SYNTHESIS, SPECTRAL CHARACTERIZATION AND DNA BINDING PROPERTIES OF LANTHANIDE (III) COMPLEXES WITH 2-FORMYLPYRIDINE NICOTINOYLHYDRAZONE

SYNTHESIS, SPECTRAL CHARACTERIZATION AND DNA BINDING PROPERTIES OF LANTHANIDE (III) COMPLEXES WITH 2-FORMYLPYRIDINE NICOTINOYLHYDRAZONE

B. Moksharagni and K. Hussain Reddy^*

Department of Chemistry, Sri Krishnadevaraya University, Anantapuramu - 515003, Andhra Pradesh, India.

ABSTRACT: The lanthanide (III) complexes of general formula of [Ln(FPNH)₂(NO₃)](NO₃)₂.nH₂O (where, Ln = La, Ce, Pr, Nd, Sm; FPNH = 2-formylpyridine nicotinoylhydrazone, n = 0, 2 and 5 for La/Ce/Pr, Sm and Nd complexes, respectively) were synthesized under mild reaction conditions with excellent yields. The complexes were characterized based on elemental analysis, molar conductivity, and infrared spectroscopy. Molar conductivity data suggest that the complexes are 1:2 electrolytes. IR spectral data suggest that FPNH acts as neutral tridentate ligand. Spectral data of complexes suggest that the ligand binds metal ion through pyridine nitrogen, azomethine-nitrogen and amido-oxygen donor atoms. Electrochemical behaviour of metal complexes was investigated by using cyclic voltammetry. The complexes undergo quasi-reversible one electron reduction. The binding interaction of complexes with CT-DNA was investigated using absorption spectrophotometry. Based on spectral changes, groove binding of complexes to DNA is suggested.

Lanthanide (III) complexes, 2-Formylpyridine nicotinic acid hydrazone, Spectral characterization, DNA binding

INTRODUCTION: Investigations of lanthanide complexes with polydentate ligands have received considerable attention because of their potential applications in various fields ^1-6. Hence, investigation of newer lanthanide complexes is a potential field of research in inorganic chemistry.  The chemistry of lanthanide complexes is of interest owing to their applications in biology and Medicine. Herein we report synthesis, spectral characterization and DNA binding properties of lanthanide (III) complexes of 2-formylpyridine nicotinoylhydrazone for the first time.

Experimental: Lanthanide nitrates, 2-formylpyridine and nicotinic acid hydrazide, purchased from Aldrich chemicals were used without further purification. Lanthanide salts were stored in desiccators to prevent hydration. CT DNA was purchased from Genie Biolabs, Bangalore, India.

Synthesis of 2-formylpyridine nicotinoyl hydrazone (FPNH): A 5 mmol of 2-formylpyridine (0.56 mL) dissolved in 20 mL of methanol was added to a methanolic solution of nicotinic acid hydrazide (0.685g, 5 mmol). The contents were taken in a round bottom flask stirred for 20 min. A yellow colored product was formed. It was collected by filtration, washed with a few drops of ethanol and dried in vacuum. Yield of the product was 85%, mp: 162-164 °C.

General procedure for the synthesis of lanthanide complexes: To the ligand (0.480g, 2 mmol) dissolved in 20mL of methanol, lanthanum (III) nitrate hexahydrate [Ln(NO₃)₃.6H₂O] (1mmol) dissolved in 10mL of methanol was added. The reaction mixture (taken in 100mL round bottom flask) was heated under reflux on water bath for 2 - 4 hr. Upon cooling solid substance was formed. It was collected by filtration, washed with 10mL of hexane and dried in vacuum. Physicochemical and analytical data of lanthanide complexes are included in Table 1.

Physical measurements: The elemental analyses were performed using a Perkin-Elmer 2400 CHNS elemental analyzer. The molar conductance of the complexes in DMF (10^-3 M) solution was measured at 28 °C with a Systronic model 303 direct-reading conductivity bridge. The electronic spectra were recorded in DMF with a Perkin Elmer UV Lamda - 50 spectrophotometer. FT-IR spectra in KBr disc were recorded in the range 4000 - 400 cm^-1 with a Perkin Elmer spectrum 100 spectrometer. The cyclic voltammetry was performed with a CH instruments 660C electrochemical analyzer and a conventional three electrodes, Ag/AgCl reference electrode, glassy carbon working electrode and platinum counter electrode. Nitrogen gas was purged and measurements were made on the degassed (N₂ bubbling for 5 min) complex solution in DMF (10^-3 M) containing 0.1M tetrabutyl ammonium hexaflourophosphate (TBAHP) as the supporting electrolyte.

DNA binding experiments: Interaction of complexes with calf thymus DNA was studied by electronic absorption spectroscopy. The interaction of the complexes with DNA was carried out in tris-buffer. Solution of calf thymus - DNA (CT-DNA) in (0.5 mM NaCl/5 mM Tris-HCl; pH = 7.0) buffer gave absorbance ratio at 260 nm and 280 nm of 1.89 indicating that the DNA was sufficiently free of proteins. The DNA concentration per nucleotide was determined by absorption coefficient (6600 dm³ mol^-1 cm^-1) at 260 nm. Stock solutions stored at 4 ºC were used after no more than four days. The electronic spectra of metal complexes in aqueous solutions were monitored in the absence and presence of CT-DNA.

Absorption titrations were performed by maintaining the metal complex concentration 20 × 10^-6M and varying the nucleic acid concentration (0-17.4 ×10^-4M). The ratio of r = [complex]/ [DNA] values vary from 8.84 to 1.10.  Absorption titration experiments were performed by maintaining the metal complex concentration constant while gradually increasing the concentration of CT-DNA within 0-80μM.

RESULTS AND DISCUSSION: The ligand, FPNH was used for the preparation of lanthanide metal complexes. The ligand was characterized based on IR, NMR and Mass Spectral data.

IR spectra: Bands at 3147, 1670, 1623, 1566, 991 and 753 cm^-1 are respectively assigned to ν(NH), ν(C=O) (Amide I), ν(C=N), ν(Py ring), ν(Py bending), and δ (Py ring oop bending) vibrations, respectively.

¹H-NMR spectra: The spectrum was recorded in deuterated DMSO. δ 7.39-7.59 (multiplet 4H), δ 8.73-8.81 (multiplet 4H), δ 8.57 (singlet 1H), δ 11.01 (singlet 1 H), are respectively assigned to pyridine, nicotine, -NH and formyl proton.

GC-MS: The spectrum (Fig. 1) shows peak at (m/z) 240 corresponding to molecular ion peak of the ligand.

FIG. 1:  GC MASS SPECTRUM OF FPNH

The complexes are obtained in good yields. The preparation of lanthanide complexes is given in Scheme 1. The complexes are stable at room temperature and non-hygroscopic and could be stored easily even at room temperature. The complexes are partially soluble in water, methanol, and ethanol and readily soluble in acetonitrile (CH₃CN), DMF and DMSO.

SCHEME 1: SYNTHESIS OF LANTHANIDE COMPLEXES WITH FPNH

Where,   M = La, Ce, Pr, Nd, Sm;

The analytical data (Table 1) are consistent with the proposed molecular formulae [Ln(FPNH)₂ (NO₃](NO₃)₂ (where, Ln = La, Ce, Pr, Nd, Sm; FPNH = 2-formylpyridine nicotinoyl hydrazone), of complexes. The molar conductivity values (87-132 Ω ^-1 cm² mol^-1) for the complexes suggest that these are 1:2 electrolytes ⁷. The presence of nitrate in the complexes was confirmed by brown ring test ⁸.

Electronic spectra: The electronic spectra of ligand and their corresponding lanthanide (III) complexes are recorded in DMF. Typical electronic spectra (a) FPNH ligand (b) [La(FPNH)₂(NO₃)](NO₃)₂ complex are given in Fig. 2. In the electronic spectra of complexes a broad peak is observed in the high energy region at 33230 – 33560 cm^-1 assigned to π - π* transition. A strong peak is observed in the low energy region (26690 – 27180 cm^-1) is assigned to charge transfer transition. Electronic spectral data are given in Table 2.

TABLE 1: ANALYTICAL AND PHYSICO-CHEMICAL PROPERTIES OF APINH LIGAND AND ITS LANTHANIDE(III) COMPLEXES

* Decomposes, ^†Units:  Ω^-1 cm² mol^-1 in DMF.

FIG. 2: ELECTRONIC SPECTRA OF (A) FPNH LIGAND AND (B)[SM(FPNH)₂(NO₃)](NO₃)₂ COMPLEX

The weak f-f transitions occurring in the region due to lanthanide ion in the complexes are obscured by the intense ligand bands. Also the ligand band remains largely unaffected on complexation and insensitive to the lanthanide ion. No absorption band due to f-f transition of lanthanide (III) ions could be located in the visible region in the spectra of complexes. This is probably due to the fact that the f-f bands are weak and obscured by the intense (M L) charge transfer band ⁹.

TABLE 2: ELECTRONIC SPECTRAL DATA (CM-1) OF LANTHANIDE (III) COMPLEXES IN SOLUTION STATE*

* Spectra of the complexes were recorded in DMF solvent.

† Molar absorptivity Units, L. mol^-1 cm^-1.

Infrared spectra: IR spectrum of FPNH is compared with its lanthanide complexes to determine donor atoms of ligand. IR spectral data are given Table 3. The IR spectrum of the ligand has several prominent peaks due to ν_N-H, ν_C=O and ν_C=N stretching modes, respectively. In the spectra of complexes, ν_N-H and ν_C=O bands are present suggesting that the hydrazone acts as neutral ligand. The ν_C=N in the spectrum of free hydrazone is shifted to lower frequency in the spectra of complexes suggesting the involvement of azomethine nitrogen in chelation. IR data suggest that the hydrazone ligands act as neutral tridentate ligand.

TABLE 3: INFRARED SPECTRAL DATA (CM^-1) FOR THE FPNH LIGAND AND ITS LANTHANIDE (III) COMPLEXES

The IR spectrum of the complexes demonstrates the presence of coordinated nitrate. The two strong bands are observed in complexes due to the presence of coordinated nitrates. The two strong bands associated with asymmetric and symmetric stretch of coordinated to NO₃^- (C_2v) group appear in the range 1417- 1468 (ν₁) and 1086 - 1089 (ν₄) cm^-1. The frequency separation ∆ν (ν₁- ν₄) increases as the coordination of nitrate group increases from monodentate to bidentate and/or bridging. The magnitude of ∆ν is used to establish the type of nitrate coordination. In the present complexes the ∆ν is about 180 cm^-1 and is typical of bidentate bonding of nitrate. The other vibrational band is observed around 1380 cm^-1 in IR spectra of complexes indicating the presence of ionic nitrate ¹⁰. (D_3h symmetry, free NO₃^- ion). The data indicate that the complexes contain free ionic nitrate and coordinated bidentate nitrate groups. The new bands in 416 - 418 and 522 - 524 cm^-1 region are assigned to ν (Ln–O) and ν (Ln-N) vibration respectively.

Based on molar conductivity, magnetic moments, electronic spectral data, a tentative and general structure for the complexes is given in Fig. 3.

Electrochemical studies: Redox behavior of the lanthanide (III) complexes has been investigated by cyclic voltammetry in DMF using 0.1M tetrabutyl ammonium hexaflourophosphate (TBAHEP) as supporting electrolyte. The cyclic voltammetric profile of [Sm(FPNH)₂(NO₃)](NO₃)₂ complex  is shown Fig. 4.

FIG. 3: GENERAL STRUCTURE FOR LANTHANIDE (III) COMPLEXES.   (M =   LA, CE, PR, ND AND SM)

FIG. 4: CYCLICVOLTAMMETRIC PROFILE OF [SM(FPNH)₂(NO₃)](NO₃)₂ COMPLEX AT DIFFERENT SCAN RATES 25, 50, 75, 100 MV. S^-1 

The electrochemical data of complexes are presented in Table 5. Repeated scans at various scan rates suggest the presence of stable redox species in solution. E_1/2 values are incorporated in Table 4.

TABLE 4:  CYCLIC VOLTAMETRIC DATA OF LANTHANIDE(III) COMPLEXES

 ^a log K_c =0.434ZF/RT∆E_p;  ^b ∆G = -2.303RT log K_c

TABLE 5:  ELECTRONIC ABSORPTION DATA UPON ADDITION OF CT-DNA TO THE COMPLEX

It may be inferred that Ln(III) complexes undergo reduction to their respective Ln(II) complexes ¹¹. The complexes have large separation (134- 274 mV) between anodic and cathodic peaks indicating quasi-reversible character. The log K_c and ∆G ̊ values suggest that the complexes are stable in solution state.

DNA binding studies: Electronic absorption spectroscopy is an effective method for examining the interaction of DNA with metal complexes. The binding interaction of complexes with CT-DNA was monitored by comparing their absorption spectra with and without CT-DNA. All the complexes exhibit an intense absorption band due to π→π* transition. Absorption spectra of [Ce(FPNH)₂(NO₃)](NO₃)₂ in the absence and in  presence of CT-DNA are shown Fig. 5.

FIG. 5: ABSORPTION SPECTRA OF [PR(FPNH)₂ (NO₃)](NO₃)₂ IN THE ABSENCE AND IN THE PRESENCE OF INCREASING CONCENTRATION OF CT-DN; INSET IS A PLOT OF  [DNA] / (Ε_A –Ε_F) VERSUS [DNA]

The intrinsic binding constants (K_b), were determined by using the equation:

[DNA]/(ε_a-ε_f) = [DNA]/(ε_b-ε_f) + 1/K_b (ε_b-ε_f)  ---- (1)

Where [DNA] is the concentration of DNA in base pairs, ε_a, ε_b and ε_f are apparent extinction coefficient (A_obs/ [M]), the extinction coefficient for the metal (M) complex in the fully bound form and the extinction coefficient for free metal (M) respectively.

A plot of [DNA]/(ε_a-ε_f) versus [DNA] gave a slope of 1/ (ε_b-ε_f), and vertical intercept equal to 1/ K_b (ε_b–ε_f); K_b was calculated from these values. The binding constants (Table 5) suggest that the complexes bind DNA strongly. On addition of DNA, the absorbance of the complexes decreases (hyperchromism) and absorption maximum of all complexes is shifted to higher wavelength (bathochromism). A sharp isobestic point (317 nm) for the absorption titration curves (Fig. 5) suggests that there is only one mode of binding ¹². Small bathochromic shift (Δλ = 2 nm) low binding constants (K_b) values and bulky nature of complexes suggest groove binding of complexes to DNA.

CONCLUSION: Lanthanide complexes of 2- formylpyridine nicotinic acid hydrazone have been synthesized and characterized based on physicochemical spectral data. A general structure is proposed for the complexes. DNA binding properties of lanthanide complexes are investigated using absorption spectroscopy.

ACKNOWLEDGEMENTS: One of the authors (B. Moksharagni) is thankful to UGC, New Delhi for the award of BSR Junior Research Fellowship. The authors are thankful to UGC, New Delhi [Sanction No.Lr.No.F 40-80/2011(SR)] for financial support. The authors also thank UGC and DST for providing equipment facility under SAP and FIST programs respectively. KHR is thankful to UGC for the sanction of one-time grant [sanction Lr. No. F.19-106/2013(BSR)] for financial support.

CONFLICTS OF INTEREST: Nil

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    How to cite this article:

    Moksharagni B and Reddy KH: Synthesis, spectral characterization and DNA binding properties of lanthanide(III) complexes with 2-formylpyridine nicotinoylhydrazone. Int J Pharm Sci Res 2017; 8(8): 3379-84.doi: 10.13040/IJPSR.0975-8232.8(8).3379-84.

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