micelle enhanced spectrofluorimetric determination of norfloxacin using terbium as fluorescent probe
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Micelle EnhancedSpectrofluorimetricDetermination of NorfloxacinUsing Terbium as FluorescentProbeZuyun Huang a , Ruxiu Cai a , Ke Zhang a , HoupingHuang a & Yune Zeng aa Department of Chemistry , Wuhan University ,Wuhan, 430072, P.R. ChinaPublished online: 22 Aug 2006.
To cite this article: Zuyun Huang , Ruxiu Cai , Ke Zhang , Houping Huang & YuneZeng (1997) Micelle Enhanced Spectrofluorimetric Determination of NorfloxacinUsing Terbium as Fluorescent Probe, Analytical Letters, 30:8, 1531-1539, DOI:10.1080/00032719708001673
To link to this article: http://dx.doi.org/10.1080/00032719708001673
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ANALYTICAL LETTERS, 30(8), 1531-1539 (1997)
MICELLE ENHANCED SPECTROFLUORIMETRIC
DETERMINATION OF NORFLOXACIN USING TERBIUM
AS FLUORESCENT PROBE
KEY WORDS: Norfloxacin, Terbium(lIl), fluorescent probe
Zuyun Huang, Ruxiu Cai, Ke Zhang, Houping Huang, Yune Zeng
Department of Chemistry,Wuhan University , Wuhan,430072, P.R. China
ABSTRACT
Terbium(II1) forms highly fluorescent complex with norfloxacin. The
presence of surfactant can enhance greatly fluorescence intensity. In this paper, the
fluorescence characteristics of complex in presence of surfactant has been
investigated. It was found that norfloxacin could form stable complex with terbium
in the pH 6.5-8.5 media. The fluorescence intensity of complex in SLS media is
three folds more than complex itself. A new micelle enhancement
spectrofluorimetry for the determination of norfloxacin has been developed .The
detection limit of proposed method is 0.017pg/ml. It has been satisfactory for the
determination of trace norfloxacin in serum with 95.2% -10 1.2% recoveries.
1531
Copyright 0 1997 by Marcel Dekker, Inc.
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1532 HUANG ET AL.
INTRODlJCTION
Norfloxacin[ 1 -ethyl-6-fluoro- I ,4-dihydro-4-oxo-7-( piperazin- 1 -y l)quinoline-
3-carboxylic acid] is the third representative of antibacterial medicine. namely
quinolone group, used extensively in the clinical treatment with higher potential and
a broad antibacterial spectrum, lower side effects. It killed bacteria through
inhibiting cell DNA-gyrase and prohibiting DNA-replication. I t is thus chosen
commonly as antibacterial medicine in the treatment of urinaq-tract infections.
gonorrhea and bacterial enteritis'. Previous literature have reported some method
for the determination of norfloxacin(NFX). such as HPLC. spectrophotonietiy.
microbioassay etc.2-S. However. most of them are based on intrinsic florescence.
The sensitivity of method is low. time-consuming and more back~round- in te r~~r in~ .
Recently, D. Perez-Bendito6 reported a method for deteiinination of
norfloxacin. which is based on a suitability of stopped-flow technique with
lanthanide-sensitized fluorescence for the determination of nalidixic acid and
norfloxacin. However, sensitivity of the method is still low. In this paper. we
combined h e advantages of micelle enhancement and lanthanide fluorescent probe.
A simple method with improved sensitivity and stability is proposed. The method
is based on terbium(lI1) forming a lughly fluorescent complex with norfloxacin. The
presence of surfactants (SLS) can enhance greatly the fluorescence of the complex.
The detection limit of the proposed method is 0 . 0 ~ 7 p ~ i i i l . Linear range is 0.05-
lpdml, the analybcal recoveries are between 95.2%-10 1.2%. The method has been
successfully applied to determine trace norfloxacin i n serum.
EXPERIMENTAL
Apparatus
All fluorescence intensity measurements were made on a SMF-25
specwofluorimeter (Kontron Instrument) with 1.Ocm quartz cells. Values of pH were
measured with a model of digital display pH meter ( Wuhan precision Insh-urnent
Corp., China).
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TERBIUM AS FLUORESCENT PROBE 1533
Reagents
Analytic-reagent grade chemicals were used. solutions were prepared in
doubly distilled water.
The lanthanide oxides were obtained from Yuelong Chemical Corp.(Shanghai,
China) in purity of more than 99.99%. Stock solutions of the lanthanide ion( Img/ml
Tb,O,) were prepared by dissolving known amounts of the appropriate rare earth
oxide in hydrochloric acid.
Norfloxacin (NFX) was obtained from Medicine and Bioproducts Test
Institute of Health Ministry. Stock solution of lmg/ml NFX solution was prepared
in 0. Imol/L sodium hydroxide and stored in a refrigerator (4OC). Standard solutions
were prepared fresh daily by diluting the stock solution. The sodium lauryl
sulfonate was obtained from Beijing Chemical Corp.( 1 . 0 ~ lO-*mol/L). The buffer
solution used was 0.2moVL Tris-HC1. The pH values were adjusted to 8 with
0.2moVL hydrochloric acid. A11 water used was doubly distilled water from quartz
vessels.
Procedure
To a lOml test tube, known amounts of NFX, Tb3+ standard solution, SLS
(1 x 10-2mol/L), 0.2mol/L Tris-HC1 buffer solution were added sequentially. The
volume was diluted to l h l with doubly distilled water, mixed well, and allowed
to stand for 5min at room temperature. The relative fluorescence intensity was
measured by setting the excitation and emission wavelengths at 325 MI and 545 tun
respectively.
RESULTS AND DISCUSSION
P c e spectrum
Fig. 1 shows the typical fluorescence spectrum of NFX,Tb3+-NFX,Tb3+-NFX-
SLS system at pH 8.0 obtained under experimental condition. The excitation of
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1534 HUANG ET AL.
A.excitation spectra
Fig. 1.
0 Wavelength (nm)
B. emission spectra
Fluorescence spectra
( I ) 0.5pg/ml NFX(pH=8.0)
(2) 0.5pg/ml NFX+4.92* 10-smol/LTb3+(pH=8.0)
( 3 ) 0.5 pdml NFX+4.92 x I 0-'mol/LTb3++9x 1 O4mol/LSLS(pH=8.0)
NFX is at 280nm and 332nm, its emission peak is 410nm. The excitation peak of
Tb3+-NFX complex moves to long wavelengths of 288nm and 332nm. The emission peaks of complex at 49Onm, 545nm, 590nm and 625nm are attributed
to the sD,-7D,(n=6,5,4,3) transition of Terbium. The most intense peaks are the
5D,-7D, transition at 49Onm and the sD,-7Ds transition at 545nm. Its luminescent
mechanism belongs to L-M; the emission peak(4lOnm) of NFX decreases. In the
presence of surfactant, the fluorescence intensity of the complex is enhanced
greatly,but the position of peak of the complex doesn't change.
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TERBIUM AS FLUORESCENT PROBE 1535
Effect of Tb3+ concentration
Lanthanide ions. especially europium(ll1) and terbium (111) , emit the energy
as narrow-banded line-type fluorescence spectrum with long Stockes shifts(over
250nm) and exceptionally long fluorescence decay times(0. I-lms). They are widely
used as fluorescence probe. In tlus work, the effect of terbium(lI1) concentration on
fluorescence intensity of system was studied. It was found that a maximum and
stable fluorescence intensity was obtained between 3.280~10-"-4.373 x 1 O-'mol/L.
Effect of surfactants
In the presence of surfactants, equilibrium, kinetic and spectral properties can
be modified and this has been used to improve the characteristics of analytical
procedures. The use of surfactant media can also enhance fluorescence intensity in
the spectra of drug molecules. Perhaps due to the formation of a stable mixed-
ligands complex above(CMC) critical micelle concentration, the solubility of
complex increases greatly. Hence the fluorescence intensity is enhanced greatly.
Fig.2. shows that the fluorescent intensity of system increases dramatically below
8 . 0 ~ 10-4mol/L of SLS, then reaches maximum at 9.0. IO-'mol/L SLS.(see Fig.2.)
Effect of DH and species of buffer solutions
The influence of pH on fluorescence intensity was investigated in the NFX
and NFX-Tb3+ system (see Fig.3). It was well known that the NFX molecular is
monoatomic acid with emitting intensive fluorescence at the acid
medium@H=l-3.5). The fluorescence emission body is the state of molecular in the
acidic media, whereas the ionic state is the weak fluorescent emission body in basic
media. Hence the fluorescence intensity Tb-NFX complex reaches maximum at pH
8.0. The optimum pH range is 7.0-8.5.
The fluorescence intensity of Tb3+-NFX systems was measured at the pH 8.0
with some species of the buffer solution. The results obtained are shown in Table2.
The experiments show that the 0.2M Tris-HCI buffer solution (pH=8.0) is best.
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1536 HUANG ET AL.
Fig.2.Effect of SLS on fluorescence intensity
0.5pg/ml NFX+4.92 1 0-smol/LTb3+(pH=8.0)
Fig.3. Effect of pH on the fluorescence intensities of NFX and Tb3+-NFX
a: [NFX]= 0.5pg’ml he, /h,,,=332/440nm
b: 0.5 pg/ml NFX+4.92 x 1 O-smol/LTb3+ hey /h,,,=3 32/545nm
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TERBIUM AS FLUORESCENT PROBE 1537
Table 1. Surfactants effect
surfactant no Bnj35( 1%) TX-100( 1%) CTAB( 1 x I04M) TPC*(0.2%) SDBS(O.2%) SLS( I x 10"M) If 27.3 37.3 33.5 25 .8 17.8 51.5 72.5
'Tetradecyl pyridine chloride
Table 3. The effect of various substances on the fluorescence intensity
species Ca2'(Mg2') Zn2+ Fe3+ cu2+ v c ............................................................................................... concentration(mol/l) 1 x lo" 1 . 2 ~ 1 . 6 ~ 5 . 5 ~ lo-* I x lo-'
Table 4. Determination of NFX in serum samples
sample concentration of NFX added in serum average value recovery h i d mu (Pg/mu (%)
1 0.800 0.762 95.3
2 0.500 0.495 99.0
3 0.300 0.298 99.2
4
5
0.100
0.0750
0.1013 101.3
0.0759 101.2
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1538 HUANG ET AL.
The stability of complex
The experiment shows that the fluorescence intensity of the system reaches a
maximum for ~ m i n at room temperature containing I x IO-’~OI/L, I x 1u6mol/L
NFX, and the complex stablizes for 24h.
Effect of interference SD . ecies in body
Table 3 shows the infuence of some metal ions and Vc in biological body
on fluorescence intensity. The following amounts of substances don’t interfere in
the determination for the concentration of 4 . 9 2 0 ~ 10-5mol/LTb3+ and O.l&ml
NFX:
Determination of NFX in s u
Under the experimental conditions, The fluorescence intensity with the
concentration of NFX is a linear relation in the range 0.05-lpg/ml,the
determination results deviated slightly from a linear relationship at high
concentration.The detection limit is O.O17pg/ml. The new method was applied
successfully to determine NFX in serum by standard addition method (see Table 4).
ACKNOWLEDGEMENTS
The authors gratefully acknowedge financial support from the National
Science Foundation of China.
REFERENCES
1. L.F. Wu, J.P.Zhang; Handbook of Import Medicine; Chinese Medical
Science and Technology Press, Beijing, 100- 10 1 ( 199 1).
G.Y.Zhao, H.Z.Fan, J.H.Pan, Chinese J. Pharm. Anal., 15(5)48-60 (1995). 2.
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TERBIUM AS FLUORESCENT PROBE 1539
3. J.R.Zhang, H.Pang, X.Y.Liu, Sh.J.Pan, H.S.Wang, Chines J . Pharm. Anal..
30(7)428-429 (1995).
4.
5 .
J. Georges, S. Ghazarian, Anal. Chim. Acta. 276:401-409 (1993).
P. Izquierdo, A. Gomez-hens, D. Perez-bendito. Anal. Chim. Acta. 292: 137-
139 (1994).
S. Panadero, A. Gomez-hens, D. Perez-bendito, Anal. Chim. Acta. 303:39-
45 (1995).
6 .
Received: November 1. 1996 Accepted: March 10. 1997
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