On-the-Fly Fluorescence Lifetime Determination with Total Emission Detection in HPLC

Area of Science:

• Analytical Chemistry
• Spectroscopy
• Chromatography

Background:

• Accurate detection and quantification of polycyclic aromatic hydrocarbons (PAHs) are crucial in environmental and chemical analysis.
• Traditional fluorescence detection methods in High-Performance Liquid Chromatography (HPLC) have limitations in sensitivity and specificity, especially for complex mixtures.
• Characterizing fluorescence decay profiles offers a potential avenue for improved analytical performance.

Purpose of the Study:

• To develop and demonstrate a novel method for measuring the entire fluorescence decay profile on-the-fly during HPLC elution.
• To achieve higher sensitivity in fluorescence detection of PAHs compared to existing literature methods.
• To explore the utility of fluorescence lifetime analysis for resolving coeluting PAHs.

Main Methods:

• Utilized a pulsed Nd:YAG laser system (266 nm excitation) for fluorescence excitation.
• Employed a photomultiplier tube and digital storage oscilloscope for broadband fluorescence detection.
• Analyzed time-integrated fluorescence and fluorescence decay profiles for individual PAHs and mixtures.

Main Results:

• Achieved detection limits in the 1-10 parts per billion (ppb) range for individual PAHs.
• Successfully analyzed chromatograms of PAH mixtures containing 8-10 species using phase plane analysis of fluorescence decay.
• Demonstrated the determination of fluorescence lifetimes under coelution conditions, indicating potential for component resolution.

Conclusions:

• The direct, on-the-fly measurement of fluorescence decay profiles in HPLC offers significantly enhanced sensitivity for PAH analysis.
• Fluorescence lifetime analysis provides a valuable tool for resolving and identifying PAHs, even when they elute simultaneously.
• This advanced spectroscopic approach holds promise for more comprehensive and sensitive characterization of complex PAH samples.