A quantum-mechanical study on the complexation

of b-cyclodextrin with quercetin

Chunli Yan a, Xiaohui Li a, Zhilong Xiu a,*, Ce Hao b

a Department of Bioscience and Biotechnology, School of Environmental and Biological Science and Technology,

Dalian University of Technology, Dalian 116024, Liaoning, People’s Republic of Chinab Department of Chemistry, School of Chemical Science and Engineering, Dalian University of Technology,

Dalian 116024, Liaoning, People’s Republic of China

Received 26 September 2005; received in revised form 15 February 2006; accepted 19 February 2006

Available online 5 April 2006

Abstract

The inclusion process involving b-cyclodextrin (b-CD) and quercetin has been investigated by using the PM3 quantum-mechanical semi-

empirical method. In the b-CD quercetin inclusion complex, a large portion of the flavonoid skeleton is included in the b-CD cavity and the bond

connected ring B with ring C is inclined to the molecular axis of b-CD. The orientation in which the B ring of the guest molecule located near the

secondary hydroxyls of the b-CD cavity is preferred in energy. One intermolecular hydrogen bond is formed. The molecular modeling results are

in agreement with the NMR observations and molecular dynamics (MD) simulations. The statistical thermodynamic calculations at 1 atm and

298.15 K by PM3 demonstrate that 1:1 quercetin/b-CD complex is favored by a negative enthalpy change.

q 2006 Elsevier B.V. All rights reserved.

Keywords: Cyclodextrin; Quercetin; PM3; Semi-empirical method; Inclusion complexation

1. Introduction

Naturally occurring cyclodextrins (CDs) are a-1,4-linkedcyclic oligomers of D-glucopyranose which possess the

remarkable property of forming inclusion complexes with a

variety of small molecules appropriate size through the

influences of noncovalent interactions. The resultant inclusion

complexes can induce modification of the physicochemical

properties of the ‘guest’ molecules, particularly in terms of

water solubility and solution stability [1–3]. Therefore, it is

important to clarify the structures of the inclusion complexes

from a viewpoint of enzyme-substrates within the hydrophobic

cavities of CDs [4]. The most common CDs are a-, b- andg-CDs which consist of 6–8 glucopyranose units, respectively.

Nevertheless, b-CD (Fig. 1(a)) is by far the most widely used

compound owing to the optimal size of its internal cavity (8 A),

the most accessibility, and the lowest price for the encapsula-

tion of molecules [5].

Quercetin (3, 3 0, 4 0, 5, 7-penthydroxy flavone, Fig. 1(b)) is

an important constituent of the flavonoid family and is found

0166-1280/$ - see front matter q 2006 Elsevier B.V. All rights reserved.

doi:10.1016/j.theochem.2006.02.008

* Corresponding author. Tel./fax: C86 411 84706369.

E-mail address: zhlxiu@dlut.edu.cn (Z. Xiu).

in many fruits and vegetables, as well as olive oil, red wine,

and tea [6–8]. It has been demonstrated to possess many

biological effects that are considered beneficial to health,

including antioxidation by scavenging free radicals, antic-

ancer, antiviral prevention of atherosclerosis, and chronic

inflammation activities [9,10]. In spite of this wide spectrum

of pharmacological properties, its use in pharmaceutical field

is limited by its low aqueous solubility [11]. In recent years,

CD complexation has been successfully used to improve

solubility, chemical stability and bioavailability of such

polyphenolic flavonoids [12–15] and other active ingredient

of Chinese herbal medicine, such as alkaloids [16],

terpennoids [17] and so on. The major driving forces for

the complex formation have been proposed to include the

release of entropy-rich water molecules from the cavity, van

der Waals interactions, hydrophobic interactions, hydrogen

bonding and release of ring strain in the CD molecule.

However, the relative contributions and even the nature of

the different forces are not well-known [18]. Due to the

limitations of the experimental methods, molecular modeling

is frequently used to rationalize experimental findings

concerning molecular and chiral recognition by CDs.

Molecular modeling methods of CD complexes are powerful

tools for deriving information on the geometry and the

interaction energy of the inclusion compounds.

Journal of Molecular Structure: THEOCHEM 764 (2006) 95–100

www.elsevier.com/locate/theochem

Fig. 1. Molecular structures of b-cyclodextrin (a), quercetin (b) and PM3-optimized quercetin (c).

C. Yan et al. / Journal of Molecular Structure: THEOCHEM 764 (2006) 95–10096

Intensive theoretical works have been performed over the

past few years on CDs. Owing to their conformational

flexibility and large size, most computations on CD-effective

constituents of Chinese medicine are carried out with

molecular mechanics (MM) [19] or molecular dynamics

(MD) methods [20,21]. These methods are unable to fully

account for the inherent quantum-mechanical effects, as the

molecular mechanics and dynamics methods are based on the

classical physics of balls and springs. Nevertheless, in recent

years, with increased availability of computer resources, the

number of studies reporting the use of quantum mechanics

(QM) for investigating the 3D structure and properties of such

complexes has increased dramatically. QM method can

provide information about the electronic structure of the

system and hence, better understanding of the inclusion

structure. There have been a series of QM studies on the

complexation of CDs with small organic compounds, and these

methods have been successfully applied to these CD systems.

Again, most guests considered were simple, disubstituted

benzenes [18,22]. But very few calculations were performed

for CD complexation with herbal constituents based on QM

methods [16], and the optimum positions of complexation were

determined by only trying several starting points rather than by

a global search.

The inclusion of quercetin with native and modified b-CDshas been studied by other authors and an inclusion complex

with a stoichiometry of 1:1, in solid-state and in aqueous

solution, was attained by different preparation techniques: free-

drying, co-evaporation and kneading method [13,20,23].

Furthermore, a spatial configuration of the b-CD with quercetin

was proposed on the basis of NMR and molecular dynamics

simulations [20]. However, to our best knowledge, quantum

mechanics studies on the inclusion complexation of b-CD with

quercetin, a rather flexible molecule, have been scarcely

reported by far. Furthermore, the studies of noncovalent

interactions involving aromatic substrates are pivotal to both

chemical and biological recognition, as reviewed and evaluated

by Meyer et al. [24].

As applied by Liu and Guo [22], Parametric Model 3 (PM3)

[25] has been chosen to study host–guest complexes between

b-CD and quercetin. Due to the molecular size, PM3 is a

powerful technique which can be currently applied and

performs better than Austin model 1 (AM1) in biochemical

systems due to its improved description of the interactions

between nonbonded atoms, e.g. hydrogen bond and steric

effects [26]. It has high computational efficiency and its

precision is comparable to that of ab initio with medium-sized

basis sets. Conformational flexibility of the quercetin molecule

includes the orientation of the linkage between ring B and C.

Previous NMR studies and MD simulation on the inclusion of

quercetin by b-CD yield ambiguous results for the configur-

ation of quercetin in the complexes and the driving forces

contributing to complexes formation have not been known

clearly yet [20]. The aim of the present work is to re-investigate

their difference in energy, conformations of quercetin in the

complex and the effect of hydroxyl attached to quercetin on the

complexation process with b-CD using the PM3 method.

2. Computational methodology

All theoretical calculations were performed using GAUSSIAN

03 software package [27]. For all the systems, a full geometry

optimization was done at the PM3 level, and a harmonic

frequency analysis was followed to characterize the located

stationary points as true minima (all eigenvalues of the Hessian

matrix are positive).

C. Yan et al. / Journal of Molecular Structure: THEOCHEM 764 (2006) 95–100 97

The initial structures of b-CD and quercetin were

constructed with the help of CS Chem3D Ultra (Version 6.0,

CambridgeSoft.com) from the crystal structure [28,29] and

were fully optimized with PM3. Calculations were carried out

in the gas phase, and solvation effects were not considered.

However, the obtained results are qualitatively useful. As

shown in Fig. 1(c), a conformation for quercetin was obtained

with a minimum energy, where the dihedral angle 6 0-1 0-2-3

(Fig. 1(b)) is equal to 64.58. Experimental evidence suggests

that the B ring of the substrate is preferentially inserted in the

cavity of b-CD [20]. So we constructed substrate/b-CDcomplexes by manually introducing the reactive guest in a

vertical position into the b-CD cavity through the secondary or

primary rim of the latter and perpendicular to the rim diameter.

The coordinate system used to define the process of

complexation is shown in Fig. 2. The method used by Liu

and Guo [22] was referred. The glycosidic oxygens of b-CDwere placed onto the XY plane and their center was defined as

the center of the coordination system. The 1 0-2 bond of

quercetin (Fig. 1(b)) was coincident with the Z-axis, and the

relative position between the host and the guest was measured

by the Z-coordinate of the labeled carbon atom of the guest

(Fig. 2). For the complexation process, the host b-CD was kept

in this position while the guest approached along the Z-axis

toward the wide or narrow edge of the b-CD torus. Fig. 2

depicts the two possible approaches, which will be denoted by

head down and head up approaches, respectively.

The guest was initially located at a Z-coordinate of 8 A and

was moved through the host cavity along the Z-axis to K3 A

at with a stepwise 1 A. For each step, the geometry of

OHO

HO

(a)

(b)

O H

O H OHO

O

Y

X

Z*

OHO

HO

OH

OHOHO

O

Y

X

Z*

Fig. 2. Coordinate systems used to define the process of complexation for: (a)

head up and (b) head down.

the complex was fully optimized by PM3 without imposing any

symmetrical restrictions. In order to find an even more stable

structure of the complex, each guest molecule was calculated

for all of the structures obtained by scanning q, circling around

Z-axis, at 208 intervals from 0 to 3608 and scanning

Z-coordinate at 1 A intervals from K2 to C2 A. Here, q is

the angles circling around the Z-axis of the system and the 1 0-2

bond of quercetin (Fig. 1(b)) is coincident with the Z-axis. The

angle, qZ0, between XZ and the B ring plane of quercetin is

defined as the reference point (Fig. 2). Stabilization energy

(DE) upon complexation between quercetin and the b-CD was

calculated for the minimum energy structure according to

Eq. (1) as a previous work [30]

DEZEcomplxKðEsubst CEbKCDÞ (1)

where Ecomplx, Esubst and Eb-CD represent HF energies (heats

of formation) of the complex, the free substrate and the free

b-CD, respectively. The magnitude of the energy change

would be a sign of the driving force towards complexation. The

more negative the stabilization energy is, the more thermo-

dynamically favorable is the inclusion complex.

3. Results and discussions

The potential energy surfaces of DE versus q and Z for all

the optimized structures obtained by scanning the q angle, from

0 to 3608 at 208 intervals, and the coordinate Z, fromK2 to 2 A

at 1 A intervals, were used to find the most favorable approach

of the quercetin to the b-CD. Two DE minima were found at

qZ40 and 608 for head up and head down, respectively. All

calculations reported thereinafter use the approach of quercetin

into b-CD where the Z coordinate ranged from 8 to K3 A at a

fixed q of 408 (head up) and 608 (head down).

The optimum position and angle for quercetin into b-CD at

head up and head down can be, respectively, determined

according to stability energy as shown in Figs. 3 and 4. Other

possible locations and angle of quercetin were examined using

PM3 method, but were shown to be energetically less favorable

and so they were not listed. As shown in Figs. 3 and 4, the

complexation process is energetically favorable. The stable

structure is reached at approximately ZZK1 A for both head

up at qZ408 and head down at qZ608, respectively.

The most stable structure can be obtained by comparing the

characteristic energies of the complexes between head up and

head down orientation at the optimum position and angle. As

listed in Table 1, the negative DE changes upon complexation

clearly demonstrate that b-CD can form stable complex with

quercetin. The changes in the binding energy are of similar

magnitude to those reported in most MM studies on CD

systems [31], which is reasonable for nonbonded supramole-

cular complexation. The (ELUMOKEHOMO) gap is an important

stability index [32], and chemicals with larger (ELUMOKEHOMO) values tend to have higher stability. Therefore, with

the increase of the (ELUMOKEHOMO) gap, the complexes

formed are more stable, which agrees with the calculated

results of the stability energies. It can also be seen that the head

–4 –2 0 2 4 6 8

Z (Å)

–60

–50

–40

–30

–20

–10

0

10

Stab

ility

Ene

rgy

(kJ/

mol

)

–60

–50

–40

–30

–20

–10

0

10(a)

(b)

–4 –2 0 2 4 6 8

Z (Å)

Stab

ility

Ene

rgy

(kJ/

mol

)

Fig. 3. Stability energies of the inclusion complexation of quercetin into b-CD

at different positions (Z) and orientations: (a) head up; (b) head down. The

position of the guest was determined by the Z-coordinate of the carbon atom in

the phenyl group from the center of the glycosidic oxygens.

–60

–50

–40

–30

–20

–10

0(a)

(b)

0 50 100 150 200

q250 300 350 400

Stab

ility

ene

rgy

(kJ/

mol

)

–60

–50

–40

–30

–20

–10

0

0 50 100 150 200 250 300 350 400

q

Stab

ility

ene

rgy

(kJ/

mol

)

Fig. 4. Stability energies of the inclusion complexation of quercetin into b-CD at

different scanning q (ZZK1 A) and orientations: head up (a) and head down (b).

Table 1

Interaction energies and thermodynamic properties for the inclusion

complexation of b-CD with quercetin

Species Quercetin b-CD Head up Head

down

Ea(kJ molK1) K946.39 K6096.50 K7096.57 7090.59

DEa(kJ molK1) K53.68 K47.70

Hf(kJ molK1) K270.36 K2792.00 K3111.18 K3108.82

DH8b(kJ molK1) K48.82 K46.46

Gf(kJ molK1) K447.12 K3298.87 K3718.60 K3720.54

DG8b(kJ molK1) 27.39 25.45

DS8c(J molK1 KK1) K255.61 K241.19

EHOMOd(ev) K8.98 K10.83 K9.06 K9.04

ELUMOe(ev) K0.89 1.47 K0.79 K1.00

EHOMOKELUMO gap (ev) K8.89 K12.30 K8.27 K8.04

a E is the HF energy, DE is the stabilization energy upon complex, DEZEcomplxK(EsubstCEb-CD).b DA8ZAcomplxK(AsubstCAb-CD); AZH, G.c DS8Z(DH8KDG8)/T.d Energy of the highest occupied molecular orbital.e Energy of the lowest unoccupied molecular orbital.

C. Yan et al. / Journal of Molecular Structure: THEOCHEM 764 (2006) 95–10098

up orientation was significantly more favorable than the head

down orientation by an energy difference of K5.98 kJ molK1.

The driving forces for the stable structures can also be

analyzed by using the PM3 method. Fig. 5 presents the

proposed favorable structures for the 1:1 inclusion complex of

both the head down and head up orientations at each energy

minimum. At the head up orientation, the aromatic B-ring of

quercetin entered fully into cavity of b-CD so that the

7-hydroxy group (OH) of quercetin was close to 6-OH of b-CD. The O/O PM3 calculated distance (of cyclodextrin) was

2.73 A. It is suggested that an intermolecular hydrogen bond

which contributes a stabilization energy of 16–25 kJ molK1

[33] to the complex is formed between quercetin and b-CD.The head down orientation does not seem to be stabilized by

such an interaction. Even though we work in the theoretical

treatment in a vacuum, it is known from the literature that

secondary OH groups form intramolecular hydrogen bonds and

the rate of exchange of protons for head down orientation is

much lower (stronger bonding) than that of head up orientation.

The hydrogen bond here is defined as an O–H/O interaction

in which the O/O distance is less than or equal to 3.2 A and

the angle at H is greater than 908 [34]. Possible hydrogen

bonding is shown in Fig. 5(a and b) as a dotted line. On

the other hand, in the head up structure, the quercetin B-ring

projects onto the 2-OH/3-OH face of b-CD, and the A-ring

projects from the 6-OH face. The four phenolic hydroxyl

groups of the A and B rings of quercetin are left out of the

hydrophobic cavity of b-CD because of their hydrophilic

(a) (b)

(c) (d)

Fig. 5. Structures of the b-CD-quercetin complex with minimum energies obtained by PM3 calculations at different orientations: (a) head up perpendicular to the

cavity axis; (b) head up along the cavity axis from the primary side; (c) head down perpendicular to the cavity axis; (d) head down along the cavity axis from the

secondary side.

C. Yan et al. / Journal of Molecular Structure: THEOCHEM 764 (2006) 95–100 99

characters. There is a good agreement between the experimen-

tal NMR and MD simulations about the overall orientation of

the quercetin ligand in the b-CD cavity: the B-ring, C-ring, and

part of the A-ring of quercetin display favorable interaction

with the hydrophobic cavity of the b-CD [20]. By MD

calculation (MM2 empirical force field), Ding et al. [19] also

confirmed that the phenyl group of rutin (quercetin-3-

rhamnosylglucoside) penetrates into the cavity from the

primary edge of b-CD for the rutin-b-CD complex.

The conformation of quercetin was significantly altered

during complexation (seen in Fig. 5). The dihedral angle (6 0-1 0-

2-3) was changed to 84.8 and 66.48 for the head up and head

down orientation, respectively. It is suggested that, in b-CD-quercetin complexes, the B ring of quercetin can rotate around

the molecular axis of b-CD because of inclusion interaction.

And so the bond connected C1 0 with C2 is inclined to the

molecular axis of b-CD, which results in one hydrogen bond

formation between 7-OH of quercetin and 6-OH of b-CD. Asimilar experiment was conducted by using NMR for

complexation of (C)-catechin into b-CD [35]. The results

revealed that upon inclusion complex formation with (C)-

catechin, the axis of the C1 0–C4 0 of the B ring of (C)-catechin

inclined to the molecular axis of b-CD in the cavity. This

indicates that the guest conformation is easily affected during

CD complexation. Therefore, the assumption in many MM

studies that inclusion complexation did not affect the structure

of the guest molecule should be taken cautiously [31].

To investigate the thermodynamics of the binding process,

the statistical thermodynamic calculation were carried out at

1 atm and 298.15 K in vacuo by PM3. The thermodynamic

quantities, the enthalpy change, the thermal Gibbs free energy

(DG) and entropy contribution (DS) are given in Table 1. FromTable 1, we can be seen that the complexation reactions of

quercetin with b-CD are exothermic judged from the negative

enthalpy changes. And the negative enthalpy changes suggest

that both the inclusion processes are enthalpically favorable in

nature. On the other hand, the enthalpy changes for the head up

orientation is more negative than the head down orientation,

which is surely attributed to the more tightly van der Waals

interactions. The enthalpy changes are of similar magnitude

to the experimental data DHZK26.70 kJ molK1, showing that

the structure and the reaction energy considered are well

described by the PM3 semi-empirical method. However, one

very important aspect of the inclusion process relative to the

entropy contribution should be pointed out. According to the

experimental study [20], the DS for the quercetin-b-CDinclusion process was found to be K32.20 J molK1 KK1. The

PM3 calculated DS was K255.61 J molK1 KK1, which is

C. Yan et al. / Journal of Molecular Structure: THEOCHEM 764 (2006) 95–100100

completely different from the expected value. As a conse-

quence, the standard DG is K17.20 (experimental) and

C27.39 kJ molK1 (PM3 for head up orientation). This

discrepancy can be attributed, in part, to the neglect of the

hydrophobic effect that involves a gain in entropy due to the

assimilation of the solvation water molecules by the medium

after the inclusion takes place [36]. That step of the inclusion

process should be more important in the complexation of

cyclodextrin with the hydrophobic substrate. Therefore, the

Gibbs free energy obtained from static methods (as applied

here) has no absolute meaning and should be considered only

in a relative way.

4. Conclusions

The stable structures and the inclusion process for

quercetin-b-CD inclusion complexes were studied by use of

quantum mechanics PM3 method. One intermolecular hydro-

gen bond is formed, a driving force responsible for its stability.

Its calculations on the complexation of b-CD with quercetin

support the simulations by molecular dynamics (MD) and the

experimental observation by NMR. The orientation in which

the B ring of the guest molecule located near the secondary

hydroxyls of the b-CD is preferred according to the

characteristic energies. The statistical thermodynamic calcu-

lations suggest that both of the complex processes are

enthalpically favorable in nature.

Acknowledgements

We are grateful to the supports from National Natural

Science Foundation of China (20176005) and a project of ‘973’

plans (2003CB716000). We also appreciate Prof. C. Hao from

Dalian University of Technology, who generously offered the

GAUSSIAN 03W software package for the whole computational

tasks.

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