New Pyrazole-Hydrazone Derivatives

14 Pages·2017·2.53 MB·English

International Journal o f
Molecular Sciences
Article
New Pyrazole-Hydrazone Derivatives: X-ray Analysis,
Molecular Structure Investigation via Density
Functional Theory (DFT) and Their High In-Situ
Catecholase Activity
KhalidKarrouchi1,2,3,ElBekkayeYousfi4,NadaKheiraSebbar5,YoussefRamli1,
JamalTaoufik1,YounesOuzidan6,M’hammedAnsar1,YahiaN.Mabkhot7,* ID,
HazemA.Ghabbour8 ID andSmaailRadi2,* ID
1 LaboratoiredeChimieThérapeutique,FacultédeMédecineetdePharmacie,UniversitéMohammedV,
P.O.Box8007,Rabat10100,Morocco;[email protected](K.K.);[email protected](Y.R.);
jataoufi[email protected](J.T.);[email protected](M.A.)
2 LaboratoiredeChimieAppliquéeetEnvironnement(LCAE),FacultédesSciences,UniversitéMohamedI,
P.O.Box524,Oujda60000,Morocco
3 LaboratoireNationaldeContrôledesMédicaments,DirectionduMédicamentetdelaPharmacie,
MinistèredelaSanté,P.O.Box6206,Rabat10100,Morocco
4 InstitutionSupérieuredesProfessionsInfirmièresetTechniquesdeSanté,P.O.Box4806,Oujda60000,
Morocco;yousfi@netcourrier.com
5 LaboratoiredeChimieOrganiqueHétérocyclique,Pharmacochimie,Facultédessciences,
UniversitéMohammedV,P.O.Box8007,Rabat10100,Morocco;[email protected]
6 LaboratoiredeChimieOrganiqueAppliquée,FacultédesSciencesetTechniques,UniversitéSidiMohamed
BenAbdellah,P.O.Box2202,Fès30000,Morocco;[email protected]
7 DepartmentofChemistry,FacultyofScience,KingSaudUniversity,P.O.Box2455,
Riyadh11451,SaudiArabia
8 DepartmentofPharmaceuticalChemistry,CollegeofPharmacy,KingSaudUniversity,P.O.Box2457,
Riyadh11451,SaudiArabia;[email protected]
* Correspondence:[email protected](Y.N.M.);[email protected](S.R.);Tel.:[phone](S.R.)
Received:18September2017;Accepted:21October2017;Published:25October2017
Abstract:Thedevelopmentoflow-costcatalyticsystemsthatmimictheactivityoftyrosinaseenzymes
(Catecholoxidase)isofgreatpromiseforfuturebiochemistrytechnologicdemands.Herein,wereport
thesynthesisofnewbiomoleculessystemsbasedonhydrazonederivativescontainingapyrazole
moiety (L1–L6) with superior catecholase activity. Crystal structures of L1 and L2 biomolecules
weredeterminedbyX-raysinglecrystaldiffraction(XRD).Optimizedgeometricalparameterswere
calculatedbydensityfunctionaltheory(DFT)atB3LYP/6–31G(d,p)levelandwerefoundtobein
goodagreementwithsinglecrystalXRDdata. Copper(II)complexesofthecompounds(L1–L6),
generated in-situ, were investigated for their catalytic activities towards the oxidation reaction
of catechol to ortho-quinone with the atmospheric dioxygen, in an attempt to model the activity
of the copper containing enzyme tyrosinase. The studies showed that the activities depend on
four parameters: the nature of the ligand, the nature of counter anion, the nature of solvent and
the concentration of ligand. The Cu(II)-ligands, given here, present the highest catalytic activity
(72.920µmol·L−1·min−1)amongthecatalystsrecentlyreportedintheexistingliterature.
Keywords: pyrazole;hydrazone;crystalstructure;DFT;catecholaseactivity
Int.J.Mol.Sci.2017,18,2215;doi:10.3390/ijms18112215 www.mdpi.com/journal/ijms
Int.J.Mol.Sci.2017,18,2215 2of14
1. Introduction
Bioorganometallic compounds are reputed for their remarkable applications in the field of
catalysis;however,muchlessisknownabouttheirpotentialinchemicalbiology[1,2]. Animportant
goal in organometallic chemistry is the synthesis of biomolecules that exhibit catalytic activity
analogoustotheactivityofenzymes. Anumberofcatalystshavingbiomimeticactivityfordifferent
enzymes have been designed by chemists [3,4]. The preparation and etude of efficient template
complexesformetalloenzymeswithoxidaseoroxygenaseactivityarethusmostimportantforthe
elaborationofnewveritablecatalystsforoxidationreactions[5]. Copperions,ascentersofpotent
siteofvariousmetalloproteins,playanessentialroleinseveralbiologicalprocesses: electrontransfer,
oxidation,transportofdioxygen,etc.[6,7]. Coppercomplexesoflowmolecularweightarestudiedas
structuralandfunctionalmodelsofactivecentersofenzymeswithcopper[8–11].
Catecholoxidasesperformtheoxidationof1,2-diphenols,suchascatechol,too-quinones,using
dioxygen(O ). Hydrogenatomsremovedfromcatecholcombinewithoxygentoformwater. The
2
crystalstructuresofdifferentformsofcatecholoxidasehaveimprovedtheaccordanceofthemechanism
of catecholase activity of catechol oxidase. Several workers proposed the mechanism of catechol
oxidationbynaturalenzyme,includingimportantproposalsbySolomonandkerbs[12,13].Inaddition
tothehighnumberofCu(II)complexesreported,theircatecholaseactivityhasalsobeendemonstrated,
althoughtheirmechanisticaspectsarenotaswellunderstoodasthoseofCu(II)complexes[14–18].
HydrazonesaremembersoftheSchiffbasesfamily,whicharebuiltwitharomaticacidhydrazides
and carbonyl compounds. They are quite interesting in coordination chemistry as they present a
combinationofdonorsites,suchasaprotonated/deprotonatedamideoxygenatom,iminenitrogen
atom of the hydrazone moiety and an additional donor site (usually N or O) provided from the
aldehydeorketone[19,20]. Hydrazonesformwidevarietyofcomplexeswithchemical,structural,
biologicalandindustrialimportance[21–23]. Theseproprietiesareattributedtotheformationofstable
chelatecomplexeswithtransitionmetalswhichcatalyzephysiologicalprocesses[24,25].
Inthisstudy,wereportthesynthesisofsixnewhydrazonederivativescontainingpyrazolemoiety
withagoodyield(Scheme1). TheX-raycrystalstructuresofcompoundsL1andL2weredetermined,
andtheirgeometricalparameterswerecomparedwiththeoreticalDFTcalculationsattheB3LYPlevel
oftheory. Theinvestigationofcatalyticactivitiesofcopper(II)-ligandcomplexestowardsoxidation
of catechol to o-quinone was studied. All of the parameters that can affect the catalytic efficiency
wereIsntt.u J.d Mieold. S.ci. 2017, 18, 2215 3 of 15
O R
1
R3 R1
a R H
HN OEt HN NHNH2 2 HN N N
N N b N R2
O O O
R
3
1 2
L1 R =H;R =OH;R =OCH
1 2 3 3
L2 R =H;R =CH;R =H
1 2 3 3
L3 R =H;R =Cl;R =H
1 2 3
L4 R =H;R =F;R =H
1 2 3
L5 R =CH;R =H;R =H
1 3 2 3
L6 R1=Ph;R2=H,R3=H
Scheme 1. The synthetic routes of compounds L1–L6: (a) hydrazine hydrate (80%), ethanol, reflux 5h;
Scheme1.ThesyntheticroutesofcompoundsL1–L6:(a)hydrazinehydrate(80%),ethanol,reflux5h;
and (b) ethanol, acetic acid, reflux, 2–5 h.
and(b)ethanol,aceticacid,reflux,2–5h.
2.2. X-Ray Crystal Structures Description
Compounds L1 and L2 were analyzed by X-ray diffraction. Refinement parameters and crystal
data are listed in Table S1. Supplementary data are deposited with the Cambridge Crystallographic
Data Centre (CCDC) under deposition numbers 1522882 and 1523265.
The selected bond lengths and bond angles and hydrogen bonds are listed in Tables S2–S5 in the
Supplementary Materials. The asymmetric unit of L1 contains one independent molecule with DMF
and water molecules as mixed crystallizing solvent, as shown in Figure 1. All the bind lengths and
angles are in normal ranges. In the crystal wrap, molecules are connected via many classical and non-
classical intermolecular hydrogen bonds (Table S3). Figure 2 shows the crystal structure of L2. In the
crystal structure, the pyrazole ring (N1/N2/C7–C9) makes dihedral angles with the phenyl ring (C1–
C6) and tolyl ring (C12–C17), at 23.39° and 36.16°, respectively. In the crystal wrap, molecules are
linked via two classical intermolecular hydrogen bonds between N1—H1N1···O1i and N3—
H1N3···N2ii, Symmetry codes: (i) x, −y+1, z+1/2; and (ii) −x+1, y, −z+3/2 (Table S5).

Figure 1. Asymmetric unit of L1 (CCDC 1522882).
Int.J.Mol.Sci.2017,18,2215 3of14
Int. J. Mol. Sci. 2017, 18, 2215 3 of 15
O R
1
2. Results
2.1. Synthesis
R3 R1
SynHtNhesis oOfEthydraazonesHN(L1–L6)NHiNsH2outlined Rin2 SchemeHN1. SHNtarNting compound,
ethyl 3-phNenyl-1H-pyrazole-5-carbNoxylate (1), was rbeadily synthesiNzed by the reactiRo2n of
O O O
ethyl-2,4-dioxo-4-phenyl-butanoate, obtained from acetophenone, and diethyl oxalRate, with
3
1 2
hydrazine in the presence of sulfuric acid at room temperature. The reaction of
L1 R =H;R =OH;R =OCH
ethyl 3-phenyl-1H-pyrazole-5-carboxylate (1) with hydrazine hy1drate2 in et3hanol3 afforded
L2 R =H;R =CH;R =H
1 2 3 3
3-phenyl-1H-pyrazole-5-carbohydrazide (2). Finally, the novel dLe3siRred=Hh;yRd=rCalz;oRn=eHs (L1–L6) were
1 2 3
obtainedbycondensingcompound(2)witharomaticaldehydesatLr4eflRu1x=Hof;Re2t=hFa;nRo3l=uHsingaceticacid
L5 R =CH;R =H;R =H
asreportedinourpreviousprocedure[26–28]. Theexpectedproduct1swer3ei2solate3dascrystalline
L6 R1=Ph;R2=H,R3=H
materialswithreliabletoexcellentyields.
Scheme 1. The synthetic routes of compounds L1–L6: (a) hydrazine hydrate (80%), ethanol, reflux 5h;
2.2. X-RayanCdr y(bs)t aelthSatnroul,c atucerteics aDcieds,c rreifplutixo, n2–5 h.
C2.o2m. Xp-Rouayn CdrsysLt1al aSntrducLtu2rwes eDreescarnipatlioynz edbyX-raydiffraction. Refinementparametersandcrystal
dataarelistedinTableS1. SupplementarydataaredepositedwiththeCambridgeCrystallographic
Compounds L1 and L2 were analyzed by X-ray diffraction. Refinement parameters and crystal
DatadCaetna tarree( lCisCteDd Cin) Tuanbdlee rS1d. eSpuopspitleiomnenntuamryb deartsa 1a5re2 2d8e8p2osaintedd 1w5i2th3 2t6h5e .Cambridge Crystallographic
TDhaetas Celeencttreed (CbCoDnCd) luenndgetrh dseapnodsitbioonn nduamnbgelress 1a52n2d88h2y adnrdo 1g5e2n32b6o5n. dsarelistedinTablesS2–S5in
theSupplTehmee snelteacrtyedM boantedr lieanlsg.thTsh aenda sbyomndm aentgrliecs uanndit hoyfdLro1gceonn btoanindss aornee lisintedde ipne Tnadbelenst Sm2–oS5le icnu tlhee with
DMFSaunpdplwemateenrtamryo Mlecautelreisalas.s Tmheix aesdymcrmyesttrailcl iuzninitg ofs oLl1v ceonntt,aainsss ohnoew inndienpeFnigduenret m1.oAlecllutleh ewbitihn DdMleFn gths
and aanngdl ewsaaterre minolnecourlmesa alsr manixgeeds .crIynsttahlleizcinrgy sstoallvewnrt,a aps, smhoowlenc uinle Fsigaurree c1o. nAnlle tchtee dbivnida lemnagnthys calnads sical
angles are in normal ranges. In the crystal wrap, molecules are connected via many classical and non-
and non-classical intermolecular hydrogen bonds (Table S3). Figure 2 shows the crystal structure
classical intermolecular hydrogen bonds (Table S3). Figure 2 shows the crystal structure of L2. In the
of L2. In the crystal structure, the pyrazole ring (N1/N2/C7–C9) makes dihedral angles with the
crystal structure, the pyrazole ring (N1/N2/C7–C9) makes dihedral angles with the phenyl ring (C1–
phenylring(C1–C6)andtolylring(C12–C17),at23.39◦ and36.16◦,respectively. Inthecrystalwrap,
C6) and tolyl ring (C12–C17), at 23.39° and 36.16°, respectively. In the crystal wrap, molecules are
moleculesarelinkedviatwoclassicalintermolecularhydrogenbondsbetweenN1—H1N1···O1iand
linked via two classical intermolecular hydrogen bonds between N1—H1N1···O1i and N3—
N3—HH11NN33·····N·N2i2i,i iS,ySmymmemtreyt croydceosd: (ei)s :x,( i−)yx+,1−, zy+1+/12,; za+nd1/ (i2i); −axn+d1,( yii,) −−z+x3+/21 (,Tya,b−lez S+53)/. 2(TableS5).

FigFuirgeur1e. 1A. sAysmymmmeetrtricicu unniitt ooff LL11 ((CCCCDDCC 1512522828828).2 ).
Int.J.MInot.l .J.S Mcio.l2. S0c1i7. ,201187,,2 1281, 52215 4 of 154 of14

Figure2.AsymmetricunitofL2(CCDC1523265).
Figure 2. Asymmetric unit of L2 (CCDC 1523265).
2.3. C2o.m3. pCuotmatpiuotnaatiloSntaul Sdtieusdies
ComCpoamrispoanrisoofnt hofe tthhee tohreeotriectaiclavl avlauleusesw witihth tthhee eexxppeerriimmeenntatal olnoense isndinicdaitceast tehsatth aallt tahlel othpetimopizteimd ized
bondbloenndg tlehnsgathres acrloe scelorsteor ttho etheex epxepreimrimenentatall vvaalluueess.. IInn ththee cacsaes eofo Xf-Xra-yra sytrustcrtuurcet uorfe coomfcpoomunpdo Lu1n, dL1,
theobthsee rovbesdervbeodn dbolnedn gletnhgsthosf oCf1 C–1C–1C01,0O, O33––CC1155 aanndd NN33––NN44 bboonndds sini nfivfiev-me-emmebmerbeedr peydrapzyorlaez roinleg arirne gare
1.227(3) Å, 1.363(3) Å and 1.383(3) Å, respectively. The calculated bond lengths, through DFT method,
1.227(3)Å,1.363(3)Åand1.383(3)Å,respectively. Thecalculatedbondlengths,throughDFTmethod,
of same pyrazole ring are 1.24394 Å, 1.38125 Å and 1.37144 Å, respectively, which are very close to
ofsamepyrazoleringare1.24394Å,1.38125Åand1.37144Å,respectively,whichareveryclosetothe
the actual values. In Table 1, it is clear that actual C-C and C-H bond lengths are also in close
actualvalues. InTable1,itisclearthatactualC-CandC-Hbondlengthsarealsoincloseagreement
agreement with calculated values. The calculated bond angles for O1–C10–C9, C14–O2–C18 and N4–
withcalculatedvalues. ThecalculatedbondanglesforO1–C10–C9,C14–O2–C18andN4–C11–C12
C11–C12 bond angles of L1 are 122.22°, 118.52°, 121.37°, respectively, which are close to the
bondcaonrrgelsepsonodfiLng1 aacrteua1l2 a2n.g2l2e◦s, o1b1ta8i.n5e2d◦ ,fr1o2m1. X37-r◦a,yr. eTshpee accttiuvaell yv,awluehsi cohf aabroevcel boosnedt oantghleesc aorrer 1e2s1p.o3°n, ding
actua1l1a8n.3g°,l easndo b12ta2i.9n°e, drefsrpoemctivXe-lrya. y. Theactualvaluesofabovebondanglesare121.3◦, 118.3◦, and
122.9◦,respectively.
Table 1. Selected structural parameters by X-ray and theoretical calculations of compound L1.
BoTnadb Lleen1g.tShe lectEexdpesrtirmucetnutraal lparaCmalectuelrastebdy X-rayandtheoreticalcEaxlcpuerlaimtioenntsalo fcomCpoalucnudlatLe1d.
Bond Angle (°)
(Å) Bond Lengths Bond Lengths Bond Angles Bond Angles
BonOd1L–eCng1t0h Exp1e.2ri2m(3e)n tal Ca1l.c2u4l ated NBo2n–dCA9–nCg8le (◦) E1x0p6e.0r(i2m) ental C10a9lc.2u lated
O(Å2–)C14 Bon1d.3L6e(3n)g ths Bon1d.3L9e ngths O1–C10–N3 B12o2n.d8(A2)n gles B1o2n5d.0A ngles
OO1–2C–1C018 11..4212((43)) 1.14.52 4 N3N–C2–1C0–9C–C9 8 1161.006(.20)( 2) 1121.079 .2
OO2–3C–1C415 11..3366((33)) 1.13.83 9 O1O–1C–1C01–0C–9N 3 1211.232(.28)( 2) 1221.225 .0
O2N–1C–1N82 11..3431((34)) 1.13.74 5 N4N–C3–1C1–1C0–1C2 9 1221.196(.20)( 2) 1211.132 .7
O3–C15 1.36(3) 1.38 O1–C10–C9 121.3(2) 122.2
N1–C7 1.33(3) 1.38 O2–C14–C13 125.7(2) 126.0
N1–N2 1.33(3) 1.37 N4–C11–C12 122.9(2) 121.3
N2–C9 1.34(3) 1.36 C14–O2–C18 118.3(2) 118.5
N1–C7 1.33(3) 1.38 O2–C14–C13 125.7(2) 126.0
NN2–3C–N94 11..3384((33)) 1.13.73 6 NC21–4N–1O–2C–7C 18 1051.138(.23)( 2) 1131.128 .5
NN33––NC410 11..3348((33)) 1.13.83 7 N1N–2N–2N–1C–9C 7 1121.065(.23)( 2) 1051.103 .2
NN3–4C–1C011 11..2374((33)) 1.12.93 8 N4N–N1–3N–C2–1C0 9 1191.152(.26)( 2) 1211.005 .0
N4–C11 1.27(3) 1.29 N4–N3–C10 119.5(2) 121.0
The optimized geometry of compounds L1 and L2 were obtained at B3LYP/6-31G* level. Some
optimized geometric parameters are also listed in Figure 3, and Tables 2 and 3.
TheoptimizedgeometryofcompoundsL1andL2wereobtainedatB3LYP/6-31G*level. Some
optimizedgeometricparametersarealsolistedinFigure3,andTables2and3.
Int.J.Mol.Sci.2017,18,2215 5of14
Int. J. Mol. Sci. 2017, 18, 2215 5 of 15

L1
L2
FigFiugruere3 .3.O Oppttiimmiizzeedd ggeeoommeetrtyry oof Lf1L 1anadn Ld2L. 2.
The total energy, energy of HOMO and energy of LUMO, as well as other parameters for
Table2.SelectedstructuralparametersbyX-rayandtheoreticalcalculationsofcompoundL2.
structures L1 and L2 are obtained theoretically and listed in Table 3. The HOMO and LUMO electron
density distributions of L1 and L2 are given in Figures 4 and 5, respectively. After the analysis of the
BthoenodreLteicnaglt rhesultsE oxpbetariinmeedn,t wale can Csaaylc tuhlaatt emdolecuBleosn dL1A anngdle L(◦2) havEe xap neroinm-penlatanlar struCctaulcrue.l aDteFdT
(Å) BondLengths BondLengths BondAngles BondAngles
calculation gives an idea about the substance reactivity and site selectivity of the frameworks.
O1–C10 1.22(2) 1.21 N2–N1–C7 113.4(15) 113.6
EHOMO; ELUMO, which clarifies the inevitable charge exchange collaboration inside the studied
N1–N2 1.34(2) 1.34 N1–N2–C9 103.9(15) 105.4
material; electronegativity (χ); hardness (η); potential (μ); electrophilicity (ω); softness (S); and
N1–C7 1.34(3) 1.37 N4–N3–C10 119.2(17) 117.2
sofNtn2e–sCs 9(σ) are recor1d.e3d3( 2in) Table 3. The1 s.3ig4nificance oNf 3η– Nan4d–C ϭ1 i1s to eval1u1a5t.e6 (b1o8t)h the reactiv11it4y.2 and
staNbi3li–tNy.4 1.38(2) 1.41 N1–C7–C6 122.7(17) 122.39
N3–C10 1.33(3) 1.36 N1–C7–C8 105.3(18) 103.59
N4–CT11able 2. Select1e.d2 6st(r3u)ctural parame1te.2rs7 by X-ray anNd 2th–eCo9r–etCic8al calculat1io1n1s.4 o(1f 6c)ompound L21. 09.21
C15–C18 1.512(4) 1.50 N2–C9–C10 119.4(17) 119.83
BoNn1d– LHength Exp0e.r8i6m(2e)ntal Calc1u.l0a0ted BonNd3 A–Cng10le– C(°9) Exper1i1m4e.9n(t1a6l) Calculate1d1 1B.0o5nd
(Å) Bond Lengths Bond Lengths Bond Angles Angles
O1–C10 1.22(2) 1.21 N2–N1–C7 113.4(15) 113.6
TheNt1o–tNal2 energy, 1e.n34e(r2g)y of HOM1O.34a nd enerNg1y–Nof2–LCU9 MO, a1s03w.9(e1l5l) as other1p0a5r.4a meters for
structureNs1L–1Ca7n dL2are1.o3b4(t3a)i nedtheore1t.i3c7a llyandNlis4t–eNd3i–nCT1a0b le3.1T1h9.e2(H17O) MOand1L1U7.M2 Oelectron
density dNi2s–trCib9 utions of1.L331(2a)n d L2 are g1.i3v4e n in FigNu3r–eNs44–Can11d 5, re1s1p5e.6c(t1iv8)e ly. After1t1h4e.2 analysis of
the theorNe3t–icNa4l results o1b.3t8a(i2n)e d, we can1.s4a1y that moNl1e–cCu7le–sC6L 1 and1L222.7h(1a7v)e a non-p1l2a2n.3a9r structure.
DFTcalcNu3l–aCti1o0n givesa1n.3i3d(3e)a aboutthe1.s3u6 bstancerNe1a–cCti7v–iCty8 andsi1t0e5s.3e(l1e8c)t ivityoft1h0e3.5fr9a meworks.
N4–C11 1.26(3) 1.27 N2–C9–C8 111.4(16) 109.21
EHOMO;ELUMO,whichclarifiestheinevitablechargeexchangecollaborationinsidethestudied
C15–C18 1.512(4) 1.50 N2–C9–C10 119.4(17) 119.83
material;electronegativity(χ);hardness(η);potential(µ);electrophilicity(ω);softness(S);andsoftness
N1–H 0.86(2) 1.00 N3–C10–C9 114.9(16) 111.05
(σ)arerecordedinTable3. Thesignificanceofηandσistoevaluateboththereactivityandstability.
Int. J. Mol. Sci. 2017, 18, 2215 6 of 15
Table 3. Calculated energies of L1 and L2.
Molecular Energy (a.u.)(eV) L1 L2
TE −31008.6 −26911.4
EHOMO −5.8186 −6.4850
ELUMO −1.0152 −0.7349
Gap ΔE 4.8034 5.7500
Chemical potential µ (D) 5.8871 6.3122
Ionization potential (IP) 5.8186 6.4850
Electron affinity (EA) 1.0152 0.7349
Electron negatiity (χ) 3.4169 3.6100
Int.J.Mol.Sci.2017,18,2215 Global hardness (η) 2.4017 2.875 6of14
Global electrophilicity (ω) 2.4306 2.2664
Table3.CalculatedenergiesofL1andL2.
The analysis of the wave function indicates that the energy space between the molecular orbit
HOMO and LUMO determines the chemical stability and the electrical transport properties of the
MolecularEnergy(a.u.)(eV) L1 L2
molecule. The red and green colors of the molecular orbital ridge, respectively, represent the positive
and negative phasesT. E −31008.6 −26911.4
The HOMO oEfH LO1M sOhows the charge density lo−ca5li.8z1e8d6 on the 4-hydroxy-3-m−e6t.h4o8x5y0benzaldehyde
E −1.0152 −0.7349
ring, while LUMO LiUs MchOaracterized by a charge distribution on the hydrazone function, indicating
Gap∆E 4.8034 5.7500
that this moiety can influence the electron transition. The HOMO of L2 has a localized charge density
Chemicalpotentialµ(D) 5.8871 6.3122
on the pyrazole and hydrazone function, but LUMO is characterized by a charge distribution on the
Ionizationpotential(IP) 5.8186 6.4850
4-methylbenzaldehyde ring and the hydrazone function. The energy difference between HOMO and
Electronaffinity(EA) 1.0152 0.7349
LUMO of LE1l eacntrdo Ln2n iesg aabtioituyt (4χ.3)8 and 5.75 eV, resp3ec.4t1iv6e9ly. 3.6100
The enGerlgoyba olfh athrde nsemssa(llηe)r band space increa2s.e4s0 1th7e stability of the mole2c.u87le5. The molecular
boundarGy loorbbailtaellse cotfr oLp1h ainlidci tLy2 ((ωH)OMO–LUMO) ar2e.4 s3h0o6wn in Figures 4 and 5, 2r.e2s6p6e4ctively.

Figure4.HOMO–LUMOenergydiagramofL1.
Int. J. Mol. Sci. 2017, 18, 2215 Figure 4. HOMO–LUMO energy diagram of L1. 7 of 15

Figure5.HOMO–LUMOenergydiagramofL2.
Figure 5. HOMO–LUMO energy diagram of L2.
Theanalysisofthewavefunctionindicatesthattheenergyspacebetweenthemolecularorbit
2.4. Catecholase Activity: Spectrophotometric Study
HOMOandLUMOdeterminesthechemicalstabilityandtheelectricaltransportpropertiesofthe
Catechol oxidation reaction catalyzed by copper complexes with ligands L1–L6 is followed by
molecule. Theredandgreencolorsofthemolecularorbitalridge,respectively,representthepositive
the evolution of o-quinone absorbance measured at 390 nm with a UV-vis spectrometer (Scheme 2).
andnegativephases.
1/2O2 H2O
OH O
OH O
RT
Catechol ortho-Quinone
Ligand/Metal
(L1-L6)/Cu
Scheme 2. Catecholase reaction.
The catalytic oxidation rate variations from one complex to another are shown in Figures S1–S5.
On the other hand, catechol oxidation rates were calculated and collected in Table 4. According to
these results, we find that all copper complexes, formed in situ from ligands L1–L6 and salts of
copper, catalyze the oxidation reaction of catechol to o-quinone, but with different rates. The catalytic
activities depend highly on the ligand concentration, the nature of solvent and the type of inorganic
anion (Figures S1–S5).
Table 4. Oxidation rate of catechol oxidation in methanol (μmol·L−1·min−1).
Ligand/Metallic Salt Cu(NO3)2 CuCl2 Cu(CH3COO)2 CuSO4
L1 10.57 10.28 9.80 9.27
L2 15.52 0.05 22.92 16.06
L3 37.89 9.19 24.58 21.01
L4 17.71 11.43 15.02 6.57
L5 3.10 8.07 19.62 10.61
L6 27.77 40.27 60.50 72.92
It appears plainly that catalytic activity varies from one complex to another. All of the copper
complexes formed by the hydrazone derivatives catalyze the oxidation reaction of catechol to o-
quinone. The best is the L6 ligand with highest oxidation rate for all copper salts with maximum
Int.J.Mol.Sci.2017,18,2215 7of14
TheHOMOofL1showsthechargedensitylocalizedonthe4-hydroxy-3-methoxybenzaldehyde
ring,whileLUMOischaracterizedbyachargedistributiononthehydrazonefunction,indicating
thatthismoietycaninfluencetheelectrontransition. TheHOMOofL2hasalocalizedchargedensity
onthepyrazoleandhydrazonefunction,butLUMOischaracterizedbyachargedistributiononthe
4-methylbenzaldehyderingandthehydrazonefunction. TheenergydifferencebetweenHOMOand
LUMOofL1andL2isabout4.38an d5.75eV,respectively.
The ener1g yToypfe otfh thee Psampera (Allreticrle,b Raevniewd, Csopmamcuneicaitnionc,r eetca.) ses the stability of the molecule. The molecular
boundaryorb2i talTsiotflLe1 andL2(HOMO–LUMO)areshowninFigures4and5,respectively.
2.4. Catecholas3e AFcitrisvtniatmye: LSapstencamtreo 1p, Fhirosttnoammee tLraisctnSamtue d2 aynd Firstname Lastname 2,*
4 1 Affiliation 1; [email protected]
Catechol5o xid2 aAtfifoilinatiorne 2a; [email protected] zedbycoppercomplexeswithligandsL1–L6isfollowedbythe
6
evolutionofo7- qui noneabsorbancemeasuredat390nmwithaUV-visspectrometer(Scheme2).
8
9
10
11
12 Scheme2.Catecholasereaction.
13
14
15
Thecata16ly tic oxidationratevariationsfromonecomplextoanotherareshowninFiguresS1–S5.
17
Ontheother18h and ,catecholoxidationrateswerecalculatedandcollectedinTable4. Accordingto
theseresults,19w efi ndthatallcoppercomplexes,formedinsitufromligandsL1–L6andsaltsofcopper,
20
catalyzetheo21x ida tionreactionofcatecholtoo-quinone,butwithdifferentrates. Thecatalyticactivities
22
depend high2l3y on* Cthorreesploigndaennced: [email protected]; iToel.n: +,xxt-hxxex-xnxxa-xtxxux re of solvent and the type of inorganic anion
(FiguresS1–S245 ). Academic Editor: name
25 Received: date; Accepted: date; Published: date
26 TaAbblsetr4ac.t:O Ax siindgalet ipoanragrraatpeh ooff acbaoutet 2c0h0o wloordxsi dmaaxtiimounmi.n Fomr reetsheaarcnho alrt(icµlems, oabls·tLra−ct1s ·smhoiunld− 1).
27 give a pertinent overview of the work. We strongly encourage authors to use the following style of
28 structured abstracts, but without headings: 1) Background: Place the question addressed in a broad
Liga29n d/Mcoenttaexllt iacnd highlight the purpose of the study; 2) Methods: Describe briefly the main methods or
Cu(NO ) CuCl Cu(CH COO) CuSO
30 Salttreatments applied; 3) Results:3 Su2mmarize the article's m2ain findings; and 4) C3onclusion2s: Indicate 4
31 the main conclusions or interpretations. The abstract should be an objective representation of the
32 L1article, it must not cont1ai0n. r5e7sults which are not 1pr0e.s2en8ted and substantiate9d. 8in0 the main text and 9.27
33 L2should not exaggerate t1he5 m.5a2in conclusions. 0.05 22.92 16.06
34 L3Keywords: keyword 1; 3k7ey.8w9ord 2; keyword 3 (Li9st. 1th9ree to ten pertinent k2e4yw.5o8rds specific to the 21.01
35 L4article; yet reasonably c1om7.m7o1n within the subject1 d1is.4ci3pline.) 15.02 6.57
36 L 5 3.10 8.07 19.62 10.61
L6 27.77 40.27 60.50 72.92
Animals 2017, 7, x; doi: FOR PEER REVIEW www.mdpi.com/journal/animals
Itappearsplainlythatcatalyticactivityvariesfromonecomplextoanother. Allofthecopper
complexesformedbythehydrazonederivativescatalyzetheoxidationreactionofcatecholtoo-quinone.
ThebestistheL6ligandwithhighestoxidationrateforallcoppersaltswithmaximumreactionrate
equalto72.92µmol·L–1·min–1forCuSO followedbyL3withmaximumrateof37.89µmol·L–1·min–1
4
for Cu(NO ) . Lowest recorded oxidation rates, whose values are less than 10 µmol·L–1·min–1,
3 2
correspond to complex L2-CuCl , followed by L5-(CuNO ) and L4-CuSO , and then L5-CuCl .
2 3 2 4 2
The L1 ligand seems to have nearly the same activity for all copper salts determined at about
10µmol·L–1·min–1.
Basically,foranyligandusedexceptL6,Cu(CH COO) giveshighestactivitiesandCuCl gives
3 2 2
lowestactivities(Figure6).ConcerningCH COO−andligandsL3,L4andL6,thevaluesofabsorbance
3
increaseatthebeginningofthereactionandthenremainconstantafteracertaintime. Thishasbeen
Int. J. Mol. Sci. 2017, 18, 2215 8 of 15
reaction rate equal to 72.92 μmol·L–1·min–1 for CuSO4 followed by L3 with maximum rate of 37.89
μmol·L–1·min–1 for Cu(NO3)2. Lowest recorded oxidation rates, whose values are less than 10
μmol·L–1·min–1, correspond to complex L2-CuCl2, followed by L5-(CuNO3)2 and L4-CuSO4, and then
L5-CuCl2. The L1 ligand seems to have nearly the same activity for all copper salts determined at
Int.J.Maobl.oSucit. 1200 1μ7,m18o,l·2L2–115·min–1. 8of14
Basically, for any ligand used except L6, Cu(CH3COO)2 gives highest activities and CuCl2 gives
lowest activities (Figure 6). Concerning CH3COO− and ligands L3, L4 and L6, the values of absorbance
explaininecdreiansep aret vthioeu bsegstinundiinegs obfy thaep rreeaccitpioitna tainodn tohfenth reemcoaminp cloenxstthanatt acfatnert aa kceerptalianc etimdue.r Tinhgist hhaiss dbeeecnre ase
inabsoexrbpalaninceed. in previous studies by a precipitation of the complex that can take place during this
decrease in absorbance.

FigurFeig6u.rCe a6t.e Cchatoelchooxli doxaitdioantioinn itnh tehpe rperseesnenceceo offc cooppppeerr ccoommpplelexxese sfofromrmede wdiwthi tLh6.L 6.
2.4.1. Effect of Ligand Concentration on the Catecholase Activity
2.4.1. EffectofLigandConcentrationontheCatecholaseActivity
The effect of ligand concentration on catecholase activity is studied by varying the ratio of
The effect of ligand concentration on catecholase activity is studied by varying the ratio of
equivalent ligand L6: metallic salt Cu(CH3COO)2. Three tests were carried out in ratios: 1:1, 2:1 and
equivalentligandL6: metallicsaltCu(CH COO) . Threetestswerecarriedoutinratios: 1:1,2:1and
1:2. The results obtained show that the3 test wi2th the ratio L6: Cu(CH3COO)2 = 1:2 leads to higher
1:2. Thoexidreastiuolnts roatbet avianluede. sThhoewreftohrea,t tthhee Lt6e sctowmiptlhext hteharta ctioontLa6in:sC tuw(oC cHu3pCriOc Oion)2 C=u1(I:I2) lheaasd as tboethteirg her
oxidatcioatnalryattiec avcatliuviet.yT. Hheorweefovreer,, rtahteioLs6 2:c1o amndp l1e:x1 ltehaadt tcoo lnotwaienrs otxwidoatciuonp rriacteio vnalCueu,( aIIn)dh raastiao b2e:1t teexrhcibaittasl ytic
activitym.aHxiomwuemv eart, 3r5a tmioisn,2 a:1nda nadfte1r: 1slligeahdtlyt odelocrweaesreos,x pidroabtiaobnlyr adtueev taol cuoem,apnledx rparteiocip2i:t1ateixohni (bFiitgsumrea 7x)i.m um
Int. J. Mol. Sci. 2017, 18, 2215 9 of 15
at35min,andafterslightlydecreases,probablyduetocomplexprecipitation(Figure7).

FigurFeig7u.re 7C. aCtaetcehchooll ooxxiiddaattiioonn ini nmemtheatnhoal,n ionl p,riensenpcree soef nfocremoedf Lfo6 rcmopepderL c6omcpolpepxeesr wciothm dpiflfeexreenst with
concentrations.
differentconcentrations.
2.4.2. Solvent Effect
The effect of three solvents (MeOH, CH3CN and DMF) on the oxidation reaction with the ligand
L6 and the Cu(CH3COO)2 salt is studied in similar thermodynamic conditions. The results of that
study are presented by the rate values obtained for the oxidation reaction (Figure 8). Using a polar
protic solvent such as methanol promotes oxidation reaction much better than the other two aprotic
solvents, i.e. acetonitrile and DMF. Our results seem to be in perfect agreement with previous studies
and show that solvatation of copper by the aprotic and polar solvents such as DMF and CH3CN
decreases the catalytic activity of the metal cation in the oxidation reaction of catechol to o-quinone.

Figure 8. Catechol oxidation in different solvents and in the presence of formed L6 copper complexes
(1 Equivalent of L6 for 1 Equivalent of Cu(CH3COOH)2).
Int. J. Mol. Sci. 2017, 18, 2215 9 of 15
Int.J.Mol.Sci.2017,18,2215 9of14
Figure 7. Catechol oxidation in methanol, in presence of formed L6 copper complexes with different
concentrations.
2.4.2. SolventEffect
2.T4.h2.e Seoflfveecntto Efftfhecrte esolvents(MeOH,CH CNandDMF)ontheoxidationreactionwiththeligand
3
L6andtheCu(CH COO) saltisstudiedinsimilarthermodynamicconditions. Theresultsofthat
The effect of 3three so2lvents (MeOH, CH3CN and DMF) on the oxidation reaction with the ligand
studLy6 aanred pthree sCeun(tCedH3bCyOtOh)e2 rsaaltte ivs asltuudesieodb itna isnimedilafro trhtehrmeoodxiydnaatmioicn croenadcittiioonns(. FTigheu rrees8u)l.tsU osfi nthgata polar
prostitcudsoy lavreen ptrseusecnhteads bmye tthhea rnaotel pvarolumeso otebstaoinxeidd aftoiro tnhere oaxcitdioatniomn urecahctbioent t(eFrigthuraen 8t)h. eUsoitnhge ar tpwoloara protic
solvpernottsic, is.oel.v,eancte stounchit raisl emaenthdanDoMl pFr.oOmuorterse osuxildtsatsieoenm retaoctbioeni nmpucehrf beectttearg trheaenm tehne towthietrh tpwroe vapioruotsics tudies
andsoslhvoenwts,t hi.ea.t ascoetlovnaittaritlieo anndo fDcMopF.p Oeurrb ryestuhltes saepermo ttioc baen idn ppeorfleacrt saoglrveeemntesnts wucithh apsreDviMouFs satnuddieCs H CN
3
decarenads eshsotwhe tchaatta sloyltvicataactitoivni toyf ocofpthpeerm beyt athlec aatpiorontiicn atnhde pooxliadra stoiolvnernetsa cstuiochn aosf DcaMteFc haonldt oCHo-3qCuNin one.
decreases the catalytic activity of the metal cation in the oxidation reaction of catechol to o-quinone.

Figure 8. Catechol oxidation in different solvents and in the presence of formed L6 copper complexes
Figure8.CatecholoxidationindifferentsolventsandinthepresenceofformedL6coppercomplexes
(1 Equivalent of L6 for 1 Equivalent of Cu(CH3COOH)2).
(1EquivalentofL6for1EquivalentofCu(CH3COOH)2).
2.4.3. ComparisonwithAlternativeCatalysts
Table5showsthecatalyticactivitybyothercatalystsreportedintheliterature. Itisclearthat
thehydrazone–pyrazolederivatives,inparticularligandL6,describedinthisworkpresentfurther
improvementandshowbettervaluesandhigheractivityfortheeffectiveaerobicoxidationofthe
catecholintoo-quinone.

Table5.Comparisonofthecatalyticactivityofvariouscatalyststowardoxidationofthecatecholinto
o-quinone,establishedinthesameconditions,asgiveninpreviousliterature.
OxidationRate
Cu(II)-Ligands Cu(II)SaltUsed (µmol·L−1·min−1) Ref.
ligandL6 CuSO 72.920 -
4
ligandL6 Cu(CH COO) 60.500 -
3 2
ligandL6 CuCl 40.270 -
2
C,N-bipyrazole Cu(CH COO) 4.440 [29]
3 2
bipyrazolic
Cu(CH COO) 11.825 [30]
tripode-prop-2-ylacetate 3 2
bipyrazolic
CuCl 1.458 [31]
tripode-4-hydroxyphenyl 2
bipyrazolic
CuSO 28.990 [32]
tripode-3-hydroxypropyl 4
bipyrazolic
CuCl 4.378 [33]
tripode-3-hydroxypropyl 2
indole-3-chalcone Cu(CH COO) 31.780 [34]
3 2
[(3,5-dimethyl-pyrazol-1-ylmethyl)-amino]
CuSO 8.710 [35]
-propionitrile 4
Int.J.Mol.Sci.2017,18,2215 10of14
The superior catalytic activity observed for ligand L6 is probably due to the stability of
corresponding copper complex (catalyst) favored by the organic conjugate π bonds of the three
benzeneringcontainedintheligandandbytheintensecoordinationbondsoftheSchiffbase. Toour
knowledge,thecatalyticactivityobservedforligandL6(72.920µmol·L–1·min–1)isthemostimportant
amongthecatalystsdescribedintheliterature.
3. Experimental
3.1. GeneralMethods
ThechemicalreagentsusedinsynthesiswerepurchasedfromMerck,Darmstadt,Germanyor
Aldrich,St.Louis,MO,USA;wereofthehighestcommerciallyavailablepurity;andwereusedwithout
previouspurification. MeltingpointsweremeasuredusingaBüchiB-545digitalcapillarymelting
pointapparatus(BUCHILabortechnikAG,Flawil, CH,Switzerland)andusedwithoutcorrection.
ReactionswerecheckedwithTLCsilicagel60F254(MACHEREY-NAGEL,Neumann-Neander,Drun,
Germany). SpectraIRwererecordedonaVERTEX70FT-IRspectrometer(Perkin-Elmer, Billerica,
MA, USA) and frequencies are reported in cm−1. The spectra of 1H NMR and 13C NMR were
recordedinsolutioninDMSO-d onaBrukerAvance300NMRspectrometer(Bruker,MA,USA).The
6
chemicalshiftsareexpressedinpartspermillion(ppm)byusingtetramethylsilane(TMS)asinternal
referenceandcouplingconstants(J)aregiveninHz. MassspectrawerecollectedusingaAPI3200
LC/MS/MSsystem(ABMDSSciex,Ontario,Canada)equippedwithanESIsource. DRXdatawere
collected on a Bruker APEX-II D8 Venture area diffractometer (Bruker, MA, USA) equipped with
graphitemonochromaticMoKαradiation,λ=0.71073Åat293(2)and296(2)K,respectively. The
electronicspectraoftheligandanditsmetalcomplexeseremeasuredonaLambda35ESUV/VIS
spectrophotometer(PerkinElmer,Waltham,MA,USA)intherangeof200–900nm.
3.2. Synthesis
3.2.1. Synthesisof3-phenyl-1H-pyrazole-4-carbohydrazides(2)
Toastirredsolutionof1mmolofthe3-phenyl-1H-pyrazole-4-carboxylate(1)inethanol(10mL),
2mLof80%hydrazinemonohydratewasadded.Thereactionmixturewasmaintainedunderrefluxfor
5h,untilTLCindicatedtheendofreaction. Afterwards,thereactionmixturewaspouredontoiceand
thesolidformedwascollectedbyfiltration,washedwithcoldwaterandrecrystallizedfromethanol.
Yield: 66%;m.p: 207–209◦C;IR(ν(cm−1)): 3296–3203(NH,NH ),1629(C=O);1HNMR:(300MHz,
2
DMSO-d ,δ(ppm)): 4.45(s,2H,NH ),7.05(1H,s,CH-pyrazole),7.17–7.60(5H,m,Ar-H),9.38(s,1H,
6 2
NHCO),13.61(1H,s,NH-pyrazole);ESI-MS:m/z=203.3[M+H]+,225.1[M+Na]+. TheFT-IR,1HNMR
andmassspectraofcompound2areshowninFiguresS6–S8.
3.2.2. GeneralProcedurefortheSynthesisofLigands(L1–L6)
Toasolutionof5-phenyl-1H-pyrazole-4-carbohydrazide(2)(1mmol)in10mLofethanol,an
equimolaramountoftheappropriatebenzaldehydederivativewasaddedinthepresenceofacetic
acid. Themixturewasmaintainedunderrefluxfor2–5h, untilTLCindicatedtheendofreaction.
Then,thereactionmixturewascooledto25◦C,andtheprecipitateformedwasfilteredoutwashed
withethanolandrecrystallizedfromethanol. TheFT-IR,(1H,13C)NMRandmassspectraofLigands
(L1–L6)areshowninFiguresS9–S30.
3.2.3. N’-(4-hydroxy-3-methoxybenzylidene)-5-phenyl-1H-pyrazole-3-carbohydrazide(L1)
Yield: 85%; m.p: 229–231 ◦C; IR (ν(cm−1)): 3276 (NH), 1681 (C=O), 1568 (C=N); 1H NMR:
(300MHz, DMSO-d , δ(ppm)): 3.81 (3H, s, -OCH ), 6.83 (d, J = 8.1 Hz, 1H, H-Ar), 7.18 (1H, s,
6 3
CH-pyrazole),7.26–7.49(5H,m,Ar-H),7.54(s,1H,H-Ar),7.82(d,J=8.1Hz,1H,H-Ar),8.39(1H,s,
N=CH),11.51(s, 1H,OH),11.72(s, 1H,NHCO),13.72(1H,s, NH-pyrazole); 13CNMR:(300MHz,