DR.
CATHERINE NGAMBI
FACULTY OF SCIENCE
DEPARTMENT OF BIOCHEMISTRY AND MOLECULAR BIOLOGY
BIOC 203; INTRODUCTION TO LABORATORY TECHNIQUES I
ASSIGNMENT
NAME: LINOX OCHIENG WASONGA
REGISTRATION NUMBER: S15/03116/23
COLLECTION DATE: 4TH MARCH,2025
DESCRIBE THE ATOMIC ABSORPTION SPECTROPHOTOMETRY UNDER THE
FOLLOWING SUB-HEADINGS;
1. BASIC PRINCIPLES OF ATOMIC SPECTROMETERS
2. INSTRUMENTATION
3. APPLICATION
Atomic Absorption Spectrophotometry
Atomic Absorption Spectrophotometry (AAS) is an analytical technique used to determine the
concentration of metal elements in a sample by measuring the absorption of light by free atoms in the
gaseous state. It is widely used in chemistry, environmental science, clinical analysis, and industrial
applications.
1. Basic Principle of Atomic Spectrometers
Atomic spectrometers operate based on the interaction of electromagnetic radiation with free atoms in
the gaseous state. The fundamental principle relies on the fact that each element has a unique
electronic structure, meaning it absorbs or emits radiation at characteristic wavelengths.
1. Atomization – The sample is converted into free atoms using a heat source, such as a flame, graphite
furnace, or plasma. This step is essential to ensure that the atoms are in their ground state for
absorption or excitation.
2. Excitation or Absorption – Depending on the spectroscopic technique used:
Atomic Absorption Spectroscopy (AAS): Atoms absorb light of a specific wavelength corresponding to an
electronic transition, reducing the intensity of incident radiation.
Atomic Emission Spectroscopy (AES): Atoms, after excitation, emit light at characteristic wavelengths
when returning to their ground state.
Atomic Fluorescence Spectroscopy (AFS): Atoms absorb radiation and re-emit it at a longer wavelength.
3. Wavelength Selection – A monochromator isolates the specific wavelength of interest, ensuring only
relevant radiation reaches the detector.
4. Detection and Measurement – A photodetector records the intensity of absorbed or emitted light,
and the concentration of the element is determined using a calibration curve based on Beer-Lambert’s
law.
2. Instrumentation of Atomic Absorption Spectrophotometry (AAS)
Atomic Absorption Spectrophotometry (AAS) consists of several key components that work together to
measure the absorption of light by free atoms in a sample. The main components include:
1. Radiation Source (Hollow Cathode Lamp or Electrodeless Discharge Lamp)
Emits light at a specific wavelength corresponding to the element being analyzed.
The hollow cathode lamp (HCL) contains a cathode made of the element of interest, ensuring selective
emission.
Electrodeless discharge lamps (EDL) are used for elements requiring higher intensity radiation.
2. Atomizer (Flame or Graphite Furnace)
Converts the sample into free atoms in the gaseous state.
Flame Atomization: The sample is aspirated into a flame (e.g., air-acetylene or nitrous oxide-acetylene)
for atomization.
Graphite Furnace Atomization: A graphite tube electrically heats the sample to higher temperatures,
enhancing sensitivity for trace metal analysis.
3. Monochromator
Isolates the specific wavelength absorbed by the element of interest.
Helps eliminate interference from other wavelengths.
4. Detector (Photomultiplier Tube or Photodiode)
Measures the intensity of light before and after absorption by the sample.
Converts light signals into electrical signals for quantification.
5. Data Processing System (Computer or Digital Readout System)
Analyzes and processes the electrical signals.
Displays the concentration of the element based on a calibration curve.
Working Principle
The hollow cathode lamp emits element-specific radiation.
The sample is atomized in a flame or graphite furnace.
Atoms absorb radiation at their characteristic wavelengths.
The monochromator isolates the desired wavelength.
The detector measures the light absorption, which is proportional to the element concentration.
3. Applications of Atomic Absorption Spectrophotometry (AAS)
Atomic Absorption Spectrophotometry (AAS) is widely used for the quantitative and qualitative analysis
of metal elements in various fields. Its high sensitivity, selectivity, and precision make it suitable for
detecting trace metals in complex samples. Below are the major applications of AAS:
1. Environmental Analysis
Water and wastewater analysis: Detects heavy metals such as lead (Pb), mercury (Hg), arsenic (As),
cadmium (Cd), and chromium (Cr) in drinking water, industrial effluents, and wastewater.
Soil and sediment analysis: Determines metal contamination in agricultural and industrial soils.
Air quality monitoring: Measures metal pollutants in atmospheric particles and emissions from
industries.
2. Clinical and Biomedical Applications
Blood and urine analysis: Measures metal concentrations such as iron (Fe), copper (Cu), zinc (Zn),
magnesium (Mg), and lead (Pb) for diagnosing deficiencies or toxicities.
Toxicology studies: Detects heavy metal poisoning (e.g., lead poisoning in children or arsenic exposure).
Nutritional studies: Analyzes trace elements essential for human health.
3. Pharmaceutical Industry
Quality control of drugs and supplements: Ensures compliance with regulatory standards by detecting
metal impurities in raw materials and finished pharmaceutical products.
Bioavailability studies: Measures the absorption of metal-based drugs in the human body.
4. Food and Beverage Industry
Food safety testing: Detects toxic metals like lead, cadmium, and mercury in food products such as
vegetables, fruits, seafood, and dairy.
Nutritional analysis: Measures essential minerals like calcium (Ca), iron (Fe), and zinc (Zn) in food and
beverages.
Contaminant monitoring: Ensures compliance with food safety regulations by testing for metal residues
in agricultural products.
5. Industrial and Manufacturing Applications
Metal and alloy analysis: Determines the composition of metals and alloys in metallurgy, welding, and
automotive industries.
Petroleum and fuel analysis: Detects metal impurities in crude oil, gasoline, and lubricants that could
affect engine performance.
Cosmetics and personal care products: Ensures that cosmetic products comply with safety regulations
regarding heavy metal contamination.
6. Geology and Mining
Ore and mineral analysis: Determines the concentration of metals in ores and geological samples for
mining and exploration.
Metal recovery and recycling: Evaluates the efficiency of metal extraction and recycling processes.
7. Agriculture and Veterinary Science
Soil fertility studies: Measures trace metals in soil to optimize crop nutrition.
Animal feed analysis: Ensures appropriate mineral content in livestock feed.
Plant tissue analysis: Monitors nutrient uptake and heavy metal accumulation in plants.
8. Forensic Science
Crime scene investigations: Detects metal traces in forensic evidence (e.g., gunshot residue, poisons).
Poisoning cases: Identifies toxic metal exposure in criminal investigations.
9. Nuclear and Energy Sector
Radioactive metal analysis: Measures trace metals in nuclear reactor materials and waste management.
Battery and renewable energy research: Analyzes metal content in solar panels, batteries, and fuel cells.
4. Sample Preparation and Analysis in Atomic Absorption Spectrophotometry (AAS)
The accuracy and precision of Atomic Absorption Spectrophotometry (AAS) depend heavily on proper
sample preparation and analytical procedures. The process involves several key steps to ensure reliable
results.
1. Sample Preparation
A. Sample Collection and Handling
Ensure that samples are collected in clean, contamination-free containers.
Use plastic or acid-washed glass containers for metal analysis.
If necessary, store samples in a refrigerator to prevent degradation.
B. Sample Digestion (for Solid and Biological Samples)
Objective: Convert solid samples into a liquid form by dissolving or digesting them.
Common Digestion Methods:
Acid Digestion: Strong acids (e.g., HNO₃, HCl, H₂SO₄) break down solid samples.
Microwave Digestion: Uses microwave energy for efficient sample decomposition.
Dry Ashing: Samples are heated in a furnace at high temperatures to remove organic matter, leaving
behind metal oxides.
Wet Ashing: A combination of acids and heat is used to dissolve metals while removing organic matter.
C. Filtration and Dilution
After digestion, samples may be filtered to remove undissolved particles.
Dilute samples to bring metal concentrations within the detection range of AAS.
D. Matrix Modification (for Graphite Furnace AAS)
Some samples require matrix modifiers to prevent interference and improve sensitivity.
2. Analysis Using AAS
A. Instrument Calibration
Prepare standard solutions of the metal being analyzed.
Generate a calibration curve by measuring the absorbance of standard solutions at known
concentrations.
Verify calibration accuracy using quality control samples.
B. Sample Introduction into the AAS Instrument
1. Flame AAS (FAAS)
The sample is aspirated into a flame (air-acetylene or nitrous oxide-acetylene).
The solvent evaporates, and metal atoms are converted into free atoms in the gaseous state.
Absorption of light from the hollow cathode lamp (HCL) is measured.
2. Graphite Furnace AAS (GFAAS)
A small volume of sample (typically 5–20 µL) is injected into a graphite tube.
The furnace heats the sample in stages (drying, ashing, atomization).
The absorbance of the element at its characteristic wavelength is recorded.
C. Measurement of Absorbance
The monochromator selects the specific wavelength absorbed by the target element.
The detector measures the reduction in light intensity, which is proportional to the metal concentration.
D. Data Analysis and Interpretation
Compare the sample’s absorbance with the calibration curve to determine metal concentration.
Apply dilution factors if necessary to obtain the final concentration.
3. Quality Control and Validation
Use Certified Reference Materials (CRMs): To verify accuracy.
Perform Blank Sample Analysis: To detect contamination.
Repeat Measurements: To ensure precision.
Check for Interference Effects: Matrix interferences, spectral interferences, and ionization effects should
be controlled.
5. Types of Atomic Absorption Spectrophotometry (AAS)
Atomic Absorption Spectrophotometry (AAS) is classified based on the method of atomization used to
convert the sample into free atoms for absorption measurement.
1. Flame Atomic Absorption Spectrophotometry (FAAS)
Principle:
The sample solution is nebulized and introduced into a flame (air-acetylene or nitrous oxide-acetylene).
The heat of the flame converts metal ions into free atoms.
The absorption of light from a hollow cathode lamp (HCL) at a specific wavelength is measured.
Features:
Fast and widely used for routine analysis.
Suitable for detecting moderate to high concentrations (ppm level).
Requires large sample volumes (several mL).
Applications:
Environmental analysis (water, soil).
Food and beverage industry (trace metals in food).
Clinical and biomedical applications (blood, urine).
2. Graphite Furnace Atomic Absorption Spectrophotometry (GFAAS)
Principle:
A small volume (5–20 µL) of the sample is injected into a graphite tube (electrothermal atomizer).
The sample undergoes stepwise heating (drying, ashing, atomization).
The absorption of light at a specific wavelength is measured.
Features:
Higher sensitivity than FAAS (detects ppb levels).
Requires smaller sample volumes.
More prone to matrix interferences, requiring careful optimization.
Applications:
Toxicology (lead, arsenic, cadmium in biological fluids).
Pharmaceutical industry (trace metal impurities in drugs).
Food safety (low-level contaminants in food).
3. Hydride Generation Atomic Absorption Spectrophotometry (HGAAS)
Principle:
Used for elements that form volatile hydrides (e.g., arsenic, selenium, antimony, tin).
A chemical reaction generates volatile metal hydrides, which are transported into the atomization
chamber.
The hydrides decompose, releasing free atoms for absorption measurement.
Features:
Highly selective and sensitive for hydride-forming elements.
Detects metals at ultra-trace levels (ppt to ppb).
Requires special reagents and sample preparation.
Applications:
Environmental monitoring (arsenic in drinking water).
Food safety (selenium in grains, fish).
Clinical toxicology (arsenic poisoning).
4. Cold Vapor Atomic Absorption Spectrophotometry (CVAAS)
Principle:
Designed specifically for mercury (Hg) analysis.
Mercury is reduced to its elemental form (Hg⁰) using a reducing agent (e.g., stannous chloride).
The vaporized mercury is transported into an absorption cell, where its absorbance is measured.
Features:
Extremely sensitive for mercury detection (ppt level).
Requires a closed system to prevent mercury loss.
Specific to mercury and not applicable to other metals.
Applications:
Environmental analysis (mercury in water, soil, and air).
Food and pharmaceutical testing (mercury in fish and medicines).
Occupational health monitoring (mercury exposure in workers).
6. Advantages of AAS
1. High Sensitivity and Selectivity
AAS can detect trace amounts of metals (ppb to ppm levels), making it useful for environmental, clinical,
and industrial applications.
The use of element-specific hollow cathode lamps (HCL) enhances selectivity for individual metals.
2. Rapid and Simple Analysis
AAS provides quick results with minimal sample preparation, especially in Flame AAS (FAAS).
Routine analysis of multiple samples is possible with automated systems.
3. Wide Range of Element Detection
AAS can analyze over 70 metal elements, including essential and toxic metals like lead (Pb), mercury
(Hg), and arsenic (As).
Specialized techniques like Hydride Generation AAS (HGAAS) and Cold Vapor AAS (CVAAS) enhance
detection for specific elements.
4. Cost-Effective and Accessible
Compared to other techniques like Inductively Coupled Plasma (ICP), AAS is more affordable and
requires less complex instrumentation.
Maintenance and operation costs are lower.
5. Minimal Matrix Interference (with Proper Optimization)
Graphite Furnace AAS (GFAAS) allows precise measurement with small sample volumes.
Use of matrix modifiers can reduce interference effects.
7. Limitations of AAS
1. Limited to Metal Analysis
AAS cannot detect non-metal elements like carbon, nitrogen, or sulfur.
It is restricted to analyzing only free metal atoms in a sample.
2. Single-Element Analysis at a Time
Unlike ICP-MS or ICP-OES, which can analyze multiple elements simultaneously, AAS measures one
element per analysis cycle using different lamps.
3. Matrix Interference and Sensitivity Issues
Chemical and spectral interferences can affect accuracy, especially in complex samples.
Requires careful calibration and matrix modification to reduce interferences.
4. Flame AAS Has Lower Sensitivity
FAAS detection limits are in ppm range, making it less suitable for ultra-trace metal analysis.
GFAAS or ICP-MS is preferred for ppb or ppt-level detection.
5. Time-Consuming for Large Sample Batches
Since each metal is analyzed separately, analyzing multiple elements in different samples can be time-
intensive.
Requires frequent recalibration when switching between elements.
