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Lecture Chapter 7 - Fundamentals of Spectros

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22 views26 pages

Lecture Chapter 7 - Fundamentals of Spectros

Uploaded by

Alysa Parole
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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Foundations of

Spectrophotometry
Chapter 7 | BIO 1103

Far Eastern University, BIO1103/4 Cluster AY 2021 – 2022


Discussion Outline
• The Electromagnetic Radiation (EMR)
• Emission Spectra
• Fluorescence and Phosphorescence
• Research Application of Spectroscopy
Electromagnetic Radiation (EMR)

THE ELECTROMAGNETIC SPECTRUM


Electromagnetic Radiation (EMR)

• James Maxwell – proposed that visible light consist of


electromagnetic waves (1873)
Electromagnetic Radiation (EMR)

• Isaac Newton – showed that visible light can be separated into


different colors using a prism
Electromagnetic Radiation (EMR)

wavelength ()

frequency ()

Equation:  = c
Quantum Theory (1900)

• The Quantum theory states that energy


can be gained or lost by matter only in
multiples of the quantity h

E =
h
where h is the Planck’s constant, 6.63 x
10-34 J⦁s
Max Planck

“energy is
quantized”
Photoelectric Effect (1905)

• light exists as stream of particles


called photons

(dual nature of light)

Einstein
Spectroscopy vs Spectrometry

• Spectroscopy is the study of the interaction between


electromagnetic radiation and matter.
• Spectrometry is the measurement of electromagnetic radiation
to obtain information about a system
Emission Spectroscopy

• methods in which the stimulus for the


analyte is the application of heat or
electrical energy

* the radiant power emitted by the analyte as


it returns to the ground gives information
about its identity and concentration
Emission Spectroscopy

Chemiluminescence refers to excitation of the analyte by


chemical reaction
Emission Spectroscopy

Bioluminescence
Absorption Spectroscopy

• the amount of light absorbed as a function of wavelength is


measured, which can give qualitative and quantitative
information about the sample
Photoluminescence Spectroscopy

• the emission of photons is measured following absorption


• fluorescence and phosphorescence spectroscopy
Absorption Spectra

Atomic Absorption
• When UV or VIS radiation is passed
through a medium containing gaseous
atoms, the outermost electron of the atom
is promoted from the ground state to
higher energy level orbitals.

• As a result, the absorption spectra consist


only of specific wavelengths that
correspond to the energies of the
electronic transitions between orbitals.
Absorption Spectra

Molecular Absorption
• Aside from electronic transitions, molecules also undergo
vibrational and rotational transitions.
Absorption Spectra

Molecular Absorption
• Infrared radiation is not energetic
enough to cause electronic transitions,
but can induce vibrational and rotational
states.
• UV and VIS radiations are more
energetic.
Emission Spectra

Line Spectra
• occur when the radiating species
are individual atoms or ions in
the gas phase
Emission Spectra

Band Spectra
• produced due to presence of
gaseous radicals or small
molecules
Emission Spectra

Continuous Spectra
• produced when solids, such
as tungsten, are heated to
incandescence
Fluorescence Spectra

Atomic Fluorescence
• gaseous atoms fluoresce when exposed to radiation that has a
wavelength that exactly matches that of one of the absorption
(or emission) lines of the element (resonance fluorescence)
Fluorescence Spectra

Molecular Fluorescence
Non-radiative relaxation
• vibrational deactivation occurs during collision between
excited molecules and molecules of the solvent ina series of
steps, with an average lifetime of 10-15 s
• internal conversion occurs between two vibrational levels,
but less efficient than vibrational relaxation with an average
lifetime of 10-9 to 10-6 s
Fluorescence Spectra

Molecular Fluorescence
Fluorescence requires structural features that slow the rate of the non-radiative
relaxation process and enhance the rate of fluorescence emission.
Research Application of
Spectrophotometry
APPLICATION OF SPECTROPHOTOMETRY
• Most commonly used in biomedical and life science research,
which includes both academic and industrial research.
• Typical spectrophotometry applications are the measurements of
nucleic acids, proteins and bacterial density.
• It is widely utilized in a variety of industries including
biotechnology, diagnostic and clinical testing, drug discovery,
pharmaceutical research, chemical engineering, material science
and agricultural research.
Research Application of
Spectrophotometry
APPLICATION OF SPECTROPHOTOMETRY
• Chemical classification methods distinguish compounds
according to their structural properties, commonly by assessing
physical properties (molecular weight, electrical charge,
solubility, pH) or chemical or biological function.
• It quantifies the concentration of compounds at one or a variety
of wavelengths. Thus, the researcher would select between
chemical classification methods or spectrophotometry depending
on the type of information that they want to obtain from the
substance.

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