5-Fluoro-1H-indole-2,3-dione-triazoles- synthesis, biological activity, molecular docking, and DFT study

Journal Pre-proof 5-Fluoro-1H-indole-2,3-dione-triazoles- synthesis, biological activity, molecular docking, and DFT study Sonal Deswal, Naveen, Ram Kumar Tittal, D. Ghule Vikas, Kashmiri Lal, Ashwani Kumar PII:

S0022-2860(20)30307-0

DOI:

https://doi.org/10.1016/j.molstruc.2020.127982

Reference:

MOLSTR 127982

To appear in:

Journal of Molecular Structure

Received Date: 4 December 2019 Revised Date:

14 February 2020

Accepted Date: 26 February 2020

Please cite this article as: S. Deswal, Naveen, R.K. Tittal, D. Ghule Vikas, K. Lal, A. Kumar, 5Fluoro-1H-indole-2,3-dione-triazoles- synthesis, biological activity, molecular docking, and DFT study, Journal of Molecular Structure (2020), doi: https://doi.org/10.1016/j.molstruc.2020.127982. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2020 Published by Elsevier B.V.

Credit Author Statement Sonal Deswal: Validation, Formal analysis, Investigation Naveen: Molinspiration Physicochemical Parameters Software, Data Curation Ram Kumar Tittal: Conceptualization, Methodology, Resources, Data Curation, Visualization, Writing -Original Draft, Writing- Review & Editing, Supervision Kashmiri Lal: Conceptualization, Methodology, Resources, Data Curation, Ghule Vikas D.: Software, Data Curation Ashwani Kumar: Docking study.

5-Fluoro-1H-indole-2,3-dione-triazoles- Synthesis, Biological Activity, Molecular Docking, and DFT Study Sonal Deswala, Naveen,a Ram Kumar Tittala,*, Ghule Vikas D.a Kashmiri Lalb,*, Ashwani Kumarc a

Department of Chemistry, National Institute of Technology, Kurukshetra, Haryana-136119 (India). b Department of Chemistry, GJUS&T Hisar, Haryana-12500 (India). c Department of Pharmaceutical Sciences, GJUS&T Hisar, Haryana-12500 (India).

New antibacterial and antifungal 1-((1-aryl)-1H-1,2,3-triazol-4-yl)methyl-5-fluoroindoline-2,3-dione molecules were synthesized under environment benign green condition using Cell-CuI-NPs heterogeneous catalyst in water. All synthesized molecules were successfully characterized by IR, 1H NMR, 13C NMR, mass and 19F NMR for the representative compounds. The antibacterial and antifungal activities are very much comparable to commonly used antibacterial and antifungal drugs Fluconazole and Ciprofloxacin, respectively. The pharmacological experimental results are in good agreement with molecular docking and DFT results.

5-Fluoro-1H-indole-2,3-dione-triazoles- Synthesis, Biological Activity, Molecular Docking, and DFT Study Sonal Deswala, Naveen,a Ram Kumar Tittala,*, Ghule Vikas D.a Kashmiri Lalb,*, Ashwani Kumarc a

Department of Chemistry, National Institute of Technology, Kurukshetra, Haryana-136119 (India). Department of Chemistry, GJUS&T Hisar, Haryana-12500 (India). c Department of Pharmaceutical Sciences, GJUS&T Hisar, Haryana-12500 (India). b

Abstract: An environmental friendly heterogeneous catalyst Cell-CuI-NPs was employed for the synthesis of biologically promising 1-((1-aryl)-1H-1,2,3-triazol-4-yl)methyl-5-fluoroindoline-2,3-diones via CuAAC click reaction of 5-fluoro-(1-prop-2-ynyl)indoline-2,3-dione, an alkyne with various organic azides in aqueous medium. Compounds 4b, and 4c, with MIC values 0.0075, 0.0075, 0.0082, 0.0164, for S. Epidermidis and B. Subtilis, respectively and compound 7b with MIC value 0.0156 for each S. Epidermidis, E. Coli, & P. Aeruginosa bacterial strains exhibited considerable antibacterial potency with the reference drug Ciprofloxacin (MIC: 0.0047 µmol/mL). On antifungal activity investigation, compound 4a, 4d, and 7c (MIC: 0.0075, 0.0082, and 0.0092 µmol/mL, respectively) for A. Niger exhibited better potency than reference drug Fluconazole (MIC: 0.0102 µmol/mL). Also, compound 4a, 4d, and 4e (MIC: 0.0075, 0.0082, and 0.0090 µmol/mL, respectively) for C. Albicans demonstrated considerable potency with respect to reference antifungal drug Fluconazole (MIC: 0.0051 µmol/mL). Antibacterial and antifungal activity results showed that incorporation of triazole unit in an alkyne 2 improved the potency. Molinspiration physicochemical parameters revealed that all the synthesized 5fluoroisatin-triazole molecules 4a-e and 7a-e have possessed good drug like properties. Further, antimicrobial activity results were supported by the molecular docking on alkyne 2 and its active triazole 4a as well as DFT study by B3PW91 level with 6-311G(d,p) basis set. The FMOs also revealed that incorporation of triazole moiety on to alkyne 2 has improved the pharmacological activities of the resulted triazoles 4a-e and 7a-e. Keywords: 5-Fluoro-isatin-triazole; Cell-CuI-NPs; Antimicrobial; Antifungal; DFT study

Introduction The versatility of 1H-indole-2,3-dione scaffold have been extensively reported in the literature[1-3]. Isatin, the trivial name for 1H-indole-2,3-dione, was found in medicinal plants of genus Isatis and Couroupitaguianensis aubl[1]. It has been explored by both synthetic and medicinal chemists for antimicrobial[4], anticancer[5], anti-HIV[6], anticonvulsants[7], antiinflammatories[8], antitubercular[9], and analgesic properties[10]. Sutent (SU11248) (generic name Sunitinib(I)) is a drug derived from 5-fluoro indoline-2,3-dione and is used for targeted therapy for gastrointestinal stromal tumour, advanced renal cancer pancreatic neuroendocrine carcinoma[11]. Some other related biologically active molecules shown in Figure 1 containing O

O

N H

N

N

NH HN

HO

OH

H

F

O

N N

CF3 F N H

NH O

(I)

(II)

(III)

Figure 1. Some biologically active indoline-2,3-dione derivatives as anticancer agents. Page 1 of 19

indoline-2,3-dione derivatives have recently been used for cancer treatment[12]. The development of resistance to drugs used for microbial pathogens is a serious problem which needs to be addressed. Hence, extensive efforts have been made for the synthesis and study of safe, effective and broad-spectrum antimicrobial agents[13]. Triazoles are among the most studied pharmacological agents due to their broad-spectrum activity, and high therapeutic index[14]. 1,2,3-Triazoles are reported for potential biological applications since they displayed anticancer[15],

antifungals[16-17],

antibacterial[18],

antituberculosis[19],

and

antiviral

activities[20]. Recently, Dawid Zych et al have explored the synthesis, optical and biological activity of heteroaryl substituted 4ˈ-(1,2,3-triazol-4-yl)phenyl-2,2ˈ:6ˈ,2ˈˈ-terpyridine. Also, Zych group synthesized six derivatives of 4′‐phenyl‐2,2′:6′,2′′‐terpyridine containing glycidyl, cyclohexylmethyl, benzyl, 4‐tert‐butylbenzyl, pentafluorobenzyl, and 2,6‐dichlorobenzyl groups. The chelating and physicochemical properties of terpyridine enabled one of the derivative more promising anticancer substitutes than doxorubicin anticancer drug[21-22]. A strategy of linking two bioactive molecules together to improve the biological and pharmacological potency of the resulted molecules has been a common practice nowadays in pharmaceutical and medicinal chemistry[23-25]. Recently Jacob P. Macdonald et al[26]. and Raghu Raj et al[27]. have employed N-propargyl-5-fluoroindoline-2,3-dione moiety to obtain isatin-linked triazoles with enhanced biological potency[28-29]. Our group has also reported some isatione-oxime, trifluoromethyl benzoate and semicarbazone linked-triazoles as potential antimicrobial agents[30-32]. Some environment-friendly methods of CuAAC reaction have reported the selective synthesis of 1,4-disubstituted-1,2,3-triazoles by using solid-supported (like cellulose) copper nanoparticles (Cell-CuI-NPs) as a heterogeneous-catalyst to enable quick and easy separation of reaction products in high yield from catalysts[33-35]. With these viewpoints in our mind, we are herewith reporting the synthesis of 5-fluoroindoline-2,3-dione-1-aryl- 1H[1,2,3]triazol-4-ylmethylhybrid molecules by a well-known CuAAC reaction using Cell-CuI-NPs as a heterogeneous catalyst in aqueous condition. The synthesized triazole conjugates were tested for their biological potency against some fungal and bacterial strains. Also, computational DFT studies were done for synthesized 5-fluoroindoline-2,3-dione-linked triazoles and results were optimized at the (B3PW91) level with 6311G(d,p) basis set. Further, frontiers molecular orbitals (FMO) and molecular electrostatic potential (MESP) calculations were done for their structural Page 2 of 19

exploration. Thereafter, reactivity parameters were predicted and recorded for these synthesized 5-fluoroindoline-2,3-dione-linked triazole conjugates. Results and Discussions: In order to obtain desired 5-fluoroindoline-2,3-dione-triazole conjugates, terminal alkyne 2 was prepared via N-propargylation of commercially available 5-fluoroindoline-2,3-dione 1 with propargyl bromide in presence of K2CO3 in DMF as per the reported procedure[36]. The resulting alkyne 2 was obtained as bright red colour precipitate which was recrystallized with chloroform/hexane (8:2 v/v ratio) to get a pure N-propargyl-5-fluoroindoline-2,3-dione in appreciable yield of 83%. Under method A, 5-fluoroindoline-2,3-dione-triazole conjugates 4a-e were synthesized by Cu(I) catalyzed 1,3-dipolar cycloaddition reaction between N-propargyl-5fluoroindoline-2,3-dione 2 with in situ generated benzyl azides from corresponding benzyl bromides 3 (Scheme 1)[37]. The desired representative 5-fluoroindoline-2,3-dione-triazole 4a was formed in 72% yield. However, in method B, the other desired 1,4-disubstituted-1,2,3triazole 7a-e (Scheme 2) were synthesized from readily prepared aryl azides 6 with already prepared alkyne 2 in higher yield and lesser reaction time as compared to method A. The aryl azides were prepared by the diazotization of corresponding anilines 5. The structure of the synthesized triazole 7a was fully characterized and established by FT IR, 1H NMR, 19

13

C NMR,

F NMR and mass spectral analysis. Wherein, the characteristic νmax peak due to =C–H

stretching of triazole ring appeared at 3146 cm-1 in FT IR spectrum of triazole 7a. Two C=O stretching bands appeared at νmax 1748 cm-1 and 1732 cm-1. In the 1H NMR spectrum, characteristic singlet due to triazolyl proton appeared at δ 8.09 ppm. The protons present in the

Scheme 1 (method A): Synthesis of 5-fluoroindoline-2,3-dione-triazoles 4a-e. Page 3 of 19

Scheme 2 (method B): Synthesis of 5-fluoroindoline-2,3-dione-triazoles 7a-e. aromatic ring resonated in the region δ 7.78-7.35 ppm. One sharp singlet at δ 5.13 ppm demonstrated the presence of two methylene protons of NCH2. In the 13C NMR spectrum of 7a, the two carbonyl carbons appeared at δ 182.47 and 157.78 ppm. Peaks at δ 142.21 and 121.25 ppm assigned to the carbon atom at C-4 and C-5 position of the triazole ring, respectively. The methylene carbon attached to N-1 of indoline exhibited a peak at δ 35.43 ppm. The multiplets appeared in the 13C NMR spectrum of 7a due to 13C-19F coupling, doublet at δ 160.78 (d, 1JCF = 244.9 Hz) appeared due to 13C-19F coupling of the carbon-bearing fluorine atom in indole C-5. In addition to this, doublets also appeared due to two bond and three bond coupling at δ 125.01 (d, 2

JCF = 23.9 Hz, indole C-6), 118.16 (d, 3JCF = 6.7 Hz, indole C-7), 112.94 (d, 3JCF = 7.1 Hz,

indole C-7a) and 112.40 (d, 2JCF = 24.3 Hz, indole C-4). Further, mass spectrum of triazole 7a showed a signal at m/z 345.0689 corresponding to [M+Na]+. The calculated value for [M+Na]+ is 345.0764. On the basis of spectral patterns and peaks observed from FT IR, 1H NMR, 13C NMR and mass spectra, the chemical structure of 7a was deduced and named as 5-fluoro-1-((1-phenyl1H-1,2,3-triazol-4-yl)methyl)indoline-2,3-dione. Efforts were made to improve the yield and standardized the reaction condition with different catalyst systems (CuSO4.5H2O/Na-ascorbate, CuI/DIPEA, Cu(OAc)2/Na-ascorbate, and Cell-CuI-NPs) along with variation in the choice of

suitable solvent system (THF:H2O (3:1 v/v), DMF:H2O (4:1 v/v), only THF, t-BuOH/H2O (4:1 v/v), and H2O only) for the formation of representative 1,4-disubstituted-1,2,3-triazoles 4a and 7a in

lesser reaction time, as detailed in Table 1. It was standardized that the use of heterogeneous catalyst Cell-CuI-NPs in H2O under reflux resulted to the synthesis of triazoles 4a and 7a in higher yield of 88 and 85%, respectively. Also, under this reaction condition both electronwithdrawing and electron-donating functional groups (NO2, Br, Cl, F, and CH3) at each o, m and p-positions remained unaffected as highlighted in Table 2.

Page 4 of 19

Table 1. Standardized reaction conditions for the synthesis of representative triazoles 4a and 7aa S. No.

Condition

T (h)

Compound

Yieldb

1

NaN3, CuSO4.5H2O, Na ascorbate, THF:H2O (3:1)

7

4a

72

2

NaN3, CuSO4.5H2O, Na ascorbate, THF:H2O (3:1)

5

7a

78

3

NaN3, CuI, DIPEA, THF

9

4a

70

4

NaN3,CuI, DIPEA, THF

6

7a

76

5

NaN3, Cu(OAc)2, Na ascorbate, t-BuOH/H2O (4:1)

8

4a

73

6

NaN3, Cu(OAc)2, Na ascorbate, BuOH:H2O (4:1)

5

7a

79

7

NaN3, CuSO4.5H2O, Na ascorbate, DMF:H2O (4:1)

5

4a

75

8

NaN3, CuSO4.5H2O, Na ascorbate, DMF:H2O (4:1)

4

7a

80

9

NaN3, Cell-CuI-NPs, H2O, 70 °C

2

4a

88

10

NaN3, Cell-CuI-NPs, H2O, 70 °C

1.5

7a

85

a

Reaction condition: All benzyl bromide, sodium azide-alkyne 2 and aryl azide (1 mmol). bYield is after purification by column chromatography.

Table 2. Synthesis of alkyne 2a, and 5-fluoroindoline-2,3-dione-triazoles 4a-e & 7a-e via CuAAC using Cell-CuI- NPs as catalyst in water as solvent.b S. No.

R1 or R2

Product

T (h)

M. P. (°C)

Yield (%)c

1

--

2

8

128-130

83

2

R1: 3-BrC6H4CH2

4a

2

187-189

82

3

R1: 2-BrC6H4CH2

4b

2

184-186

76

4

R1: 3-NO2C6H4CH2

4c

1.5

193-195

87

5

R1: 4-NO2C6H4CH2

4d

1.5

199-201

86

6

R1: 2-CH3C6H4CH2

4e

2

205-207

72

7

R2: H

7a

1.5

174-176

85

8

R2: 4-Br

7b

1.5

196-198

79

9

R2: 4-F

7c

1.5

200-202

83

10

R2: 4-Cl

7d

1.5

201-203

84

11

R2: 4-NO2

7e

1.5

209-211

87

a

5-Fluoroisatin (1.0 mmol), anhy. K2CO3 (1.5 mmol), propargyl bromide (80% in toluene, 1.5 mmol), 8h reflux. Sodium azide (3.0 mmol), benzyl bromide (1.0 mmol), N-propargyl-5-fluoroindoline-2,3-dione 2 (1.0 mmol), CellCuI-NPs (0.1g), 1-2h at 70 °C. cYield after purification by column chromatography. b

Page 5 of 19

Pharmacological Study (Antibacterial Activity) All the synthesized 5-fluoroindoline-2,3-dione-triazole hybrids were tested for in vitro antibacterial activity against two gram-positive bacteria (Staphylococcus Epidermidis MTCC 6880 and Bacillus Subtilis MTCC 441) and two gram-negative bacteria (Escherichia Coli MTCC 16521 and Pseudomonas Aeruginosa MTCC 424) by standard serial dilution method (Spooner and Sykes, 1972) [38]. The antibacterial efficiency of synthesized triazoles compared with standard drug Ciprofloxacin. The test results were reported by recording minimum inhibitory concentrations (MIC) in µmol/mL and listed in Table 3. From antibacterial screening results, it was revealed that most of the synthesized 1,4-disubstituted-1,2,3-triazole hybrids (4a-e, 7a-e) exhibited good to high activity against tested bacterial strains. Similar to the case of antifungal activity evaluation, the activity of the synthesized 5-fluoroindoline-2,3-dione-triazole hybrids was found better than 5-fluoro-1-(prop-2-ynyl)indoline-2,3-dione 2, highlighting that introduction of triazole moiety increases the antibacterial activity. Triazoles 4b (MIC, 0.0075 µmol/mL) and 4c (0.0082 µmol/mL) having 2-Br and 3-NO2 groups at benzyl ring displayed very good antibacterial activity against S. epidermidis. Their activity was found comparable to that of reference drug Ciprofloxacin with MIC value of 0.0047 µmol/mL. Moreover, compound 4b (0.0075 µmol/mL) exhibited antibacterial potency comparable to the standard B. Subtilis also. Table 3. In vitro antibacterial activity of 4a-e & 7a-e, MIC in µmol/mL. S. No.

Comp.

R1/R2

Ac

Bd

Ce

Df

1

2

---

0.0308

0.0615

0.0615

0.0615

2

4a

3-Br-C6H4CH2

0.0151

0.0301

0.0301

0.0301

3

4b

2-Br-C6H4CH2

0.0075

0.0075

0.0301

0.0301

4

4c

3-NO2-C6H4CH2

0.0082

0.0164

0.0328

0.0164

5

4d

4-NO2-C6H4CH2

0.0164

0.0328

0.0328

0.0164

6

4e

2-CH3-C6H4CH2

0.0178

0.0357

0.0357

0.0357

7

7a

H

0.0194

0.0388

0.0388

0.0388

8

7b

4-Br

0.0156

0.0312

0.0156

0.0156

9

7c

4-F

0.0184

0.0367

0.0367

0.0367

10

7d

4-Cl

0.0175

0.0350

0.0350

0.0350

11

7e

4-NO2

0.0170

0.0340

0.0340

0.0340

---

0.0047

0.0047

0.0047

12

Ciprofloxacin

Page 6 of 19

c

A: S. epidermidis; dB: B. subtilis; eC: E. coli; fD: P. aeruginosa

Further, it was observed in the case of E. Coli that the compound 7b having 4-Br group attached to benzene ring shows appreciable potency with MIC value of 0.0156 µmol/mL. It was inferred that 4c with the NO2 group displayed broad-spectrum activity with good inhibition effects and are active against all strains under test. On comparing triazoles with benzyl ring (4a-e) it was observed that compound with electron-withdrawing groups (4a-d) exhibited better antibacterial activity than the compound containing electron releasing methyl group in 4e. Antifungal activity The biological potency of all the synthesized 5-fluoroindoline-2,3-dione-triazole hybrid compounds was tested by screening them for in vitro antifungal activity against two fungal strains viz. Aspergillus Niger (MTCC 8189) and Candida Albicans (MTCC 227) by standard serial dilution method (Spooner and Sykes, 1972) [38]. Fluconazole was used as a standard drug and minimum inhibitory concentrations (MIC in µmol/mL) were recorded as shown in Table 4. The activity data revealed moderate to the excellent antifungal activity of the synthesized 1,4Table 4. In vitro antifungal activity of compounds 4a-e, 7a-e (MIC in µmol/mL). S. No.

R1, R2

Comp.

A. Niger

C. Albicans

1

2

---

0.0308

0.0308

2

4a

3-Br-C6H4-CH2

0.0075

0.0075

3

4b

2-Br-C6H4-CH2

0.0151

0.0151

4

4c

3-NO2-C6H4-CH2

0.0164

0.0164

5

4d

4-NO2-C6H4-CH2

0.0082

0.0082

6

4e

2-CH3-C6H4-CH2

0.0178

0.0090

7

7a

-H

0.0194

0.0194

8

7b

4-Br

0.0156

0.0156

9

7c

4-F

0.0092

0.0184

10

7d

4-Cl

0.0175

0.0175

11

7e

4-NO2

0.0170

0.0170

--

0.0102

0.0051

12

Fluconazole31

disubstituted-1,2,3-triazoles (4a-e, 7a-e). The antifungal activity of the synthesized 5fluoroindoline-2,3-dione-triazole

hybrids

was

found

better

than

5-fluoro-1-(prop-2-

ynyl)indoline-2,3-dione 2, highlighting that the addition of triazole moiety increases the Page 7 of 19

antifungal activity of triazoles. Also, triazole hybrids 4a, 4d and 7c displayed better activity with MIC values of 0.0075, 0.0082 and 0.0092 µmol/mL, respectively than classically used antifungal drug Fluconazole (MIC, 0.0102 µmol/mL) for A. Niger. However, in the case of C. Albicans triazole hybrids 4a, 4d and 4e displayed comparable activity (MIC, 0.0090-0.0075 µmol/mL) to the reference drug Fluconazole (MIC, 0.0051µmol/mL). Drug Likeness (Molinspiration Physicochemical Parameters) The physicochemical parameters of the alkyne 2 and triazole derivatives (4a-e & 7a-e) to predict the drug-likeness behavior were calculated by using Molinspiration software[39]. The drug-likeness was measured to check whether the synthesized compounds are similar to some existing drugs. The Lipinski’s rules of five have been used to measure the drug-likeness. The drug-likeness data are useful to analyze the pharmacokinetic parameters like absorption, distribution, metabolism, and excretion from the living body. The computed values of the synthesized compounds are presented in Table, SI-1-2 (SI File). The Molinspiration physicochemical parameters showed that all the synthesized triazoles have sufficient number of rotational bonds and would show good flexibility. Also, the sufficient number of H-acceptor and H-donor bonds in the synthesized triazoles showed strong binding with the target molecule. The computed data revealed the good absorption values for all the synthesized triazoles and thus, can be easily absorbed in the living systems. The %ABS was calculated by using the formula %ABS = 109-(0.345 x TPSA) [39]. All the synthesized triazole compounds showed good %absorption i.e. %ABS= 69.11 - 84.92 which ranges from considerable to good range i.e. towards cent percent absorption. Further, the values of octanol-water partition coefficient (milogP) determine the hydrophilicity of any drug which decides toxicity, absorption, and drug-receptor interactions. The data of milogP for the synthesized triazoles ranges from 1.05 to 2.19. This range is much less than 5.0 and showed good agreement as per Lipinski’s rule. Also, the number of H-bond acceptor ranges from 6-9 for triazoles which is less than 10 and the number of H-bond donors for all synthesized triazoles is 0 and thus, less than 5 as per the rule. According to the Lipinski rule of five, also called Pfizer’s rule of five any chemical compound as an oral drug would be biologically active if it does not violate more than one rule out of the proposed rules wherein, first rule said, the octanol-water partition coefficient (milogP) must be ≤5; the second rule said, the molecular weight of the probable drug must be <500 daltons; the third rule said, the number

Page 8 of 19

of H-bond acceptors in the molecule under consideration must be ≤10 and the last rule said, the number of H-bond donors must be ≤5. The bioactivity results listed in the tables revealed that the parameters of alkyne and all the synthesized triazoles are within limits of Lipinski’s rule of five with violation number equal to zero. Thus, all the synthesized 5-fluoroisatin-triazole molecules 4a-e and 7a-e have possessed good drug like property. Docking Studies The 5-fluoroindoline-2,3-dione-linked triazole molecules were synthesized with a view to increase the antimicrobial activity of the existing biomolecule 5-fluoroisatin. The experimental results have proved that the intention was right. To further investigate the binding conformations of these compounds, alkyne 2 and its most active triazole 4a were docked in the active site of E. Coli DNA Gyrase (PDB ID: 1KZN) using Autodock Vina tool [40]. The 3D structures of 2 and 4a were prepared with Marvin sketch [41] and the protein preparation part was completed with UCSF Chimera [42]. The Autodock Vina was utilized for docking simulations while discovery studio was used for visualization work. The most stable anchoring conformations of these compounds along with interacting residues are shown in Figure 2 and 3 created with the help of discovery studio visualizer [43]. Alkyne 2 exhibited one π-donor H-bond with Asn46 and hydrophobic interactions with Ala47 & Ile78. The triazole 4a displayed more number of interactions and introduction of substituted triazole ring caused better fitting of the compound i.e. binding affinity of 4a was 8.1 kcal/mol while for 2 it was 7.0 kcal/mol. The fluorine group helped specially in affixing to the binding site residues by making one H-bond with Ala96 and two halogen bonds with Val93 and His95. Other parts of the molecule showed hydrophobic interactions with Ala47, Ile78 and Ile90. These results further confirmed our experimental antimicrobial results.

Page 9 of 19

Figure 2 Binding mode of 2 docked with E. Coli DNA gyrase topoisomerase II (1 kzn); [Green: H-bond, Pink: hydrophobic interactions].

Figure 3 Binding mode of 4a docked with E. Coli DNA gyrase topoisomerase II (1 kzn); [Green: H-bond, Cyan: halogen bond, Pink: hydrophobic interactions]. Computational Study The optimized geometries of the synthesized 5-fluoroindoline-2,3-dione-triazole hybrid compounds (4a-e, 7a-e) were obtained by DFT (B3PW91) level with 6-311G (d, p) basis set. The FMO analysis of the optimized geometries was computed at the same level. Frontier molecular orbital energies along with the energy gap between HOMO and LUMO is given in Table 5. The distribution patterns of FMOs (HOMOs and LUMOs along with energies) of synthesized triazole hybrids at ground states are shown in Figure 4. HOMO has the ability to donate electrons while LUMO accepts electrons[44]. In order to investigate the bioactivity behaviour of molecules; FMOs are studied. As per FMO theory, both HOMO and LUMO, are very useful factors to understand molecular reactivities, electronic transitions, and intermolecular interactions, thus providing insight for bioactivity[45-47]. Further, the location of FMOs on the same side of the molecule strongly decreases the biological activity[48]. The π-electron cloud of HOMOs and LUMO of all triazoles is distributed over the 5-fluoroindoline-2,3-dione ring except in 7e where the π-electron cloud of LUMO is concentrated over the whole molecule, for reference alkyne 2, triazole 4a and 7a are given in Figure 4. Addition of substituents on the benzyl or benzene ring has little effect on the π-electron cloud of HOMO and LUMO. Interestingly, the synthesized 5-fluoroindoline-2,3-dione-triazole hybrid compounds show a small difference in the energy gap (∆ELUMO–HOMO) value with all derivatives lying in the range of Page 10 of 19

3.57-3.58 eV. The lower ∆ELUMO–HOMO reveals the higher biological activity[48]. It is also observed that the energy gap of the synthesized 5-fluoroindoline-2,3-dione-triazole hybrid was found smaller than 5-fluoro-1-(prop-2-ynyl)indoline-2,3-dione 2, it revealed that incorporation of 1,2,3-triazole unit lowers the energy gap between FMOs and hence increasing the chemical reactivity and antimicrobial activity of these synthesized triazole hybrids. Similar results were obtained with antifungal as well as antibacterial activity data. Table 5. FMO energy of 5-fluoroindoline-2,3-dione-triazole hybrid compounds (4a-e, 7a-e). EHOMO (eV)

ELUMO (eV)

∆ELUMO–HOMO (eV)

2

-6.78

-3.15

3.63

2

4a

-6.58

-3.01

3.57

3

4b

-7.22

-2.07

5.15

4

4c

-7.35

-2.06

5.29

5

4d

-7.73

-2.96

4.77

6

4e

-7.18

-2.07

5.11

7

7a

-7.11

-2.12

4.99

8

7b

-7.17

-2.10

5.07

9

7c

-7.17

-2.12

5.05

10

7d

-7.73

-3.13

4.60

11

7e

-6.45

-2.33

4.12

S.No.

Compound

1

HOMO (2)

LUMO (2)

Page 11 of 19

HOMO (4a)

LUMO (4a)

HOMO (7a)

LUMO (7a)

Figure 4. HOMO & LUMO of alkyne 2, and triazoles 4a & 7a. The MESP of synthesized 5-fluoroindoline-2,3-dione-triazole hybrid compounds (4a-e, 7a-e) has been computed at B3PW91/6-311G(d,p) level of DFT study. The computational mapping of MESP explores the molecular reactivity of compounds under investigation. By means of MESP, electrophilic as well as nucleophilic sites in the target molecules are predicted[49]. During MESP mapping the electrophilic site is shown by blue colour code (region of more negative potential than -12 kcal/mol) which is electron deficient region and the nucleophilic site shown by the red colour code (region of much greater potential than 16 kcal/mol) which is electron-rich as shown in Figure 5. This helps in understanding the type of action of the whole target molecule[50]. As a result, MESP strongly applied for understanding the interactions between enzyme and substrate[51], energy change profile that take place upon interaction, determining local reactivity site of large target molecules, H-bonding[52], drug-receptor and interactions between ligand & substrate. The synthesized triazole hybrids follow the usual trend of having deep red electronrich region lies mainly on 5-F substituent of indoline-2,3-dione ring and over the active functional group attached to benzyl or phenyl ring of the conjugated triazole molecules. Hence, for triazoles in the study, concerned atoms are most reactive points for the electrophilic attack also ideal sites for positive points of Page 12 of 19

2

4a

7a

Figure 5. Calculated electrostatic potentials on the 0.001 electrons/bohr3 molecular surface of the synthesized compounds. Positions of atoms in a molecule are shown as grey circles and colour ranges (in kcal/mol): red greater than 16; yellow between 16 and 14; green between 14 and 12; blue more negative than 12. receptor molecules. Hence, it is observed that these sites of molecule easily interact with the amino acid residues of receptor molecules. However, the deep blue colour region, an electrondeficient region is mainly located over indoline-2,3-dione moiety, triazole unit and benzyl or phenyl ring of synthesized molecules. These sites are the most reactive site for a nucleophilic reaction. In order to gain insight into the stability and chemical reactivity of synthesized molecular conjugates, we decided to study some chemical reactivity parameters. These include total energy, electrophilicity (ω), chemical hardness (η) and electronic chemical potential (µ) [53]. These indices depend on the HOMO-LUMO energy gap of molecules. Chemical hardness (η) can be defined by equation (1) as follows: η = (EHOMO-ELUMO)/2

(1)

Along with chemical hardness, molecular stability is another feature that is determined and correlated by the HOMO-LUMO energy gap. It was inferred that chemically harder and stable molecules have higher HOMO-LUMO energy gap than softer and less stable molecules. Another parameter, electrophilicity index (ω) measures the energy changes associated with the flow of electrons from HOMO to LUMO and is defined by the mathematical equation (2) given below[54]. ω = µ2/2η

(2)

where η represents chemical hardness and µ represents the electronic chemical potential which is negative of electronegativity of molecules[55]. Electronic chemical potential is given by the equation shown below (3): Page 13 of 19

µ = (EHOMO + ELUMO)/2

(3)

It depicts the transfer of charge that takes place within the molecule in the ground state and describes the tendency of electrons to escape from the equilibrium state. Hence, chemically reactive molecules show greater chemical potential. Table 6 depicts the above-described chemical reactivity parameters of synthesized triazole conjugates. On comparing the reactivity parameters of compound 2 with other synthesized triazole hybrids we concluded that introduction of the 1,2,3-triazole moiety in the structure decreases the HOMO-LUMO energy gap along with decreasing the chemical hardness and increasing chemical potential of a molecule. Hence, ultimately this results in increasing the chemical reactivity and bioactivity of molecules by 1,2,3-triazole moiety. On comparison, all triazoles show almost similar hardness in the range of 1.78-1.79 eV. Similarly, the electronic potential range is (-4.74)-(-4.99) eV while 7e is most electrophilic in nature with the highest electrophilicity index (ω) i.e. 6.97 eV. Table 6: Reactivity indices of 5-fluoroindoline-2,3-dione-triazole hybrids (4a-e, 7a-e). S. No.

Compound

ɳg (ev)

-µh (ev)

ωi (ev)

1

2

1.82

4.97

6.78

2

4a

1.79

4.74

6.43

3

4b

1.79

4.79

6.28

4

4c

1.79

4.85

6.57

5

4d

1.79

4.89

6.69

6

4e

1.78

4.74

6.30

7

7a

1.78

4.80

6.44

8

7b

1.79

4.86

6.62

9

7c

1.79

4.84

6.56

10

7d

1.78

4.87

6.62

11

7e

1.79

4.99

6.97

g

Chemical hardness. hElectronic potential. iElectrophilicity

Conclusions In conclusions, a library of 5-fluoroindoline-2,3-dione-triazole hybrid compounds was synthesized by CuAAC reaction using cell-CuI-NPs as a heterogeneous catalyst in environmentally benign aqueous condition. The synthesized derivatives exhibited moderate to excellent activity against tested fungal and bacterial strains. Compounds 4a, 4d and 7c (MIC: Page 14 of 19

0.0075, 0.0082, 0.0092 µmol/mL, respectively) displayed better potency than antifungal drug Fluconazole (MIC: 0.0102 µmol/mL) against A. Niger. Compounds 4b, 4d, and 4e exhibited activity comparable to the standard drug against C. Albicans with MIC values in the range of 0.0090-0.0075 µmol/mL. Triazoles 4b and 4c with 2-Br and 3-NO2 groups attached to benzyl and phenyl ring, respectively displayed activity comparable to that of reference drug Ciprofloxacin against S. Epidermidis and B. Subtilis. Also, compound 4c with the NO2 group displayed broad-spectrum antibacterial activity with good inhibition effects against all the tested strains. The DFT results by (B3PW91)/6-311G (d,p) basis set studied FMO, MESP and reactivity parameters. It was observed that the introduction of triazole moiety into 5-fluoroisatin linked alkyne increases biological potency of synthesized molecules by lowering the energy gap between FMOs. The results obtained from biological activity data were in good agreement to the theoretical DFT results. The synthesized 5-fluoroisatin linked triazole molecules may find potential application in bioactive natural products, pharmaceutical and synthetic chemistry. Author Information Corresponding Author: [email protected] Declaration of competing interest: The authors declare no competing financial interest. Acknowledgements We thank Dr A. Bhankhar, Department of Bio & Nano Technology, Hisar, GJUS&T, Haryana (India) for rendering Pharmacological facilities. http://dx.doi.org/10.4172/2161-0444.1000173 References [1] G. Brahmachari Chemistry and pharmacology of naturally occurring bioactive compounds 2013 (CRC Press) Boca Raton. [2] G. Mathur, S. Nain Recent advancement in synthesis of isatin as anticonvulsant agents: a review Med Chem, 2014, 4, 417-427. http://dx.doi.org/10.4172/2161-0444.1000173 [3] M. Prudhomme Advances in anticancer agents in medicinal chemistry 2013 (Bentham Science Publishers), USA.

Page 15 of 19

[4] S. Tiwari, P. Pathak, R. Sagar Efficient synthesis of new 2, 3-dihydrooxazole-spirooxindoles hybrids as antimicrobial agents Bioorg Med Chem Lett, 2016, 26, 2513-2516. https://doi.org/10.1016/j.bmcl.2016.03.093

[5] A. Cane, M. C. Tournaire, D. Barritault, M. Crumeyrolle-Arias The endogenous oxindoles 5hydroxyoxindole and isatin are antiproliferative and proapoptotic Biochem Biophys Res Commun, 2000, 276, 379-384. https://doi.org/10.1006/bbrc.2000.3477 [6] P. Selvam, N. Murugesh, M. Chandramohan, Z. Debyser, M. Debyser, M. Witvrouw Design, synthesis and antiHIV activity of novel isatine-sulphonamides Indian J Pharm Sc, 2008, 70, 779782. https://doi.org/10.4103/0250-474X.49121 [7] M. Verma, S. N. Pandeya, K. N. Singh, P. S. James Anticonvulsant activity of Schiff bases of isatin derivatives Acta Pharm, 2004, 54, 49-56. https://pdfs.semanticscholar.org/be94/13ea3407e88e5003df77fefb2ce2d6d1bd0b.pdf [8] P. K. Sharma, S. Balwani, D. Mathur, S. Malhotra, B. K. Singh, A. K. Prasad, C. Len, E. V. V. Eycken, B. Ghosh, N. G. Richards, V. S. Parmar Synthesis and anti-inflammatory activity evaluation of novel triazolyl-isatin hybrids J Enzyme Inhib Med Chem, 2016, 5, 1520-1526. https://doi.org/10.3109/14756366.2016.1151015

[9] K. Kumar, S. Carrère-Kremer, L. Kremer, Y. Guérardel, C. Biot, V. Kumar 1H-1,2,3Triazole-Tethered Isatin–Ferrocene and Isatin–Ferrocenylchalcone Conjugates: Synthesis and in Vitro Antitubercular Evaluation Organometallics, 2013, 32, 5713-5719. https://doi.org/10.1021/om301157z [10] A. V. Lastovka, V. P. Fadeeva, I. V. Il'Ina, S. Yu. Kurbakova, K. P. Volcho, N. F. Salakhutdinov, Study of physicochemical properties and development of the technique for quantitative determination of (2,4,4a,7,8a)-4,7-dimethyl-2-(thiophen-2-yl)octahydro-2-chromen4-ol which exhibits high analgesic activity Zavodsk. Lab. 2017, 83, 11-17. [11] P. Bianca, B. Mauro, G. Luciana, C. Sara, D. Sara, T. Pietro, A. Marco, M. Elena, T. Giuseppe, Analytical aspects of sunitinib and its geometric isomerism towards therapeutic drug monitoring in clinical routine J. Pharm. Biomed. Anal. 2018, 160, 360-367. https://doi.org/10.1016/j.jpba.2018.08.013

[12] N. M. Evdokimov, I. V. Magedov, D. McBrayer, A. Kornienko satin derivatives with activity against apoptosis-resistant cancer cells Bioorg Med Chem Lett, 2016, 26, 1558-1560. https://doi.org/10.1016/j.bmcl.2016.02.015

[13] P. L. White, R. B. Posso, R. A. Barnes Analytical and clinical evaluation of the PathoNostics AsperGenius assay for detection of invasive aspergillosis and resistance to azole antifungal drugs during testing J Clin Microbiol, 2015, 53, 2115-2121. doi: 10.1128/JCM.00667-15. [14] T. R. K. Reddy, C. Li, X. X. Guo, P. M. Fischer, L. V. Dekker Design, synthesis and SAR exploration of tri-substituted 1, 2, 4-triazoles as inhibitors of the annexin A2–S100A10 protein interaction Bioorg Med Chem, 2014, 22, 5378-5391. https://doi.org/10.1016/j.bmc.2014.07.043 [15] D. K. Dalvie, A. S. Kalgutkar, S. C. Khojasteh-Bakht, R. S. Obach, J. P. Donnell Biotransformation reactions of five-membered aromatic heterocyclic rings Chem Res Toxicol, 2002, 15, 269-299. https://doi.org/10.1021/tx015574b [16] K. Motahari, H. Badali, S. M. Hashemi, H. Fakhim, H. Mirzaei, A. Vaezi, M. Shokrzadeh, S. Emami Discovery of benzylthio analogs of fluconazole as potent antifungal agents Future Med Chem, 2018, 10, 987-1002. https://doi.org/10.4155/fmc-2017-0295 [17] G. C. Brandão, F. C. R. Missias, L. M. Arantes, L. F. Soares, K. K. Roy, R. J. Doerksen, A. B. Oliveira, G. R. Pereira Antimalarial naphthoquinones. Synthesis via click chemistry, in vitro Page 16 of 19

activity, docking to PfDHODH and SAR of lapachol-based compounds Eur J Med Chem, 2018, 145, 191-205. https://doi.org/10.1016/j.ejmech.2017.12.051 [18] O. A. Phillips, E. E. Udo, M. E. Abdel-Hamid, R. Varghese Synthesis and antibacterial activity of novel 5-(4-methyl-1H-1,2,3-triazole) methyl oxazolidinones. Eur J Med Chem, 2009, 44, 3217-3227. https://doi.org/10.1016/j.ejmech.2009.03.024 [19] N. Pokhodylo, O. Shyyka, V. Matiychuk Synthesis and anticancer activity evaluation of new 1,2,3-triazole-4-carboxamide derivatives Med Chem Res, 2014, 23, 2426-2438. DOI 10.1007/s00044-013-0841-8 [20] R. Kharb, M. S. Yar, P. C. Sharma New insights into chemistry and anti-infective potential of triazole scaffold. Curr Med Chem, 2011, 18, 3265-3297. DOI : 10.2174/092986711796391615 [21] D. Zych, A. Slodek, D. Zimny, S. Golba, K. Malarz, A. M. Wilczkiewicz Influence of the substituent D/A at the 1,2,3-triazole ring on novel terpyridine derivatives: synthesis and properties RSC Adv., 2019, 9, 16554-16564. https://doi.org/10.1039/C9RA02655J [22] D. Zych, A. Slodek, S. Krompiec, K. Malarz, A. M. Wilczkiewicz, R. Musio 4′Phenyl2,2′:6′,2′′terpyridine Derivatives Containing 1Substituted2,3Triazole Ring: Synthesis, Characterization and Anticancer Activity ChemSelect 2018, 3, 7009-7017. https://doi.org/10.1002/slct.201801204 [23] Y Zou, Q. J. Zhao, J. Liao, H. G. Hu, S. C. Yu, X. Y. Chai, M. J. Xu, Q. Y. Wu New triazole derivatives as antifungal agents: synthesis via click reaction, in vitro evaluation and molecular docking studies Bioorg Med Chem Lett, 2012, 22, 2959-2962. https://doi.org/10.1016/j.bmcl.2012.02.042 [24] C. Q. Sheng, X. Y. Che, W. Y. Wang, S. Z. Wang, Y. B. Cao, J. Z. Yao, Z. Y. Miao, W. N. Zhang StructureBased Design, Synthesis, and Antifungal Activity of New Triazole Derivatives Chem Biol Drug Des, 2011, 78, 309-313. https://doi.org/10.1111/j.1747-0285.2011.01138.x [25] N. S. Vatmurge, B. G. Hazra, V. S. Pore, F. Shirazi, M. V. Deshpande, S. Kadreppa, S. Chattopadhyay, R. G. Gonnade Synthesis and biological evaluation of bile acid dimers linked with 1,2,3-triazole and bis-β-lactam Org Biomol Chem, 2008, 6, 3823-3830. https://doi.org/10.1039/B809221D [26] J. P. MacDonald, J. J. Badillo, G. V. Arevalo, A. Silva-García, A. K. Franz Catalytic Stereoselective Synthesis of Diverse Oxindoles and Spirooxindoles from Isatins ACS Comb Sci, 2012, 14, 285-293. https://doi.org/10.1021/co300003c [27] R. Raj, P. Singh, P. Singh, J. Gut, P. J. Rosenthal, V. Kumar Azide-alkyne cycloaddition en route to 1H-1,2,3-triazole-tethered 7-chloroquinoline-isatin chimeras: synthesis and antimalarial evaluation. Eur J Med Chem, 2013, 62, 590-596. https://doi.org/10.1016/j.ejmech.2013.01.032 [28] V. V. Rostovtsev, L. G. Green, V. V. Fokin, K. B. Sharpless A stepwise huisgen cycloaddition process: copper(I)-catalyzed regioselective "ligation" of azides and terminal alkynes Angew Chem Int Ed, 2002, 41, 2596-2599. https://doi.org/10.1002/15213773(20020715)41:14<2596::AID-ANIE2596>3.0.CO;2-4 [29] F. Himo, T. Lovell, R. Hilgraf, V. V. Rosrovtsev, L. Noodleman, K. B. Sharpless, V. V. Fokin Copper(I)-Catalyzed Synthesis of Azoles. DFT Study Predicts Unprecedented Reactivity and Intermediates J Am Chem Soc, 2005, 127, 210-216. https://doi.org/10.1021/ja0471525 [30] P. Yadav, K. Lal, A. Kumar, S. K. Guru, S. Jaglan, S. Bhushan Green synthesis and anticancer potential of chalcone linked-1,2,3-triazoles. Eur J Med Chem, 2017, 126, 944-953. https://doi.org/10.1016/j.ejmech.2016.11.030 Page 17 of 19

[31] S. Deawal, R. K. Tittal, P. Yadav, K. Lal, V. D. Ghule, N. Kumar CelluloseSupported CuINanoparticlesMediated Green Synthesis of TrifluoromethylbenzoateLinked Triazoles for Pharmacological & DFT study ChemistrySelect, 2019, 4, 759-764. https://doi.org/10.1002/slct.201803099 [32] Naveen, R. K. Tittal, P. Yadav, K. Lal, V. D. Ghule, A. Kumar Synthesis, molecular docking and DFT studies on biologically active 1,4-disubstituted-1,2,3-triazole-semicarbazone hybrid molecules New J Chem, 2019, 43, 8052-8058. https://doi.org/10.1039/C9NJ00473D [33] F. Y. Alonso, M. G. Radivoy Copper Nanoparticles in Click Chemistry Acc Chem Res, 2015, 48, 2516-2528. https://doi.org/10.1021/acs.accounts.5b00293 [34] K. Lal, P. Rani Recent developments in copper nanoparticle-catalyzed synthesis of 1,4 disubstituted 1,2,3-triazoles in water Arkivoc, 2016, (i), 307-341. http://dx.doi.org/10.3998/ark.5550190.p009.593 [35] K. Lal, P. Yadav, A. Kumar, A. K. Paul Design, synthesis, characterization, antimicrobial evaluation and molecular modeling studies of some dehydroacetic acid-chalcone-1,2,3-triazole hybrids Bioorg Chem, 2018, 77, 236-244. https://doi.org/10.1016/j.bioorg.2018.01.016 [36] K. Lal, P. Yadav, A. Kumar Synthesis, characterization and antimicrobial activity of 4-((1benzyl/phenyl-1H-1,2,3-triazol-4-yl)methoxy)benzaldehyde analogues Med Chem Res, 2016, 25, 644-652. DOI 10.1007/s00044-016-1515-0 [37] S. Xu, X. Zhun, X. Pan, Z. Zhang, L. Duan, Y. Liu, L. Zhang, X. Ren, K. Ding 1-Phenyl-4benzoyl-1H-1,2,3-triazoles as orally bioavailable transcriptional function suppressors of estrogen-related receptor α. J Med Chem, 2013, 56, 4631-4640. https://doi.org/10.1021/jm4003928 [38] F. D. Spooner, G. Sykes 1972, (Academic Press), London. [39] Molinspiration Chemoinformatics Brastislava, Slovak Republic, available from: http://www.molinspiration.com/ cgibin/properties, 2014. [40] O. Trott, A. J. Olson, AutoDock Vina: improving the speed and accuracy of docking with a new scoring function, efficient optimization and multithreading J. Comput. Chem. 2010, 31, 455461. DOI 10.1002/jcc.21334 [41] Marvin Sketch 19.19. 0, 2019, Chem Axon, http://www.chemaxon.com. [42] E. F. Pettersen, T. D. Goddard, C. C. Huang, G. S. Couch, D. M. Greenblatt, E. C. Meng, T. E. Ferrin. UCSF Chimeraa visualization system for exploratory research and analysis. J. Comput. Chem. 2004, 25, 1605-12. https://doi.org/10.1002/jcc.20084 [43] Dassault Systèmes BIOVIA, Discovery studio visualizer v17.2.0.16349, San Diego: 2016. [44] X. H. Liu, J. Q. Weng, C. X. Tan, L. Pan, B. L. Wang, Z. M. Li Antibacterial action on the crude solvent extract of Piper Officenerum Asian J Chem, 2011, 23, 4031-4038. [45] N. B. Sun, J. Q. Fu, J. Q. Weng, J. Z. Jin, C. X. Tan, X. H. Liu Microwave Assisted Synthesis, Antifungal Activity and DFT Theoretical Study of Some Novel 1,2,4-Triazole Derivatives Containing the 1,2,3-Thiadiazole Moiety Molecules, 2013, 18, 12725-12739. https://doi.org/10.3390/molecules181012725 [46] M. N. Arshad, T. Mahmood, A. F. Khan, M. Zia-Ur-Rehman, A. M. Asiri, I. U. Khan, K. Ayub, A. Mukhtar, M. T. Saeed Synthesis, Crystal Structure and Spectroscopic Properties of 1,2Benzothiazine Derivatives: An Experimental and DFT Study Chin J Struct Chem 2015, 34, 1525. https://doi.org/10.3390/molecules20045851 [47] M. N. Arshad, A. M. Asiri, K. A. Alamry, T. Mahmood, M. A. Gilani, K. Ayub, A. S. Birinji Synthesis, crystal structure, spectroscopic and density functional theory (DFT) study of Page 18 of 19

N-[3-anthracen-9-yl-1-(4-bromo-phenyl)-allylidene]-N-benzenesulfonohydrazine Spectrochim Acta Part A Mol Biomol Spectrosc, 2015, 142, 364-374. https://doi.org/10.1016/j.saa.2015.01.101 [48] R. Galeazzi, C. Marucchini, M. Orena, C. Zadra Stereoelectronic properties and activity of some imidazolinone herbicides: a computational approach J Mol Struc-Theochem. 2003, 640, 191-200. https://doi.org/10.1016/j.theochem.2003.08.134 [49] L. Szabo, V. Chiş, A. Pirnau, N. Leopold, O. Cozar, S. Orosz Spectroscopic and theoretical study of amlodipine besylate J Mol Struct, 2009, 924, 385-392. doi:10.1016/ j.molstruc.2009.02.003 [50] M. Bartošková, Z. Friedl The Relationship Between the Heats of Formation and the Molecular Electrostatic Potentials of Polyazaarenes Cent Eur J Energy Mater, 2013, 10, 103112. [51] B. Honig, A. Nicholls Classical electrostatics in biology and chemistry Science 1995, 268, 1144-1149. DOI: 10.1126/science.7761829 [52] P. Politzer, J. S. Murray The fundamental nature and role of the electrostatic potential in atoms and molecules Theor Chem Acc, 2002, 108, 134-142. DOI 10.1007/s00214-002-0363-9 [53] C. A. Mebi DFT study on structure, electronic properties, and reactivity of cis-isomers of [(NC5H4-S)2Fe(CO)2] J Chem Sci, 2011, 123. 727-731. [54] N. Sukumar A Matter of Density: Exploring the Electron Density Concept in the Chemical, Biological, and Materials Sciences 2012, (Wiley, Hoboken, NJ) USA. [55] S. Špirtović-Halilović, M. Salihovic, H. Džudžević-Čančar, S Trifunovic, S. Roca, D. Softic, D. Završnik DFT study and microbiology of some coumarin-based compounds containing a chalcone moiety J Serb Chem Soc, 2014, 79, 435-443. DOI: 10.2298/JSC130628077S

Page 19 of 19

Highlights of the study: 1) A new library of antimicrobial 5-fluoroindoline-2,3-dione-linked triazoles synthesized under environmentally benign aqueous condition. 2) Compounds 4b, 4c, and 7b exhibited considerable antibacterial potency with the reference drug Ciprofloxacin. 3) Compound 4a, 4d, and 7c exhibited better potency for A. Niger than reference drug Fluconazole. 4) Molecular docking and Molinspiration data revealed that 5-fluoroisatin-triazoles 4a-e and 7a-e possessed good drug like properties. 5) In DFT study, FMOs revealed that incorporation of triazole in alkyne 2 improved the pharmacological activities of the resulted triazoles 4a-e and 7a-e.

Yours truly, Ram Kumar Tittal

Declaration of interests ☒ The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. ☐The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:

The authors declare no conflict of interest.