UV spectrophotometric method for determination of Bilastine in bulk and pharmaceutical formulation

 

Peethala Prathyusha, Raja Sundararajan*

Department of Pharmaceutical Analysis & Quality Assurance, Gandhi Institute of Technology and Management,

GITAM Institute of Pharmacy, GITAM (Deemed to be University), Visakhapatnam, Andhra Pradesh-530045

*Corresponding Author E-mail: sraja61@gmail.com

 

ABSTRACT:

Three simple, fast and economic UV-spectrophotometric methods were developed for the estimation of Bilastine in bulk and pharmaceutical formulation. For zero order (D0) λmax was measured at 281.60 nm. For first order method (D1) amplitude was measured at 280.20 nm (maxima) and 283.60 nm (minima). Bilastine obeyed Beer’s-Lambert’s law in the concentration range of 10-140 µg/ml for both zero order and first order methods. The % RSD value for both intra-day and inter-day precision was less than 2%. The methods were validated as per ICH guidelines for linearity, accuracy, precision and were found to be linear, accurate and precise.

 

KEYWORDS: Spectrophotometric method, Bilastine, Beer’s-Lambert’s law, ICH guidelines.

 

 


INTRODUCTION:

Bilastine is a new highly selective peripheral histamine H1- receptor antagonist, chemically named as 2-[4-(2-{4-[1-(2-ethoxyethyl)-1H-1, 3-benzimidazol-2-yl] piperidin-1-yl} ethyl) phenyl]-2-methylpropanoic acid (Figure 1).

 

Bilastine is a white crystalline powder with molecular formula C28H37N3O3, molecular mass of 463.61g/mole and melting point greater than 1950 C 1. It belongs to piperidine antihistamine class of drugs. It is a H1 receptor inverse agonist like other antihistamines 2, used for treating allergic disorders such as rhinoconjunctivitis and urticarial 3. Histamine plays a major role in the allergic reaction and is released by mast cell degranulation4. This histamine binds with H1 receptors, activates the receptors and causes allergic reactions. Bilastine binds with H1 receptor and prevents the activation of H1 receptor by histamine. Thus, it acts as an antagonist for histamine. Bilastine shows no cardiotoxic, sedative side effects and undergoes minimal or no first pass metabolism 5. It has less chance to undergo drug-drug interactions. Therefore, it is useful for treating patients suffering with renal/ hepatic dysfunction 6.

 

 

Bilastine, a piperidine class antihistamine medication used for the treatment of allergic rhinitis and chronic urticaria. From the review of literature, it was found that very few methods such as LC-MS/MS 7, HPLC-fluorescence 8 in biological sample, RP-HPLC 9, HILIC 10 and UV- spectrophotometry 11 are available for estimation of Bilastine. The aim and objective of the present work was to develop and validate as per ICH guidelines 12 a simple, fast, accurate, precise, economic and sensitive method for estimation of Bilastine using UV- spectrophotometry, in both bulk and pharmaceutical formulation, which can be used for routine analysis in QC laboratories.

 

MATERIALS AND METHODS:

Chemicals

Bilastine of reference standard was obtained as gift sample from Spectrum Pharma Research Solutions (Hyderabad) and Bilasure (20mg) pharmaceutical formulation from local pharmacy manufactured by Sun Pharmaceuticals Private Limited. Methanol (AR) grade, NaOH pellets and HCl were obtained from Thermo Fisher Scientific India Private Limited. Potassium dihydrogen phosphate was obtained from Finar Limited. Throughout the analytical process distilled water was used.

 

Instrumentation

A Shimadzu double beam UV spectrophotometer (UV-1800) along with an even set of 10 mm quartz cuvettes were used for measurement of absorbance. All analytical weight measurements were done on a Shimadzu model electronic balance. A computer software UV solutions 2.42 was used. All the glassware used in the study was of borosilicate glass. Microsoft excel was used for all statistical calculations.

 

Preparation of drug stock solution

25 mg of Bilastine was weighed and dissolved using few ml of methanol in 25 ml volumetric flask. The volume was made upto the mark with methanol to produce 1000µg/ml solution.

 

Preparation of 0.1M NaOH

1g of Sodium hydroxide was dissolved in 200 ml of distilled water in a 250 ml volumetric flask and was made upto the mark with distilled water.

 

Preparation of phosphate buffer (pH 2.0)

0.136 g of Potassium dihydrogen phosphate was dissolved in 800 ml of distilled water and the pH was adjusted to (pH 2.0) with HCl and sufficient amount of distilled water was added to produce 1000 ml.

 

Zero order (D0) and first order(D1) UV-spectrophotometric methods were developed using 0.1M NaOH, Phosphate buffer (pH 2.0) and distilled water.

 

Preparation of calibration curve

Twenty four numbers of volumetric flasks (10 ml each) were taken and divided into 3 sets thus each set containing eight flasks. From the drug stock solution, 0.1, 0.2, 0.4, 0.6, 0.8, 1.0, 1.2 & 1.4 ml samples were pipetted thrice and transferred into the 10 ml volumetric flasks to produce 10-140 µg/ml. First set of stock solutions were diluted with 0.1M NaOH, second set with phosphate buffer (pH 2.0) and the third set with distilled water upto the mark. All the three sets of solutions were scanned from 200-400 nm UV range using UV- double beam spectrophotometer.

 

Assay procedure

20 commercial Bilastine tablets were weighed and triturated into powder. Tablet powder weight equivalent to 10 mg was taken into a 10 ml volumetric flask, dissolved in methanol, sonicated for 10 minutes and filtered using whatman filter paper. The filtered solution was made upto the volume with methanol. Aliquots of sample solutions were taken in triplicate and were made upto the volume separately with 0.1 M NaOH, phosphate buffer (pH 2.0) and distilled water. Absorbance was measured at 281.60 nm for each sample.   

 

Method validation:

Linearity, Accuracy and Precision

Bilastine standard solutions were prepared in the concentration range of 10-140 µg/ml in 0.1 M NaOH, phosphate buffer (pH 2.0) and distilled water. The absorbance of each solution was measured and calibration curves were established by plotting absorbance versus concentration graph.

 

Accuracy of the methods was analyzed by the percentage recovery of the standard drug that was added to the fixed concentration of sample solutions. The study was carried out by adding three different percentage levels of standard drug i.e. 50%, 100%, 150% to the sample concentration. Each solution was prepared in triplicate and the absorbance was noted to find out the percentage recovery.

 

Precision means repeatability of results for a particular method. Bilastine stock solutions (1.0, 2.0, 4.0ml) were taken in 10 ml volumetric flask and were made upto the mark with respective diluents. The absorbance of these solutions were measured individually thrice within a day and the absorbance was noted (Intraday precision). Similarly, the absorbances of these solutions were individually measured thrice in three days and recorded (Interday precision).

 

RESULTS AND DISCUSSION:

The developed methods were validated as per ICH guidelines. Three UV- spectrophotometric methods were developed using 0.1M NaOH, phosphate buffer (pH 2.0) and distilled water. The λmax of drug was observed at 281.60 nm in zero order method (D0). The spectra produced for the above methods were derivatized to first order spectra. The maximum and minimum absorbance of Bilastine was measured at 280.20 nm and 283.60 nm, respectively. The methods obeyed Beer’s-Lambert’s law in the concentration range of 10-140µg/ml in both zero order and first order method. The regression equation of drug in 0.1 M NaOH was found to be: y = 0.0178x + 0.0252 (R2=0.9991) for zero order and y = 0.0062x + 0.0106 (R2=0.9994) for first order. The regression equation of drug in Phosphate buffer (pH 2.0) was found to be: y = 0.0124x + 0.007 (R2=0.9995) for zero order and y = 0.0043x - 0.0023 (R2=0.9993) for first order. Similarly, the regression equation of drug in distilled water was found to be: y = 0.0142x + 0.0153 (R2=0.9994) for zero order and y = 0.0049x + 0.0068 (R2=0.9992) for first order. Overlain spectra and calibration graphs of the methods were shown in Figure 2-3.  The RSDs were found to be less than 2% which specifies that the methods are accurate. Table 1 shows the results of %recovery along with their RSDs for accuracy studies. Mean of intra-day and inter-day precision were found to be in between 98-101%. The RSDs were found to be less than 2% which indicates the methods are precise. Table 2-4 shows the results of %recovery along with their RSDs for precision studies.  A summary of all validation parameters were depicted in Table 5.


 

 

                                               

Table 1a: Accuracy data of Bilastine in 0.1M NaOH

% Level of recovery

Drug in tablet (µg/ml)

Pure drug added

(µg/ml)

Drug recovered

(µg/ml)

*Recovery (%) ± SD

RSD (%)

D0

D1

D0

D1

D0

D1

50%

10

5

14.95

14.83

99.3 ± 0.46

98.8 ± 0.64

0.46

0.65

100%

10

10

19.98

19.74

99.6 ± 0.77

98.3 ± 0.84

0.77

0.85

150%

10

15

25.13

24.99

100 ± 0.98

99.64 ± 1.06

0.98

1.07

*Mean of three determinations at each level

 

Table 1b: Accuracy data of Bilastine in Phosphate (pH 2.0)

%Level of recovery

Drug in tablet (µg/ml)

Pure drug added

(µg/ml)

Drug recovered

(µg/ml)

*Recovery (%) ± SD

RSD (%)

D0

D1

D0

D1

D0

D1

50%

10

5

15.09

15.02

100.58 ± 0.55

100.13 ± 0.49

0.55

0.49

100%

10

10

19.78

19.76

98.8 ± 0.31

98.8 ± 0.36

0.32

0.37

150%

10

15

25.02

25.07

99.93 ± 0.72

100 ± 0.69

0.72

0.69

*Mean of three determinations at each level

Table 1c: Accuracy data of Bilastine in Distilled water

%Level of recovery

Drug in tablet (µg/ml)

Pure drug added

(µg/ml)

Drug recovered

(µg/ml)

*Recovery (%) ± SD

RSD (%)

D0

D1

D0

D1

D0

D1

50%

10

5

15.29

14.89

101.3 ± 0.55

99.28 ± 0.46

0.54

0.46

100%

10

10

19.96

19.96

99.8 ± 0.26

99.83 ± 0.30

0.26

0.30

150%

10

15

24.96

24.96

99.64 ± 0.61

99.78 ± 0.72

0.61

0.73

*Mean of three determinations at each level

Table 2: Precision results of Bilastine in 0.1M NaOH  

Conc.

(µg/ml)

Intra-day precision

Inter-day precision

Amount found (µg/ml) (n=3)

*Mean ± SD, RSD (%)

Amount found (µg/ml) (n=3)

*Mean ± SD, RSD (%)

D0

D1

D0

D1

D0

D1

D0

D1

20

20.04

19.8

100.4 ± 0.23, 0.22

99.3 ± 0.25, 0.25

19.93

19.91

99.3 ± 0.38, 0.38

99 ± 0.27, 0.27

40

39.72

40

99 ± 0.68, 0.68

100 ± 0.4, 0.4

40.58

40.35

101 ± 0.54, 0.53

101 ± 0.52, 0.51

80

79.88

79.75

99.69 ± 0.33, 0.33

99 ± 0.36, 0.36

80.43

80.22

101.08 ± 0.93, 0.92

100.55 ± 0.85, 0.85

*Mean of three determinations at each level

Table 3: Precision results of Bilastine in Phosphate (pH 2.0)

Conc.

(µg/ml)

Intra-day precision

Inter-day precision

Amount found (µg/ml)(n=3)

*Mean ± SD, RSD (%)

Amount found (µg/ml)(n=3)

*Mean ± SD, RSD (%)

D0

D1

D0

D1

D0

D1

D0

D1

20

19.92

19.97

99.2 ± 0.32, 0.32

99.57 ± 0.15, 0.15

19.89

20.04

98.96 ± 0.18, 0.18

100.4 ± 0.14, 0.14

40

39.86

39.39

99.33 ± 0.23, 0.23

98.73 ± 0.34, 0.34

39.84

40.34

99.2 ± 0.23, 0.23

101.71 ± 0.61, 0.60

80

79.40

79.52

98.97 ± 0.42, 0.42

98.8 ± 0.52, 0.52

79.58

80.21

98.95 ± 0.52, 0.52

100.54 ± 0.69, 0.68

*Mean of three determinations at each level

Table 4: Precision results of Bilastine in Distilled water

Conc. (µg/ml)

Intra-day precision

Inter-day precision

Amount found (µg/ml)(n=3)

*Mean ± SD, RSD (%)

Amount found (µg/ml)(n=3)

*Mean ± SD, RSD (%)

D0

D1

D0

D1

D0

D1

D0

D1

20

19.86

19.86

99 ± 0.20, 0.21

98.6 ± 0.23, 0.23

19.99

19.87

99.96 ± 0.2, 0.2

98.7 ± 0.25, 0.25

40

39.88

39.65

99.43 ± 0.19, 0.19

98.26 ± 0.11, 0.11

39.64

39.65

98.2 ± 0.45, 0.46

98.26 ± 0.59, 0.60

80

80.18

79.90

100.43± 0.41,0.41

100.54 ± 0.54, 0.54

80.20

79.90

100.5 ± 0.61, 0.61

100.54 ± 0.73, 0.73

*Mean of three determinations at each level

Table 5: Summary of validation parameters/ Optical characteristics of Bilastine

Parameters

 

Obtained values

0.1M NaOH

0.1M NaOH

Phosphate buffer (pH 2.0)

Phosphate buffer (pH 2.0)

Water

Water

D0

D1

D0

D1

D0

D1

λmax /Maxima-Minima (nm)

281.60

280.20-280.60

281.60

280.20-280.60

281.60

280.20-280.60

Beer’s Law limit (µg/ml)

10-140

10-140

10-140

10-140

10-140

10-140

Molar absorptivity (l mol-1 cm-1)

68622.6249

24057.148

46519.554

15799.829

54365.933

18785.478

Sandell’s sensitivity (μg/cm2 /0.001)

0.053

0.151

0.079

0.244

0.062

0.185

Intercept(c)

0.0252

0.0106

0.007

0.0023

0.0153

0.0068

Slope (m)

0.0178

0.0062

0.0124

0.0043

0.0142

0.0049

Correlation coefficient (r2)

0.9991

0.9994

0.9995

0.9993

0.9994

0.9992

Accuracy ( %RSD)

0.46-0.98

0.65-1.07

0.32-0.72

0.37-0.69

0.26-0.61

0.30-0.73

Precision (%RSD)

Intra-day (n=3)

0.22 - 0.68

0.25-0.4

0.23-0.42

0.15-0.52

0.19-0.41

0.11-0.54

Precision (%RSD)

Inter-day (n=3)

0.38-0.92

0.27-0.85

0.18-0.52

0.14-0.68

0.19- 0.60

0.25-0.73

LOD (µg/ml)

0.079

0.298

0.095

0.334

0.069

0.206

LOQ (µg/ml)

0.240

0.903

0.246

1.013

0.211

0.624

Assay (%)

98.2

99.1

99.7

 

 


CONCLUSION:

Three simple, fast, accurate, precise, economic and sensitive UV-spectrophotometric methods were developed for the estimation of Bilastine in both bulk and pharmaceutical formulation. Therefore, the methods can be successfully employed for routine analysis of Bilastine in QC laboratories.

 

ACKNOWLEDGEMENTS:

The authors are thankful for Spectrum Pharma Research Solutions Hyderabad, Telangana, India and the management of GITAM (Deemed to be University), Visakhapatnam, Andhara Pradesh, India, for providing necessary facilities to carry out the research work.

 

CONFLICT OF INTEREST:

There is no conflict of interest.

 

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Received on 22.01.2020            Modified on 05.02.2020

Accepted on 10.02.2020           © RJPT All right reserved

Research J. Pharm. and Tech 2020; 13(2):933-938.

DOI: 10.5958/0974-360X.2020.00176.6