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Original Article
Development and validation of a simple and economical spectrofluorimetric
method for estimation of ciprofloxacin in pharmaceutical dosage forms
Serigne Omar SARR1,2,*, Serigne Momar NDIAYE1, Djibril FALL2,3, Adama DIEDHIOU2,3, Amadou DIOP2, Bara
NDIAYE2, Yrim Mbagnick DIOP1,2
1
Laboratoire National de Contrle des Mdicaments, 39, Avenue Pasteur, BP 6303, Dakar-toile, Sngal
2
Laboratoire de Chimie Analytique et Bromatologie, Facult de Mdecine, de Pharmacie et dOdontologie, Universit
Cheikh Anta DIOP de Dakar, BP 5005 Dakar-Fann, Sngal
3
Laboratoire de Chimie Organique et Thrapeutique, Facult de Mdecine, de Pharmacie et dOdontologie, Universit
Cheikh Anta DIOP de Dakar, BP 5005 Dakar-Fann, Sngal
*
Corresponding author : sosarr1@yahoo.fr
Received 23 June 2013; accepted 16 July 2013
Abstract
A new simple, precise, accurate and green spectrofluorimetric method for the determination of ciprofloxacin both as a
bulk drug and in pharmaceutical formulations was developed and validated using water as solvent. At a predetermined
excitation wavelength (280 nm) and emission wavelength (450 nm), it was proved linear in the concentration range of 20250 ng/mL, exhibited good correlation coefficient (R2= 0.9993) and excellent mean recovery (98.40-102.26%). The results
of the recoveries studies showed that the method was not affected by the presence of common excipients. The method was
applied for the analysis of the drug in the pure, tablet and injectable forms. The method was validated for various
parameters according to ICH guidelines. LOD and LOQ for ciprofloxacin were found to be 4.53 ng/mL and 5.67 ng/mL
respectively. The method has been successfully applied for the analysis of marketed formulations available in Senegal.
2013 Universal Research Publications. All rights reserved
KEY WORDS: Spectrofluorometric analysis, validation, ciprofloxacin, green method.
1. INTRODUCTION
Ciprofloxacin is a synthetic antibiotic of the
fluoroquinolone drug class and is chemically cyclopropyl6-fluoro-4-oxo-7-(piperazin-1-yl)-quinoline-3-carboxylic
acid. It is a second-generation-fluoroquinolone anti-bacterial.
Literature survey revealed that ciprofloxacin is very
sensitive to ultraviolet light (UV) and will fluoresce, due to
its highly conjugated resonance structure as shown with the
chemical structure in Fig. 1 [1].
72
73
3.4. Precision
Repeatability results indicate the precision under the same
operating conditions over a short interval of time and interassay precision. Intermediate precision expresses withinlaboratory variations in different days and in different
instruments.
In this work, intra-day precision (repeatability) and interday precision study (intermediate precision) of the method
were assessed at three concentration levels (100, 200 and
250 ng/mL) (n=3) against a qualified reference standard.
The inter-day precision study was performed using the
same protocol on three different days i.e day 1, day 2 and
day 3 at different concentration levels (100, 200 and 250
ng/mL) (n=3). These inter-day studies form part of
robustness. The relative standard deviations (% RSD) of
the obtained assay values at three different concentration
levels were calculated.
3.5. Stability studies
Samples prepared for repeatability study were preserved for
24h at room temperature and were analyzed on the
following day to test for short-term stability [11]. Mean
percentage recovery was determined.
3.6. Assay of content of ciprofloxacin in selected
marketed brands
Five market brands of ciprofloxacin tablets or bolus were
randomly selected and analyzed using the newly developed
and validated method. An exact amount of powder or
Or
100
331.34
367.81100
331.34
74
Table I: Recovery/accuracy for five different concentration of ciprofloxacin by the proposed method (n=8).
Dosage
Label claim
Pre-formulated
granulated
100 mg
Amount foundSD
157.44 0.25
178.90 0.40
201.70 0.30
224.99 0.41
244.58 0.25
RSD (%)
0.16
0.23
0.14
0.18
0.10
% recovered
98.40
99.40
100.80
102.26
101.91
Table II: Repeatability and intermediate precision determined for three different concentrations of ciprofloxacin (n=3).
Repeatability and intermediate precision determined for three different concentrations of ciprofloxacin (n=3)
Repeatability
Intermediate precision
Declared
concentration
(Mean SD
RSD
Average
RSD
Average
(Mean SD ng/mL)
(ng/mL)
ng/mL)
(%)
potency (%)
(%)
potency (%)
100
102.86 0.20
0.25
102.86
102.25 0.30
0.29
102.25
200
192.53 0.20
0.14
96.26
192.77 0.31
0.21
96.38
250
255.21 0.24
0.12
102.08
254.69 0.29
0.15
101.88
The slope of 1.041 and the low limits of detection and
quantization reaching both nanogram per liter indicate
clearly that the developed method is sensitive. These results
are in accordance with the sensitivity of fluorimetric
analytical techniques [17].
No significant difference was observed in the slopes of
calibration plots prepared on different days (ANOVA, P >
0.05). It can be concluded that the developed method is
sensitive.
4.2.3. Accuracy/recovery studies
The excellent % recovery (nearly 100%) and their low
standard (<0,5) represent accuracy. In water, the mean
percentage recoveries for concentration were found to be in
the range of 98.40 to 102.26% (Table 1). The % RSD in
accuracy studies was calculated at each concentration
levels and was found to be less than 0.5% (Table 1). Also
relative error (inaccuracy) values ranged from -1.87% to
2.27% as shown in table 1.
Since Ccalc (0.32) < C0,99 (0.63), the Cochrans test revealed
no high standard deviations. Also, since Gcalc < G0,99 (1.97)
(for all means at each concentration level), the Grubbs test
showed no outlying means.
No significant difference was observed between the mean
values of amounts added and found (p > 0.05).
These results revealed that any change in the drug
concentration in the solution can be accurately determined
by these proposed methods [18].
The high percentage recoveries obtained in Table 1 for
various amounts of ciprofloxacin in formulated mixture
with excipients suggested that there is no interference from
any of the excipients (such as starch, gelatin, gum arabic
and talc) as evidenced by the lack of absorbance and
emission at the specified max for the excipients and blank
solutions [11].
4.2.4. Precision
The RSD in precision studies was found to be 0.12-0.25 %
for repeatability (Intra-day precision) and 0.15-0.29% for
intermediate precision (Inter-day precision) (Table 2).
In all cases, R.S.D. values were within the acceptable range
indicating that this method has excellent repeatability and
intermediate precision.
The RSD in precision studies was found to be 0.12-0.25 %
(Intra-day) and 0.15-0.29 % (Inter-day) (Table 2). The
intra-day and inter-day precision study (Table 2) of the
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Table IV: Content of ciprofloxacin in five marketed products determined by the proposed method (n=3)
Concentration found
Drug found (%)
(mg)
Sample N Brand name (code) Label claim (mg)
Mean
SD
Mean
SD
1
A
750
743.62
0.19
99.15
0.03
2
B
500
475.99
0.01
95.19
0.05
3
C
500
495.21
0.05
99.04
0.02
4
D
500
511.03
0.01
102.20
0.05
5
E
100
101.30
0.01
101.30
0.01
RSD (%)
0.50
0.21
0.08
0.19
0.50
Table V: Application of the USP reference method (method A) and the proposed method (method B) for the determination
of ciprofloxacin in four pharmaceutical preparations (n=2).
Method A (HPLC)
Method B (Fluorimetry)
relative
Sample code
T- value F-value
error (%)
MeanStandard deviation (%)
MeanStandard deviation (%)
E
96.81 0.91
97.08 2.77
1.09a
9.31b
0.28
a
F
103.20 1.30
103.48 6,03
0.06
21.51b
0.27
G
103.88 2.22
100.52 1.06
0.46a
4.41b
-3.24
H
108.04 0.57
110.26 2.15
1.70a
13.94b
2.04
a
critical T-value (p=0,05) : 4,30
b
critical F-value (p=0,05) : 161
is the only ciprofloxacin assay method using water as
Stationery Office: London, 2009.
exclusive solvent without any trace of organic solvent. 6. USP, The United States Pharmacopeia, National
Such a method can be called green analytical technique.
Formulary NF29, vol. 2, Rockville, 2011.
7. WHO, World Health Organization, The International
5. Conclusion
The proposed method is simple, sensitive, cheap, less time
Pharmacopoeia, Quality Specifications, third ed, vol.5,
consuming and reliable (RSD <1%) with good precision
World Health Organization, Geneva, 2003.
and accuracy. The proposed method is specific while 8. ICH Q2B, International Conference of Harmonization,
estimating the commercial formulations without
Validation of Analytical procedure: Methodology, ICH
interference of excipients and other additives. Hence, this
guideline, 2005.
method can be used for the routine determination of 9. S. Bolton:, in: Pharmaceutical Statistics: Practical and
ciprofloxacin in pure samples and pharmaceutical
clinical application, Third ed., Marcel Dekker, New
formulations in quality control.
York,1997, pp216-264.
10. M.E. Swartz, I.S. Krull, Handbook of Analytical
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