We have developed a variety of microplanar amperometric biosensors for measuring glucose, glutamate, and lactate concentrations in food. The microplanar electrodes of the sensor chip were manufactured by sputtering Pt and Ag onto the substrate surface. The reference electrode was a AgCl electrode pretreated by electrolysing Ag on the substrate in FeCl3 solution. Four membrane layers were fabricated on the electrode surface of the sensor chip by spin-coating a γ-aminopropyl triethoxy silane solution, a Nafion dispersion, an enzyme (glucose oxidase, glutamate oxidase, or lactate oxidase) and glutaraldehyde solution, and a fluorocarbon resin dispersion. The membrane layers provided a fast response time, low interference from other substances in the samples, and a wide dynamic range. These biosensors enable accurate measurements of the target substances.