A new method for the microextraction and electrochemical determination of insulin is presented based on the following steps: (1) extraction of insulin from an aqueous phase (the donor phase) at pH 8.0 into a sodium dodecyl sulfate (SDS) supported liquid membrane phase to form an insulin-dodecyl sulfate complex, (2) re-extraction of insulin into a small volume of an acceptor phase at pH 3.0 (leading to preconcentration), and (3) detection by differential pulse voltammetry (DPV) using a new kind of electrode. The effects of the pH values of the donor and acceptor phases, SDS concentration, percentage of organic solvent, extraction time and ionic strength on the microextraction were optimized. A pencil graphite electrode modified with graphene oxide and ruthenium dioxide was used as an electrochemical sensor. The DPV responses to the concentration of insulin was linear in therange 0.0008 – 0.020 and 0.02 – 1.00 μ mol L− 1, with a detection limit of 24 pmol L− 1 . The technique was successfully applied to the determination of insulin in human urine and plasma.