ОПТИКА И СПЕКТРОСКОПИЯ, 2013, том 115, № 2, с. 349-352
ГЕОМЕТРИЧЕСКАЯ И ПРИКЛАДНАЯ ОПТИКА
УДК 535.8
NEW SPECTROFLUORIMETRIC METHOD FOR THE DETERMINATION
OF NIZATIDINE IN BULK FORM AND IN PHARMACEUTICAL PREPARATIONS
© 2013 г. Ayga Karasakal* and Sevgi Tatar Ulu**
* Department of Chemistry, Faculty of Science and Letters, Namik Kemal University, Tekirdag, Turkey ** Department of Analytical Chemistry, Faculty of Pharmacy, Istanbul University, Istanbul, 34416 Turkey Received November 15, 2012; in final form, January 24, 2013
A simple, accurate and highly sensitive spectrofluorimetric method has been developed for determination of nizatidine in pure form and in pharmaceutical dosage forms. The method is based on the reaction between nizatidine and 1-dimethylaminonaphthalene-5-sulphonyl chloride in carbonate buffer, pH 10.5, to yield a highly fluorescent derivative peaking at 513 nm after excitation at 367 nm. Various factors affecting the fluorescence intensity of nizatidin-dansyl derivative were studied and conditions were optimized. The method was validated as per ICH guidelines. The fluorescence concentration plot was rectilinear over the range of 25—300 ng/mL. Limit of detection and limit of quantification were calculated as 11.71 and 35.73 ng/mL, respectively. The proposed method was successfully applied to pharmaceutical preparations.
DOI: 10.7868/S0030403413080035
INTRODUCTION
Nizatidine (NIZ) is a histamine H2-receptor antagonist [1]. Chemically, it is N-(2-[(2-[(dimethy-lamino)methyl]thiazol-4yl)methylthio]ethyl)-^-me-thyl-2-nitroethene-1,1-diamine [2]. Several methods have been reported concerning the analysis of NIZ in pharmaceutical preparations including — kinetic spectrophotometry [3, 4], spectrophotometry and titrime-try [5], spectrophotometry [6—10], voltammetry and spectrophotometry [11], derivative spectrophotometry [12], high performance liquid chromatography (HPLC) with UV detection [13], capillary electrophoresis [14], polarography [15], potentiometry [16], and culometry [17]. Some analytical methods for the determination of NIZ in biological fluids have been developed. These mainly include voltammetry [18], (HPLC) [19-25].
This paper describes, for the first time, the deriva-tization of NIZ with 1-dimethylaminonaphthalene-5-sulphonyl chloride (dansyl chloride). The proposed method is sensitive, accurate, simple and selective, and was applied for the determination of NIZ in pharmaceutical preparations and in pure.
EXPERIMENTAL
Apparatus
Model RF-1501 Shimadzu spectrofluorimeter (Kyoto, Japan) equipped with xenon lamp and 1 cm quartz cuvette (Japan).
Reagents and Materials
NIZ was kindly provided by Avantis (Istanbul, Turkey). Axid capsules (150 mg) (Avantis, Istanbul, TR) was purchased from a local pharmacy. Dansyl chloride was purchased from Fluka (Buchs, Switzerland). All solvents used were analytical grade (Merck Darmstadt, Germany). Water was purified by aquaMAXTM ultra, Young instrument (Korea) ultra water purification system.
PREPARATION OF STANDARD, REAGENT, BUFFER AND SAMPLE SOLUTIONS
Nizatidine Standard Solution
NIZ stock solution (0.001%, w/v) was prepared by dissolving 10 mg of NIZ in 10 mL of methanol. 0.0001% standard solution (w/v) was freshly prepared from the stock solution by appropriate dilutions with water.
Dansyl Chloride Solution
Reagent solution of (0.2 mg/mL) was prepared by dissolving 2 mg of dansyl chloride in 10 mL of acetone and protected from light.
Buffer Solutions
Sodium bicarbonate (0.1 M) solution was prepared in water and adjusted to pH 10.5 with 0.1 M sodium hydroxide. All solutions were kept refrigerated.
H3C
CH i
,N
V
Nizatidine Cl
O=S=O
CH3
no2
1000
500
Ö
<D
(a)
H3C CH3 Dansyl chloride
Fig. 1. Chemical structures of nizatidine with dansyl chloride.
Capsules Sample Solution
The contents of ten capsules were weighed and finely powdered. A quantity of the powder equivalent to 50 mg of NIZ was transferred into a 50 mL volumetric flask, dissolved in methanol, and sonicated for 30 min, the volume was then completed with methanol. The contents were mixed well and filtered. The working solution of 100 ^g/mL for NIZ was prepared by an appropriate dilution (with water).
DERIVATIZATION PROCEDURE
An aliquot of NIZ working solution equivalent to 25—300 ng/mL of NIZ, 100 ^L of carbonate buffer solution at pH 10.5 and 100 ^L of dansyl chloride solution were placed into 12 mL volumetric flasks. The solutions were left for 30 min at room temperature. The dansyl derivatives were extracted three times with 3 mL of chloroform after shaking for 1 min. The dansyl derivatives were collected in 10 mL calibrated flask and then were made to volume with chloroform. The derivatives fluorescence intensity was measured at 513 nm using ^ex 367 nm.
RESULTS AND DISCUSSION
Excitation and Emission Spectra
Dansyl chloride (Fig. 1) is widely utilized as a fluorescent derivatizing agent for amino compounds. NIZ was found to react with dansyl chloride to form a highly fluorescent derivative. The fluorescence exhibits its highest fluorescence intensities of excitation and emission wavelengths of 367 and 513 nm, respectively (Fig. 2).
<D O
§ 1000 <D
o Ë
500
0 220
A
452
684
(b)
, A
220
452
684
Wavelength, nm
Fig. 2. Emission spectra of the blank (a) and 200 ng/mL nizatidine-dansyl derivative in chloroform (b) at room temperature for 30 min.
OPTIMIZATION OF EXPERIMENTAL CONDITIONS
The experimental conditions for the derivatization reaction were optimized.
Effect of Dansy Chloride Concentration and Volume
The influence of the concentration of dansyl chloride was examined by addition of different volumes of 0.02% reagent in the range of 25—800 ^L. The maximum fluorescence intensity was obtained when 100 ^L dansyl chloride solution was utilized (Fig. 3).
Effect of pH
The pH dependence of the system was studied over the range of 9.0—11.0 using 0.1 M carbonate buffer. Maximum fluorescence intensity was obtained at pH 10.5 (Fig. 4).
Effect of Heating Temperature and Heating Time
In this study, the reaction between NIZ and dansyl chloride was performed at different temperatures (room temperature (20°C), 40°C, 50°C and 60°C) for various time intervals. As it is seen in Fig. 5 NlZ-dan-syl derivative was found that the reaction proceeds speedy at room temperature for 30 min.
OOTHKA H CnEKTPOCKOnHa tom 115 № 2 2013
NEW SPECTROFLUORIMETRIC METHOD
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Effect of Solvents
Fluorescence spectral characteristics of NIZ with dansyl chloride in different solvents were examined. Dichloromethane, ethylacetate and chloroform were tested for this investigation. Experimental results indicated that chloroform gave the maximum intensities and stable fluorescence emission among the studied solvents.
Effect of Time on the Stability
The NIZ-dansyl derivative was found to be stable for at least 2 hours at room temperature and 3 days at 4°C in chloroform.
Fluorescence intensity
Mikroliter added of 0.02% dansyl chloride
Fig. 3. Effect of the volume of dansyl chloride on the de-rivatization reaction.
METHOD VALIDATION
The validity of the proposed spectrofluorimetric method was tested regarding linearity, limit of detection, limit of quantification, accuracy, precision, recovery and robustness according to ICH Q2(R1) [26] recommendations.
Linearity
The calibration sets of NIZ-dansyl derivative with related compound were prepared in the concentration range NIZ 25—300 ng/mL. Then the calibration curves were constructed (fitting to the equation of F= = aC + b) by plotting concentration versus fluorescence intensities, where F is the fluorescence intensity, C is the concentration of the drug, a is the slope and b is the intercept of the calibration equation.
Limit of Detection (LOD) and Limit of Quantification (LOQ)
LOD and LOQ were calculated according to the following equation: LOD = 3.3ct/S and LOQ = 1Oct/S, where ct is the standard deviation of the intercept of regression line, S is the slope of the calibration curve. LOD and LOQ were calculated as 11.71 and 35.13 ng/mL, respectively.
Precision and Accuracy
Precision and accuracy of the method were investigated to determine intra-day and inter-day fluorescence intensity, expressed as relative standard deviation (RSD%) and relative mean error (RME%) for a series of measurements. Three different concentrations of the relevant drug were analyzed in six replicates during the same and three consecutive days. The relative mean error (%) and relative standard deviation (%) were found to be 1.2% and 0.79% indicating the high accuracy and precision of the spectrofluorimetric method.
ОПТИКА И СПЕКТРОСКОПИЯ том 115 № 2
Fig. 4. Effect of pH on the derivatization reaction.
Fluorescence intensity
Time, min
Fig. 5. Effect of heating temperature and heating time on the derivatization reaction: 1 - 20°C, 2 - 40°C, 3 - 50°C, 4 - 60°C.
Recovery
The recovery study was performed by adding known amounts of the related compounds to the known concentration of their commercial pharmaceutical capsule contents (standard addition method). Mean recovery was found to be 95.33 (Table 1).
2013
Table 1. Linearity range, regression equation, correlation coefficient, LOD, LOQ, intraday and interday variations (n = 3)
Parameters
Linearity range (ng/mL) 25-3GG
Regression equation F = 3.G9G4C + 63.227
Correlation coefficient G.9956
LOD (ng/mL) 11.71
LOQ (ng/mL) 35.13
Mean recovery (%) 95.33
Table l. Assay results of capsule
Axid (15G mg)
Mean 153.35
Recovery 1G2.23
SD 8.6 X 1G-2
Robustness
The robustness of the spectrofluorimetric method was evaluated by making small changes in the volume of dansyl chloride (100 ± 5 ^L), reaction time (30 ± 1 min), and volume of buffer (100 ± 5 ^L) and the effect of the changes were studied. It can be concluded that this method is robust, because slight variation of these experimental conditions have little or no effect on the results. Precision values expressed as RSD % (<0
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