Q analysis of Montelukast sodium and Fexofenadine
hydrochloride in Tablet Formulation by Derivative Spectrophotometry
Dr. Rajeev Kumar P1*, Rekha Rajeev Kumar2
1Department
of Pharmaceutics, School of Pharmacy, PRIST University, Manamai-Nallur, ECR,
Near Mahabalipuram, Chennai, Tamilnadu -603102.
2Department
of Pharmaceutical Analysis, School of Pharmacy, PRIST University,
Manamai-Nallur, ECR, Near Mahabalipuram, Chennai, Tamilnadu -603102
*Corresponding Author E-mail: rajeevmuthup@gmail.com
Objective:
The aim of this study is to present
the simple and sensitive method for the analysis of montelukast and
fexofenadine hydrochloride and apply the proposed method for the analysis of
said drugs in pharmaceutical dosage forms. Methods: Montelukast and
fexofenadine hydrochloride are used in combination for management of allergy.
The procedure of the first order derivative spectrophotometry allowed
simultaneous determination of montelukast sodium and fexofenadine hydrochloride
in fixed dose combination product. Results: The method employed first
order derivative spectroscopy for estimation of λmax by taking 10
μg/ml each of montelukast sodium and fexofenadine hydrochloride were
scanned in 200-400 nm range. The absorbance values at 340 nm and 212.6 nm of
the first derivative spectrum was used for the determination of montelukast and
fexofenadine hydrochloride respectively without related interference.
Conclusion: This method obeyed Beer’s law in the concentration range of
4-24 µg/ml for montelukast and 4-24 µg/ml for fexofenadine hydrochloride . The
results of analysis have been validated statistically and recovery studies
established the accuracy of the proposed method and low values of standard
deviation confirmed correctness of the used method. The method was validated as
per ICH guidelines.
KEYWORDS: Montelukast (MLKT), Fexofenadine hydrochloride (FXD),
Ultraviolet Spectrophotometry, Derivative Spectrophotometry.
INTRODUCTION:
Montelukast sodium (MTK),
1-[({(R)-m-[(E)-2-(7-chloro-2-quinolyl)
vinyl]-α-[o-(1-hydroxyl-1-methylethyl)phenethyl]benzyl}thio)methyl]cyclopropaneacetate
sodium [1,2] and shown in Figure 1. Montelukast (trade name Singulair)
is a leukotriene receptor antagonist (LTRA) used for the maintenance treatment
of asthma and to relieve symptoms of seasonal allergies. It is usually
administered orally.Leukotriene receptor
antagonist (LTRA) used
for the maintenance treatment of asthma and
to relieve symptoms of seasonal allergies [3].
Figure 1: Structure of Montelukast sodium.
Fexofenadine
[4-7], (Allegra, Telfast, Fastofen, Tilfur,
Vifas, Telfexo, Allerfexo) is an antihistamine drug used in the management of
hay fever and related allergy symptoms. It was developed as a beneficiary of
and substitute to terfenadine (brand names include Triludan and Seldane), an
antihistamine with possibly serious contraindications. α,
α-dimethyl-4-[1-hydroxy-4-[4-(hydroxydiphenylmethyl)-1-piperidinyl] butyl]
benzeneneacetic acid, is the most vital terfenadine metabolite shown in Figure
2.H1-receptor antagonist used
in patients with allergic rhinitis[8].Fexofenadine, like other second and
third-generation antihistamines, does not freely cross the blood-brain barrier,
and so causes less drowsiness than first-generation histamine-receptor
antagonists. It works by being an antagonist to the H1 receptor. It
has been termed as both second-generation and third-generation [3], [9].
Figure 2: Structure of Fexofenadine.
To the best of our knowledge no method is reported in
literature for simultaneous determination of Montelukast sodium and
Fexofenadine by high performance liquid chromatography (HPLC) using the
specified chromatographic condition. The survey of literature showed few UV
Spectrophotometric [10], HPLC [11-13], LC-ESI–MS/MS method [14], are available
for the estimation of Montelukast sodium in pharmaceutical preparation and in
biological fluids. Literature survey also showed that available for the
estimation of UV Spectrophotometric [15]-17] and HPLC [18] for the estimation
of Fexofenadine in pharmaceutical preparation. This paper presents simple,
rapid, reproducible and economical methods for the simultaneous analysis of Montelukast
sodium (MLT) and fenofibrate in tablet dosage forms. The proposed method was
successfully applied to a solid dosage form of Montelukast sodium and
Fexofenadine and analyzed in presence of commonly used tablet excipients. This
method can also be employed for quality control during manufacture of drug
product.
MATERIALS AND METHODS:
Material:
Montelukast sodium (MLKT) and fexofenadine
hydrochloride (FXD) were obtained as gift sample from Cadila pharmaceutical,
Ahmedabad and Ami Life science, Baroda, Gujarat. The marketed fixed dose
combination product (MONTAIR FX comprising 10 mg of MLKT and 120 mg of FXD) was
obtained from local market. HPLC grade acetonitrile, methanol and analytical
grade potassium dihydrogen ortho phosphate, ortho phosphoric acid,
triethylamine was procured from S.D. Fine chemicals Ltd. (Mumbai, India).
Hydrochloric acid, sodium hydroxide pellets and 3% v/v hydrogen peroxide
solution were acquired from Ranbaxy Fine Chemicals, New Delhi (India). High
purity water was obtained by Millipore Milli-Q plus purification system
(Millipore, Bedford, USA).
Equipment:
Shimadzu UV-1700; UV-visible spectrophotometer with
1cm matched quartz cells was used for the measurement of absorbance. Analysis
was achieved by direct method over a wavelength range from 200–400 nm. The
apparatus settings were zero order and first derivative mode and band width of
2 nm in the range of 200-400 nm. Shimadzu electronic balance was used for
weighing the sample. Class ‘A’ volumetric glassware were used.
Procedure:
Preparation
of Stock Solution:
Accurately
weighed 10mg of montelukast sodium equivalent to montelukast sodium and
fenofibrate hydrochloride was transferred to 100mL volumetric flask. Drug was
then dissolved and made up to the volume to 100mL with acetonitrile: buffer of
pH4.5: methanol (50:30:20) %v/v. It was further diluted to a concentration of
100µg/mL with solvent, which was then used as the stock solution for the further
dilutions.
Development
of the Method:
The correct dilutions of montelukast sodium and
fexofenadine hydrochloride were prepared separately with solvent acetonitrile:
buffer: methanol in the ratio of 50:30:20%v/v at a concentration of 1 mg/ml and
then it was diluted with sufficient quantity of solvent to make the
concentration of 12µg/ml. They were scanned in the wavelength range of 200-400
nm. Data were recorded at the interval of 1nm. Then the spectra of two drugs
were derivatised to obtain first derivative spectra. Wavelength was selected
where one drug showed zero crossing and other drug showed substantial
absorbance. The wavelength selected for montelukast sodium and fexofenadine was
340nm where fexofenadine hydrochloride has zero absorbance and the wavelength
selected for fexofenadine hydrochloride analysis was 212.6 nm where the
absorbance of montelukast sodium is zero.
Linearity:
Standard stock solutions were prepared by dissolving
100 mg of each drug sample using acetonitrile: buffer: methanol 50:30:20 %v/v
in 100 ml volumertic flask separately and the volume is made up with same
solvent to get the concentration of 1 mg/ml. From this, suitable dilution was
made in water to get the working standard solution of 4-24 µg/ml for montelukast
sodium and 4-24µg/ml for fexofenadine hydrochloride separately. The absorbance
of derivatised spectra was measured at 340 nm and 212.6 nm for montelukast
sodium and fexofenadine hydrochloride respectively. Six replicate of the
analysis were carried out. Absorbance Vs concentration were plotted to obtain
the calibration graph.
Limit of Detection and limit of Quantification:
LOD and LOQ were calculated from the data obtained
from the linearity studies (ICH guidelines)[19]. The slope of the linearity
plot was determined. For each of the six replicate determinations, y intercept
was calculated and standard deviation of the y intercept was computed. From the
values, LOD and LOQ were calculated as follows,
And 
Where, σ = standard deviation of response
S = average of slope
Analysis of Marketed Formulation:
Twenty tablets were weighed accurately and powdered. An
precisely weighed quantity of powder equivalent to 10 mg of MLKT and FXD was
taken in a 100 ml volumetric flask and dissolved the drug in acetonitrile:
buffer: methanol 50:30:20 %v/v as solvent by sonication. Then the solution is
prepared up to the volume with the solvent and filtered. This solution
comprises 10μg/ml of montelukast sodium and 12 μg/ml of fexofenadine
hydrochloride sample solution. The mixed sample solutions were analyzed to
obtain spectra’s and absorbance values were measured at selected wavelengths
for montelukast sodium and fexofenadine hydrochloride.
Recovery Studies:
To determine the accuracy of the method, recovery
studies were achieved using the method of addition. Recovery studies were
approved by adding a known amount of standard to the pre analyzed sample. The
tablet analysis obtained by suggested method was validated by statistical
evaluation and data are showed in results.
Validation of Spectrophotometric Method:
All
the validation parameters were calculated same as pronounced in methods and
data attained by linearity, sensitivity, precision, accuracy and ruggedness
studies were shown in results[20-25].
RESULTS AND DISCUSSION:
UV
spectrum of both the drugs (MLKT and FXD) were derivatised to first order with Dl = 1 for the
entire spectrum. Zero crossing points for MLKT and FXD was found to be 340 and
212.6 nm respectively. Figure 3 shows overlain first derivative spectra of MLKT
and FXD at a concentration of 12µg/ml each.
Figure
3. Zero Crossing Point for RC at 233.5 nm and Fenofibrate at 254 nm
The
absorption in first derivative mode of both MLKT and FXD at respective selected
wavelength 340nm and 212.6 nm were linear in concentration range of 4-24 µg/ml
and 4-24 µg/ml for MLKT and FXD, respectively shown in figure 4 and 5.
Fig
4. Calibration curve for MLKT at
340 nm by First order derivative spectroscopy.
Fig 5. Calibration
curve for FXD at 212.6nm by First order derivative spectroscopy.
The
R2 value was found to be 0.9998 for MLKT and 0.9997 for FXD.As per
ICH guidelines, LOD and LOQ can be determined using visual evaluation, signal
to noise ratio or from slope of linearity plot and standard deviation. Visual
evaluation may be used in non instrumental methods and signal to noise ratio is
normally possible with chromatographic methods. Hence, the method based on
determination of slope of linearity plot and standard deviation of y intercept
of linearity was used for the determination of LOD and LOQ and found to be
0.312µg/ml and 0.105µg/ml for MLKT and 0.512µg/ml and 0.578µg/ml for FXD,
respectively illustrated in Table 1.
Commercial
formulation containing MLKT and FXD were analyzed by proposed method. Six
replicate analysis of formulation were carried out and the mean MLKT content
was 9.92 mg/tablet and the mean content of FXD, was 119.79 mg/tablet. The
corresponding standard deviation was found to be 0.630 for MLKT and 0.640 for
FXD indicating that the method has required precision. The accuracy of the
method was determined by recovery studies. MLKT and FXD were added to pre
analyzed tablet powder at four different levels viz., 50%, 100% and 200%.Six
replicate analysis was carried out at each level. The recovery was 98.90 for
MLKT and 99.65% for FXD indicating that the method has required accuracy. To
study the robustness of the method, analysis of formulation was carried out on
three different days. The % RSD for inter day and intraday is varying from
0.202 to 0.164 and 0.200 to 0.139 for MLKT and FXD, respectively. The validation
results are given in Table 2.
Table:
1. Optical characteristics and
statistical data of the regression equation by first order derivative
spectroscopy.
|
Parameter
|
Montelukast sodium (MLKT)
|
Fexofenadine
hydrochloride(FXD)
|
|
Detection wavelength (nm)
|
340 nm
|
212.6 nm
|
|
Beer’s law limits (µg/ml)
|
4-24
|
4-24
|
|
Molar extinction coefficient
(L mol-1 cm-1)
|
0.144 × 103
|
0.252 × 103
|
|
Regression equation (y*)
|
y = 0.0009 x + 0.0004
|
y = 0.0016 x + 0.0002
|
|
Slope (b)
|
0.0009
|
0.0016
|
|
Intercept (a)
|
0.0004
|
0.0002
|
|
Correlation coefficient (R2)
|
0.9998
|
0.9997
|
|
Limit of detection (µg/ml)
|
0.312
|
0.512
|
|
Limit of quantification
(µg/ml)
|
0.105
|
0.578
|
*y
= b x + a where x is the concentration of montelukast sodium and Fexofenadine
hydrochloride in µg/ml and y is the absorbance at the respective wavelength.
Table
2: Analysis of formulation and recovery studies
|
Dugs
|
Labeled amount (mg)
|
*Amount found (mg)
|
%label claim
|
*%Recovery
|
*Precision# (%RSD)
|
|
Inter day
|
Intraday
|
|
MLKT
|
10
|
9.92
|
99.20
|
98.8±0.896
|
0.200
|
0.202
|
|
FXD
|
120
|
119.79
|
99.78
|
99.7±0.652
|
0.139
|
0.164
|
*Each
value is a mean of six observations.
CONCLUSION:
Thus
the developed method is simple, accurate and precise and can be used for
routine analysis of MLKT and FXD simultaneously form combined dosage forms.
ACKNOWLEDGEMENT:
The
authors wish to thank School of Pharmacy, PRIST University, Chennai, Tamilnadu
for providing necessary facilities and to carry over the work. The authors wish
to express their gratitude to Ami Life Science, Baroda, Gujarat and Cadila
pharmaceutical, Ahmedabad for providing the authentic sample (Montelukast and
Fexofenadine).
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