![]() ![]() ![]() Experimental parameters: λ ex = 340 nm, Δλ ex = 1nm, Δλ em= 1 nmįluorescence emission spectra show the change in fluorescence intensity as a function of the wavelength of the emission light (Figure 5), and are measured using a spectrofluorometer. Excitation spectra can therefore be thought of as fluorescence detected absorption spectra.įigure 4: Schematic of the measurement of excitation spectra in a spectrofluorometer.įigure 5: Fluorescence emission spectrum of anthracene in cyclohexane measured using the FS5 Spectrofluorometer. If the sample obeys Kasha’s Rule and Vavilov’s Rule then the excitation spectrum and absorption spectrum will be identical (compare Figures 1 and 3). The wavelength of emission monochromator is set to a wavelength of known fluorescence emission by the sample, and the wavelength of the excitation monochromator is scanned across the desired excitation range and the intensity of fluorescence recorded on the detector as a function of excitation wavelength. Experimental parameters: λ em = 420 nm, Δλ em = 1 nm, Δλ ex = 1 nm.įluorescence excitation spectra show the change in fluorescence intensity as a function of the wavelength of the excitation light (Figure 3), and are measured using a spectrofluorometer. The absorbance is linearly proportional to the molar concentration of the sample which enables the concentration of the sample to be calculated from the absorption spectrum using the Beer-Lambert Law.įigure 2: Schematic of the measurement of absorption spectra in a spectrophotometer.įigure 3: Fluorescence excitation spectrum of anthracene in cyclohexane measured using the FS5 Spectrofluorometer. ![]() The intensity of light transmitted through the sample, I Sample, (such as an analyte dissolved in solvent) and the intensity of light through a blank, I Blank, (solvent only) are recorded and the absorbance of the sample calculated using: Absorption spectra are measured by varying the wavelength of the incident light using a monochromator and recording the intensity of transmitted light on a detector. Experimental parameters: Δλ = 1 nm.Ībsorption spectra (also known as UV-Vis spectra, absorbance spectra and electronic spectra) show the change in absorbance of a sample as a function of the wavelength of incident light (Figure 1), and are measured using a spectrophotometer. In this Spectral School tutorial, the differences between molecular absorption, excitation and emission spectra are explained.įigure 1: Absorption spectrum of anthracene in cyclohexane measured using the FS5 Spectrofluorometer. ![]()
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