A new amperometric glucose sensor based on the glucose oxidase immobilized on pyrolytic graphite (PG) modified with tetraammineplatinum(II) chloride (TAPtCl) and 5,10,15,20-tetrakis (4-methoxy-phenyl)-21H,23H-porphine cobalt(II) (TMPPCo) as well as Nafion was studied. The performances amongst the glucose sensors with or without TAPtCl or/and TMPPCo measured with oxygen present in the solution were compared. The compositions of the membranes of the glucose sensors were optimized by a new orthogonal experimental design technique-sequential level elimination method according to chemometric approaches. Our studies show that the prepared sensor with optimal membrane composition in this study gives satisfactory performance in terms of long-term stability, fast amperometric response, good detection limits and satisfactory recovery. The study provides a useful basis for developing other sensors with corresponding optimal membranes.