Development of a sensitive HPLC method to measure in vitro permeability of E- and Z-isomeric forms of thiosemicarbazones in Caco-2 monolayers

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• 作者：Zufan Debebe ; Sergei Nekhai ; Meseret Ashenafi ; David B. Lovejoy ; Danuta S. Kalinowski ; Victor R. Gordeuk ; W. Malcolm Byrnes ; Des R. Richardson ; Pradeep K. Karla
• 关键词：HIV-1 iron chelator ; HPLC method development ; Inter-convertible Z and E isomers ; Caco2 drug permeability
• 刊名：Journal of Chromatography B, Analytical Technologies in the Biomedical and Life Sciences
• 出版年：2012
• 出版时间：1 October, 2012
• 年：2012
• 卷：906
• 期：Complete
• 页码：25-32
• 全文大小：346 K

文摘

In the current study, we developed a HPLC method to quantitatively measure the permeability of the BpT-based chelators, 2-benzoylpyridine 4-ethyl-3-thiosemicarbazone (Bp4eT) and 2-benzoylpyridine 4-allyl-3-thiosemicarbazone (Bp4aT), across human colorectal adenocarcinoma (Caco-2) monolayers as a model of gut absorption. In aqueous solution, Bp4eT and Bp4aT formed inter-convertible Z and E isomers that were resolved by HPLC. Peak area was linear with respect to chelator concentration. Acceptable within-day and between-day precision (<22 % ) and accuracy (85-115 % of true values) were obtained over a range of 1.0-100 ¦ÌM for Bp4eT and 1.5-300 ¦ÌM for Bp4aT. Limits of detection were 0.3 ¦ÌM and 1 ¦ÌM for Bp4eT and Bp4aT, respectively, while corresponding limits of quantification were 1 ¦ÌM and 5 ¦ÌM. Both chelators showed significant ability to chelate iron in THP-1 cells using a calcein-based assay and no apparent cytotoxicity was observed within 24 h. Ratios of the apical to basolateral and basolateral to apical transport for Bp4eT were 1.10 and 0.89 at 100 ¦ÌM and 300 ¦ÌM respectively, indicating equal bi-directional movement of the compounds. Similarly, ratios were 0.77 and 0.92 for Bp4aT, respectively. This study demonstrates that Bp4eT and Bp4aT can be efficiently transported through Caco-2 cells and can potentially be formulated for oral delivery.