NON-FLAME AAS
3
3.1 Limitations of flame AAS
In conventional atomic absorption spectroscopy (FAAS) a flame is used as the means of atomisation.
The advantages of this method of atomisation are:

simplicity

speed

most metals atomise in readily available flames

sensitivity suitable for general analyses (mg/L)
Flame AAS was developed in the 1950s, and at this time, the environmental significance of ug/L
levels of metals such as lead and mercury, was not really appreciated. However, this would soon
change, and improvements in sensitivity were an ongoing part of instrumental development (think
about the changes in polarographic instrumentation). A number of factors limit the sensitivity of
flame AAS.
CLASS EXERCISE 3.1
What factors or processes in flame AAS could contribute to sensitivity losses?
To achieve sensitivity improvements of 100 times or more (to get accurate analysis of 10 ug/L levels)
from atomic absorption spectrophotometry would require a considerably different approach: a
system AA system based around a flame and nebuliser will never achieve the sensitivities required.
There are two very different approaches, both of which dispense with the flame:

vapour generation – where the analyte is converted into a volatile form

electrothermal – where electrical heating is used
3.2 Basic principles of electrothermal atomisation
As we have seen, the two major obstacles to improved sensitivity in FAAS were the flame and the
nebuliser. In EAAS each of these is replaced. The sample is delivered as a single aliquot, rather than
a constant flow, into a sample holder, which is heated electrically. As in other forms of spectroscopy,
the absorption beam passes through the sample holder to the monochromator and detector. Table
3.1 summarises the key differences between FAAS and EAAS.
TABLE 3.1 Improvements in EAAS
Problem in FAAS
Approach in EAAS
Flame
No flame; heat is provided electrically
Nebuliser
Aliquot of sample delivered to sample cell, rather than
constant flow; nebuliser and burner is replaced by workhead
3. Non-flame AAS
CLASS EXERCISE 3.2
How are the problems identified in Exercise 3.1 addressed by these changes?
FAAS Problem
Solution in EAAS
3.3 Equipment
The burner/nebuliser in FAAS instruments is removed and the electrothermal workhead put in its
place. Most modern instruments allow the two units to be interchangeable, because the rest of the
instrument is identical:

hollow cathode lamp

monochromator

photomultiplier detector

signal processing and readout
Electrothermal-capable instruments must have computer control to automate the sample delivery,
and to monitor the heating and signal measurement.
The workhead
The workhead contains the sample cell and provides the services for controlled heating of the
sample. The sample cell is a hollow graphite tube approximately 28 mm in length by 8mm in
diameter. The other common name for EAAS is graphite furnace AAS, deriving from the graphite
tube in which the sample is heated. The interior of the cylinder is coated with a layer of pyrolytic
graphite, which has a greater resistance to heat than normal graphite. The sample is delivered into
the graphite tube through a small hole in the barrel of the tube, as seen in
Figure 3.1.
There are two types of graphite tubes – partition and platform. The former
has raised sections one-third of the length along from each end. These act like
dam walls, preventing the droplet of sample spreading out too far. The
platform is a separate piece of pyrolytic graphite sitting in the centre of tube.
The droplet sits in this and supposedly is vaporised more predictably, because
the heating only occurs by radiation (the gas inside) than by conduction (from
the walls).
The tube is positioned inside the workhead so that the delivery hole is
FIGURE 3.1
pointing upwards. It rests on electrodes which are connected to a low-voltage
Graphite furnace
high current power supply which delivers upwards of 3.5kW to the cylinder
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3.2
3. Non-flame AAS
walls. Because graphite is only a moderate conductor of electricity, the tube heats up, providing the
conditions to vaporise and atomise the sample.
The metal housing around the furnace is water cooled to enable rapid restoration of the
furnace to ambient temperature after atomisation. Inert gas (generally argon) is flushed through and
around the tube from the ends. This has three purposes:

to remove steam and smoke vaporised during the heating

to prevent oxidation of the graphite

assist with cooling
Figure 3.2 shows a typical workhead.
Ar flows through and around tube
cooling
water coils
HCL beam
quartz windows
FIGURE 3.2 Workhead
Autosamplers
Modern electrothermal atomisation units almost always use an autosampler to place the sample in
the graphite tube. This is because the volumes involved are typically 5-20 uL, and therefore manual
loading of graphite tubes (even with micropipettes) would be unreliable, with precision being greatly
reduced compared to that obtainable with autosamplers. These also allow multiple samples to be
run without an operator in attendance.
Frequently, different solutions (blank, sample, standard, matrix modifier) are used in
combination for the analysis. These are kept apart in the plastic tubing by air gaps. The sampler arm
then swings over to the workhead and delivers the sample to the graphite tube.
3.4 The heating process
The heating of the sample (or standard) in the graphite furnace does not simply involve an instant
transition to the temperatures encountered in the flame 1800-3000C. Because there is a droplet of
liquid in the tube, such heating would be disastrous for the analysis.
CLASS EXERCISE 3.3
What do you expect would happen to a droplet of milk inside the graphite furnace it is was suddenly
exposed to 2200C?
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3.3
3. Non-flame AAS
Therefore, the heating process must occur gradually, and involves a number of carefully programmed
steps, such as those shown in Table 3.2 and Figure 3.5:
1.
Drying – removal of all volatiles at temperatures slightly above the boiling point of the solvent,
and for 30-60 seconds
2.
Ashing – the temperature is increased to 500-1000C and the organic matter is burnt away,
like in a muffle furnace, for 5-40 seconds
3.
Atomisation – now that all the non-elemental matter is gone, the analyte and any other
inorganic elements in the matrix are heated to the temperatures used in FAAS (1800-3000C)
for 3-4 seconds; measurement of absorbance occurs here
TABLE 3.2 & FIGURE 3.5 Temperature program for Cu in water
Step
1
2
3
4
5
6
7
8
9
Temp. (C)
85
95
120
800
800
800
2300
2300
2300
Time (s)
5
40
10
5
1
2
1.1
2
2
Gas
ON
ON
ON
ON
ON
OFF
OFF
OFF
ON
Designing the temperature program
Each step in the cycle of dry/ash/atomise is important and the correct time and temperature
conditions must be carefully selected. Both the analyte and the matrix will affect the times and
temperatures that are needed. The instrument manual will provide a standard program for all
elements, which is fine for that element in pure water or dilute acid, such as that for copper in Table
3.2. This can be used as a starting point for a real sample. In most instrument manuals, there will
also be sections covering selected real analyte/sample situations, and these give guidance about the
type of changes to the temperature that are needed.
CLASS EXERCISE 3.4
Does the nature of the analyte/matrix have any effect on the steps in the temperature program?
Analyte
Drying time
Drying temperature
Matrix
No
Yes
Ashing time
Ashing temperature
Atomisation time
Atomisation temperature
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3.4
3. Non-flame AAS
These notes are intended only give some general guidelines about the effect on the temperature
program of the analyte, but more importantly, the sample matrix. When setting up a method for a
new sample, it is very much a matter of trial and error until the most efficient sequence of times and
temperatures is determined.
Drying stage
This step is designed to remove all volatiles without losing analyte by over-vigorous boiling, which
could cause the mixture to splatter.
CLASS EXERCISE 3.5
List some factors that would influence the temperature required for drying.
List some factors that would influence the time required for drying.
Usually the drying stage is divided into a number of steps:

increase from room temperature to boiling point of solvent

remain at this temperature for a time

increase up to about 50ºC above b.p. to remove last volatiles
The main problem likely in the drying stage is incomplete drying which can lead to splattering of the
remaining material as the temperature increases to ashing point. You will hear the fizz as the
temperature goes up.
CLASS EXERCISE 3.6
Describe the differences in the drying stage, compared to the standard program for lead in water,
where the matrix is:
(a) petrol
(b)
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milk
3.5
3. Non-flame AAS
Ashing stage
This step is designed to remove the non-volatile organic components of the matrix by burning them
away as in a muffle furnace.
CLASS EXERCISE 3.7
List some factors that would influence the temperature required for ashing.
List some factors that would influence the time required for ashing.
The ashing stage will normally be divided up into at least three steps:

increase from the final drying temperature to the ashing temperature (normally 5-10 seconds)

remain at ashing temperature for required time

in the last few seconds before atomisation, the argon gas flow is turned off so that the
atmosphere inside the tube is still for atomisation
Chemical modifiers are used where the analyte is relatively volatile and would be lost during the
ashing stage, eg lead which is volatile above 500ºC. If a modifier wasn’t used, then the ashing
temperature would have to be quite low, meaning a much longer time would be required. The
modifier for lead is phosphate, since lead phosphate has a much higher boiling point than other lead
salts, such as chloride and nitrate. However, the modifier must not hold onto the analyte during
atomisation!
Some of the volatile elements are lead, cadmium, mercury (of course), antimony and arsenic.
The instrument documentation will indicate whether a modifier is needed.
The main problem encountered with incomplete ashing is the generation of smoke during
atomisation, which causes a high background reading due to scattering. You can observe this at the
start of the atomisation step as a puff of smoke coming out through the delivery hole.
CLASS EXERCISE 3.8
Describe the differences in the ashing stage, compared to the standard program for copper in water,
where the analyte/matrix is:
(a) lead in water
(b)
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aluminium in milk
3.6
3. Non-flame AAS
Atomisation stage
This step is designed to drive the analyte atoms into an atomic vapour in the path of the HCL beam.
CLASS EXERCISE 3.9
List some factors that would influence the time required for atomisation.
List some factors that would influence the temperature required for atomisation.
The argon gas is turned on after the brief atomisation step to flush the tube and avoid it turning into
CO2.
3.5 Signal measurement
This is much more complex than in flame AAS, because the analyte atoms are only present for a few
seconds once per run, rather than being present whenever the nebuliser is in the flask. Therefore,
the signal is a short-lived peak (like in a GC or HPLC), as in Figure 3.6.
This means that signal capture by the computer is essential. The alternative is to wait through
two minutes of drying and ashing and then make sure you are watching when the atomisation step
occurs. To make matters more difficult, the steam and ash generated in the first two stages cause
scattering of the beam, and an apparent absorbance which varies over time, and has nothing to do
with the real analyte absorbance. In the software, this is avoided by not having the detector
responding until the atomisation steps.
Absorbance
EAAS
FAAS
Time
FIGURE 3.6 Signals in electrothermal and flame AAS
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3.7
3. Non-flame AAS
3.6 Practical aspects
Checking instrument performance
In flame AAS, the sensitivity actually had a numerical value: it was the concentration that would give
an absorbance of 0.0044. In electrothermal AAAS, where a discrete volume of sample is analysed,
concentration is only one of the factors that affects absorbance: the other is the volume. Therefore,
the mass of analyte (concentration x volume) is the variable that determines the absorbance.
The characteristic mass is the mass that produces the absorbance of 0.0044. This can be used
as shown below to determine the expected absorbance of a solution, and also the working
performance of the instrument.
A exp 
0.0044ms
mc
Eqn 3.1
where mc is the characteristic mass, most often in pg, ms the mass of analyte in the sample/standard
(in the same units) and As the expected absorbance of the standard.
Calculating masses in pg (10-12 g) from uL and ug/L might seem a good candidate for getting
the decimal place wrong, but in fact, nothing could be easier, as shown in Equation 3.2: no unit
conversion required at all!
pg  uL x ug / L
Eqn 3.2
CLASS EXERCISE 3.10
(a) Calculate the expected absorbance for 10 uL of 20 ug/L Fe, if the characteristic mass for Fe is
1.2 pg.
(b)
Comment on the instrument performance, if a 10 uL aliquot of 50 ug/L Pb gives an absorbance
of 0.24, given the characteristic mass is 5.5 pg.
Memory effects
If a metallic element is present in the analysed aliquot at concentrations above 250 ug/L, the
atomisation stage will not be sufficient to atomise and then remove all of it. This will occur whether
the species is the analyte or in the matrix. If it is the analyte, then some proportion will remain in the
tube when the next run commences. This will produce a higher absorbance than it should, and is
known as a memory effect.
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3.8
3. Non-flame AAS
The solution to this problem is a number of tube cleans, which take the temperature up to above
2000C very rapidly. Depending on the element and the concentration, it may require three or four
tube cleans before continuing.
CLASS EXERCISE 3.11
How could you check whether the tube is ready to be used after a number of tube cleans?
Solutions
A typical analysis run may use at least three different solutions: sample, added standard, blank,
modifier. The total volume of liquid should be between 15 and 30 uL, and should be consistent for all
solutions.
Tube performance
The graphite tube has a finite lifetime of around 500 normal temperature cycles. This will be reduced
by various factors.
CLASS EXERCISE 3.12
To be done in your own time. Download the Varian publication Factors affecting the analytical
performance and lifetime of graphite tube atomizers from the AIT webpage. Read it and answer the
following questions. They are examinable.
(a)
(b)
(c)
(d)
(e)
(f)
What should be checked when changing tubes?
What should be done after installing the new tube?
What problems are encountered with solutions of different surface tension? Of high viscosity?
What circumstances would require the use of air in the ashing stage? What limitations should
be made on its use?
Why is argon preferred to nitrogen?
Why is a consistent liquid volume recommended?
Background correction
Background correction is a means of correcting for sources of non-analyte reduction in beam
intensity. These sources include:

smoke particles

molecular species
These are more significant (especially the smoke) in EAAS than FAAS. When it occurs, the measured
absorbance is the sum of analyte and background absorbance (as in Eqn 3.3). An ideal system would
measure the total absorbance, and also the background by itself.
At = Aa + Ab
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Eqn 3.3
3.9
3. Non-flame AAS
There are two commonly employed systems for background correction:

deuterium lamp – by aid of a chopper, the signal reaching the detector alternates between the
HCL, which measures At, and a deuterium lamp (like in a UV-VIS) which only detects Ab; this is a
simple system, but requires that the HCL and D2 beams are exactly aligned through the tube,
which is difficult

Zeeman effect – a magnetic effect which changes the characteristics of the analyte absorption
line, meaning that when the magnet is on, the analyte no longer absorbs the HCL; this only
requires the HCL lamp, but is much more expensive
CLASS EXERCISE 3.13
Why would the analyte absorption not be detected in the deuterium beam?
3.7 Vapour generation
The vapour generation technique relies on the formation of a volatile species containing the analyte
metal. This may be the pure metal itself, as in the case of mercury (cold vapour), or the readily
decomposed hydride of metals, such as arsenic, antimony and tin (hydride generation).
The volatile form of the metal is created by chemical reaction with the sample in a reaction
vessel which has a stream of gas (eg nitrogen) passing through it. The volatile species is carried along
in the gas into the measurement part of the AAS where a quartz tube is seated (where the burner
would be in a flame AAS). It has optical windows at either end to allow the beam from the hollow
cathode lamp to pass through and out to the detector (see Figure 3.7). The analyte absorbs the
radiation in the normal way.
heated quartz tube
Hollow
Cathode
Lamp
Monochromator
& Detector
Gas stream in
Gas stream out
(with analyte)
Reaction vessel
(sample + reagents)
FIGURE 3.7 Schematic diagram of vapour generation apparatus
In the case of mercury, the analyte is already in the chemical form required – Hg(g) – after chemical
treatment in the reaction vessel by reduction of ionic mercury by tin chloride, so no heating of the
quartz tube is required, hence the name “cold vapour”.
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3.10
3. Non-flame AAS
In the hydride generation method, the analyte is reduced, usually by sodium borohydride (NaBH4),
producing the molecular hydride. When this is flushed out in the gas stream, it is not in the atomic
form, and requiring heating in the tube to decompose the hydride into the atomic metal. This
procedure is applicable to As, Se, Te, Sb, Bi and Sn. The tube may be heated electrically or with a
flame.
What You Need To Be Able To Do













define important terminology
explain reasons for limitations of flame AAS sensitivity
outline the changes in NFAAS that allow improved sensitivity
draw a schematic diagram of the workhead of an EAAS instrument and explain the function of
the components, including the various services (eg water)
describe a typical temperature program
outline factors affecting the design of the temperature program
outline how the nature of the analyte and matrix affects the temperature cycle
explain the purpose of a modifier, and give an example of an analysis when it may be necessary
explain the purpose of background correction, and give an example of an analysis when it may
be necessary
describe the principle of one type of background correction
perform calculations related to characteristic mass
list optimisation steps for routine EAAS analysis
explain how vapour generation AAS works
Revision Questions
1.
(a) Give THREE factors that limit the sensitivity of flame AAS.
(b) Explain how electrothermal AAS overcomes these.
2.
What services other than electricity are required for EAAS? What function(s) do they serve?
3.
Why is an autosampler an essential part of an EAAS instrument?
4.
Why is the gas switched off at the end of the ashing stage?
5.
What changes would you make to the basic “analyte in water” temperature program for the
following analyses:
(a) aluminium in milk
(b) lead in petrol
6.
Why is a phosphate modifier recommended for lead analyses? How does it work?
7.
The characteristic mass for aluminium is 6.1 pg. What is the expected absorbance for a 5 uL
aliquot of 50 ug/L standard?
8.
What type of samples would most require background correction?
9.
Explain the following for measurements of lead in a wine sample. Using the standard
temperature program for lead in water, the sample gives an absorbance of 0.6
(a) When the ashing time is increased from 5 to 60 seconds, the absorbance decreases to 0.3.
(b) When the drying time is reduced from 60 seconds to 5, the absorbance decreases to 0.1.
(c) When the modifier is not used, there is no absorbance.
10. Why is tin chloride required in mercury analysis by vapour generation AAS?
11. Why is heat needed in the vapour generation analysis of arsenic?
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3.11
3. Non-flame AAS
Answers to revision questions
Where the answer can be found directly in your notes, only a reference to it will be provided.
1.
(a) inefficiency of nebuliser, reaction in the flame, formation of oxides, background colour of
flame
(b) Table 3.1
2.
p3.3
3.
p3.4
4.
top of p3.7
5.
(a) increase drying & ashing times (b) decrease drying temp, small increase in ashing time
6.
p3.7
7.
mass in aliquot is 50 x 5 pg. Abs = 0.0044 x 250 ÷ 6.1 = 0.18
8.
High organic content, high levels of elements giving molecular species eg Al, Ca
9.
(a) significant amounts of smoke in the standard conditions run causing higher absorbance,
this is removed during the longer ashing
(b) insufficient drying time means that when the temperature goes up to the ashing point, the
remaining moisture is rapidly lost, causing the sample to fizz and spray out of the tube.
(c) the lead becomes volatile during the ashing stage and is lost before measurement
10.
For this analysis, mercury must be in the elemental form (Hg0) so the tin chloride reduces the
+1 and +2 ionic forms.
11.
The volatile form of arsenic is AsH3, which must be decomposed (by heat) to form As to absorb
the HCL radiation.
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3.12