Molecular complexes of l -phenylalanine with substituted 1,4-benzoquinones in aqueous medium: Spectral and theoretical investigations

Accepted Manuscript Molecular complexes of L-phenylalanine with substituted,4-benzoquinones in aqueous medium: Spectral and theoretical investigations K. Ganesh, E.H. El-Mossalamy, A. Satheshkumar, C. Balraj, K.P. Elango PII: DOI: Reference:

S1386-1425(13)00763-4 http://dx.doi.org/10.1016/j.saa.2013.07.026 SAA 10770

To appear in:

Spectrochimica Acta Part A: Molecular and Biomo‐ lecular Spectroscopy

Received Date: Revised Date: Accepted Date:

24 April 2013 29 June 2013 16 July 2013

Please cite this article as: K. Ganesh, E.H. El-Mossalamy, A. Satheshkumar, C. Balraj, K.P. Elango, Molecular complexes of L-phenylalanine with substituted,4-benzoquinones in aqueous medium: Spectral and theoretical investigations, Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy (2013), doi: http:// dx.doi.org/10.1016/j.saa.2013.07.026

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Molecular complexes of L-phenylalanine with substituted 1,4-benzoquinones in aqueous medium: Spectral and theoretical investigations K. Ganesh1, E. H. El-Mossalamy2, A. Satheshkumar1, C. Balraj1 and K. P. Elango1* 1 Department of Chemistry, Gandhigram Rural Institute (Deemed University), Gandhigram-624 302, India. 2

Chemistry Department, Faculty of Science, King Abdulaziz University, P.O.Box. 80203, 21589- Saudi Arabia.

Abstract Various spectral techniques such as UV-Vis, FT-IR, and fluorescence have been employed to investigate the charge transfer interaction of L-phenylalanine (LPA) with substituted 1,4-benzoquinones (MQ1-4). Kinetic and thermodynamic properties of the complexes were determined in aqueous medium at physiological condition (pH =7). The interaction of MQ1-4 with L-phenylalanine (LPA) was found to proceed through the formation of donor-acceptor complex, yielding a radical anion. The stoichiometry of the complexes was determined by Jobs continuous variation method and was found to be 1:1 in all the cases. Fluorescence quenching studies showed that the interaction between the donor and the acceptors is spontaneous. The results indicated that the progressive replacement of chlorine atom (-I effect) by methoxy group (+M effect) in the quinone decreased the electron acceptor property of the quinone. The order of the experimentally measured association constant of these complexes was well supported by DFT/B3LYP calculations.

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Keywords: Charge transfer; L-phenylalanine; substituent effect; Fluorescence; Theoretical studies. * Corresponding author. Tel.: +91 451 245 2371; Fax: +91 451 2454466 E-mail address: [email protected] (Dr. K.P. Elango) 1. Introduction Due to its interesting optical and electronic properties, the studies on the charge transfer (CT) or electron donor acceptor complexes (EDA) between the organic donor and acceptor molecules are enhanced nowadays [1-4]. CT phenomenon governs a number of pivotal processes in nature such as photosynthesis and vision [5]. CT has an extensive range of applications in the field of organic photovoltaics [6], sensors [7], organic field effect transistor [8], non linear optics [9], magnetic materials [9], organic solar cells [10] xerogel nano particles [11] and in the quantitative estimation of drugs [12] etc. In some cases CT complexes plays a driving force for variety of organic and organametallic reactions such as aromatic electrophilic substitution reaction [13]. For their wide applications, extensive studies on CT-complexes of variety of acceptors have been performed and almost entire work has been done only in non aqueous solvents [1-4]. Drug-receptor mechanism could be explained by means of CT phenomenon in addition to weak forces of non- covalent interactions [14]. As a primary step to determine whether CT phenomenon at any level involved, the ability of the donor drugs and related compounds to form charge transfer complexes with acceptors in physiological condition in aqueous medium should be studied. In spite a large number of studies, the real understanding of drug-receptor interactions is still a far cry as the matter is complex as no consideration of water molecule in drug-receptor interaction is made till now. Water

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constitutes 80% of our body weight and is the predominant biological solvent. Unless the role of water in drug-receptor interactions is properly understood, otherwise the understanding the possibility of drug-receptor interaction is remote. Quinones such as ubiquinone, menaquinone and plastoquinone are non-covalently bound enzyme cofactors that function as oxidation/reduction intermediates between assemblies of the membrane bound proteins of the energy conversion systems of photosynthetic and respiratory systems [15, 16]. The protein/amino acid environment plays a major role in modulating the thermodynamic property, kinetics of the electron and proton transfer and redox chemistry of the quinones [17, 18]. Naturally occurring ubiquinones possess variable number of methoxy substituents in their units and the orientation of methoxy group substantially influences its electron affinity, redox cofactor and vibrational spectroscopy [19]. Hence as a model study CT interaction between 1,4benzoquinones possessing varying the number of chloro/methoxy substituents with a chosen amino acid donor has been attempted. The objective, therefore, of the present article is to study the spectral, thermodynamic and kinetic aspects of the interaction of 1,4-benzoquinones possessing varying number of methoxy/chloro substituents as electron acceptors (MQ1-4) with Lphenylalanine (LPA) amino acid with an aim to investigate the mechanism of these interactions and to characterize the structure of the complexes formed in these interactions. L-phenylalanine is chemically known as (S)-2-amino-3-phenylpropanoic acid which is an essential amino acid used in the treatment of endogenic depression symptoms [20], attention deficit disorder [21] and as an enhancer of opium analgesics used in the conditions of chronic pain [22]. In the presence of phenylalanine hydroxylase

3

enzyme, LPA is biologically converted into another amino acid, tyrosine. LPA has also been reported to reduce the calories with its ability to burn fat and suppress appetite [23]. Carbon labeled LPA has been used to assess basal muscle protein synthesis rates in vivo in humans by single biopsy approach [24]. Thus the pharmaceutical and biological importance of LPA emphasizes the need to study the actual site of attack during the formation of charge transfer interaction. Such a study would undeniably shed some light on the mechanism of the drug action in real pharmacokinetic study. Though these 1,4benzoquinones are known to organic chemists as intermediates, it is the first systematic attempt to utilize them as acceptors with an amino acid in aqueous medium. Such a structural variation of the quinones would certainly helps to tune the redox chemistry of them and hence its biological activity. In the frame of Density Functional Theory (DFT) we have also performed the complete optimization of the geometry for these quinones, LPA and their CT complexes.

2. Experimental 2.1 Material and methodology The electron acceptors viz. varying number of methoxy/chloro substituted 1,4benzoquinones were synthesized and purified by the reported method [25, 26]. The electron donor amino acid L-phenylalanine was obtained from Merck, India and was used as received. The purity of the amino acid was checked by its melting point (observed 274 o

C; literature 273 °C) and FT-IR spectra. Disodium-monohydrogenphosphate and

monosodium-dihydrogen phosphate buffer (AR grade, Merck, India) was used to adjust

4

for the physiological pH 7. The structures of the amino acid and the acceptors are shown below.

O

H2N

OH

Chemical structure of phenylalanine

O R4

6

1

2

R1

R3

5

4

3

R2

O

Acceptor

R1

R2

R3

R4

MQ1

OCH3

Cl

Cl

Cl

MQ2

OCH3

Cl

OCH3

Cl

MQ3

OCH3

OCH3

OCH3

Cl

MQ4

OCH3

OCH3

OCH3

OCH3

5

Solutions for the spectroscopic measurements were prepared by dissolving accurately weighed amounts of donor (D) and acceptor (A) in the appropriate volume of double distilled water immediately before running the spectra. The electronic absorption spectra were recorded on a JASCO (V 630, Japan) UV-Vis double beam spectrophotometer using 1 cm matched quartz cells. The temperature of the cell holder was controlled with a water flow (±0.2oC). The steady state fluorescence spectra were obtained on a JASCO (FP 6200, Japan) spectrofluorimeter. The emission slit width (5 nm) and the scan rates (250 nm) were kept constant for all of the experiments. FT-IR spectra were recorded in a JASCO (FT-IR 460 Plus, Japan) spectrometer in KBr pellet.

2.2 Kinetic procedure The kinetics of the interaction of LPA with the acceptors (MQ1-4) was followed at three different temperatures in water under pseudo-first-order conditions, keeping [D] >> [A]. The increase in absorbance of the new peak at 541 nm for (LPA-MQ1), 536 nm for (LPA-MQ2), 533 nm for (LPA-MQ3) and 527 nm (LPA-MQ4) with elapse of time were recorded. The pseudo-first-order rate constants (k1) were calculated from the gradients of log (A∞ – At) against time plots, where A∞ and At represent the absorbance at infinity and time t, respectively. The second order rate constants were calculated by dividing k1 by [D] [27]. 3. Results and discussion 3.1 Stoichiometry of the interaction The stoichiometry of the CT complex formed, in all the cases, was determined by applying Job’s continuous variation method using absorption spectral data [28]. In all the 6

cases (LPA-MQ1-4) the symmetrical curve with a maximum at 0.5 mole fraction indicated the formation of a 1:1 (D:A) CT complex (Fig. 1). The photometric titration measurements were also performed for the determination of the stoichiometry in these interactions. The results of the photometric titration studies (Fig. 2) confirmed the observed stoichiometry of the interaction [29]. The stoichiometry of the LPA-MQ1-4 systems is further confirmed by Job’s continuous variation method using emission studies also [30] [(Fig. 1Sa) Supplemental information]. << Figure 1 >> << Figure 2 >> The observed stoichiometries of the CT complexes were also supported by the elemental analysis (CHN) of the complexes. The results obtained are: Found (cal.) %, LPA-MQ1 C 46.74 (47.26), H 3.02 (3.47), N 3.12 (3.44); LPA-MQ2 C 50.04 (50.76), H 3.98 (4.26), N 3.11 (3.48); LPA-MQ3 C 53.97 (54.35), H 4.56 (5.07), N 3.14 (3.52); LPA-MQ4 C 57.23 (58.01), H 5.35 (5.89), N 3.04 (3.56). 3.2 FT-IR spectral studies In all the cases, the pink colored CT complexes were obtained by allowing the reactants (5 mmol of LPA and 5 mmol of corresponding acceptor in water) to react for 24 h under stoichiometric conditions. The FT-IR spectra of the pure LPA, acceptors and their CT complexes were recorded and the peak assignments for important peaks are given in Tables 1-4. The results indicated that the shifts in positions of some of the peaks could be attributed to the symmetry and electronic structure modifications in both donor and acceptor units in the formed CT complex relative to the free molecules.

7

In the FT-IR spectrum of free LPA molecule, the broad and overlapped bands observed in the region of 3300-2300 cm-1 is the characteristic common feature when the spectrum was recorded in KBr pellet. The main band at 3066 cm-1 is due to the aggregation or association of molecules in the crystal state. The peaks at 1625 and 1492 cm-1 are due to asymmetric and symmetric bending vibrations of the –NH3+ group of the LPA. The strong bands in the spectrum at 1562 and 1409 cm-1 are assigned to the asymmetric and symmetric stretching vibrations of the carboxyl group ( –COO-). These observations suggest that phenylalanine is in the zwitter ionic form. The two intense bands at 744 and 698 cm-1 is the pattern of the mono substituted benzene ring [31]. As a representative case, the FT-IR spectra of LPA, MQ1, LPA-MQ1 complex are show in Figure 3. In this spectrum, a strong peak at 1737 cm-1 corresponds to the presence of free carboxyl group (–COOH). This indicates that LPA is not in Zwitter ionic form during complexation, as it is carried out at pH 7. The isoelectric point of LPA is pH = 5.49. The asymmetric and symmetric bending vibration of –NH2 group occurred at 1602 and 1482 cm-1, respectively. The ν(C=O), ν(O-CH3) and ν(C-Cl) stretching vibrations in the free MQ1 species appeared at 1684, 1270 and 909 cm-1, respectively. In the complex these stretching vibrations occurred at 1679, 1229 and 823 cm-1, respectively. Such a bathochromic shift could be indicative of a higher charge density on the carbonyl and chloro groups of the MQ1 molecule [32]. Two intense peaks corresponds to mono substituted benzene ring found at 744 and 698 cm-1 in the free LPA exhibited no significant shifts in the CT complexes indicating non participation of benzene ring in CT complex formation with these acceptors. These observations

8

suggested that the NH2 moiety of LPA participated in the complex formation with the quinones through charge transfer transition.

<< Figure 3 >> 3.3 Electronic spectral studies The electronic spectrum of MQ1 in the presence of large excess of the donor i.e. [D]/[A] > 100 were recorded as a function of time in water (Fig. 4). Immediately after mixing aqueous solutions of colorless LPA and pale yellow colored MQ1, a pink colored solution resulted whose electronic spectrum showed absorption band in 430-540 nm range. This is the characteristic absorption bands of quinone radical ion [33, 34]. It is observed that with elapse of time the intensity of band at 291 nm due to pure LPA has decreased (Fig.4) with a concurrent increase in intensity of the new bands at 360 and 540 nm. A clear isosbestic point is observed at 302 nm. These observations indicated that the initial reactants were converted into CT complex. For comparison the electronic spectrum of the complex is also shown in figure 4. Similar electronic spectral behavior observed for other systems also. [(Fig. 3Sa-c) Supplemental information] << Figure 4 >>

The kinetics of interaction of LPA with MQ1-4 has been followed by monitoring the increase in absorbance of new peak in water as a function of time under pseudo-firstorder conditions i.e. [D] >> [A]. The pseudo-first-order rate constant (k1) values for the formation of the complex as a function of [D] and [A] are collected in Table 5. It is evident from the results that in all the cases the rate is independent of initial concentration

9

of A indicating first order dependence on [A]. A plot of log k1 versus log [D] is linear with a slope of unity (r > 0.995; slope range 0.95 – 1.09) [(Fig. 4S) supplemental information] indicating unit order dependence on [D]. This was further supported by the constancy in k2 values [34]. The pseudo first-order rate constants, for all the systems, were measured at three different temperatures and the thermodynamic parameters computed are collected in Table. 6. A large negative value of entropy of activation indicated the involvement of a polar transition state. This may be due to the fact that there is some charge separation in the transformation of reactants to complex. Also the negative entropy of activation indicated a greater degree of ordering in the transition state than in the initial state, due to an increase in solvation during the activation process. Based on the foregoing results and discussions the following plausible mechanism (scheme 1) for the interaction LPA with MQ1-4 has been proposed.

10

O

O

NH2

R4

R1

R3

R2

R4

+

NH2

O O

OH

O

R1

R3

R2 O

OH

MQ1-4

CT- Complex

O R4 NH2 O

R1

R3

OH

R2 O

radical ion pair

Scheme 1. Mechanism of interaction of LPA with MQ1

In all the LPA-MQ1-4 systems, an attempt was made to characterize the CT complexes formed in these reactions. For that the absorbance of the new bands were measured using constant acceptor concentration in water and varying concentrations of the donor but always [D]>>[A]. A representative spectra is shown in [(Figure 5S) supplemental information]. The nature of the spectra indicated that the interaction 11

between the donor and the acceptor is of CT type. The formation constants (K) and molar extinction coefficients (ε) of the CT complexes were determined spectrophotometrically using the Scott equation [35]. [D][A]/d = [D]/ε + (1/Kε)

(1)

where [D] and [A] are the initial molar concentrations of the donor and the acceptor, respectively and d the absorbance. The values of K and ε are determined from the gradient and intercept of the linear plots of [D][A]/d against [D]. Representative Scott plots are shown in [(Figure 6S) Supplemental information] and the values of K and ε thus determined are given in Table 7. The observed high values of K suggested that the formed CT complexes are of strong type [34] and the linearity of the Scott plots further supported this result.

3.4 Fluorescence studies The nature and magnitude of the interaction of drugs with receptors play an important role in the pharmacokinetics of the drugs. CT interaction is one of the non covalent binding forces in the drug–receptor mechanism. In the present study, an attempt was made to study the CT interaction of LPA with MQ1-4 by means of fluorescence study. Fluorescence spectra were recorded at room temperature in water in the range of 300-700 nm using an excitation at 255 nm for LPA. It was observed that the fluorescence of LPA was quenched by MQ1-4 at 287 nm as a result of formation of CT complex. The experimental results indicated that the quenching efficiency increased with increasing concentration of the electron acceptor (Figures 5-8) and with increasing time. The

12

fraction of acceptor bound to the donor was determined by using the following equation 2. θ = (Fo-F)/ Fo

(2)

Where F and Fo denote the fluorescence intensities of donor in the presence of acceptor and in the absence of acceptor, respectively. From the resulting values of θ, the association constant Kf, for LPA-MQ1-4 systems, was computed using the method described by Ward [36]. It has been shown that for equivalent and independent binding sites (Eq. 3): 1/(1- θ) Kf = [AT]/ θ-n [DT]

(3)

Where n is the number of binding sites, [AT] is the total acceptor concentration and [DT] is the total donor concentration. The plot 1/(1- θ) versus [AT]/θ is linear (r > 0.97, for LPA-MQ1-4 systems) indicating that under the experimental conditions all the binding sites are equivalent and independent. The values of Kf obtained, from the plots, for LPAMQ1, LPA–MQ2, LPA–MQ3 and LPA–MQ4 systems are found to be 1.7 x 105, 1.5 x 105, 2.2 x 104, and 5.6 x 103 mol L-1, respectively. The standard Gibbs energy change ΔGo was calculated from Kf values using the relation ΔGo = -2.303 RT log Kf. The ΔGo values for LPA-MQ1-4 systems were found to be -28.8, -28.5, -24.0 and -21.0 kJ mol-1 respectively, indicating that the interaction between the drug and the acceptors is spontaneous in nature.

<< Figure 5-8 >>

13

Fluorescence quenching can occur by different mechanisms viz. static or dynamic or both. Stern–Volmer equation (Eq. 4) is useful in understanding the mechanism of fluorescence quenching. Fo/F = 1 + KSV [Q]

(4)

Where Fo is the initial fluorescence intensity measured in the absence of quencher and F is that in the presence of quencher. The Stern–Volmer constant KSV is obtained by plotting Fo/F against [Q]. In all the cases, curvature Stern-Volmer relationship was observed (Fig. 9) which indicated that the quenching of fluorescence of LPA by the acceptors can occur via both static and dynamic quenching mechanism [37, 38]. << Figure 9 >> The relationship between the fluorescence quenching intensity and the concentration of quenchers can be described by the following equation. log (Fo – F)/F = log KA + n log [Q]

(5)

Where KA is the binding constant and n is the number of binding sites per donor molecule [39]. In the present study, in all the cases, a plot of log (Fo–F)/F versus log [Q] is linear (Fig. 10, r > 0.98) and the binding constant values computed are collected in Table 8. The results indicated that, the binding constant value decreased from MQ1 to MQ4, i.e. the magnitude of binding constant is in the order of LPA-MQ1 > LPA-MQ2 > LPA-MQ3 > LPA-MQ4. These observations are in corroboration with the results of absorption spectral studies as enumerated earlier in this paper. That is the formation constant computed using absorption spectral data follows the same order (Table 6). Also, the rate constant for the interaction, in these cases, decreased with an increase in the number of methoxy group in

14

the acceptor (Tables 6 and 8). The value of n, for the systems, is nearly constant in water indicating the presence of equivalent binding sites. << Figure 10 >> The results of the redox potentials of these acceptors reported by us [26] indicated that the E1/2 values become more negative from MQ1 to MQ4. That is the electron accepting property (reduction) of MQ4 is relatively low when compared to MQ1 or in other words MQ4 is comparatively a weaker acceptor. This may be due to the fact that progressive replacement of electron withdrawing chlorine atom (-I effect) by electron releasing methoxy group (+M effect) rendered the quinone increasingly electron rich and consequently make it as a weak acceptor. This observation corroborates well with the results obtained in the spectral and kinetic studies enumerated above.

3.5. Theoretical Calculations To understand the foregoing experimental observations on the CT complex formed between LPA and the acceptors, we have performed the optimization of LPA and MQ1-4 using Density Functional Theory with the Backle3LYP hybrid functional, by using a basis set of 6-31G. Computations have been performed using the Gaussian 03 Revision D.01 program package [40]. The optimized geometry of the donor along with HOMO and the acceptors along with LUMO are depicted in Figure 11. In the case of LPA, the HOMO is concentrated on the –NH2 group and in the case of the acceptors the LUMO resides on the quinone ring []. << Figure 11 >> << Figure 12 >>

15

<< Figure 13 >>

The energies of the frontier orbitals of the donor and the acceptors along with the energy corresponds to the CT transition, ∆E (=HOMOLPA – LUMOacceptor) [41, 42], for all the systems are shown in Figure 12. It is evident from the figure that the ∆E depends on the nature of the substituent present in the quinone. Also, a good linear correlation obtained between the theoretical energy gap values and the experimentally determined stability constant values (Fig. 13) indicated that the strength of the acceptor decrease in the order: O

O Cl

Cl

O

> Cl

Cl O

MQ1

O

O O

Cl

O

Cl

O

O

>

O

> O

O

O O

MQ2

MQ3

O

O O

MQ4

As a representative case, the Mulliken charges of various atoms of MQ1, LPA and LPA-MQ1 CT complex were calculated using DFT [B3LYP- 631G++(d,p)] basis set in the gas phase [43] [(Figure 7S Supplemental information)]. The electronic charges thus computed, for selected atoms are given in Table 9. It is in teresting to compare the charge distribution in MQ1 with that reported chloranil [43]. In the case of chloranil the four carbon atoms attached to Cl-atom (C-Cl) are similar and likewise the two carbonyl carbon atoms (C=O) possess exactly similar electronic charges. However in the case of MQ1 the charges on all the atoms are found to be different. The electronic charges on the carbonyl atom is relatively less (-0.200, -0.197 a.u) when compared to that of chloranil (0.734 a.u). These observations may be due to the presence of an electron donating 16

methoxy substituent in the case of MQ1. On complexation with LPA charges of LPA were found to vary from atom to atom as shown in Table 9. The results of spectral and MO studies indicated that the –NH2 group of LPA acts as the donor in the CT complexation with MQ1. The results depicted in Table 9 indicated the charge of N-atom of LPA decreased drastically from – 0.374 to – 0.328 a.u. on complexation with MQ1. Consequently the charges on the carbonyl atoms of MQ were found to increase from – 0.200 to – 0.226 and – 0.197 to – 0.457 a.u. This clearly indicated that appreciable amount of electronic charge has been transferred from –NH2 group of LPA to MQ1 through CT complex formation. This result is also well supported by the calculated dipole moments of the donor and acceptor. The dipole moments of MQ1, LPA and LPA-MQ1 CT complex were calculated to be 3.1450, 1.7609 and 1.5922 D respectively. The dipole moment of the chloranil is zero. The higher dipole moment of MQ1 suggested that the electronic charge distribution in this molecule is highly asymmetric. A decrease in dipole moment of both donor and acceptor, observed on complexation is a clear indication of charge transfer from LPA to MQ1. 4. Conclusions The charge transfer properties of 1,4-benzoquinones possessing varying number of chloro and methoxy substituents with LPA were investigated in aqueous medium. Various spectral techniques have been employed to characterize the interactions. In all the cases, the stoichiometry of the CT interaction was found to be 1:1. The trends in the rate constants and formation constants showed that the strength of the complex formation is in the order of LPA-MQ1>LPA-MQ2>LPA-MQ3>LPA-MQ4. The half wave potential values (E1/2) for the one electron reduction of these acceptors indicated that the 17

electronegativity of the acceptors increased from MQ1 to MQ4. The observed equilibrium, kinetic and electrochemical properties of these acceptors were found to be well supported by ab initio DFT calculations. Acknowledgement The authors thank the University Grants Commission, New Delhi for its financial assistance to carry out this research work.

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M.J. Frisch, G.W. Trucks, H.B. Schlegel, G.E. Scuseria, M.A. Robb, J.R. Cheeseman, J.A.J. Montgomery, T. Vreven, K.N. Kudin, J.C. Burant, J.M. Millam, S.S. Iyengar, J. Tomasi, V. Barone, B. Mennucci, M. Cossi, G. Scalmani, N. Rega, G.A. Petersson, H. Nakatsuji, M. Hada, M. Ehara, K. Toyota, R. Fukuda, J. Hasegawa, M. Ishida, T. Nakajima, Y. Honda, O. Kitao, H. Nakai, M. Klene, X. Li, J.E. Knox, H.P. Hratchian, J.B. Cross, C. Adamo, J. Jaramillo, R. Gomperts, R.E. Stratmann, O. Yazyev, A.J. Austin, R. Cammi, C. Pomelli, J.W. Ochterski, P.Y. Ayala, K. Morokuma, G.A. Voth, P. Salvador, J.J. Dannenberg, V.G. Zakrzewski, S. Dapprich, A.D. Daniels, M.C. Strain, O. Farkas, D.K. Malick, A.D. Rabuck, K. Raghavachari, J.B. Foresman, J.V. Ortiz, Q. Cui, A.G. Baboul, S. Clifford, J. Cioslowski, B.B. Stefanov, G. Liu, A. Liashenko, P. Piskorz, I. Komaromi, R.L. Martin, D.J. Fox, T. Keith, M.A. Al-Laham, C.Y. Peng, A. Nanayakkara, M. Challacombe, P.M.W. Gill, B. Johnson, W. Chen,

21

M.W. Wong, C. Gonzalez, J.A. Pople, Gaussian 03W Revision D.01, Gaussian, Inc., Wallingford CT, 2004. [41]. N.Cho, J.Kim, K.Song, J.K.Lee, J. Ko. Tetrahedron., 68 (2012) 4029-4036. [42]. B.Tejerina, C.M.Gothard, B.A. Grzybowski, Chem.Eur.J., 18 (2012) 5606-5611. [43]. M.Shukla, N.Srivastava, S.Saha, J. Mol. Struc., 1021 (2012) 153-157.

22

Table 1. FT-IR wave numbers (cm-1), intensitya and tentative band assignments for the LPA and its complex with MQ1. ________________________________________________________________________ LPA

MQ1

3066w

1684s 1671 1615

LPA-MQ1 complex

Assignments

3029b

ν(N-H) ; association

1737

ν(C=O)

1679s 1629

ν(C=O)

δ(-NH3+); asymmetric

1625 1602

ν(N-H) ; asymmetric

1562

ν(C=O); asymmetric

1492

δ(-NH3+); symmetric 1482

ν(C=O); symmetric

1409

744 698

a

ν(N-H) ; symmetric

1270m 1099s

1229m 1079m

ν(O-CH3)

909m 778m 722m

823w 732

ν(C-Cl)

744 700

mono substituted benzene

s, strong; m, medium; w, weak; ν, stretching; δ, bending.

23

Table 2. FT-IR wave numbers (cm-1), intensitya and tentative band assignments for the LPA and its complex with MQ2. ________________________________________________________________________ LPA

MQ2

3066w

1663s

LPA-MQ2 complex

Assignments

3029b

ν(N-H) ; association

1736

ν(C=O)

1631m

ν(C=O) δ(-NH3+); asymmetric

1625 1602

ν(N-H) ; asymmetric

1562

ν(C=O); asymmetric

1492

δ(-NH3+); symmetric 1481

ν(C=O); symmetric

1409

744 698

a

ν(N-H) ; symmetric

1270m 1060s

1207m 1041m

ν(O-CH3)

908m 734m

829w 732

ν(C-Cl)

742 700

mono substituted benzene

s, strong; m, medium; w, weak; ν, stretching; δ, bending.

24

Table 3. FT-IR wave numbers (cm-1), intensitya and tentative band assignments for the LPA and its complex with MQ3. ________________________________________________________________________ LPA

MQ3

3066w

1674s 1662 1634

LPA-MQ3 complex

Assignments

3027b

ν(N-H) ; association

1737

ν(C=O)

1683s 1631

ν(C=O)

δ(-NH3+); asymmetric

1625 1604

ν(N-H) ; asymmetric

1562

ν(C=O); asymmetric

1492

δ(-NH3+); symmetric 1483

ν(C=O); symmetric

1409

744 698

a

ν(N-H) ; symmetric

1277m 1052s

1205m 1049m

ν(O-CH3)

940m 747m

923w 732

ν(C-Cl)

741 701

mono substituted benzene

s, strong; m, medium; w, weak; ν, stretching; δ, bending.

25

Table 4. FT-IR wave numbers (cm-1), intensitya and tentative band assignments for the LPA and its complex with MQ4. ________________________________________________________________________ LPA

MQ4

3066w

1666s

LPA-MQ4 complex

Assignments

3002b

ν(N-H) ; association

1737

ν(C=O)

1683m 1626

ν(C=O) δ(-NH3+); asymmetric

1625 1604

ν(N-H) ; asymmetric

1562

ν(C=O); asymmetric

1492

δ(-NH3+); symmetric 1483

ν(C=O); symmetric

1409

1276m 1059s 744 698

a

ν(N-H) ; symmetric

1226m 1049m

ν(O-CH3)

740 702

mono substituted benzene

s, strong; m, medium; w, weak; ν, stretching; δ, bending.

26

Table 5. Effect of concentration of donor and acceptors on the rate of the interaction at 298 K. k1 (10-4), s-1

[D] [A] (10-3 M) (10-5 M)

k2(10-1) s-1mol-1dm3

__________________________________________________________________

LPA-MQ1 LPA-MQ2 LPA-MQ3 LPA-MQ4

__________________________________________________________________

LPA-MQ1 LPA-MQ2 LPA-MQ3 LPA-MQ4

1.3

3.1

1.52

1.19

0.97

0.39

1.1

0.9

0.7

0.3

1.9

3.1

2.07

1.82

1.39

0.61

1.1

0.9

0.7

0.3

2.5

3.1

2.84

2.41

1.98

0.72

1.1

0.9

0.8

0.3

3.1

3.1

3.41

3.09

2.39

0.91

1.1

0.9

0.7

0.3

3.1

2.5

2.92

2.54

1.45

0.74

3.1

1.9

2.91

2.54

1.45

0.74

3.1

1.3

2.94

2.52

1.43

0.71

27

Table 6. Kinetic and thermodynamic parameters for the interaction of LPA with MQ-4 in water.

Systems

λ

k1 (10-4) sec-1 ___________________________

ΔH#a

-ΔS#b

ΔG#c

nm

298

305

LPA-MQ1

541

3.41

4.16

5.85

25

226

93

LPA-MQ2

536

3.09

3.97

4.48

20

245

93

LPA-MQ3

533

2.39

3.68

4.25

27

223

93

LPA-MQ4

527

0.91

1.43

1.63

27

230

94

a

kJ mol-1; bΔS# J K-1 mol-1; cΔG# kJ mol-1

28

313 K

Table 7. Spectral properties of the CT complexes formed between phenylalanine with MQ1-4 in water at 298 K.

Property Formation constant K (dm3 mol-1) Abs. method

LPA-MQ1

LPA-MQ2

4150

1610

Extinction coefficient log ε 2.65 (dm3 mol-1 cm-1) Abs. method

2.63

LPA-MQ3 1110

2.92

LPA-MQ4 689

2.98

Association constant Kf (mol L-1) Emission method

1.7 x 105

1.5 x 105

2.2 x 104

5.6 x 103

Stern-Volmer constant KSV Emission method

95240

83366

24590

24118

29

Table 8. Binding constants (KA) and number of binding sites (n) for LPA–MQ1-4 systems in water medium. Acceptors

KA (mol-1 L)

n

LPA–MQ1

8.1 x 106

1.2

LPA–MQ2

6.6 x 106

1.2

LPA–MQ3

2.1 x 105

1.2

LPA–MQ4

2.8 x 104

1.0

30

Table 9. Mulliken electronic charge (a.u) on various atoms of MQ1, LPA and LPA-MQ1 complex using DFT (6-31G++ (d,p) basis set calculation. Atom number C10 C11 C12 C13 C14 C15 Cl1 Cl2 Cl3 O3 O4 O5

MQ1

LPA

0.459 -0.166 -0.200 0.133 -0.093 -0.197 0.319 0.304 0.267 -0.423 -0.404 -0.174

C7 C8 C9 N1 O1 O2

0.200 -0.364 -0.226 0.521 0.154 -0.457 0.363 0.288 0.267 -0.387 -0.402 -0.327

-0.252 -0.831 0.327 -0.374 -0.421 -0.402

31

LPA-MQ1

-0.328 -0.826 0.355 -0.328 -0.418 -0.441

LPA-MQ1 LPA-MQ2 LPA-MQ3 LPA-MQ4

0.28 0.26 0.24

Absorbance

0.22 0.20 0.18 0.16 0.14 0.12 0.2

0.3

0.4

0.5

0.6

0.7

0.8

mole fraction of the donor

Fig. 1. Job's Continuous variation plots for LPA with acceptors in water at 298 K.

32

0.25

Absorbance

0.20

0.15

0.10

LPA-MQ1 LPA-MQ2 LPA-MQ3 LPA-MQ4

0.05

0

1

2

3

4

Volume of acceptor (ml)

Fig. 2. Photometric titration plots for LPA with MQ1-4 in water at 298 K

33

LPA-MQ1

% T (A.U)

MQ1

LPA

4000

3500

3000

2500

2000

1500

1000

-1

Wavenumber (cm )

Fig. 3. FT-IR spectra of LPA, MQ1 and LPA-MQ1 complex

34

500

1.2

1.0

0.8

0.6

Absorbance

1.0

0.2

D

0.8

0.4

Absorbance

0.0 200

300

400

500

600

700

800

Wavelength (nm)

0.6

0.4

0.2

A C

0.0 200

400

600

800

Wavelength (nm)

Fig. 4. Electronic spectra of LPA with MQ1 in water at 298 K D: LPA; A: MQ1; C: CT complex; Inset Electronic spectrum of synthsised LPA-MQ1 complex

35

D

Fluorescence intensity (AU)

a b c d e f

300

400

500

600

700

Wavelength (nm)

Fig. 5. Fluroscence spectra for LPA-MQ1 system in water at -4

fixed concentration of [D]= {7.8125x10 (curve D)} and variable concentrations of -6 [A] x10 ={7.8125 (curve a), 1.5625 (curve b), 2.3437 (curve c), -1 3.125 (curve d), 3.9063 (curve e), 4.6875(curve f)} mol L at 298 K

36

D a

Fluorescence intensity (AU)

b c d e f g

300

400

500

600

700

Wavelength (nm)

Fig. 6. Fluroscence spectra for LPA-MQ2 system in water at -2

fixed concentration of [D]= {7.8125x10 (curve D)} and variable concentrations of -6 [A] x10 ={7.8125 (curve a), 1.5625 (curve b), 2.3437 (curve c), -1 3.125 (curve d), 3.9063 (curve e), 4.6875(curve f), 5.4688(curve g)} mol L at 298 K

37

D a

Fluorescence intensity (AU)

b c d e f

300

400

500

600

700

Wavelength (nm)

Fig. 7. Fluroscence spectra for LPA-MQ3 system in water at -4

fixed concentration of [D]= {7.8125x10 (curve D)} and variable concentrations of -6 [A] x10 ={7.8125 (curve a), 1.5625 (curve b), 2.3437 (curve c), -1 3.125 (curve d), 3.9063 (curve e), 4.6875(curve f)} mol L at 298 K

38

D

Fluorescence intensity (AU)

a b c d e

300

400

500

600

700

Wavelength (nm)

Fig. 8. Variation of fluorescence spectra of LPA-MQ4 system -4

in water at fixed concentration [D]= {7.8125x 10 (curve D)} and -6 variable concentrations of [A] x10 ={7.8125 (curve a),1.5625 (curve b), -1 2.3437 (curve c),3.125 (curve d),3.9063 (curve e) mol L at 298 K

39

6 LPA-MQ1 r=0.87 LPA-MQ2 r=0.97 LPA-MQ3 r=0.94 LPA-MQ4 r=0.98

5

F0/F

4

3

2

1

0 0.00000

0.00001

0.00002

0.00003

0.00004

0.00005

0.00006

[Q]

Fig. 9. Stern-Volmer plots for the fluorescence quenching of LPA with the acceptors MQ1-4 in water at 298 K

40

0.8

LPA-MQ1 r=0.99 LPA-MQ2 r=0.98 LPA-MQ3 r=0.98 LPA-MQ4 r=0.98

log (Fo-F/F)

0.4

0.0

-0.4

-0.8

-5.2

-5.0

-4.8

-4.6

-4.4

-4.2

log [Q]

Fig.10. Plot of log (Fo-F/F) versus log [Q] of LPA with the acceptors in water at 298 K

41

LPA

MQ1

HOMO (-6.3481 eV)

LUMO (-4.4017 eV)

MQ2

MQ3

MQ4

LUMO (-3.6120 eV)

LUMO (-3.3998 eV)

LUMO (-3.1608 eV)

Fig. 11. The optimized structures of the LPA and MQ1-4 along with frontier orbitals 42

-3.0

MQ4 MQ3

-3.5

LUMO

-5.5

ΔΕ = 2.7361

-5.0 ΔΕ = 1.9464

Energy (eV)

-4.5

ΔΕ = 2.9483

MQ1

ΔΕ = 3.1873

-4.0

MQ2

-6.0 HOMO of LPA

-6.5

Fig. 12. Energy of HOMO of the donor and LUMO of the acceptors ΔΕ = HOMOLPA-LUMOacceptor

43

LPA-MQ1

180 160

LPA-MQ2

140

100 80

3

10 Kf, mol L

-1

120

60 40

LPA-MQ3

20

LPA-MQ4 0 1.8

2.0

2.2

2.4

2.6

2.8

3.0

3.2

ΔΕ (eV)

Fig.13. Correlation between association constant Kf and ΔE

44

* Novel substituted 1,4-benoquinones were employed as acceptors in the CT interaction with a amino acid L-phenylalanine. * The mechanism of the interaction was studied using various spectral techniques. * Progressive replacement of –Cl (-I effect) by –OMe (+M effect) makes the acceptor weaker.

45

Molecular complexes of L-phenylalanine with substituted 1,4-benzoquinones in aqueous medium: Spectral and theoretical investigations

1.2

1.0

D

Absorbance

0.8

0.6

0.4

0.2

A C

0.0 200

400

600

800

Wavelength (nm)

Electronic spectra of LPA with MQ1 in water at 298 K D: LPA; A: MQ1; C: CT complex;

UV-Vis, FT-IR, and fluorescence spectral techniques were employed to investigate the mechanism of interaction of L-phenylalanine with 1,4-benzoquinone possessing varying number of methoxy/chloro substituents (MQ1-4). K. Ganesh, E. H. El-Mossalamy, A.Satheshkumar, C.Balraj and K. P. Elango

46