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Fluorescence Experiment 2: Stern-Volmer Quenching

Background

Certain molecules (i.e., quenchers) can deactivate fluorescence from fluorophores in solution by collision. This process can, in some instances, be described by Stern-Volmer kinetics, which relates the efficiency of quenching at some concentration of quencher to the bimolecular rate constant. The rate at which the quencher diffuses through the medium is governed by the viscosity of the solvent. Consequently, Stern-Volmer quenching provides an experimental method to directly measure molecular diffusion rates. Our simulation software allows the student to prepare a fluorophore in two solvents of very different viscosity. The effect of viscosity on the quenching rate constant can then be assessed by measuring fluorescence decays as a function of quencher concentration in each solvent.

Fluorescence Experiment 2, Stern-Volmer Quenching

In the first part of this experiment, stock solutions and dilutions will be prepared for anthracene in solvents of varying viscosity using the virtual flask module. Fluorescence and UV-Vis absorption spectra will also be recorded:

Virtual Flask Module & Absorption & Fluorescence Spectrometer Simulators

A fluorescence decay will be recorded at room temperature using the single photon counter simulator with temperature control. These decay measurements will be repeated at several different temperatures.

Virtual solutions with varying concentrations of fluorescence quencher will also be prepared.

Decay Measurement using the Simulator

Virtual solutions with varying concentrations of fluorescence quencher will also be prepared.

A fluorescence decay will be recorded at room temperature using the single photon counter simulator. These decay measurements will be repeated at several different quencher concentrations.

Decay Measurement using the Simulator

A corresponding instrument profile (prompt) will be recorded following each decay curve measurement:

Prompt Measurement using the Simulator

A fluorescence lifetime will be derived at each quencher concentration using our decay curve analyses software:

Decay Curve Analyses Software

The derived lifetimes will then be plotted as a function of concentration in a Stern-Volmer plot to derive the bimolecular quenching constant in each solvent.

Any questions? Please contact our experts who will be happy to help.

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We cover a variety of topics using a UV-Vis Absorption spectrometer simulator: Everything from studying the kinetics of the crystal violet/sodium hydroxide reaction to determining the level of aspirin in a blood sample. Take a look at our product page.