INTRODUCTION:

Dasatinib is anticancer agent used to treat cancers like chronic myeloid leukemia and acute lymphoid leukemia¹. It is an oral medication. Dasatinib is an inhibitor of multiple tyrosine kinases. Dasatinib is chemically designated as N-(2-Chloro-6-methylphenyl)-2-({6-[4-(2-hydroxyethyl)piperazin-1-yl]-2-methylpyrimidin-4-yl}amino)-1,3-thiazole-5 carboxamide. Its molecular formula is C₂₂H₂₆ClN₇O₂S.H₂O and its molecular weight is 488g/mol². Dasatinib is poorly soluble in water and freely soluble in acetonitrile, methanol, ethanol and DMSO³.

Fig 1: Chemical Structure of Dasatinib

The literature survey reveals that various analytical methods were reported for estimation of dasatinib (UV^3,4, RP-HPLC^5-8, UPLC⁹, HPTLC¹⁰). This work introduces a new UV-Spectrophotometric method for estimation of dasatinib in bulk and pharmaceutical formulation.

MATERIALS AND METHODS:

Chemicals and reagents:

Dasatinib was obtained as gift sample from Aadhar life Sciences, Solapur. Acetonitrile was obtained from Research Lab Fine CHEM Industries, Mumbai and Dasanat 70mg Tablets were purchased from local Market.

Instruments:

Analytical balance (Shimadzu AY220), Ultra Sonicator (OSCAR) and UV-Vis Double beam Spectrophotometer (Systronics) instruments were used.

Method:

Preparation of standard stock solution:

10mg of dasatinib weighed and transferred to 100ml volumetric flask, 50ml of acetonitrile was added, sonicated for 10mins and volume was made up to the mark with acetonitrile. (Conc. of Dasatinib: 100µg/ml)

Working Standard solution:

1.5ml of standard stock solution was transferred into 10ml volumetric flask, 5ml of acetonitrile was added and volume was made up to the mark with acetonitrile. (Conc. of Dasatinib: 15µg/ml)

Selection of analytical wavelength was done by scanning above solution in the range 200-400nm.

Preparation of sample solution:

Weigh accurately and finely powdered 10 tablets. Transfer the powder equivalent to 10mg of dasatinib into 100ml volumetric flask. Then add 50ml acetonitrile and sonicate for 10min, volume was make up to the mark with acetonitrile (100µg/ml). Pipette out 1.5ml of above solution in 10ml volumetric flask, add 5ml acetonitrile and make up the volume with acetonitrile (15µg/ml).

Method Validation:

The developed method was validated as per ICH guidelines^11,12. The parameters assessed were specificity, linearity, range, accuracy, precision (repeatability), system suitability, LOD and LOQ.

1. Linearity:

0.5, 1, 1.5, 2 and 2.5ml of standard stock solution (Conc. of Dasatinib=100µg/ml) were transferred to 10ml volumetric flask separately and the volume was made up to the mark with acetonitrile to obtain concentration of 5, 10, 15, 20 and 25µg/ml of Dasatinib. The solutions were prepared and analyzed at wavelength 315nm and absorbances were noted. Calibration curve of dasatinib was constructed by plotting the absorbance v/s conc. of dasatinib. The correlation coefficient (r²) of least square linear regression for dasatinib was calculated.

2. Range:

The range of analytical method was decided for dasatinib (5-25µg/ml).

3. Accuracy:

The accuracy was determined by calculating % recovery of dasatinib. It was carried out by adding known amounts of analyte corresponding to three concentration levels 80, 100and 120% and results were expressed as % recovery.

4. Precision:

The precision of analytical method was studied by performing repeatability. Repeatability studies were carried out by estimating responses of working standard solution (Conc. of Dasatinib: 15µg/ml) for 5 times. The results were reported in terms of percentage relative standard deviation (% RSD).

5. Robustness:

By deliberate change in wavelength that is 313nm and 317nm of concentration 15µg/ml of dasatinib was analyzed, in the same environmental condition.

6. Limit of Detection ( LOD):

LOD is the lowest amount of analyte in sample that can be easily detected but not necessarily quantified. LOD was calculated by following formula.

LOD = 3.3*σ /S

Whereas, σ = Standard deviation and S= Slope of regression coefficient.

7. Limit of Quantification (LOQ):

LOQ is the lowest amount of analyte in sample that can be easily detected and quantified with suitable precision and accuracy. LOD was calculated by following formula.

LOD = 10*σ /S

Whereas, σ = Standard deviation and S= Slope of regression coefficient.

Assay:

Assay of tablets (Sample solution) was also calculated.

RESULTS AND DISCUSSION:

The absorption spectrum shows λ_max of dasatinib at 315nm.

Fig 2: UV scan of Dasatinib

Method Validation:

1. Linearity:

The absorbance is proportional to the concentration and linear in the range of 5-25µg/ml (Tab: 1).The value of r² was 0.9992 which is well within acceptance limit (r²< 1) (Fig: 3).

Table 1: Linearity for Dasatinib

Concentration (µg/ml)	Absorbance
5	0.350
10	0.625
15	0.927
20	1.186
25	1.505

2. Range:

The range is from 80-120% of test concentration. (Dasatinib: 5-25µg/ml) (Fig: 3)

Fig 3: Linearity curve for Dasatinib

3. Accuracy:

The percentage recoveries of the results indicate that the recoveries are well within acceptance range (RSD<2), therefore method is accurate.

Standard absorbance: 0.927(Tab: 2)

4. Precision :

The % RSD < 2 values obtained shows that method developed is precise. (Tab: 3)

5. Robustness:

The deliberate change in wavelength, in the same environmental condition, gave the reliable results. (Tab: 4)

6. LOD:

LOD of dasatinib was found to be 0.908µg/ml.

7. LOQ:

LOQ of dasatinib was found to be 2.752µg/ml.

Assay:

Amount of drug present in marketed formulation Dasanat 70 Tablets (Nacto Pharma Ltd) was calculated. Amount was found to be 98.60% of labeled claim. (Absorbance= 0.914)

Table 2: Accuracy Results of Dasatinib

% Level	Reps	Spiked conc. (µg/ml)	Absorbance	Amount Recovered (µg/ml)	% Recovery	Average	Std. deviation	% RSD
80	1	11.96	0.7510	12.12	101.27	101.31	0.08	0.08
	2		0.7510	12.12	101.27
	3		0.7520	12.13	101.40
100	1	14.96	0.9270	14.96	100	99.93	0.12	0.12
	2		0.9250	14.92	99.78
	3		0.9270	14.96	100
120	1	17.95	1.0900	17.58	97.99	98.02	0.05	0.05
	2		1.0910	17.60	98.08
	3		1.0900	17.58	97.99

Table 3: Precision Results of Dasatinib

Reps	Absorbance
1	0.927
2	0.925
3	0.924
4	0.927
5	0.927
Avg	0.926
STDEV	0.001414
RSD	0.15

Table 4: Robustness Results of Dasatinib

Maximum wavelength (nm)	313	317
Absorbance	0.922	0.935
	0.923	0.935
	0.920	0.935
Average	0.921	0.935
STDEV	0.001	0
% RSD	0.165	0

Summary of Validation Parameters:

Table 5: Summary of validation parameters

Parameters	Values
Maximum wavelength (nm)	315
Beers law (µg/ml)	5-25
Regression equation	Y= 0.0574x + 0.0573
Slope	0.0574
Intercept	0.0573
Regression coefficient (r²)	0.9992
Repeatability(n=5) % RSD	0.15
LOD (µg/ml)	0.908
LOQ (µg/ml)	2.752

CONCLUSION:

An attempt has been made to develop UV- Spectrophotometric method for estimation of dasatinib in bulk and pharmaceutical formulation. Acetonitrile was used as solvent. The developed method has been validated according to ICH guidelines for linearity, range, accuracy, precision (repeatability), Robustness, LOD and LOQ parameters. The results showed that the developed method is simple, reliable, accurate, reproducible and robust. Therefore, the method can be applied for routine analysis of dasatinib in bulk and pharmaceutical formulation.

ACKNOWLEDGEMENT:

The authors are thankful to Aadhar Life Sciences, Solapur for providing gift sample of dasatinib API.

CONFLICT OF INTEREST:

The authors declare no conflict of interest.

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Received on 20.02.2021 Modified on 17.04.2021

Asian J. Pharm. Ana. 2021; 11(3):203-206.

DOI: 10.52711/2231-5675.2021.00036