The electrochemical biosensor is an integrated receptor-transducer device, which can convert a biological response into a measurable electrical signal. Because of its important features like selectivity, sensitivity, stability, reproducibility, linearity, and low cost, biosensors have a wide range of applications. The aim of this paper is to review our achievements in the development of electrochemical enzyme biosensors and the possibility of their applications in food industry, agriculture and environmental control. The specific structural features of transducers in the developed amperometric, potenciometric and conductometric enzyme biosensors are discussed. The laboratory prototypes were fabricated and systematically tested for the determination of glycoalkaloids, mycotoxins, heavy metal ions and pesticides in the agriculture and environment samples as well as some components of food and dietary supplements. They demonstrated fast, highly selective, sensitive and correct analitycal operation and also the possibility to modulate main characteristics to comply with the specific requirements for the practical purposes. It is important that the manufacture of such miniaturized electrochemical biosensors may be adapted to the technologies of large-scale production.