Principle
of
Atomic Absorption Spectrophotometry
Mr. Charnchai Suracheep
Introduction
Atomic Absorption Spectrophotometry,
which are standard instruments for
the determination of metal elements,
are widely applied of samples, such as
agriculture chemical, clinical and
biochemistry, minerals, food and
drugs, environmental and other.
Principle of Atomic Absorption
Spectrophotometer
Principle of the Atomic Absorption
Method
Atomized elements each absorb energy of a
wavelength that is peculiar to that element. The
atomic absorption method uses as its light source a
hollow cathode lamp which emits light of a
wavelength that is peculiar to each element.
Elements within a solution are heated in a flame or
electrically (2000K to 3000K) and subsequently
determined using the fact that the degree of
absorption will vary with its concentration.
Light absorption
process of atoms
Principle of Atomic Absorption
Spectrophotometer
Atomic Absorption Spectroscopy, AAS
Excited state E1
Absorption
e
Ground state E0
e
Atomic Emission Spectroscopy, AES
Excited state E1
Emission
e
Ground state E0
Electronic Transition
Sodium (Na) energy states
Excited state (II) 3.6 eV
Excited state (I) 2.2 eV
330.3 nm
589.0 nm
Ground state 0.0 eV
Relation between light absorption
and density
• When light of a certain intensity is given to
many atom in ground state, part of this light is
absorbed by atoms.
Density C
I
I0
l
Relation between light absorption
and density
Lambert-beer’s Law
Density C
I
I0
l
I = I0 e-k .l .C
Abs = -logI/I0 = k .l. C
k : proportional constant
l : path length
C : density (concentration)
Relation between light absorption
and density
Calibration curve
Absorbance
• Graph show the relation between absorbance and concentration
Absorbance of
unknown sample
Concentration of
unknown sample
Concentration (ppm)
Atomization method
• Atomic absorption spectrometry measures
absorption of free atom.
• “Free atom” means an atom not combined with
other atoms.
• Elements in the sample to be analyzed are not in
the free state, and are combined with other
elements invariably to make a so-called molecule.
Atomization method
• The combination must be cut off by some means to
free the atoms.
• This is called “atomization”
• 2 types:
- Flame method
- Flameless method
Flame Method
With the Flame Method, the sample
solution is converted into mist-form using a
nebulizer, and then introduced into the
flame. It is atomized by the temperature of
the flame.
Measurement time: A few dozen seconds
Flame Atomization Method
Optical diagram of Flame Atomic
Absorption Spectrometers
Flame Method
Flame selection
• These flames vary in temperature, reducibility and
transmission characteristics.
• Selected according to the element being analyzed,
and properties of the sample.
• Argon-hydrogen
:
Max. temp. 1,577 0C
• Air-hydrogen
:
Max. temp. 2,045 0C
• Air-acetylene
:
Max. temp. 2,300 0C
•Nitrous oxide-acetylene :
Max. temp. 2,955 0C
(For elements are hard to combine with oxygen (Al, Si, V, Ti, etc.))
Flame Method
Flame selection
Flameless Method (Graphite Furnace)
Graphite holder
Graphite cap
Cooling block
Sample
inlet
Aperture
plate socket
シール
Seal
Graphite tube
Graphite tube
Eject arm
Spring
Fixing knob
Flameless Method (Graphite Furnace)
• Sample is injected in the formed graphite tube.
• An electric current of 300 ampere (maximum) is
applied to the tube.
Flameless Method (Graphite Furnace)
• In an actual measurement heating is done in 3 stage.
- Drying stage (100oC)
- Ashing stage (400-1000oC)
- Atomizing stage (1400-3000oC)
Other atomic absorption methods
• Methods having higher sensitivity than
normal flame atomic absorption or electrothermal atomic absorption
• Used for special elements including arsenic,
selenium and mercury.
• Use chemical reactions in the process of
atomization to vaporize in the form of an
atom or simple molecule.
Hydride Vapor Generation
Technique
• As, Se, Sb, Sn, Te, Bi, Hg and other metals produce a metal hydride by this method
Elements
Concentration (ppb)
As
5~20
Sb
5~20
Te
5~20
Bi
5~20
Se
10~40
Hg
20~80
Sn
30~90
6BH4- +As3++ 3H+
3B2H6+3H2 +AsH3 (gas)
Absorption Cell
Peristaltic
Pump Manifold
Burner Head
of AAS
Gas Liquid
Separator
Reaction
Coil
Drain
Sample HCl NaBH4
Carrier Gas Ar
Structural Diagram of Hydride Vapor Generator
Cold Vapor Technique
SnCl2 + Hg2+
reduce
 253.6 nm
Hgo(gas)
Ho
SnCl2
5%KMnO4
5%H2SO4
Limit of Quantitative
Element
Detection Limit
Flame (ppm)
Furnace (ppb)
Ag
0.04
0.01
Al
0.5
0.03
As
0.02 ppb (HVG)
0.2
As
0.4
-
Cd
0.012
0.003
Cr
0.08
0.015
Cu
0.04
0.008
Hg
0.01 ppb (cold vapor)
-
Hg
0.2 ppb (HVG)
-
Mg
0.0035
0.003
Mn
0.025
0.01
Ni
0.08
0.13
Pb
0.2
0.06
Se
0.3 ppb (HVG)
0.2
Sn
2 N2O-C2H2
2
Zn
0.01
0.01
Interference effects
• Physical interference
• Spectral interference
• Chemical interference
Physical interference
• Flame
– Spray efficiency fluctuations due to difference in
viscosity and surface tension between the standard
and sample.
• Furnace
– Sample dispersion ;
Measurement value fluctuations due to tube temperature distribution
– Viscosity within the graphite furnace ;
Adherence to sample tip causing errors in collection quantity.
• Example: samples, such as blood or juice, containing numerous
organic components.
Spectral interference
• Spectral absorption line overlapping with the
absorption line of the target element.
• Absorption and scattering by molecules
Spectral interference
Spectral absorption line overlapping with the absorption
line of the target element.
Target element
Al
Ca
Cd
Co
Cu
Fe
Ga
Hg
Mn
Sb
Si
Zn
Spectral line
(nm)












Interfering
element
V
Ge
As
In
Eu
Pt
Mn
Co
Ga
Pb
V
Fe
Spectral line
(nm)












Spectral interference
• Absorption and scattering by molecules
– Molecules absorption
• Alkaline metals + Halogens = Alkali halides
(Na, K)+(F, Cl, Br, I) = (Ex: NaCl, KI)
Chemical interference
• Generation of non-separable compounds by coexisting
matrices
–Example : influence of PO4-, SO4-, SiO2 relative to Ca, Mg
in flame analysis
• (generation of Ca2PO4)
• Generation of low boiling point compounds by
coexisting matrices
–Example: influence of chloride ions relative to Cd in
furnace analyses
• (generation of CdCl2)
Matrix modifier effect
• Masking of obstructing matrices
• Influence of phosphate on Ca is masked by La
• Conversion of obstructing matrices to compounds
that easily undergo sublimation or evaporation
– Sublimation agent
• Example: removal of chloride ion by ammonium salt of nitric
acid or phosphoric acid
• Conversion of measured elements to stable oxides
or metallic intermediary compounds
– Stabilizing agent:
• Example: creation of measured element alloy using white
metals (Pd, Pt, Rh)
Application
examples of the matrix modifier method
Standard Addition Method
No.1
No.4
No.2
No.3
10 ml
20 ml
10 ml
10 ml
10 ml
10 ml
X
X+0.1
X+0.2
X+0.3
100 ml
Mg concentration
after filled up
10 ml
Unknown sample
30 ml
Solvent
1.0 ppm X Standard solution (ppm : mg/1000ml)
Standard Addition Method
Calibration Curve of Standard Addition Method
Concentration of
unknown sample
Background Correction
2-Way Background Correction is
Standard
•D2 lamp method ( 190-430 nm) – Molecular absorption
•Self-Reversal (SR) method – Spectra interference
Background Correction
Spectral interference
Target element
Al
Ca
Cd
Co
Cu
Fe
Ga
Hg
Mn
Sb
Si
Zn
Spectral line
(nm)












Interfering
element
V
Ge
As
In
Eu
Pt
Mn
Co
Ga
Pb
V
Fe
Elements/ wavelengths where spectral
interference becomes problematic
Spectral line
(nm)












Background Correction
Self-Reversal Method
Background Correction
Self-Reversal Method
Background
100 mA
10 mA
Signal
Atomic Absorption Spectrophotometer
AA-6300
High Performance Optical System
Optical diagram of Double Beam System
Easy Switching between Flame and Furnace
Flame -> Furnace: All that is involved is to remove the burner head,
place the furnace unit, and fix it with the screw. No tools are required.
Remove the burner head.
Fit the furnace.
Fit the burner head.
Remove the furnace.
New Flame Atomizer
For chemical resistance
• Neburizer w/ Ceramic
made Impact Bead
• Polypropylene-made
Chamber
• Solid Titanium-made
Burner Head
High Productivity
• Full Auto ASC
- Auto measurement up to 60 samples
- Reagent addition 8 position
- Automatic dilution
• Optimize Flame analysis
- Automatic search the best fuel gas flow rate
- Automatic search the Optimize Flame
analysis best burner height
Enhanced Safety
•
•
•
•
Auto Gas Leak Check
Gas pressure monitoring to prevent flashback
Automatic flame monitoring
Automatic flame extinguish when
power failure
High Temp. Burner
• Safety interlock for burner misuse
• Auto Air/N2O flame changeover
• Drain level sensor
Drain level sensor
Wizard Software System
* Select
elements
*Set the calibration curve
and samples condition
*Connect to PC
*Set the spectrophotometer
* Set the atomizer
Automated/ Optimized
Effectiveness of the automatic
Line Search/Beam Balance
Automated/ Optimized
Effectiveness of the automatic
burner height
(Cr : 4ppm standard solution used)
Burner height & Sensitivity (Cr)
Automated/ Optimized
Search for the optimal fuel flow rate
(Cu : 4ppm standard solution used)
Screen during measurement
Signals in real-time
The 4 newest
signals
Display of
saved signal
Calibration
curve
User Management
• The Login ID and password need to be entered when the software is started up.
• Records of who logged in at what time are preserved in the “Event Log”.
User Management
Authority can be set in detail for each user
Initial Validation Screen
Summary Validation Report
Application
of
Atomic Absorption Spectrophotometry
Application of AAS
AAS
Pretreatment (dissolution) is required for solid samples.
Pretreatment
Precautions for pretreatment:
 Dissolve all the elements into the same solution evenly.
(Check with certified reference material.)
 Ensure that elements are not lost in the solution. i.e., due to vaporization
or sedimentation (Check with recovery test.)
 Contamination : Purified water, reagent (e.g., acid), container,
environment. (Check with blank operation.)
 Ensure that the solution to be analyzed is stable for a long time (i.e., no
hydrolysis or sedimentation).
 Consider the interference effect of the reagent on the analysis values.
Types of Pretreatment
 Dilution
Dilute the sample with purified water, dilute acid, or organic solvents.
Examples: food products (e.g., dairy products), pharmaceuticals, and biological
samples (e.g., blood, urine).
 Dry Decomposition
Heat the sample to a high temperature (400 to 500C), Decomposition is possible in a
short time (a few hours) and operation is simple.
Elements with low boiling points (e.g., Hg, As, Se, Te, and Sb) will vaporize
 Wet Decomposition
Heat the sample together with acid to a low temperature (approx. 300C). Suitable for
volatile elements.
A long time is required for the decomposition of organic substances.
 Microwave Decomposition
Decompose the sample at high pressure by heating it together with acid to a
temperature in the range 100 to 200C in a sealed Teflon container.
The decomposition process is sealed; there is little vaporization of elements with low
boiling points; the decomposition time is short; there is little contamination from the
operating environment and the reagent; and only a small amount of acid is required.
Examples: Sediment, soil, dust, ceramics, living organisms, food products, etc.
Wet Decomposition Method
Kjeldahl flask wet decomposition method
Waste
gas
Cooling
tube
Nitric
acid
Simple method
(no cooling)
Sample+
Sulfuric acid
Heating
Pretreatment
Microwave Decomposition
 Decompose the sample together with an acid in a sealed container.
 Decomposition possible in a short time with little vaporization or contamination.
- Ideal for the pretreatment of trace elements and trace samples.
- Food products, living organisms, pharmaceuticals, airborne dust, soil, etc.
High-pressure Decomposition Container
Microwave Digestion
Sample Preparation using Pressure Digestion
with Microwave heating
Digestion Vessels 1 - 12
Microwave
power
Pressure
measurement
Temperature
measurement
Control by
Tmax and Pmax
internal PC or
Controller
Real-Time Display
Pretreatment
Solubility of Elements in Samples
Total Content
Simple
water-soluble
ions
Simple soluble metals
& compounds
Organic
compounds
Inorganic compounds
with low solubility
Sulfides, oxides,
silicates, etc.
Carbonates, oxides, etc.
Pretreatment Methods
Dilution, Elution
Wet Decomposition
Purified water,
Hydrochloric acid,
nitric acid, etc.
solvents, etc.
Dry/Wet Decomposition
Microwave Decomposition
Nitric acid,
sulfuric acid, etc.
Wet/High-pressure
Decomposition
Hydrofluoric acid, nitric
acid, etc.
Example
Application of AAS
EU Regulation for Hazardous Substances
EU Regulation for Hazardous Substances
IEC Recommendation for RoHS
RoHS : Restriction of Hazardous Substance in Electrical and Electronic equipment.
Substances
PBB/PBDE :
1000 ppm
Cr6+ : 1000 ppm
Polymers
Metals
Electronics
GC-MS
NA
GC-MS
Colorimetric Method
(Spectrophotometer)
Hg : 1000 ppm
Pb : 1000 ppm
Cd : 100 ppm
Spot-test
procedure/boiling=waterExtraction procedure
(Clause8)
Colorimetric Method
(Spectrophotometer)
Cold Vapor-AAS, ICP
AAS, ICP
AAS, ICP
AAS, ICP
Preparation of circuit boards
Pre-Cutting with the
Heavy Duty Cutting Mill
, bottom sieve 6 mm
Heavy Duty Cutting Mill SM 2000
after a grinding time of 2 min.
endfineness 90 % < 125 µm
Vibratory Disc Mill RS 100
Sample Preparation
Target
Element
Polymer
Pb
Electronics
Microwave digestion (HNO3+ HBF4+ H2O2)
Hg
Cd
Pretreatment Methods
Metals
Microwave digestion
(HNO3+H2O2)
(If contain ing Si, Ti add
HF)
a) Common method
(HCl : HNO3 : water ; 2 : 1 : 2)
Microwave digestion Step
b) If containing Zr, Hf, Ti, Ta, Nb, W A (HNO3+HBF4+H2O2)
(HNO3 : HF ; 1 : 3)
Microwave digestion Step B
(add HCl)
c) If containing Sn
(HCl :HNO3 ; 3 : 1)
Pretreatment method, which follow by IEC 62321
Analyzing Cadmium (Cd) in Rice
Pretreatment Using Wet Decomposition
Level suggested by FAO/WHO Codex Committee ; 0.2 ppm max. in polished rice (proposed)
Put 5 g of the sample in a beaker.
Add 30 mL of nitric acid (1+1) and 0.5 mL of sulfuric acid.
Warm on a hot plate until the violent reaction subsides.
↓
Perform thermal decomposition until the contents approach a hardened and
dried state.
When the contents turn dark brown, add 1 mL of nitric acid. Repeat this process.
When the contents turn light yellow or become transparent, expel the white smoke of
the sulfuric acid and leave to cool.
Add nitric acid.
Heat on the hot plate to dissolve the salt content.
Leave to cool.
Dilute for measurement.
Results of Quantitative
Analysis of Cd in Rice
The following 2 methods can be used to analyze unpolished and polished rice decomposed using acid:
Flame method
Furnace method
Polished rice:
0.118 ppm
0.1 ppm
0.5 ppm
Unpolished rice:
0.070 ppm
Polished rice :
0.118 ppm
Unpolished rice :
0.073 ppm
Air-C2H2
Injected amount: 10 µL
Interference inhibitor: Pd 50ppm 5 µL
Ashing: 400C; Atomization: 1,800C
Summary of Methods for
Analyzing Cd in Rice
Comparison of Pretreatment Methods
Wet oxidation: 3 to 5 hours; Dry ashing: 5 to 10 hours;
Microwave: 1 hour; Acid extraction: 2 hours
Expected Lower Limits for Quantitative Measurement
Flame method : 0.100 ppm
Furnace method : 0.001 ppm
Comparison of Measurement Times for Each Measurement
Method (n=3)
Flame method
: 30 s
Furnace method : 360 s
Preventive Maintenance / Calibration
of
Atomic Absorption Spectrophotometry
Preventive Maintenance/Calibration
Maintenance Weekly check
1. Cleaning the Burner head
Clogged
(by carbide or salt etc.)
Normal
Preventive Maintenance/Calibration
Maintenance Weekly check
2. Cleaning the Chamber with diluted water or alcohol
Burner Head
Nebulizer Construction
Sample Solution
O-ring
Chamber
Disperser
O-ring Fixing Plate
Air
Nebulizer
Drain
Sample Suction
Preventive Maintenance/Calibration
Maintenance Weekly check
3. Cleaning the Nebulizer
Cleaning wire
Nebulizer
Do not apply the ultrasonic cleaner to the nebulizer
Hardware Validation
Preventive Maintenance/Calibration
Calibration (12 Month)
1. Wavelength Accuracy
- Using Hg hollow cathode lamp set at Emission mode
- Measure peaks should be within + 0.7 nm
(253.6nm 365.0nm 435.8nm 546..1nm 585.2 640.2nm)
2. Noise Level
- Using Se hollow cathode lamp ( 196 nm)
- NON-BGC Noise level should be < 0.015 Abs.
- BGC-D2 Noise level should be < 0.035 Abs.
3. Baseline Drift
- Using Cu hollow cathode lamp ( 324.8 nm)
- Measuring time 1800 sec
- Measured value less than 0.006 Abs
Preventive Maintenance/Calibration
Calibration (12 Month)
4. Absorption
- Using Cu hollow cathode lamp
- Standard Cu 2 ppm
- Measured value more than 0.23 Abs
5. Repeatability
- Using Cu hollow cathode lamp
- Standard Cu 2 ppm
- Measure 5 time and CV value < 2%
Preventive Maintenance/Calibration
Calibration (12 Month)
6. Detection Limit
- Using Cu hollow cathode lamp
- Standard Cu 2 ppm
- Measure Standard is 3-5 time and calculate the mean value (A)
- Measure Blank solution is 3-5 time and calculate the standard
deviation (S)
- Take the obtained value as the detection limit < 0.004 Abs.
Detection limit = (2.0 x 3 x S) / A
Preventive Maintenance/Calibration
Calibration (12 Month)
7. Stability
- Using Se and Cu hollow cathode lamp
- Standard Cu 2 ppm
- Measure std. Cu around 5 sec (B)
- Measure std. Cu continuous around 30 sec and measure
amplitude of Abs. value (W)
- Take the ratio of W to B
Stability = W/B < 6.0 %
Preventive Maintenance/Calibration