Basic Principles of Spectrophotometry - BASIC...

Basic Principles of Spectrophotometry
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Unformatted text preview: BASIC SPECTROPHOTOMETRYBASIC PRINCIPLES OFSPECTROPHOTOMETRY1Principles of LightMeasurement Properties of light and energy (Photons) Wavelength Amplitude (intensity, brightness) Frequency (waves / second) Relationships: E = hf; f = c/!231BASIC SPECTROPHOTOMETRY4 Energies of atoms and molecules562BASIC SPECTROPHOTOMETRY78 Transmission/Absorption of light Reflection = bouncing Refraction = bending Diffusion = spreading Diffraction = combination of all three wavelength isolation filters Prisms -- non-linear Diffraction gratings -- linear93BASIC SPECTROPHOTOMETRY1011124BASIC SPECTROPHOTOMETRY13Absorption / Emission of light Atoms Molecules Solids14155BASIC SPECTROPHOTOMETRYLaws of Absorbance Beer / Lambert Laws Absorbance vs. %Transmittance Formulas1617186BASIC SPECTROPHOTOMETRY1920217BASIC SPECTROPHOTOMETRY2223248BASIC SPECTROPHOTOMETRY25Spectrophotometer Components Power supply Light Source -- halogen quartz Collimating lenses -- organize parallel lines Focusing lenses -- narrow or intensify beam Monochromator Band pass Stray light= band width at band height(room light, scratches, dust, fingerprints, widebandpass) Slit width -- exit slit26 Sample cell -- quartz or sliica Photodetector Photomultiplier tube: Read-out device --cathode, dynodes, anodedigital, recorder, printer279BASIC SPECTROPHOTOMETRY28293010BASIC SPECTROPHOTOMETRY31323311BASIC SPECTROPHOTOMETRY34353612BASIC SPECTROPHOTOMETRY37Analytical Problems Absorbance Variation Check photometric accuracy with Neutral Densityfilters Noise and Drift Recorder or oscilloscope Stray Light Check with sharp cut-off filters Linearity Dye solutions Wavelength Filters: holmium oxide or didymium3813...
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