ATOMIC FLUORESCENCE SPECTROSCOPY

Introduction

Atomic fluorescence spectroscopy is an analytical quantitative technique employed for the

determination of a concentration of elements present in a sample. It is used mostly in the
analysis of more than 65 metals in biological samples, agricultural samples, pharmaceuticals,
environmental, water, and industrial oils. The sample is converted to gaseous atoms, and the
element of interest is excited to a higher electronic energy level by a light source. Following
excitation, the atoms are deactivated by the emission of a photon. Atomic fluorescence
spectroscopy makes use of the same basic instrumental components as atomic absorption
spectroscopy; however, it measures the intensity of the light emitted by atoms that have been
excited from their ground state by the absorption of light of shorter wavelength than that
emitted.

Origin

The origin of atomic fluorescence can be traced back to the early experiments on atomic
spectroscopy by scientists such as Gustav Kirchoff and Robert Bunsen in mid-19 th century. In
year 1920, Wood observed for the very first time of the atomic fluorescence from excited
atoms. The development of atomic fluorescence spectroscopy was invented by Winefordner
and Vickers in 1964 to analyze the chemical concentration of a sample. The idea was to excite
a sample vapor with the appropriate UV radiation, and by measuring the emitting radiation,
the amount of the specific element being measured could be quantified.

Basic Principle

The emission of radiation energy in the UV-visible range from gas-phase atoms that have been
stimulated to higher energy levels by absorbing radiant energy is known as atomic fluorescence
spectroscopy, or AFS. The atom is typically obtained in a gaseous condition by means of a
flame. The process of emission is radiative and moves from the singlet ground state (S 0) to the
lowest singlet (S1).

In simple wording, a sample absorbs photons and becomes more excited when it is exposed to
flame or other radiation sources. Since excited state atoms are unstable, they release a photon
to return to the ground state. The amount of each element contained in a sample can be
ascertained by measuring the photon that is released.

Page 1 of 7
 .

Figure 1: Principle of AFS

Fluorescence Intensity and Analyte Concentration:

There are two major processes includes in fluorescence emissions that are given below.

 Absorption of radiation to generate excited atoms

 De-excitation of excited atoms by emission of radiation

The Beer-Lambert law is used to calculate the relationship between fluorescence intensity and
concentration. As we know that according to Beer's law, while a radiation of intensity P0 is
passed through an analyte of concentration (c) mol dm-3 taken in a cell of thickness (b) cm, an
intensity of transmitted radiation (P) by an analyte is given by Lambert-Beer's law,

viz., P = P0.e -εcb

Steps:

The following things happen quickly after a solution containing a sufficient compound of the
metal under investigation is inhaled into a flame (atomization can also be done without a
flame).

Desolvation: Desolvation involves drying a sample in a solution. The metal particles in the
solvent are dehydrated by the flame and thus solvent is evaporated.

Vaporization: The metal particles in the sample are also dehydrated. This also led to the
evaporation of the solvent.

Page 2 of 7
 Atomization: Atomization is the separation of all atoms in a chemical substance. The metal
ions in the sample are reduced to metal atoms by the flame.

Excitation: The electrostatic force of attraction between the electrons and nucleus of the atom
helps them to absorb a particular amount of energy. The atoms then jump to the higher energy
state when excited.

Emission: Since the higher energy state is unstable the atoms jump back to the ground state or
low energy state to gain stability. This jumping of atoms emits radiation with characteristic
wavelength. The radiation is measured by the photo detector.

Types of Atomic Fluorescence Transitions:

 Resonance fluorescence
 Stokes direct line fluorescence
 Stepwise line fluorescence
 Two-step excitation or double resonance
 Sensitized fluorescence
Their explanation is as follows.
 Resonance Fluorescence
When the excited states emit a spectral line of the same wavelength as that of excitation,
resonance fluorescence occurs. It is mostly employed for analytical determination.
 Stokes direct line fluorescence
It is observed when an atom that has been excited to higher radiation absorption emits radiation
and returns to a lower intermediate level. It lowers to the ground state by a radiation-less
process from this intermediate level. As a result, the wavelength at which direct line
fluorescence occurs will never be the same as the wavelength of the resonance line that excites
it. It is also called Stokes fluorescence. Resonance fluorescence encounters interference from
scattered radiation, which can be eliminated by utilizing direct line fluorescence.
 Stepwise line fluorescence
In this kind of fluorescence, an atom that has been excited to a higher energy state by the
absorption of radiation is deactivated to a lower excited state through a radiation less process,
where it emits radiation to return to the ground state.
 Two-step excitation or double resonance

Page 3 of 7
 Two dye lasers are used in a two-step excitation procedure for the double resonance
fluorescence. The analyte is excited by the first laser from its ground state to an excited state,
from which it is excited by the second laser to an even higher excited state. Fluorescence
emission occurs along with the de-excitation of this more highly excited state to a lower energy
state. This is called double resonance fluorescence.
 Sensitized fluorescence
Sensitized fluorescence occurs when an excited atom transfers its energy to another atom,
which is then excited and relaxes again while emitting fluorescence. However, sensitized
fluorescence is generally not employed for analytical purposes.

Figure 2: Types of AFS Transitions

Instrumentation of Atomic Fluorescence Spectroscopy:

The basic layout of an AFS instrument is similar to those for AAS and AES except that the
light source and the detector are located at a right-angle. Although the configuration of an AFS
instrument can be modified for different purposes, the basic apparatus consists of:

Light Source: Employs a high-intensity lamp emitting resonance lines of the analyte for
efficient excitation.

Page 4 of 7
Sample Introduction System: Transports the sample, utilizing techniques like flame or
graphite furnace atomization to generate free atoms.

Monochromator: Selects a specific fluorescence wavelength with precision, enhancing

analytical specificity. AFS contains two monochromators i.e.; excitation and emission
monochromators.

Sample Cells: They are used to hold sample, basically cuvettes.

Detector: Utilizes sensitive photomultiplier tubes to measure the intensity of the selected
fluorescence, ensuring accurate signal detection.

Amplifier: Collects, processes, and records signals from the detector for subsequent analysis.

Readout Device: Manages overall instrument operation, including parameter settings and
coordination of the different components.

Figure 3: Instrumentation of AFS

Working:
The light source produces light photons over a broad energy spectrum, typically ranging from
200 to 900 nm. Photons impinge on the excitation monochromator, which selectively transmits
light about the specified excitation wavelength. The transmitted light passes through adjustable
slits, then into the sample cell causing fluorescent emission by fluorophores within the sample.
Emitted light enters the emission monochromator, which is positioned at a 90° angle from the
excitation light path to eliminate background signal and minimize noise due to stray light.

Page 5 of 7
 Finally, the emitted light entering the photomultiplier tube which the signal is amplified and
creates a voltage that is proportional to the measured emitted intensity.

Figure 4: Working process of atomic fluorescence Spectrometer.

Advantages of Atomic Fluorescence Spectroscopy:

 It is a relatively simple and more sensitive technique than atomic absorption spectroscopy.
 Precision up to 1% can be achieved.
 More sensitivity even at low concentrations.
 Without the use of chemical separation methods, two substances that are stimulated at the same
wavelength but emit at distinct wavelengths can be easily distinguished.

Disadvantages of Atomic Fluorescence Spectroscopy

 It has the drawback of quenching and background noise brought on by radiation dispersion by
flame particulate matter, especially with refractory samples in high-temperature flame.
 Not applicable for all compounds due to fluorescence issues.
 Not ideal for identification.

Page 6 of 7
 Applications of Atomic Fluorescence Spectroscopy

 More than sixty distinct elements contained in a wide range of samples have been analyzed
using atomic fluorescence spectrometry. AFS is typically used to determine the concentration
of elements in samples.
 Applied in industries like metallurgy and semiconductor manufacturing for quality control,
ensuring precise metal content in materials.
 AFS is employed in pharmaceutical laboratories for quality control and analyzing trace metal
impurities in drug formulations.
 For the determination of sulphide ion in sewage waters.
 In petrochemical for the quantitative determination of Pb, Hg, Cd, As, Sn, Zn in fuels,
lubricant, and crude oil.
 Determination of Hg, Pb, As, and Se in active ingredients and fillers (Pharmaceutical industry).

Page 7 of 7