sampling, preparation and analysis of heavy metal in foods
TRANSCRIPT
Sampling, preparation and analysis of Heavy Metal in
Foods
Wee Siew MoiChemical Contaminants Expert
Nestlé Quality Assurance Center, Asia Oceania Africa
CII, 11th Food Safety & Quality Summit 6-7 December 2016
A. Heavy (Toxic) metals analysis
B. Types of Equipment (Décision criterions)
C. Good Laboratory Practice (GLP) for sample preparation
D. Sample preparation
- digestion techniques/preparation of test solutions
E. Détermination
F . Quality Control
G. Method Performance
2
Outline of lecture
Heavy metals (Toxic metals)
Metal are elements that are typically hard, opaque, shiny, and has good electrical and
thermal conductivity. Metals are generally malleable, as well as fusible.
About 91 of the 118 elements in the periodic table are metals (some elements appear in
both metallic and non-metallic forms).
In food/feed, metals are usually
found in ionic or covalent form (not
metallic).
The species (oxididation degree,
inorganic versus organic) may
impact the toxicity, e.g.
Cr (III) versus Cr (VI)
As inorganic versus As organic
Toxic metals - food point of view
Metals can be
Essential macronutrients (Ca, Mg, K, Na)
Essential micronutrients (Cr, Co, Cu, Ni, Se)
Toxic metals
But some essential metals may become toxic!
«Heavy metals» is often used to describe toxic metals.
Strictly speaking it should be used only for element > 200Da.
In practice it refers to metals with high gravity and which have high attraction for
biological tissues.
5
Heavy Metal Analysis
Heavy
Metals
Analysis
Sampling and Sample
Preparation
Determination
Calibration
Digestion
Preparation of
Test solutions
• Avoid contamination when
preparing a test portion
• Obtain homogenized
representative test sample
• Use the appropriate digestion
program according to the type and
amount of sample.
• Avoid contamination of acids and dust
• Carry out blank test in parallel by the
same procedure
• Check if digestion is complete
• Prepare the test solution with similar
HNO3, HCl, Au and IPA concentrations as
those of calibrants depending on ICP-MS
technologies.
• ICP-MS instrument set-up
• Check ICP-MS performance
Critical Points
A. Heavy (Toxic) metals analysis
B. Types of Equipment (Décision criterions)
C. Good Laboratory Practice (GLP) for sample preparation
D. Sample preparation
- digestion techniques/preparation of test solutions
E. Détermination
F . Quality Control
G. Method Performance
6
Outline of lecture
B. Types of Equipment (Atomic Spectroscopy)
7
Flame AASGraphite
Furnace AAS
AAS instruments can be flame only, furnace only, or combined (switchable)
ICP-OES ICP-MS
Photos taken from Agilent Technologies as illustration purposes
ICP-MS versus Atomic Absorption Spectroscopy(AAS)
For higher sample throughput
10
Figure 2. ICP-MS detection limit ranges and orders of magnitude of
signal intensity compared to other atomic spectroscopic techniques
Figure 2 shows that ICP-MS is selected as the technique) than Flame
(FAA)
or Graphite Furnace (GFAA), and (Hydride Generation) Inductively
Coupled Plasma–Optical Emission Spectroscopy ((HG)-ICP-OES).
- Allow samples with varying analyte concentrations to be
analyzed together
due to its wide analytical working range (9 orders of magnitude):
Combination of wide analytical working range and excellent
sensitivity:
Provides short ICP-MS analysis times.
Can reduce sample-handling requirements.
Minimizes potential analytical errors.
Avoids to frequently recalibrate the system of choice to:
- Get better instrumental detection limits in solution (ng/L
Major limitations to use ICP-MS equipment is the high initial
investment and cost of consumable supplies / Gases / Power
compared to other atomic spectroscopic techniques
What is ICP-MS?
11
ICP-MS stands for Inductively Coupled Plasma Mass Spectrometry.
ICP-MS Instrument comprises five basic analytical parts as shown in Figure 1:
Sample introduction generating an aerosol of the liquid (or solid) sample
Plasma source ionizing the aerosol
Sampling interface extracting ions from ICP
Ion optics and mass spectrometer focusing and separating ions
Ion detector converting ions into an electronic signal processed by
the data handling system
Figure 1- Analytical parts of ICP-MS instrumentation
Determination
A. Heavy (Toxic) metals analysis
B. Types of Equipment (Décision criterions)
C. Good Laboratory Practice (GLP) for sample preparation
D. Sample preparation
- digestion techniques/preparation of test solutions
E. Détermination
F . Quality Control
G. Method Performance
12
Outline of lecture
C. GLP for sample preparation
13
Food and beverage samples should be stored in their typical commercial
storage conditions (either frozen, refrigerated, or at room temperature)
until analysis. Samples should be analyzed within 6 months of
preparation.
If food or beverage samples are subsampled from their original storage
containers, ensure that containers are free from contamination for the
elements of concern.
Well-homogenized samples and small reproducible aliquots help
minimize interferences. Particle size and distribution should be
normalized using blenders, grinders and mixers.
Sampling and Sample
Preparation
C. GLP for sample preparation
14
• Management of analytical blanks (Cleanliness of laboratory environment)During sample preparations for trace element analysis by ICP-MS, the main requirement
is the management of analytical blanks.
Analytical blank s the measure of all external sources of elemental contamination and is used
to make a correction to the measured sample correction.
Contamination for trace analysis can occur from the:
• Laboratory environment (clean air facilities, air filters and flow, hoods and bench tops)
• Materials (plastic ware for storage and handling)
• Reagents (standard solutions, water, acids, hydrogen peroxide, TMAH as alkaline buffer,
enzymes, isopropanol as carbon buffer)
• Apparatus (auto-sampler, plastic ware, nebulizer, spray chamber and torch, peristaltic,
transfer and drain tubings, sampler and skimmer cones and lens)
• Analyst (bare hand, cosmetics, hair, wearing jewels and watches)
Sampling and Sample
Preparation
A. Heavy (Toxic) metals analysis
B. Types of Equipment (Décision criterions)
C. Good Laboratory Practice (GLP) for sample preparation
D. Sample preparation
- digestion techniques/preparation of test solutions
E. Détermination
F . Quality Control
G. Method Performance
15
Outline of lecture
D. Digestion techniques
16
For accurate decomposition (destruction of organic matter) of food sample.
Digestion
Why decomposition is required?
• Conversion from solids into liquids
• Destruction of matrix
• Separation of interfering substances
• Isoformation of sample and standard
• Homogenization
• Preconcentration of analytes
Pros:
Leads to a representative sample
Reduces problems in the
measurement step
Easy to standardize
Cons:
Labor intensive cost factor
Bottle neck in analytical process
Risk of contamination or losses of
analyte
Microwave
Digestion
High
Pressure
Asher
D. Digestion techniques
17
Decomposition may be safe
1. Analytically accurate
2. Economically efficient
3. Safe and easy to perform
Digestion
Analytical accurate
No contamination
No loss of elements
Complete decomposition
Reliable equipment
Reproducible
analytical results
Economically efficient
Low consumption of
chemicals
Ease of handling
Low investment/operating
costs
Automation
Reduce cost of faulty
analytical results
Safety
Low amounts of
hazardous chemicals
Simple handling
Spontaneous reactions
Reduce operator error
Instrument safety
(design/Manufacture)
Safety First
18
Preparation of sample solution
15
.09
.201
3
Reagent Blank The blank test must be carried out in parallel with the
determination by the same procedure but omitting the test
portion
Preparation of digested
samples
Check if the digestion of test portion is complete. Prepare the
test solution by diluting the digested solution with ultrapure
water to a known volume.
After diluting to volume, the test solution should be clear and
colorless to slightly yellow.
Turbidity and/or a deep color usually indicate an incomplete
digestion
Reagents High-purity reagents should always be used. Each reagent
lot should be tested and certified to be low in the elements of
interest before use.
Standard (Stock/Internal) Elements must be compatible and stable in solutions
together. Concentrations need to be verified before use.
For analysis of As, Cd, Pb, and Hg in food matrices, internal
standard solution of rhodium (Rh), indium (In), and thulium
(Tm) is recommended
Calibration standard Fresh calibration standards should be prepared every day, or
as needed.
Preparation
of Test
solutions
A. Heavy (Toxic) metals analysis
B. Types of Equipment (Décision criterions)
C. Good Laboratory Practice (GLP) for sample preparation
D. Sample preparation
- digestion techniques/preparation of test solutions
E. Détermination
F . Quality Control
G. Method Performance
19
Outline of lecture
20
ICP-MS Instrument Setup/Optimization
15
.09
.201
3
Determination
(1) Instrument startupTune ICP-MS instrument using suitable optimizing / tuning solutions and
according to manufacturer’s instructions
Note : Tuning of the ICP-MS instrument (i.e. optimization for best performance)
usually while aspirating a prescribed element solution called tuning solution is
started using default or recommended settings by manufacturer for plasma power,
sampling depth, gas flow rates and sample introduction rate (plasma gas flow rate
is set at a fixed value and not optimized at all). Ensure that ICPMS passes all
check criteria of tuning after stabilization of plasma during 30 min.
(2) OptimizationCheck mass resolution, mass calibration, sensitivity and stability of the system
- Adjust ICP-MS instrument daily with an optimizing solution to achieve
maximum ion signals and both low oxide rates (e.g. < 2 %) and low rates of
doubly charged ions (e.g. < 2 %).
Note: The optimizing solution should contain elements that cover the whole mass
range giving a high rate of oxides and doubly charged ions. The solutions
recommended by the manufacturer of the ICP-MS instrument may be used.
21
Calibration1
5.0
9.2
01
3
A set of at least seven different calibration solutions including blank solution
(i.e. zero member compensation) with evenly spaced concentrations must be
used for external standard calibration so that the concentration range should
be chosen with respect to the concentrations expected in samples and with
respect to the linear dynamic range. It is important that the concentration of
acids and IPA in the calibration solutions and in the sample solutions are the
same.
Calibration
Prepare calibration solutions with similar HNO3 ,IPA, HCl (or Au) concentrations
as those of sample digests depending on ICP-MS / digestion system.
R2 must be > 0.995 using recommended weighted least squares
regression
A. Heavy (Toxic) metals analysis
B. Types of Equipment (Décision criterions)
C. Good Laboratory Practice (GLP) for sample preparation
D. Sample preparation
- digestion techniques/preparation of test solutions
E. Détermination
F . Quality Control
G. Method Performance
22
Outline of lecture
23
Quality Control
15
.09
.201
3
• The correlation coefficients of the weighted-linear calibration curves
for each element must be ≥0.995 to proceed with sample analysis.
• The percent recovery of the ICV standard should be 90-110% for each
element being determined.
• Perform instrument rinses after any samples suspected to be high in
metals, and before any method blanks, to ensure baseline sensitivity
has been achieved.
• Run rinses between all samples in the batch to ensure a consistent
sampling method.
• Each analytical or digestion batch must have at least three
preparation (or method) blanks associated with it if method blank
correction is to be performed. The blanks are treated the same as the
samples and must go through all of the preparative steps. If method
blank correction is being used, all of the samples in the batch should
be corrected using the mean concentration of these blanks.
QC samples (certified, P-test, in-house reference samples or spiked
samples) must be regularly included and analysed in duplicate.
Quality
Control
24
Summary of quality control samples(Ref: AOAC Official Method 2015.01 Heavy Metals in Food)
15
.09
.201
3
Quality
Control
25
Summary of quality control samples(Ref: AOAC Official Method 2015.01 Heavy Metals in Food)
15
.09
.201
3
Quality
Control
A. Heavy (Toxic) metals analysis
B. Types of Equipment (Décision criterions)
C. Good Laboratory Practice (GLP) for sample preparation
D. Sample preparation
- digestion techniques/preparation of test solutions
E. Détermination
F . Quality Control
G. Method Performance
26
Outline of lecture
27
Method Performance (Ref: AOAC Official Method 2015.01 Heavy Metals in Food)
Performance
Characteristics
Definition
Linearity A coefficient of determination R2 ≥ 0.995 should be generally obtained for ten
standards using weighted linear regression
Limit of Detection (LoD) Limit of detection (LOD) and LOQ were determined through the analysis
method blanks. LOD was calculated as 3 times the SD of the results of the
blanks, and LOQ was calculated as 2 times the value of the LOD, except
where the resulting LOQ would be less than the lowest calibration point, in
which case LOQ was elevated and set at the lowest calibration point and LOD
was calculated as 1/3 of the LOQ. All LOQs achieved are ≤10 μg/kg for all
food matrices and ≤8 μg/kg for liquid matrices, such as infant formula. Limit of Quantification (LoQ)
Repeatability The absolute difference between two independent single test results obtained
using the same method on identical test material in the same laboratory by the
same operator using the same equipment within a short interval of time and
calculated as should not be greater than 20 % (25 % for values close to
PLOQ) which corresponds to the repeatability limit, r, at 95 % confidence
level.
Intermediate reproducibility The absolute difference between two independent single test results obtained
using the same method, on identical test material by different operators using
different equipments at different days for intermediate reproducibility test and
calculated as should not be greater than 35 % (40 % for values close to
PLOQ) which corresponds to the reproducibility limit, iR, at 95 % confidence
level.
28
Method Performance (Ref: AOAC Official Method 2015.01 Heavy Metals in Food)
Sample-specific LOQs for several matrices, based on LOQs determined by
the default method, and adjusted for changes in sample mass for particular
samples, are shown. Values have been rounded up to the nearest
part-per-billion.
Sample-specific LOQs
Sample LOQ, µg/kg As Cd Pb Hg
Infant Formula 2 1 4 3
Chocolate 4 2 8 6
Rice Flour 4 2 8 6
Fruit juice 1 1 2 2
29
In summary
Heavy
Metals
Analysis
Sampling and Sample
Preparation
Determination
Calibration
Digestion
Preparation of
Test solutions
• Avoid contamination when
preparing a test portion
• Obtain homogenized
representative test sample
• Use the appropriate digestion
program according to the type and
amount of sample.
• Avoid contamination of acids and dust
• Carry out blank test in parallel by the
same procedure
• Check if digestion is complete
• Prepare the test solution with similar
HNO3, HCl, Au and IPA concentrations as
those of calibrants depending on ICP-MS
technologies.
• ICP-MS instrument set-up
• Check ICP-MS performance
Critical Points