Development and validation of an analytical method for the quantification of cytochrome c in skin transport studies

作者：Bachhav Yogeshwar G; Kalia Yogeshvar N^*

来源：Biomedical Chromatography, 2010, 24(7): 732-736.

DOI：10.1002/bmc.1356

• Cytochrome c
• HPLC
• transdermal delivery
• permeation
• skin deposition
• laser poration
• quantification

A simple isocratic HPLC method for the quantification of Cytochrome c in skin permeation samples was developed and validated. The mobile phase comprised a 41:59 mixture of an organic phase A (0.1% trifluoroacetic acid in a 90:10 mixture of MeCN-H(2)O) and an aqueous phase B (0.1% trifluoroacetic acid in H(2)O). The Cytochrome c retention and run times were 2.62 and 8.0 min, respectively much shorter than those for existing gradient methods. The response was accurate, precise and linear from 2.5 to 25 mu g/mL. The mean recoveries for intra-day and inter-day analysis ranged from 88.5 to 103.8% and the RSD varied from 0.05 to 1.55%. The assay was used to quantify transport of Cytochrome c across intact and laser-microporated porcine skin in vitro. Cytochrome c permeation and the amount of protein retained within the membrane over 24 h were quantified as a function of the number of micropores. Although no Cytochrome c permeation was observed across intact skin, laser microporation enabled delivery of 22.9 +/- 3.3 and 56.0 +/- 15.9 mu g/cm(2) of the protein across skin samples with 300 and 1800 micropores, respectively. In conclusion, the HPLC method provided a fast, efficient means to quantify Cytochrome c in samples from skin transport studies.

• 出版日期2010-7