The Good, The Not So Good Along with E-64
The extracted UV spectra indicate that the entire degradation products are derived from gemcitabine or its intermediates. In case of acidic stress the degradation products were observed at 4.953 (as d-5), 6.082 (d-7), and 7.131?min E-64 (d-8) (Figure 6). Jansen et al. [3] have reported the presence of a coeluted degraded product with gemcitabine which does not possess UV absorption at 275?nm. The hydrolytic product d-5 (4.95?min) was observed in alkaline as well as in aqueous stress condition. On exposure to hydrogen peroxide (5%, 60��C, 1?h), gemcitabine produces only one minor degradation products having retention time 3.772?min (Table 6, Figure 7). The percentage of unoxidized gemcitabine was 97.1%. Gemcitabine was completely degraded on exposure to drastic oxidative condition (50%, 60��C, 1?h). However, these degraded compounds have no ultraviolet (UV) absorbance at 275?nm, the wavelength used to monitor the gemcitabine concentrations. Borisagar et al. [5] reported the oxidative degradation [13.8%] of gemcitabine utilizing HPTLC. The present results indicate that, using appropriate chromatographic conditions, the structurally related degraded products of gemcitabine can be separated which were not studied earlier (Table 8). Figure 5 Typical HPLC chromatogram of (a) gemcitabine exposed to alkaline stress (1?N NaOH, 60��C, 1?h), (b) contour plot, (c) peak purity index, and extracted UV spectra of (d) gemcitabine, (e) degraded product d-1, (f) d-2, (g) d-4, (h) ... Figure 6 Typical HPLC chromatogram of (a) gemcitabine exposed to acidic stress Selleckchem JQ1 (1?N HCl, 60��C, 1?h), (b) contour plot, (c) peak purity index, and extracted UV spectrum of (d) degraded product d-5, (e) d-7, and (f) d-8. Figure 7 Typical HPLC chromatogram of gemcitabine exposed to (a) hydrolytic (H2O, 60��C, 1?h) and (b) oxidative stress (5%, 60��C, 1?h). Table 6 Stability data under different stressed conditions. Table 8 Comparison between analytical methods. 3.2.8. Assay The proposed method was applied to the determination gemcitabine in injectable formulations. The results of these assay yielded 99.6% (RSD, 0.26%) of labeled claimed. Low value of precision indicates that the method can be used precisely for R428 mouse the estimation of drug in formulations (Table 7). Table 7 Assay of formulations. 4. Conclusion A validated HPLC method has been developed for determination of gemcitabine in formulations. The proposed stability indicating method is simple, economical, accurate, precise, specific, and robust. Method is capable of separating different degraded products of drug which can be estimated separately. The experimental design was found to be very useful in testing the robustness of chromatographic separation during the validation step. Hence this method can be easily and conveniently adopted for the routine analysis of gemcitabine in pharmaceutical dosage form.
