EFFECTS OF RESOLUTION, PEAK RATIO AND SAMPLING FREQUENCY IN DIODE-ARRAY FLUORESCENCE DETECTION IN LIQUID-CHROMATOGRAPHY

In this work, different methods of quantification that use both fluorescence spectral and retention time data were evaluated for accuracy and precision in diode-array fluorescence detection in liquid chromatography. The methods evaluated include the peak height and peak area methods of quantification, an adaptive Kalman filter and the generalized rank annihilation method. All of these methods were used to estimate the concentration of a model led component in the presence of other partially resolved unmodelled components. The peak area and peak height methods of quantification gave the largest errors as the chromatographic resolution was decreased. The generalized rank annihilation method gave the most reliable results when the chromatographic resolution was greater than 0.3. However, this method was affected by retention time shifts between the calibration and unknown samples and the rate at which the spectra were acquired.