Second derivative synchronous fluorimetric method for simultaneous determination of harman and norharman in coffee samples.
The simultaneous determination of harman and norharman using second derivative synchronous fluorescence method has been developed based on their natural fluorescence. Due to their similar molecular structures, it is difficult to determine them simultaneously in the mixture using conventional fluorimetry. Overlapping of fluorescence spectra was resolved by using a constant second derivative synchronous fluorimetry. The derivative synchronous spectrum, maintaining a constant difference of Δλ150 nm between emission and excitation for both the compounds, has been selected for the analysis. The range of application is between 0.182 and 18.2 μgmL (correlation coefficient, R0.9982) for harman and between 0.504 and 16.8 μgmL (correlation coefficient, R0.9962) for norharman. The recovery ranges of 98.5-101.1% for harman and 97.5-99.1% for norharman from their synthetic mixture was reported. The detection limits are 0.016 μgmL and 0.038 μgmL for harman and norharman, respectively. Similarly, the quantitation limit of the two compounds was found to be 0.049 and 0.109 μgmL, respectively. The method was applied to the simultaneous determination of both compounds in coffee samples with satisfactory results.