New Methods for the Characterization of Different Gelatin Based Nanoparticulate Drug Carrier Systems

Peak 1 displays the adaptation of the fractionation conditions as described above to the siRNA oligonucleotide. With an increased initial cross-flow intensity the separation from the void peak, inherent to AF4 systems, will be enhanced (peak 2). The higher MW of the siRNA oligonucleotide compared to the ssDNA oligonucleotide permit the application of a 5 kDa cut-off ultrafiltration membrane Similar studies to those using CHDF were performed with AF4. Optimal conditions for nanoparticle / oligonucleotide fractionation were developed using model proteins with a MW similar to those of the siRNA and DNA oligonucleotides, respectively (data not shown). With the combination ssDNA oligonucleotide and gelatin nanoparticles, a positive drug loading reaction could be characterized. The results of the drug loading process are shown in Fig. 4. The level of oligonucleotide load on the cholamin modified nanoparticles can be determined by interpretation of the according UV-signals. The difference in AUC of loaded nanoparticles and AUC of unloaded nanoparticles corresponds to the differences of the oligonucleotide AUCs. The required linear correlation between the AUC of the separated analytes and their varying concentration was verified experimentally (data not shown). Fig. 4 Fractionation of ssDNA oligonucleotide (____ 100 μg/ml) and gelatin nanoparticles (____ 500 μg/ml) before and after (____) drug loading; initial cross-flow 65%, cross-flow duration 780 sec (cp. Fig. 2 ____), 1 kDa cut-off ultrafiltration membrane