Ultraviolet and visible (UV-Vis) absorption spectroscopy is the measurement of the attenuation of a beam of light after it passes through a sample or after reflection from a sample surface. Absorption measurements can be at a single wavelength or over an extended spectral range. Overview. Source: Laboratory of Dr. B. Jill Venton - University of Virginia. Ultraviolet-visible (UV-Vis) spectroscopy is one of the most popular analytical. Visible and Ultraviolet Spectroscopy. 1. Background. An obvious difference between certain compounds is their color. Thus, quinone is yellow; chlorophyll is.


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Absorbance follows Beer's Law, which states absorbance equals the molar attenuation coefficient times the path length and concentration.

The molar attenuation coefficient is related to the individual compound's ability to absorb light of a specific wavelength. Path length uv and visible spectroscopy to the distance traveled by light through the sample, which is typically 1 cm for standard cuvettes.

Beer's law can be used to calculate sample concentration, if the absorptivity is known, or a calibration curve can be used. UV-Vis is often called a general technique, as most molecules uv and visible spectroscopy light in the UV-visible wavelength range.

The UV range extends from — nm, and the visible spectrum ranges from — nm. However, most spectrophotometers do not operate in the deep UV range of — nm, as light sources in this range are expensive.

Most UV-Vis spectrophotometers use a deuterium lamp for the UV range, which uv and visible spectroscopy light from — nm, and a tungsten filament lamp for the visible range, which produces light from —2, nm. Since the light source is usually a lamp with broad wavelength ranges, the specific absorbance wavelength is selected using filters or a monochromator.

uv and visible spectroscopy

Ultraviolet-Visible (UV-Vis) Spectroscopy | Protocol

A monochromator is a device that separates the wavelengths of light spatially, and then places an exit slit where the desired wavelength of light is. The monochromator can be scanned over a wavelength range uv and visible spectroscopy provide an entire absorbance spectrum.

This makes the technique useful for quantifying and identifying a wide range of molecules. Now that the basics uv and visible spectroscopy UV-Vis spectroscopy have been outlined, lets take a look at a simple UV-Vis experiment in the laboratory.


Before beginning the measurement, turn on the spectrophotometer, and allow the lamps to warm up for uv and visible spectroscopy appropriate period of time to stabilize them.

Ensure that the cuvette is aligned properly with any grooved sides out of the beam-path, and insert it into the spectrophotometer.

Ultraviolet–visible spectroscopy - Wikipedia

Secure the lid to prevent ambient light from entering the system. Measure the absorbance of the blank at one wavelength, or over a wavelength range.

Record or save the absorbance, as it must be subtracted from the absorbance of the sample. Next, discard the blank and rinse the cuvette twice with sample.

Ultraviolet-Visible (UV-Vis) Spectroscopy

Wipe the outside of the cuvette again, to ensure that it is clean and free of fingerprints. Place the cuvette in the spectrophotometer in the correct orientation, and secure the lid.

Collect an absorbance measurement or spectrum at the same wavelength uv and visible spectroscopy wavelength range as the blank. Subtract the blank spectrum or measurement, if the instrument does not automatically do so.

Ultraviolet–Visible Spectroscopy (UV-Vis)

To quantify the amount of analyte in the sample, create a calibration curve using a range of known analyte concentrations. For more information on how to construct and use a calibration curve, please watch this collection's video "Calibration Curves".

The absorbance measurement can also be used to calculate reaction kinetics by measuring the uv and visible spectroscopy or decrease in a compounds concentration throughout the reaction.