Coupling of microextraction and voltammetry techniques for the determination of hemin in human serum samples

Extra concentrations of hemin (ferriprotoporphyrin) are toxic since it may intercalate in lipid membranes and catalyze the formation of hydroxyl radicals and the oxidation of low density lipoproteins, damage mitochondrial DNA of hepatocytes, inhibit cytochrome P450-catalyzed reactions of human liver microsome, cause the oxidation of membrane components which may promote cell lysis and death (1, 2). So, determination of hemin concentration is of importance. However, a sample preparation step seems to be inevitable prior to the determination due to the complexity of the biological samples. Dispersive liquid–liquid microextraction (DLLME) is a very simple and rapid method for the extraction and separation of various analytes from complex samples (3). Ionic liquids (ILs) are green solvents comprised of asymmetric organic cation and organic or inorganic anion, so are liquids which represent a new class of non-aqueous but polar solvents with high ionic conductivity (4). In the present work, an ammonium-based task-specific ionic liquid containing salicylate as anion was synthesized and used for the extraction of hemin Hemin is extracted into the ionic liquid after interaction with the anion of the ionic liquid. Voltammetric determination followed the extraction process. For this purpose, the IL-rich phase separated from the sample was placed on the surface of a glassy carbon electrode with the help of nafion and ethanol. In this way, the IL was used as both extracting solvent (due to its polarity and water-immiscibility) and electrode modifying agent (due to its ionic structure). The peak current obtained from the electrochemical reduction of hemin was used as the analytical signal correlated to the concentration of hemin. Different experimental parameters affecting both the extraction and electrochemical processes were optimized. Under the optimized conditions, a linear concentration range of 0.020-2.60 μM with a detection limit of 3.16×10¯³ μM was obtained. The 327 presented methodology was successfully applied to the determination of hemin in human serum samples indicating its applicability to complex media.