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Experiment 6:
Determination of TraceLevels of Copper inVegetable Samples Using
the Atomic AbsorptionSpectrophotometerArgamino.Buenaseda.Gajigan
Chem 127.1 Maam Cruz
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INTRODUCTION
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AAS
one of the mostcommoninstrumental
methods foranalyzing formetals and somemetalloids
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PARTS
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Hollow Cathode Lamp
Provide the analytical light line for theelement of interest
Provide a constant yet intense beam ofthat analytical line
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Nebulizer
Suck up liquid sample at a controlledrate
Create a fine aerosol for introductioninto the flame
Mix the aerosol and fuel and oxidant
thoroughly for introduction into theflame
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Flame
Destroy any analyte ions and
breakdown complexes Create atoms (the elemental form) of
the element of interest
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Monochromator Isolate analytical lines' photons passing
through the flame
Remove scattered light of otherwavelengths from the flame
In doing this, only a narrow spectral line
impinges on the PMT.
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Photomultiplier Tube (PMT)
As the detector, the PMT determinesthe intensity of photons of the analytical
line exiting the monochromator.
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KANGKONG
Ipomoea aquatica
A smooth, widely spreading vine,
with the stems trailing on mud orfloating on water.
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KAMOTE Ipomoea batatas
A herbaceous vine with alternateheart-shaped or palmately lobedleaves and sympetalous flowers.
The edible root is long and taperedwith a smooth skin of varying colorsfrom red, purple, brown and white.
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Copper Content Standards
Kangkong
37.68 mg/kg (dry weight) (Wang, 2010)
3.34-3.95 mg/100g (Umar, 2007) Kamote
0 mg/100g (Antia, 2006)
Root: 0.34 mg/100g (www.whfoods.org)
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METHODOLOGY
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Preparation of StockSolutions
0.5 g of coppermetal weredissolved in 20 mL
of 1:1 nitric acid diluted to mark in
250-mL volumetricflasks
5 mL aliquot wereobtained from theprepared solution
diluted to a 100-mL
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Preparation of Standard
Five clean, 50-mLvolumetric flaskswere prepared and
labeled withnumbers 1 to 5.
0.50, 1.25, 2.50,5.00 mL of the
stock solution wereplaced to flasks 1,2, 3, and 4respectively
to re are 1.00,
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Preparation of Sample
The leaves fromthe vegetablesamples were
rinsed and dried inthe oven
The temperature ofthe oven was
maintained at 100to 150C for 45minutes.
Two (2) grams of
the dried sample
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Preparation of Sample
The mixture wasboiled slowly atlow setting for 20
minutes. 10 mL distilled
water was addedto the cooled set-
up.
It was thenfiltered to a 50-
mL volumetric
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Analysis of theVegetable Sample
The absorbanceof the standardsolutions and the
sample weremeasured usingthe requiredinstrumental
parameters (forCu) of the atomicabsorptionspectrophotomet
er.
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Standard AdditionMethod
Five clean 50-mLvolumetric flaskswere prepared and
labeled withnumbers 1 to 5.
To each flask,10 mLof the digested
sample were added.
0.00, 0.50, 1.25,2.50, 5.00 mL of the
stock copper
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Standard AdditionMethod
The absorbancesof the solutions
were measuredusing the requiredinstrumentalparameters for
Cu.
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RESULTS
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DIRECT CALIBRATION
StandardConcentration
(ppm)Absorbance
CorrectedAbsorbance
Blank 0.00 0.0123 0
1 1.00 0.1937 0.1814
2 2.50 0.3685 0.3562
3 5.00 0.6789 0.6666
4 10.00 1.1751 1.1628
0 2 4 6 8 10 12
0
2
4
6
8
10
12f(x) = 1.#NANxR = 1.#NAN
Concentration (ppm)
Corrected Absorbance
Kangkong 0.036 = 0.1089
[Cu] + 0.088
[Cu] = -0.4775ppm
Accounting for
dilution:
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STANDARD ADDITIONMETHOD
Kangkong
Set-upVolume ofStandard
(mL)
Absorbance
Trial 1 Trial 1 Trial 3
1 0.00 0.0119 0.0127 0.01332 0.50 0.1619 0.1632 0.16123 1.25 0.3350 0.3311 0.32824 2.50 0.6165 0.6147 0.61485 5.00 1.0855 1.0820 1.0820
Set-up
A/Ao
(Vx+Vs/Cs)
(A/Ao) (Vx +Vs/Cs)
Trial 1 Trial 1 Trial 3 Trial 1 Trial 1 Trial 3
1 0 0 0 0 0 0 02 14 13 12 0.105 1.4285 1.3493 1.27263 28 26 25 0.1125 3.1670 2.9330 2.77614 52 48 46 0.125 6.4758 6.0502 5.77825 91 85 81 0.15 13.683 12.780 12.203
A/Ao= 0.1619/0.0119
=14
(Vx +Vs)/Cs=(10+0.50)/100= 0.105
(A/Ao )(Vx+Vs)/Cs= 14 X 0.105= 1.4285
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Kangkong
0 2 4 6 8 10 12
0
2
4
6
8
10
12f(x) = 1.#NANxR = 1.#NANf(x) = 1.#NANxR = 1.#NANf(x) = 1.#NANxR = 1.#NAN
LinearRegressionfor
LinearRegressionfor
LinearRegressionfor
Volume of Standard (mL)
(A/Ao) (Vx +Vs/Cs)
Cx = (1/m)Linear Equations:
y = 2.7261x + 0.0018Cx=0.3668y = 2.5459x + 0.0019Cx=0.3928y = 2.4333x + 0.0021Cx=0.4110
Average of Cx = 0.3902ppmAccounting for dilution:(0.3902mg/L)(50mL/10mL)
(50mL)(1L/1000mL)/2g= 0.0488 mg/g
STANDARD ADDITIONMETHOD
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Kamote
Set-upVolume ofSolution
(mL)
Absorbance
Trial 1 Trial 1 Trial 3
1 0.00 0.0272 0.0261 0.02762 0.50 0.1696 0.1679 0.17333 1.25 0.3308 0.3315 0.33174 2.50 0.6200 0.6191 0.62025 5.00 1.1073 1.1078 1.1088
Set-up
A/Ao(Vx
+Vs/Cs)
(A/Ao) (Vx +Vs/Cs)
Trial 1 Trial 1 Trial 3 Trial 1 Trial 1 Trial 31 0 0 0 0 0 0 0
2 6 6 60.105 0.6547 0.6755 0.6593
3 12 13 120.113 1.3682 1.4289 1.3520
4 23 24 22 0.125 2.8493 2.9650 2.8089
A/Ao= 0.1696/ 0.027=6
(Vx +Vs)/Cs= (10+0.50)/100= 0.105
(A/Ao )(Vx +Vs)/
= 6 X 0.105= 0.6547
STANDARD ADDITIONMETHOD
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Kamote
0 2 4 6 8 10 12
0
2
4
6
8
10
12f(x) = 1.#NANxR = 1.#NANf(x) = 1.#NANxR = 1.#NANf(x) = 1.#NANxR = 1.#NAN
LinearRegressionfor
LinearRegressionfor
LinearRegressionfor
Volume of Standard (Vs)
(A/Ao) (Vx +Vs/Cs)
Cx = (1/m)Linear Equations:
y = 1.2152x + 0.0524Cx=0.8229y = 1.2673x + 0.0573Cx=0.7891y = 1.1979x + 0.0469Cx=0.8348
Average of Cx = 0.8156 ppm
Accounting for dilution:(0.8156mg/L)(50mL/10mL)(50mL)
(1L/1000mL)/2g= 0.1019 mg/g
STANDARD ADDITIONMETHOD
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discussion
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Atomic AbsorptionSpectrophotometry
AAS Simple, low cost
Application: effective determination of
trace amounts of heavy metals inbiological samples
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Atomic AbsorptionSpectrophotometry
Cathodelamp
made ofmetal underinvestigation
emit the line
spectrumcharacteristic of theparticularelement
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Sample Preparation
Air dried
Advantage: ease of homogenization
[Cu] reported per dry mass Rinsed
remove surface contaminants
Oven dried Further drying
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Sample Preparation
Solid samples should be brought intosolution
acid attack
alkali fusion and subsequent acid attack
dissolution in solvent
Acid attack important not to add components that
interfere with the copper determination
nitric acid causes no interference in the
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Direct Calibration Method
Plot the measured absorbanceagainst the concentration of thestandard solution (ppm)
copper atoms in the flame
follow the Beers Lambert Equation
A = bc
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Direct Calibration Method
Blank determination exhibits little absorbance
distilled water used as blank contains
traces of copper ions
Standard Solution
copper metal
high purity: minimize chemicalinterference effects due to chemicalimpurities present in some inorganic
salts available in the laboratory
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Standard Addition Method
Absorbance of several solutionscontaining a constant volume ofdigested kangkong and kamote
sample and varying volume ofstandard copper solution wasmeasure
Derivation:
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Standard Addition Method
Standard Addition
accounts for the matrix effects
used when sample matrix affects theresponse of the analyte.
usually necessary for complicatedsamples such as leaves
some components in the leaves canaffect the response of the copper
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Interference
Spectral Interference
presence of very few atoms
Interference from flame emissions whichcan be reduced by modulating lamp
Interference from molecular speciesabsorbing lamp photons that occurs at
shorter wavelengths which can beremoved by using a deuterium lampwhich has a broader band light source.
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InterferenceChemical Interference Solution
Matrix effect standard addition method
Poor nebulization because ofhigh viscosity
Use hotter flames
Poor volatility Use hotter flames
Formation of metal oxides andhydroxides
Using fuel rich flame
Ionization of analyte atoms Add an ion that has a lowerionization potential
Light scattering by particles inthe flame
Background correction ORSelecting the optimum temperaturewhere the background signal andanalyte can be effectivelyseparated ORUsing broad band light source (i.e.
deuterium)
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Interference
Other methods in overcoming someinterference
using higher temperature flame such as
nitrous oxide which sufficiently causecomplete dissociation of some refractorycompounds
extracting the analyte element extracting the interferent
use of releasing agents that will
combine with the interferents.
I t f i C
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Interference in Cudetermination
High Zn/Cu ratio which lowers theobserve absorption.
Some papers suggest the use of nitrous
oxide can lessen the interference.
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Possible Sources of Errors
Low concentration of standardssolutions
solns deteriorate due to the absorption
of the analyte in the walls of thecontainer
Oxide coating of the metal standard
(Cu metal) was not properly removed Differences in the amount of sample
and standard that reach the flame
because of the differences in the
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CONCLUSION
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CONCLUSION
Trace amounts of severalnutritionally important elements invegetable samples could be
successfully determined throughAtomic AbsorptionSpectrophotometry.
The analyte concentration wasdetermined from the absorption ofits solution using the Beer-Lambert
equation.
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CONCLUSION
In the experiment, two methods wereused to determine the copperconcentration in leaf samples: direct
calibration method and the standardaddition method.
In direct calibration method, the
equation y = 0.1089x + 0.088 wasobtained as standard curve. Thecalculated concentrations of Copperwere -0.0112 mg/g and 4.76E-3 mg/g for