Thermo Scientific Evolution 300 UV-Vis Spectrophotometer
The Thermo Evolution 300 UV-Visible Spectrophotometer, is used for optical absorbance measurements between 190 and 1100 nm, covering the UV and visible ranges and part of the near IR range.
It is a double-beam instrument with separate positions for sample and reference cuvettes. The light source is a xenon lamp and the detector is a silicon photodiode. A monochromator based on a diffraction grating is used to separate the different wavelengths of light. The instrument is very fast, permitting data acquisition at a rate of 50 data points per sec.
To measure absorbance, a beam of light with intensity I0 is aimed at the tested solution placed in a cuvette. The intensities of the entering beam - I0 and the emerging beam I1 are measured, and the absorbance - A - is calculated from the ratio of the two.
A = -log (I1/I0)
Absorbance measurement in a cuvette
Three modes of measurement are offered:
Concentration measurements - The concentration of the analyte in the sample is calculated from the measured absorbance, using Beer's law. The instrument is calibrated using standard solutions with known analyte concentrations.
Time Scan - Changes in absorbance are measured as a function of time at a chosen wavelength. This mode is used to monitor reaction rates, where changes in reactant or product concentration involve changes in optical absorbance.
Wavelength Scan - An Absorbance spectrum of a sample can be obtained by monitoring the absorbance as a function of wavelength. 'Absorbance Spectra' of chlorophyll a and chlorophyll b solutions are presented below.
Absorbance Spectrum of chlorophyll a and b