A portable biochemical testing apparatus and operating method thereof are disclosed. The portable biochemical testing apparatus includes a light source module, a sample module, a photoconductive material layer, a touch module, and a control module. At least one sample is disposed in the sample module. The photoconductive material layer is disposed between the sample module and the light source module. The touch module generates a driving signal according to a touch action of the user to drive the light source module to emit a light. When the light is emitted to the photoconductive material layer, the photoconductive material layer will generate a photoelectric driving effect. The at least one sample is affected by the photoelectric driving effect and generates a change corresponding to the touch action.