Computational and Theoretical Chemistry 1023 (2013) 51–58

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Computational and Theoretical Chemistry

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Electronic structure analysis of isomeric preferences of canonicaland zwitterionic forms of lornoxicam

2210-271X/$ - see front matter � 2013 Published by Elsevier B.V.http://dx.doi.org/10.1016/j.comptc.2013.09.011

⇑ Corresponding author. Tel.: +91 172 2292018, mobile: +91 9417503172; fax:+91 172 2214692.

E-mail address: pvbharatam@niper.ac.in (P.V. Bharatam).

Zankhana P. Nathavad a, Sonam Bhatia b, Devendra K. Dhaked b, Prasad V. Bharatam b,⇑a Department of Pharmacoinformatics, National Institute of Pharmaceutical Education and Research (NIPER), Sector-67, S.A.S. Nagar, Punjab 160 062, Indiab Department of Medicinal Chemistry, National Institute of Pharmaceutical Education and Research (NIPER), Sector-67, S.A.S. Nagar, Punjab 160 062, India

a r t i c l e i n f o a b s t r a c t

Article history:Received 30 May 2013Received in revised form 25 August 2013Accepted 9 September 2013Available online 19 September 2013

Keywords:LornoxicamPolymorphTautomersZwitterion (O)Zwitterion (N)Density functional theory

Lornoxicam, is a non-steroidal anti-inflammatory drug (NSAID) and has analgesic, anti-inflammatory andantipyretic activity. Various polymorphic forms of drugs belonging to oxicam class are known in the lit-erature and study of their polymorphic behavior has become a research interest over the past few years.Due to the differences in the conformational arrangement of the molecules in the crystal lattice, polymor-phic forms of a drug substance can exhibit different physicochemical properties–solubility, density, dis-solution, pKa, etc. Density functional (DFT) study has been carried out on various canonical andzwitterionic forms of lornoxicam. The electronic level details revealed that, the existence of polymor-phism in lornoxicam can be traced to the prototropic exchange which helps in the interconversion ofone polymorph to the other. Electronic structures of all the probable isomers of lornoxicam at HF,B3LYP and M06L levels using 6-31 + G(d) basis set have been analyzed. The comparative analysis of theirrelative Gibbs free energies in the gas, solution and explicit water phase revealed that the form of globalminimum structure differs with respect to the varied conditions. Microsolvation calculations show thatthree water (3W) molecules are sufficient to stabilize the zwitterion ZO. Therefore, transition of canonicalto zwitterionic form can happen under the influence of explicit water molecules. Further, transition statestudies point out easy conversion between the two zwitterionic states (ZN M ZO). This phenomenon canbe attributed to the observed polymorphism in lornoxicam.

� 2013 Published by Elsevier B.V.

1. Introduction

Lornoxicam is a new non-steroidal anti-inflammatory drug(NSAID) from the oxicam class. It exhibits anti-inflammatory activ-ity and is a potent analgesic agent in post-operative pain, rheuma-toid arthritis, osteoarthritis and acute lumbar-sciatica conditions[1]. These effects are as a result of non-selective inhibition of cy-clo-oxygenase-1 and -2. Additionally, it plays an important roleas chemo-preventive and chemo-suppressive agent [2]. It has animproved safety profile owing to its shorter half-life [3]. The crystalstructure of two polymorphic forms (I and II) of lornoxicam wasestablished by Zhang et al. using various characterization tech-niques such as; FTIR spectroscopy, DSC experiments, X-ray powderdiffractometry (XPRD), and thermogravimetric analysis [4]. Form Iis found to have a triclinic space group while form II belongs toorthorhombic lattice system dominated by intermolecular andintramolecular hydrogen bonds, respectively. In addition to this,lornoxicam also known to exist as a zwitterion in acidic media ow-ing to keto-enol tautomerism [5]. Fig. 1 shows the three important

structures of lornoxicam in canonical and zwitterionic forms (ZOand ZN).

Zwitterions of amino acid are extensilvely studied using exper-imental and theoretical methods [6–14], but the same in drugs arerarely exposed: (i) Relative energies of canonical form and zwitter-ions (ii) the number of water molecules required to make canonicaland zwitterionic form isoenergetic (iii) the charge solvated state(canonical) vs. the salt bridged (zwitterionic) states of amino acidsare being extensilvely studied using quantum chemical studies.Drug molecules which are known to act in their zwitterionic stateare governed by their electric field which is believed to be the driv-ing force for their drug action [15,16]. Acknowledging the impor-tance of such concepts and the fact that such studies ontherapeutic drug molecules hitherto remained unexplored, we in-tended to implement similar studies for the identification of elec-tronic structure of the anti-inflammatory drug, lornoxicam.

Many of the drugs belonging to oxicam class are known to existin different polymorphic forms [17–21]. Several studies explainingthe details of various polymorphic states of piroxicam are reportedin the literature [18,20]. Among them a few reports highlight theimportance of proton transfer during the interconversion of poly-morphic forms, which mostly involves tautomeric conversion[21,22]. Distinctive molecular interactions are shown by canonical

S

SN

O O

O

N

O

NCl

S

SN

O O

O

HNO NH

Cl

S

SN

O O

O

NOH NH

Cl

Canonical (L-1) Zwitterion-N (ZN) Zwitterion-O (ZN)

1

23

H4

56

7

H

8

9

1011

12

1314

15H16

Fig. 1. 2-D structures of canonical (L-1), zwitterion O (ZO) and zwitterion N (ZN) in place of isomers write forms of lornoxicam.

52 Z.P. Nathavad et al. / Computational and Theoretical Chemistry 1023 (2013) 51–58

and zwitterionic states underlying the differences in their hydro-gen bonding pattern. Quantum chemical studies in the context ofunderstanding acid–base behavior of oxicams is studied by Hoet al. [5]. They have introduced a proton-exchange scheme to pre-dict the microscopic pKa values corresponding to the possibledeprotonation pathways. They combined these microscopic pKa

values to predict the macroscopic pKa values of oxicam com-pounds. Similarly, Franco-Pérez et al. studied several microspeciesinvolved in the prototropic equilibria of oxicams using time-dependent density functional theory (TD-DFT) [23].

Lornoxicam has not received much exploration in the context ofphysical and quantum chemical studies. There are no reports dis-playing the energetics of canonical and zwitterionic forms of lorn-oxicam and relating the thermodynamic stabilities and theprototropic exchanges associated with this drug molecule. Consid-ering the fact that polymorphism effect the physiochemical behav-ior of drug molecules which ultimately effect their bioavailability,we decided to carry out DFT [24] study to analyze the most stableform among several canonical and zwitterionic forms of lornoxi-cam under the gas and solvent phase conditions. QM/MM studiescarried out by Tahan et al. states that, the varied dielectric constanteffect the energy profiles of canonical and zwitterionic species [25].Hence, it is worth exploring the stabilities of different forms oflornoxicm under the conditions of varied polarity. We have alsoenvisaged the thermodynamical outcome of different forms oflornoxicam under the microsolvation conditions by consideringthe explicit water molecules. Further, the proton transfer pathwayfor the interconversion of canonical to the zwitterionic form oflornoxicam has also been explored under different operatingconditions to examine their effect on the energetics of thisinterconversion.

2. Methods and computational details

Complete geometry optimizations were performed on variouscanonical and zwitterioinc forms of lornoxicam using Hartree–Fock (HF) [26], Becke-Lee–Yang-Parr (B3LYP) [27,28] and M06L[29] method with 6-31 + G(d) basis set in both the gas and solutionphases (IEFPCM model [30]) respectively using Gaussian 09 [31]suite of programs. Crystal structure of lornoxicam (CCSD code:EHIGUX [32]) which was used in a study on metal-oxicam coordi-nation compounds by Tamasi et al. [2] has been taken for thisstudy as a reference structure for generating 11 possible canonicalforms along with the zwitterionic states (ZN and ZO) of lornoxi-cam. Intramolecular H-bonding for the most significant (L-1, ZN,ZO) forms were confirmed by AIM (atom in molecule) calculationsusing the AIM2000 software [33–36]. Different solvents (ethanol,acetone, dichloroethane, dichloromethane and tetrahydrofuran)were used in order to study the effect of their polarity on the rela-tive energy trends of canonical and zwitterionic forms of lornoxi-cam. Their choice was in accordance to their respective dielectricconstants (K = 80.00, 24.55, 21.00, 10.50, 9.10 and 7.50) with thevalues in the decreasing order. The pathway for the conversion ofcanonical into zwitterionic forms was also explored by taking ex-

plicit water molecules (n = 1, 2) in consideration. Frequencies werecomputed analytically to characterize each point as a minimum ora transition state, and also to estimate the zero point vibrationalenergies (ZPE)-corrected to Gibbs free energy. The calculated ZPE[37] values (at 298.15 K) were scaled by a factor of 0.9806,0.9153 and 0.9780 for B3LYP, HF and M06L levels respectively[38–40].

3. Results and discussion

3.1. Isomeric and zwitterionic preferences

Lornoxicam is a monoprotic weak acid [3] and it exists in canon-ical and zwiterionic states (Fig. 1). In the canonical state 11 differentisomers are in principle possible for this drug candidate. DFT calcu-lations [24,41] were carried out to scan the potential energy surface(PES) [42] of lornoxicam by taking into consideration all the possibleisomeric and zwitterionic forms. Geometrically optimized struc-tures with their respective relative energies (Fig. 2, Table 1 andFig. S1, Supporting Information) in the gas and solution phases wereanalyzed. The results have indicated that the isomeric form L-1 is themost stable form in the gas phase condition, at three different levelsof theory namely B3LYP, HF and M06L. It is characterized by threeintramolecular H-bonds, however the other isomeric forms are>5 kcal/mol less favorable than this global minimum energy struc-ture. ZN and ZO are two important zwitterionic forms of lornoxicamwhich are characterized by the presence of two intramolecularhydrogen bonds at their negative and positive centers. These areabout �7 kcal/mol less stable on the potential energy surface com-pared to the most stable canonical form (L-1). However, under thesolvent phase conditions ZO gains thermodynamic stability andturns out to be a predominant form. In comparison, ZN form comesout to be relatively less stable by 5.03 kcal/mol in the solvent phasecondition. Its lower stability in the solvent phase is attributed to itsburied negative nitrogen center as compared to openly accessedoxygen (negative center) of ZO. The isomeric form L-1 can intercon-vert to ZO form via rotation across C2–C7 with exchange of hydrogenfrom the amide nitrogen to the pyridine ring followed by a transfer ofhydrogen from enolic oxygen to amide nitrogen. The relative stabil-ity of N or O zwitterionic forms in an oxicam class of drugs is gov-erned by the nature of substituents. The stability of ZO can berationalized due to the presence of electron withdrawing 2-chloro-thieno ring which markedly enhances the acidity of enolic proton.This is because of low gas phase deprotonation energies of lornoxi-cam (3.5–5.25 kcal/mol) as compared to other drug candidates(0.07 to 3.48 kcal/mol) belonging to the oxicam class [5]. To furtheranalyze the correlation between the geometrical and topologicalparameters of intramolecular H-bonds in three important forms oflornoxicam (L-1, ZO and ZN), the theory of atoms in molecules(AIMs) was used. The correlation between the properties of bondcritical points (BCPs) and H-bond energies both in the conventionaland unconventional H-bonds were found in the literature [43–46].The results are mentioned in Table 2 and the corresponding molec-ular graphs are provided in Fig. S2 (Supporting Information). All

Fig. 2. B3LYP/6-31 + G(d) optimized geometries of first three lower energy canonical isomers (L-1 to L-3) and zwitterionic states (ZO and ZN) of lornoxicam. Hydrogenbonding distances are given in Angstrom (Å) units and angles in degrees (�) shown as dashed lines. Note: Optimized geometries for isomers (L-4 to L-11) is provided in Fig. S1(Supporting Information).

Table 1Relative Gibbs free energy (kcal/mol) of various canonical isomeric and zwitterionic forms of lornoxicam in the gas and aqueous phase conditions.a,b

Molecule Gas phase Solventd

HF B3LYP M06L B3LYP

Isomers L-1 00.00 00.00 00.00 02.57L-2 05.01 05.60 05.88 05.60L-3 11.56 07.91 08.00 06.98L-4 13.34 13.93 14.53 12.93L-5 17.50 13.77 – 15.52L-6 17.52 14.72 – 01.58L-7 13.49 15.15 – 02.37L-8 20.75 17.46 – 18.47L-9 28.35 28.36 – 22.20L-10 33.01 32.41 – 29.74L-11 –c 23.10 - 18.72

Zwitterions ZN 11.31 07.14 07.57 05.03ZO 11.74 06.94 07.30 00.00

Note: Absolute energies were corrected by taking thermal correction to Gibbs free energy into consideration.a All energies are corrected for zero-point vibrational energy.b Basis set 6-31 + G(d) is used for optimizations.c During complete optimization structure get converted to ZN.d Implicit solvent analysis performed using IEFPCM method.

Table 2Property functions like electron density (qc) and Laplacian of electron density (r2q) of canonical, ZO and ZN forms to obtain Bond Critical Points (BCP) and Ring Critical Point(RCP) calculated by using AIM2000 software.

Canonical qc r2q ZO qc r2q ZN qc r2q

O6� � �H16 0.016 0.068 O6� � �H9 0.052 0.184 O6� � �H9 0.046 0.208RCP-C 0.02 0.12 RCP-C 0.02 0.12 RCP-C 0.02 0.11RCP-D 0.02 0.09 – – – RCP-D 0.01 0.05RCP-E 0.01 0.06 RCP-E 0.02 0.12 RCP-E 0.02 0.12

Z.P. Nathavad et al. / Computational and Theoretical Chemistry 1023 (2013) 51–58 53

the three forms show resonance assisted intramolecular hydrogenbond (RAHBs) due to p electron delocalization [47]. The effects ofp-electron delocalization in the L-1 system are as follows (a) short-ening of O� � �O contact; (b) increase in the strength of H-bond; (c)shift of proton towards the center of O� � �O contact; (d) equally com-parable C@C and C–C bond lengths and C@O and C–O bond lengths.All these effects are direct consequences of p-electron delocalizationon the geometry of L-1 isomeric form. The topological properties ofthe electron density distributions (qBCP) for H-bridges in L-1, ZN andZO forms is slightly higher than the maximum threshold value of0.40 a.u. suggested by Koch and Popelier while defining the criteriafor detecting hydrogen bonding interactions [44,45]. The higher

(qBCP) values (0.046–0.0594) suggest the presence of partial covalentcharacter in the concerned intramolecular hydrogen bond (IMHB)interactions, thus supporting our hypothesis of existence of RAHBinteraction in this drug molecule. Fig. 3 displays the correlationcurve between (qBCP) and O� � �H IMHB distance for three differentforms of lornoxicam (L-1, ZN and ZO) with an excellent regressioncoefficient (R2) of 0.957. Thus, the strength of IMHB interactionscan be measured by analyzing the electron density parameter atBCP between the hydrogen atom and acceptor atom.

Owing to RAHBs effect, L-1 form also carries three ring criticalpoints (RCP) with r2qRCP ranging from 0.010–0.020 a.u. Similarly,zwitterionic forms also show clear RCPs (Table 2) which provide

Fig. 3. Correlation graph showing qBCP as a function of intramolecular hydrogenbond distance R(O� � �H) for L-1, ZN and ZO forms of lornoxicam.

54 Z.P. Nathavad et al. / Computational and Theoretical Chemistry 1023 (2013) 51–58

resonance assistance in the IMHB interactions in these forms. Com-putational estimation of the dipole moments of canonical andzwitterionic forms of lornoxicam revealed that the ZO is having amuch higher dipole moment value in the gas, solution and explicitwater phases (12.50, 17.24 and 10.92 Debye respectively) as com-pared to two other forms (3.55–13.35 Debye) (Table 3). The stabil-ity of ZO in the aqueous phase is due to its higher dipole momentwhich helps in better interaction with the polar solvents. On thecontrary the canonical form is significantly less stable in these con-ditions as its dipole moment doesn’t support its solvation in thepolar media.

NBO analysis, at B3LYP/6-31 + G(d) level, was employed to esti-mate second-order interactions in these systems. In our previousreports, we have used similar calculations in order to understandthe delocalization and strength of hydrogen bonding in differentheteroaromatic systems by analyzing second-order interactionenergies [48–51]. In the most stable canonical form (L-1) the sec-ond order-interaction energies (E(2)) due to nN8 ? p⁄C7–O6 andnO4 ? p-C2–C3 is 73.82 and 43.23 kcal/mol, respectively which givestrong stabilization to this isomer by the lone pair delocalization ofthe central nitrogen atom (LP(N8)r = 1.63 e). While in comparisonto L-1, the delocalization is greatly reduced in the case of ZO andZN with nO4 ? p⁄C2–C3 and nN8 ? p⁄C10–N11 (2.75 and 14.05 kcal/mol). Thus, localization of electron density at O4 in ZO and N8 inZN increases the nucleophilicity of these forms which is supportedby higher lone pair occupancies of ZO (LP(O4)r = 1.96 e) and ZN(LP(N8)r = 1.83 e). This clearly shows charge accumulation in zwit-terionic forms at O and N centers.

As mentioned earlier, L-1, ZO and ZN are also characterized bystrong intramolecular hydrogen bonding interactions with a bondlength ranging from 1.64–2.28 Å. The presence of strong H-bondswere also characterized by second order delocalization energiesof one of the lone pairs of oxygen nO4 ? r⁄N8–H5 (ZO) and nitrogennN8 ? r⁄O4–H5 (ZN) with energy stabilization of 35.28 and

Table 3NBO charges (electrons) and dipole moments (Debye) of canonical (L-1), ZO and ZN form

Molecule NBO charge (q)

N8

Gas L-1 �0.62ZO �0.63ZN �0.67

Solution L-1 �0.61ZO �0.66ZN �0.68

Explicit water L-1 �0.63ZO �0.62ZN �0.67

42.02 kcal/mol respectively (Table 4). Thus, from the above analy-sis, it can be interpreted that L-1 is stabilized by greater chargedelocalization while on the other side ZO and ZN forms are charac-terized by localization of electron density on oxygen (O4) andnitrogen (N8) centers.

3.2. Effect of solvent polarity

The polarity of solvent effects the strength of intramolecularhydrogen bonding; thus the relative energy trends of differentforms (canonical (L-1) and zwitterionic) of lornoxicam in solventsof varied polarity have been studied. Selection of solvents wasmade on the basis of their dielectric constant ranging fromK = 7.00–80.00 for non-polar to polar solvents respectively. First,the optimization was carried out with the non-polar solvents fol-lowed by the other solvents with the increasing order of polarity.Interestingly, ZO was found to be predominantly stable form underthe conditions of varied polarity indicating the effect of solvationon the relative stablitity of zwitterion. This is due to the stabiliza-tion of its negative ion center by solvent molecules. In comparisonto ZN, better stabilization is noticed in ZO; this can be envisagedfrom its molecular structure, as the negative center (O4) in ZO iseasily exposed to solvent molecules in comparison to buried nega-tive ion center (N8) of ZN where solvent access is slightly difficult.NBO charges at N8 and O4 centers are listed in Table 3. The largedifference in the charge distribution (0.10 e) at O4 in ZO in thegas and solution phase (aqueous) is clearly evident from Table 3whereas, in contrast there is a minor change in this value for N8negative center in ZN. This explanation is also supported by im-proved thermodynamic stabilities of canonical (L-1) and ZN formsunder the organic solvents (THF, DCM, DCE) in comparison to theirstabilities in polar solvents. From Fig. S3 of Supporting Informationit is clear that there is a decrease in the energy difference (espe-cially between L-1 and ZO) with an increase in the organic charac-ter (K = 7.50–10.50) of the solvent. However, as the non-polarnature of the solvent increases the electrostatic interactions be-came highly favourable thus the effect of solvent is clearly evidentin the non-polar environment also.

3.3. Effect of microsolvation on canonical and zwitterionic formspreference

There are several reports which indicated that the relative pref-erences of canonical and zwitterionic forms is modulated by thesurrounding molecules, cations, anions, etc. in amino acids [11–13]. Lornoxicam posses (sulfonyl, amide, pyridyl/pyridinium,enol/enolate) functional groups which have hydrogen bondingability with water molecules. The structures of canonical and zwit-terionic form are influenced by the number of water molecules andtheir arrangement with the functional groups. In order to see the

s in the gas, solution and explicit water phase conditions.

Dipole moment

O4 N11

�0.68 �0.48 03.55�0.69 �0.54 12.50�0.70 �0.55 09.60

�0.69 �0.50 05.11�0.79 �0.12 17.24�0.71 �0.53 13.35

�0.69 �0.47 02.27�0.74 �0.53 10.92�0.70 �0.54 07.80

Table 5Relative Gibbs free energy (kcal/mol) of lornoxicam-nH2O clusters with canonical andzwitterionic forms.a,b

Explicit water Explicit continuum solventc

n = 0 n = 1 n = 2 n = 3 n = 0 n = 1 n = 2

L-1 0.00 0.00 0.00 0.00 0.00 0.00 0.00ZN 7.14 6.42 4.07 �0.09 3.26 3.21 1.30ZO 6.94 4.70 2.11 �2.23 �1.51 �2.24 �4.14

a All energies are at B3LYP/6-31 + G(d) level of theory.b Absolute energies were corrected by taking thermal correction to Gibbs free

energy into consideration.c Implicit solvent analysis of microsolvated molecules, performed using IEFPCM

method.

Fig. 5. The relative energetic preferences of zwitterionic configurations (ZN and ZO)compared to the canonical configuration as a function of number of watermolecules.

Table 4Second order interaction energies (E(2)) of canonical (L-1), ZO and ZN forms calculatedat B3LYP level with 6-31 + G(d) basis set (E(2) in kcal/mol; Ei � Ej, Fij values in a.u.).a

Structure Interaction E(2) Ei � Ej Fij

L-1 nO6 ? p⁄C2–C7 11.85 0.74 0.086nO6 ? p⁄C7–N8 22.66 0.77 0.121nN8 ? p⁄C7–O6 73.82 0.25 0.122nN8 ? p⁄C10–C15 37.30 0.29 0.092nO4 ? p⁄C2–C3 43.23 0.33 0.110

ZN nN8 ? p⁄O4–H5 42.02 0.75 0.162nN8 ? p⁄C10–N11 14.05 0.78 0.096nO4 ? p⁄C2–C3 41.47 0.33 0.108

ZO nO6 ? p⁄N11–H9 81.53 0.12 0.091nO4 ? p⁄N8–H9 35.28 0.17 0.069nO4 ? p⁄C2–C3 2.75 1.18 0.072

a All calculations are carried out at B3LYP/6-31 + G(d) level of theory.

Z.P. Nathavad et al. / Computational and Theoretical Chemistry 1023 (2013) 51–58 55

effect of explicit water molecules on the relative stability of lornox-icam isomers (L-1, ZN and ZO) microsolvation study is carried outin the presence of 1–3 water molecules. For each isomer about 4–6microsolvated structures were identified using chemical intuitionat B3LYP/6-31 + G(d) level of which optimized geometries of thelowest energy clusters are displayed in Fig. 4 and their relativeenergies are shown in Table 5. The other alternative structuresand their relative energy are given in supporting information(Figs. S4–S6 and Supplementary Table S1).

In one water complex of L-1-1W, a water molecule is localizedbetween amide NH and pyridyl N atom (Fig. 4). In ZN-1W, a watermolecule bridges carbonyl group with pyridinium ring; whereas inZO-1W, a water molecule is located above enolate group by threeintermolecular hydrogen bonds. In the presence of one water mol-ecule ZN and ZO zwitterions, are about 6.42 and 4.07 kcal/mol lessstable respectively than the canonical L-1-1W cluster.

In two water containing canonical cluster L-1-2W, a chain oftwo water molecules bridges sulfonyl oxygen atom with pyridylnitrogen and amide NH. In the zwitterionic complexes ZN-2Wand ZO-2W, a two water chain is located between carbonyloxygen and pyridinium ring. The relative stability of ZN-2Wand ZO-2W complexes is improved by about 2.4 and 2.6 kcal/

Fig. 4. B3LYP/6-31 + G(d) optimized geometries of lornoxicam-nH2O clusters isomers ounits).

mol compared to their one water clusters (ZN-1W and ZO-1W).The zwitterion ZO-2W is about 2 kcal/mol less stable than L-1-2W. This indicates that two water molecules are not sufficient

f canonical and zwitterionic forms (Hydrogen bonding distances in Angstrom (Å)

Fig. 6. Potential energy surface (at B3LYP/6-31 + G(d)) for the conversion of canonical (L-1) to zwitterions (ZN, ZO) with transition states barriers (Note: Rate determiningsteps were taken into consideration).

Table 6Relative Gibbs free energy (kcal/mol) of isomers and transition states of lornoxicam.a,b

Gas Sol.c Explicit waterd

n = 1 n = 2

L-1 0.00 0.00 0.00 0.00TS1 40.67 39.21 17.09 16.31ZN 7.14 2.46 6.42 1.96TS2 7.54 1.37 6.49 1.94ZO 6.94 �2.57 4.70 2.11

a All energies are at B3LYP/6-31 + G(d) level of theory.b Absolute energies were corrected by taking thermal correction to Gibbs free

energy into consideration.c Implicit solvent phase analysis was carried out using water as a solvent med-

ium in IEFPCM solvent model.d Consideration of one (1 W) and two (2 W) water molecules for explicit solvent

calculations.

56 Z.P. Nathavad et al. / Computational and Theoretical Chemistry 1023 (2013) 51–58

to make zwitterionic isomers energetically competitive withcanonical isomer.

In L-1-3W, two water molecules (chain) span across sulfonyloxygen atoms and a additional water molecule connects the pyri-dyl moiety to a chain of two water molecules. Whereas in both ofthe zwitterionic configurations (ZN-1W and ZO-1W), a chain ofthree water molecules bridges one of oxygen atom of sulfonylgroup to pyridyl nitrogen by three intermolecular hydrogen bonds.Data from Table 5, indicates that the zwitterionic form ZN-3W isisoenergetic with the canonical form L-1-3W, in the presence ofthree microsolvating water molecules. On the other hand ZO-3Wis preferred by about 2 kcal/mol over ZN-3W. The relative energydifference (DE) of zwitterionic structures (ZN and ZO) with thecanonical L-1, is given in Fig. 5 as a function of number of watermolecules. This indicates that the energy difference betweenunsolvated canonical and zwitterionic structures is high (about7 kcal/mol) and as the number of microsolvating water moleculesincrease, DE gradually decrease. As per microsolvation calculationit can be concluded that transition of canonical form to zwitter-ionic form starts with 2–3 water molecules.

125.78

(a) (bFig. 7. 3D optimized geometries of TS1 in three different conditions (a) gas (b) one explcibonding distances in Angstrom (Å) units and angles in degrees (�) shown as dashed line

3.4. Proposed pathway for the canonical M zwitterion transformation

The pathway for the conversion of canonical conformer (L-1) tozwitterionic forms involves prototropic exchanges (1,3 and 1,5-Hshift) along with the rotational processes, however the rate deter-mining step in this reaction pathway is the shift of proton (H9)from N8 to N11 center (1,3-H shift) leading to the formation ofZN which is followed by 1,5-H shift and transforms into ZO whichis thermodynamically stable product. Three different conditionswere employed to study this pathway (Fig. 6). The energy valuesand 3-D structures for the optimized geometries of TS1 were pro-vided in Table 6 and Fig. 7 (Figs. S7–S9 of Supporting Information).In the gas phase condition, the energy barrier for the conversion ofL-1, primarily to the ZN form is �41 kcal/mol, on the contrary, theconversion of ZN to ZO (1,5-H shift) is highly favorable which canhappen with a negligible barrier of �0.40 kcal/mol owing to its sixmembered catalyzed transition state (TS2). In the implicit watermedium the barrier for the 1,3-H shift is reduced only marginallyby 1.46 kcal/mol. The 1,5-H shift (TS2) leads to spontaneous trans-fer of ZN to ZO on the potential energy curve without any reactionbarrier in the implicit solvent condition.

In the presence of one explicit (1W) water molecule the energybarrier for the conversion from L-1 to ZN is 17.09 kcal/mol. This va-lue is much smaller than the corresponding gas phase value(�41 kcal/mol) indicating that zwitterion formation is facilitatedin presence of explicit water. Thus, proton exchange drives the for-mation of zwitterion. The effect of the second water molecule(2W), on the energy barriers was also examined. The data showsthat there is only a marginal gain in the energy barrier(16.31 kcal/mol) under 2W catalyzed condition. Thus, by examin-ing the energetics of the pathway, we can conclude that conversionof L-1 to ZN is catalyzed by explicit (n = 1, 2) water moleculeswhich makes a difference of 23.58 and 24.36 kcal/mol in the en-ergy barriers of TS1 in comparison to gas phase condition. Theinterconversion of zwitterionic states (ZN to ZO) via 1,5-H shiftgoes by a barrierless transition under the explicit solvent phasecondition also. Hence, the ease of prototropic exchange betweenthe various forms of lornoxcicam marked the existence of poly-morphism in this drug molecule.

1.61

1.18

1.341.39

119.81124.62

) (c) t water (1 W) catalyzed and (c) two explicit (2 W) water catalyzed phase. (Hydrogens).

Z.P. Nathavad et al. / Computational and Theoretical Chemistry 1023 (2013) 51–58 57

4. Conclusions

Quantum chemical analysis was carried out to understand thecanonical isomeric preferences and the stability of solvated zwit-terions of lonoxicam. In the gas phase condition, the canonical iso-mer (L-1) is more stable at HF, B3LYP and M06L levels of ab initioand DFT calculations. This form is characterized by resonance as-sisted intramolecular hydrogen bond (RAHB), due to p electrondelocalization. However, the stability of zwitterionic form (ZO) isnoticeable on the PES of lornoxicam under the solvent phase con-dition (polar and non-polar solvents). Thus, lornoxixcam is anexample of a drug where the stability of its zwitterionic form isgoverned by solvent molecules. In addition to this, tautomerismbetween the polymorphic forms is justified based on their isomericstability. Microsolvation calculations illustrate that ZO form startsto dominant other two forms (canonical (L-1) and ZN) under theinfluence of three water molecules. This study provided clues forthe explicit water mediated conversion of canonical to zwitterionicform (L-1 M ZN) form. The prototropic exchange (1,5-H shift) be-tween the two zwitterionic forms is exothermic and is almost bar-rierless on the reaction path. Thus, this study revealed theimportance of prototropic exchanges which governs the existenceof polymorphism in drug molecules.

Acknowledgements

SB thanks Department of Science and Technology (DST), NewDelhi, for providing INSPIRE fellowship. DD thanks to Council ofScientific and Industrial Research (CSIR), New Delhi, India forproviding financial support.

Appendix A. Supplementary data

Supplementary data associated with this article can be found, inthe online version, at http://dx.doi.org/10.1016/j.comptc.2013.09.011.

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