Accepted Manuscript Synthesis, structure, spectral characterization, electrochemistry and evaluation of antibacterial potentiality of a novel oxime-based palladium(II) compound Nirmalya Bandyopadhyay, Miaoli Zhu, Liping Lu, Debmalya Mitra, Mousumi Das, Piu Das, Amalesh Samanta, Jnan Prakash Naskar PII:
S0223-5234(14)00962-3
DOI:
10.1016/j.ejmech.2014.10.035
Reference:
EJMECH 7442
To appear in:
European Journal of Medicinal Chemistry
Received Date: 10 July 2014 Revised Date:
3 September 2014
Accepted Date: 12 October 2014
Please cite this article as: N. Bandyopadhyay, M. Zhu, L. Lu, D. Mitra, M. Das, P. Das, A. Samanta, J.P. Naskar, Synthesis, structure, spectral characterization, electrochemistry and evaluation of antibacterial potentiality of a novel oxime-based palladium(II) compound, European Journal of Medicinal Chemistry (2014), doi: 10.1016/j.ejmech.2014.10.035. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
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Graphical abstract Synthesis, structure, spectral characterization, electrochemistry and evaluation of
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antibacterial potentiality of a novel oxime-based palladium(II) compound Nirmalya Bandyopadhyaya, Miaoli Zhub, Liping Lub, Debmalya Mitrac, Mousumi Dasc, Piu
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Dasc, Amalesh Samantac, Jnan Prakash Naskara*
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Highlights Synthesis, structure, spectral characterization, electrochemistry and evaluation of
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antibacterial potentiality of a novel oxime-based palladium(II) compound Nirmalya Bandyopadhyaya, Miaoli Zhub, Liping Lub, Debmalya Mitrac, Mousumi Dasc, Piu Dasc, Amalesh Samantac, Jnan Prakash Naskara*
Synthesis of oxime-based ligand and its Pd(II) complex.
Spectroscopic aspects and X-ray single crystal structure of the complex.
Quasi-reversible Pd(II)/Pd(I) reduction couple.
This complex exhibits satisfactory bactericidal as well as bacteriostatic activity.
The bioactivity was substantiated by SEM study.
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Synthesis, structure, spectral characterization, electrochemistry and evaluation of
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antibacterial potentiality of a novel oxime-based palladium(II) compound
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Nirmalya Bandyopadhyaya, Miaoli Zhub, Liping Lub, Debmalya Mitrac, Mousumi Dasc, Piu
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Dasc, Amalesh Samantac, Jnan Prakash Naskara*
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a
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India.
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Fax: +(91)(33)2414 6223 E-mail: [email protected]
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b
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Engineering of the Education Ministry, Shanxi University, 92 Wucheng Road, Taiyuan, Shanxi
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Department of Chemistry, Inorganic Chemistry Section, Jadavpur University, Kolkata 700 032,
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Institute of Molecular Science, Key Laboratory of Chemical Biology and Molecular
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030006, People's Republic of China.
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c
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Kolkata 700 032, India.
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E-mail: [email protected]
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Abstract
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The title monomeric Pd(II) compound, [Pd(L)(Cl)], was synthesized in moderate yield out of the
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reaction of equimolar proportion of Na2[PdCl4] and 3-[(5-bromo-2-hydroxy-benzylidene)-
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hydrazono]-butan-2-one oxime (LH) in tetrahydrofuran milieu. LH is a 1:1 Schiff-base
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condensate
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[Pd(L)(Cl)] has been characterized by C, H and N microanalyses, 1H and
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FT-IR, Raman spectra, UV-Vis spectra and molar electrical conductivity measurements.
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[Pd(L)(Cl)] is diamagnetic. Structural elucidation reveals that the palladium center in [Pd(L)(Cl)]
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is nested in ‘N2OCl’ coordination environment. The geometry around Pd in [Pd(L)(Cl)] is
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distorted square-planar. The redox behavior of [Pd(L)(Cl)] in DMF shows a reduction couple,
2,3-butanedionemonoxime
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monohydrazone
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C NMR, FAB-MS,
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Pd(II)/ Pd(I) at –0.836 V versus Ag/AgCl. The in vitro antimicrobial activity of [Pd(L)(Cl)] was
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screened against both Gram-positive and Gram-negative human pathogenic bacteria. This
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bioactivity was substantiated with SEM study. [Pd(L)(Cl)] exhibits satisfactory bactericidal as
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well as bacteriostatic activity.
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Key words: Oxime; Palladium; Crystal structure; Redox; Antibacterial; SEM study.
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30 1. Introduction
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Synthesis, characterization and evaluation of biological activities of novel metal-based
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compounds are a rapidly growing area of contemporary research [1–3]. Drugs based on transition
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metal compounds are superior to organic-based drugs so much so that medicinal inorganic
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chemistry now is an emerging area of research [4]. Again, bacterial drug resistance potentiality is
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increasing alarmingly [5]. Consequently, development of new therapeutic agents to combat
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bacterial infection warrants current attention [6,7]. Synthesis of novel antibacterial drugs with
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properly known structure is important to understand their behavior in biological environment and
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subsequent possible mode of action. The transition metal compounds of hetero donor ligands
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show promising biological activities with reduced toxicity [8]. Palladium, an important platinum
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group metal [9], is noteworthy in this perspective. The compounds of palladium manifest
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diverse biological activities [10]. Palladium-based compounds are well-known antitumor [11],
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antiviral [12], antifungal [13], antiinflammatory [13], antimalarial [14] and antibacterial [15]
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agents. The intense and diverse bioactivities of various palladium compounds on bacterial strains
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are well-documented in the literature [10,15,16]. Pd(II) being a soft Lewis acid has an inherent
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tendency to bind soft donor centers like N, S in its d8 square-planar structural disposition [17].
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Accordingly, most of the antibacterial palladium(II) compounds are from N- and S-based ligands
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like thiosemicarbazones [16–18], thiocarbazates-thioamides [10], thiodiamines [19] etc.
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Palladium(II) compounds, synthesized from ligands with N donor only, are also known to
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exhibit antimicrobial activities against Gram-negative strains like E. coli, P. aeruginosa, S.
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marcescens along with Gram-positive strains like S. aureus, B. subtilis [20]. Thus it seems that
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for showing bioactivity, palladium complexes of N, S donor ligands are of obvious choice.
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Examples are also known from ligands having N, O donor ligands [21]. Palladium complexes
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from the alkyl derivative of thiosalicylic acids having S, O donor sides also exhibit moderate
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antimicrobial activity [22]. On the contrary, such type of bioactivity of monomeric Pd(II)
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compound in a N, O donor oxime-based Schiff-base ligand is rare indeed [23]. For quite some
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time, we are interested to develop novel oxime-based Pd(II) compounds as potential antibacterial
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agents with pronounced activity. Herein we wish to report our endeavor along that line – the
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synthesis, spectroscopic aspects, X-ray structure and electrochemical behavior of a novel Pd(II)
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complex. The antibacterial activities of the ligand, 3-[(5-bromo-2-hydroxy-benzylidene)-
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hydrazono]-butan-2-one oxime (LH) and its palladium(II) compound, [Pd(L)(Cl)], was screened
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against a good number of human pathogenic bacterial cell lines. The bioactivity was also
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substantiated with Scanning Electron Microscopic (SEM) studies in proper cases.
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2. Results and discussion
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2.1. Chemistry
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2.1.1. Synthesis and characterization
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A Schiff-base ligand, 3-[(5-bromo-2-hydroxy-benzylidene)-hydrazono]-butan-2-one oxime
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(LH), has been used for the present study. The ligand has been prepared by the Schiff-base
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condensation of equimolar proportion of 2,3-butanedionemonoxime monohydrazone and 5-
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bromosalicylaldehyde in tetrahydrofuran. LH was spectroscopically characterized by 1H,
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NMR and ESI-MS (Supporting Information, Figs. S1–S3). Subsequent reaction in 1:1 molar
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proportion of LH and Na2[PdCl4] in tetrahydrofuran enables us to isolate the orange monomeric
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Pd(II) compound, [Pd(L)(Cl)] (1) in moderate yield. FAB-MS peaks in m-nitrobenzyl alcohol
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matrix at 402.1 (for
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Information, Fig. S4). This is a testimony to the formation of 1. C, H and N micro analytical data
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of the single crystals of 1 also substantiate the above formulation. In Fig. 1, the synthetic scheme
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of LH along with 1 is outlined.
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The diamagnetic nature of the compound suggests a square-planar disposition of the palladium
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center in 1. This diamagnetic nature enables us to detect all the carbon nuclei present in 1
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(Supporting Information, Fig. S5). 1H NMR spectral data of 1 were also recorded in DMSO-d6
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for the evaluation of the number and nature of all the protons relative to TMS with chemical
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shifts given in ppm (Supporting Information, Fig. S6).
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The IR spectrum of 1 (Supporting Information, Fig. S7) is consistent with the structure
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confirmed by single crystal X-ray diffraction studies. Free LH have infrared spectral bands at ν
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1612(s) [ν(C=N) of imine], 1477(s) [ν(C=N) of oxime)] and 1184(s) [ν(N–O)]. In 1, these bands
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appear respectively at 1624, 1475 and 1182 cm–1. The strong band at 1612 cm–1 is assigned to
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the C=N stretching vibration, characteristic for a Schiff-base ligand. The ring skeletal vibration
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(C=C) of free ligand was assigned at 1547 cm–1. In 1, this band undergoes a shift towards higher
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wavenumber by 17 cm–1 and appears at 1564 cm–1. The Ar–O stretching frequency [ν(C–O) of
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phenolic –OH] appears as a strong band at 1271 cm–1 in the free ligand, and in 1 it appears at
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1267 cm–1. Thus, the lower wavenumber shift of the Ar–O stretching by ca. 4 cm–1 indicates that
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Pd–O (phenolic) bond is formed [24,25]. The Pd–Cl stretching frequency, [ν(Pd–Cl)] is
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Br) correspond to [Pd(L)]+ for 1 (Supporting
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generally observed in the range 300–400 cm–1 [26]. Here we could not observe it due to the
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limitation of the window (4000–400 cm–1) of the spectrometer used. The ν(Pd–N) modes can be
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identified at 445 cm–1 [27] in the FT-IR of 1. However, we have discerned the ν(Pd–Cl), ν(Pd–N)
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and ν(Pd–O) vibrations in the Raman spectrum of 1, shown in Fig. S8 (Supporting Information).
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The Raman shift values at 300, 440 and 647 cm–1 are assigned to the ν(Pd–Cl), ν(Pd–N) and
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ν(Pd–O) vibrations respectively [28–30].
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In the electronic spectrum of 1 in DMSO and DMF, only one d–d band was observed at 440 and
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443 nm respectively. The broad band in this region is assigned to the dz2 to dx2–y2 transition [20].
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This transition brings about the orange hue to the compound. This general feature pertains to the
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low-spin d8 metal ion complexes with square-planar geometry. The molar electrical
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conductivities of ~10–3 M solutions of 1 in DMF is 57 Ω–1 cm2 mol–1. This value in DMSO is
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only 10 Ω–1 cm2 mol–1.Thus 1 behaves as a non-electrolyte both in DMF and DMSO [31].
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2.1.2. Molecular structure of [Pd(L)(Cl)] (1)
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The X-ray crystal structure (Fig. 2) of 1 was determined. In Table 1, a summary of data
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collection, structure refinement for 1 is given. Selected bond lengths, bond angles and hydrogen-
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bond geometry are provided in Tables S1 (Supporting Information) and Table 2 respectively.
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The palladium center in 1 is nested in ‘N2OCl’ core. The Pd1–N1 and Pd1–N3 bond distances
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are almost the same, having the values of 1.976(5) and 1.978(5) Ǻ respectively. Pd1–O1 bond
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distance [1.963(4) Ǻ] is quite similar to Pd–N bonds. The bond distance of Pd1–Cl1 is 2.326(2)
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Ǻ. This bond is much longer than Pd–N and Pd–O bonds. These bond length values correlate
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well with those reported earlier in similar ‘N2OCl’ core for mononuclear Pd(II) compounds [32].
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The cis-angles, O1–Pd1–Cl1, N1–Pd1–N3, N3–Pd1–Cl1 and O1–Pd1–N1 are of values 84.8,
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90.2, 91.3 and 93.7° respectively. The trans-angles, O1–Pd1–N3 and N1–Pd1–Cl1 respectively
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are of 176.0 and 177.1°. Thus the range (84.8–93.7°) of the cis-angles is in copious deviation
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from 90°. Alike are the deviations of the trans-angles from 180°. Consequently, the geometry
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around the palladium(II) center in 1 experiences distortion. The palladium center is deviated by
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0.015(2) Ǻ from the average square-based basal plane defined by N1, N3, O1 and Cl1. Thus the
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core no longer retains its planarity.
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To augment the geometric shape of 1, we have taken recourse to estimate the Addison angular
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structural parameter (4) for four-coordinated complexes [33,34]. By definition, 4 = [360°–
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(α+β)] /141° where α and β respectively are the largest and second largest angles around the
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metal center. The 4 value is equal to 0 for a perfectly square-planar geometry and 1 for a regular
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tetrahedral core [34]. For 1, the angle N1–Pd1–Cl1 (177.1°) is α and O1–Pd1–N3 (176.0°) is β.
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Accordingly, the 4 value obtained here is 0.049. This value closely approaches to 0. Thus the
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geometry around the palladium center in 1 is almost square-planar. However, the sum of the
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subtended cis-angles around the palladium centre is 360°. The rationale behind this is that the Pd
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center gains some stability due to the chelate effects of two 6-membered planar rings.
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The intermolecular hydrogen bonding exclusively consists of the classical OH···Cl bond. Fig.
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S9 (Supporting Information) represents the packing diagram along with the H-bonded network in
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1.
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2.1.3. Electrochemistry
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The electrochemical aspect of 1 has been studied by CV in DMF at Glassy Carbon (GC) working
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electrode (Supporting Information, Fig. S10).
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1 exhibits one electrochemical response at E½ value of –0.737 V vs Ag/AgCl (Couple I). Under
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the same experimental condition, the ferrocene-ferrocenium (Fc/Fc+) couple appears at 0.44 V vs
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Ag/AgCl. Comparison of the cyclic voltammetric peak currents of 1 with those of the standard
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redox couple, Fc/Fc+, establishes that couple I involves only one electron. LH is
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electrochemically dormant in the potential range of interest here. Consequently, this response can
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be assigned as Pd(II) to Pd(I) reduction. Peak potential of couple I is –0.836 V with 39.80 A
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peak–current. In the reverse cycle, the subsequent oxidative response can be observed at –0.637
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V vs Ag/AgCl. The corresponding anodic peak current, ipa is 1.86 A. The ipc to ipa ratio is
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21.40, a value grossly deviated from unity. Again ipc, the cathodic peak current, increases with
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the square root of the sweep rate (ν1/2), though the increase is not in proportionality. Moreover,
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Epc, the cathodic peak potential shifts more negatively with the increase in sweep rate (ν). Thus
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the electron transfer at couple I occurs quasi-reversibly [35]. Pd(II) to Pd(I) reductions in DMF
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are known to occur at ~ –0.9 V vs Ag/AgCl [23,36].
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2.2. Antimicrobial Activity
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Various pathogenic bacteria pose serious threats to human kind. Virulent Gram-negative strains
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of E. coli can cause gastroenteritis, urinary tract infection, infection of wounds and neonatal
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meningitis; while P. aeruginosa causes infection of wounds and septicemia. Gram-positive
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strains like M. luteus can cause skin infections that produce pruritic eruptions sometimes even
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with central ulceration. Again Gram-positive B. subtilis is responsible for food poisoning [16,
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37–39]. Harmful bacteria often develop resistance to the existing antidotes. For this, novel
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compounds with pronounced cytotoxic/cytostatic effects are the need of the day. In our present
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study we have incorporated specifically the above strains also.
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2.2.1. Antimicrobial assay
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1 has moderate antimicrobial properties. LH does not have such efficacies. As shown in Table
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S2 and Table S3 (Supporting Information), the compound was screened against bacteria using
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Well-diffusion method. Among the listed organisms in Table S2, our compound 1 was active
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against 12 tested organisms (Fig. 3).
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The compound displays moderate activity with high MIC value against Salmonella typhi 62,
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Vibrio cholerae 20, Shigella dysenteriae 1, Bacillus cereus 11778 , Bacillus subtilis 6633,
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Micrococcus luteus 10240, Bordetella bronchiseptica 4617, Streptococcus epidermidis 12228,
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Shigella dysenteriae 37 (400 µg/ml; diameters of inhibition zone ranges 7.0 to 14 mm) and low
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value against Escherichia coli 25938, Salmonella typhimurium NTCC 74, Pseudomonas
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aeruginosa 25619 (200 µg/ml; inhibition zone ranges: 10 to 15 mm) (Supporting Information,
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Fig. S11).
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Effectiveness of an antibacterial drug can be judged on the basis of its zone of inhibition. The
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data obtained here shows that 1 has the ability to inhibit the metabolic growth of our tested
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organisms (Fig. 4).
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2.2.2. Growth kinetic studies
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The results of 6 selected microorganisms have been shown graphically. The graph depicts the
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respective numerical logarithmic values of number of colonies after incubation. By analysing the
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bacterial growth in Fig. S12 and Fig. S13 (Supporting Information), it is clear that compound 1
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had its effect as a bactericidal on Pseudomonas aeruginosa 25619 and Micrococcus luteus 10240
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respectively. Compound 1 had its effect as bacteriostatic on the rest of the organisms as shown in
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Figs. S14–S17 (Supporting Information). In Table S4 (Supporting Information) the respective
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data are tabulated.
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2.2.3. Scanning Electron Microscopic (SEM) observation
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Scanning electron microscopy was carried out to assess the morphological changes in bacterial
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cells following exposure to the compound. SEM studies were carried out on Bacillus cereus
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11778 and Pseudomonas aeruginosa 25619 strains due to their higher and moderate zone of
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inhibition. Untreated Bacillus cereus 11778 (Fig. 5A1) and Pseudomonas aeruginosa 25619
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(Fig. 5B1) cells had apparently shown normal rod-shape and smooth surface with no visible
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damage. After treatment with compound 1, the cell permeability of Bacillus cereus (Fig. 5A2)
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was increased leading to an accumulation of fluid, resulting in swelling and cell deformation.
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However, in case of Pseudomonas aeruginosa (Fig. 5B2), the cell walls were digested at various
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junctions leading to the destruction of cells and subsequent death.
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The possible mode of bioactivity of our palladium complex may be explained by Tweedy’s
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chelation theory [40]. Generally the polarity of any central metal ion is lessened considerably
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due to chelation. This is due to the partial sharing of metal’s positive charge with the ligand and
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possible π-electron delocalization over the chelate ring. Consequently chelation increases the
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lipophilic character of the central metal ion enabling the subsequent permeation of the compound
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through the lipid layer of the microbial cell membrane. After penetration, the possible mode of
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action involves the formation of hydrogen bond preferably through the azomethine group of the
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compound with the active sites of the microbial cell constituents. As a result, the normal
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physiological cell process of the microbes gets potential interference from the drug [41].
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From the perspective of bactericidal activity, the palladium-based metallo-drugs are generally
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more toxic towards Gram-positive strains than Gram-negative strains [42]. The reason is due to
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the inherent difference in the structure of the cell walls. The walls of Gram-negative cells are
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more complex than those of Gram-positive cells [38]. The outer cell membrane of Gram-
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negative bacteria is constituted of a complex lipopolysaccharide (LPS) that acts as an endotoxin;
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protecting these bacteria from the extraneous control agents even from lysozyme and penicillin
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[43]. Interestingly, 1 is more effective to three of our tested Gram-negative strains (Escherichia
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coli 25938, Salmonella typhimurium NTCC 74, Pseudomonas aeruginosa 25619). Again, 1 is
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moderately active to Gram-positive strains under study. Thus 1 behaves as a broad spectrum
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antibacterial agent. The result obtained here is of great significance for biological and
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pharmaceutical control applications.
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3. Conclusions
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Here we have been able to synthesize and structurally characterize a novel Pd(II) compound
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[Pd(L)(Cl)] from an oxime-based ligand, 3-[(5-bromo-2-hydroxy-benzylidene)-hydrazono]-
214
butan-2-one oxime (LH). Diamagnetic [Pd(L)(Cl)] is in square-planar ‘N2OCl’ coordination
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core. Redox studies in DMF show a quasi-reversible Pd(II)/Pd(I) reduction couple at –0.836 V
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versus Ag/AgCl. [Pd(L)(Cl)] shows promising broad spectrum antimicrobial activity against a
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good number of both Gram-positive and Gram-negative human pathogenic bacteria.
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4. Experimental
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4.1. Chemistry
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4.1.1. Materials
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All chemicals and solvents employed in this work were of analytical reagent grade. They were
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used as received without any further purification. Palladium chloride, 2,3-butanedione
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monoxime, 98% and 5-bromosalicylaldehyde, 98% were procured from Sigma-Aldrich
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Chemicals Pvt. Ltd; while hydrazine hydrate, tetrahydrofuran and methanol were from Merck,
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India. 2,3-butanedionemonoxime monohydrazone was prepared by literature method [44].
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4.1.2. Physical measurements
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An electro-thermal digital melting point apparatus (SUMSIM, India make) was used to know the
228
melting points and were uncorrected. C, H and N microanalyses were carried out on a Perkin-
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Elmer 2400II elemental analyzer. FT-IR spectra (KBr pellet) of LH and 1 were recorded with a
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Nicolet Magna-IR spectrophotometer (Series II). The presence of ν(Pd–Cl), ν(Pd–N) and ν(Pd–
231
O) vibrations in 1 have been confirmed with the help of a Raman spectrometer (WITEC alpha,
232
300 R) in the range 2000–200 cm–1 (wave length 532 nm). Electronic spectra (UV-visible) of
233
both the ligand and complex were recorded on a Shimadzu UV-160A spectrophotometer in
234
DMSO and DMF solvents. 300 MHz 1H NMR spectra for both the ligand and complex and 13C
235
NMR spectrum only for ligand were recorded on a Bruker DPX300 spectrometer using DMSO-
236
d6 as a solvent. 500 MHz
237
DPX500 spectrometer. All chemical shifts (δ) are given in parts per million (ppm) relative to
238
TMS, used as an internal reference. Molar electrical conductivities of 1 in DMSO and DMF were
239
measured at room temperature on a Mettler Toledo FiveEasy Plus Conductivity meter. Prior to
240
each acquisition, the cell constant was calibrated with 0.1 M aqueous KCl solution. Magnetic
241
susceptibility measurements were done at 300 K with a PAR 155 vibrating sample magnetometer
242
fitted with a walker scientific L75FBAL magnet. Hg [Co(SCN)4] was used as the calibrant. The
243
ESI-Mass spectrum of LH in the positive ionization mode was recorded in acetonitrile (MeCN)
244
solution on a Xevo G2 Q-Tof Micromass spectrometer. A JEOL Mass spectrometer (Model:
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JMS-700), Japan was employed to record the FAB-Mass spectrum of 1 in the positive ionization
246
mode in m-nitrobenzyl alcohol matrix. Electrochemical cyclic voltammetric (CV) experiments
247
were done with a Bioanalytical Systems Inc. Epsilon electrochemical workstation (Model: CV-
248
50) on a C3 cell stand at 298 K under dry and degassed DMF which contained 1.0 mM of analyte
249
and 0.10 M tetra-n-butyl ammonium perchlorate (TBAP) as supporting electrolyte. The base
250
electrolyte was saturated with nitrogen for 15 min prior to each acquisition. A blanket of nitrogen
251
gas was maintained throughout the measurements. The measurements were carried out with a
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three-electrode assembly consisting of a Glassy Carbon (GC) working electrode, a platinum wire
C NMR spectrum (in DMSO-d6) of 1 was recorded on a Bruker
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counter electrode and a Ag/AgCl reference electrode. The working electrode was polished before
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each experiment with alumina slurry. All potentials reported herein are referenced to Ag/AgCl.
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Scanning Electron Microscopy was done in a JEOL JSM 6360 to find out the mode of action of
256
compound 1.
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4.1.3. Preparation of 3-[(5-bromo-2-hydroxy-benzylidene)-hydrazono]-butan-2-one oxime, LH
258
2,3-butanedionemonoxime monohydrazone (0.230 g, 2.0 mmol) was dissolved in 25 mL of
259
tetrahydrofuran to get a colorless solution. To it, 5-bromosalicylaldehyde (0.402 g, 2.0 mmol)
260
was added all at a time. The resulting colorless solution was heated under reflux for 5 h. A dark
261
yellow solution was obtained after refluxing. It was left in the air for slow evaporation. After 48
262
h, yellow amorphous compound separated thereby. It was filtered, washed thoroughly with
263
diethyl ether and subsequently dried in a vacuum desiccator over fused CaCl2.
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Yield: 0.465 g (80%). m.p. 180–182 °C. C11H12BrN3O2 (298.125): Anal. Calc. (%) for
265
C11H12BrN3O2: C, 44.31; H, 4.05; N, 14.09; Found (%): C, 44.51; H, 3.94; N, 13.92. FT-IR
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(KBr): ν [cm–1] = 3253(vb) [ν(OH)], 1612(s) [ν(C=N)], 1547(s) [ν(C=C) of phenyl ring
267
skeletal)], 1477(s) [ν(C=N) of oxime)], 1271(s) [ν(C–O) of phenolic –OH)], 1184(s) [ν(N–O)].
268
1
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to phenyl ring), 11.30 (1H, s, hydrogen of –OH group attached to oxime moiety), 8.65 (1H, s,
270
methylene proton of the –CH=N group attached to phenyl ring), 7.89 (1H, s, ortho-hydrogen of
271
the phenyl group), 7.53 (1H, d, para-hydrogen of phenyl group), 6.95 (1H, d, meta-hydrogen of
272
phenyl group), 2.20 (3H, s, methyl protons near to hydrazono moiety), 2.04 (3H, s, methyl
273
protons close to oxime moiety).
274
158.4(C10), 158.1(C8), 155.2(C1), 135.5(C3), 132.6(C5), 121.2(C6), 119.2(C2), 111.0(C4),
275
13.4(C9), 9.8(C11). UV-Vis (DMSO): λmax[nm] (εmax[M–1cm–1]) = 348 (1.105 × 104), 291 (2.036
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AC C
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H NMR (300 MHz, DMSO-d6, TMS) δ [ppm]: 11.97 (1H, s, hydrogen of –OH group attached
13
C NMR (76 MHz, DMSO-d6, TMS) δ [ppm]: 164.8(C7),
12
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× 104). UV-Vis (DMF): λmax[nm] (εmax[M–1cm–1]) = 347 (1.295 × 104), 291 (2.987 × 104). ESI-
277
MS (positive ion mode in MeCN) (m/z): 298.022 (Calc. 298.229) (79Br) and 300.019 (Calc.
278
300.229) (81Br) for [LH+H]+.
279
4.1.4. Preparation of [Pd(L)(Cl)](1)
280
0.030 g (0.1 mmol) of ligand (LH) was dissolved in 10 mL of THF to get a light yellow solution.
281
Na2[PdCl4] (0.030 g, 0.1 mmol), dissolved in 10 mL of THF, was added dropwise to the ligand
282
solution with constant stirring at room temperature. The stirring was continued for 3 h. A dark
283
orange yellow reaction mixture was obtained thereafter. It was kept in a refrigerator for slow
284
evaporation. After 10 days shining, air-stable, diamond-shaped orange single crystals were
285
deposited. These were filtered, washed thoroughly with diethyl ether and dried in a vacuum
286
desiccator over fused CaCl2. Some of the orange crystals were fit for X-ray structure
287
determination.
288
Yield: 0.016 g (36 %). m. p. > 240 °C. C11H11BrClN3O2Pd (438.99): Anal. Calc. (%) for
289
C11H11BrClN3O2Pd: C, 30.10; H, 2.53; N, 9.57; Found (%): C, 30.23; H, 2.42; N, 9.44 %. FT-IR
290
(KBr): ν [cm–1]: 3070 (br) [ν(OH)], 1624(vs) [ν(C=N) of imine], 1564(s) [ν(C=C) of phenyl ring
291
skeletal)], 1475(s) [ν(C=N) of oxime)], 1267(s) [ν(C–O) of phenolic –OH)], 1182(s) [ν(N–O)],
292
445(s) [ν(Pd–N)]. Raman: ν [cm–1]: 1260 [ν(C–O) of phenolic –OH)], 647 [ν(Pd–O)], 440 [ν(Pd–
293
N)] , 300 [ν(Pd–Cl)]. 1H NMR (300 MHz, DMSO-d6, TMS) δ [ppm]: 11.15 (1H, s, hydrogen of
294
–OH group attached to oxime moiety), 8.95 (1H, s, methylene proton of the –CH=N group
295
attached to phenyl ring), 7.91 (1H, s, ortho-hydrogen of the phenyl group), 7.57 (1H, d, para-
296
hydrogen of phenyl group), 6.98 (1H, d, meta-hydrogen of phenyl group), 1.24 (3H, s, methyl
297
protons near to hydrazono moiety), 1.14 (3H, s, methyl protons close to oxime moiety).
298
NMR (126 MHz, DMSO-d6, TMS) δ [ppm]: 160.6(C7), 157.5(C10, C8), 151.0(C1), 135.3(C3),
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13
C
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131.4(C5), 120.4(C6), 118.8(C2), 110.4(C4), 13.0(C9), 11.8(C11). UV-Vis (DMSO): λmax[nm]
300
(εmax[M–1cm–1]) = 440 (9.854 × 102), 365 (1.184 × 104), 291 (1.666 × 104). UV-Vis (DMF):
301
λmax[nm] (εmax[M–1cm–1]) = 443 (9.858 × 102), 377 (1.399 × 104), 296 (3.294 × 104). FAB-MS
302
(positive ion mode in m-nitrobenzyl alcohol matrix) (m/z): 402.1 (Calc. 402.63) (79Br) and 404.1
303
(Calc. 404.63) (81Br) for [Pd(L)]+. ΛM (DMSO): 10 Ω–1 cm2 mol–1 (Non-conducting). ΛM (DMF):
304
57 Ω–1 cm2 mol–1 (Non-conducting). μ = Diamagnetic.
305
4.1.5. Crystal structure determination
306
An appropriate single crystal of 1, suitable for X-ray crystallography, was selected following
307
examination under a microscope. Intensity data were collected at 298(2) K using a graphite
308
monochromated Mo-Kα radiation (λ = 0.71073 Å) on a Bruker Smart Apex II diffractometer
309
equipped with 1K CCD instrument. SMART software [45] was used to determine cell
310
parameters. Using SAINTPLUS [45], data reduction and corrections were made. Empirical
311
absorption corrections were performed with SADBAS program [46]. The structure was solved
312
by direct methods with the program SHELXS-97 and was refined with SHELXL-97 [46] by full-
313
matrix least-squares methods on all F2 data. All non-hydrogen atoms were refined
314
anisotropically. Hydrogen atoms attached with C, N and O atoms were placed in calculated
315
positions and were constrained to ride on their parent atoms. On the basis of observed reflections
316
and variable parameters, the final cycle of full-matrix least-squares refinement was executed.
317
4.2. Studies of antimicrobial activity
318
In vitro testing was undertaken to examine the antimicrobial activity of both LH and 1.
319
4.2.1. Microbial strains
320
The in vitro antimicrobial activity of 1 was evaluated against 15 different pathogenic bacteria
321
belonging to Gram-negative genera like Escherichia coli 25938, Escherichia coli 397,
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Salmonella typhi 62, Vibrio cholerae 20, Shigella dysenteriae 1, Shigella dysenteriae 37,
323
Salmonella typhimurium NTCC 74, Pseudomonas aeruginosa 25619 and Gram-positive genera
324
like Staphylococcus aureus 29737, Bacillus cereus 11778, Bacillus subtilis 6633, Streptococcus
325
epidermidis 12228,
326
bronchiseptica 4617 respectively. All these microbial strains were collected from Division of
327
Microbiology, Department of Pharmaceutical Technology, Jadavpur University, Kolkata, India.
328
4.2.2. Preparation of Inoculums
329
The bacterial strains were grown in nutrient broth (HI-MEDIA) at 37 0C for 24 h. Then the
330
strains were adjusted to 0.5 McFarland standards [47] and diluted with sterile normal saline to
331
give initial bacterial suspension of 2×106 CFU/mL.
332
4.2.3. Reagent, chemicals and culture media
333
Amoxicillin (Sigma-Aldrich) was used as reference antibiotics for the evaluation of bacterial
334
inhibition. All the media used in antimicrobial study were purchased from Oxoid Ltd., UK.
335
4.2.4. Determination of MIC
336
The minimum inhibitory concentration (MIC) of the test compound was determined by agar
337
dilution method following NCCLS 2006 protocol [48]. Briefly, the compound was first dissolved
338
in 2% DMSO and then sterile water was added to give rise a final concentration of 1000 μg/mL.
339
This mother solution was then serially diluted to obtain different concentrations — 10, 25, 50,
340
100, 200 and 400 μg/mL. Preliminary analysis revealed that any inhibition on tested
341
microorganism was not shown up to 4% DMSO (v/v). Standardized inoculum (2×106 CFU/mL)
342
was spotted on the agar plates and incubated at 37 °C for 24 h. Amoxicillin was used as a
343
reference standard antibiotic. Lowest concentration which completely inhibited visible growth of
344
microorganism was recorded as MIC [49].
Bordetella
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Micrococcus luteus 10240, Bacillus pumilus 14884,
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4.2.5. Antimicrobial assay
346
Antimicrobial activity was determined by Well-diffusion method according to the NCCLS 2004
347
guidelines [50]. Nutrient Agar (NA) plates were prepared in 90 mm sterile Petri dishes (20 mL
348
NA) and 100 μL bacterial cell suspension (2×106 CFU/mL) was spread on solid NA plates using
349
spread plate techniques. Wells were dug by sterile borer and appropriate concentration of drug
350
was added to the wells. After a while, all the plates were incubated at 37 0C for 24 h. Amoxicillin
351
was used as a reference standard to compare antimicrobial activity of the test sample against the
352
tested microorganisms. The sensitivity was measured by clear inhibitory zones (mm) formed
353
around the wells. Each experiment was repeated in triplicate and the average of three trials were
354
reported [23,51].
355
4.2.6. Growth kinetic studies
356
Time-kill assay was performed in Mueller-Hinton Broth (MHB) containing the test sample.
357
Briefly, tubes containing liquid broth with and without (growth control) test drug were seeded
358
with 1 mL of log-phase inoculum (2×106 CFU/mL) to a final volume of 10 mL. These were
359
incubated at 37 °C. At different time intervals (h) after inoculation, a 0.1 mL volume was taken
360
from each tube and was subjected to 1:10 dilution. An aliquot of 10 μL of each dilution was
361
taken and plated on NA plates. Then the plates were incubated at 37 °C for 24–48 h and the
362
numbers of colony forming units (CFU) were counted. By using the viable counts at each
363
interval, a 24 h time-kill curve was recorded for each. The experiment was performed in
364
duplicate. The log CFU/mL was plotted on a graph as a function of time. It was used to compare
365
the rate and extent of killing-effect in various concentrations of the test drug [52,53].
366
4.2.7. Scanning Electron Microscopy
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Scanning electron microscopic observations were carried out on bacterial cells after 16 h
368
incubation in MHB at 37 0C. Test compound at 2 fold of MIC concentration was added to the
369
cell and was incubated for 24 h at 37 0C. A 24 h incubated normal culture was used as a control.
370
After that, bacterial cells were fixed with 4% glutaraldehyde in 0.1 M phosphate buffer for 2 h at
371
room temperature and then were washed four times in 0.1 M phosphate buffer (pH 7.2–7.4).
372
They were dehydrated in a graded alcohol series and were mounted onto stubs using double-
373
sided carbon tape. Then these were coated with a thin layer of gold using a microscopic sputter
374
coater for 1 min at 20 mA and were observed under the scanning electron microscope [54,55].
375
Acknowledgements
376
Financial support [F. No. 41–220/2012 (SR)] received from the University Grants Commission
377
(UGC), New Delhi, India, is thankfully acknowledged. M.L.Z and L.P.L. gratefully appreciates
378
the NSFC (Grant Nos. 21171109 & 21271121), SRFDP (Grant Nos. 20111401110002 &
379
20121401110005), the Natural Science Foundation of Shanxi Province of China (Grant Nos.
380
2010011011–2 & 2011011009–1) and the Shanxi Scholarship Council of China (2012–004 &
381
2013–026). Special thanks are also due to the revered reviewers for their valuable and
382
constructive suggestions during revision.
383
Appendix A. Supplementary data
384
CCDC 960808 contains the supplementary crystallographic data for 1. These data can be
385
retrieved free of charge via http://www.ccdc.cam.ac.uk/conts/retrieving.html, or from the
386
Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK; fax: (+44)
387
1223–336–033; or e–mail: [email protected].
388
References
389
[1]
AC C
EP
TE D
M AN U
SC
RI PT
367
D. Gaynor, D.M. Griffith, J. Chem. Soc., Dalton Trans. 41 (2012) 13239–13257.
17
ACCEPTED MANUSCRIPT
390
[2]
C. Orvig, M.J. Abrams, Chem. Rev. 99 (1999) 2201–2204.
391
[3]
B.T. Khan, J. Bhatt, K. Najmuddin, S. Shamsuddin, K. Annapoorna, J. Inorg. Biochem.
393
44 (1991) 55–63. [4]
394
J. Zhang, L. Li, L. Ma, F. Zhang, Z. Zhang, S. Wang, Eur. J. Med. Chem. 46 (2011)
RI PT
392
5711–5716. [5]
C. Rîmbu, R. Danac, A. Pui, Chem. Pharm. Bull. 62 (1) (2014) 12–15.
396
[6]
D.R. Ilić, V.V. Jevtić, G.P. Radić, K. Arsikin, B. Ristić, L. Harhaji-Trajković, N.
SC
395
Vuković, S. Sukdolak, O. Klisurić, V. Trajković, S.R. Trifunović, Eur. J. Med. Chem. 74
398
(2014) 502–508. [7]
400 401
(2014) 93–98. [8]
402 403
M. Kalita, P. Gogoi, P. Barman, B. Sarma, A.K. Buragohain, R.D. Kalita, Polyhedron 74
N.M. Aghatabay, M. Somer, M. Senel, B. Dulger, F. Gucin, Eur. J. Med. Chem. 42 (2007) 1069–1075.
[9]
404
TE D
399
M AN U
397
J. Traeger, T. Klamroth, A. Kelling, S. Lubahn, E. Cleve, W. Mickler, M. Heydenreich, H. Müller, H. Holdt, Eur. J. Inorg. Chem. (2012) 2341–2352.
[10]
A. Garoufis, S.K. Hadjikakou, N. Hadjiliadis, Coord. Chem. Rev. 253 (2009) 1384–1397.
406
[11]
M. Jagadeesh, H.K. Rashmi, Y.S. Rao, A.S. Reddy, B. Prathima, P.U.M. Devi, A.V.
408
Reddy, Spectrochim. Acta A 115 (2013) 583–587.
[12]
409 410
A. Karaküçük–İyidoğan, D. Taşdemir, E.E. Oruç–Emre, J. Balzarini, Eur. J. Med. Chem. 46 (2011) 5616–5624.
[13]
411 412
AC C
407
EP
405
F. Shaheen, A. Badshah, M. Gielen, C. Gieck, M. Jamil, D. de Vos, J. Organomet. Chem. 693 (2008) 1117–1126.
[14]
P.F. Salas, C. Herrmann, C. Orvig, Chem. Rev. 113 (2013) 3450–3492.
18
ACCEPTED MANUSCRIPT
[15]
414
Siddiq, A. Hameed, S.Z. Haider, Inorg. Chim. Acta 363 (2010) 3261–3269. [16]
416 417
V.V. Padma, R. Renganathan, K. Natarajan, Metallomics 4 (2012) 101–113. [17]
G.P. Vasić, V.V. Glodjović, I.D. Radojević, O.D. Stefanović, L.R. Čomić, V.M. Djinović, S.R. Trifunović, Inorg. Chim. Acta 363 (2010) 3606–3610.
418 419
P. Kalaivani, R. Prabhakaran, F. Dallemer, P. Poornima, E. Vaishnavi, E. Ramachandran,
RI PT
415
S. Nadeem, M. Bolte, S. Ahmad, T. Fazeelat, S.A. Tirmizi, M.K. Rauf, S.A. Sattar, S.
[18]
P. Kalaivani, R. Prabhakaran, E. Ramachandran, F. Dallemer, G. Paramaguru, R.
SC
413
Renganathan, P. Poornima, V.V. Padma, K. Natarajan, J. Chem. Soc., Dalton Trans. 41
421
(2012) 2486–2499.
M AN U
420
422
[19]
A.K. Mishra, N.K. Kaushik, Eur. J. Med. Chem. 42 (2007) 1239–1246.
423
[20]
M.N. Patel, P.A. Dosi, B.S. Bhatt, Inorg. Chem. Commun. 21 (2012) 61–64.
424
[21]
J. Chakraborty, H. Mayer–Figge, W.S. Sheldrick, P. Banerjee, Polyhedron 25 (2006)
[22]
428
A. Valkonen, K. Rissanen, S.R. Trifunović, Polyhedron 31 (2012) 69–76. [23]
429 430
(2013) 211–220. [24]
431
L. Zhao, X.T. Dong, Q. Cheng, J.X. Zhao, L. Wang, Synth. React. Inorg. Met–Org. Nano–Met. Chem. 43 (2013) 1241–1246.
432
[25]
433
[26]
434
C. Biswas, M. Zhu, L. Lu, S. Kaity, M. Das, A. Samanta, J.P. Naskar, Polyhedron 56
EP
427
G.P. Radić, V.V. Glođović, I.D. Radojević, O.D. Stefanović, L.R. Čomić, Z.R. Ratković,
AC C
426
3138–3144.
TE D
425
M. Asadi, K.A. Jamshid , A.H. Kyanfar, Inorg. Chim. Acta 360 (2007) 1725–1730. M. Espinal, J. Pons, J. García–antón, X. Solans, M. Font–bardia, J. Ros, Inorg. Chem. Commun. 12 (2009) 368–370.
19
ACCEPTED MANUSCRIPT
435
[27]
436 437
A. Castiñeiras, N. Fernández–Hermida, I. García–Santos, L. Gómez–Rodríguez, J. Chem. Soc., Dalton Trans. 41 (2012) 13486–13495.
[28]
K. Nakamato, Infrared and Raman Spectra of Inorganic and Coordination Compounds, Part B: Application in Coordination, Organometallic and Bioinorganic Chemistry, 5th
439
ed., Wiley–Interscience, New York, 1997.
RI PT
438
[29]
G.W. Simmons, Y.N. Wang, J. Marcos, K. Klier, J. Phys. Chem. 95 (1991) 4522–4528.
441
[30]
M. Hakimi, Z. Mardani, K. Moeini, F. Mohr, M.A. Fernandes, Polyhedron 67 (2014) 27–
442
35.
SC
440
[31]
W.J. Geary, Coord. Chem. Rev. 7 (1971) 81–122.
444
[32]
T. Motoyama, Y. Shimazaki, T. Yajima, Y. Nakabayashi, Y. Naruta, O. Yamauchi, J.
445
Am. Chem. Soc. 126 (2004) 7378–7385. [33]
447
A.W. Addison, T.N. Rao, J. Reedijk, J. van Rijn, G.C. Verschoor, J. Chem. Soc., Dalton Trans. 7 (1984) 1349–1356.
TE D
446
M AN U
443
448
[34]
L. Yang, D.R. Powell, R.P. Houser, J. Chem. Soc., Dalton Trans. 9 (2007) 955–964.
449
[35]
R. Greef, R. Peat, L.M. Peter, D. Pletcher, J. Robinson, Instrumental Methods in
[36]
452 453
Polyhedron 16 (1997) 3625–3633.
[37]
454 455 456
D. Kovala–Demertzi, A. Domopoulou, M.A. Demertzis, A. Papageorgiou, D.X. West,
AC C
451
Electrochemistry, Ellis Horwood Limited, England, 1985, pp. 188–195.
EP
450
K. Todar, Pathogenic E. coli. Online Textbook of Bacteriology, University of Wisconsin– Madison, Department of Bacteriology.
[38]
S. Shujha, A. Shah, Z. Rehman, N. Muhammad, S. Ali, R. Qureshi, N. Khalid, A. Meetsma, Eur. J. Med. Chem. 45 (2010) 2902–2911.
20
ACCEPTED MANUSCRIPT
457
[39]
458
A.V. Krishnaraju, T.V.N. Rao, D. Sundararaju, M. Vanisree, H.S. Tsay, G.V. Subbaraju, Int. J. Appl. Sci. Eng. 4 (2006) 115–125.
[40]
B.G. Tweedy, Phytopathology 55 (1964) 910–914.
460
[41]
R. Prabhakaran, A. Geetha, M. Thilagavathi, R. Karvembu, V. Krishnan, H. Bertagnolli,
[42]
463
Spectrochim. Acta A 107 (2013) 108–116. [43]
465 466
Rehim, RSC Adv. 4 (2014) 24131–24141. [44]
J.P. Naskar, B. Guhathakurtha, L. Lu, M. Zhu, R. Bhattacharya, J. Ind. Chem. Soc. 90 (2013) 1301–1309.
467 468
F.A. Bassyouni, S.M. Abu–Baker, K. Mahmoud, M. Moharam, S.S. El–Nakkady, M.A.
[45]
469
M AN U
464
K.S. Prasad, L.S. Kumar, S. Chandan, R.M.N. Kumar, H.D. Ravanasiddappa,
Bruker, SMART (Version 5.0) and SAINT (Version 6.02), Bruker AXS Inc., Madison, Wisconsin, USA, 2000.
TE D
462
K. Natarajan, J. Inorg. Biochem. 98 (2004) 2131–2140.
SC
461
RI PT
459
[46]
G.M. Sheldrick, Acta Crystallogr. A 64 (2008) 112–122.
471
[47]
J. McFarland, J. Am. Med. Assoc. 14 (1907) 1176–1178.
472
[48]
National Committee for Clinical Laboratory Standards, approved standard M2–A9,
474
NCCLS, Wayne, PA; 9th Edn. USA, 2006. [49]
475
Ngadjui, B.M. Abegaz, BMC Complementary and Alternative Medicine 11 (2011) 42 (1–
476 477 478
V. Kuete, P.Y. Ango, G.W. Fotso, G.D.W.F. Kapche, J.P. Dzoyem, A.G. Wouking, B.T.
AC C
473
EP
470
5).
[50]
National Committee for Clinical Laboratory Standards, approved standard M44–A. NCCLS, Wayne, PA, USA, 2004.
21
ACCEPTED MANUSCRIPT
[51]
480 481
Spiteller, J. Natural Products 74 (2011) 272–278. [52]
482 483
M.D. Awouafack, P. Spiteller, M. Lamshöft, S. Kusari, B. Ivanova, P. Tane, M.
C. Bantar, C. Schell, G. Posse, A. Limansky, V. Ballerini, L. Mobilia, Diagn. Microbiol. Infect. Dis. 61 (2008) 309–314.
[53]
484
RI PT
479
E.E. Roling, E.M. Klepser, A. Wasson, R.E. Lewis, E.J. Ernst, M.A. Pfaller, Diagn. Microbiol. Infect. Dis. 43 (2002) 13–17.
[54]
I. Kaya, N. Yigit, M. Benli, Afr. J. Trad. CAM 5 (4) (2008) 363–369.
486
[55]
S. Kaneshima, S. Koga, K. Tanaka, J. Electron Microsc 26 (1977) 355–358.
SC
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M AN U
487 488 489 490
494 495 496 497 498 499
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500 501 502 22
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List of the Captions
504
Figure Captions
505
Fig. 1. Synthetic scheme of LH and 1
506
Fig. 2. The structure of [Pd(L)(Cl)] (1) with ellipsoids at 30% probability and H-bonding motif.
507
Fig. 3. Minimum inhibitory concentration (MIC) of 1.
508
Fig. 4. Zone of inhibition of 1.
509
Fig. 5. Scanning electron micrographs of Bacillus cereus 11778 and Pseudomonas aeruginosa
510
25619 (A1, B1) before and (A2, B2) after treatment with compound 1.
511
Table Captions
512
Table 1 Crystal data and structure refinement for the compound, 1
513
Table 2 Hydrogen bonds (Å and o) for 1
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Table 1
Compound
1
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Crystal data and structure refinement for the compound, 1
960808
Color/shape
Orange/Diamond
SC
CCDC No.
Empirical formula
C11H11BrClN3O2Pd 438.99
M AN U
Formula weight Temperature [K]
298(2)
Wavelength [Å]
0.71073
Crystal system
Triclinic
P–1
a [Å] b [Å] c [Å]
AC C
β [o]
EP
Unit cell dimensions
TE D
Space group
7.1744(1) 8.7770(2) 11.2552(2) 79.4400(10)
Volume [Å3]
672.93(2)
Z
2
ρCalcd [g/cm3]
2.167
Absorption coefficient [mm–1]
4.546
F(000)
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0.35 × 0.25 × 0.2
range for data collection [deg]
1.84 to 25.05
Limiting indices
–8
S0223-5234(14)00962-3
DOI:
10.1016/j.ejmech.2014.10.035
Reference:
EJMECH 7442
To appear in:
European Journal of Medicinal Chemistry
Received Date: 10 July 2014 Revised Date:
3 September 2014
Accepted Date: 12 October 2014
Please cite this article as: N. Bandyopadhyay, M. Zhu, L. Lu, D. Mitra, M. Das, P. Das, A. Samanta, J.P. Naskar, Synthesis, structure, spectral characterization, electrochemistry and evaluation of antibacterial potentiality of a novel oxime-based palladium(II) compound, European Journal of Medicinal Chemistry (2014), doi: 10.1016/j.ejmech.2014.10.035. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
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Graphical abstract Synthesis, structure, spectral characterization, electrochemistry and evaluation of
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antibacterial potentiality of a novel oxime-based palladium(II) compound Nirmalya Bandyopadhyaya, Miaoli Zhub, Liping Lub, Debmalya Mitrac, Mousumi Dasc, Piu
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Dasc, Amalesh Samantac, Jnan Prakash Naskara*
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Highlights Synthesis, structure, spectral characterization, electrochemistry and evaluation of
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antibacterial potentiality of a novel oxime-based palladium(II) compound Nirmalya Bandyopadhyaya, Miaoli Zhub, Liping Lub, Debmalya Mitrac, Mousumi Dasc, Piu Dasc, Amalesh Samantac, Jnan Prakash Naskara*
Synthesis of oxime-based ligand and its Pd(II) complex.
Spectroscopic aspects and X-ray single crystal structure of the complex.
Quasi-reversible Pd(II)/Pd(I) reduction couple.
This complex exhibits satisfactory bactericidal as well as bacteriostatic activity.
The bioactivity was substantiated by SEM study.
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Synthesis, structure, spectral characterization, electrochemistry and evaluation of
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antibacterial potentiality of a novel oxime-based palladium(II) compound
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Nirmalya Bandyopadhyaya, Miaoli Zhub, Liping Lub, Debmalya Mitrac, Mousumi Dasc, Piu
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Dasc, Amalesh Samantac, Jnan Prakash Naskara*
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a
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India.
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Fax: +(91)(33)2414 6223 E-mail: [email protected]
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b
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Engineering of the Education Ministry, Shanxi University, 92 Wucheng Road, Taiyuan, Shanxi
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Department of Chemistry, Inorganic Chemistry Section, Jadavpur University, Kolkata 700 032,
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Institute of Molecular Science, Key Laboratory of Chemical Biology and Molecular
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030006, People's Republic of China.
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c
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Kolkata 700 032, India.
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E-mail: [email protected]
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Abstract
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The title monomeric Pd(II) compound, [Pd(L)(Cl)], was synthesized in moderate yield out of the
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reaction of equimolar proportion of Na2[PdCl4] and 3-[(5-bromo-2-hydroxy-benzylidene)-
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hydrazono]-butan-2-one oxime (LH) in tetrahydrofuran milieu. LH is a 1:1 Schiff-base
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condensate
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[Pd(L)(Cl)] has been characterized by C, H and N microanalyses, 1H and
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FT-IR, Raman spectra, UV-Vis spectra and molar electrical conductivity measurements.
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[Pd(L)(Cl)] is diamagnetic. Structural elucidation reveals that the palladium center in [Pd(L)(Cl)]
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is nested in ‘N2OCl’ coordination environment. The geometry around Pd in [Pd(L)(Cl)] is
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distorted square-planar. The redox behavior of [Pd(L)(Cl)] in DMF shows a reduction couple,
2,3-butanedionemonoxime
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monohydrazone
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5-bromosalicylaldehyde. 13
C NMR, FAB-MS,
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Pd(II)/ Pd(I) at –0.836 V versus Ag/AgCl. The in vitro antimicrobial activity of [Pd(L)(Cl)] was
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screened against both Gram-positive and Gram-negative human pathogenic bacteria. This
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bioactivity was substantiated with SEM study. [Pd(L)(Cl)] exhibits satisfactory bactericidal as
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well as bacteriostatic activity.
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Key words: Oxime; Palladium; Crystal structure; Redox; Antibacterial; SEM study.
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30 1. Introduction
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Synthesis, characterization and evaluation of biological activities of novel metal-based
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compounds are a rapidly growing area of contemporary research [1–3]. Drugs based on transition
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metal compounds are superior to organic-based drugs so much so that medicinal inorganic
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chemistry now is an emerging area of research [4]. Again, bacterial drug resistance potentiality is
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increasing alarmingly [5]. Consequently, development of new therapeutic agents to combat
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bacterial infection warrants current attention [6,7]. Synthesis of novel antibacterial drugs with
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properly known structure is important to understand their behavior in biological environment and
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subsequent possible mode of action. The transition metal compounds of hetero donor ligands
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show promising biological activities with reduced toxicity [8]. Palladium, an important platinum
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group metal [9], is noteworthy in this perspective. The compounds of palladium manifest
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diverse biological activities [10]. Palladium-based compounds are well-known antitumor [11],
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antiviral [12], antifungal [13], antiinflammatory [13], antimalarial [14] and antibacterial [15]
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agents. The intense and diverse bioactivities of various palladium compounds on bacterial strains
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are well-documented in the literature [10,15,16]. Pd(II) being a soft Lewis acid has an inherent
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tendency to bind soft donor centers like N, S in its d8 square-planar structural disposition [17].
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Accordingly, most of the antibacterial palladium(II) compounds are from N- and S-based ligands
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like thiosemicarbazones [16–18], thiocarbazates-thioamides [10], thiodiamines [19] etc.
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Palladium(II) compounds, synthesized from ligands with N donor only, are also known to
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exhibit antimicrobial activities against Gram-negative strains like E. coli, P. aeruginosa, S.
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marcescens along with Gram-positive strains like S. aureus, B. subtilis [20]. Thus it seems that
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for showing bioactivity, palladium complexes of N, S donor ligands are of obvious choice.
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Examples are also known from ligands having N, O donor ligands [21]. Palladium complexes
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from the alkyl derivative of thiosalicylic acids having S, O donor sides also exhibit moderate
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antimicrobial activity [22]. On the contrary, such type of bioactivity of monomeric Pd(II)
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compound in a N, O donor oxime-based Schiff-base ligand is rare indeed [23]. For quite some
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time, we are interested to develop novel oxime-based Pd(II) compounds as potential antibacterial
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agents with pronounced activity. Herein we wish to report our endeavor along that line – the
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synthesis, spectroscopic aspects, X-ray structure and electrochemical behavior of a novel Pd(II)
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complex. The antibacterial activities of the ligand, 3-[(5-bromo-2-hydroxy-benzylidene)-
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hydrazono]-butan-2-one oxime (LH) and its palladium(II) compound, [Pd(L)(Cl)], was screened
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against a good number of human pathogenic bacterial cell lines. The bioactivity was also
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substantiated with Scanning Electron Microscopic (SEM) studies in proper cases.
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2. Results and discussion
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2.1. Chemistry
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2.1.1. Synthesis and characterization
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A Schiff-base ligand, 3-[(5-bromo-2-hydroxy-benzylidene)-hydrazono]-butan-2-one oxime
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(LH), has been used for the present study. The ligand has been prepared by the Schiff-base
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condensation of equimolar proportion of 2,3-butanedionemonoxime monohydrazone and 5-
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bromosalicylaldehyde in tetrahydrofuran. LH was spectroscopically characterized by 1H,
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NMR and ESI-MS (Supporting Information, Figs. S1–S3). Subsequent reaction in 1:1 molar
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proportion of LH and Na2[PdCl4] in tetrahydrofuran enables us to isolate the orange monomeric
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Pd(II) compound, [Pd(L)(Cl)] (1) in moderate yield. FAB-MS peaks in m-nitrobenzyl alcohol
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matrix at 402.1 (for
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Information, Fig. S4). This is a testimony to the formation of 1. C, H and N micro analytical data
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of the single crystals of 1 also substantiate the above formulation. In Fig. 1, the synthetic scheme
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of LH along with 1 is outlined.
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The diamagnetic nature of the compound suggests a square-planar disposition of the palladium
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center in 1. This diamagnetic nature enables us to detect all the carbon nuclei present in 1
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(Supporting Information, Fig. S5). 1H NMR spectral data of 1 were also recorded in DMSO-d6
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for the evaluation of the number and nature of all the protons relative to TMS with chemical
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shifts given in ppm (Supporting Information, Fig. S6).
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The IR spectrum of 1 (Supporting Information, Fig. S7) is consistent with the structure
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confirmed by single crystal X-ray diffraction studies. Free LH have infrared spectral bands at ν
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1612(s) [ν(C=N) of imine], 1477(s) [ν(C=N) of oxime)] and 1184(s) [ν(N–O)]. In 1, these bands
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appear respectively at 1624, 1475 and 1182 cm–1. The strong band at 1612 cm–1 is assigned to
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the C=N stretching vibration, characteristic for a Schiff-base ligand. The ring skeletal vibration
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(C=C) of free ligand was assigned at 1547 cm–1. In 1, this band undergoes a shift towards higher
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wavenumber by 17 cm–1 and appears at 1564 cm–1. The Ar–O stretching frequency [ν(C–O) of
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phenolic –OH] appears as a strong band at 1271 cm–1 in the free ligand, and in 1 it appears at
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1267 cm–1. Thus, the lower wavenumber shift of the Ar–O stretching by ca. 4 cm–1 indicates that
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Pd–O (phenolic) bond is formed [24,25]. The Pd–Cl stretching frequency, [ν(Pd–Cl)] is
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Br) correspond to [Pd(L)]+ for 1 (Supporting
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generally observed in the range 300–400 cm–1 [26]. Here we could not observe it due to the
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limitation of the window (4000–400 cm–1) of the spectrometer used. The ν(Pd–N) modes can be
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identified at 445 cm–1 [27] in the FT-IR of 1. However, we have discerned the ν(Pd–Cl), ν(Pd–N)
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and ν(Pd–O) vibrations in the Raman spectrum of 1, shown in Fig. S8 (Supporting Information).
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The Raman shift values at 300, 440 and 647 cm–1 are assigned to the ν(Pd–Cl), ν(Pd–N) and
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ν(Pd–O) vibrations respectively [28–30].
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In the electronic spectrum of 1 in DMSO and DMF, only one d–d band was observed at 440 and
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443 nm respectively. The broad band in this region is assigned to the dz2 to dx2–y2 transition [20].
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This transition brings about the orange hue to the compound. This general feature pertains to the
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low-spin d8 metal ion complexes with square-planar geometry. The molar electrical
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conductivities of ~10–3 M solutions of 1 in DMF is 57 Ω–1 cm2 mol–1. This value in DMSO is
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only 10 Ω–1 cm2 mol–1.Thus 1 behaves as a non-electrolyte both in DMF and DMSO [31].
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2.1.2. Molecular structure of [Pd(L)(Cl)] (1)
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The X-ray crystal structure (Fig. 2) of 1 was determined. In Table 1, a summary of data
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collection, structure refinement for 1 is given. Selected bond lengths, bond angles and hydrogen-
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bond geometry are provided in Tables S1 (Supporting Information) and Table 2 respectively.
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The palladium center in 1 is nested in ‘N2OCl’ core. The Pd1–N1 and Pd1–N3 bond distances
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are almost the same, having the values of 1.976(5) and 1.978(5) Ǻ respectively. Pd1–O1 bond
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distance [1.963(4) Ǻ] is quite similar to Pd–N bonds. The bond distance of Pd1–Cl1 is 2.326(2)
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Ǻ. This bond is much longer than Pd–N and Pd–O bonds. These bond length values correlate
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well with those reported earlier in similar ‘N2OCl’ core for mononuclear Pd(II) compounds [32].
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The cis-angles, O1–Pd1–Cl1, N1–Pd1–N3, N3–Pd1–Cl1 and O1–Pd1–N1 are of values 84.8,
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90.2, 91.3 and 93.7° respectively. The trans-angles, O1–Pd1–N3 and N1–Pd1–Cl1 respectively
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are of 176.0 and 177.1°. Thus the range (84.8–93.7°) of the cis-angles is in copious deviation
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from 90°. Alike are the deviations of the trans-angles from 180°. Consequently, the geometry
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around the palladium(II) center in 1 experiences distortion. The palladium center is deviated by
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0.015(2) Ǻ from the average square-based basal plane defined by N1, N3, O1 and Cl1. Thus the
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core no longer retains its planarity.
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To augment the geometric shape of 1, we have taken recourse to estimate the Addison angular
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structural parameter (4) for four-coordinated complexes [33,34]. By definition, 4 = [360°–
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(α+β)] /141° where α and β respectively are the largest and second largest angles around the
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metal center. The 4 value is equal to 0 for a perfectly square-planar geometry and 1 for a regular
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tetrahedral core [34]. For 1, the angle N1–Pd1–Cl1 (177.1°) is α and O1–Pd1–N3 (176.0°) is β.
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Accordingly, the 4 value obtained here is 0.049. This value closely approaches to 0. Thus the
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geometry around the palladium center in 1 is almost square-planar. However, the sum of the
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subtended cis-angles around the palladium centre is 360°. The rationale behind this is that the Pd
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center gains some stability due to the chelate effects of two 6-membered planar rings.
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The intermolecular hydrogen bonding exclusively consists of the classical OH···Cl bond. Fig.
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S9 (Supporting Information) represents the packing diagram along with the H-bonded network in
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1.
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2.1.3. Electrochemistry
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The electrochemical aspect of 1 has been studied by CV in DMF at Glassy Carbon (GC) working
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electrode (Supporting Information, Fig. S10).
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1 exhibits one electrochemical response at E½ value of –0.737 V vs Ag/AgCl (Couple I). Under
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the same experimental condition, the ferrocene-ferrocenium (Fc/Fc+) couple appears at 0.44 V vs
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Ag/AgCl. Comparison of the cyclic voltammetric peak currents of 1 with those of the standard
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redox couple, Fc/Fc+, establishes that couple I involves only one electron. LH is
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electrochemically dormant in the potential range of interest here. Consequently, this response can
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be assigned as Pd(II) to Pd(I) reduction. Peak potential of couple I is –0.836 V with 39.80 A
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peak–current. In the reverse cycle, the subsequent oxidative response can be observed at –0.637
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V vs Ag/AgCl. The corresponding anodic peak current, ipa is 1.86 A. The ipc to ipa ratio is
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21.40, a value grossly deviated from unity. Again ipc, the cathodic peak current, increases with
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the square root of the sweep rate (ν1/2), though the increase is not in proportionality. Moreover,
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Epc, the cathodic peak potential shifts more negatively with the increase in sweep rate (ν). Thus
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the electron transfer at couple I occurs quasi-reversibly [35]. Pd(II) to Pd(I) reductions in DMF
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are known to occur at ~ –0.9 V vs Ag/AgCl [23,36].
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2.2. Antimicrobial Activity
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Various pathogenic bacteria pose serious threats to human kind. Virulent Gram-negative strains
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of E. coli can cause gastroenteritis, urinary tract infection, infection of wounds and neonatal
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meningitis; while P. aeruginosa causes infection of wounds and septicemia. Gram-positive
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strains like M. luteus can cause skin infections that produce pruritic eruptions sometimes even
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with central ulceration. Again Gram-positive B. subtilis is responsible for food poisoning [16,
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37–39]. Harmful bacteria often develop resistance to the existing antidotes. For this, novel
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compounds with pronounced cytotoxic/cytostatic effects are the need of the day. In our present
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study we have incorporated specifically the above strains also.
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2.2.1. Antimicrobial assay
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1 has moderate antimicrobial properties. LH does not have such efficacies. As shown in Table
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S2 and Table S3 (Supporting Information), the compound was screened against bacteria using
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Well-diffusion method. Among the listed organisms in Table S2, our compound 1 was active
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against 12 tested organisms (Fig. 3).
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The compound displays moderate activity with high MIC value against Salmonella typhi 62,
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Vibrio cholerae 20, Shigella dysenteriae 1, Bacillus cereus 11778 , Bacillus subtilis 6633,
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Micrococcus luteus 10240, Bordetella bronchiseptica 4617, Streptococcus epidermidis 12228,
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Shigella dysenteriae 37 (400 µg/ml; diameters of inhibition zone ranges 7.0 to 14 mm) and low
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value against Escherichia coli 25938, Salmonella typhimurium NTCC 74, Pseudomonas
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aeruginosa 25619 (200 µg/ml; inhibition zone ranges: 10 to 15 mm) (Supporting Information,
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Fig. S11).
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Effectiveness of an antibacterial drug can be judged on the basis of its zone of inhibition. The
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data obtained here shows that 1 has the ability to inhibit the metabolic growth of our tested
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organisms (Fig. 4).
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2.2.2. Growth kinetic studies
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The results of 6 selected microorganisms have been shown graphically. The graph depicts the
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respective numerical logarithmic values of number of colonies after incubation. By analysing the
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bacterial growth in Fig. S12 and Fig. S13 (Supporting Information), it is clear that compound 1
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had its effect as a bactericidal on Pseudomonas aeruginosa 25619 and Micrococcus luteus 10240
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respectively. Compound 1 had its effect as bacteriostatic on the rest of the organisms as shown in
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Figs. S14–S17 (Supporting Information). In Table S4 (Supporting Information) the respective
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data are tabulated.
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2.2.3. Scanning Electron Microscopic (SEM) observation
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Scanning electron microscopy was carried out to assess the morphological changes in bacterial
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cells following exposure to the compound. SEM studies were carried out on Bacillus cereus
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11778 and Pseudomonas aeruginosa 25619 strains due to their higher and moderate zone of
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inhibition. Untreated Bacillus cereus 11778 (Fig. 5A1) and Pseudomonas aeruginosa 25619
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(Fig. 5B1) cells had apparently shown normal rod-shape and smooth surface with no visible
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damage. After treatment with compound 1, the cell permeability of Bacillus cereus (Fig. 5A2)
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was increased leading to an accumulation of fluid, resulting in swelling and cell deformation.
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However, in case of Pseudomonas aeruginosa (Fig. 5B2), the cell walls were digested at various
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junctions leading to the destruction of cells and subsequent death.
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The possible mode of bioactivity of our palladium complex may be explained by Tweedy’s
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chelation theory [40]. Generally the polarity of any central metal ion is lessened considerably
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due to chelation. This is due to the partial sharing of metal’s positive charge with the ligand and
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possible π-electron delocalization over the chelate ring. Consequently chelation increases the
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lipophilic character of the central metal ion enabling the subsequent permeation of the compound
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through the lipid layer of the microbial cell membrane. After penetration, the possible mode of
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action involves the formation of hydrogen bond preferably through the azomethine group of the
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compound with the active sites of the microbial cell constituents. As a result, the normal
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physiological cell process of the microbes gets potential interference from the drug [41].
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From the perspective of bactericidal activity, the palladium-based metallo-drugs are generally
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more toxic towards Gram-positive strains than Gram-negative strains [42]. The reason is due to
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the inherent difference in the structure of the cell walls. The walls of Gram-negative cells are
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more complex than those of Gram-positive cells [38]. The outer cell membrane of Gram-
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negative bacteria is constituted of a complex lipopolysaccharide (LPS) that acts as an endotoxin;
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protecting these bacteria from the extraneous control agents even from lysozyme and penicillin
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[43]. Interestingly, 1 is more effective to three of our tested Gram-negative strains (Escherichia
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coli 25938, Salmonella typhimurium NTCC 74, Pseudomonas aeruginosa 25619). Again, 1 is
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moderately active to Gram-positive strains under study. Thus 1 behaves as a broad spectrum
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antibacterial agent. The result obtained here is of great significance for biological and
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pharmaceutical control applications.
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3. Conclusions
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Here we have been able to synthesize and structurally characterize a novel Pd(II) compound
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[Pd(L)(Cl)] from an oxime-based ligand, 3-[(5-bromo-2-hydroxy-benzylidene)-hydrazono]-
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butan-2-one oxime (LH). Diamagnetic [Pd(L)(Cl)] is in square-planar ‘N2OCl’ coordination
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core. Redox studies in DMF show a quasi-reversible Pd(II)/Pd(I) reduction couple at –0.836 V
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versus Ag/AgCl. [Pd(L)(Cl)] shows promising broad spectrum antimicrobial activity against a
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good number of both Gram-positive and Gram-negative human pathogenic bacteria.
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4. Experimental
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4.1. Chemistry
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4.1.1. Materials
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All chemicals and solvents employed in this work were of analytical reagent grade. They were
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used as received without any further purification. Palladium chloride, 2,3-butanedione
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monoxime, 98% and 5-bromosalicylaldehyde, 98% were procured from Sigma-Aldrich
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Chemicals Pvt. Ltd; while hydrazine hydrate, tetrahydrofuran and methanol were from Merck,
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India. 2,3-butanedionemonoxime monohydrazone was prepared by literature method [44].
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4.1.2. Physical measurements
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An electro-thermal digital melting point apparatus (SUMSIM, India make) was used to know the
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melting points and were uncorrected. C, H and N microanalyses were carried out on a Perkin-
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Elmer 2400II elemental analyzer. FT-IR spectra (KBr pellet) of LH and 1 were recorded with a
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Nicolet Magna-IR spectrophotometer (Series II). The presence of ν(Pd–Cl), ν(Pd–N) and ν(Pd–
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O) vibrations in 1 have been confirmed with the help of a Raman spectrometer (WITEC alpha,
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300 R) in the range 2000–200 cm–1 (wave length 532 nm). Electronic spectra (UV-visible) of
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both the ligand and complex were recorded on a Shimadzu UV-160A spectrophotometer in
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DMSO and DMF solvents. 300 MHz 1H NMR spectra for both the ligand and complex and 13C
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NMR spectrum only for ligand were recorded on a Bruker DPX300 spectrometer using DMSO-
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d6 as a solvent. 500 MHz
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DPX500 spectrometer. All chemical shifts (δ) are given in parts per million (ppm) relative to
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TMS, used as an internal reference. Molar electrical conductivities of 1 in DMSO and DMF were
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measured at room temperature on a Mettler Toledo FiveEasy Plus Conductivity meter. Prior to
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each acquisition, the cell constant was calibrated with 0.1 M aqueous KCl solution. Magnetic
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susceptibility measurements were done at 300 K with a PAR 155 vibrating sample magnetometer
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fitted with a walker scientific L75FBAL magnet. Hg [Co(SCN)4] was used as the calibrant. The
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ESI-Mass spectrum of LH in the positive ionization mode was recorded in acetonitrile (MeCN)
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solution on a Xevo G2 Q-Tof Micromass spectrometer. A JEOL Mass spectrometer (Model:
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JMS-700), Japan was employed to record the FAB-Mass spectrum of 1 in the positive ionization
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mode in m-nitrobenzyl alcohol matrix. Electrochemical cyclic voltammetric (CV) experiments
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were done with a Bioanalytical Systems Inc. Epsilon electrochemical workstation (Model: CV-
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50) on a C3 cell stand at 298 K under dry and degassed DMF which contained 1.0 mM of analyte
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and 0.10 M tetra-n-butyl ammonium perchlorate (TBAP) as supporting electrolyte. The base
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electrolyte was saturated with nitrogen for 15 min prior to each acquisition. A blanket of nitrogen
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gas was maintained throughout the measurements. The measurements were carried out with a
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three-electrode assembly consisting of a Glassy Carbon (GC) working electrode, a platinum wire
C NMR spectrum (in DMSO-d6) of 1 was recorded on a Bruker
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counter electrode and a Ag/AgCl reference electrode. The working electrode was polished before
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each experiment with alumina slurry. All potentials reported herein are referenced to Ag/AgCl.
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Scanning Electron Microscopy was done in a JEOL JSM 6360 to find out the mode of action of
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compound 1.
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4.1.3. Preparation of 3-[(5-bromo-2-hydroxy-benzylidene)-hydrazono]-butan-2-one oxime, LH
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2,3-butanedionemonoxime monohydrazone (0.230 g, 2.0 mmol) was dissolved in 25 mL of
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tetrahydrofuran to get a colorless solution. To it, 5-bromosalicylaldehyde (0.402 g, 2.0 mmol)
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was added all at a time. The resulting colorless solution was heated under reflux for 5 h. A dark
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yellow solution was obtained after refluxing. It was left in the air for slow evaporation. After 48
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h, yellow amorphous compound separated thereby. It was filtered, washed thoroughly with
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diethyl ether and subsequently dried in a vacuum desiccator over fused CaCl2.
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Yield: 0.465 g (80%). m.p. 180–182 °C. C11H12BrN3O2 (298.125): Anal. Calc. (%) for
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C11H12BrN3O2: C, 44.31; H, 4.05; N, 14.09; Found (%): C, 44.51; H, 3.94; N, 13.92. FT-IR
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(KBr): ν [cm–1] = 3253(vb) [ν(OH)], 1612(s) [ν(C=N)], 1547(s) [ν(C=C) of phenyl ring
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skeletal)], 1477(s) [ν(C=N) of oxime)], 1271(s) [ν(C–O) of phenolic –OH)], 1184(s) [ν(N–O)].
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1
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to phenyl ring), 11.30 (1H, s, hydrogen of –OH group attached to oxime moiety), 8.65 (1H, s,
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methylene proton of the –CH=N group attached to phenyl ring), 7.89 (1H, s, ortho-hydrogen of
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the phenyl group), 7.53 (1H, d, para-hydrogen of phenyl group), 6.95 (1H, d, meta-hydrogen of
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phenyl group), 2.20 (3H, s, methyl protons near to hydrazono moiety), 2.04 (3H, s, methyl
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protons close to oxime moiety).
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158.4(C10), 158.1(C8), 155.2(C1), 135.5(C3), 132.6(C5), 121.2(C6), 119.2(C2), 111.0(C4),
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13.4(C9), 9.8(C11). UV-Vis (DMSO): λmax[nm] (εmax[M–1cm–1]) = 348 (1.105 × 104), 291 (2.036
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H NMR (300 MHz, DMSO-d6, TMS) δ [ppm]: 11.97 (1H, s, hydrogen of –OH group attached
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C NMR (76 MHz, DMSO-d6, TMS) δ [ppm]: 164.8(C7),
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× 104). UV-Vis (DMF): λmax[nm] (εmax[M–1cm–1]) = 347 (1.295 × 104), 291 (2.987 × 104). ESI-
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MS (positive ion mode in MeCN) (m/z): 298.022 (Calc. 298.229) (79Br) and 300.019 (Calc.
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300.229) (81Br) for [LH+H]+.
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4.1.4. Preparation of [Pd(L)(Cl)](1)
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0.030 g (0.1 mmol) of ligand (LH) was dissolved in 10 mL of THF to get a light yellow solution.
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Na2[PdCl4] (0.030 g, 0.1 mmol), dissolved in 10 mL of THF, was added dropwise to the ligand
282
solution with constant stirring at room temperature. The stirring was continued for 3 h. A dark
283
orange yellow reaction mixture was obtained thereafter. It was kept in a refrigerator for slow
284
evaporation. After 10 days shining, air-stable, diamond-shaped orange single crystals were
285
deposited. These were filtered, washed thoroughly with diethyl ether and dried in a vacuum
286
desiccator over fused CaCl2. Some of the orange crystals were fit for X-ray structure
287
determination.
288
Yield: 0.016 g (36 %). m. p. > 240 °C. C11H11BrClN3O2Pd (438.99): Anal. Calc. (%) for
289
C11H11BrClN3O2Pd: C, 30.10; H, 2.53; N, 9.57; Found (%): C, 30.23; H, 2.42; N, 9.44 %. FT-IR
290
(KBr): ν [cm–1]: 3070 (br) [ν(OH)], 1624(vs) [ν(C=N) of imine], 1564(s) [ν(C=C) of phenyl ring
291
skeletal)], 1475(s) [ν(C=N) of oxime)], 1267(s) [ν(C–O) of phenolic –OH)], 1182(s) [ν(N–O)],
292
445(s) [ν(Pd–N)]. Raman: ν [cm–1]: 1260 [ν(C–O) of phenolic –OH)], 647 [ν(Pd–O)], 440 [ν(Pd–
293
N)] , 300 [ν(Pd–Cl)]. 1H NMR (300 MHz, DMSO-d6, TMS) δ [ppm]: 11.15 (1H, s, hydrogen of
294
–OH group attached to oxime moiety), 8.95 (1H, s, methylene proton of the –CH=N group
295
attached to phenyl ring), 7.91 (1H, s, ortho-hydrogen of the phenyl group), 7.57 (1H, d, para-
296
hydrogen of phenyl group), 6.98 (1H, d, meta-hydrogen of phenyl group), 1.24 (3H, s, methyl
297
protons near to hydrazono moiety), 1.14 (3H, s, methyl protons close to oxime moiety).
298
NMR (126 MHz, DMSO-d6, TMS) δ [ppm]: 160.6(C7), 157.5(C10, C8), 151.0(C1), 135.3(C3),
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131.4(C5), 120.4(C6), 118.8(C2), 110.4(C4), 13.0(C9), 11.8(C11). UV-Vis (DMSO): λmax[nm]
300
(εmax[M–1cm–1]) = 440 (9.854 × 102), 365 (1.184 × 104), 291 (1.666 × 104). UV-Vis (DMF):
301
λmax[nm] (εmax[M–1cm–1]) = 443 (9.858 × 102), 377 (1.399 × 104), 296 (3.294 × 104). FAB-MS
302
(positive ion mode in m-nitrobenzyl alcohol matrix) (m/z): 402.1 (Calc. 402.63) (79Br) and 404.1
303
(Calc. 404.63) (81Br) for [Pd(L)]+. ΛM (DMSO): 10 Ω–1 cm2 mol–1 (Non-conducting). ΛM (DMF):
304
57 Ω–1 cm2 mol–1 (Non-conducting). μ = Diamagnetic.
305
4.1.5. Crystal structure determination
306
An appropriate single crystal of 1, suitable for X-ray crystallography, was selected following
307
examination under a microscope. Intensity data were collected at 298(2) K using a graphite
308
monochromated Mo-Kα radiation (λ = 0.71073 Å) on a Bruker Smart Apex II diffractometer
309
equipped with 1K CCD instrument. SMART software [45] was used to determine cell
310
parameters. Using SAINTPLUS [45], data reduction and corrections were made. Empirical
311
absorption corrections were performed with SADBAS program [46]. The structure was solved
312
by direct methods with the program SHELXS-97 and was refined with SHELXL-97 [46] by full-
313
matrix least-squares methods on all F2 data. All non-hydrogen atoms were refined
314
anisotropically. Hydrogen atoms attached with C, N and O atoms were placed in calculated
315
positions and were constrained to ride on their parent atoms. On the basis of observed reflections
316
and variable parameters, the final cycle of full-matrix least-squares refinement was executed.
317
4.2. Studies of antimicrobial activity
318
In vitro testing was undertaken to examine the antimicrobial activity of both LH and 1.
319
4.2.1. Microbial strains
320
The in vitro antimicrobial activity of 1 was evaluated against 15 different pathogenic bacteria
321
belonging to Gram-negative genera like Escherichia coli 25938, Escherichia coli 397,
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Salmonella typhi 62, Vibrio cholerae 20, Shigella dysenteriae 1, Shigella dysenteriae 37,
323
Salmonella typhimurium NTCC 74, Pseudomonas aeruginosa 25619 and Gram-positive genera
324
like Staphylococcus aureus 29737, Bacillus cereus 11778, Bacillus subtilis 6633, Streptococcus
325
epidermidis 12228,
326
bronchiseptica 4617 respectively. All these microbial strains were collected from Division of
327
Microbiology, Department of Pharmaceutical Technology, Jadavpur University, Kolkata, India.
328
4.2.2. Preparation of Inoculums
329
The bacterial strains were grown in nutrient broth (HI-MEDIA) at 37 0C for 24 h. Then the
330
strains were adjusted to 0.5 McFarland standards [47] and diluted with sterile normal saline to
331
give initial bacterial suspension of 2×106 CFU/mL.
332
4.2.3. Reagent, chemicals and culture media
333
Amoxicillin (Sigma-Aldrich) was used as reference antibiotics for the evaluation of bacterial
334
inhibition. All the media used in antimicrobial study were purchased from Oxoid Ltd., UK.
335
4.2.4. Determination of MIC
336
The minimum inhibitory concentration (MIC) of the test compound was determined by agar
337
dilution method following NCCLS 2006 protocol [48]. Briefly, the compound was first dissolved
338
in 2% DMSO and then sterile water was added to give rise a final concentration of 1000 μg/mL.
339
This mother solution was then serially diluted to obtain different concentrations — 10, 25, 50,
340
100, 200 and 400 μg/mL. Preliminary analysis revealed that any inhibition on tested
341
microorganism was not shown up to 4% DMSO (v/v). Standardized inoculum (2×106 CFU/mL)
342
was spotted on the agar plates and incubated at 37 °C for 24 h. Amoxicillin was used as a
343
reference standard antibiotic. Lowest concentration which completely inhibited visible growth of
344
microorganism was recorded as MIC [49].
Bordetella
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Micrococcus luteus 10240, Bacillus pumilus 14884,
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4.2.5. Antimicrobial assay
346
Antimicrobial activity was determined by Well-diffusion method according to the NCCLS 2004
347
guidelines [50]. Nutrient Agar (NA) plates were prepared in 90 mm sterile Petri dishes (20 mL
348
NA) and 100 μL bacterial cell suspension (2×106 CFU/mL) was spread on solid NA plates using
349
spread plate techniques. Wells were dug by sterile borer and appropriate concentration of drug
350
was added to the wells. After a while, all the plates were incubated at 37 0C for 24 h. Amoxicillin
351
was used as a reference standard to compare antimicrobial activity of the test sample against the
352
tested microorganisms. The sensitivity was measured by clear inhibitory zones (mm) formed
353
around the wells. Each experiment was repeated in triplicate and the average of three trials were
354
reported [23,51].
355
4.2.6. Growth kinetic studies
356
Time-kill assay was performed in Mueller-Hinton Broth (MHB) containing the test sample.
357
Briefly, tubes containing liquid broth with and without (growth control) test drug were seeded
358
with 1 mL of log-phase inoculum (2×106 CFU/mL) to a final volume of 10 mL. These were
359
incubated at 37 °C. At different time intervals (h) after inoculation, a 0.1 mL volume was taken
360
from each tube and was subjected to 1:10 dilution. An aliquot of 10 μL of each dilution was
361
taken and plated on NA plates. Then the plates were incubated at 37 °C for 24–48 h and the
362
numbers of colony forming units (CFU) were counted. By using the viable counts at each
363
interval, a 24 h time-kill curve was recorded for each. The experiment was performed in
364
duplicate. The log CFU/mL was plotted on a graph as a function of time. It was used to compare
365
the rate and extent of killing-effect in various concentrations of the test drug [52,53].
366
4.2.7. Scanning Electron Microscopy
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Scanning electron microscopic observations were carried out on bacterial cells after 16 h
368
incubation in MHB at 37 0C. Test compound at 2 fold of MIC concentration was added to the
369
cell and was incubated for 24 h at 37 0C. A 24 h incubated normal culture was used as a control.
370
After that, bacterial cells were fixed with 4% glutaraldehyde in 0.1 M phosphate buffer for 2 h at
371
room temperature and then were washed four times in 0.1 M phosphate buffer (pH 7.2–7.4).
372
They were dehydrated in a graded alcohol series and were mounted onto stubs using double-
373
sided carbon tape. Then these were coated with a thin layer of gold using a microscopic sputter
374
coater for 1 min at 20 mA and were observed under the scanning electron microscope [54,55].
375
Acknowledgements
376
Financial support [F. No. 41–220/2012 (SR)] received from the University Grants Commission
377
(UGC), New Delhi, India, is thankfully acknowledged. M.L.Z and L.P.L. gratefully appreciates
378
the NSFC (Grant Nos. 21171109 & 21271121), SRFDP (Grant Nos. 20111401110002 &
379
20121401110005), the Natural Science Foundation of Shanxi Province of China (Grant Nos.
380
2010011011–2 & 2011011009–1) and the Shanxi Scholarship Council of China (2012–004 &
381
2013–026). Special thanks are also due to the revered reviewers for their valuable and
382
constructive suggestions during revision.
383
Appendix A. Supplementary data
384
CCDC 960808 contains the supplementary crystallographic data for 1. These data can be
385
retrieved free of charge via http://www.ccdc.cam.ac.uk/conts/retrieving.html, or from the
386
Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK; fax: (+44)
387
1223–336–033; or e–mail: [email protected].
388
References
389
[1]
AC C
EP
TE D
M AN U
SC
RI PT
367
D. Gaynor, D.M. Griffith, J. Chem. Soc., Dalton Trans. 41 (2012) 13239–13257.
17
ACCEPTED MANUSCRIPT
390
[2]
C. Orvig, M.J. Abrams, Chem. Rev. 99 (1999) 2201–2204.
391
[3]
B.T. Khan, J. Bhatt, K. Najmuddin, S. Shamsuddin, K. Annapoorna, J. Inorg. Biochem.
393
44 (1991) 55–63. [4]
394
J. Zhang, L. Li, L. Ma, F. Zhang, Z. Zhang, S. Wang, Eur. J. Med. Chem. 46 (2011)
RI PT
392
5711–5716. [5]
C. Rîmbu, R. Danac, A. Pui, Chem. Pharm. Bull. 62 (1) (2014) 12–15.
396
[6]
D.R. Ilić, V.V. Jevtić, G.P. Radić, K. Arsikin, B. Ristić, L. Harhaji-Trajković, N.
SC
395
Vuković, S. Sukdolak, O. Klisurić, V. Trajković, S.R. Trifunović, Eur. J. Med. Chem. 74
398
(2014) 502–508. [7]
400 401
(2014) 93–98. [8]
402 403
M. Kalita, P. Gogoi, P. Barman, B. Sarma, A.K. Buragohain, R.D. Kalita, Polyhedron 74
N.M. Aghatabay, M. Somer, M. Senel, B. Dulger, F. Gucin, Eur. J. Med. Chem. 42 (2007) 1069–1075.
[9]
404
TE D
399
M AN U
397
J. Traeger, T. Klamroth, A. Kelling, S. Lubahn, E. Cleve, W. Mickler, M. Heydenreich, H. Müller, H. Holdt, Eur. J. Inorg. Chem. (2012) 2341–2352.
[10]
A. Garoufis, S.K. Hadjikakou, N. Hadjiliadis, Coord. Chem. Rev. 253 (2009) 1384–1397.
406
[11]
M. Jagadeesh, H.K. Rashmi, Y.S. Rao, A.S. Reddy, B. Prathima, P.U.M. Devi, A.V.
408
Reddy, Spectrochim. Acta A 115 (2013) 583–587.
[12]
409 410
A. Karaküçük–İyidoğan, D. Taşdemir, E.E. Oruç–Emre, J. Balzarini, Eur. J. Med. Chem. 46 (2011) 5616–5624.
[13]
411 412
AC C
407
EP
405
F. Shaheen, A. Badshah, M. Gielen, C. Gieck, M. Jamil, D. de Vos, J. Organomet. Chem. 693 (2008) 1117–1126.
[14]
P.F. Salas, C. Herrmann, C. Orvig, Chem. Rev. 113 (2013) 3450–3492.
18
ACCEPTED MANUSCRIPT
[15]
414
Siddiq, A. Hameed, S.Z. Haider, Inorg. Chim. Acta 363 (2010) 3261–3269. [16]
416 417
V.V. Padma, R. Renganathan, K. Natarajan, Metallomics 4 (2012) 101–113. [17]
G.P. Vasić, V.V. Glodjović, I.D. Radojević, O.D. Stefanović, L.R. Čomić, V.M. Djinović, S.R. Trifunović, Inorg. Chim. Acta 363 (2010) 3606–3610.
418 419
P. Kalaivani, R. Prabhakaran, F. Dallemer, P. Poornima, E. Vaishnavi, E. Ramachandran,
RI PT
415
S. Nadeem, M. Bolte, S. Ahmad, T. Fazeelat, S.A. Tirmizi, M.K. Rauf, S.A. Sattar, S.
[18]
P. Kalaivani, R. Prabhakaran, E. Ramachandran, F. Dallemer, G. Paramaguru, R.
SC
413
Renganathan, P. Poornima, V.V. Padma, K. Natarajan, J. Chem. Soc., Dalton Trans. 41
421
(2012) 2486–2499.
M AN U
420
422
[19]
A.K. Mishra, N.K. Kaushik, Eur. J. Med. Chem. 42 (2007) 1239–1246.
423
[20]
M.N. Patel, P.A. Dosi, B.S. Bhatt, Inorg. Chem. Commun. 21 (2012) 61–64.
424
[21]
J. Chakraborty, H. Mayer–Figge, W.S. Sheldrick, P. Banerjee, Polyhedron 25 (2006)
[22]
428
A. Valkonen, K. Rissanen, S.R. Trifunović, Polyhedron 31 (2012) 69–76. [23]
429 430
(2013) 211–220. [24]
431
L. Zhao, X.T. Dong, Q. Cheng, J.X. Zhao, L. Wang, Synth. React. Inorg. Met–Org. Nano–Met. Chem. 43 (2013) 1241–1246.
432
[25]
433
[26]
434
C. Biswas, M. Zhu, L. Lu, S. Kaity, M. Das, A. Samanta, J.P. Naskar, Polyhedron 56
EP
427
G.P. Radić, V.V. Glođović, I.D. Radojević, O.D. Stefanović, L.R. Čomić, Z.R. Ratković,
AC C
426
3138–3144.
TE D
425
M. Asadi, K.A. Jamshid , A.H. Kyanfar, Inorg. Chim. Acta 360 (2007) 1725–1730. M. Espinal, J. Pons, J. García–antón, X. Solans, M. Font–bardia, J. Ros, Inorg. Chem. Commun. 12 (2009) 368–370.
19
ACCEPTED MANUSCRIPT
435
[27]
436 437
A. Castiñeiras, N. Fernández–Hermida, I. García–Santos, L. Gómez–Rodríguez, J. Chem. Soc., Dalton Trans. 41 (2012) 13486–13495.
[28]
K. Nakamato, Infrared and Raman Spectra of Inorganic and Coordination Compounds, Part B: Application in Coordination, Organometallic and Bioinorganic Chemistry, 5th
439
ed., Wiley–Interscience, New York, 1997.
RI PT
438
[29]
G.W. Simmons, Y.N. Wang, J. Marcos, K. Klier, J. Phys. Chem. 95 (1991) 4522–4528.
441
[30]
M. Hakimi, Z. Mardani, K. Moeini, F. Mohr, M.A. Fernandes, Polyhedron 67 (2014) 27–
442
35.
SC
440
[31]
W.J. Geary, Coord. Chem. Rev. 7 (1971) 81–122.
444
[32]
T. Motoyama, Y. Shimazaki, T. Yajima, Y. Nakabayashi, Y. Naruta, O. Yamauchi, J.
445
Am. Chem. Soc. 126 (2004) 7378–7385. [33]
447
A.W. Addison, T.N. Rao, J. Reedijk, J. van Rijn, G.C. Verschoor, J. Chem. Soc., Dalton Trans. 7 (1984) 1349–1356.
TE D
446
M AN U
443
448
[34]
L. Yang, D.R. Powell, R.P. Houser, J. Chem. Soc., Dalton Trans. 9 (2007) 955–964.
449
[35]
R. Greef, R. Peat, L.M. Peter, D. Pletcher, J. Robinson, Instrumental Methods in
[36]
452 453
Polyhedron 16 (1997) 3625–3633.
[37]
454 455 456
D. Kovala–Demertzi, A. Domopoulou, M.A. Demertzis, A. Papageorgiou, D.X. West,
AC C
451
Electrochemistry, Ellis Horwood Limited, England, 1985, pp. 188–195.
EP
450
K. Todar, Pathogenic E. coli. Online Textbook of Bacteriology, University of Wisconsin– Madison, Department of Bacteriology.
[38]
S. Shujha, A. Shah, Z. Rehman, N. Muhammad, S. Ali, R. Qureshi, N. Khalid, A. Meetsma, Eur. J. Med. Chem. 45 (2010) 2902–2911.
20
ACCEPTED MANUSCRIPT
457
[39]
458
A.V. Krishnaraju, T.V.N. Rao, D. Sundararaju, M. Vanisree, H.S. Tsay, G.V. Subbaraju, Int. J. Appl. Sci. Eng. 4 (2006) 115–125.
[40]
B.G. Tweedy, Phytopathology 55 (1964) 910–914.
460
[41]
R. Prabhakaran, A. Geetha, M. Thilagavathi, R. Karvembu, V. Krishnan, H. Bertagnolli,
[42]
463
Spectrochim. Acta A 107 (2013) 108–116. [43]
465 466
Rehim, RSC Adv. 4 (2014) 24131–24141. [44]
J.P. Naskar, B. Guhathakurtha, L. Lu, M. Zhu, R. Bhattacharya, J. Ind. Chem. Soc. 90 (2013) 1301–1309.
467 468
F.A. Bassyouni, S.M. Abu–Baker, K. Mahmoud, M. Moharam, S.S. El–Nakkady, M.A.
[45]
469
M AN U
464
K.S. Prasad, L.S. Kumar, S. Chandan, R.M.N. Kumar, H.D. Ravanasiddappa,
Bruker, SMART (Version 5.0) and SAINT (Version 6.02), Bruker AXS Inc., Madison, Wisconsin, USA, 2000.
TE D
462
K. Natarajan, J. Inorg. Biochem. 98 (2004) 2131–2140.
SC
461
RI PT
459
[46]
G.M. Sheldrick, Acta Crystallogr. A 64 (2008) 112–122.
471
[47]
J. McFarland, J. Am. Med. Assoc. 14 (1907) 1176–1178.
472
[48]
National Committee for Clinical Laboratory Standards, approved standard M2–A9,
474
NCCLS, Wayne, PA; 9th Edn. USA, 2006. [49]
475
Ngadjui, B.M. Abegaz, BMC Complementary and Alternative Medicine 11 (2011) 42 (1–
476 477 478
V. Kuete, P.Y. Ango, G.W. Fotso, G.D.W.F. Kapche, J.P. Dzoyem, A.G. Wouking, B.T.
AC C
473
EP
470
5).
[50]
National Committee for Clinical Laboratory Standards, approved standard M44–A. NCCLS, Wayne, PA, USA, 2004.
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[51]
480 481
Spiteller, J. Natural Products 74 (2011) 272–278. [52]
482 483
M.D. Awouafack, P. Spiteller, M. Lamshöft, S. Kusari, B. Ivanova, P. Tane, M.
C. Bantar, C. Schell, G. Posse, A. Limansky, V. Ballerini, L. Mobilia, Diagn. Microbiol. Infect. Dis. 61 (2008) 309–314.
[53]
484
RI PT
479
E.E. Roling, E.M. Klepser, A. Wasson, R.E. Lewis, E.J. Ernst, M.A. Pfaller, Diagn. Microbiol. Infect. Dis. 43 (2002) 13–17.
[54]
I. Kaya, N. Yigit, M. Benli, Afr. J. Trad. CAM 5 (4) (2008) 363–369.
486
[55]
S. Kaneshima, S. Koga, K. Tanaka, J. Electron Microsc 26 (1977) 355–358.
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List of the Captions
504
Figure Captions
505
Fig. 1. Synthetic scheme of LH and 1
506
Fig. 2. The structure of [Pd(L)(Cl)] (1) with ellipsoids at 30% probability and H-bonding motif.
507
Fig. 3. Minimum inhibitory concentration (MIC) of 1.
508
Fig. 4. Zone of inhibition of 1.
509
Fig. 5. Scanning electron micrographs of Bacillus cereus 11778 and Pseudomonas aeruginosa
510
25619 (A1, B1) before and (A2, B2) after treatment with compound 1.
511
Table Captions
512
Table 1 Crystal data and structure refinement for the compound, 1
513
Table 2 Hydrogen bonds (Å and o) for 1
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Table 1
Compound
1
RI PT
Crystal data and structure refinement for the compound, 1
960808
Color/shape
Orange/Diamond
SC
CCDC No.
Empirical formula
C11H11BrClN3O2Pd 438.99
M AN U
Formula weight Temperature [K]
298(2)
Wavelength [Å]
0.71073
Crystal system
Triclinic
P–1
a [Å] b [Å] c [Å]
AC C
β [o]
EP
Unit cell dimensions
TE D
Space group
7.1744(1) 8.7770(2) 11.2552(2) 79.4400(10)
Volume [Å3]
672.93(2)
Z
2
ρCalcd [g/cm3]
2.167
Absorption coefficient [mm–1]
4.546
F(000)
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0.35 × 0.25 × 0.2
range for data collection [deg]
1.84 to 25.05
Limiting indices
–8
