See discussions, stats, and author profiles for this publication at: https://www.researchgate.net/publication/281589601

Saccharin salts of biologically active hydrazone derivatives

Article  in  New Journal of Chemistry · September 2015

DOI: 10.1039/C5NJ01532D

CITATIONS

12READS

88

6 authors, including:

Some of the authors of this publication are also working on these related projects:

Monte Carlo Studies of Drug Nucleation View project

Nitrosyl carboxylate palladium complexes as catalysts for aerobic oxidation of alcohols View project

Artem Surov

Institute of Solution Chemistry of RAS

37 PUBLICATIONS   296 CITATIONS   

SEE PROFILE

Alexander Voronin

Institute of Solution Chemistry of RAS

20 PUBLICATIONS   160 CITATIONS   

SEE PROFILE

Andrei V. Churakov

Russian Academy of Sciences

305 PUBLICATIONS   2,046 CITATIONS   

SEE PROFILE

German L Perlovich

Institute of Solution Chemistry of RAS

183 PUBLICATIONS   2,153 CITATIONS   

SEE PROFILE

All content following this page was uploaded by Artem Surov on 31 October 2015.

The user has requested enhancement of the downloaded file.

8614 | New J. Chem., 2015, 39, 8614--8622 This journal is©The Royal Society of Chemistry and the Centre National de la Recherche Scientifique 2015

Cite this: NewJ.Chem., 2015,

39, 8614

Saccharin salts of biologically active hydrazonederivatives†

Artem O. Surov,a Alexander P. Voronin,a Anna A. Simagina,a Andrei V. Churakov,b

Sophia Y. Skachilovac and German L. Perlovich*a

The crystal structures of two saccharin salts with derivatives of an anti-tubercular drug isoniazid, namely

vanillin isoniazid saccharinate (salt I) and salinazid saccharinate (salt II), were obtained in a X-ray

diffraction experiment. The pattern of intermolecular interactions in the crystals was quantified by solid-

state DFT followed by the Bader analysis of periodic electron density. It was established that ca. 42% of

lattice energy is contributed by C–H� � �O contacts, while conventional hydrogen bonds have only ca. 28%.

Salt I was found to show a 12-fold aqueous solubility improvement compared to pure API, whereas salt II

is approximately 20 times more soluble than the starting salinazid. The standard thermodynamic functions

of the salt formation were determined. The Gibbs energy change of the process was found to be

negative, indicating that the formation of the salts from individual components is a spontaneous

process. The most significant contribution to the Gibbs energy is provided by the enthalpy term, while

the entropy change of the process has a negative value, introducing a positive contribution to DG�f .

1. Introduction

Hydrazones represent an important class of Schiff base com-pounds that exhibit different types of biological activity.1 Thereare a lot of data in the literature suggesting that hydrazonederivatives have great potency against resistant forms of tubercu-losis and can be used as an inexpensive substitution for isoniazid(an important first-line antituberculosis drug).2 It has been foundthat the combination of isoniazid with some hydroxyaldehydesleads to the formation of stable hydrazones that display conservedactivity and less toxicity due to the inactivation of the NH2 group ofisoniazid.3 In addition, hydroxy- and methoxy-derivatives of hydra-zones possess antioxidant activity due to their ability to capturefree radicals.4 The presence of hydroxyl groups on the benzenering also plays an important role in the anticancer activity ofhydrazones, especially when it is located in the ortho-position.5

Although the biological activity is undoubtedly the keyfeature of potent active pharmaceutical ingredients (APIs),other factors may be equally important for application in vivo

such as solubility. Unfortunately, in many cases this importantaspect is subject to later studies in drug discovery and drugdevelopment research. This is a serious drawback for a drugcandidate on its way to become a useful pharmaceutical agent asit is hard to compensate for weak solubility properties. It hasbeen reported that currently almost 40% of marketed drugs facethe major problem of poor aqueous solubility.6 One of the bestapproaches to overcoming the solubility challenge without mod-ification of the pharmacophore structure of an API is developingnew crystalline forms such as salts or co-crystals. In fact, saltformation is the most common method for improving solubilityand today more than 50% of APIs are marketed as salts.7

There are various pharmaceutically relevant organic counterions for salt formation. In this work, well-known artificialsweeter called saccharin was employed as a salt former forthe development of new crystalline forms of hydrazone deriva-tives. Saccharin is currently approved by FDA for use in food asa non-nutritive sweetener, which is a significant advantage interms of further biopharmaceutical studies of API sacchari-nates. It has been reported in the literature that salt orco-crystal formation with saccharin often leads to a considerableenhancement of the aqueous solubility of APIs.8 The presence ofsaccharin as a potent sweetener may improve organolepticproperties of a formulation, masking the bitter taste of a drug.9

In addition, the pH of the saccharinate solutions is higher thanthat of, for example, the corresponding hydrochlorides, whichmakes them more suitable for injections and drops. Therefore,saccharin is one of the most frequently chosen co-formers/saltformers for preparation of new solid forms of APIs.10

a Institution of Russian Academy of Sciences, G.A. Krestov Institute of Solution

Chemistry RAS, 153045, Ivanovo, Russia. E-mail: glp@isc-ras.rub Institute of General and Inorganic Chemistry RAS, Leninskii Prosp. 31, 119991,

Moscow, Russiac Russian Research Center for Safety of Bioactive Substances, 142450,

Staraya Kupavna, Russia

† Electronic supplementary information (ESI) available: Details of the DFTcalculations, packing arrangements of the pure APIs, results of PXRD analysis.CCDC 1407124 and 1407125. For ESI and crystallographic data in CIF or otherelectronic format see DOI: 10.1039/c5nj01532d

Received (in Montpellier, France)17th June 2015,Accepted 25th August 2015

DOI: 10.1039/c5nj01532d

www.rsc.org/njc

NJC

PAPER

Publ

ishe

d on

26

Aug

ust 2

015.

Dow

nloa

ded

by G

azi U

nive

rsite

si o

n 29

/10/

2015

09:

18:1

0.

View Article OnlineView Journal | View Issue

This journal is©The Royal Society of Chemistry and the Centre National de la Recherche Scientifique 2015 New J. Chem., 2015, 39, 8614--8622 | 8615

In this work, we report crystal structures, thermal analysis,aqueous solubility and formation thermodynamics of the saccharinsalts with two biologically active hydrazone compounds, namelyvanillin isoniazid ((2-hydroxy-3-methoxybenzaldehyde)isonicotinoylhydrazone) and salinazid ((2-hydroxybenzaldehyde)isonicotinoylhydrazone) (Fig. 1).

Additionally, the DFT computations complemented with theBader analysis of the periodic electron density were performedto describe quantitatively the patterns of the intermolecularinteractions in the saccharin salts as well as to estimate theirlattice energies.

2. Materials and methods2.1 Compounds and solvents

Vanillin isoniazid ((2-hydroxy-3-methoxybenzaldehyde)isonicotinoylhydrazone, C14H13N3O3, MW 271.27, 99%) and salinazid ((2-hydroxy-benzaldehyde)isonicotinoyl hydrazone, C13H11N3O2, MW 241.25,99%) were obtained from Interbioscreen Ltd. Saccharin(C7H5NO3S, MW 183.18, 99%) was purchased from AcrosOrganics. All the solvents were of analytical grade and usedas received without further purification.

2.2 Salt synthesis

Solvent-drop grinding experiments were performed using aFritsch planetary micro mill, model Pulverisette 7, in 12 mL agategrinding jars with ten 5 mm agate balls at a rate of 600 rpm for60 min. The experiments were carried out with stoichiometricamounts of vanillin isoniazid or salinazid and saccharin and afew drops of solvent (methanol) were added using a micropipette.For slurry experiments, 100 mg of the hydrazone and an equi-molar amount of saccharin were stirred in methanol for 12 h.

2.3 Crystallization procedure

For each salt, 30 mg of the hydrazone and an equimolar amountof saccharin were dissolved in methanol and stirred at 50 1C. Theresulting clear solution was slowly cooled. The solution was keptin a fume hood at room temperature. Diffraction quality yellowcrystals of saccharin salts were grown over a period of severaldays. Crystals obtained from the crystallisation batches were airdried before being subjected to further analysis.

2.4 X-ray diffraction experiments

Single-crystal X-ray diffraction data were collected on a BrukerSMART APEX II diffractometer using graphite-monochromatedMoKa radiation (l = 0.71073 Å). The structures were solved bydirect methods and refined by full matrix least-squares on F2

with anisotropic thermal parameters for all non-hydrogenatoms.11 Absorption corrections based on measurements ofequivalent reflections were applied.12 All hydrogen atoms werefound from the difference Fourier map and refined isotropi-cally. X-ray powder diffraction (PXRD) data were recorded underambient conditions in Bragg–Brentano geometry by a BrukerD8 Advance diffractometer with CuKa1 radiation (l = 1.5406 Å).

2.5 Differential scanning calorimetry (DSC)

Thermal analysis was carried out using a Perkin Elmer DSC4000 differential scanning calorimeter with a refrigerated cool-ing system (USA). The sample was heated in sealed aluminumsample holders at a rate of 10 K min�1 in a nitrogen atmo-sphere. The unit was calibrated with indium and zinc stan-dards. The accuracy of the weighing procedure was �0.01 mg.

2.6 Solubility experiments and formation thermodynamicsstudy

Dissolution measurements with the solids were made using theshake-flask method at 298.2 � 0.1 K. The samples were sus-pended in 10 mL of degassed water in pyrex glass tubes. Theamount of the substance dissolved was measured by takingaliquots of 1 mL of the respective media. The solid phase wasremoved by filtration (Rotilabos syringe filter, PTFE, 0.2 mm),and the concentration was determined by UV-vis spectroscopy(Varian Cary 50). The following reference wavelengths wereused: 300 nm for vanillin isoniazid and salt I, 330 nm forsalinazid and salt II.

The solubility of the compounds was also measured at 293.2,298.2, 303.2 and 308.2� 0.1 K. An excess of the solid was placedin an Eppendorf tube and 2 mL of water was added. After 24 hthe suspension was filtered through a Rotilabos syringe filter(PTFE, 0.2 mm), and the concentration in the supernatant wasdetermined by UV-vis spectroscopy as described above. Theresults are stated as the average of at least three replicatedexperiments. Concentrations were calculated according to anestablished calibration curve.

In the case of 1 : 1 stoichiometry, the formation reaction of amulti-component compound from a pure API (A) and a pureco-former (B) may be described as

Asolid + Bsolid - ABsolid (1)

It has been established in the literature that the standard free-energy change, DG�f , for the above reaction may be expressedthrough consideration of the solubility data of each of thematerials.13 In the case of salt dissolution, however, ionisedspecies are usually formed. Therefore, the dissociation con-stants of A (pKa,A) and B (pKa,B) have to be taken into account.Hence, all the solubility experiments should be conducted inwater solutions to ensure proximity of the intrinsic pKa value of

Fig. 1 Molecular structures of the studied hydrazones and saccharin.

Paper NJC

Publ

ishe

d on

26

Aug

ust 2

015.

Dow

nloa

ded

by G

azi U

nive

rsite

si o

n 29

/10/

2015

09:

18:1

0.

View Article Online

8616 | New J. Chem., 2015, 39, 8614--8622 This journal is©The Royal Society of Chemistry and the Centre National de la Recherche Scientifique 2015

each compound. If A is a weak acid and B is a weak base, thenthe Gibbs energy of formation of a (1 : 1) salt may be given bythe following equation:14

DG�f ¼ �RT � ln10pKa;B�pKa;ASp

A � SpB

SsaltA � Ssalt

B

!(2)

where SpA and Sp

B are the solubility of pure A and B in a solvent,while Ssalt

A and SsaltB are the solubility of the salt components in a

solution when in equilibrium with the pure salt. For the sake ofsimplicity, the activities of the components are approximated bymolar concentrations. The product of Ssalt

A and SsaltB is generally

known as Ksp of a salt. Thus, eqn (2) may be written as:

DG�f ¼ �RT � ln10pKa;B�pKa;AS

pA � S

pB

Ksp

� �(3)

In addition, comparison of eqn (2) and/or (3) with the expressionfor the standard free-energy change for the isothermal reaction

DG1 = �RT�ln K (4)

shows that the ratio appearing in the logarithmic term ofeqn (2) and/or (3) has the character of an equilibrium constantat the given temperature. Therefore, this quantity can bedefined as Kf. If the Kf values are known at different tempera-tures, the van’t Hoff relation may be used to derive the enthalpyof formation, DH�f , of a multi-component compound:

d lnKf

dð1=TÞ ¼ �DH�fR

(5)

Finally, the entropy change of the formation process can beestimated from the general relationship relating different thermo-dynamic functions:

DG�f ¼ DH�f � T � DS�f (6)

In order to verify the salt stability in aqueous media, a composi-tion of the saturated solutions of salts at each temperature wasalso analysed by HPLC. HPLC was performed on a ShimadzuProminence model LC-20AD equipped with a PDA detector and aC-18 column (150 mm � 4.6 mm ID, 5 mm particle size and 100 Åpore size). Elution was achieved by a mobile phase made of 0.1%trifluoroacetic acid in water (55%) and methanol (45%) by theisocratic method. An injection volume of 20 mL was used with aneluent flow rate of 1 mL min�1.

2.7 Solid-state DFT calculations and energy of intermolecularinteractions

The DFT computations with periodic boundary conditions(solid-state DFT calculations) were performed using the Crys-tal14 program.15 Details of the DFT computation proceduresare given in the ESI†. All the calculations were carried out usingthe B3LYP/6-31G** approximation and a Grimme modifiedempirical dispersion correction (f (R)C6/R6).16 The quantumtheory of atoms in molecule and crystal (QTAIMC) analysis17

of the periodic electron density obtained from the crystallinewave function was performed with TOPOND 14.18 The calcula-tion methodology is presented elsewhere.19 The followingelectron-density features at the (3;�1) bond critical point

(BCP) are computed: (i) the values of the electron density, rb,(ii) the Laplacian of the electron density, r2rb, and (iii) thepositively-defined local electronic kinetic energy density, Gb.Within the QTAIMC, the particular noncovalent intermolecularinteraction is associated with the existence of the bond path (i.e.the bond critical point) between the pair of atoms. The absence ofthe bond critical point implies that the two atoms do not interact.The network of the bond paths yields a comprehensive bondpicture, the energy of each specific interaction (in our case itis the intermolecular hydrogen bonds, C–H� � �O contact, etc.) isconsidered to be totally independent of the others. The effects ofthe crystal environment, long-term electrostatic effects, etc. aretaken into account implicitly, via the periodic electronic wave-function, and are coded in the bond critical point features. Theenergy of the particular noncovalent interaction, Eint, is evaluatedaccording to Mata et al.20 as

Eint = 0.429�Gb (in atomic units) (7)

Eqn (7) yields reasonable Eint values for molecular crystals withH-bonds, C–H� � �O and p-stacking contacts, etc.21

In addition, this approach gives an opportunity to estimatethe lattice energy of a crystal as a sum of the energies ofnoncovalent interactions between the considered moleculeand its neighbors:22

Elatt ¼Xi

Xjo i

Eint;j;i (8)

where j and i denote the atoms belonging to different mole-cules. Eqn (8) is BSSE free. For the sake of simplicity, indexes jand i will be omitted below.

It has to be pointed out that Eint,j,i values are commonlydetermined using atom-atom potentials23 or the PIXEL methodbased on semi-classical density sums.24 In the case of crystalswith proton transfer, however, the applicability of theseschemes for Eint and Elatt evaluation is not straightforward, atleast for the Gavezzotti model, as it was originally calibratedonly for molecular crystals containing neutral atoms. Switchingto ions thus requires preliminary parametrization of the forcefield. On the other hand, the QTAIMC concept is successfullyused both for salts and neutral co-crystals,21e,25 and eqn (7) wasderived using the interaction data of both neutral and charge-transfer heterodimers.20 Thus, Bader analysis of the calculatedwavefunction was chosen in the present work as superior to theGavezzotti model in estimating the energy of particular non-covalent interactions.

3. Results and discussion3.1 The salt formation and the pKa rule

The saccharinate salts with two substituted hydrazones withpotential antitubercular activity, vanillin isoniazid and salina-zid, were obtained by solvent-drop grinding, slurry conversionand crystallization from solution. DSC and PXRD analysis ofresulting solids revealed that different preparation procedureshave led to identical phases.

NJC Paper

Publ

ishe

d on

26

Aug

ust 2

015.

Dow

nloa

ded

by G

azi U

nive

rsite

si o

n 29

/10/

2015

09:

18:1

0.

View Article Online

This journal is©The Royal Society of Chemistry and the Centre National de la Recherche Scientifique 2015 New J. Chem., 2015, 39, 8614--8622 | 8617

Vanillin isoniazid and salinazid are weak bases. Their pKa

values based on pyridine nitrogen (3.57 and 3.60, respectively)26

are quite close to that of isoniazid (3.50), which is the structuralprecursor of the studied molecules. Therefore, both com-pounds are able to form either co-crystals or salts with differentorganic co-formers/counterions. It has been established in theliterature that the reaction of an acid with a base is expectedto form a salt if the difference in DpKa = pK(base)

a � pK(acid)a is

greater than 3, and it is most widely known as a ‘‘rule ofthree’’.27 This states that salt formation generally requires adifference of at least three pKa units between a conjugate baseand a conjugate acid. Cruz-Cabeza has conducted a study onthe relationship between the proton transfer probability andthe DpKa value of 6465 acid–base crystalline complexes takenfrom CSD.28 It has been shown that components with DpKa o�1 form mostly co-crystals, while systems with DpKa 4 4 tendto form exclusively salts. This ‘‘rule’’ has been also adjusted tovarious drug/co-former pairs,29 despite the fact that DpKa rangebetween 0 and 3 remains hardly predictable. For the studiedAPIs and saccharin (pKa = 2.32) the DpKa value equals ca. 1.3,which is in the middle of the range of the salt-co-crystalcontinuum. Therefore, the formation of either a salt or aco-crystal between the components is almost equally probable.On the other hand, the analysis of the Cambridge StructuralDatabase (CSD)30 has revealed that saccharin has a greaterpropensity for salt formation. A search of the CSD (Version5.36, 2014 release with Nov 2014 update) yields 68 two-component crystal structures of saccharinates and only 18co-crystals (saccharin saccharinates were not considered).

3.2 Crystal structures and thermal analysis

The crystal structures of pure vanillin isoniazid and salinazidhave been described earlier. CSD30 contains two records forvanillin isoniazid (ref. codes CANCOK and CANCOK01) and

four records for salinazid (ref. codes WEHFEU, WEHFEU01-03).All of these structures were found to be identical, whereasdifferent polymorphs were not observed. Crystal packingarrangement for the pure APIs is shown in Fig. S1 (see ESI†).Crystal structures of the compounds are quite similar andconsist of distinct zigzag layers of the molecules packed in a‘‘head-to-head’’ and ‘‘tail-to-tail’’ manner. Inside each layer, themolecules are linked by N–H� � �N hydrogen bonds to forminfinite chains with graph set notation C(7).31

Crystallographic data for the saccharin salts of vanillinisoniazid and salinazid are summarized in Table 1, and thepacking arrangements of the salts are shown in Fig. 2. Tosimplify the discussion of the salts, the following nomenclaturewill be applied: salt I (saccharinate of vanillin isoniazid), salt II(saccharinate of salinazid). The single crystal X-ray diffractiondata confirmed the protonation of the pyridine ring of the APIs,as evidenced by the proton location and bond length analysis.Both salts are found to be isostructural, and they crystallize inthe monoclinic P21/n space group with one API cation and onesaccharinate anion in the asymmetric unit.

The asymmetric unit contains API and saccharin moleculesconnected by N+3–H3� � �O6 hydrogen bonds (H-bonds) invol-ving the pyridine ring of the API and the carbonyl oxygen atomattached to the thiazole fragment of the saccharin (Fig. 2a).

In both structures, the pyridine ring of the API molecule liesapproximately in the same plane as the saccharin molecule towhich it is hydrogen-bonded (the angles between least-squaresplanes ca. 9.91 for salt I and ca. 8.91 for salt II). This spatialarrangement is probably stabilized by weak C1–H1� � �N�4 inter-action (green dash line in Fig. 2a), which completes a closed-ring dimeric unit. The second N1–H21� � �O4 hydrogen bond(Fig. 2a) connects two neighboring [API + saccharin] units toform a hydrogen bonded chain consisting of alternating APIand saccharinate ions along the c-axis, while the API molecules

Table 1 Crystallographic data for the API saccharinates

Compound reference Salt I Salt II

Chemical formula C14H14N3O3�C7H4NO3S C13H12N3O2�C7H4NO3SFormula Mass 454.45 424.43Crystal system Monoclinic Monoclinica/Å 6.1904(9) 6.1217(11)b/Å 26.105(4) 24.779(5)c/Å 12.4015(18) 12.246(2)b/1 100.464(2) 98.345(3)Unit cell volume/Å3 1970.8(5) 1838.0(6)Temperature/K 173(2) 173(2)Space group P21/n P21/nNo. of formula units per unit cell, Z 4 4Absorption coefficient, m/mm�1 0.215 0.220No. of reflections measured 16673 14883No. of independent reflections 4746 4404Rint 0.0553 0.0469Final R1 values (I 4 2s(I)) 0.0463 0.0549Final wR(F2) values (I 4 2s(I)) 0.0974 0.1114Final R1 values (all data) 0.0850 0.0794Final wR(F2) values (all data) 0.1112 0.1198Goodness of fit on F2 1.013 1.071Largest diff. peak & hole, e Å�3 0.280/�0.410 0.311/�0.420CCDC 1407124 1407125

Paper NJC

Publ

ishe

d on

26

Aug

ust 2

015.

Dow

nloa

ded

by G

azi U

nive

rsite

si o

n 29

/10/

2015

09:

18:1

0.

View Article Online

8618 | New J. Chem., 2015, 39, 8614--8622 This journal is©The Royal Society of Chemistry and the Centre National de la Recherche Scientifique 2015

are not directly connected with each other by hydrogen bonds(Fig. 2b). In addition, nearly planar [API + saccharin] in a chainforms an angle of ca. 681 with each other (Fig. 2c). Thispromotes a number of C–H� � �O and other weak interactionsbetween the neighboring molecules in the crystal. The descrip-tion and energies of these contacts will be discussed below.

The conformations of each API molecule can be defined interms of at least three torsion angles, one defining the con-formation of the central spacer unit between the two rings (t2)and two defining the orientation of the rings themselves(t1 and t3) (see Fig. 1). The values of the selected torsion anglesfor the pure APIs and the API saccharinates are representedin Table 2. In addition, we introduced an angle between the

aromatic rings referenced below as – b (the acute angle betweenthe least-squares planes through the two rings).

Table 2 shows that conformations of the API molecules inthe salts with saccharin are similar to those in the crystals ofthe pure APIs. In each molecule, the t2 torsion angle deviates byno more than �71 from 1801. In the case of t3, all the valuesare generally located around 01. However, the orientation of thepyridine ring (t1) is twisted from planarity relatively moresignificantly than the rest of the molecule, increasing the bvalues up to ca. 201.

The DSC traces for the salts, APIs and saccharin are shownin Fig. 3, and the thermal data are presented in Table 3. Thebulk materials were inspected by X-ray powder diffraction

Fig. 2 (a) Schematic representation of the hydrogen bonds (red dashlines) occurring in the salt crystals. Flexible torsion angles in the hydrazonemolecules are numbered and indicated by t1, t2 and t3; (b) molecularpacking projections for salt I along the a-axis and (c) salt II along the c-axis.

Table 2 Selected torsion angles and the dihedral angles between planesof aromatic rings, b, in the pure API molecules and the corresponding APIsaccharinates

t1 (C2–C3–C6–N1),1

t2 (C6–N1–N2–C7),1

t3 (N2–C7–C8–C9),1 b,1

Vanillin isoniazid 157.05 �176.92 2.68 15.57Salt I �169.17 173.71 �8.02 5.70

Salinazid �153.63 179.91 �4.15 21.28Salt II 169.17 �175.35 7.90 3.42

Fig. 3 DSC curves of the salts, saccharin, vanillin isoniazid and salinazidrecorded at a 10 K min�1 heating rate.

Table 3 Thermophysical data for the salts, compared to saccharin,vanillin isoniazid and salinazid

Tfus, K (onset) DHTfus

a, kJ mol�1 DSTfus

a, J mol�1 K�1

Vanillin isoniazid 507.4 � 0.3 49.2 � 1.0 96.9Salinazid 522.2 � 0.4 42.5 � 1.5 81.9Saccharin 500.2 � 0.2 30.3 � 1.0 60.5

Salt I 477.5 � 0.3 72.7 � 1.0 152.2Salt II 468.9 � 0.3 64.9 � 1.0 138.5

a For the salts, the values correspond to a mole of molecules in theasymmetric units.

NJC Paper

Publ

ishe

d on

26

Aug

ust 2

015.

Dow

nloa

ded

by G

azi U

nive

rsite

si o

n 29

/10/

2015

09:

18:1

0.

View Article Online

This journal is©The Royal Society of Chemistry and the Centre National de la Recherche Scientifique 2015 New J. Chem., 2015, 39, 8614--8622 | 8619

before being subjected to thermal analysis. The experimentalPXRD pattern was found to be in good agreement with thosecalculated from the single crystal analysis (Fig. S2, ESI†). As Fig. 3indicates, DSC thermograms have only one endotherm for all thecompounds which corresponds to the melting process.

The melting temperature of vanillin isoniazid is found to beca. 15 K lower than that of salinazid, however, the opposite trendis observed for the fusion enthalpies. Commonly, salt formationis applied to improve stability of APIs by increasing their meltingpoint, but according to the DSC experiments (Fig. 3), the saltformation of the API compounds decreases their melting point.The melting temperature of salt I is ca. 30 K lower compared tothat of the pure API (vanillin isoniazid), while for salt II, Tfus

decreases by ca. 53 K. It should be noted that the difference inmelting points of the salts is less than that of the pure APIs. Inaddition, an inversion of thermal stability occurs, i.e. vanillinisoniazid saccharinate (salt I) melts at the temperature of 9 Khigher than salinazid saccharinate (salt II) does.

3.3 Pattern of intermolecular interactions in the salts

As mentioned above, there are a number of relatively short contactsbetween the neighboring molecules in the salt crystals. However,the existence of short intermolecular contact does not imply theexistence of the intermolecular (noncovalent) interaction, e.g. seeref. 32. The Bader analysis of the computed periodical electrondensity enables us to detect and to quantify the noncovalentinteractions (besides conventional N–H� � �O bonds) in the salts.The obtained results are collected in Table S1 (see ESI†).

According to the QTAIMC analysis, the charge assisted N+3–H3� � �O6 hydrogen bond is characterized by rb 4 0.05 a.u. andbelongs to the intermediate type of interactions.33 Its energyreaches 57.1 kJ mol�1 in salt I and 58.8 kJ mol�1 in salt II(Fig. 4). This is the strongest intermolecular interaction in thecrystals of both salts. It should be noted that the energyobtained for the N+3–H3� � �O6 H-bond is in good agreementwith the value for the 3-hydroxypyridine–benzoic acid complex,where a similar proton transfer forms an N+–H� � �O bond.21e

The QTAIMC analysis also confirms the existence of a weak C1–H1� � �N�4 contact (Eint = 9.4–9.5 kJ mol�1), which completes aclosed-ring heterodimer between the molecules (Fig. 4). There-fore, the total energy of the supramolecular synthon formed bythese two interactions is calculated to be 66.5 kJ mol�1 for salt Iand 68.3 kJ mol�1 for salt II. Analysis of CSD showed that thistype of synthon is quite robust, and it is readily formed in thesaccharin salts with various pyridine derivatives.

The second N1–H21� � �O4 H-bond between API and saccharinmolecules is characterized by small positive r2rb values and theelectron density rb at the bond critical point lower than 0.02 a.u.According to Gatti, this intermolecular H-bond corresponds tothe closed-shell interactions.34 The energy of this H-bond isslightly larger in salt II (16.3 kJ mol�1) compared to that in salt I(14.6 kJ mol�1). In addition, the QTAIMC analysis reveals that theO4 and O5 atoms of saccharin act as acceptors of four C–H� � �Ointeractions from the neighboring API molecule (Fig. 4). The ener-gies of these contacts vary from 6.1 to 13.5 kJ mol�1 (Table S1, ESI†).It should be noted that the energy of the strongest contact

(C7–H7� � �O5) is found to be comparable to that of the conven-tional N1–H21� � �O4 H-bond. It suggests an important role ofC–H� � �O interactions in the lattice energy balance of the salts. Infact, the total energy of intermolecular interactions between APIand saccharin molecules linked by the N1–H21� � �O4 H-bondequals 53.2 kJ mol�1 for salt I and 55.3 kJ mol�1 for salt II, andthe contribution of C–H� � �O contacts is approximately 72%.

A number of the (3;�1) bond critical points correspondingto weak C–H� � �O contacts between the basic molecule of APIand the neighboring saccharin molecules was located by theQTAIMC analysis (Fig. S3, ESI†). The Eint values of thesecontacts vary from B4 to 11 kJ mol�1 (Table S1, ESI†). Thestrongest interactions were found between the carbonylO1-atom and H17-atom, which is attached to the phenyl ringof saccharin. The adjusted vanillin isoniazid molecules in salt Iwere also observed to interact with each other via the C14–H14B� � �O3 and C14–H14B� � �O2 contacts between the methoxyand hydroxy groups to form centrosymmetric dimers (Fig. S4and Table S1, ESI†). In addition, a weak C14–H14C� � �O2interaction was found between the API molecules related bysimple translation along the a-axis. Interestingly, such packingfeatures are not seen in the crystal structure of the pure vanillinisoniazid compound.

It might be reasonable to assume that the nearly planarorientation of the API molecules (b angle in Table 2 is close tozero) should promote a p-stacking in the salt crystals. Indeed, thefact that there were a set of BCPs in the structures can be attributed

Fig. 4 Intermolecular N–H� � �O hydrogen bonds (blue) and C–H� � �Ointeractions (green) in the crystals of (a) salt I and (b) salt II. The interactionenergies are given in kJ mol�1.

Paper NJC

Publ

ishe

d on

26

Aug

ust 2

015.

Dow

nloa

ded

by G

azi U

nive

rsite

si o

n 29

/10/

2015

09:

18:1

0.

View Article Online

8620 | New J. Chem., 2015, 39, 8614--8622 This journal is©The Royal Society of Chemistry and the Centre National de la Recherche Scientifique 2015

to p-stacking interactions between API–API and API–saccharinmolecules. It was previously established that the energy of thistype of intermolecular interaction may be confidently evaluatedin the same manner as H-bonds or C–H� � �O contacts, e.g. usingeqn (7).21b Calculations for salt I resulted in the following valuesof the p-stacking interaction energy: ca. 38 kJ mol�1 for API–APIstacks and ca. 15 kJ mol�1 for API–saccharin contacts (Fig. S5a,S6a and Table S1, ESI†). Similar values were obtained for salt II(Fig. S5b, S6b and Table S1, ESI†).

The values of the lattice energies of the salts evaluated usingeqn (8) were found to be 263.9 kJ mol�1 and 260.4 kJ mol�1 for saltsI and II, respectively. Therefore, in both structures, conventionalhydrogen bonds comprise ca. 28% of the lattice energy, while themost significant contribution is made by C–H� � �O(N) interactions,the value of which reaches ca. 42%. The rest of the packing energycorresponds to the p-stacking contacts (ca. 19%) and other weakintermolecular interactions (B11%). It would also be interesting toanalyze the sums of the intermolecular interaction energiesbetween different types of molecules in the crystal structure (TableS2, ESI†). It is evident that the API–saccharin interactions providethe largest contribution to the lattice energy (more than 70%).The API–API interactions comprise approximately a quarter of thetotal energy in salt I and ca. 16% in salt II, while there is almost nointeraction between the saccharin molecules.

3.4 Dissolution study

It is known that solubility in aqueous media is a key parameteramong other physicochemical properties for pharmaceuticalsolids. The dissolution profiles of vanillin isoniazid, salinazid

and the corresponding salts in water at 298.2 K are shown inFig. 5. The solubility values are shown in Table 4.

As Table 4 shows, the solubility of vanillin isoniazid at298.2 K is about 17% higher than that of salinazid. This maybe caused by the influence of the hydrophilic methoxy group inthe vanillin isoniazid molecule. In the case of the salts, anopposite tendency is observed: the solubility of salinazid sac-charinate (salt II) at 298.2 K is higher than the solubility ofvanillin isoniazid saccharinate (salt I) by approximately aquarter. It should be noted that the solubility values of thepure APIs as well as the saccharinates qualitatively agree withthe trend in their melting points, i.e. the compound with alower solubility melts at a higher temperature. Therefore, salt Idemonstrates a 12-fold solubility improvement compared topure API (vanillin isoniazid), whereas salt II is approximately20 times more soluble than the starting salinazid.

3.5 Thermodynamics of the salt formation

In spite of great interest in the structure, preparation and proper-ties of multi-component compounds such as co-crystals, salts,solvates, etc., there is relatively little data on their thermodynamiccharacteristics of formation, which are fundamental measures oftheir stability.35

The solubility data of vanillin isoniazid, salinazid, saccharinand the corresponding salts in water from 293.2 K to 308.2 K areshown in Table 4. Congruent solubility of the salts was observedat each temperature, and the solid phase recovered after theexperiment was identified by PXRD as the starting material(Fig. S7, ESI†). The 1 : 1 API–saccharin molar composition in thesaturated solutions was also confirmed by the HPLC analysis.

As Table 5 shows, the Gibbs energy of formation of salt Icalculated by eqn (3) is more negative than that of salt II, whichindicates its greater thermodynamic stability. Negative valuesof DG�f also suggest that the formation of the salts fromindividual components is a spontaneous process. The for-mation enthalpy of the salts was derived from eqn (5) (Fig. 6).It is evident that for both salts the enthalpy term providesthe most significant contribution to the driving force of the

Fig. 5 Dissolution profiles of vanillin isoniazid, salinazid and the salts inwater at 298.2 K.

Table 4 Temperature dependence of solubility, S0 (mol L�1), of the APIs, saccharin and the corresponding salts

Temperature

Vanillin isoniazid Salinazid Saccharin Salt I Salt II

S0 � 105 S0 � 105 S0 � 102 S0a � 104 ln Ksp S0

a � 104 ln Ksp

293.2 5.4 � 0.1 4.4 � 0.2 1.5 � 0.08 6.7 � 0.2 �14.6 8.7 � 0.3 �14.1298.2 6.9 � 0.2 5.7 � 0.2 1.7 � 0.06 8.4 � 0.2 �14.2 11.3 � 0.2 �13.6303.2 8.9 � 0.2 7.2 � 0.3 1.9 � 0.08 10.5 � 0.3 �13.7 14.2 � 0.3 �13.1308.2 11.3 � 0.3 9.4 � 0.3 2.1 � 0.09 13.1 � 0.4 �13.3 18.1 � 0.3 �12.6

a The numbers represent concentration (in mol L�1) of each of API and saccharin in a stoichiometric solution in equilibrium with the salt.

Table 5 Values of Kf and standard thermodynamic functions of the saltformation

Kf DG�f , kJ mol�1 DH�f , kJ mol�1 DS�f , J mol�1 K�1

Salt I 39.5 �9.1 �13.5 �14.7Salt II 19.2 �7.3 �18.4 �37.0

NJC Paper

Publ

ishe

d on

26

Aug

ust 2

015.

Dow

nloa

ded

by G

azi U

nive

rsite

si o

n 29

/10/

2015

09:

18:1

0.

View Article Online

This journal is©The Royal Society of Chemistry and the Centre National de la Recherche Scientifique 2015 New J. Chem., 2015, 39, 8614--8622 | 8621

formation process (Table 5). For salt II, the value of DH�f isfound to be ca. 5 kJ mol�1 larger than that for salt I.

It has to be pointed out that the enthalpy of formation is anintegral parameter which can be considered as the differencebetween the crystal lattice energy of the salt and the individualcomponents. However, the DFT calculations accompanied bythe QTAIMC analysis showed that the lattice energies of salts Iand II are closely comparable. Thus, the difference in theformation enthalpies of salts should be mostly caused bythe difference in the crystal lattice energy of the pure APIs,since the lattice energy of saccharin is a constant. Small valuesof the formation enthalpies are in good agreement with theresults of the DFT calculations, which indicates that the saltcrystals are mainly stabilized by C–H� � �O(N), p-stacking andother weak intermolecular interactions, while the hydrogenbonds comprise only 30% of the lattice energy.

The negative entropy change of the process indicates a moreefficient packing of the molecules in the salt compared to theirnative crystal structures. For salt II, the absolute value of DS�f isca. 2.5 times greater than that for salt I, while the difference in theformation enthalpies is only ca. 25%. As a result of this competitionbetween DH�f and DS�f , the Gibbs energy of salt II increases (i.e.becomes less negative) more appreciably compared to that of salt I.

Since the thermodynamic parameters of the formation of amulti-component compound are not a function of the solvent,it would be interesting to analyze the thermodynamic dataavailable for saccharin co-crystals. For example, it has beenreported by Oliveira et al.35a that the formation process of thecarbamazepine–saccharin (1 : 1) co-crystal is characterized bythe following values at 306 K: DG�f ¼ �4:4 kJ mol�1, DH�f ¼�5:9 kJ mol�1 and DS�f ¼ �4:9 J mol�1 K�1. Similar to salts Iand II, all the thermodynamic parameters of the co-crystalappeared with the negative sign. Their absolute values, however,are considerably lower than those of the salts. On the other hand,the co-crystal of saccharin with adefovir dipivoxil36 shows that theGibbs energy of formation equals ca. �12.8 kJ mol�1 at 303 K.35d Itindicates a greater affinity between adefovir dipivoxil and saccharinin the co-crystal compared to that of the APIs and saccharin in salts Iand II, despite the fact that no charge transfer occurs in the ade-fovir dipivoxil–saccharin system. The enthalpy and entropy terms of

the formation of the adefovir dipivoxil–saccharin co-crystal(DH�f � �40 kJ mol�1, DS�f � �90 J mol�1 K�1) are also foundto be significantly larger than those of salts I and II. Hence, asimilar competition between DH�f and DS�f contributions isobserved for the mentioned co-crystal. These examples demonstratethat thermodynamic parameters of the formation process may varyover a wide range. Nevertheless, a number of regularities have to bepointed out: (i) the largest contribution to the Gibbs energy offormation is provided by the enthalpy part, which characterizes thecrystal packing energy gain via different intermolecular interactionsbetween the components; (ii) the entropy change of the process hasa negative value, introducing a positive contribution to DG�f . As aresult, the formation of a co-crystal/salt is seen to be a consequenceof the competition between DH�f and DS�f terms.

4. Conclusions

The saccharin salts with two biologically active hydrazone com-pounds were obtained and their crystal structures were determined.The Bader analysis of the periodic electron density computed by thesolid-state DFT methods was performed to quantify the pattern ofintermolecular interactions in the salts. It was found that conven-tional N–H� � �O hydrogen bonds comprise ca. 28% of the latticeenergy, while the most significant contribution is made byC–H� � �O(N) interactions, the value of which reaches ca. 42%.The rest of the packing energy corresponds to p-stacking contacts(ca. 19%) and other weak intermolecular interactions (B11%).

The dissolution study of the compounds in water has shownthat salt formation of the drug molecules with saccharin sub-stantially increases their solubility. The saccharine salt of vanillinisoniazid demonstrates a 12-fold solubility improvement com-pared to the pure base, whereas salinazid saccharinate is approxi-mately 20 times more soluble than the starting drug. Both saltswere found to be stable and to dissolve congruently in water.

With the solubility of the salts and the corresponding solubilityof the pure compounds in water determined at different tempera-tures, the thermodynamic functions of the formation of the saltswere estimated. The Gibbs energy change of the process (DG�f ) wasfound to be �9.1 kJ mol�1 for vanillin isoniazid saccharinate and�7.3 kJ mol�1 for salinazid saccharinate, which indicates that theformation of the salts from individual components is a spontaneousprocess. The most significant contribution to the Gibbs energy isprovided by the enthalpy of formation, which characterizes thecrystal packing energy gain via different intermolecular interactionsbetween components. The entropy change of the process has anegative value, introducing a positive contribution to DG�f . As aresult, the formation of the salts is seen to be a consequence of thecompetition between enthalpy and entropy terms.

Acknowledgements

This work was supported by a grant from the President of theRussian Federation no. MK-67.2014.3. We thank ‘‘The UpperVolga Region Centre of Physicochemical Research’’ for thetechnical assistance of XRPD experiments.

Fig. 6 The van’t Hoff plots of ln Kf against reciprocal temperature of salt Iand salt II.

Paper NJC

Publ

ishe

d on

26

Aug

ust 2

015.

Dow

nloa

ded

by G

azi U

nive

rsite

si o

n 29

/10/

2015

09:

18:1

0.

View Article Online

8622 | New J. Chem., 2015, 39, 8614--8622 This journal is©The Royal Society of Chemistry and the Centre National de la Recherche Scientifique 2015

Notes and references

1 (a) S. Rollas and S- . G. Kuçukguzel, Molecules, 2007, 12, 1910;(b) R. Narang, B. Narasimhan and S. Sharma, Curr. Med.Chem., 2012, 19, 569.

2 (a) E. Vavrıkova, S. Polanc, M. Kocevar, J. Kosmrlj, K. Horvati,S. B+osze, J. Stolarıkova, A. Imramovsky and J. Vinsova, Eur.J. Med. Chem., 2011, 46, 5902; (b) F. Martins, S. Santos,C. Ventura, R. Elvas-Leitao, L. Santos, S. Vitorino, M. Reis,V. Miranda, H. F. Correia, J. Aires-de-Sousa, V. Kovalishyn,D. A. R. S. Latino, J. Ramos and M. Viveiros, Eur. J. Med. Chem.,2014, 81, 119; (c) R. Maccari, R. Ottana and M. G. Vigorita,Bioorg. Med. Chem. Lett., 2005, 15, 2509.

3 P. H. Buu-Hoi, D. Xuong, H. Nam, F. Binon and R. Royer,J. Chem. Soc., 1953, 75, 1358.

4 C. Vanucci-Bacque, C. Carayon, C. Bernis, C. Camare,A. Negre-Salvayre, F. Bedos-Belval and M. Baltas, Bioorg.Med. Chem., 2014, 22, 4269.

5 F. A. R. Rodrigues, A. C. A. Oliveira, B. C. Cavalcanti, C. Pessoa,A. C. Pinheiro and M. V. N. de Souza, Sci. Pharm., 2014, 82, 21.

6 (a) D. L. Prohotsky and F. Zhao, J. Pharm. Sci., 2012, 101, 1;(b) A. M. Thayer, Chem. Eng. News, 2007, 85, 17.

7 M. Pudipeddi, A. T. M. Serajuddin, D. J. W. Grant andP. H. Stahl, in Handbook of pharmaceutical salts, properties,selection and use, ed. P. H. Stahl and C. G. Wermuth, Wiley-VCH, Weinheim, Germany, 2002, pp. 19–40.

8 C. L. Cooke and R. J. Davey, Cryst. Growth Des., 2008, 8, 3483.9 R. Banerjee, P. M. Bhatt, N. V. Ravindra and G. R. Desiraju,

Cryst. Growth Des., 2005, 5, 2299.10 P. M. Bhatt, N. V. Ravindra, R. Banerjee and G. R. Desiraju,

Chem. Commun., 2005, 1073.11 G. M. Sheldrick, Acta Crystallogr., Sect. A: Found. Crystallogr.,

2008, 64, 112.12 G. M. Sheldrick, SADABS, Program for scaling and correction of

area detector data, University of Gottingen, Germany, 1997.13 R. R. Schartman, Int. J. Pharm., 2009, 365, 77.14 T. Rager and R. Hilfiker, Z. Phys. Chem., 2009, 223, 793.15 R. Dovesi, R. Orlando, A. Erba, C. M. Zicovich-Wilson,

B. Civalleri, S. Casassa, L. Maschio, M. Ferrabone, M. dela Pierre, P. D’Arco, Y. Noel, M. Causa, M. Rerat andB. Kirtman, Int. J. Quantum Chem., 2014, 114, 1287.

16 (a) B. Civalleri, C. M. Zicovich-Wilson, L. Valenzano andP. Ugliengo, CrystEngComm, 2008, 10, 405; (b) S. Grimme,J. Comput. Chem., 2006, 27, 1787.

17 (a) R. F. W. Bader, Atoms in Molecules – A Quantum Theory, OxfordUniversity Press, Oxford, 1990; (b) V. G. Tsirelson, in The QuantumTheory of Atoms in Molecules: From Solid State to DNA and DrugDesign, ed. C. Matta and R. Boyd, Wiley-VCH, Berlin, 2007, ch. 10.

18 C. Gatti and S. Casassa, TOPOND User’s Manual, CNR-ISTMof Milano, Milano, 2014.

19 (a) L. Bertini, F. Cargnoni and C. Gatti, Theor. Chem. Acc., 2007,117, 847; (b) A. V. Churakov, P. V. Prikhodchenko, O. Lev,A. G. Medvedev, T. A. Tripol’skaya and M. V. Vener, J. Chem.Phys., 2010, 133, 164506; (c) M. V. Vener, A. G. Medvedev,A. V. Churakov, P. V. Prikhodchenko, T. A. Tripol’skaya andO. Lev, J. Phys. Chem. A, 2011, 115, 13657.

20 I. Mata, I. Alkorta, E. Espinosa and E. Molins, Chem. Phys.Lett., 2011, 507, 185.

21 (a) M. V. Vener, A. N. Egorova, A. V. Churakov and V. G.Tsirelson, J. Comput. Chem., 2012, 33, 2303; (b) A. V. Shishkina,V. V. Zhurov, A. I. Stash, M. V. Vener, A. A. Pinkerton andV. G. Tsirelson, Cryst. Growth Des., 2013, 13, 816; (c) M. V. Vener,A. V. Shishkina, A. A. Rykounov and V. G. Tsirelson, J. Phys.Chem. A, 2013, 117, 8459; (d) A. N. Manin, A. P. Voronin, N. G.Manin, M. V. Vener, A. V. Shishkina, A. S. Lermontov andG. L. Perlovich, J. Phys. Chem. B, 2014, 118, 6803; (e) M. V.Vener, E. O. Levina, O. A. Koloskov, A. A. Rykounov, A. P. Voroninand V. G. Tsirelson, Cryst. Growth Des., 2014, 14, 4997.

22 P. M. Dominiak, E. Espinosa and J. Angyan, in ModernCharge Density Analysis, ed. C. Gatti and P. Macchi, Springer,Heidelberg, London, New York, 2012, pp. 387–433.

23 (a) A. J. Pertsin and A. I. Kitaigorodsky, The Atom–Atom PotentialMethod. Application to Organic Molecular Solids, Springer-Verlag,New York, 1987; (b) A. Gavezzotti, New J. Chem., 2011, 35, 1360.

24 (a) A. Gavezzotti, J. Phys. Chem., 2003, B107, 2344;(b) A. Gavezzotti, Mol. Phys., 2008, 106, 1473.

25 V. R. Hathwar, R. Pal and T. N. G. Row, Cryst. Growth Des.,2010, 10, 3306.

26 Advanced Chemistry Development (ACD/Laboratories) Soft-ware V11.02.

27 (a) K.-S. Huang, D. Britton, M. C. Etter and S. R. Byrn, J. Mater.Chem., 1997, 7, 713; (b) H. Stahl and C. G. Wermuth, Hand-book of Pharmaceutical Salts: Properties, Selection, and Use,Wiley-VCH, Zurich, 2002; (c) B. G. Bhogala, S. Basavoju andA. Nangia, CrystEngComm, 2005, 7, 551; (d) S. L. Childs,G. P. Stahly and A. Park, Mol. Pharmaceutics, 2007, 4, 323.

28 A. J. Cruz-Cabeza, CrystEngComm, 2012, 14, 6362.29 (a) A. C. Kathalikkattil, S. Damodaran, K. K. Bisht and

E. Suresh, J. Mol. Struct., 2011, 985, 361; (b) C. C. P. da Silva,R. de Oliveira, J. C. Tenorio, S. B. Honorato, A. P. Ayala andJ. Ellena, Cryst. Growth Des., 2013, 13, 4315; (c) P. Sanphui,S. Tothadi, S. Ganguly and G. R. Desiraju, Mol. Pharmaceu-tics, 2013, 10, 4687; (d) E. Elacqua, D.-K. Bucar, R. F. Henry,G. G. Z. Zhang and L. R. MacGillivray, Cryst. Growth Des.,2013, 13, 393; (e) L. H. Thomas, A. R. Klapwijk, C. Wales andC. C. Wilson, CrystEngComm, 2014, 16, 5924.

30 F. H. Allen, Acta Crystallogr., Sect. B: Struct. Sci., 2002, 58, 380.31 (a) M. C. Etter, Acc. Chem. Res., 1990, 23, 120; (b) J. Bernstein,

R. E. Davis, L. Shimoni and N.-L. Chang, Angew. Chem., Int.Ed. Engl., 1995, 34, 1555.

32 M. V. Vener, A. N. Egorova, D. P. Fomin and V. G. Tsirelson,J. Phys. Org. Chem., 2009, 22, 177.

33 M. V. Vener, A. V. Manaev, A. N. Egorova and V. G. Tsirelson,J. Phys. Chem. A, 2007, 111, 1155.

34 C. Gatti, Z. Kristallogr., 2005, 220, 399.35 (a) M. A. Oliveira, M. L. Peterson and R. J. Davey, Cryst. Growth

Des., 2011, 11, 449; (b) S. Zhang and A. C. Rasmuson, Cryst-EngComm, 2012, 14, 4644; (c) S. Zhang and A. C. Rasmuson,Cryst. Growth Des., 2013, 13, 1153; (d) K. Ma, Y. Zhang, H. Kan,L. Cheng, L. Luo, Q. Su, J. Gao, Y. Gao and J. Zhang, Pharm.Res., 2014, 31, 1766.

36 Y. Gao, H. Zu and J. Zhang, J. Pharm. Pharmacol., 2011, 63, 483.

NJC Paper

Publ

ishe

d on

26

Aug

ust 2

015.

Dow

nloa

ded

by G

azi U

nive

rsite

si o

n 29

/10/

2015

09:

18:1

0.

View Article Online

View publication statsView publication stats