A microscale biosensor for acetate, propionate, isobutyrate, and lactate is described. The sensor is based on the bacterial respiration of low-molecular-weight, negatively charged species with a concomitant reduction of NO(-)(3) to N(2)O. A culture of denitrifying bacteria deficient in N(2)O reductase was immobilized in front of the tip of an electrochemical N(2)O microsensor. The bacteria were separated from the outside environment by an ion-permeable membrane and supplied with nutrients (except for electron donors) from a medium reservoir behind the N(2)O sensor. The signal of the sensor, which corresponded to the rate of N(2)O production, was proportional to the supply of the electron donor to the bacterial mass. The selectivity for volatile fatty acids compared to other organic compounds was increased by selectively enhancing the transport of negatively charged compounds into the sensor by electrophoretic migration (electrophoretic sensitivity control). The sensor was susceptible to interference from O(2), N(2)O, NO(2)(-), H(2)S, and NO(-)(3). Interference from NO(-)(3) was low and could be quantified and accounted for. The detection limit was equivalent to about 1 microM acetate, and the 90% response time was 30 to 90 s. The response of the sensor was not affected by changes in pH between 5.5 and 9 and was also unaffected by changes in salinity in the range of 2 to 32 per thousand. The functioning of the sensor over a temperature span of 7 to 30 degrees C was investigated. The concentration range for a linear response was increased five times by increasing the temperature from 7 to 19.5 degrees C. The life span of the biosensor varied between 1 and 3 weeks after manufacturing.