Modified dual lifetime referencing method for simultaneous optical determination and sensing of two analytes.

Simultaneous fluorometric sensing of two analytes becomes possible using a modified dual lifetime referencing (m-DLR) method. In this scheme, two luminescent indicators are needed that have overlapping absorption and emission spectra but largely different decay times. They are excited by a single light source, and both emissions are measured simultaneously. In the frequency domain m-DLR method, the phase of the short-lived fluorescence of a first indicator is referenced against that of the long-lived luminescence of the second indicator. The analytical information is obtained by measurement of the phase shifts at two modulation frequencies. The method is demonstrated to work for the case of dually sensing oxygen and carbon dioxide. It benefits from simple instrumentation and optical setup. The approach is perceived to be of wide applicability. Examples include (1) analysis of two luminescent analytes, (2) analytical determinations that make use of two probes, and (3) sensing of two species such as carbon dioxide and oxygen (as demonstrated here), or oxygen and chlorophyll, provided the luminophores meet the condition of having largely different decay times and overlapping absorption and emission spectra.

Characterization of a reservoir-type capillary optical microsensor for pCO(2) measurements.

A reservoir type of capillary microsensor for pCO(2) measurements is presented. The sensor is based on the measurement of the fluorescence intensity of the anionic form of the pH indicator 1-hydroxy-3,6,8-pyrenetrisulfonate in the form of its ion pair with a quaternary ammonium base in an ethyl cellulose matrix. The glass capillary containing the reservoir sensor was prepared by immersing the tip of the optical fiber into the sensing agent very close to the sensor tip thus providing a very small volume for the sensing reaction. The purpose of the sensing approach is to regenerate the dye/buffer system by diffusion, which may be poisoned by interfering acids, or bleach by photolysis. The fresh cocktail from the reservoir takes the place of protonated form of the dye. The internal buffer system also makes the protonation-deprotonation equilibria reversible. The distal tip of the internal buffer containing reservoir is coated with a gas-permeable but ion-impermeable teflon membrane. The dynamic range for the detection of pCO(2) is between 1 and 20 hPa, which corresponds to the range of dissolved CO(2) in water. The response time is 15 s and the detection limit is 1 hPa of pCO(2.) The recovery performance of this sensor can be improved by means of mechanical adjustment of the sensor tip in a micrometric scale.