The non-invasive detection of blood glucose with the photoacoustic technique has attracted many attentions in the world. To study the effects of multiple factors on the photoacoustic detection of glucose, a Nd: YAG 532nm pumped optical parameters oscillator (OPO) pulsed laser induced photoacoustic detection system were established in this paper. The lateral model was used to capture the photoacoustic signals of glucose by using the ultrasonic transducer. In the system, several components about the effects of the influence factors were considered and designed. In the experiments of glucose aqueous solutions, the photoacoustic experiments about four different influence factors, i.e., concentrations, temperatures, laser energies and flow velocities were performed. Not only the relationships between each factor and the photoacoustic detection of glucose were built, but also the coupled relationship model between the multiple factors on the concentration of glucose was also established by using the partial least square algorithm. Results show that the photoacoustic peak-to-peak value of glucose linearly increases with the glucose’s concentration, laser’s energy and temperature, but decreases with the increasing of flow velocity. Moreover, the glucose concentration prediction accuracy is improved with the increasing of laser’s energy. At the same time, it was found that the influences of glucose concentration and laser energy on the photoacoustic detection of glucose are larger than that of the temperature and flow velocity. Finally, the concentration prediction model of coupled several factors has good prediction effect.