Here, we provide an extremely simple, rapid, direct, sensitive and specific method to quantify, for the first time in human colon carcinoma cells (HT29) and epidermoid human larynx carcinoma cells (HEp-2), the intracellular glutathione (GSH) using an electrochemical detector coupled to a high performance liquid chromatograph (HPLC-ECD). GSH determination in cancer cells are of valuable interest because GSH has been implicated in the development of resistance to several anticancer drugs. Separation of GSH was performed on a Alltima C18 column using a binary pre-mixed mobile phase of methanol-NaH2PO4 (10 mM, pH 3.0 with phosphoric acid) (2:98, v/v). GSH was detected with an oxidation potential of +500 mV. The limit of detections was 0.30 pMol and good linearity (r = 0.9988) was found. The intra-day and the inter-day recoveries were 97.1–101.3% and 98.2–101.8% respectively, whereas the accuracies were less than 2.56% and 3.02% respectively. The usefulness of the method was established by subjecting HT29 and HEp-2 cells to an oxidant stress generated by menadione, a substance able to decrease GSH level in cells. The experiments show that the HPLC-ECD method developed could represent an useful alternative to the existing procedures since its simplicity and rapidity.
|Titolo:||SIMPLE ANALYSIS OF GLUTATHIONE IN HUMAN COLON CARCINOMA CELLS AND EPIDERMOID HUMAN LARYNX CARCINOMA CELLS BY HPLC WITH ELECTROCHEMICAL DETECTION|
|Autori interni:||SANTAGATI, Natale Alfredo|
|Data di pubblicazione:||2005|
|Appare nelle tipologie:||1.1 Articolo in rivista|