FARMACIA, 2009, Vol. 57, 2
247
SCANNING ELECTRONIC MICROSCOPY STUDY
OF NEW PYRAZINAMIDE COMPOUNDS WITH
METALLIC IONS
TÜNDE JURCA*, ELEONORA MARIAN
Oradea University, Faculty of Medicine and Pharmacy, Speciality of
Pharmacy, Department of Inorganic Chemistry, 29 Nicolae Jiga Street,
410028, Oradea, România
*corresponding author: [email protected]
Abstract
In this study it is presented the preparation method of two pyrazinamide
compounds with transitional metal ions, their physico-chemical properties, the results of the
elementary analysis, as well as the examination by scanning electronic microscopy of these
compounds’ particles form. The FT-IR (4000-400 cm-1) spectra of pyrazinamide (PZA),
[Cu(PZA)2]Cl2 and [Cu(PZA)2(C6H5COO)2 were recorded. The observed wavenumbers were
analyzed and assigned to different normal modes of vibration of the molecule.
Pyrazinamide (PZA) is among the first-line drugs for the treatment of
tuberculosis, which seems to reappear as a frequent disease.
With the assistance of the scanning electronic microscopy, the granules’ surface
aspect may be observed, the X-ray spectra that show the presence of the elements on the
analyzed micro area and the chemical composition diagram per granule for each newly
synthesized compound.
The examinations were performed using the scanning electronic microscope
SEM type Jeol 5600 LV, equipped with an X-ray spectrometer type Oxford Instrument.
Details concerning the spatial architecture and the crystalline structure will be
obtained by vibration spectroscopy of magnetic resonance and X-ray diffraction.
Rezumat
În acest studiu este prezentată prepararea a două combinaţii complexe ale
pirazinamidei cu ionii metalelor tranziţionale, proprietăţile fizico-chimice, rezultatele analizei
elementale, respectiv examinarea compuşilor sintetizaţi prin microscopie electronică de
baleiaj. Au fost înregistrate spectrele FT-IR (4000-400 cm-1) ale pirazinamidei (PZA),
[Cu(PZA)2]Cl2 şi [Cu(PZA)2(C6H5COO)2 şi au fost stabilite benzile caracteristice.
Pirazinamida este un tuberculostatic bine cunoscut având o acţiune
asemănătoare izoniazidei.
Prin microscopie electronică de baleiaj a fost observat aspectul suprafeţei
granulelor, iar spectrele de raze X au arătat prezenţa elementelor componente şi diagrama
compoziţiei chimice per granulă, pentru fiecare compus nou sintetizat.
Măsurătorile s-au efectuat cu un Microscop electronic cu baleiaj tip JEOL5600LV, cu mărire maximă de 300000x, dotat cu spectrometru cu raze X tip Oxford
Instrument, rezoluţia fiind 3,5 nm. Detaliile privind structura moleculară şi cristalină s-au
obţinut în urma studierii spectrelor de vibraţie şi de raze X.
Keywords: pyrazinamide, scanning electronic microscopy, crystalline structure,
FT-IR
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Introduction
Although they represent mere 3% of the human body, the metallic
ions (especially those of the transitional metals from 3d series), under the
form of some natural chelate compounds, they are present in all the
fundamental biologic processes, solar energy capture, photosynthesis, oxygen
and nitrogen fixation and transportation, coordination of all metabolic
reactions and controlled discharge of the energy in the biologic systems.
Furthermore, the formation of complex combinations has implications in the
explanation of some biologic processes or pathological conditions and may
constitute models for the study of the drugs’ pharmacodynamic effects.
In this study it is presented the preparation method of two
pyrazinamide compounds with transitional metal ions, their physico-chemical
properties, the results of the elementary analysis, as well as the examination
by scanning electronic microscopy of these compounds’ particles form.
Pyrazinamide (pyrazine carboxamide, PZA) is a nicotinamide
analogue that has been used for almost 50 years as a first-line drug to treat
tuberculosis [3]. PZA is bactericidal for mycobacteria and reduces the total
treatment time [4]. The parent molecule PZA and some of its complexes are
widely used, due to their antimycrobial properties [6].
The chemical structure is presented in Figure 1.
N
O
C
NH2
N
Figure 1
Chemical structure of pyrazinamide
Materials and methods
The pyrazinamide used is a reagent provided by Trans Medical
Pharma GmbH, Germany.
The FT-IR spectra of pyrazinamide and their complexes with
cooper (II) were recorded in the region 4000-400 cm-1 by a Bruker
EQUINOX 55 spectrometer, using a diamond ATR unit.
The examinations were performed using the scanning electronic
microscope SEM [7,8] type Jeol 5600 LV, being equipped with a X-ray
spectrometer type Oxford Instrument, with the following characteristics:
• resolution: 3.5nm with secondary electrons;
• enlargement: 300,000 x
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FARMACIA, 2009, Vol. 57, 2
•
•
the non-conducting samples’ examination (ceramics, biologic,
medical, etc) may be performed in poor vacuum (up to 130 Pa) with
backscattered electrons (maximum enlargement: 5,000 x );
local quantitative chemical analyses based upon the X-ray
characteristic spectrum (EDS) for the elements listed between boron
and uranium, with the detection limit of 0.01%.
Results and discussion
Synthesis of [Cu(PZA)2(C6H5COO)2: an aqueous solution (30 mL)
of CuSO4.5H2O (1.25 g) was added to 80 mL solution of pyrazinamide (2.4
g) and sodium benzoate(1.5g). The formed blue precipitate was removed by
filtration, then washed with H2O and EtOH, and finally dried at room
temperature; yields 65-70%.
Elemental analysis: found: C, 52.92%; H, 3.86%; N, 15.85%;
calculated: C, 52.18%; H, 3.62%; N, 15.22%; M = 551.91
Synthesis of [Cu(PZA)2Cl2]: an aqueous solution (30 mL) of CuCl2
(1.7 g) was added to a solution of pyrazinamide (2.4 g). The mixture was
stirred ½ h. The filtrate was set aside at room temperature one day, in order
to form a large quantity of green crystals, which was collected by filtration,
then washed with H2O and EtOH; yields 65-72%.
Elemental analysis: found: C, 30.1%; H, 3.04%; N, 21.07%;
calculated: C, 30.6%; H, 3.15%; N, 22.04; M = 398.66; color: green
The accentuated coloration of the internal compounds shows a sudden
deformation of the electronic layers of the atoms forming the mole [9,10].
From the results of the elemental analysis it results that the Me:L
combination ratio is 2:1 for [Cu(PZA)2(C6H5COO)2 and Cu(PZA)2]Cl2.
The assignment of some of the most characteristic IR bands of the
complex and ligand is shown in Table I.
Table I
The assignment of some of the most characteristic IR bands
Tent.
Assig.
νas NH2
νs NH2
νCH
νC=O(1)
δNH2(2)
νring
νring
νCN(III)
δCH
ΔNH2t
δring
ΔNH2w
PZA
3410s
3140m
3080
1705s
1600m
1570
1530
1375s
1150w
1090m
870w
665w
Me-PZA complexes
Cu(C6H5-COOH)2
3610w
3170m
3065m
1915w
1590m
1585m
1545s
1380s
1180w
1085w
850w
680m
Me-PZA complexes
CuCl2
3430s
3110m
3070m
1700s
1590m
1585m
1510w
1385s
1170m
1080w
870m
670w
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PZA can coordinate through the pyrazine ring nitrogens, the –C=O
and/or – NH2 groups. IR technique is a diagnostic of the coordination mode
of PZA in complexes.
When the aromatic ring nitrogen is involved in complex formation,
certain ring modes, particularly the ring breathing mode and the two ring
stretching modes around 1400-1600 cm-1, increase in value both due to the
coupling with Me-N (ligand) [4,5] bond vibrations and due to alterations of
the ring force field [6]. On the other hand, when amino nitrogen is involved
in coordination, drastic changes occur in amino group vibrational
wavenumbers [7,8,9], and when coordination occurs through the oxygen of
the carbonyl group, a negative shift of the νc=o mode of the coordinated
molecule with respect to the free ligand is expected [3].
Since it is well known that a coordinated amino group shows a
negative shift, Δ= 150-220 cm-1, in the NH2 stretching modes, in comparison
to the free ligand [7- 9], we can conclude that this group does not take part in
coordination. The IR data show that the amide·NH2 of PZA does not take part
in coordination, but forms weak hydrogen bonds, probably with guest water
molecules. In the IR spectra the ring-breathing mode of PZA is observed at
1015±10 cm-1 and around 1035 cm-1 for Me-PZA complexes. The lack of blue
shift of ring breathing suggests that the PZA molecule does not coordinate to
the metal (II) in Me-PZA complexes through the heterocyclic ring nitrogen.
We also did not observe any blue shift in certain ring modes due to formation
of Me-PZA complexes, which are known to be very sensitive to complex
formation through the ring nitrogen.
Typical scanning electronic microscopy (SEM) pictures showing
the morphology and the crystal structure of this combination were taken.
The complexes are crystalline, non-hygroscopic and insoluble in
common organic solvents. However, they are soluble to limited extent in
dimethylformamide (DMF) and dimethyl sulfoxide DMSO.
The proposed structures are depicted in figures 2 and 3.
O
N
N
C
Cu
NH2
C
N
N (C6H5COO)2
O
NH2
Figure 2
The proposed structure of [Cu(PZA)2](C6H5COO)2
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O
Cl
N
N
C
N
Cu
O
NH2
C
N
Cl
NH2
Figure 3
The proposed structure of [Cu(PZA)2]Cl2
For the [Cu(PZA)2](C6H5COO)2 compound, typical SEM pictures
(figure 4 and 5) show the morphology and the crystal shape of this
combination: irregular conglomeration with different shapes and
dimensions, some of them being lamellar. The analyzed segment is 10μm.
Figure 4
[Cu(PZA)2](C6H5COO)2 granule
surface’s aspect
Figure 5
Energy-dispersive spectroscopy (EDS)
spectrum (X-rays) of the elements on the
[Cu(PZA)2](C6H5COO)2 compound
analyzed micro area
For the compound Cu(PZA)2]Cl2 (figure 6 and 7) the particles have
an elongated acicular form, and the fibers of an average size of 1.5 microns
are predominant. The analyzed segment is 10 µm in size. The distribution of
the component elements has a high degree of uniformity. This may be
confirmed by the X-ray diffraction.
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Figure 6
A Cu(PZA)2]Cl2 granule surface’s aspect
Figure 7
EDS spectrum (X-rays) of the elements on
the compound Cu(PZA)2]Cl2 analyzed
micro area
Conclusions
By studying the two pyrazinamide, newly synthesized compounds
with transitional metallic ions, using the elemental analysis, it was
discovered that the ratio of Me:L is 1:2 for the Cu(PZA)2]Cl2 and
[Cu(PZA)2](C6H5COO)2 compounds. We may conclude that the
coordination through the – C=O groups and pyrazine ring nitrogen in the
Me-PZA complexes is implied by the FT-IR spectra.
Some of typical pyrazinamide bands are only slightly displaced
after coordination, remaining in their characteristic absorption regions.
With the assistance of the scanning electronic microscopy, the
granules’ surface aspect may be observed, the X-ray spectra that show the
presence of the elements on the analyzed micro area and the chemical
composition diagram per granule for each newly synthesized compound.
Details concerning the spatial architecture and the crystalline
structure will be obtained by vibration spectroscopy of magnetic resonance
and X-ray diffraction.
1.
2.
3.
4.
References
E.Akalin, S.Akyuz, J.Mol.Struct. 2001, 463/464, 579
E. Akalin, S. Akyuz, Journal of Moleculare Structure, 2001, 463-464, 579
C.Rao, Chemical Applications of Infrared Spectroscopy, Academic Press, London
1963
Grecu, I., Curea, E., Identificarea substanţelor medicamentoase, Editura Dacia,
Cluj-Napoca, 1980, 305
FARMACIA, 2009, Vol. 57, 2
5.
6.
7.
8.
9.
253
Kellner, R., Mermet, M.J., Otto, M., Valcarel, M., Widmer, M.H., Analytical
chemistry. A modern approach to analytical science, Second Edition, Wiley-Vch.,
2004, 911-951
Marian, E., Jurca, T., Banica, F., Vicas, L., Revista de Medicină şi Farmacie, 50,
suplimentul II, Târgu-Mureş, 2004, 171
S.Akyuz, J.E.D. Davies, J.Mol. Struct. 1982, 95 157
Skoog, A. D., Holler, F.J., Niemann, A.T., Principles of instrumental analysis,
Fifth Edition, Harcourt Brace College Publishers, 1998, 543-553
T.Jurca, S. Cavalu, L.Vicaş, F. Bănica, Synthesis and vibrational analysis of new
complexes of pyrazinamide with transition metal (II) Farmacia, 2005, LIII, 2 , 3846
Manuscript received: 20.10.2008