Name: spectrophotometry for absorption spectra
Uploaded: 2024-12-05T18:32:03.000Z
Duration: 1 min 16 s
Channel: JoVE

K12 Standards

Pennsylvania State Academic Standards

9-12

Earth and Space Science

Space Systems

3.3.9-12.C

Spectrophotometry: Introduction

Spectrophotometry is the quantitative method of evaluating the radiation interacting with a material by measuring its relative intensity.
This includes radiation transmitted through, reflected from, diffracted within, absorbed by, or emitted by the material.
In an absorption spectrometer, electromagnetic radiation of a particular wavelength is passed through the sample to a detector. Radiation absorbed by the sample yields a reduced radiation intensity at the detector. This process is then repeated for a range of wavelengths.
An absorption spectrum is the plot of the intensity lost to absorbed radiation versus the radiation wavelength.
Similarly, plotting the diffracted or transmitted radiation intensities against the wavelength generates the diffraction and transmission spectra.
In a molecule, the energy gaps for different types of transitions fall into distinct ranges.
So, the range of wavelengths indicates the transition type that occurred, and the specific wavelength of absorption can give an idea about which functional group may be in the molecule.

spectrophotometry: introduction

Spectrophotometry is the quantitative measurement of the absorption, reflection, diffraction, or transmission of electromagnetic radiation through a material as a function of the intensity and wavelength of the radiation. A spectrophotometer is a device used to measure the change in the radiation intensity caused by its interaction with the material.
The essential components of a spectrophotometer include a source of electromagnetic radiation, a slot for placing a material to be analyzed, and a detector to measure the intensity of the incident radiation from the source and the radiation exiting from the material (transmitted, reflected, or diffracted radiation). The difference between the intensities of incident radiation and transmitted radiation reaching the detector is used to calculate the intensity of radiation absorbed by the material. The sample is exposed to radiation of different wavelengths, often one at a time. The spectrometer provides a series of data with absorption details in each attempt, a process known as scanning. Since the material energy levels are finite, absorption occurs only at specific wavelengths.
The plot of absorbed radiation against the corresponding wavelength is known as the absorption spectrum. Similarly, the plots of reflected, diffracted, and transmitted radiation as a function of wavelength are the reflection, diffraction, and transmission spectra, respectively. These spectra provide information about the specific wavelength at which the interaction between the material and radiation occurs.
The wavelength range of radiation determines the type of transition that occurs in the material. For example, UV&#8211;vis spectra show UV&#8211;vis radiation absorption by the material. The absorbed radiation causes an electronic excitation in the material. In contrast, infrared spectra show the excitation of vibration levels of a specific bond in the material. The specific absorption wavelength details the molecule&#39;s chemical structure inside a range of wavelengths. For example, the specific wavelength of infrared radiation absorbed by a hydroxyl bond in a molecule is different from the wavelength of radiation absorbed by the carbonyl bond. So, infrared spectra can be used to identify functional groups in the material.
Unlike other chemical characterization methods like titration, spectroscopy is a nondestructive characterization technique, and sample material can be recovered after analysis.

Spectrophotometry is the quantitative measurement of the absorption, reflection, diffraction, or transmission of electromagnetic radiation through a ...

Education

JoVE Core

Analytical Chemistry

Unit 1

Introduction to Molecular Spectroscopy

KEY TERMS AND CONCEPTS

spectrophotometry for absorption spectra

Spectrophotometry measures how a material absorbs, reflects, diffracts, or transmits electromagnetic radiation. It compares the intensity of radiation before and after it interacts with the material. The result shows how the material responds to different wavelengths.

A spectrophotometer is the device used for this measurement. It includes a source of electromagnetic radiation, a slot for the sample, and a detector. The detector measures the radiation that leaves the material, including transmitted, reflected, or diffracted light. The difference between the incoming and outgoing intensities is used to find how much radiation the material absorbed.

During analysis, the sample is exposed to different wavelengths, often one at a time. The instrument collects data during this scanning process. Because energy levels in a material are finite, absorption happens only at specific wavelengths. A graph of absorbed radiation versus wavelength is called an absorption spectrum.

Spectrophotometry can also produce reflection, diffraction, and transmission spectra. These plots show how the material interacts with radiation at specific wavelengths. The wavelength range helps identify the kind of transition taking place in the material. In UV-vis spectra, UV-vis radiation is absorbed and causes electronic excitation.

Infrared spectra show excitation of vibration levels in a specific bond. The absorbed wavelength can reveal details about molecular structure. For example, a hydroxyl bond absorbs a different infrared wavelength than a carbonyl bond. This is why infrared spectra can help identify functional groups in a material.

Unlike titration and some other chemical tests, spectroscopy is nondestructive. The sample can be recovered after analysis. That makes spectrophotometry useful for studying materials without destroying them.

Spectrophotometry for Absorption Spectra

dual nature of electromagnetic (em) radiation

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EM Spectrum, Waves, and Photon Energy

Electromagnetic radiation has both wave and particle features. It includes electric and magnetic field components that oscillate in planes perpendicular to each other. These fields are also perpendicular to the direction the radiation moves through space. As a wave, electromagnetic radiation is described by wavelength and frequency. Wavelength is the distance between two consecutive peaks or troughs. Frequency is the number of wave cycles that pass a point in a fixed time. The electromagnetic...

interaction of em radiation with matter: spectroscopy

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EM Radiation and Matter in Spectroscopy

Electromagnetic radiation and matter interact in ways that reveal details about atoms and molecules. In spectroscopy, scientists study how EM radiation is absorbed, emitted, transmitted, reflected, or scattered by a substance. This helps identify its chemical composition and properties. EM radiation behaves as both a wave and a particle. As a wave, it has intensity and amplitude. Intensity, also called irradiance, is the number of photons that strike a given area in a certain time. Amplitude...

molecular spectroscopy: absorption and emission

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Molecular Energy Levels in Spectroscopy

Molecular spectroscopy describes how molecules absorb and emit light by moving between energy levels. These levels are called quantum states. Unlike atoms, molecules also have rotational and vibrational energy levels, so their energy structure is more complex. Each level is separated by an energy gap. The gaps between electronic, vibrational, and rotational levels are very different in size. A diatomic molecule can be used to show these three kinds of energy levels. A molecule can absorb a...

Spectrophotometry measures how a material absorbs, reflects, diffracts, or transmits electromagnetic radiation. It compares the intensity of radiation before and after it interacts with the material. The result shows how the material responds to different wavelengths. A spectrophotometer is the device used for this measurement. It includes a source of electromagnetic radiation, a slot for the sample, and a detector. The detector measures the radiation that leaves the material, including...

ultraviolet and visible (uv&ndash;vis) spectroscopy: overview

ultraviolet-and-visible-uvvis-spectroscopy-overview

UV-Vis Spectroscopy for Light Absorption

UV-Vis spectroscopy measures how materials absorb ultraviolet and visible light. It is a useful analytical method because it is versatile, simple to use, and gives results quickly. Scientists use it for both qualitative and quantitative analysis. When UV-Vis radiation passes through a sample, molecules absorb specific wavelengths based on the energy needed for electronic transitions. This absorption moves atoms or molecules from a ground state to an excited state. If a molecule absorbs UV...

uv&ndash;vis spectroscopy: molecular electronic transitions

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UV-Vis Spectroscopy and Molecular Absorption

UV-Vis spectroscopy shows how molecules absorb ultraviolet and visible light. It is used to study electronic structure by tracking how electrons move between molecular orbitals. The absorbed light must match the energy gap between the ground state and an excited state. In this process, an electron moves from the highest occupied molecular orbital, or HOMO, to the lowest unoccupied molecular orbital, or LUMO. This jump raises the electron to a higher energy level. The energy difference between...

uv&ndash;vis spectrometers

UV-Vis Spectrophotometer Parts and Samples

A UV-visible spectrophotometer measures how much UV and visible light a sample absorbs. It uses a light source, a wavelength selector, a sample holder, and a detector to build the spectrum. This makes it useful for checking liquid samples in the lab. The light source depends on the region being measured. A deuterium lamp provides UV radiation, and a tungsten lamp provides visible radiation. A monochromator or prism acts as a diffraction grating and splits the incoming light into different...

uv&ndash;vis spectrum

How Molecular Structure Shapes UV-Vis Bands

UV-Vis spectroscopy shows how a molecule absorbs light in the ultraviolet and visible ranges. When light passes through a substance, some of it is absorbed and the rest is reflected or transmitted. Molecules that absorb between 180 and 400 nm give a UV spectrum, while absorption in the 400 to 780 nm range gives a visible spectrum. A UV-Vis spectrum is a plot of absorbance versus wavelength. The y-axis shows molar absorptivity, ε, or log ε, and the x-axis shows wavelength. Absorbance describes...

uv&ndash;vis spectroscopy: beer&ndash;lambert law

Using Absorbance to Find Concentration

The Beer-Lambert law links absorbance, concentration, and path length in a solution. It is based on ideas from Johann Heinrich Lambert and August Beer. Their work helps explain how light changes as it passes through a medium. Lambert's law says that when light passes through a medium, the loss in intensity is directly proportional to the original intensity and the path length. Beer’s law adds that the transmittance of a solution stays constant if the product of concentration and path length...

photoluminescence: fluorescence and phosphorescence

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Light Emission from Excited Molecules

Photoluminescence is light emission from molecules after they absorb light energy. A substance takes in photons, which raises electrons to excited energy levels. The electrons then relax back to the ground state and release light. This process appears in many materials, including semiconductors, organic compounds, and inorganic compounds. The main role of electron spin helps separate photoluminescence into fluorescence and phosphorescence. In a singlet spin state, two electrons share the same...

variables affecting phosphorescence and fluorescence

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Factors That Change Light Emission in Molecules

Fluorescence and phosphorescence depend on several factors that change how molecules emit light. These effects matter in analytical chemistry, biological imaging, and materials science because they help scientists detect molecular properties and see cellular structures. Molecular structure is one of the main influences. Aromatic compounds with low-energy pi-bonding to pi-non-bonding transitions often give strong fluorescence. Aliphatic and alicyclic carbonyl compounds, as well as highly...

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Jablonski Diagram: Light Loss Pathways

Jablonski diagrams show how excited molecules lose energy after absorbing light. They are used to track both light-emitting and non-light-emitting steps in luminescence. The diagram gives a simple picture of the energy states a molecule can move through. The lowest line in a Jablonski diagram is the ground-state energy of the molecule. This state is usually a singlet state, and it represents the condition of most molecules in solution at room temperature. The lines above it show vibrational...

photoluminescence: applications

Fluorescent and Phosphorescent Analysis

Photoluminescence is useful in chemical analysis because it is sensitive and selective. It can support both direct and indirect measurements of an analyte, which is the substance being tested. When an analyte has a strong quantum yield, it can be measured directly by fluorescence or phosphorescence. Indirect analysis is used when the analyte is not naturally fluorescent or phosphorescent, or when its quantum yield is poor. One approach is to react the analyte with a reagent to make a...

fluorescence and phosphorescence: instrumentation

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Measuring Fluorescence and Phosphorescence

Fluorometers and spectrofluorometers are instruments used to measure molecular fluorescence. They do this by choosing specific excitation and emission wavelengths. The two instruments use different light sources and wavelength-selection methods. A fluorometer uses absorption interference filters to select the excitation and emission wavelengths. Its excitation source is usually a low-pressure mercury vapor lamp. This lamp gives intense lines across the ultraviolet and visible regions.

key terms and concepts

Learn how spectrophotometry measures absorption, reflection, transmission, and diffraction, and how UV-vis and infrared spectra reveal material properties.

Spectrophotometry for Absorption Spectra | JoVE K12

analytical chemistry

JoVE Core Analytical Chemistry simplifies complex chemical principles by breaking them down into clear, engaging animations, making topics from SI Units to NMR spectroscopy more accessible and straightforward. Additionally, the scientist-in-action videos highlight the application of these concepts in real-world research conducted in laboratories globally.

Physical Science

Waves and Electromagnetic Radiation

3.2.9-12.X

Louisiana Student Standards for Science

High School

Earth Science

Earth’s Place In The Universe

HS-ESS1-2

Chemistry

motion-and-stability-forces-and-interactions

Motion And Stability: Forces And Interactions

HS-PS2-6

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3.3.9-12.A

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Atomic Absorption Spectroscopy: Radiation and Light Sources

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Students who demonstrate understanding can construct an explanation of the Big Bang theory based on astronomical evidence of light spectra, motion of distant galaxies, and composition of matter in the universe.  Clarification Statement: Emphasis is on the astronomical evidence of the red shift of light from galaxies as an indication that the universe is currently expanding, the cosmic microwave background as the remnant radiation from the Big Bang, and the observed composition of ordinary matter of the universe, primarily found in stars and interstellar gases (from the spectra of electromagnetic radiation from stars), which matches that predicted by the Big Bang theory (3/4 hydrogen and 1/4 helium).  Assessment Boundary:

3.3.9-12.B

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3.3.9-12.D

Spectrophotometry for Absorption Spectra

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