{A survey on the characterization and biological activity of isatin derivatives} J. Serb. Chem. Soc. 85 (8) 979–1000 (2020) UDC 547.756:57–188:615.28+542.9+ JSCS–5353 547.571’551:539.16+519.677 Review 979 REVIEW A survey on the characterization and biological activity of isatin derivatives SAŠA Ž. DRMANIĆ1#, PREDRAG PETROVIĆ2, DOMINIK R. BRKIĆ3, ALEKSANDAR D. MARINKOVIĆ1# and JASMINA B. NIKOLIĆ1*# 1Department of Organic Chemistry, Faculty of Technology and Metallurgy, University of Belgrade, Karnegijeva 4, 11000 Belgrade, Serbia, 2Department of Chemical Engineering, Faculty of Technology and Metallurgy, University of Belgrade, Karnegijeva 4, 11000 Belgrade, Serbia and 3Belgrade Polytechnic, Brankova 17, 11000 Belgrade, Serbia (Received 20 March, accepted 19 April 2020) Abstract: The derivatives of isatin have already been known to display a vari- ety of biological activities. Therefore, the studies on their activity and its rel- ation to structure have recently become a popular subject for investigation. The examined compounds were synthesized by the reaction of isatin and substituted primary amines and characterized by spectroscopic methods. The investigation of the antimicrobial and antioxidative activity of the synthesized compounds was performed by broth microdilution method. As for the characterization of the investigated isatin based Schiff bases, the linear solvation energy relation- ships (LSER) were used to analyze the solvent influence on the UV absorption maxima shifts (νmax), using the well known Kamlet–Taft model and taking geometrical isomers into consideration when possible. Linear free energy relat- ionships (LFER) were used to analyze substituent effect on pKa, as well as NMR chemical shifts and νmax values. The antimicrobial activity and charac- terization were related using both experimental and theoretical methods. Keywords: antimicrobial activity; E/Z isomers; solvent effects; substituent effects; TD-DFT; 3D QSAR. CONTENTS 1. INTRODUCTION 2. ANTIMICROBIAL AND ANTIOXIDATIVE ACTIVITY OF ISATIN SCHIFF BASES 3. SOLVATOCHROMISM OF ISATIN BASED SCHIFF BASES: LSER AND LFER STUDY 4. DETAILED STRUCTURAL AND QUANTUM CHEMICAL STUDY RELATED TO ANTIMICROBIAL ACTIVITY 5. CONCLUSIONS * Corresponding author. E-mail: jasmina@tmf.bg.ac.rs # Serbian Chemical Society member. https://doi.org/10.2298/JSC200320020D ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. 980 DRMANIĆ et al. 1. INTRODUCTION The derivatives of isatin (indole-2,3-dione), its Schiff and Mannich bases, have been reported to show a variety of biological activities, such as anti- bacterial,1 antifungal2 and anti-HIV3,4 activities. The wide spectrum of isatin derivatives and their various chemical properties have led to their expanded use as precursors for the preparation of many biologically active compounds.5–11 Isatin and its metabolites are components of many natural substances and display a wide range of the already mentioned activities such as antiviral, anticancer, antibacterial, antituberculosis, antifungal and anticonvulsants.5–11 Our recently published study on the antimicrobial activity, as well as on the antioxidative activity of isatin derivatives showed their moderate to significant antimicrobial activity.12 To the best of our knowledge not much has been reported on the solvato- chromic effects of isatin derivatives, therefore the studies were performed by our team recently.13,14 It is already known that the absorption spectra in different solvents are often used for investigation of the solvatochromic effect of organic molecules. When absorption spectra are measured in solvents of different polar- ity, it is usually found that the positions, intensities, and shape of absorption bands are influenced by the solvent. Mostly used spectra which can provide information about solute-solvent are: UV–Vis, IR, 1H- and 13C-NMR spectra.15 In here presented studies,13,14 UV–Vis and NMR data were analysed by the use of LSER and LFER models, in order to evaluate the influence of the solvent/ /solute interactions and substituent effects, respectively. Quantification of the sol- vent effects: dipolarity/polarizability and the hydrogen-bonding ability on the UV spectral shifts (νmax), were interpreted by means of the Kamlet–Taft (LSER)16 Equation: νmax = ν0 + sπ* + aα + bβ (1) Later on, LFER analysis was applied to the UV and NMR data in of studied compounds. The transmission of substituent effects from the substituent R (see Fig. 1) to the carbon atoms of interest were studied using equation:17–19 s hρσ= + (2) Again, the transmission of substituent effects, i.e., LFER study, was dis- cussed in relation to the geometry of the molecules obtained optimized by density functional theory (DFT) calculations.13 Furthermore, it is known that Schiff bases show E/Z isomerisation caused by the presence of imino group, which was analysed in the next part of our inves- tigation.14 For example, β-phenylethylamines in the reaction with isatin form the mixtures of the two stereoisomers E and Z, which can be thermodynamically or kinetically controlled, but their ratios may vary, depending on compounds struc- ture and conditions.20–24 It was of interest to find out which of all the mentioned ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. A SURVEY ON ISATIN DERIVATIVES 981 characteristics of the isatin derivatives have the impact on their biological activity. Fig. 1. The chemical structure of isatin derivatives with the labelling of π-electronic units.13 2. ANTIMICROBIAL AND ANTIOXIDATIVE ACTIVITY OF ISATIN SCHIFF BASES The examined Schiff bases were synthesized by the reaction of isatin and a corresponding primary amine,12 Scheme 1, and the general formula is given in Fig. 1.13 The list of synthesized compounds is given in Table I. N H O N R N H O O + NH2R CH3OH Scheme 1. Isatin derivatives synthesis. TABLE I. The synthesized derivatives of isatin12 No. R Compound name 2.1 N N S SH 1,3-Dihydro-3-[(2,4,5-thiadiazolone-2(3H)-thione)imino]-2H-indol-2-on 2.2 N S 1,3-Dihydro-3-[(2-benzothiadiazole) imino]-2H-indol-2-on 2.3 CN 1,3-Dihydro-3-[(4-cyanophenyl)imino]-2H-indol-2-one 2.4 N S NO2 1,3-Dihydro-3-[(5-nitro-2-thiazolyl)imino]-2H-indol-2-one 2.5 N CH3 1,3-Dihydro-3-[(4-methyl-2-pyridyl)-2-imino]-2H-indol-2-on 2.6 NO2 1,3-Dihydro-3-[(4-nitrophenyl)imino]-2H-indol-2-one ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. 982 DRMANIĆ et al. The compounds 2.1 and 2.5 were not synthesized before.2 Compound 2.2 has already been used for metal complexes synthesis,25 compound 2.3* is com- mercially available, compounds 2.426 and 2.627 have also already been used for research of their antimicrobial activity. The structure of all synthesized com- pounds (2.1–2.6) from Table I was confirmed by melting points, FTIR, 1H- and 13C-NMR spectra and also by elemental analysis for new compounds (2.1 and 2.5).12 The antimicrobial activity of all synthesized compounds 2.1–2.6 was deter- mined on a wide range of different microorganisms (Table II) by the broth micro- dilution method.28 TABLE II. The examined bacteria and fungi types12 No. Microorganism ATCC No. 1 Staphylococcus aureus 6538 2 Lysteria monocytogenes 19115 3 Enterococcus faecalis 29212 4 Shigella sonnei 29930 5 Salmonella enteritidis 13076 6 Yersinia enterolitica 27729 7 Escherichia coli 35150 8 Proteus hauseri 13315 9 Pseudomonas aeruginosa 27853 10 Candida albicans 10259 All examined compounds have shown considerable activity against all tested microorganisms except for compound 2.3, which has shown rather weak activity against E. coli, P. aeruginosa and C. albicans, in the range of investigated con- centrations. Generally, the examined activity can be described as moderate with some selectivity against Gram-positive (G+) or Gram-negative (G–) strains of bacteria, or yeast C. albicans. The selectivity to G– bacteria is of importance, as it enables the antibiotic agent based on a G– selective compound to be taken without the support of an agent that recovers the gastrointestinal tract, because it contains G+ natural bacteria. The activity of certain examined isatin derivatives against fungi is also important for they can be applied as antifungal agents. The overall results of the antimicrobial screening are given in Table III. Some of the isatin derivatives synthesized in this research have displayed significant activity against various examined bacteria and fungi, while the others were only moder- ately or even weakly active. Compound 2.1 has shown the most prominent overall activity on both G+ (S. Aureus and L. monocytogenes) and G– bacterial strains, (Y. enterolitica and P. hauseri). The highest activity of this compound was noticed against the S. Aureus * Scientific Exchange, Inc., 105 Pine River Road, P.O. Box 918, Center Ossipee, NH 03814, USA. ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. A SURVEY ON ISATIN DERIVATIVES 983 (MIC value of 0.16 mM) and close to it against L. monocytogenes (MIC value of 0.32 mM). However, compound 2.1 has displayed only moderate activity on Y. enterolitica, unlike compound 2.5, which was significantly active against the same strain (MIC value of 0.33 mM). Except the above mentioned, the overall antimicrobial activity of compound 2.5 is not as strong as that of compound 2.1. Besides Y. enterolitica, it has also shown some moderate activity on S. sonei and somewhat slighter on P. hauseri. Compound 2.6 behaved somewhat similarly to compound 2.5 but without any really prominent antimicrobial activity. It has displayed only moderate activity to Y. enterolitica and S. sonei. Compounds 2.2 and 2.3 have generally shown comparably weak antimicrobial activity. The only observation that can be of interest is the relative selectivity of compound 2.3 to L. monocytogenes (MIC value of 0.63 mM). Compound 2.4 can be noticed for cer- tain moderate activity against S. aureus, E. Faecalis and also S. sonei.12 TABLE III. Antimicrobial activity of examined compounds (MIC / mM)12 Microorganism Compound 2.1 2.2 2.3 2.4 2.5 2.6 1. S. aureus 0.16 2.24 2.53 0.57 5.27 4.68 2. L. monocytogenes 0.32 1.12 0.63 2.28 2.64 1.17 3. E. faecalis 2.56 2.24 2.53 0.57 2.64 4.68 4. S. sonnei 1.28 1.12 5.06 0.57 0.66 0.59 5. S. enteritidis 2.56 1.12 >5.06 1.14 5.27 4.68 6. Y. enterolytica 0.64 1.12 2.53 1.14 0.33 0.59 7. E. coli 2.56 1.12 >5.06 1.14 2.64 4.68 8. P.Hauseri 0.64 1.12 5.06 1.14 0.99 1.17 9. P.Aeruginosa 5.12 2.24 >5.06 2.28 2.64 2.34 10. C. Albicans 2.56 2.24 >5.06 2.28 2.64 1.17 Furhermore, all the synthesized compounds were screened for antioxidative activity by the diphenylpycrilhydrazyl radical (DPPH).29 The results of DPPH analysis have shown that the most prominent antioxidative activity displays com- pound 2.1, while the other investigated specimens, including pure isatin, have shown very slight if any activity. With the increase of the concentration of compound 2.1, the absorbance of DPPH was decreased, displaying linear dependence of DPPHred in the range of examined concentrations (c / mM), which are described by the following equat- ion:12 DPPHred / % = 5.099 + (101.02±5.24)c (3) r= 0.995, s= 3.17, n = 6 DPPHred is actually the percent of DPPH reduction and c is the concen- tration of compound 2.1, given in mM. This equation enables the precise deter- mination of the concentration which reduces 50 % of DPPH concentration ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. 984 DRMANIĆ et al. (DC50). Compound 2.1 showed prominent antioxidative activity, with DC50 value of 0.444 mM.12 3. SOLVATOCHROMISM OF ISATIN BASED SCHIFF BASES: LSER AND LFER STUDY Another series of eleven isatin derivatives (1,3-dihydro-3-arylimino-2H- -indol-2-one), including the compound 2.5 from the previous one (compound 3.7 in this series) was synthesized and the list of compounds is given in Table IV.13 The characterisation of the synthesized isatin derivatives is described in the ori- ginal paper.13 TABLE IV. The list of synthesized isatin derivatives13 Compd. Compound Substituent R 3.1 1,3-Dihydro-3-(phenylimino)-2H-indol-2-one 3.2 1,3-Dihydro-3-[(2-bromophenyl)imino]-2H-indol-2-one Br 3.3 1,3-Dihydro-3-[(3-bromophenyl)imino]-2H-indol-2-one Br 3.4 1,3-Dihydro-3-[(4-bromophenyl)imino]-2H-indol-2-one Br 3.5 1,3-Dihydro-3-[(3-methyl-2-pyridinyl)imino]-2H-indol-2-one N CH3 3.6 1,3-Dihydro-3-[(4-methyl-2-pyridinyl)imino]-2H-indol-2-one N CH3 3.7 1,3-Dihydro-3-[(5-methyl-2-pyridinyl)imino]-2H-indol-2-one N CH3 3.8 1,3-Dihydro-3-[(6-methyl-2-pyridinyl)imino]-2H-indol-2-one N CH3 3.9 1,3-Dihydro-3-(3-quinolinylimino)-2H-indol-2-one N 3.10 1,3-Dihydro-3-(6-quinolinylimino)-2H-indol-2-one N 3.11 1,3-Dihydro-3[2-(8-hydroxy)quinolinyl imino]-2H-indol-2-one N OH The UV absorption spectra were recorded in the range from 200 to 600 nm in 22 solvents of different polarity, in order to study substituent effect on the ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. A SURVEY ON ISATIN DERIVATIVES 985 solvatochromism of the investigated compounds. The absorption spectra of the isatin derivatives 3.1–3.11, indicate the presence of two bands, corresponding to different electronic transition of the investigated compounds. The examples of the UV–Vis absorption spectra of compounds 3.1–3.11 in acetone, acetonitrile (AcN), benzyl alcohol (BzOH) and N,N-dimethylformamide (DMF) are given in Fig. S-1 of the Supplementary material to this survey,13 and the complete results are given in the original paper.13 The shifts of νmax in UV absorption spectra showed relatively weak dependence on the both solvent and substituent effects compared to those of the parent compound 3.1. The data13 indicate that the values of absorption frequencies of the investigated compounds depend on the electronic effect of the substituent present on C3 position of indole-2-one core. The introduction of Br as a substituent in C4 position of the phenyl ring (com- pound 3.4) and all quinoline derivatives (compounds 3.9–3.11) bring about the positive solvatochromism, comparing to the unsubstituted compound. The other compounds showed νmax shifting to higher wavelength in 2-chloroethanol, 2-methoxyethanol, DMF, DMAc and AcN, while in other solvents no rule could be observed (irregular behaviour regarding both substituent and solvent). The highest batochromic shifts was found for compoubd 3.9 in almost all solvents used. The Kamlet–Taft solvent parameters are taken from literature30 as well as substituent constants used in LSER and LFER correlations.19 The presented data confirm that the positions of the UV–Vis absorption frequencies depend on the nature of the solvent used and the substituent present at the aryl or phenyl ring at the imino group of the examined isatin derivatives.13 The results are presented in the original paper.13 According to the correlation results, obtained by the use of Kamlet–Taft equation,13 alternation of solvatochromic coefficients with respect to solvent/sub- stituent effects exists. The positive sign of s and a coefficients for isatin derivatives (compounds 3.1, 3.3–3.5 and 3.9–3.11) indicates a hypsochromic shifts with increasing solvent dipolarity/polarizability and hydrogen-bond donor capability. Largest hypsochromic shift of the absorption maxima, regarding coefficient s and a, were found for the compounds 3.5 and 3.9, respectively. The positive values of s and a coefficients suggest better stabilization of the ground state relative to the electronic excited state with the increasing solvent polarity, i.e., higher dipolar properties of the molecule in the ground state, with more pronounced HBA (hyd- rogen bond acceptor) properties of solvated molecules.13 The percentage contri- bution of solvatochromic parameters for all isatin derivatives, showed the highest Pπ value of compound 3.5 (71.01 %), with lowest Pα value (2.28 %). Oppositely could be noticed for compound 3.1 with highest value for Pβ (70.72 %) and lowest Pπ value (8.88 %). Highest Pα value was obtained for comp. 3.4 (51.28 %).13 From the correlation analysis obtained by the Kamlet–Taft equation, the negative sign of s and a coefficients, except for comp. 3.9 (positive values of ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. 986 DRMANIĆ et al. 1.57 and 1.42, respectively), were found indicating a batochromic shift with the increasing solvent dipolarity/polarizability.13 This suggests better stabilization of the electronic excited state relative to the ground state, i.e., higher dipolarity/pol- arizability and HBD (hydrogen bond properties of solvated molecules in the excited state. Alternation of the sign of coefficients b, positive values found for compounds 3.3, 3.4, 3.8 and 3.11 indicate better stabilization of the isatin derivat- ives in the ground state, in other words higher HBA solvent interaction with sol- ute molecule was found. For the other compounds the opposite behaviour was noticed. The non-specific solvent effect is a factor of the highest contribution to UV–Vis spectral shifts of all the investigated compounds, except for compound 3.9 with the highest contribution of HBA solvent effect (45.04 %), and com- pound 3.11 with the highest contribution of Pα (44.15 %). The highest value of Pπ was obtained for compound 3.5 (65.41 %). HBA solvent effect is of lower contribution and the highest values of solvent HBA effect was found for compound 3.9 (–2.45) and exceptionally low values were found for compounds 3.6–3.8 (–0.05, –0.06 and 0.02, respectively). The presented results, obtained using the Kamlet–Taft model, indicated that the solvent effects on UV–Vis absorption spectra of the investigated isatin derivatives are rather complex, due to the diversity of the contribution of both solvent and substituent effect in studied compounds. This also indicated that the electronic behaviour of the nitrogen atoms in indol-2-on moiety is significantly different between derivatives with high contribution of localized HBA effect, that originates from the electron- accepting quinoline and the methyl substituted pyridyl groups.13 The LFER concept was applied to the νmax and SCS values of isatin derivat- ives with the aim to get an insight into substituent electronic effect on the absorp- tion maxima shifts and NMR chemical shifts. An analysis, using LFER principles in the form of the Eq. (2) was performed. The Hammett substituent constants are given in the literature,13 and the obtained correlation results are presented in the original paper.13 The observed ρ values indicate different susceptibilities of the chemical shifts to substituent effects. The correlations were of reasonably good to high quality, which means that the SCS values reflect the electronic substituent effects correctly. Evidently the chemical shifts of C1’ showed an increased sus- ceptibility and normal substituent effect. The presented results indicate that the contribution of both substituent effects, electron-donating and electron-accepting, have significant influence on electron-density shift in overall investigated molecule. The effectiveness of the transmission of substituent effects was determined by the conformational change of the investigated molecules which originate from the out-of-plane rotation of the aryl substituent, with respect to indole-2-one plane, defined by the torsion angle θ (Fig. 1). The reverse substituent effect was observed at H1 and C2 car- bon. The existence of these correlations was interpreted as an evidence of sub- ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. A SURVEY ON ISATIN DERIVATIVES 987 stituent effect on the change of the electronic density over investigated molecule. The presence of an electron-accepting –C=O group at C2 position, and electron- donating amino group (N1-H), as a part of π1-electronic system of amide group, contribute to the intensive electronic interactions which exists in 3-(substituted imino)-2-pyrrolidone moiety.31 The correlations for C2 showed negative values of correlation coefficients ρ. The negative sign of the reaction constant, ρ, indicates a certain reverse behaviour, i.e., the value of SCS decreases when the electron-withdrawing ability of the substituents, measured by σ, increases. Such finding clearly reflect the behaviour opposite to normal polarization in keto group which comes from the contribution of two opposite effects: electron- accepting C2=O group and aryl substituents with the participation of the elec- tron-donating effect of the indole nitrogen. Magnitude of this effect strongly dep- ends on the electron-accepting character of aryl substituents which cause the ade- quate/proportional reverse π-electron density shift from C2=O keto group. The contribution of the resonance interaction depends on spatial arrangement of the aryl moiety, and thus, the favourable conformation provides the effective trans- mission of the resonance effect to C2 and C3 carbons. According to that, it is expected that the presence of the electron-donating group support normal polar- ization of C2=O, and the electron-accepting group exert opposite effect, which is reflected in the reverse polarization. The normal polarization at C3 carbon is a reflection of the contribution of electronic effects in both π-electronic units (Fig. 1), and primarily dictated by polarization of C=N imino bond which is more or less disturbed by change of substituent at phenyl (aryl) moiety. It is obvious that the chemical shifts of C1' show an increased susceptibility and normal substituent effect related to the substituent constant.13 The correlation results of UV–Vis absorption maxima of the investigated compounds, obtained by the use of LFER principles indicate the influences of both solvent and substituent effect on UV–Vis absorption maxima shift. These results indicate that the solvent effects: dipolarity/polarizability, HBD and HBA abilities cause the appropriate sensitivity of the position of absorption maxima (νmax) to substituent effect.13 Highly dipolar aprotic solvent DMF, and solvent with low HBD effect, AcN, as well as those with high HBD effect, like ethanol, contribute to the opposite behaviour of the absorption frequencies shift at higher wavelength with respect to the substituent effects. Aprotic solvents do not stabilize anions well, while they usually stabilize larger and more dispersible positive charges better. Lower con- tribution of substituent effects in a solvent with higher relative permittivity can be explained by the fact that highly dipolar surrounding medium suppresses elec- tron density shift inducing lower susceptibility of the absorption maxima shift to electronic substituent effects. Similar behaviours were found for AcN and etha- nol, and it could be observed that solvent dipolarity/polarizability is the most sig- ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. 988 DRMANIĆ et al. nificant effect which contribute to the reverse relationship of νmax to substituent effects. The opposite substituent effect on νmax change was found in THF.13 In the two solvents, which showed HBD properties, namely AcN and etha- nol, the negative correlation slopes of lower wavelength peak was obtained for the former. Somewhat higher sensitivity of νmax to substituent effect was found for AcN (first set of solvents), and THF. In the DMF, ethanol and THF negative solvatochromism was obtained, indicating better stabilization of ground state of investigated compounds. Somewhat lower values and similar trend of correlation slope was found for ethanol, due to lower contribution of solvent dipolarity/pol- arizability effect. This result suggests that the transmission of electronic substi- tuent effects significantly depends on the conformation of studied molecules and solvent properties. The presented results showed that the transmission of sub- stituent electronic effects through π-resonance units takes place by balanced con- tribution of two modes: through localized π-electronic unit and overall conjug- ated system of investigated compounds. Their contribution depends on substitut- ion pattern, as well as solvent properties. This fact implies that the electron den- sity change is of localized/extended delocalization phenomena in compounds with the electron-acceptor substituents. The consequence of this is a lower sub- stituent effect in a solvent with higher hydrogen bond accepting ability.13 An additional analysis of solvent and substituent effects on the measured absorption frequencies and the conformational changes of the studied com- pounds, needed theoretical calculations, i.e., geometry optimization and therefore the molecular electrostatic potential (MEP) analysis were performed.13 The ground state geometries of compounds 3.1–3.11 were fully optimized with DFT method. The theoretical absorption spectra of compound 3.1 were calculated in acetone, acetonitrile, ethanol, tetrahydrofuran, dimethylsulfoxide, formamide and toluene with time-dependent density functional theory (TD-DFT) method.13 Geometry optimization of the investigated molecules was performed using B3LYP functional with 6-311G(d,p) basis set.13 The most stable conformations of compounds 3.1–3.11 are presented in Fig. S-2 of the Supplementary material. The elements of optimized geometries of calculated compounds are given in the original paper. The theoretical absorption spectra of compound 3.113 were calcul- ated in acetone, acetonitrile, ethanol, tetrahydrofuran, dimethylsulfoxide, form- amide and toluene by TD-DFT method, and showed very good agreement with the experimental data. The calculation of the optimal geometry, with the focus on determination of the value of torsion angle θ (Fig. 1), gives valuable results required for better understanding of the transmission of substituent effects, i.e., electron density dis- tribution. In the investigated molecules, these values are different and mostly depend on substituent present. Somewhat larger deviation of θ was noticed for compounds 3.2, 3.5–3.8 and 3.11, indicating the significance of the extended ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. A SURVEY ON ISATIN DERIVATIVES 989 resonance interaction in the electron-donor substituted compounds. Oppositely, in electron-acceptor substituted compounds the appropriate contribution of n,π- -conjugation (nitrogen lone pair participation) to overall electronic interaction with π-electronic system of pyridone unit causes the perturbation of π-electron density. The geometric elements of the substituted isatin derivatives turned out to be similar to those for the unsubstituted one. The introduction of mainly electron- accepting substituent causes a decrease of C3=N bond length, i.e., a substituent supports normal polarization in the imino group, causing the appropriate shift of electron density to nitrogen. The presence of an electron-donating group attached to a complex structure of aryl moiety contributes to the electron density shift from the phenyl ring to the indol moiety, causing the increased molecule planar- ization. A decrease of the C2=O lengths is caused by the superposition of two effect: normal polarization in C2=O bond and the opposite effect of the electron- -withdrawing group at C3 position; the consequence of this is a slight shifting of π-electron density to carbonyl C atom. Therefore the length of N1–C2 gets longer, comparing to compound 3.1 as a result of the suppression of amide type of resonance.13 The larger deviation from the planarity was found for compounds 3.2, 3.7, 3.8 and 3.11 which is associated with ortho-effect of 2-Br in compound 3.2, presence of the electron-donating methyl group situated at pyridyl in differ- ent position (compounds 3.7 and 3.8) as well as the strong electron-donating hydroxyl group in compound 3.11. The two opposite electron accepting effects operate all in investigated compounds: the electron-accepting aryl substituents and indole-2-one core which influence the appropriate geometrical adjustment of these molecules, as a response to the electronic demand of the electron deficient environment. Except for the vicinity of the electron density at C2=O and nitrogen atom in compounds 3.5–3.8 and 3.11, which are also contributing factor to the increased deviation from planarity, due to the repulsion of negative potential pre- sent at these molecular fragments.13 The variation of substituent properties clearly indicates that the contributions of both conformations and donor–acceptor character are involved in the electronic transition of investigated compounds. The electron density of the investigated compounds is presented using MEP analysis, which was used to evaluate the charge distribution over investigated compounds and to illustrate the three dimensional charge distributions overall the investig- ated molecules. MEP potential, at a point in space around a molecule, gives the information about the net electrostatic effect produced at that point by the total charge distribution (electron + proton) of the molecule and correlates with dipole moments, electro-negativity, partial charges and chemical reactivity of the mole- cules. An electron density iso-surface mapped with electrostatic potential surface depicts the size, shape, charge density and the site of chemical reactivity of the molecules. MEP shown in Fig. S-3 of the Supplementary material illustrates the three dimensional charge distributions overall the investigated molecules. As it ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. 990 DRMANIĆ et al. can be seen from the Fig. S-3, the different values of the electrostatic potential at the surface are represented by different colours; red represents regions of the most electronegative electrostatic potential, it indicates the region of high elec- tron density, i.e., the sites favourable for electrophilic attack; blue represents regions of the most positive electrostatic potential, i.e., region of low electron density favourable for nucleophilic attack, and green represents regions of zero potential. The potential increases in the order: red1250 1250 1250 1250 1250 >1250 1250 1250>1250 1250 L. mono- cytegenes 1250 313 >1250>1250 1250 1250 313 1250 1250 1250 313 1250 313 S. sonnei 625 313 1250 625 625 625 313 625 1250 >1250 313 1250 156 Y. entero- colytica 1250 156 >1250 313 1250 625 313 625 1250 1250 156 1250 156 E. coli 1250 1250>1250>1250>1250 1250 1250>1250>1250>12501250 1250 1250 P. hauseri 1250 313 >1250 1250 1250 1250 313 1250 1250 156 313 1250 313 P. aeru- ginosa 625 1250>1250>1250>1250>12501250>1250 1250 1250 625 1250 625 C. neofor- mans 156 19 39 39 156 >1250 39 156 156 19 39 156 39 C. albicans 1250 313 >1250>1250>1250>1250 313 625 1250 1250 1250 1250 313 ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. A SURVEY ON ISATIN DERIVATIVES 997 vatives against most of the strains. While former are electron-donors and latter an electron-acceptors, they are all electron-negative groups, which could be an imp- ortant property for the antimicrobial activity. The difference is most pronounced in the case of compound 4.2 (2-OH) and compound 4.3 (3-OH) activity against Y. enterocolitica, which is more than 10 times greater in former (MIC = 156 mM for 2-OH and MIC > 1250 mM for 3-OH).14 There is no difference in activity against G+ and G– bacteria in general, but some strains are more susceptible, as L. monocytogenes, S. sonnei, Y. entero- colitica and P. hauseri.14 As for fungal strains, C. neoformans was shown to be far more sensitive to the tested compounds than C. albicans, while the MIC values for C. albicans were similar to those for bacterial strains. The MIC values obtained for C. neo- formans were, in some cases, even 100 times greater (compound 4.2; 2-OH and compound 4.11; 2-I, MIC = 0.019 mM). Compounds 4.3 (3-OH), 4.4 (4-OH), 4.8 (2-Cl), 4.14 (2-NO2), 4.16 (4-NO2) also showed prominent antifungal activity against this human pathogen (MIC = 0.038 mM).14 The overall activity of the compounds was most pronounced against the fore mentioned C. neoformans. Compounds 4.2 (2-OH) and 4.8 (2-Cl) showed the best activity against C. albi- cans (MIC = 0.313 mM) of all investigated compounds. The obtained results sug- gest that the other substituents present on phenyl or heterocyclic ring of the isatin derivatives should be tested.14 In order to explore the structural properties important for the antimicrobial activity of compounds 4.1–4.16, QSAR models were generated, taking the MIC values for S. sonnei, Y. enterocolitica as well as C. neoformans but the statis- tically significant model was obtained only for the activity of compounds toward C. neoformans.14 Principal component analysis (PCA) was performed using the whole set of GRIND-2 descriptors. Model with 3 principal components (PC) explained 77.78 % of X sum of squares (SSXacc), and 70.22 % of X variance (VarXacc).14 Partial least squares regression model (PLS) was created in order to correlate he structural features of compounds with antimicrobial potency toward C. neoformans. After filtering of descriptors through 2 cycles of FFD, two latent variables (LV) model with r2 = 0.89, q2 (leave-one-out, LOO cross-validation) = = 0.69, and standard deviation of error of prediction (SDEP) = 0.29.14 From the PLS coefficients plot (Fig. S-6 of the Supplementary material) several variables important for the activity were identified. The most informative variables in PLS model are the one that clearly separate the more potent com- pounds from the less. Structural motifs associated with the most informative vari- ables were found.14 The positively correlated variables are depicted on com- pound 4.2 as a model for the more potent compounds (Fig. S-7 of the Supple- mentary material). The variables with the negative influence on potency are ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. 998 DRMANIĆ et al. depicted on the least potent compound 4.6 (Fig. S-8 of the Supplementary mat- erial).14 5. CONCLUSIONS A series of 30 isatin derivatives, some of them new compounds, were syn- thesised in order to examine their properties and the application potential. Their structures were confirmed by spectral methods. The antimicrobial screening was performed, and some compounds showed relative selectivity to certain bacterial strains. In order to test antioxidative activity, DPPH reduction test was performed on some of the compounds, which should be continued. From the solvatochromic and characterization point of view UV–Vis and NMR data were analysed by the use of LSER and LFER principles and it could be concluded that solvent effects have significant influence on the transmission mode of substituent effects. The quantum chemical calculations, performed next, showed that the present substi- tuents significantly change the extent of conjugation. Then, the E/Z isomer ratios of different investigated isatin compounds were estimated from the point of NMR data, theoretical calculations and UV–Vis spectra. The inclusion of solvent effects in the TD-DFT calculations demonstrated that substituents, depending on their position in molecules, and solvent effects significantly affect the ICT char- acter. The testing of antimicrobial activity was continued further on isatin deri- vatives, with the aim of relating the activity to the compound structure. The most of the investigated isatin derivatives exhibited moderate activity against bacterial strains, but compounds with hydroxy, nitro group and chlorine in ortho position as well as para nitro showed considerable activity. The 3D QSAR model created pointed on hydroxyl groups as substituents important for the potency of com- pounds toward fungal strain C. neoformans. This complex research was performed to study the examined compounds for their potential biological activity as well as to link activity with molecular struc- ture. The aim was to determine compounds that should be submitted to further investigation, since they have a considerable chance for practical use, particularly as active compounds of medicines. Acknowledgement. This work was supported by the Ministry of Education, Science and Technological development of the Republic of Serbia (Contract No.451-03-68/2020-14/ /200135). SUPPLEMENTARY MATERIAL Additional data are available electronically at the pages of journal website: https:// //www.shd-pub.org.rs/index.php/JSCS/index, or from the corresponding author on request. ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. A SURVEY ON ISATIN DERIVATIVES 999 И З В О Д ПРЕГЛЕД КАРАКТЕРИЗАЦИЈЕ И БИОЛОШКЕ АКТИВНОСТИ ДЕРИВАТА ИСАТИНА САША Ж. ДРМАНИЋ1, ПРЕДРАГ ПЕТРОВИЋ2, ДОМИНИК Р. БРКИЋ3, АЛЕКСАНДАР Д. МАРИНКОВИЋ1 и ЈАСМИНА Б. НИКОЛИЋ1 1Катедра за органску хемију, Технолошко–металуршки факултет, Универзитет у Београду, Карнегијева 4, 11000 Београд, 2Катедра за хемијско инжењерство, Технолошко–металуршки факултет, Универзитет у Београду, Карнегијева 4, 11000 Београд и 3Београдска политехника, Бранкова 17, 11000 Београд Деривати исатина су познати по потенцијалној биолошкој активности, па су постали интересантни за проучавање везе између наведене активности и њихових структурних карактеристика. Наш тим их је годинама истраживао са оба аспекта и у овом преглед- ном раду су приказани сви резултати од значаја до којих смо дошли. Испитивани дери- вати исатина су синтетисани реакцијом између исатина и различитих примарних амина, окарактерисани су спектроскопским методама (FTIR, NMR) као и елементалном ана- лизом. На њима су извршена и разна теоријска проучавања као и одређивање антимик- робне активности, како би се стекла што потпунија слика о вези између њихове струк- туре и активности и одредила једињења за даља испитивања због њиховог потенцијала за примену. (Примљено 20. марта, прихваћено 19. априла 2020) REFERENCES 1. R. W. Daisley, V. K. Shah, J. Pharm. Sci. 73 (1984) 407 (https://doi.org/10.1002/jps.2600730333) 2. E. Piscopo, M. V. Diurno, R. Gogliardi, M. Cucciniello, G. Veneruso, Boll. Soc. Ital. Biol. Sper. 63 (1981) 827 3. S. N. Pandeya, D. Sriram, E. De. Clercq, C. Pannecouque, M. Witvrouw, Ind. J. Pharm. Sci. 60 (1999) 207 (http://www.ijpsonline.com/articles/antihiv-activity-of-some-mannich- bases-of-lsatin-derivatives.pdf) 4. V. A. Muthukumar, H. C. Nagaraj, D. Bhattacherjee, S. George, Int. J. Pharm. Pharm. Sci. 5 (Suppl. 3) (2013) 95 5. R. S. Varma, I. A. Khan, Ind. J. Med. Res. 67 (1978) 315 6. F. D. Popp, H. J. Pajouhesh, Pharm. Sci. 17 (1988) 1052 7. R. S. Varma, W. L. Nobles, J. Pharm. Sci. 64 (1975) 881 (https://doi.org/10.1002/jps.2600640539) 8. F. D. Popp, R. Parson, B. E. Donigan, J. Heterocycl. Chem. 17 (1980) 1329 (https://doi.org/10.1002/jhet.5570170639) 9. F. Kontz, Sci. Pharm. 41 (1973) 123 10. F. D. Popp, F. P. Silver, A. C. Noble, J. Med. Chem. 10 (1967) 986 (https://pubs.acs.org/doi/pdf/10.1021/jm00317a074) 11. P. Pakravan, S. Kashanian, M. M. Khodaei, F. J. Harding, Pharmacol. Rep. 65 (2013) 313 (https://doi.org/10.1016/S1734-1140(13)71007-7) 12. G. M. Šekularac, J. B. Nikolić, P. Petrović, B. Bugarski, B. Đurović, S. Ž. Drmanić, J. Serb. Chem. Soc. 79 (2014) 1347 (https://doi.org/10.2298/JSC140709084S) 13. D. R. Brkić, A. R. Božić, V. D. Nikolić, A. D. Marinković, H. Elshaflu, J. B. Nikolić, S. Ž. Drmanić, J. Serb. Chem. Soc. 81 (2016) 979 (https://doi.org/10.2298/JSC160119049B) 14. D. R. Brkić, A. R. Božić, A. D. Marinković, M. K. Milčić, N. Ž. Prlainović, F. H. Assaleh, I. N. Cvijetić, J. B. Nikolić, S. Ž. Drmanić, Spectrochim. Acta, A 196 (2018) 16 (https://doi.org/10.1016/j.saa.2018.01.080) ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. 1000 DRMANIĆ et al. 15. C. Reichardt, Solvents and Solvent Effects in Organic Chemistry, Wiley-VCH Verlag GmbH & Co. KGaA, Wienheim, 2004, p. 329 16. M. J. Kamlet, J. L. M. Abbound, R. W. Taft, in Progress in Physical Organic Chemistry Vol. 13, R.W. Taft, Ed., Wiley, New York, 1981, p. 485 (https://doi.org/10.1002/9780470171929.ch6) 17. L. P. Hammett, J. Am. Chem. Soc. 59 (1937) 96 (https://pubs.acs.org/doi/pdf/10.1021/ja01280a022) 18. O. Exner, in Advances in linear free energy relationship, N. B. Champan, J Shorter, Eds., Plenum Press, London, 1972, pp. 1–69 (ISBN 978-1-4615-8660-9) 19. C. Hansch, A. Leo, D. Hoekman, J. Med. Chem. 39 (1996) 1189 (https://doi.org/10.1021/jm950902o) 20. Y. Vélez, C. Díaz-Oviedo, R. Quevedo, J. Mol. Struct. 1133 (2017) 430 (https://doi.org/10.1016/j.molstruc.2016.12.039) 21. Q. X. Guo, Y. W. Liu, X. C. Li, L. Z. Zhong, Y. G. Peng, J. Org. Chem. 77 (2012) 3589 (https://doi.org/10.1021/jo202585w) 22. P. Davidovich, D. Novikova, V. Tribulovich, S. Smirnov, V. Gurzhiy, G. Melino, A. Garabadzhiu, J. Mol. Struct. 1075 (2014) 450 (https://doi.org/10.1016/j.molstruc.2014.07.008) 23. K. Jakusová, M. Cigáň, J. Donovalová, M. Gáplovský, R. Sokolík, A. Gáplovský, J. Photochem. Photobiol., A 288 (2014) 60 (https://doi.org/10.1039/C5RA06625E) 24. M. Cigáň, M. Gáplovský, K. Jakusová, J. Donovalová, M. Horváth, J. Filo, A. Gáplovský, RSC Adv. 5 (2015) 62449 (https://doi.org/10.1039/C5RA06625E) 25. Z. H. Chohana, H. Perveza, A. Raufb, K. M. Khanc, C. T. Supurand, J. Enzym. Inhib. Med. Chem. 19 (2004) 417 (https://doi.org/10.1080/14756360410001710383) 26. E. Piscopo, M. V. Diurno, F. Imperadrice, M. Cucciniello, G. Veneruso, Boll. – Soc. It. Biol. Sper. 62 (1986) 1441 27. J. Panda, V. J. Patro, B. Sahoo, J. Mishra, J. Nanoparticles (2013), Article ID 549502, http://dx.doi.org/10.1155/2013/549502 28. A. Espinel-Ingroff, A. Fothergill, M. Ghannoum, E. Manavathu, L. Ostrosky-Zeichner, M. Pfaller, M. Rinaldi, W. Schell, T. Walsh, J. Clin. Microbiol. 43 (2005) 5243 (https://doi.org/10.1128/JCM.43.10.5243-5246.2005) 29. K. Shimada, K. Fujikawa, K. Yahara, T. Nakamura, J. Agri. Food Chem. 40 (1992) 945 (https://pubs.acs.org/doi/pdf/10.1021/jf00018a005) 30. M. J. Kamlet, J. L. M. Abboud, M. H. Abraham, R. W. Taft, J. Org. Chem. 48 (1983) 2877 (https://pubs.acs.org/doi/pdf/10.1021/jo00165a018) 31. F. H. Assaleh, A. D. Marinković, J. Nikolić, N. Ž. Prlainović, S. Drmanić, M. M. Khan, B. Ž. Jovanović, Arab. J. Chem. 12 (2019) 3357 (https://doi.org/10.1016/j.arabjc.2015.08.014) 32. A. Albert, E. P. Serjeant, The Determination of Ionization Constants, 2nd ed., Chapman and Hall, London, 1971, p. 44. ________________________________________________________________________________________________________________________Available on line at www.shd.org.rs/JSCS/ (CC) 2020 SCS. << /ASCII85EncodePages false /AllowTransparency false /AutoPositionEPSFiles true /AutoRotatePages /None /Binding /Left /CalGrayProfile (Dot Gain 20%) /CalRGBProfile (sRGB IEC61966-2.1) /CalCMYKProfile (U.S. Web Coated \050SWOP\051 v2) /sRGBProfile (sRGB IEC61966-2.1) /CannotEmbedFontPolicy /Error /CompatibilityLevel 1.4 /CompressObjects /Tags /CompressPages true /ConvertImagesToIndexed true /PassThroughJPEGImages true /CreateJobTicket false /DefaultRenderingIntent /Default /DetectBlends true /DetectCurves 0.0000 /ColorConversionStrategy /CMYK /DoThumbnails false /EmbedAllFonts true /EmbedOpenType false /ParseICCProfilesInComments true /EmbedJobOptions true /DSCReportingLevel 0 /EmitDSCWarnings false /EndPage -1 /ImageMemory 1048576 /LockDistillerParams false /MaxSubsetPct 100 /Optimize true /OPM 1 /ParseDSCComments true /ParseDSCCommentsForDocInfo true /PreserveCopyPage true /PreserveDICMYKValues true /PreserveEPSInfo true /PreserveFlatness true /PreserveHalftoneInfo false /PreserveOPIComments true /PreserveOverprintSettings true /StartPage 1 /SubsetFonts true /TransferFunctionInfo /Apply /UCRandBGInfo /Preserve /UsePrologue false /ColorSettingsFile () /AlwaysEmbed [ true ] /NeverEmbed [ true ] /AntiAliasColorImages false /CropColorImages true /ColorImageMinResolution 300 /ColorImageMinResolutionPolicy /OK /DownsampleColorImages true /ColorImageDownsampleType /Bicubic /ColorImageResolution 300 /ColorImageDepth -1 /ColorImageMinDownsampleDepth 1 /ColorImageDownsampleThreshold 1.50000 /EncodeColorImages true /ColorImageFilter /DCTEncode /AutoFilterColorImages true /ColorImageAutoFilterStrategy /JPEG /ColorACSImageDict << /QFactor 0.15 /HSamples [1 1 1 1] /VSamples [1 1 1 1] >> /ColorImageDict << /QFactor 0.15 /HSamples [1 1 1 1] /VSamples [1 1 1 1] >> /JPEG2000ColorACSImageDict << /TileWidth 256 /TileHeight 256 /Quality 30 >> /JPEG2000ColorImageDict << /TileWidth 256 /TileHeight 256 /Quality 30 >> /AntiAliasGrayImages false /CropGrayImages true /GrayImageMinResolution 300 /GrayImageMinResolutionPolicy /OK /DownsampleGrayImages true /GrayImageDownsampleType /Bicubic /GrayImageResolution 600 /GrayImageDepth -1 /GrayImageMinDownsampleDepth 2 /GrayImageDownsampleThreshold 1.33333 /EncodeGrayImages true /GrayImageFilter /DCTEncode /AutoFilterGrayImages true /GrayImageAutoFilterStrategy /JPEG /GrayACSImageDict << /QFactor 0.15 /HSamples [1 1 1 1] /VSamples [1 1 1 1] >> /GrayImageDict << /QFactor 0.15 /HSamples [1 1 1 1] /VSamples [1 1 1 1] >> /JPEG2000GrayACSImageDict << /TileWidth 256 /TileHeight 256 /Quality 30 >> /JPEG2000GrayImageDict << /TileWidth 256 /TileHeight 256 /Quality 30 >> /AntiAliasMonoImages false /CropMonoImages true /MonoImageMinResolution 1200 /MonoImageMinResolutionPolicy /OK /DownsampleMonoImages true /MonoImageDownsampleType /Bicubic /MonoImageResolution 1200 /MonoImageDepth -1 /MonoImageDownsampleThreshold 1.50000 /EncodeMonoImages true /MonoImageFilter /CCITTFaxEncode /MonoImageDict << /K -1 >> /AllowPSXObjects false /CheckCompliance [ /None ] /PDFX1aCheck false /PDFX3Check false /PDFXCompliantPDFOnly false /PDFXNoTrimBoxError true /PDFXTrimBoxToMediaBoxOffset [ 0.00000 0.00000 0.00000 0.00000 ] /PDFXSetBleedBoxToMediaBox true /PDFXBleedBoxToTrimBoxOffset [ 0.00000 0.00000 0.00000 0.00000 ] /PDFXOutputIntentProfile () /PDFXOutputConditionIdentifier () /PDFXOutputCondition () /PDFXRegistryName () /PDFXTrapped /False /CreateJDFFile false /Description << /ARA /BGR /CHS /CHT /CZE /DAN /DEU /ESP /ETI /FRA /GRE /HEB /HRV (Za stvaranje Adobe PDF dokumenata najpogodnijih za visokokvalitetni ispis prije tiskanja koristite ove postavke. Stvoreni PDF dokumenti mogu se otvoriti Acrobat i Adobe Reader 5.0 i kasnijim verzijama.) /HUN /ITA /JPN /KOR /LTH /LVI /NLD (Gebruik deze instellingen om Adobe PDF-documenten te maken die zijn geoptimaliseerd voor prepress-afdrukken van hoge kwaliteit. De gemaakte PDF-documenten kunnen worden geopend met Acrobat en Adobe Reader 5.0 en hoger.) /NOR /POL /PTB /RUM /RUS /SKY /SLV /SUO /SVE /TUR /UKR /ENU (Use these settings to create Adobe PDF documents best suited for high-quality prepress printing. Created PDF documents can be opened with Acrobat and Adobe Reader 5.0 and later.) >> /Namespace [ (Adobe) (Common) (1.0) ] /OtherNamespaces [ << /AsReaderSpreads false /CropImagesToFrames true /ErrorControl /WarnAndContinue /FlattenerIgnoreSpreadOverrides false /IncludeGuidesGrids false /IncludeNonPrinting false /IncludeSlug false /Namespace [ (Adobe) (InDesign) (4.0) ] /OmitPlacedBitmaps false /OmitPlacedEPS false /OmitPlacedPDF false /SimulateOverprint /Legacy >> << /AddBleedMarks false /AddColorBars false /AddCropMarks false /AddPageInfo false /AddRegMarks false /ConvertColors /ConvertToCMYK /DestinationProfileName () /DestinationProfileSelector /DocumentCMYK /Downsample16BitImages true /FlattenerPreset << /PresetSelector /MediumResolution >> /FormElements false /GenerateStructure false /IncludeBookmarks false /IncludeHyperlinks false /IncludeInteractive false /IncludeLayers false /IncludeProfiles false /MultimediaHandling /UseObjectSettings /Namespace [ (Adobe) (CreativeSuite) (2.0) ] /PDFXOutputIntentProfileSelector /DocumentCMYK /PreserveEditing true /UntaggedCMYKHandling /LeaveUntagged /UntaggedRGBHandling /UseDocumentProfile /UseDocumentBleed false >> ] >> setdistillerparams << /HWResolution [2400 2400] /PageSize [612.000 792.000] >> setpagedevice