Phase Fluorometric Optical Carbon Dioxide Gas Sensor for Fermentation Off-Gas Monitoring

We demonstrated an optical carbon dioxide gas sensor suitable for replacement of gas chromatographs and mass spectrometers for the measurement of carbon dioxide in the off-gas of a bioreactor for fermentation and cell culture applications. The sensor is based upon the change in lifetime of a donor fluorophore, sulforhodamine 101 (SR101), induced by fluorescence resonance energy transfer to a pH-sensitive, nonfluorescent acceptor, m-cresol purple (MCP). Carbon dioxide diffusing into the sensor produces carbonic acid, changing the absorbance spectrum of the MCP, and thus its spectral overlap with the SR101, changing its lifetime. This lifetime change was measured in the frequency, rather than the time domain, as a change in the phase angle of the fluorescence relative to the modulated excitation light. The sensor was calibrated by correlating the phase response to carbon dioxide concentrations. The calibration remained valid over the life of the sensor, which has been shown to be greater than 2 weeks. The sensor was most sensitive at low CO₂ concentrations and responded to concentration changes in seconds. The sensor film is very inexpensive to produce and the light source is an inexpensive light-emitting diode. Furthermore, lower cost detection electronics can be developed since only one modulation frequency is required. In addition, this sensor can potentially be used in vivo, with a fiber optic both delivering the excitation light and collecting the emission.