The peak purity was determined by ProQuant software. Limit of detection and limit of quantitation The LOD and LOQ were obtained by calculating selleck chem Brefeldin A using the standard formula as per ICH guidelines. LOD = 3.3��/s and LOD = 10��/s. Where, �� is standard deviation of response and S is the slope of calibration curve. RESULTS Development of the optimum mobile phase TLC procedure was optimized with a view to develop a stability-indicating assay method. The standard of drug was spotted on the TLC plates and developed in different systems. Different mobile phase were tried to resolve drug and degradation products. The optimum result was obtained with Toluene : Chloroform : Methanol in the ratio of 5 : 5 : 1.5 v/v. The chamber was saturated with mobile phase at room temperature. Developed mobile phase gives Rf of ITZ of 0.
52+0.02. The representative chromatogram is given in Figure 2. Figure 2 Representative densitogram of ITZ (6000 ng/spot) Validation of developed stability-indicating method Linearity The response of the drug was found to be linear in the concentration range of 1 000 to 6 000 ng/band for ITZ with correlation coefficient of 0.9978. The linear regression equation obtained was y = 0.164 (x) + 28.732. The representative chromatogram is given in Figure 3. Figure 3 Overlain densitogram of ITZ for concentration of 1000-6000 ng/spot Precision The relative standard deviation value for intraday precision study was found to be 0.51% and for interday precision was found to be not more than 0.34% for ITZ. This reveals that method is precise. Accuracy Good recoveries between 98.
86 and 100.43% were obtained at each level of added concentrations. The results obtained (n = 3 for each 80%, 100%, 120% level) indicated as the mean recovery were between 98 to 102% for ITZ. Limit of detection The LOD as calculated by standard formula as given in ICH guidelines was found to be 180.29 ng/band. Limit of quantitation The LOQ as calculated by standard formula as given in ICH guidelines was found to be 546.34 ng/spot. Specificity The specificity of the method was ascertained by peak purity profiling studies. The purity of drug peak was ascertained by analyzing spectrum at peak start, peak end, and peak max, which showed no interference of any other excipients or impurities in peak of ITZ.
Analysis of marketed formulation Experimental results from analysis of the amount of ITZ in capsules were in good agreement with the label claims, suggesting that there was no interference from any of the excipients normally present in the capsules. The drug content was found to be 99.86 �� 0.28%. ITZ capsules were analyzed using the proposed procedure; the results obtained are summarized in Table 1. Table 1 Results of analysis of pharmaceutical Drug_discovery formulation The validation summary is given in Table 2.