Intra day precision of a method was the study of repeatability of the results. The repeatability was determined by injecting working standard (10 μg/mL) solution of famotidine five times, chromatograms were obtained, and the % RSD of the area of five replicates was calculated and found to be 0.9%. The intermediate precision of the method was the study of reproducibility of the results in different days and was determined on five replicates from same lot by spiking. The %RSD of the area of five chromatograms was evaluated and found to be 0.90%. The results thus obtained were shown in Table 1 and present within the acceptance Stem Cells antagonist criterion of NMT 2% RSD

To determine the linearity of the proposed method, a series of seven different concentrated solutions of the standard FMD were prepared and about 6 μL of each solution was injected in duplicate into the HPLC system, chromatograms were recorded under the optimum chromatographic conditions. A plot between mean peak area and concentration was found to be linear in the range of concentration 5.0–20.0 μg/mL and it was presented in Fig. 4. Slope, intercept and correlation coefficient were calculated

by least square regression method and were presented in Table 2. Preparation of 0.06% solution at specification level (0.006 μg/mL solution): To find out LOD (or LOQ) of the developed method, 0.006 μg/mL (or 0.02 μg/mL solution) solution, 1.0 mL of 10 μg/mL solution was pipetted into a 10 mL of volumetric flask and dilute up to the mark with diluent. Further PFI-2 datasheet pipetted 0.13 mL (for LOQ 0.2 mL) of above diluted solution into a 20 mL (10 mL in case of LOQ) of volumetric much flask and dilute up to the mark with diluent. Calculation of signal/noise ratio (S/N) from the average baseline noise obtained

for blank (42 μV) and signal obtained from 0.006 μg/mL and 0.02 μg/mL of target assay concentration (123 μV and 422) was found to be 2.92 and 10.0 respectively. Accuracy of the proposed method was determined by analyzing famotidine sample spiked at three different concentration levels in triplicate. To find out the accuracy a known amount of standard drug was added to the fixed amount of pre-analyzed sample solution at three different concentration levels in triplicate. Percent recovery of the drug was calculated by comparing the area before and after the addition of the standard drug. The mean recovery of the drug was found to be 99.8% and shown in Table 3. The study of robustness was performed by slight modification in chromatographic conditions such as flow rate of the mobile phase, pH of the buffer, wavelength and composition of the mobile phase. The working standard solution of FMD was analyzed under this new set of experimental conditions. Only one parameter was changed while the others were kept unaltered. The system suitability parameters were evaluated as per the test method in all the cases and found to be within limits.