研究目的
To analyze and exploit the deep ultraviolet fluorescence characteristics of cyanobacteria and microalgae for their discrimination in liquid samples, aiming for real-time detection.
研究成果
The UV autofluorescence method effectively discriminates cyanobacteria from other phytoplankton and among cyanobacterial species, with potential for real-time, in situ applications. Future improvements could include multiple excitation wavelengths and enhanced sensitivity with photodiodes and optical filters.
研究不足
The method requires systems capable of operating in deep ultraviolet, which may involve more expensive components. It was tested on a limited number of species and conditions; further investigations are needed for mixtures and interferents. The approach may be sensitive to photo-physiological variability, though tryptophan is used to mitigate this.
1：Experimental Design and Method Selection:
The study uses fluorescence spectroscopy with an excitation wavelength of 270 nm to access tryptophan fluorescence and other pigments. A ratiometric method is employed based on relative fluorescence peak amplitudes.
2：Sample Selection and Data Sources:
Eleven strains of freshwater phytoplankton were selected, including nine cyanobacteria, one diatom, and one green alga, grown in specific media under controlled conditions.
3：List of Experimental Equipment and Materials:
LEDs with 270 nm emission wavelength (2 mW each), UV Grade Silica cuvette, solarization-resistant optical fiber (600 μm diameter), mini-spectrophotometer (spectral range 200-1000 nm, resolution 8 nm), laptop for data recording, 3D-printed cuvette holder made of black ABS, and various culture media (BBM, BG11, AB, DM).
4：Experimental Procedures and Operational Workflow:
A 3 ml aliquot from each culture was placed in the cuvette. Fluorescence was excited by LEDs, and signals were collected orthogonally using the optical fiber connected to the spectrophotometer. Integration time was set to 1 s, with each spectrum averaged over five measurements.
5：Data Analysis Methods:
Fluorescence spectra were decomposed into five wavelength ranges (303-383 nm for tryptophan, 383-498 nm for NADH, 498-560 nm for FAD-Riboflavin, 560-612 nm for phycoerythrin, 612-830 nm for phycocyanin + chlorophyll). Ratios of the areas under these peaks to the tryptophan peak area were calculated and plotted for discrimination.
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