INTRODUCTION:

Rupatadine Fumarate, is chemically 8-chloro-6,11-dihydro-11-[-1-[(5-methyl-3-pyridyl)methyl]-4-piperidylidine]-3H-benzo[5,6]cyclohepta[1,2-b]pyridine Fumarate. Rupatadine Fumarate is a non-sedating H1- Antihistamine (second generation) and platelet activating factor receptor. It is potent and orally active that was developed as a therapeutic agent for the treatment of seasonal allergic rhinitis and chronic idiopathic urticaria¹.

A survey of literature revealed that very few analytical methods for this drug are available in human plasma and pharmaceutical formulations. These include HPLC^{2, 3} and LC-MS/MS⁴ methods. But there is no evidence in the literature for estimation of this drug by UV-spectrophotometer method, which is essential for routine quality control analysis of pharmaceutical products containing Rupatadine Fumarate as a fast, selective and economical method, so all attempts have been made to develop a simple, rapid and reproducible UV spectrophotometer method with greater precision, accuracy for analysis of Rupatadine Fumarate in bulk.

EXPERIMENTAL:

A CAMAG HPTLC system comprising of CAMAG Linomat IV semiautomatic sample applicator, CAMAG TLC scanner 3, CAMAG twin trough chamber(10 x 10 cm), CAMAG CATS 4 software, Hamilton syringe (100µL) were used during the study.

Tablets were purchased from local market. Acetonitrile and Water of HPLC grade (E.Merck India Ltd.) were used for preparing the mobile phase⁵.

Chromatographic conditions: Following chromatographic conditions were uniformly followed in the experiment.

Stationary Phase: HPTLC precoated Silica gel G60 F₂₅₄(Merck)

Size: 10 x 10 cm

Mode of application: Band

Band size: 4.0 mm

Separation technique: Ascending

Temperature: Ambient

Saturation time: 15 min.

Detection: UV

Scanning wavelength: 263 nm

Scanning mode: Absorbance/Reflectance

Slit dimension: 3 x 0.45 nm.

Linearity of detector response:

Aliquots of working standard solution (2, 3,4,5,6 μL) of Rupatadine Fumarate were spotted as sharp bands on the precoated TLC plate, using Camag linomat IV semiautomatic applicator under nitrogen stream. The plate was developed under chromatographic conditions mentioned above. The plate was removed from the chamber and dried in hot air dryer. Densitometric measurements were performed at 263 nm⁶ in absorbance mode. Data peak height and peak area of each band were recorded. The calibration curve was prepared by plotting peak area vs. concentration corresponding to each spot. The Densitogram of Rupatadine Fumarate was presented in figure – 1.

Assay:

Stock solution A: An accurately weighed quantity of Rupatadine Fumarate (10mg) was transferred into a 10ml volumetric flask. It was dissolved and diluted up to the mark with methanol to give a standard stock solution of 1 mg/ml. This 1mg/ml solution can be used as a working standard solution.

Preparation of sample solution:

25 tablets were accurately weighed and average weight was calculated. Accurately weighed quantity of tablet powder equivalent to 10mg of the drug was transferred to 10ml volumetric flask. To it 8ml of methanol was added and shaken for 10 min and the volume was adjusted upto the mark with methanol and then filtered through Whatmann filter paper no.40. This solution is used as the sample solution.

On the HPTLC⁷ plates spots of the standard and sample were applied. The plates were developed and after development the bands of the drugs were scanned at 263 nm. The peak height and area of the standard and sample bands were compared to obtain the concentration.

Method Validation:

Accuracy: Accuracy⁸ of the method was ascertained by performing the recovery studies using standard addition method. To a fixed amount of preanalysed drug were added at different levels. The total amount of the drug was determined by the above proposed method and the amount of pure drug was calculated. The average percent recovery was found to be nearly 100 % (Table-3).

Precision: Precision⁸ of the analytical method was expressed as SD or %RSD of series of measurements by replicate estimation of the drugs by the proposed method (Table -1).

Ruggedness: It was ascertained by analyst to analyst⁹ variation. The result was presented in Table -1

Table-1: Validation parameters of Rupatadine Fumarate

Parameters	Rupatadine Fumarate
λ_max(nm)	263
Beer’s law limit(µg/mL)	2-20
Limit of Detection (LOD)	21 ng
Limit of Quantification (LOQ)	53 ng
*Regression equation(Y)**
Slope(b)	683.1
Intercept(a)	328.4
Correlation coefficient(r)	0.997
Intra day %RSD **	0.05471
Inter day % RSD	0.07624
Analyst to analyst % RSD	0.04912

*Y = a + bx, where ‘Y’ is the absorbance and x is the concentration of the drug in µg/mL

**For six replicates

RESULTS AND DISCUSSION:

Rupatadine Fumarate was completely extracted from the tablet matrix with methanol. Combination of Acetonitrile: Water: Formic acid (50:50:3) offered optimum migration and resolution of Rupatadine Fumarate from other components of the formulation matrix.

The amount of Rupatadine Fumarate in the formulation¹⁰ was calculated on applying suitable dilution factor and comparing peak height and peak area of the standard and sample solutions. The content of the drug in the formulation was found to be within the limits. (Table - 2)

Table 2 - Estimation of Rupatadine Fumarate in formulation

S. No	Label Claim (mg)	Amount Estimated (mg)	%Label Claim
1 2 3	10 10 10	9.98 9.99 9.98	99 99.5 99
Mean*		99.1
Standard deviation*		0.2888
% RSD*		0.2912
Standard error*		0.1666

* Average of three determinations

Table 3 - Recovery studies

S. No	Amount Present (µg/µl)	Amount Added (µg/µl)	Amount Recovered (µg/µl)	% Recovery
1 2 3	10 10 10	5 10 15	15.1 19.9 25.1	100.1 99.1 99.4
Mean				100.03
Standard deviation				0.11547
% RSD				0.11505

The linearity of response was found to be in the range of 2-20µg/ml. Also the percent recovery values were found to be within the limits. Lower values of intra-day and inter-day variation on the analysis indicate that the method is precise. Different validation parameters for the proposed HPTLC method have been summarized in Table-1.

CONCLUSION:

The proposed HPTLC method was found to be simple, specific, precise and accurate. The sample recoveries in the formulations were in good agreement with their respective label claims. Hence this method can be conveniently adopted for the routine analysis of Rupatadine Fumarate in pure form and in its dosage form.

REFERENCES:

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