (12–14).

Hydrazide–hydrazone derivative 3 underwent Gewald thiophene synthesis via its reaction with malononitrile and elemental sulfur in refluxing ethanol containing TEA as a basic catalyst afforded the bis(3,5-diamino-4-cyanothiophene-2-carbohydrazide) derivative 15. The analytical and spectral data are consistent with the proposed structure. The IR spectrum of 15 displayed absorption band at 3,423 and 3,197 cm⁻¹ for NH₂/NH, an absorption band at 2,203 cm⁻¹ for the nitrile group, and the carbonyl absorption band at 1,628 cm⁻¹. The structure of 15 was mainly elucidated by the ¹H-NMR spectra which showed new two singlet signals at δ = 6.10 and 6.60 ppm for NH₂ protons. The mass spectrum of the same compound is in accordance with the proposed structure. It showed a very intense molecular ion peak at m/z 472 (M⁺ + 2), and a few fragments agree with the proposed structure.

On the other hand, the reaction of compound 3 with phenyl isothiocyanate and elemental sulfur afforded the 4-amino-N′-(3-(2-(4-amino-3-phenyl-2-thioxo-2,3-dihydrothiazole-5-carbonyl)-hydrazinyl)cyclohex-2-en-1-ylidene)-3-phenyl-2-thioxo-2,3-dihydrothiazole-5-carbohydrazide 16. The analytical and spectral data of compound 16 are consistent with the proposed structure [29,32,34]. Moreover, the reaction of compound 3 with cyclohexanone and elemental sulfur in the presence of TEA. The reaction led to the formation of bis(2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carbohydrazide) derivative 17. Formation of 17 took place according to the reaction of cyclohexanone with methylene moiety and elemental sulfur (application of Gewald’s thiophene synthesis). The analytical and spectral data of 17 were in agreement with the proposed structure (see Experimental section) [28,35] (Scheme 4).

Scheme 4

Synthesis of thiophene-3-carbohydrazide derivatives (15–17).

Next, we studied the reactivity of the hydrazide-hydrazone derivative 3 toward some electrophilic reagents under alkaline conditions. Thus, the reaction of compound 3 with acetonitrile dimer in refluxing ethanol in the presence of a catalytic amount of piperidine afforded bis(2-amino-4-cyano-5-methyl-1H-pyrrole-3-carbohydrazide) derivative 18. The structure of compound 18 was established and confirmed as the reaction product based on its elemental analyses and spectral data. The infrared spectrum revealed an absorption band at 3,450–3,207 cm⁻¹ for the amino groups, a sharp band at 2,200 cm⁻¹ for the CN function, and a strong band at 1,641 cm⁻¹ for the carbonyl group. The ¹H-NMR spectrum supports the proposed structure which reveals new a singlet signal at δ = 2.20 ppm for methyl groups and a singlet signal at δ = 6.39 ppm for NH₂ protons. The mass spectrum of the same product is by the proposed structure; it showed a molecular ion peak at 434 (M⁺).

Our research deals with the effective use of α, β-unsaturated ketones in the synthesis of 4H-pyrane derivatives. Thus, the reaction of chalcones with hydrazide-hydrazone derivative 3 in refluxing ethanol in the presence of piperidine to give substituted pyrane 19a,b. The structures of synthesized compounds were confirmed by IR, ¹H-NMR, ¹³C-NMR, and mass spectra. The IR spectrum of compound 19a as an example was characterized by the disappearance of the cyano group and showed an absorption band at υ _max = 3,416–3,220, 2,937–2,860, and 1,629 cm⁻¹ due to the amino, aliphatic and carbonyl function, respectively. The ¹H-NMR spectrum supports the proposed structure. This reveals new two doublets at δ = 3.66, 5.71 ppm characteristic for 4H-pyrane and = CH-pyrane, respectively. The mass spectrum of 19a showed a molecular ion peak m/z 690 (M⁺) with a base peak at m/z = 57 (100%) is consistent with the proposed structure.

On the other hand, 4-aryl-2-oxo-pyridine-3,5-dicarbonitrile derivatives 20a,b were synthesized in an excellent yield upon treatment of compound 3 with arylidinemalononitrile in the presence of piperidine as a catalyst. The products 20a,b are apparently formed via a Michael-type addition of the active methylene group in compound 3 to the activated double bond in arylidinemalononitrile to form non-isolable intermediate through an intramolecular cyclization and subsequent tautomerization to give 20a,b. The structure of pyridine derivative 20 was confirmed from the elemental analyses and spectral data of the isolated product. The IR spectrum of 20a displayed stretching absorption bands at 3,419–3,200, 2,939, 2,211, and 1,632 cm⁻¹ for the amino, aliphatic, cyano, and carbonyl function groups, respectively. Its ¹H-NMR spectrum displayed new a singlet signal at δ = 5.70 ppm assignable to the NH₂ protons and a singlet signal at δ = 10.30 ppm assignable to the NH proton. Moreover, an EI mass spectroscopic technique gave its correct molecular ion peak at m/z = 648 (M⁺ + 2) [36,37] (Scheme 5).

Scheme 5

Synthesis of pyran-3-carbohydr-azidederivatives (18–20).

Moreover, the proclivity of the active methylene group present in compound 3 towards diazonium salts was also investigated. Thus, compound 3 could be readily coupled with aromatic diazonium salts to yield the corresponding aryl hydrazone derivatives 21a,b. The structure of 21 was established through microanalysis (IR, ¹H-NMR, and mass spectral data) and the chemical reactivity of this molecule toward some nucleophilic reagents. So, aryl hydrazononitriles 21a,b reacted with hydroxylamine hydrochloride in ethanolic sodium acetate and yielded amidooxime derivatives 22a,b that cyclized into amino triazole derivatives 23a,b by refluxing it in DMF/piperidine. The structures of 22 and 23 were supported based on their elemental analyses and spectral data. The IR spectrum of 22a as an example showed the appearance of absorption bands at 3,372–3,183 cm⁻¹ for hydroxyl and amino groups and 1,673 cm⁻¹ for the carbonyl group and the absence of the nitrile group in IR spectra confirms the addition process. The ¹H-NMR and mass spectra of the latter reaction products are all consistent with the proposed structures. However, analytical and spectral data of compound 23a,b are in agreement with the proposed structure (see experimental section). Further elucidation of the structure 21 came from the reaction with hydrazine hydrate as a binucleophilic reagent to give the amino pyrazole products 24. Analytical and spectral data of the product agree with the proposed structure [33,35,38,39,40] (Scheme 6).

Scheme 6

Synthesis of oxoacetohydrazonoyl cyanide, oxo-propanimidamide, triazole-4-carbohydrazide and pyrazol-3-amine derivatives (21–24).

In addition, the behavior of compound 3 toward diazonium salt of heterocyclic amine was also investigated. Thus, hydrazide–hydrazone derivative 3 coupled with diazonium salt derived from some heterocyclic amine to afford the respective hydrazone 25. The structure of the product 25 was determined from spectroscopic and elemental analyses. Bis(5-aamino-2-oxo-7-aryl-1,2,3,7-tetrahydropyrano[2,3-d]imidazole-6-carbohydrazide) derivatives 26a,b was synthesized in an excellent yield upon treatment of compound 3 with 5-benzylidene imidazolidine-2,4-dione in ethanol at reflux in the presence of piperidine. In a similar manner, cyclization of 2-(benzylidene cyclohexanones with substrate 3 under the same experimental conditions produced the corresponding bis(2-amino-4-Aryl-5,6,7,8-tetrahydro-4H-chromene-3-carbohydrazide) derivatives 27a,b. The formation of 27a,b can be understood in terms of the Michael-type addition of the active methylene group in substrate 3 to the activated double bond in the unsaturated cyclohexanone followed by cyclization of the resultant intermediate adducts. The structures of 26a,b and 27a,b were the final product and it were characterized with the help of its elemental analyses (IR, ¹H-NMR, and Mass spectra). The IR spectrum of compound 26a taken as a typical example of the series prepared revealed absorption bands in the region 3,421–3,200 cm⁻¹ assignable for the NH₂, in addition to the disappearance of the cyano function band. This suggests its participation in the cyclization process. Its ¹H-NMR spectrum displayed new a singlet signal at δ = 3.90 ppm assignable to the 4H-pyrane and a singlet signal at δ = 6.40 ppm assignable to the NH₂ protons. Its mass spectrum showed a molecular ion peak at m/z = 650 (M⁺) corresponding to a molecular formula C₃₂H₃₀N₁₀O₆ (Scheme 7).

Scheme 7

Synthesis of oxoacetohydrazonoyl cyanide, imidazole-6-carbohydrazide and chromene-3-carbohydrazide derivatives (25–27).

3 In vitro antimicrobial activity

The anti-microbial activity of the synthesized compounds was tested against a panel of two Gram-negative bacteria (Escherichia coli and Pseudomonas aeuroginosa) and two Gram-positive bacteria (Staphylococcus aureus and Bacillus subtilis). The anti-fungal activities of the compounds were tested against two fungi (Candida albicans and Aspergillus flavus). Each of the compounds was dissolved in DMSO, and a solution of the concentration 1 mg/mL was prepared separately paper discs of Whatman filter paper prepared with standard size (5 cm) were cut and sterilized in an autoclave. The paper discs soaked in the desired concentration of the complex solution were placed aseptically in the Petri dishes containing nutrient agar media (agar 20 g + beef extract 3 g + peptone 5 g) seeded with Escherichia coli, Pseudomonas aeuroginosa, Staphylococcus aureus, Bacillus subtilis, Candida albicans, and Aspergillus flavus. The Petri dishes were incubated at 36°C, and the inhibition zones were recorded after 24 h of incubation. Each treatment was replicated three times [41]. The antibacterial activity of a common standard antibiotic ampicillin and antifungal Clotrimazole was also recorded using the same procedure as above at the same concentration and solvents. The % activity index for the compound was calculated by the following formula.

% Activity Index = Zone of inhibition by test compound ( diameter ) Zone of inhibition by standard ( diameter ) × 100 .

Results in Table 1 indicated that all the synthesized compounds had an obvious inhibitory effect against all tested microbial species. The synthesized compound 3 was the more sensitive compound with the weakest anti-microbial activity and had a similar effect on fungi and bacteria in this study. Despite the Gram-positive bacteria (Staphylococcus aureus) showing an activity index of 66.7% and a diameter of inhibition zone (16 mm) in response to synthesized compound 3, the other Gram-positive bacteria (Bacillus subtilis) showed the largest activity index of 91.3% and the largest diameter of inhibition zone (21 mm). In the face of that, the two Gram-negative bacteria (Escherichia coli and Pseudomonas aeuroginosa) had lower values of activity index about 52 and 82.6%, respectively. Despite its distinctive protective outer layer. On the other hand, the fungus (Candida albicans) showed no resistance to the three synthesized compounds 5, 23a, 24a, which resulted in zero activity index and no inhibition zone. The synthesized compound 24a had the largest antimicrobial effect with a 0% activity index in all species except a 2 mm diameter inhibition zone in Gram-positive bacteria (Bacillus subtilis) and 6 mm in (Aspergillus flavus). Hence, the above results indicate that the synthesized compound 24a had the highest antimicrobial activity which all microbial species under study cannot resist followed by compound 23a and the lowest antimicrobial activity belonging to the synthesized compound 3. All other compounds had a similar medium effect on all microbes.

Table 1

Antibacterial and antifungal activities of synthesized compounds

Compound	E. coli		P. aeuroginosa		S. aureus		B. subtilis		C. Albicans		A. flavus
Compound	Diameter of inhibition zone (mm)	% Activity index	Diameter of inhibition zone (mm)	% Activity index	Diameter of inhibition zone (mm)	% Activity index	Diameter of inhibition zone (mm)	% Activity index	Diameter of inhibition zone (mm)	% Activity index	Diameter of inhibition zone (mm)	% Activity index
3	13	52	19	82.6	16	66.7	21	91.3	12	44.4	17	68
5	3	12	6	26.1	7	29.2	8	34.8	NA	—	4	16
7	2	8	5	21.7	4	16.7	7	30.4	3	11.1	9	36
9	4	16	8	34.8	9	37.5	10	43.5	13	48.1	18	72
11	7	28	10	43.5	11	45.8	13	56.5	9	33.3	12	48
12	9	36	16	69.6	14	58.3	19	82.6	12	44.4	15	60
15	6	24	10	43.5	10	41.7	11	47.8	6	22.2	11	44
17	7	28	13	56.5	12	50	15	65.2	8	29.6	12	48
19a	10	40	17	73.9	13	54.2	18	78.3	10	37	14	56
23a	NA	—	3	13	2	8.3	4	17.4	NA	—	2	8
24a	NA	—	NA	—	NA	—	2	8.7	NA	—	6	24
Amp.	25	100	23	100	24	100	23	100	NA	—	NA	—
Col.	NA	—	NA	—	NA	—	NA	—	27	100	25	100

NA → no activity.

It is worth noting that many human infections are caused by Gram-negative bacteria which have an outer membrane that protects the bacterial cell against any foreign substance, in the other hand Gram-positive bacteria did not have this outer layer so it can be easily affected and destroyed by different antibiotic and antimicrobial compounds. In this study, results indicated that all 11 synthesized compounds showed high microbial activity against Gram-negative, so it can penetrate its outer layer and inhibit its growth which helped in controlling them. Furthermore, Gram-positive bacteria had great resistance against all 11 synthesized compounds, and all these compounds had weak effect on this type of bacteria. Moreover, all these compounds had medium antimicrobial effect on fungi.

3.1 Minimum inhibitory concentration (MIC) measurement

The MIC was determined using the disc diffusion technique by preparing discs containing 1.9–1,000 µg/mL of each compound against Gram-negative Escherichia coli, Pseudomonas aeuroginosa and Gram-positive Staphylococcus aureus, Bacillus subtilis bacterial and Candida albicans, Aspergillus flavus as fungal and applying the protocol. The two-fold dilutions of the solution were prepared. The microorganism suspensions at 10 CFU/mL (colony forming unit/mL) concentrations were inoculated to the corresponding wells. The plates were incubated at 36°C for 24 h. for the bacteria. The standard antibiotic Ampicillin and Antifungal Clotrimazole were also recorded using the same procedure as above at the same concentration and solvents. At the end of the incubation period, the MIC values were recorded as the lowest concentration of the substance that had shown a clear zone [42,43]. Control experiments with DMSO and un-inoculated media were run parallel to the test compounds under the same condition.

Results in Table 2 indicated that synthesized compound 3 had the lowest concentration that can result in the largest inhibition zone and the highest activity index according to Table 1 followed by compounds 19a and 12, respectively, and clearly this made them the best compounds that can be used as a strong antimicrobial agent with the MIC. Furthermore, the synthesized compound 24a showed that it had no effect on the microbes under study and no clear zone appeared despite using a high concentration of it followed by compound 23a, so both of them cannot be used as antimicrobial agents as it had a very weak effect on microbes. All other synthesized compounds had a medium antimicrobial effect with a medium MIC.

Table 2

MIC for tested compounds

Compound	E. coli	P. aeuroginosa	S. aureus	B. subtilis	C. Albicans	A. flavus
3	1	1	2	1	4	1
5	64	64	32	32	NA	64
7	16	64	64	32	64	16
9	64	64	32	16	4	1
11	32	8	8	8	16	4
12	8	2	4	2	8	2
15	32	16	16	16	32	8
17	32	4	8	4	16	8
19a	2	2	4	4	8	4
23a	NA	32	16	64	NA	16
24a	NA	NA	NA	16	NA	32
Ampicillin	0.5	2	1	4	—	—
Clotrimazole	—	—	—	—	2	1

NA → no activity.

4 Conclusion

In this work, 1,3-cyclohexanedione 1 reacted with cyanoacetic acid hydrazide to afford bis(2-cyanoaceto hydrazide) derivative 3. Cyanoacetic acid hydrazide and its analogues are especially important starting materials for the synthesis of new biologically heterocyclic compounds. Our research deals with the effective use of bis(2-cyanoaceto hydrazide) derivative 3 in the synthesis of a variety of polyfunctionally azoles and (or) azines with biological interest.

5 Experimental

The melting points, the elemental analyses, and the spectral data were recorded as reported in reference [44].

5.1 General synthesis of 2-cyano-N′-(3-(2-(2-cyanoacetyl)hydrazinyl)cyclohex-2-en-1-ylidene)acetohydrazide (3)

To a stirred solution of 2-cyanoaceto acid, hydrazide 2 (0.02 mol, 5.48 g) in glacial acetic acid (10 mL)/ethanol (30 mL) and 1,3-cyclohexadione 1 (0.01 mol, 1.12 g) was added dropwise from 5 to 10 min, and reaction mixture was stirred at room temperature for about 12 h The separated solid was filtered off washed with ethanol, dried, and recrystallized from ethanol to give (3) as pale yellow crystals: M.P.: 230–232°C, yield: 2.65 g (96%). IR (KBr) (v _max/cm⁻¹): 3,447–3,224 (3NH), 2,964–2,903 (CH-aliphatic), 2,261 (CN), 1,678 (CO). MS (EI, 70 eV): m/z (%) = 275 (M⁺ + 1). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.72–1.91 (m, 2H, CH₂), 2.23–2.32 (m, 4H, 2CH₂), 3.36 (s, 2H, CH₂), 3.95 (s, 2H, CH₂), 5.54 (s, 1H, CH-cyclohexene), 8.52 (hump, 1H, NH), 9.96 (s, 1H, NH), 10.38 (s, 1H, NH). ¹³C NMR (100 MHz, DMSO-d ₆) δ 21.86, 24.70, 26.52, 26.63, 31.45, 86.93, 116.66, 116.88, 156.79, 160.20, 162.12, 164.67. Anal. Calcd. for C₁₂H₁₄N₆O₂ (274): C, 52.55; H, 5.15; N, 30.64. Found: C, 52.62; H, 5.20; N, 30.70.

5.2 Synthesis of 2-cyano-N′-(3-(2-(2-cyano-3-(dimethylamino)acryloyl)hydrazinyl)-cyclohex-2-en-1-ylidene)-3-(dimethylamino)acrylohydrazide (4)

A mixture of 3 (0.01 mol, 2.74 g) and DMF-DMA (0.02 mol) in dioxane (30 mL) was heated under reflux for 6 h after cooling, The separated solid was filtered off, dried, and recrystallized from ethanol to give (3) as pale brown crystals: M.P.: 142–144°C, yield: 3.25 g (84%). IR (KBr) (v _max/cm⁻¹): 3,381–3,193 (NH), 2,93–2,870 (CH-aliphatic), 2,203 (CN), 1,619 (CO). MS (EI, 70 eV): m/z (%) = 384 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.77–1.80 (m, 2H, CH₂), 2.08–2.89 (m, 4H, 2CH₂), 3.10 (s, 6H, N(CH₃)₂), 3.13 (s, 6H, N(CH₃)₂), 5.15 (s, 1H, CH-cyclohexene), 7.51 (s, 1H, CH-Oleffinic), 7.72 (s, 1H, CH-Oleffinic), 8.34 (s, 1H, NH), 8.90 (s, 1H, NH), 11.45 (hump, 1H, NH). Anal. Calcd. for C₁₈H₂₄N₈O₂ (384): C, 56.24; H, 6.29; N, 29.15. Found: C, 56.30; H, 6.34; N, 29.21.

5.3 Synthesis of 3-amino-N′-(3-(2-(3-amino-1H-pyrazole-4-carbonyl)hydrazinyl)cyclohex-2-en-1-ylidene)-1H-pyrazole-4-carbohydrazide (5)

In ethanol (30 mL) compound 4 (0.5 g) and hydrazine, hydrate (3 mL) was refluxed for 12 h. Upon reaction completion, the reaction mixture was allowed to cool and poured into water/ice. The separated solid was collected, washed well with water, and recrystallized from the ideal solvent to afford (5) as brown crystals from ethanol: M.P.: 240–242°C, yield: 2.25 g (63%). IR (KBr) (v _max/cm⁻¹): 3,400–3,163 (NH₂/NH), 2,957 (CH-aliphatic), 1,617 (CO). MS (EI, 70 eV): m/z (%) = 358 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 0.84–1.99 (m, 2H, CH₂), 2.10–2.93 (m, 4H, 2CH₂), 4.70 (s, 1H, CH-cyclohexene), 6.99 (s, 4H, 2NH₂), 7.07 (s, 2H, CH-pyrazole), 7.20 (s, 2H, 2NH), 8.33–8.47 (hump, 3H, 3NH). Anal. Calcd. for C₁₄H₁₈N₁₀O₂ (358): C, 46.92; H, 5.06; N, 39.09. Found: C, 46.97; H, 5.13; N, 39.15.

5.4 Synthesis of 2-(4-hydroxythiazol-2-yl)-N′-(3-(2-(2-(4-hydroxythiazol-2-yl)acetyl)-hydrazinyl)-cyclohex-2-en-1-ylidene)acetohydrazide (7)

Thioglycolic acid (0.02 mol) and compound 3 (0.01 mol, 2.74 g) were dissolved in DMF (20 mL), and anhydrous ZnCl₂ (0.5 g) was added as a catalyst. The reaction mixture was heated under reflux with stirring for about 10 h. Upon reaction completion, the reaction mixture was allowed to cool, poured into water/ice, and triturated with an excess of 10% sodium bicarbonate solution. The solid product obtained was recrystallized from ethanol to give (7) as brown crystals: M.P.: 152–154°C, yield: 2.25 g (53%). IR (KBr) (v _max/cm⁻¹): 3,403–3,200 (OH/NH), 2,936 (CH-aliphatic), 1,620 (CO). MS (EI, 70 eV): m/z (%) = 424 (M⁺ + 2). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 0.8–1.91 (m, 2H, CH₂), 2.09–2.99 (m, 4H, 2CH₂), 3.18 (s, 2H, CH₂), 3.37 (s, 2H, CH₂), 4.50 (s, 1H, CH-cyclohexene), 7.32 (s, 2H, CH-thiazole), 8.18 (s, 1H, NH), 8.23 (s, 1H, NH), 8.35 (s, 1H, NH), 11.50 (hump, 2H, 2OH). ¹³C NMR (100 MHz, DMSO-d ₆) δ 21.90, 24.81, 34.90, 39.31, 39.52, 88.60, 128.92, 128.92, 144.73, 144.73, 157.0, 160.10, 162.20, 166.10, 171.84, 171.84. Anal. Calcd. for C₁₆H₁₈N₆O₄S₂ (422): C, 45.49; H, 4.29; N, 19.89. Found: C, 45.55; H, 4.34; N, 19.95.

5.5 Synthesis of 2-cyano-N′-(3-(2-(2-cyano-3-phenylacryloyl)hydrazinyl)cyclohex-2-en-1-ylidene)-3-phenylacrylohydrazide (8)

A solution of 3 (0.01 mol, 2.74 g) in ethanol (30 mL) was treated with (0.02 mol, 2.12 g) of benzaldehyde and a few drops of piperidine. The reaction mixture was heated under reflux for 6 h. The reaction mixture was left to cool at room temperature and poured into ice/water containing a few drops of hydrochloric acid, and the formed solids were collected by filtration and recrystallized from ethanol to give (8) as red crystals: M.P.: 160–162°C, yield: 3.85 g (85%). IR (KBr) (v _max/cm⁻¹): 3,423–3,400 (NH), 3,063 (CH-aromatic), 2,936 (CH-aliphatic), 2,191 (CN), 1,626 (CO). MS (EI, 70 eV): m/z (%) = 451 (M⁺ + 1). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.05–1.65 (m, 2H, CH₂), 2.50–2.99 (m, 4H, 2CH₂), 4.77 (s, 1H, CH-cyclohexene), 7.31–7.93 (m, 12H, CH-aromatic and CH-Oleffinic), 9.71 (s, 1H, NH), 9.93 (s, 1H, NH), 10.03 (s, 1H, NH). ¹³C NMR (100 MHz, DMSO-d ₆) δ 22.65, 25.10, 32.40, 106.0, 106.0, 108.0, 118.60, 118.60, 128.20, 128.50, 128.50, 129.64, 129.64, 129.64, 129.64, 130.47, 130.47, 130.47, 130.47, 132.10, 132.10, 136.10, 136.10, 156.10, 159.50, 168.70. Anal. Calcd. for C₂₆H₂₂N₆O₂ (450): C, 69.32; H, 4.92; N, 18.66. Found: C, 69.40; H, 4.97; N, 18.72.

5.6 Synthesis of 3-amino-N′-(3-(2-(3-amino-5-phenyl-1H-pyrazole-4-carbonyl)hydrazinyl)-cyclohex-2-en-1-ylidene)-5-phenyl-1H-pyrazole-4-carbohydrazide (9)

A mixture of 8 (0.5 g) and hydrazine hydrate (10 mL) was fused for 6 h after cooling and poured into ice/water containing a few drops of hydrochloric acid. The formed solid was filtered, washed with water, and recrystallized from ethanol to give (9) brown crystals: M.P.: 120–122°C, yield: 4.25 g (83%). IR (KBr) (v _max/cm⁻¹): 3,506–3,248 (NH₂/NH), 3,055 (CH-aromatic), 2,958–2,889 (CH-aliphatic), 1,625 (CO). MS (EI, 70 eV): m/z (%) = 510 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 0.75–1.83 (m, 2H, CH₂), 2.20–2.45 (m, 4H, 2CH₂), 4.70 (s, 1H, CH-cyclohexene), 5.90 (s, 4H, 2NH₂), 7.74–7.79 (m, 10H, CH-aromatic), 8.40 (s, 1H, NH), 8.40 (s, 1H, NH), 10.30 (hump, 1H, NH), 12.50 (hump, 2H, 2NH). Anal. Calcd. for C₂₆H₂₆N₁₀O₂ (510): C, 61.17; H, 5.13; N, 27.43. Found: C, 61.22; H, 5.19; N, 27.50.

5.7 Synthesis of 2-cyano-N′-(3-(2-(2-cyano-3-(2-hydroxyphenyl)acryloyl)hydrazinyl)-cyclohex-2-en-1-ylidene)-3-(2-hydroxyphenyl)acrylohydrazide (10)

A mixture of compound 3 (0.01 mol, 2.74 g), salicyladehyde (0.02 mol) and piperidine (3 drops) in ethanol (30 mL) was heated under reflux for 12 h after cooling and poured into ice/water containing a few drops of hydrochloric acid. The resulting precipitate was filtered off, dried, and recrystallized from ethanol to give (10) as orange crystals: M.P.: 170–172°C, yield: 3.75 g (77%). IR (KBr) (v _max/cm⁻¹): 3,420–3,380 (OH/NH), 2,936 (CH-aliphatic), 2,222 (CN), 1,609 (CO). MS (EI, 70 eV): m/z (%) = 482 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.23–1.91 (m, 2H, CH₂), 2.09–2.80 (m, 4H, 2CH₂), 4.60 (s, 1H, CH-cyclohexene), 6.99–7.17 (m, 10H, CH-aromatic and CH-Oleffinic), 8.03 (s, 1H, NH), 8.05 (s, 1H, NH), 10.00 (s, 2H, 2OH), 10.20 (hump, 1H, NH). Anal. Calcd. for C₂₆H₂₂N₆O₄ (482): C, 64.72; H, 4.60; N, 17.42. Found: C, 64.79; H, 4.65; N, 17.47.

5.8 Synthesis of 2-imino-N′-(3-(2-(2-imino-2H-chromene-3-carbonyl)hydrazinyl)cyclohex-2-en-1-ylidene)-2H-chromene-3-carbohydrazide (11)

A solution of compound 10 (0.5 g) in dioxane (10 mL) containing a few drops of piperidine was heated under reflux for 24 h. The reaction mixture was allowed to cool. The separated solid was filtered off washed with water, dried, and recrystallized from ethanol to give (11) red crystals: M.P.: 218–220°C, yield: 4.25 g (88%). IR (KBr) (v _max/cm⁻¹): 3,423–3,217 (NH), 3,050 (CH-aromatic), 2,934–2,837 (CH-aliphatic), 1,686 (CO). MS (EI, 70 eV): m/z (%) = 483 (M⁺ + 1). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.22–1.92 (m, 2H, CH₂), 2.20–2.80 (m, 4H, 2CH₂), 4.18 (s, 1H, CH-cyclohexene), 6.99–7.96 (m, 8H, CH-aromatic), 8.03 (s, 2H, CH-pyrane), 8.60 (s, 1H, NH), 9.87 (s, 1H, NH), 10.51 (s, 2H, 2NH), 11.67 (s, 1H, NH). ¹³C NMR (100 MHz, DMSO-d ₆) δ 22.47, 25.9, 27.3, 98.3, 114.3, 114.3, 115.1, 115.1, 118.3, 118.3, 123.5, 123.5, 129.7, 129.7, 129.9, 129.9, 131.4, 131.4, 155.2, 155.2, 157.2, 161.5, 161.5, 162.4, 167.2, 170.4. Anal. Calcd. for C₂₆H₂₂N₆O₄ (482): C, 64.72; H, 4.60; N, 17.42. Found: C, 64.78; H, 4.66; N, 17.50.

5.9 General procedure for the synthesis of (12,13)

To a solution of compound 3 (0.01 mol, 2.74 g), malononitrile (0.02 mol, 1.32 g) and (or) acetylacetone (0.02 mol) respectively in ethanol (20 mL) containing a few drops of piperidine was added, and the reaction mixture was heated under reflux for 12 h then poured into ice/water containing a few drops of hydrochloric acid. The formed solid product was collected by filtration, dried, and recrystallized from ethanol to give 12,13.

5.9.1 4,6-Diamino-1-((3-((4,6-diamino-3-cyano-2-oxopyridin-1(2H)-yl)amino)cyclohex-2-en-1-ylidene)amino)-2-oxo-1,2-dihydropyridine-3-carbonitrile (12)

It was obtained as brown crystals: M.P.: 180–182°C, yield: 3.65 g (89%). IR (KBr) (v _max/cm⁻¹): 3,448–3,330 (NH₂/NH), 2,950–2,865 (CH-aliphatic), 2,196 (CN), 1,631 (CO). MS (EI, 70 eV): m/z (%) = 406 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.61–1.83 (m, 2H, CH₂), 2.09–2.59 (m, 4H, 2CH₂), 3.56 (s, 1H, CH-cyclohexene), 5.77 (s, 4H, 2NH₂), 6.20 (s, 4H, 2NH₂), 6.88 (s, 2H, CH-pyridine), 8.50 (s, 1H, NH). ¹³C NMR (100 MHz, DMSO-d ₆) δ 22.69, 24.86, 33.70, 44.20, 44.20, 55.49, 55.49, 98.15, 114.45, 114.45, 128.50, 128.50, 157.20, 163.10, 168.60, 168.60, 183.80, 183.80. Anal. Calcd. for C₁₈H₁₈N₁₀O₂ (406): C, 53.20; H, 4.46; N, 34.47. Found: C, 53.27; H, 4.52; N, 34.54.

5.9.2 1-((3-((3-Cyano-4,6-dimethyl-2-oxopyridin-1(2H)-yl)amino)cyclohex-2-en-1-ylidene)-amino)-4,6-dimethyl-2-oxo-1,2-dihydropyridine-3-carbonitrile (13)

It was obtained as brown crystals: M.P.: 218–220°C, yield: 3.50 g (87%). IR (KBr) (v _max/cm⁻¹): 3,448 (NH), 3,017 (CH-aromatic), 2,959–2,838 (CH-aliphatic), 2,220 (CN), 1,627 (CO). MS (EI, 70 eV): m/z (%) = 402 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.55–1.91 (m, 2H, CH₂), 2.23 (s, 6H, 2CH₃), 2.31 (s, 6H, 2CH₃), 2.37–-2.96 (m, 4H, 2CH₂), 4.38 (s, 1H, CH-cyclohexene), 6.20 (s, 2H, CH-pyridine), 9.05 (s, 1H, NH). ¹³C NMR (100 MHz, DMSO-d ₆) δ 19.36, 19.36, 20.67, 20.67, 22.47, 25.72, 36.93, 99.50, 108.35, 108.35, 109.21, 109.21, 116.56, 116.56, 132.20, 132.20, 155.49, 155.49, 160.93, 161.44, 161.44, 196.52. Anal. Calcd. for C₂₂H₂₂N₆O₂ (402): C, 65.66; H, 5.51; N, 20.88. Found: C, 65.73; H, 5.57; N, 20.95.

5.10 Synthesis of 1-((3-((5-cyano-4-methyl-2,6-dioxo-3,6-dihydropyridin-1(2H)-yl)amino)-cyclohex-2-en-1-ylidene)amino)-4-methyl-2,6-dioxo-1,2,5,6-tetrahydro-pyridine-3-carbonitrile (14)

A mixture of compound 3 (0.01 mol, 2.74 g) and ethyl acetoacetate (0.02 mol) was fused for 12 h at 150°C. The reaction mixture was allowed to cool and then triturated with ethanol. The separated solid was filtered off washed with water, dried, and recrystallized from ethanol to give (14) as brown crystals: M.P.: 170–172°C, yield: 3.50 g (87%). IR (KBr) (v _max/cm⁻¹): 3,426 (NH), 2,980–2,935 (CH-aliphatic), 2,223 (CN), 1,727 (CO). MS (EI, 70 eV): m/z (%) = 402 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 0.90–1.70 (m, 2H, CH₂), 2.23 (s, 6H, 2CH₃), 2.27–2.70 (m, 4H, 2CH₂), 2.78 (s, 4H, 2CH₂), 5.98 (s, 1H, CH-cyclohexene), 7.28 (s, 1H, NH). Anal. Calcd. for C₂₀H₁₈N₆O₄ (406): C, 59.11; H, 4.46; N, 20.68. Found: C, 59.19; H, 4.53; N, 20.74.

5.11 Synthesis of 3,5-diamino-4-cyano-N′-(3-(2-(3,5-diamino-4-cyanothiophene-2-carbonyl)-hydrazinyl)cyclohex-2-en-1-ylidene)thiophene-2-carbohydrazide (15)

To a solution of compound 3 (0.01 mol, 2.74 g) in ethanol (20 mL) containing TEA (3 drops), elemental sulfur (0.02 mol, 0.64 g) was added followed by malononitrile (0.02 mol, 1.32 g). The reaction mixture was heated under reflux for 10 h and then poured into ice/water containing a few drops of hydrochloric acid; the formed solid product was collected by filtration, dried, and recrystallized from ethanol to give (15) as pale brown crystals: M.P.: 215–217°C, yield: 3.90 g (83%). IR (KBr) (v _max/cm⁻¹): 3,423–3,197 (NH₂/NH), 2,927–2,856 (CH-aliphatic), 2,203 (CN), 1,628 (CO). MS (EI, 70 eV): m/z (%) = 472 (M⁺ + 2). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 0.78–1.80 (m, 2H, CH₂), 2.08–2.46 (m, 4H, 2CH₂), 5.76 (s, 1H, CH-cyclohexene), 6.10 (s, 4H, 2NH₂), 6.60 (s, 4H, 2NH₂), 7.31 (s, 1H, NH), 7.34 (s, 1H, NH), 11.78 (s, 1H, NH). ¹³C NMR (100 MHz, DMSO-d ₆) δ 22.43, 25.7, 26.3, 83.2, 83.2, 99.8, 118.2, 118.2, 134.4, 158.3, 158.3, 158.7, 158.7, 160, 163.6, 163.7, 163.7, 170.2. Anal. Calcd. for C₁₈H₁₈N₁₀O₂S₂ (470): C, 45.95; H, 3.86; N, 29.77. Found: C, 46.01; H, 3.94; N, 29.84.

5.12 Synthesis of 4-amino-N′-(3-(2-(4-amino-3-phenyl-2-thioxo-2,3-dihydrothiazole-5-carbonyl)-hydrazinyl)cyclohex-2-en-1-ylidene)-3-phenyl-2-thioxo-2,3-dihydro-thiazole-5-carbohydrazide (16)

To a solution of compound 3 (0.01 mol, 2.74 g) in ethanol (20 mL) containing TEA (3 drops), elemental sulfur (0.02 mol, 0.64 g) was added followed by phenyl isothiocyanate (0.02 mol). The reaction mixture was heated under reflux for 12 h and then poured into ice/water containing a few drops of hydrochloric acid; the formed solid product was collected by filtration, dried, and recrystallized from ethanol to give (16) as brown crystals: M.P.: 148–150°C, yield: 4.95 g (81%). IR (KBr) (v _max/cm⁻¹): 3,423–3,250 (NH₂/NH), 2,936–2,858(CH-aliphatic), 1,628 (CO). MS (EI, 70 eV): m/z (%) = 610 (M⁺ + 2). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 0.84–1.91 (m, 2H, CH₂), 2.09–2.90 (m, 4H, 2CH₂), 4.49 (s, 1H, CH-cyclohexene), 5.70 (s, 4H, 2NH₂), 7.31–7–65 (m, 10H, CH-aromatic), 8.30 (hump, 1H, NH), 9.20 (s, 1H, NH), 11.20 (hump, 1H, NH). Anal. Calcd. for C₂₆H₂₄N₈O₂S₄ (608): C, 51.30; H, 3.97; N, 18.41. Found: C, 51.37; H, 4.05; N, 18.48.

5.13 Synthesis of 2-amino-N′-(3-(2-(2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carbonyl)-hydrazinyl)cyclohex-2-en-1-ylidene)-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carbohydrazide (17)

To a solution of compound 3 (0.01 mol, 2.74 g) in ethanol (20 mL) containing TEA (3 drops), elemental sulfur (0.02 mol, 0.64 g) was added followed by cyclohexanone (0.02 mol). The reaction mixture was heated under reflux for 24 h and then poured into ice/water containing a few drops of hydrochloric acid and the formed solid product was collected by filtration, dried, and recrystallized from ethanol to give (17) as brown crystals: M.P.: 110–112°C, yield: 4.25 g (85%). IR (KBr) (v _max/cm⁻¹): 3,420–3,196 (NH₂/NH), 2,931–2,857 (CH-aliphatic), 1,624 (CO). MS (EI, 70 eV): m/z (%) = 500 (M⁺ + 2). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 0.84–1.91 (m, 10H, 5CH₂), 2.09–2.90 (m, 12H, 6CH₂), 4.49 (s, 1H, CH-cyclohexene), 6.92 (s, 4H, 2NH₂), 7.23 (s, 1H, NH), 7.31 (s, 1H, NH), 7.32 (s, 1H, NH). Anal. Calcd. for C₂₄H₃₀N₆O₂S₂ (498): C, 57.81; H, 6.06; N, 16.85. Found: C, 57.87; H, 6.13; N, 16.92.

5.14 Synthesis of 2-amino-N′-(3-(2-(2-amino-4-cyano-5-methyl-1H-pyrrole-3-carbonyl)-hydrazinyl)-cyclohex-2-en-1-ylidene)-4-cyano-5-methyl-1H-pyrrole-3-carbohydrazide (18)

A mixture of compound 3 (0.01 mol, 2.74 g) and acetonitrile dimer (0.02 mol) in ethanol (20 mL) containing a few drops of piperidine was refluxed for 6 h. The reaction mixture was cooled, and the solid obtained was filtered off and recrystallized from ethanol to give (18) as brown crystals: M.P.: 190–192°C, yield: 3.70 g (85%). IR (KBr) (v _max/cm⁻¹): 3,450–3,207 (NH₂/NH), 2,931–2,863 (CH-aliphatic), 2,200 (CN), 1,641 (CO). MS (EI, 70 eV): m/z (%) = 434 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.19–1.98 (m, 2H, CH₂), 2.20 (s, 6H, 2CH₃), 2.22–2.69 (m, 4H, 2CH₂), 4.08 (s, 1H, CH-cyclohexene), 6.39 (s, 4H, 2NH₂), 7.31 (s, 1H, NH), 7.60 (s, 1H, NH), 9.40 (s, 1H, NH), 13.00 (s, 2H, 2NH). Anal. Calcd. for C₂₀H₂₂N₁₀O₂ (434): C, 55.29; H, 5.10; N, 32.24. Found: C, 55.36; H, 5.16; N, 32.31.

5.15 General procedure for the synthesis of (19a,b)

A mixture of 3 (0.01 mol, 2.74 g) and 3-phenyl-1-phenylprop-2-en-1-one, 3-(4-chlorophenyl)-1-phenylprop-2-en-1-one (0.02 mol) respectively in ethanol (30 mL) with a catalytic amount of piperidine was heated under reflux for 6 h. The reaction mixture was allowed to cool and poured into crushed ice then acidified with HCl. The separated solid was filtered off washed with water, dried, and recrystallized from the ideal solvent to give 19a,b.

5.15.1 2-Amino-N′-(3-(2-(2-amino-4,6-diphenyl-4H-pyran-3-carbonyl)hydrazinyl)-cyclo-hex-2-en-1-ylidene)-4,6-diphenyl-4H-pyran-3-carbohydrazide (19a)

It was obtained as brown crystals from ethanol: M.P.: 212–214°C, yield: 5.20 g (75%). IR (KBr) (v _max/cm⁻¹): 3,416–3,220 (NH₂/NH), 2,937–2,860 (CH-aliphatic), 1,629 (CO). MS (EI, 70 eV): m/z (%) = 690 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.59–1.91 (m, 2H, CH₂), 2.20–2.80 (m, 4H, 2CH₂), 3.66 (d, 2H, 4H-pyrane), 4.60 (s, 1H, CH-cyclohexene), 5.71 (d, 2H, = CH-pyrane), 7.48–7.53 (m, 24H, CH-aromatic and 2NH₂), 7.86 (s, 1H, NH), 8.21 (s, 1H, NH), 10.00 (hump, 1H, 1NH). ¹³C NMR (100 MHz, DMSO-d ₆) δ 22.59, 25.80, 30.11, 31.48, 44.05, 44.05, 88.30, 88.30, 99.50, 99.50, 113.24, 127.10, 127.10, 127.10, 127.10, 127.73, 127.73, 128.38, 128.38, 128.38, 128.38, 128.57, 128.57, 128.57, 128.57, 129.23, 129.23, 129.23, 129.23, 129.86, 129.86, 130.11, 139.25, 139.25, 148.50, 148.50, 156.96, 160.10, 162.90, 166.30, 168.70, 192.90. Anal. Calcd. for C₄₂H₃₈N₆O₄ (690): C, 73.03; H, 5.54; N, 12.17. Found: C, 73.10; H, 5.60; N, 12.25.

5.15.2 2-Amino-N′-(3-(2-(2-amino-4-(4-chlorophenyl)-6-phenyl-4H-pyran-3-carbonyl)-hydr-azinyl)cyclohex-2-en-1-ylidene)-4-(4-chlorophenyl)-6-phenyl-4H-pyran-3-carbohydr-azide (19b)

It was obtained as pale brown crystals from ethanol: M.P.: 160–162°C, yield: 4.80 g (63%). IR (KBr) (v _max/cm⁻¹): 3,423–3,200 (NH₂/NH), 3,026 (CH-aromatic), 2,939–2,859 (CH-aliphatic), 1,633 (CO). MS (EI, 70 eV): m/z (%) = 760 (M⁺ + 2). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.20–1.59 (m, 2H, CH₂), 2.00–2.48 (m, 4H, 2CH₂), 3.68 (d, 2H, 4H-pyrane), 4.60 (s, 1H, CH-cyclohexene), 5.70 (d, 2H, = CH-pyrane), 7.44–7.61 (m, 22H, CH-aromatic and 2NH₂), 7.87 (s, 1H, NH), 8.20 (s, 1H, NH), 10.00 (hump, 1H, 1NH). Anal. Calcd. for C₄₂H₃₆Cl₂N₆O₄ (758): C, 66.40; H, 4.78; N, 11.06. Found: C, 66.47; H, 4.85; N, 11.12.

5.16 General procedure for the synthesis of (20a,b)

To a solution of 3 (0.01 mol, 2.74 g) in ethanol (30 mL) and 2-(4-chlorobenzylidene)malononitrile, 2-(4-methoxybenzylidene)malononitrile (0.02 mol) respectively were added. The reaction mixture was treated with a few drops of piperidine, and then refluxed for 6 h. The reaction mixture was allowed to cool and poured into crushed ice then acidified with HCl. The separated solid was filtered off washed with water, dried and crystallized from the proper solvent to give 20a,b.

5.16.1 6-Amino-1-((3-((6-amino-4-(4-chlorophenyl)-3,5-dicyano-2-oxopyridin-1(2H)-yl)-amino)-cyclohex-2-en-1-ylidene)amino)-4-(4-chlorophenyl)-2-oxo-1,2-dihydro-pyridine-3,5-dicarbonitrile (20a)

It was obtained as orange crystals from ethanol: M.P.: 142–144°C, yield: 4.30 g (66%). IR (KBr) (v _max/cm⁻¹): 3,419–3,200 (NH₂/NH), 2,939–2,861 (CH-aliphatic), 2,211 (CN), 1,632 (CO). MS (EI, 70 eV): m/z (%) = 648 (M⁺ + 2). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.77–1.85 (m, 2H, CH₂), 2.21–2.78 (m, 4H, 2CH₂), 4.70 (s, 1H, CH-cyclohexene), 5.70 (s, 4H, 2NH₂), 7.05–7.63 (m, 8H, CH-aromatic), 10.30 (s, 1H, NH). ¹³C NMR (100 MHz, DMSO-d ₆) δ 23.4, 26.2, 28.3, 77.9, 77.9, 101, 117.3, 117.3, 117.8, 117.8, 117.8, 117.8, 129.8, 129.8, 129.8, 129.8, 131, 131, 131, 131, 131.8, 131.8, 135.5, 135.5, 160.2, 160.2, 160.8, 161.3, 161.7, 161.7, 171.3, 171.3. Anal. Calcd. for C₃₂H₂₀Cl₂N₁₀O₂ (646): C, 59.36; H, 3.11; N, 21.63. Found: C, 59.44; H, 3.17; N, 21.70.

5.16.2 6-Amino-1-((3-((6-amino-3,5-dicyano-4-(4-methoxyphenyl)-2-oxopyridin-1(2H)-yl)-amino)cyclohex-2-en-1-ylidene)amino)-4-(4-methoxyphenyl)-2-oxo-1,2-dihydro-pyridine-3,5-dicarbonitrile (20b)

It was obtained as pale yellow crystals from ethanol: M.P.: 170–172°C, yield: 4.90 g (76%). IR (KBr) (v _max/cm⁻¹): 3,422–3,224 (NH₂/NH), 2,938–2,860 (CH-aliphatic), 2,213 (CN), 1,636 (CO). MS (EI, 70 eV): m/z (%) = 638 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 0.84–1.86 (m, 2H, CH₂), 2.09–2.75 (m, 4H, 2CH₂), 3.85 (s, 6H, 2OCH₃), 4.60 (s, 1H, CH-cyclohexene), 5.71 (s, 4H, 2NH₂), 6.82–7.47 (m, 8H, CH-aromatic), 10.30 (s, 1H, NH). Anal. Calcd. for C₃₄H₂₆N₁₀O₄ (638): C, 63.94; H, 4.10; N, 21.93. Found: C, 64.00; H, 4.18; N, 21.99.

5.17 General procedure for the synthesis of (21a,b)

To a cold solution of 3 (0.01 mol, 2.74 g) in ethanol (20 mL) containing sodium acetate (3 g) was added with continuous stirring either of the appropriately substituted bezenediazonium salt (0.02 mol) [prepared by adding sodium nitrite (0.02 mol) in water to a cold solution of either of the appropriate substitute aniline in the appropriate amount of hydrochloric acid]. The reaction mixture was stirred for 1 h and the formed solid product, in each case, was collected by filtration, dried, and recrystallized from the proper solvent to give 21a,b.

5.17.1 N-(4-Chlorophenyl)-2-(2-(3-(2-(2-(2-(4-chlorophenyl)hydrazono)-2-cyanoacetyl)-hydrazinyl)-cyclohex-2-en-1-ylidene)hydrazinyl)-2-oxoacetohydrazonoyl cyanide (21a)

It was obtained as pale red crystals from ethanol: M.P.: 182–184°C, yield: 3.20 g (58%). IR (KBr) (v _max/cm⁻¹): 3,449–3,400 (NH), 3,058 (CH-aromatic), 2,992 (CH-aliphatic), 2,217 (CN), 1,656 (CO). MS (EI, 70 eV): m/z (%) = 552 (M⁺ + 2). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.23–1.91 (m, 2H, CH₂), 2.51–2.75 (m, 4H, 2CH₂), 4.49 (s, 1H, CH-cyclohexene), 6.88–7.58 (m, 8H, CH-aromatic), 7.80 (s, 1H, NH), 8.20 (s, 1H, NH), 11.63 (s, 1H, NH), 11.80 (s, 2H, 2NH). Anal. Calcd. for C₂₄H₂₀Cl₂N₁₀O₂ (550): C, 52.28; H, 3.66; N, 25.40. Found: C, 52.35; H, 3.72; N, 25.47.

5.17.2 2-(2-(3-(2-(2-Cyano-2-(2-(4-methoxyphenyl)hydrazono)acetyl)hydrazinyl)cyclohex-2-en-1-ylidene)hydrazinyl)-N-(4-methoxyphenyl)-2-oxoacetohydrazonoyl cyanide (21b)

It was obtained as red crystals from ethanol: M.P.: 160–162°C, yield: 4.20 g (77%). IR (KBr) (v _max/cm⁻¹): 3,449–3,200 (NH), 2,937–2,837 (CH-aliphatic), 2,213 (CN), 1,662 (CO). MS (EI, 70 eV): m/z (%) = 542 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.23–1.91 (m, 2H, CH₂), 2.60–3.00 (m, 4H, 2CH₂), 3.85 (s, 6H, 2OCH₃), 4.18 (s, 1H, CH-cyclohexene), 6.70–7.36 (m, 8H, CH-aromatic), 7.38 (s, 1H, NH), 8.30 (s, 1H, NH), 10.80 (hump, 1H, NH), 13.20 (s, 2H, 2NH). Anal. Calcd. for C₂₆H₂₆N₁₀O₄ (542): C, 57.56; H, 4.83; N, 25.82. Found: C, 57.63; H, 4.90; N, 25.90.

5.18 General procedure for the synthesis of (22a,b)

A mixture of compounds 21a,b (0.01 mol) and hydroxylamine hydrochloride (0.02 mol) in ethanol (20 mL), sodium acetate (2 g) and water (3 drops) was heated at reflux for 12 h. The reaction mixture was allowed to cool and poured into crushed ice. The separated solid was filtered off washed with water, dried, and recrystallized from the proper solvent to give 22a,b.

5.18.1 3-(2-(3-(2-(3-Amino-2-(2-(4-chlorophenyl)hydrazono)-3-(hydroxyimino)propanoyl)-hydrazinyl)-cyclohex-2-en-1-ylidene)hydrazinyl)-2-(2-(4-chlorophenyl)hydrazono)-N′-hydroxy-3-oxo-propanimidamide (22a)

It was obtained as pale brown crystals from ethanol: M.P.: 148–150°C, yield: 3.90 g (63%). IR (KBr) (v _max/cm⁻¹): 3,372–3,183 (OH/NH₂/NH), 2,928–2,860 (CH-aliphatic), 1,673 (CO). MS (EI, 70 eV): m/z (%) = 618 (M⁺ + 2). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.24–1.65 (m, 2H, CH₂), 2.51–2.64 (m, 4H, 2CH₂), 3.34 (s, 1H, CH-cyclohexene), 5.87 (s, 2H, 2OH), 7.14–7.60 (m, 12H, CH-aromatic and 2NH₂), 10.54 (s, 1H, NH), 11.18 (s, 1H, NH), 11.81 (s, 1H, NH), 12.81 (s, 2H, 2NH). Anal. Calcd. for C₂₄H₂₆Cl₂N₁₂O₄ (616): C, 46.69; H, 4.24; N, 27.22. Found: C, 46.75; H, 4.30; N, 27.29.

5.18.2 3-(2-(3-(2-(3-Amino-3-(hydroxyimino)-2-(2-(4-methoxyphenyl)hydrazono)propanoyl)-hydr-azinyl)cyclohex-2-en-1-ylidene)hydrazinyl)-N′-hydroxy-2-(2-(4-methoxyphenyl)-hydrazono)-3-oxopropanimidamide (22b)

It was obtained as red crystals from ethanol: M.P.: 170–172°C, yield: 4.40 g (72%). IR (KBr) (v _max/cm⁻¹): 3,423–3,205 (OH/NH₂/NH), 2,935–2,836 (CH-aliphatic), 1,639 (CO). MS (EI, 70 eV): m/z (%) = 608 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.24–1.64 (m, 2H, CH₂), 2.51–2.64 (m, 4H, 2CH₂), 3.35 (s, 1H, CH-cyclohexene), 3.77 (s, 6H, 2OCH₃), 5.74 (s, 2H, 2OH), 6.93–7.49 (m, 12H, CH-aromatic and 2NH₂), 10.47 (s, 1H, NH), 11.04 (s, 1H, NH), 11.66 (s, 1H, NH), 12.81 (s, 2H, 2NH). Anal. Calcd. for C₂₆H₃₂N₁₂O₆ (608): C, 51.31; H, 5.30; N, 27.62. Found: C, 51.38; H, 5.36; N, 27.69.

5.19 General procedure for the synthesis of (23a,b)

A solution of 22a,b (0.5 g) in DMF (15 mL) containing a few drops of piperidine was heated at reflux for 24 h. The reaction mixture was allowed to cool and poured into crushed ice and acidified with HCl. The separated solid was filtered off washed with water, dried, and recrystallized from the proper solvent to give 23a,b.

5.19.1 5-Amino-N′-(3-(2-(5-amino-2-(4-chlorophenyl)-2H-1,2,3-triazole-4-carbonyl)hydrazinyl)-cyclo-hex-2-en-1-ylidene)-2-(4-chlorophenyl)-2H-1,2,3-triazole-4-carbohydrazide (23a)

It was obtained as pale brown crystals from ethanol: M.P.: 136–138°C, yield: 5.30 g (91%). IR (KBr) (v _max/cm⁻¹): 3,470–3,419 (NH₂/NH), 3,050 (CH-aromatic), 2,925–2,856 (CH-aliphatic), 1,656 (CO). MS (EI, 70 eV): m/z (%) = 582 (M⁺ + 2). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.20–1.80 (m, 2H, CH₂), 2.51–2.89 (m, 4H, 2CH₂), 3.35 (s, 1H, CH-cyclohexene), 7.36–7.71 (m, 12H, CH-aromatic and 2NH₂), 7.96 (s, 1H, NH), 8.28 (s, 1H, NH), 10.30 (hump, 1H, NH). Anal. Calcd. for C₂₄H₂₂Cl₂N₁₂O₂ (580): C, 49.58; H, 3.81; N, 28.91. Found: C, 49.66; H, 3.88; N, 28.98.

5.19.2 5-Amino-N′-(3-(2-(5-amino-2-(4-methoxyphenyl)-2H-1,2,3-triazole-4-carbonyl)hydr-azinyl)-cyclohex-2-en-1-ylidene)-2-(4-methoxyphenyl)-2H-1,2,3-triazole-4-carbohydr-azide (23b)

It was obtained as brown crystals from ethanol: M.P.: 160–162°C, yield: 3.40 g (59%). IR (KBr) (v _max/cm⁻¹): 3,470–3,419 (NH₂/NH), 2,928–2,845 (CH-aliphatic), 1,655 (CO). MS (EI, 70 eV): m/z (%) = 572 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.23–1.90 (m, 2H, CH₂), 2.51–2.90 (m, 4H, 2CH₂), 3.36 (s, 1H, CH-cyclohexene), 3.82 (s, 6H, 2OCH₃), 7.09–7.92 (m, 12H, CH-aromatic and 2NH₂), 7.96 (s, 1H, NH), 8.30 (s, 1H, NH), 11.31 (s, 1H, NH). Anal. Calcd. for C₂₆H₂₈N₁₂O₄ (572): C, 54.54; H, 4.93; N, 29.36. Found: C, 54.60; H, 4.99; N, 29.43.

5.20 General procedure for the synthesis of (24a,b)

A mixture of compounds 21a,b (0.5 g) and hydrazine hydrate (10 mL) was fused for 12 h. The reaction mixture was allowed to cool and poured into crushed ice containing a few drops of hydrochloric acid. The separated solid was filtered off washed with water, dried, and recrystallized from the proper solvent to give 24a,b.

5.20.1 5-(2-(3-(2-(3-Amino-4-((4-chlorophenyl)diazenyl)-1H-pyrazol-5-yl)hydrazinyl)cyclo-hex-2-en-1-ylidene)hydrazinyl)-4-((4-chlorophenyl)diazenyl)-1H-pyrazol-3-amine (24a)

It was obtained as brown crystals from ethanol: M.P.: 178–180°C, yield: 4.60 g (79%). IR (KBr) (v _max/cm⁻¹): 3,422–3,200 (NH₂/NH), 2,937–2,830 (CH-aliphatic). MS (EI, 70 eV): m/z (%) = 580 (M⁺ + 2). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.24–1.91 (m, 2H, CH₂), 2.09–2.38 (m, 4H, 2CH₂), 4.70 (s, 1H, CH-cyclohexene), 6.00 (s, 4H, 2NH₂), 7.10–7.49 (m, 11H, CH-aromatic), 8.20 (s, 1H, NH), 8.30 (s, 1H, NH), 8.40 (s, 1H, NH), 13.00 (hump, 2H, 2NH). Anal. Calcd. for C₂₄H₂₄Cl₂N₁₄ (578): C, 49.75; H, 4.17; N, 33.84. Found: C, 49.82; H, 4.23; N, 33.90.

5.20.2 5-(2-(3-(2-(3-Amino-4-((4-methoxyphenyl)diazenyl)-1H-pyrazol-5-yl)hydrazinyl)cyclohex-2-en-1-ylidene)hydrazinyl)-4-((4-methoxyphenyl)diazenyl)-1H-pyrazol-3-amine (24b)

It was obtained as brown crystals from ethanol: M.P.: 140–142°C, yield: 4.20 g (73%). IR (KBr) (v _max/cm⁻¹): 3,470–3,204 (NH₂/NH), 3,050 (CH-aromatic), 2,928 (CH-aliphatic). MS (EI, 70 eV): m/z (%) = 570 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 0.84–1.99 (m, 2H, CH₂), 2.01–2.89 (m, 4H, 2CH₂), 3.82 (s, 6H, 2OCH₃), 4.80 (s, 1H, CH-cyclohexene), 6.99 (s, 4H, 2NH₂), 7.12–7.49 (m, 11H, CH-aromatic), 8.20 (s, 1H, NH), 8.30 (s, 1H, NH), 8.40 (s, 1H, NH), 13.30 (hump, 2H, 2NH). Anal. Calcd. for C₂₆H₃₀N₁₄O₂ (570): C, 54.73; H, 5.30; N, 34.37. Found: C, 54.80; H, 5.36; N, 34.45.

5.21 Synthesis of N-(3-amino-4-(p-tolyldiazenyl)-1H-pyrazol-5-yl)-2-(2-(3-(2-(2-(2-(3-amino-4-(p-tolyldiazenyl)-1H-pyrazol-5-yl)hydrazono)-2-cyanoacetyl)hydrazinyl)cyclohex-2-en-1-ylidene)-hydrazinyl)-2-oxoacetohydrazonoyl cyanide (25)

A cold suspension of aryl diazonium salts (0.02 mol [prepared from 0.02 mol of heteroaromatic amine with the appropriate quantities of sodium nitrite and hydrochloric acid]) was gradually added to a cold solution (0–5°C) of 3 (0.01 mol, 2.74 g) in ethanol (30 mL) containing anhydrous sodium acetate (1 g) with continuous stirring for 2 h. The resulting reaction product was filtered off, washed with water, and recrystallized from ethanol to give (25) as brown crystals: M.P.: 208–210°C, yield: 5.60 g (77%). IR (KBr) (v _max/cm⁻¹): 3,398–3,303 (NH₂/NH), 2,919–2,861 (CH-aliphatic), 2,215 (CN), 1,679 (CO). MS (EI, 70 eV): m/z (%) = 728 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 0.80–1.90 (m, 2H, CH₂), 2.29–2.36 (m, 4H, 2CH₂), 2.38 (s, 6H, 2CH₃), 4.70 (s, 1H, CH-cyclohexene), 6.50 (s, 4H, 2NH₂), 7.20–7.84 (m, 15H, CH-aromatic and 7NH). Anal. Calcd. for C₃₂H₃₂N₂₀O₂ (728): C, 52.74; H, 4.43; N, 38.44. Found: C, 52.81; H, 4.49; N, 38.50.

5.22 General procedure for the synthesis of (26a,b, 27a,b)

A solution of 3 (0.01 mol, 2.74 g) in ethanol (30 mL) containing a few drops of piperidine was treated with (0.02 mol) of 5-benzylideneimidazolidine-2,4-dione, 5-(2,4-dichlorobenzylidene)imidazolidine-2,4-dione, 2-benzylidene-cyclohexan-1-one and 2-(4-methoxybenzylidene)cyclohexan-1-one, respectively. The reaction mixture was heated under reflux for 6 h. The reaction mixture was left to cool at room temperature and poured into ice/water containing a few drops of hydrochloric acid, and the formed solids were collected by filtration, dried, and recrystallized from the proper solvent to give 26a,b, 27a,b.

5.22.1 5-Amino-N′-(3-(2-(5-amino-2-oxo-7-phenyl-1,2,3,7-tetrahydropyrano[2,3-d]imidazole-6-carbonyl)hydrazinyl)cyclohex-2-en-1-ylidene)-2-oxo-7-phenyl-1,2,3,7-tetrahydro-pyrano[2,3-d]-imidazole-6-carbohydrazide (26a)

It was obtained as brown crystals from ethanol: M.P.: 220–222°C, yield: 4.60 g (70%). IR (KBr) (v _max/cm⁻¹): 3,421–3,200 (NH₂/NH), 2,925–2,854 (CH-aliphatic), 1,770, 1,628 (2CO). MS (EI, 70 eV): m/z (%) = 650 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 0.84–1.91 (m, 2H, CH₂), 2.09–3.00 (m, 4H, 2CH₂), 3.90 (s, 2H, 4H-pyrane), 4.70 (s, 1H, CH-cyclohexene), 6.40 (s, 4H, 2NH₂), 6.92–7.66 (m, 13H, CH-aromatic and 3NH), 10.70 (s, 2H, 2NH), 11.40 (s, 2H, 2NH). Anal. Calcd. for C₃₂H₃₀N₁₀O₆ (650): C, 59.07; H, 4.65; N, 21.53. Found: C, 59.14; H, 4.70; N, 21.58.

5.22.2 5-Amino-N′-(3-(2-(5-amino-7-(2,4-dichlorophenyl)-2-oxo-1,2,3,7-tetrahydropyrano[2,3-d]-imid-azole-6-carbonyl)hydrazinyl)cyclohex-2-en-1-ylidene)-7-(2,4-dichlorophenyl)-2-oxo-1,2,3,7-tetrahydropyrano[2,3-d]imidazole-6-carbohydrazide (26b)

It was obtained as pale brown crystals from ethanol: M.P.: 180–182°C, yield: 5.20 g (66%). IR (KBr) (v _max/cm⁻¹): 3,423–3,181 (NH₂/NH), 3,066 (CH-aromatic), 2,935–2,856 (CH-aliphatic), 1,770, 1,634 (2CO). MS (EI, 70 eV): m/z (%) = 788 (M⁺ + 2). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.23–1.65 (m, 2H, CH₂), 2.09–3.00 (m, 4H, 2CH₂), 3.90 (s, 2H, 4H-pyrane), 4.49 (s, 1H, CH-cyclohexene), 6.48 (s, 4H, 2NH₂), 7.31–7.73 (m, 9H, CH-aromatic and 3NH), 10.74 (s, 2H, 2NH), 11.40 (s, 2H, 2NH). Anal. Calcd. for C₃₂H₂₆Cl₄N₁₀O₆ (786): C, 48.75; H, 3.32; N, 17.77. Found: C, 48.82; H, 3.40; N, 17.83.

5.22.3 2-Amino-N′-(3-(2-(2-amino-4-phenyl-5,6,7,8-tetrahydro-4H-chromene-3-carbonyl)-hydrazinyl)-cyclohex-2-en-1-ylidene)-4-phenyl-5,6,7,8-tetrahydro-4H-chromene-3-carbohydrazide (27a)

It was obtained as red crystals from ethanol: M.P.: 168–170°C, yield: 4.40 g (68%). IR (KBr) (v _max/cm⁻¹): 3,446–3,200 (NH₂/NH), 2,934–2,862 (CH-aliphatic), 1,628 (CO). MS (EI, 70 eV): m/z (%) = 646 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.19–1.91 (m, 10H, 5CH₂), 2.51–2.98 (m, 12H, 6CH₂), 3.68 (s, 2H, 4H-pyrane), 4.60 (s, 1H, CH-cyclohexene), 5.80 (s, 4H, 2NH₂), 7.23–7–36 (m, 13H, CH-aromatic and 3 NH). Anal. Calcd. for C₃₈H₄₂N₆O₄ (646): C, 70.57; H, 6.55; N, 12.99. Found: C, 70.64; H, 6.61; N, 13.04.

5.22.4 2-Amino-N′-(3-(2-(2-amino-4-(4-methoxyphenyl)-5,6,7,8-tetrahydro-4H-chromene-3-carbonyl)-hydrazinyl)cyclohex-2-en-1-ylidene)-4-(4-methoxyphenyl)-5,6,7,8-tetra-hydro-4H-chromene-3-carbohydrazide (27b)

It was obtained as pale brown crystals from ethanol: M.P.: 180–182°C, yield: 5.40 g (76%). IR (KBr) (v _max/cm⁻¹): 3,419–3,200 (NH₂/NH), 2,937–2,833 (CH-aliphatic), 1,656 (CO). MS (EI, 70 eV): m/z (%) = 706 (M⁺). ¹H NMR (400 MHz, DMSO-d ₆): δ (ppm) 1.59–1.73 (m, 10H, 5CH₂), 2.51–2.89 (m, 12H, 6CH₂), 3.36 (s, 2H, 4H-pyrane), 3.81 (s, 6H, 2OCH₃), 4.10 (s, 1H, CH-cyclohexene), 5.70 (s, 4H, 2NH₂), 7.01–7.59 (m, 11H, CH-aromatic and 3 NH). Anal. Calcd. for C₄₀H₄₆N₆O₆ (706): C, 67.97; H, 6.56; N, 11.89. Found: C, 68.04; H, 6.63; N, 11.96.

6 Conclusions

In this work, 1,3-cyclohexanedione 1 reacted with cyanoacetic acid hydrazide to afford bis (2-cyanoaceto hydrazide) derivative 3. Cyanoacetic acid hydrazide and its analogues are especially important starting materials for the synthesis of new biologically heterocyclic compounds. Our research deals with the effective use of bis (2-cyanoaceto hydrazide) derivative 3 in the synthesis of a variety of polyfunctionally azoles and (or) azines with biological interest.

Acknowledgements

The authors are very grateful to Prof. Dr. Mohamed. S. A. El-Gaby, Department of Chemistry, Faculty of Science, Al-Azhar University, Assiut, Egypt, for the valuable support and for reviewing this manuscript.

Funding information: Authors state no funding involved.
Conflict of interest: Authors state no conflict of interest.
Data availability statement: The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.

References

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Received: 2023-01-27

Revised: 2023-09-27

Accepted: 2023-10-06

Published Online: 2023-12-12

This work is licensed under the Creative Commons Attribution 4.0 International License.

Articles in the same Issue

https://doi.org/10.1515/hc-2022-0167

Keywords for this article

hydrazide–hydrazone; pyrazole; pyrane; pyridine; pyrole; spectral data; anti-microbial activity

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