Multiple baseline manipulation Composition of the but blank is changed to estimate different analytes form the mixture. The method has advantage over the Singular baseline manipulation (SBM) since it can be used for ternary mixtures of analytes. In these types of methods, the spectrum of sample-containing mixture of two drugs shows two different peaks, from which suitable analytical wavelength can be assigned to each analyte. This is free from inferences by other analytes in the mixture. Method validation The newly developed method was validated according to the ICH guidelines with respect to method sensitivity, linearity, range, accuracy, precision, robustness, and specificity. Linearity and method sensitivity Typically, the regression equations for the calibration curve was found to be y = �C0.
02938X + 0.691341 for DRT for a concentration range of 4�C20 ��g/mL and y = 0.066701X �C 0.39853 for ETR for a concentration range of 4.5�C22.5 ��g/mL. The correlation coefficient (��r��) values were >0.999 (n=6). In the test of linearity, i.e. Fischer variance ratio, calculated values of F (Fcal) for both the analytes were less than tabulated F values (FTab) which indicate linearity of response. Limits of detection (LOD) were found to be 0.42 ��g/mL for DRT and 0.55 ��g/mL for ETR. Limits of quantification (LOQ) were found to be 1.26 ��g/mL for DRT and 1.65 ��g/mL for ETR. Formulation analysis and accuracy studies Newly developed methodology was used to determine assay of the marketed tablets with present spectrographic conditions, and it was found to be accurate and reliable.
The average drug content was found to be 99.98% for DRT and 100.12% for ETR of the labelled claim. No interfering peaks were found in the spectrograph, indicating that the estimation of drug free from inference of excipients. The results of the recovery study were in the range of 98.40�C101.13% and % RSD was always less than 0.89. The results for formulation analysis and accuracy studies are presented in Table 1. Table 1 Optical characteristics of the method and results of formulation analysis, recovery study, and method sensitivity Precision Assay values during the precision study were in the range of 98.8�C100.7 and % RSD values were always less than 0.6. The results obtained for inter-day and analyst precision were statistically tested and presents in Table 2 for DRT.
The method precision is shown by results of the ANOVA test for both the analytes where calculated F (Fcal) values were always less than tabulated F (Ftab) values for both the analytes. Table 2 Results of precision study Robustness To determine the robustness of the method, the final experimental conditions were purposely altered and the results were examined. Effects of variation of experimental Cilengitide conditions were studied which shows that the assay and % RSD values were well within the limits [Table 3].