5.1. Linearity:
5.1.2. Method A: At the described wavelength linear relationship was obtained between the trough amplitude and the dinitolimide concentration in the range (10–150µg/ml). The linear regression equation of the trough was:
A280.4= 0.0013 C + 0.0016 (r = 0.9999)
Where A is a trough height at 280.4 in cm and C is the drug concentration in (µg /ml). This method determine the drug in presence of up to 93.34% of the acidic degradate as shown in Table (4).
5.1.3. Method B: Under the described experimental conditions, the calibration graph for the method was constructed by plotting trough height versus concentration in (µg /ml). The regression plot was found to be linear over the range of 10–150µg/ml. The linear regression equation for the graph is:
P 296.4 nm= 0.0034 C + 0.0012 (r = 0.9998)
Where C is the concentration of dinitolimide in (µg /ml), P is the trough height of the first derivative of the ratio spectrum curve at 296.4 nm and r is the correlation coefficient. This method determine the drug in presence of up to 70% of the acidic degradate as shown in Table (4).
5.1.4. Method C: Linear correlation was obtained between the differences in amplitudes at 262.0 and 301.0nm, against the corresponding concentration of dinitolimide. Good linearity is obtained in the concentration range of (10–150µg/ml). The corresponding regression equation was computed to be:
ΔP262.0 – 301.0 = 0.0384 C + 0.1214 (r = 0.9997)
Where ΔP is the amplitude difference at the selected wavelengths, C is the concentration in (µg /ml) and r = the correlation coefficient. This method determine the drug in presence of up to 70% of the acidic degradate as shown in Table (4).
5.1.5. Method D: At the described wavelength linear relationship was obtained between the trough amplitude and the corresponding concentration of dinitolimide. Good linearity is obtained in the concentration range of (10–150µg /ml)) dinitolimide. The corresponding regression equation was computed to be:
A 290 = 0.0071 C − 0.0059 (r = 0.9998)
Where A is the peak amplitude at 290nm, C is the concentration in (µg /ml) and r = the correlation coefficient, as shown in (table 2). This method determine the drug in presence of up to 70% of the acidic degradate as shown in Table (4).
5.1.6. Method E: The calibration curves were plotted over a concentration range of 10–150 µg/ml for dinitolimide.
ΔP269.0 – 280.0 = 0.0013 C + 0.0068 (r = 0.9997)
Where ΔP is the amplitude difference at the selected wavelengths, C is the concentration in (µg/ml) and r = the correlation coefficient. This method determine the drug in presence of up to 80% of the acidic degradate as shown in Table (4).
5.2. LOD and LOQ:
The limit of detection (LOD) and the limit of quantitation (LOQ) were calculated according to ICH Q2 Recommendation [22, 23] from the following equations:
LOD = 3.3 Sa / slope
LOQ = 10 Sa / slope
Where Sa is the standard deviation of the intercept of regression line.LOD was found to be 2.1570, 1.1633, 3.0958, 1.4527, 2.6570 and 1.3026 µg/ml, while LOQ was found to be 7.0516, 3.5251, 9.3811, 4.4022, 8.0516 and 3.9474 µg/ml for method A ,B,C,D,E and D respectively .The small values of LOD and LOQ indicate good sensitivity.