You should select a dye whose photophysical characteristics are similar to those of the sample in order to calculate an accurate relative quantum yield value; more specifically, the dye's absorption and emission spectra should be as similar to your sample as possible. Among the many dyes that are readily accessible are rhodamine, quinine sulphate, and fluorescein. What's best for your sample will be found through a literature search.
To confirm that you actually have two peaks, you must first subtract the solvent peak and the proper baseline. If so, you can substitute the area under the total fluorescence curve for the intensity in the formula to obtain a more accurate answer for both the sample and the standard.
Always consider the integrated spectra (excitation and emission) and not simply the intensity at maximum (especially in your case when you have two emission peaks) since the definition of QY involves the total number of photons absorbed/emitted and absorption / emission bands may not have the same shape.
- You have to be careful with concentrations since too concentrated solutions may lead to re-absorption (inner-filter effect) or to the formtion of exciplexes. Moreover, theory shows that in emission the intensity is proportional to the concentration only up to an absorbance of 0.05 at excitation wavelength! Usually you have a linear dependence upt to A = 0.05-0.1 then a levelling off and then the intensity decreases with increasing concentration.
- Concentrations must be used with caution since they can cause exciplex formation or re-absorption (the inner-filter effect). In addition, theory demonstrates that in emission, intensity and concentration are only proportional up to an absorbance of 0.05 at the wavelength of excitation! Typically, the intensity declines with increasing concentration after a levelling off at a linear dependence up to A = 0.05–0.1.
- If for some reasons (particularly to avoid dissociation of complexes) you have to have solutions with A > 0.05 then you have to correct and replace A with (1-10^-A)
- It is best to have the same excitation wavelength both both the sample and the standard and, also, to have the same absorbance for both solutions; this avoid many errors. In this case, solutions may have A up to 0.1.
- Note that emission spectra (standard and sample) have to be corrected for the instrumental function
- A good way to test if your solutions are "ideal" is to prepare several solutions of the sample (typically with A = 0.01, 0.02, 0.03, 0.04, 0.05, 0.1) and to plot the integrated emission intensity versus A. If everything is OK (no exciplex formation, no dissociation of complex, no inner-filter effect), then you should get a straight line the slope of which is the quantum yield.
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In case that your sample emits around 780nm with an excitation around 530?nm - you could go for like styryl6.
What is quantum yield in photochemical reactions?
In a chemical photodegradation process, when a molecule dissociates after absorbing a light quantum, the quantum yield is the number of destroyed molecules divided by the number of photons absorbed by the system.
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