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

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Published in Biomedical Chromatography. 2010, vol. 24, no. 7, p. 732-6
Abstract 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 microg/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 microg/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.
Keywords AnimalsChromatography, High Pressure Liquid/methodsCytochromes c/analysis/metabolismPermeabilitySkin/chemistry/metabolismSwine
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PMID: 19882748
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BACHHAV, Yogeshwar, KALIA, Yogeshvar. Development and validation of an analytical method for the quantification of cytochrome c in skin transport studies. In: Biomedical Chromatography, 2010, vol. 24, n° 7, p. 732-6. doi: 10.1002/bmc.1356 https://archive-ouverte.unige.ch/unige:22718

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Deposited on : 2012-09-03

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