chemical metrology – some basics
TRANSCRIPT
Chemical Metrology – Some Basics
Metrology and Physical Constants International School of Physics “Enrico Fermi”
Stephen A. Wise Analytical Chemistry Division
Material Measurement Laboratory National Institute of Standards and Technology (NIST)
Gaithersburg, Maryland USA
Chemical Metrology for Food, Nutrition, Environmental, and Human Health Assessment - Outline Basics of Chemical Metrology
Organic Chemical Metrology Inorganic Chemical Metrology Quantification Approaches Isotope Dilution Approach for Quantification
Chemical Metrology and CRM Development Chemical Metrology in Food and Nutrition
Assessment – examples Use of CRMs
Chemical Metrology in Environmental and Human Health Assessment - examples
Chemical Metrology – Some Basics
Chemical Identification What are we measuring? (Qualitative Analysis)
Amount of Substance – mole How much is there? (Quantitative Analysis) Generally reported as mass/mass, e.g., mg/kg
Purity of reference standards Measure trace level impurities and subtract from 100% Major focus at BIPM activities in organic metrology
Inorganic Chemical Metrology
Direct analysis of solid samples Acid digestion and analysis of resulting solution for
total element content (species lost) Chemical separation/isolation/enrichment Analysis/Detection
Inductively coupled plasma/Mass spectrometry (ICP-MS)
X-ray fluorescence Atomic Absorption Spectrometry (AAS) Neutron activation analysis (NAA) Voltammetry Others
Quantification Internal and external standard methods Isotope dilution approach
Examples: Total element content of Pb, Cu, Zn, etc.
Organic Chemical Metrology
For solid samples, solvent extraction to remove constituents of interest from the matrix
Isolation of constituents of interest/extract clean up Chromatographic separation
Gas Chromatography (GC) Liquid Chromatography (LC)
Analysis/Detection Mass spectrometry (MS) Tandem MS (MS/MS) UV-Visible and fluorescence spectrometry Electron capture detection (ECD)
Quantification Internal and external standards Isotope dilution approach (different from inorganic)
Examples: pesticides, vitamins, cholesterol, etc.
Approaches to Quantification
External Standard Calibration Approach Calibration is based on direct comparison with instrumental response (via calibration curves
or response factors)
Advantage: simple
Disadvantage: skill required to minimize biases from sample processing
Internal Standard Calibration Approach
One or more compounds are added to both calibrants and unknowns
Calibration is based on the ratio of responses for analytes and internal standards
Advantage: losses from transfers, dilutions, etc. are compensated; may compensate changes in instrumental response; less skill is required
Disadvantage: calculations more complex; internal standards must be identified and used
Standard Addition Approach
A known quantity of the analyte is added to the unknown sample; the response is compared with the unspiked sample
Advantage: quantitation is based on the sample matrix; similar advantages to using calibrants prepared in a “blank” sample matrix,
Disadvantage: calculations are complex, sample processing is labor intensive; separate calibrations needed for each sample;
External Standard Approach - need to know extract volumes - quantitative transfers required for all steps
(Area)
[ ]AC(concentration)
RAX
[ ]AX
[ ]RFR
AA
C
C=
[ ]A R RFX AX=
RF = Response factor
[ ]( )= R R AA A CX C
[ ]= ⋅RR
AAC
X
C
AXmass = [ ]AX volume (or mass) of extract
level A in sample = mass AX
mass sample extracted
RACResponse for analyte in calibrant=
[ ]AC= Concentration of analyte in calibrant
[ ]AX= Concentration of analyte in unknown
RAXResponse for analyte in unknown=
A
RAC
Internal Standard Approach - IS is added at earliest opportunity
- knowledge of volumes is not required - quantitative transfers are not required
(area ratio)
(concentration ratio or mass ratio)
RF = Response factor
level A in sample = mass AX
mass sample extracted
RACResponse for analyte in calibrant=
[ ]AC= Concentration of analyte in calibrant
[ ]AX= Concentration of analyte in unknown
RAXResponse for analyte in unknown=
RR
A
IS
C
C
[ ][ ]
A
ISC
C
RR
A
IS
X
X
[ ][ ]
A
ISX
X
[ ][ ]RF
RR
A
IS
RR
mass Amass IS
A
IS
C
C
A
IS
C
C
C
C
C
C
= =
[ ][ ]= ⋅ = ⋅
RR
IS
A
RR
mass ISmass A
A
IS
C
C
A
IS
C
C
C
C
C
C
mass Amass IS
C
C
or
Note: are concentrations in a calibration solution, so:[ ]AC [ ]ISC&
[ ][ ]
A
ISmass Amass IS
C
C
C
C= (masses of each analyte (or internal standard)
added to the calibration solution)
mass AR mass IS
R RFmass A in sample extractedX
A X
ISX
X
X
=⋅
⋅=
mass Amass IS
RR
RFX
X
A
IS
X
X
=
[ ]ISC
RISC
[ ]ISX
RISX
=
=
==
Concentration of internal standard in calibrantConcentration of internal standard in unknown
Response for internal standard in calibrantResponse for internal standard in unknown
Isotope Dilution Concept – Inorganic
How many pennies in the bottle? 1. Add 100 green cents to the bottle 2. Mix the pennies thoroughly 3. Withdraw 100 cents 4. Count how many are green & tabulate results 5. Replace cents, mix and repeat experiment 10x
Results: On average there were 5.3 green cents out of 100 So, how many cents (N) are in the bottle?
5.3100
=100
N+ 100
Sample Population
{
Total Population
{
∴N = 1787
Principle of IDMS – inorganic
x
M
sample x
x
M
spike y
blend bx x
M
( )( ) ∑
∑⋅
−
−⋅⋅⋅=
ii
ii
R
R
RRRR
MM
mm
ww,y
,x
xbx
bxy
y
x
x
yxyx
For example: Pb isotopes are 204 (1.8%), 206 (22.1%), 207 (24.2%), and 208 (52.1%)
ORGANIC ISOTOPE DILUTION MASS SPECTROMETRY
Isotope Dilution Mass Spectrometry for Organic Analysis
Use an stable isotope labeled internal standard (e.g., 13C, 2H, 15N, 18O)
Add labeled spike, allow equilibration, then compare the ratio of the natural to labeled compound
Isotope labeled internal standard added for each analyte to be measured
For organic analysis, isotope dilution is really just internal standard method with ideal internal standard
Internal Standards: Choice and Number
Stable isotope substitution preferable: 13C, 2H, 15N, 18O Typically for MS, m/z difference should be at least 3 Separation of analyte and IS is not necessary or even desirable
for MS (particularly for LC/MS)
If labeled reagent is not available, try to select a compound as similar as possible to the analyte (i.e., methyl analog or isomer)
Internal standards should be resolved from analytes and matrix components, either by chromatography and/or by selective detection.
For determination of multiple analytes, use several internal standards to cover ranges in chromatographic elution
Use of Isotopically Labeled Reagents in Quantitation Organic Analysis
• Isotope dilution mass spectrometry (IDMS) – A known quantity of a labeled form of the analyte is added to the sample – Quantitation is based on measurement of isotopic ratios
• Isotopically labeled internal standards – A labeled form of the analyte (13C or 2H) is added to the sample and to the calibrant – Quantitation is based on calibration curves – Internal standard often coelutes with analyte (preferred for LC/MS)
• May compensate for:
– Losses from sample processing – Fluctuations in instrumental response
• Does not compensate for: – Incomplete extraction – Unresolved interferences – Insufficient sensitivity
Certified Reference Materials (CRMs)
Reference Material (RM): Material, sufficiently homogeneous and stable with reference to specified properties, which has been established to be fit for its intended use in measurement or in examination of nominal properties.”
Certified Reference Material (CRM): Reference material, accompanied by documentation issued by an authoritative body and providing one or more specified property values with associated uncertainties and traceabilities, using valid procedures.
International Vocabulary of Basic and General Terms in Metrology, International Organization for Standardization (ISO), 2012 (VIM)
Standard Reference Materials (SRMs) are Certified Reference Materials (CRMs) issued by the National Institute of Standards and Technology (NIST)
Homogeneous, stable materials well-characterized for one or more
chemical and/or physical properties Assist laboratories worldwide in validating analytical measurements
of chemical composition
NIST Standards for Chemical Measurements
• High Purity Neat Chemicals • Organic Solution Standards • Inorganic Solution Standards • Gas Mixture Standards
Complex Matrix Standards • Advanced Materials • Biological Fluids/Tissues • Foods/Botanicals • Geologicals • Metals and Metal Alloys • Petroleum/Fossil Fuels • Sediments/Soils/Particulates • Cements
Chemical composition standards constitute over 2/3 of ~1,400 NIST SRM types and ~24,000 of over 33,000 NIST SRM Units sold in 2011
• Molecular Spectrometry Standards • Electrolytic Conductivity Standards • pH / Ion Activity Standards
Development of the Independent Analytical Methods Concept at NIST
One criterion for a standard sample is “Its composition should have been determined by independent and reliable methods affording agreeing results”
W.F. Hillebrand, J. Ind. Eng. Chem. 8, 466 (1916) Modes of establishing the accuracy of NBS SRMs defined as
“reference method, two independent methods, or interlaboratory comparison”
H.T. Yolken, NBS Spec. Publ. 408 (1973) “At NBS, three modes of measurement are used to assure that the
values of the SRM property(ies) are accurate. These are (a) definitive methods, (b) reference methods, (c) two or more independent and reliable methods.”
J.P. Cali and W.P. Reed, NBS Spec. Publ. 422 (1974) Adapted from M.S. Epstein, Spectrochemica Acta, 468, 1583 (1991)
SRM 1571 Orchard Leaves
First NIST natural matrix environmental SRM for trace element content
1971
Analytical Methods used for SRM 1571 Orchard Leaves
First NIST natural matrix environmental SRM for trace element content
1971
First SRM for Organic Contaminants – SRM 1580 Shale Oil
1980
Analytical Approach for Organic Contaminants in Complex Mixtures
Use Multiple Analytical Methods
exploit differences in
Extraction Isolation and Cleanup
Separation and Detection
minimizes possibility of undetected bias in resulting certified concentrations
that
Independent Analytical Methods Approach for Certification of Organic Contaminants
Use of Independent Methods for Value Assigning SRMs for Organic Constituents
Use of Independent Analytical Methods to Exploit Differences in
Extraction
Parameters
Soxhlet Ultrasonic Pressurized Fluid Supercritical Fluid Microwave-assisted Mechanically agitated
Solvent Temperature Pressure pH
Techniques
Cleanup Isolation
Enrichment
Liquid-Liquid Extraction Column Chromatography Liquid Chromatography Solid Phase Extraction (SPE) Solid Phase Microextraction (SPME)
Off-Line Approaches
Separation
Gas Chromatography (GC) Liquid Chromatography (LC) Ion Chromatography Electrophoresis Multidimensional Separation
Instrumental Approaches
Detection
MS MS/MS FID ECD Flame photometric AED
MS MS/MS Absorbance Fluorescence Electrochemical ELSD CAD
GC LC
Quantification
External Standard Internal Standard Isotope Dilution Standard Addition
Linear Regression Slope/Intercept Zero Intercept Bracketed Calibration Exact Matching Nonlinear Calibration
Model Calibration
Minimize the possibility of undetected bias in resulting certified concentrations
Use of Independent Methods for Value Assigning SRMs for Organic Constituents
Use of Independent Analytical Methods to Exploit Differences in
Extraction
Parameters
Soxhlet Ultrasonic Pressurized Fluid Supercritical Fluid Microwave-assisted Mechanically agitated
Solvent Temperature Pressure pH
Techniques
Cleanup Isolation
Enrichment
Liquid-Liquid Extraction Column Chromatography Liquid Chromatography Solid Phase Extraction (SPE) Solid Phase Microextraction (SPME)
Off-Line Approaches
Separation
Gas Chromatography (GC) Liquid Chromatography (LC) Ion Chromatography Electrophoresis Multidimensional Separation
Instrumental Approaches
Detection
MS MS/MS FID ECD Flame photometric AED
MS MS/MS Absorbance Fluorescence Electrochemical ELSD CAD
GC LC
Quantification
External Standard Internal Standard Isotope Dilution Standard Addition
Linear Regression Slope/Intercept Zero Intercept Bracketed Calibration Exact Matching Nonlinear Calibration
Model Calibration
Minimize the possibility of undetected bias in resulting certified concentrations
Isotope dilution is the preferred approach
Modes Used at NIST for Value-Assignment of Reference Materials for Chemical Composition
Cer
tifie
d Va
lue
Ref
eren
ce V
alue
In
form
atio
n Va
lue
1. Certification at NIST Using a Primary Method with Confirmation by Other Method(s)
2. Certification at NIST Using Two Independent Critically-Evaluated Methods
3. Certification/Value-Assignment Using One Method at NIST and Different Methods by Outside Collaborating Laboratories
4. Value-Assignment Based On Measurements by Two or More Laboratories Using Different Methods in Collaboration with NIST
5. Value-Assignment Based on a Method-Specific Protocol
6. Value-Assignment Based on NIST Measurements Using a Single Method or Measurements by an Outside Collaborating Laboratory Using a Single Method
7. Value-Assignment Based on Selected Data from Interlaboratory Studies
Certificate of Analysis
• Certified Values • Reference Values • Expiration Date • Storage • Instructions for Use • Material Information • Information on
Analyses • Moisture Determination
Traceability
Traceability has been defined as: a “property of a measurement result whereby the result can be related to a reference through a
documented unbroken chain of calibrations, each contributing to the measurement uncertainty”
CRM referencestandard
“A”
referencestandard
“A”
Calibrant“B”
Unknown“C”
Calibrant“B”
Measurements of unknown “C” are traceable to the CRM Measurement uncertainty increases with each comparison
CRM referencestandard
“A”
referencestandard
“A”
Calibrant“B”
Unknown“C”
Calibrant“B”
Traceability depends on an “unbroken chain of calibrations”
(suppose B is “damaged”)
Traceability
In principle, measurements that are traceable are also comparable
To assess potential breaks in the traceability chain, ask the question:
“What could go wrong with this measurement?”
Method suitability? Specificity, sensitivity, repeatability, dynamic range, sources of bias
Instrumentation? CRM commutability? Extraction efficiency Matrix interferences Analyte stability Calibration
Chemical Metrology for
Environmental Assessment
Metrology and Physical Constants
International School of Physics “Enrico Fermi”
Stephen A. Wise
Analytical Chemistry Division
Material Measurement Laboratory
National Institute of Standards and Technology (NIST)
Gaithersburg, Maryland USA
Polycyclic Aromatic Hydrocarbons
(PAHs)
Naphthalene
Acenaphthene
Acenaphthylene
Fluorene
Anthracene
Phenanthrene
Fluoranthene
Pyrene
Benz[ ]anthracenea
Chrysene
Benzo[ ]fluorantheneb
Benzo[ ]fluoranthenek
Benzo[ ]pyrenea
Benzo[ ]peryleneghi
Indeno[1,2,3- ]pyrenecd
Dibnez[ ]anthracenea,h
Products of combustion of organic
material, e.g., coal/oil/wood burning,
auto exhaust, cigarette smoke
Many are mutagenic or carcinogenic
Large number of isomeric structures
Use Multiple Analytical Methods
exploit differences in
Extraction Isolation and Cleanup
Separation and Detection
minimizes possibility of undetected bias in
resulting certified concentrations
that
Independent Analytical Methods Approach
for Certification of Organic Contaminants
Reversed-Phase LC with Fluorescence
Detection for Determination of PAHs
0
Naphthalene-d8
Phenanthrene-d10
Perylene-d12
Minutes
Benzo[a]pyrene-d12
1 2 3 4 5 6 7 8 9 10
2 = 249/362
3 = 250/400
4 = 285/450
5 = 333/390
6 = 285/385
7 = 263/358
8 = 406/440
9 = 296/405
10 = 300/500
1 = 280/340
Fluoranthene-d10
SRM 1941a Marine Sediment
Multidimensional LC Approach for
Determination of PAHs
0 10 20 30 40 50 60 70 80 90 100
278 MW Fraction 302 MW Fraction
x2
x5
x20
Time (min)
Excitation/Emission (nm)
0 5 10 15 20
1 0 3 4 2
1 2
4
5
6
7
IS
287/377 298/405 320/430 287/391 Reversed-Phase LC
Normal-Phase LC
Excitation/Emission (nm)
0
1
2
3
0
1
2
3
0 2 3 1
0 5 10 15 20 25 30
2
3
6 7
8 9 10 IS
4 370/420 395/435 334/464 312/454
Reversed-phase LC
separation of PAH isomers
Normal-phase LC separation
based on number of aromatic
carbons
SRM 1597 Coal Tar
Use of Multiple GC Columns with Different
Selectivity for PAHs (MW 228)
triphenylene and chrysene
chrysene
chrysene
triphenylene
triphenylene
benz[ ]-anthracene
a
benz[ ]-anthracene
abenz[ ]-anthracene
a
benzo[ ]-phenanthrene
c
benzo[ ]-phenanthrene
c
benzo[ ]-phenanthrene
c
50 % phenylmethylpolysiloxane
dimethyl (50 % liquid crystal)polysiloxane column
non-polar (DB-XLB)
GC/MS Analysis of SRM 2585 Organic Contaminants in House Dust
Use of Multiple GC Columns with Different
Selectivity for PAHs (MW 252)
benzo[ ]-fluoranthene
bbenzo[ ]-fluoranthene
b
benzo[ ]-fluoranthene
k
benzo[ ]-fluoranthene
k
benzo[ ]-fluoranthene
j
benzo[ ]pyrenee
benzo[ ]pyrenee
benzo[ ]pyrenea
benzo[ ]pyrenea
perylenebenzo[ ]-fluoranthene
a
benzo[ + ]-fluoranthene
a j
perylene
50 % phenylmethylpolysiloxane
dimethyl (50 % liquid crystal)polysiloxane column
GC/MS Analysis of SRM 2585 Organic Contaminants in House Dust
Molecular Mass 302 PAHs
N
N N N NN
N
NDBbeF
(#1)
?unknown
(#2)
N12bF
(#3)
N12kF
(#4)
N23jF
(#5)N23bF
(#7)
DBbkF
(#8)
DBakF
(#9)
DBjlF(#10)
N12eP(#11)
DBalP(#12)
?unknown
(#13)
N23kF
(#14)
N12aP(#15)
?unknown
(#16)
N23eP(#17)
DBaeP(#18)
N21aP(#20)
DBelP(#21)
N23aP(#22)
BbPer(#23)
DBaiP(#24)
DBahP(#25)
DBbeF
(#1)
?unknown
(#2)
N12bF
(#3)
N12kF
(#4)
N23jF
(#5)N23bF
(#7)
DBbkF
(#8)
DBakF
(#9)
DBjlF(#10)
N12eP(#11)
DBalP(#12)
?unknown
(#13)
N23kF
(#14)
N12aP(#15)
?unknown
(#16)
N23eP(#17)
DBaeP(#18)
N21aP(#20)
DBelP(#21)
N23aP(#22)
BbPer(#23)
DBaiP(#24)
DBahP(#25)
MW 302:Dibenzo/Naphthopyrenes and Fluoranthenes
GC/MS and LC/FL Determination of MW 302
PAHs in Coal Tar (SRM 1597a)
PAHs MW 302 Isomers Value Assigned
(mg/kg)
Naphtho[1,2-b]fluoranthene (N12bF) 8.6 ± 2.0
Naphtho[1,2-k]fluoranthene (N12kF) 10.7 ± 1.2
Naphtho[2,3-b]fluoranthene (N23bF) 3.52 ± 0.30
Dibenzo[b,k]fluoranthene (DBbkF) 11.2 ± 0.8
Naphtho[2,3-e]pyrene (N23eP) 4.31 ± 0.44
Dibenzo[a,e]pyrene (DBaeP) 9.08 ± 0.39
Naphtho[2,3-a]pyrene (N23aP) 4.29 ± 0.89
Dibenzo[a,h]pyrene (DBahP) 2.57 ± 0.30
80 85 90 95 100 105
X
X
X
X...Unknown
N…mutagen or
carcinogen
N DBbkF
DBaeP N
N DBahP
N DBaiP
N
DBalP
N21aPN
N23ePN
DBbeF
N12bF
N12kF
N23jF
N23bF
DBakF
DBjlF
N23kF
DBelP
BbPer
N
N12aPN
N23aP
N12eP
m/z 302
80 85 90 95 100 10580 85 90 95 100 105
X
X
X
X...Unknown
N…mutagen or
carcinogen
N DBbkF
DBaeP N
N DBahP
N DBaiP
N
DBalP
N21aPN
N23ePN
DBbeF
N12bF
N12kF
N23jF
N23bF
DBakF
DBjlF
N23kF
DBelP
BbPer
N
N12aPN
N23aP
N12eP
m/z 302
Values assigned based on combined results from
GC/MS on two different columns and LC/FL.
Use of Natural Matrix SRMs for
New Method Validation
Comparison of Extraction Methods
0
10
20
30
40
50
60
0
1
2
3
4
5
6
7
8 10
5
0
Fluoranthene BaA BeP BghiP IP
S o
x h
l e t
( D C
M )
P F
E (
D C
M )
S F
E (
C O
2 2
0 0
C )
P
F E
( T
/ M )
S o
x h
l e t
( D C
M )
P F
E (
D C
M )
S F
E (
C O
2 2
0 0
C
)
P F
E (
T / M
)
S o
x h
l e t
( D C
M )
P F
E (
D C
M )
S F
E (
C O
2 2
0 0
C
)
P F
E (
T / M
)
S o
x h
l e t
( D C
M )
P F
E (
D C
M )
S F
E (
C O
2 2
0 0
C
)
P F
E (
T / M
)
S o
x h
l e t
( D C
M )
P F
E (
D C
M )
S F
E (
C O
2 2
0 0
C
)
P F
E (
T / M
)
S o
x h
l e t
( D C
M )
P F
E (
D C
M )
S F
E (
C O
2 2
0 0
C
)
S o
x h
l e t
( D C
M )
P F
E (
D C
M )
S F
E (
C O
2 2
0 0
C
)
S o
x h
l e t
( D C
M )
P F
E (
D C
M )
S F
E (
C O
2 2
0 0
C
)
S o
x h
l e t
( D C
M )
P F
E (
D C
M )
S F
E (
C O
2 2
0 0
C )
S o
x h
l e t
( D C
M )
P F
E (
D C
M )
S F
E (
C O
2 2
0 0
C
)
Fluoranthene BaA BeP BghiP IP
m g/g m g/g
SRM 1650a SRM 1649a
B. A. Benner, Anal. Chem., 70 (1998) 4594-4601
M. M. Schantz, et al., Anal. Chem., 69 (1997) 4210-4219
Comparison of Extraction Methods
Diesel Particulate Matter Air Particulate Matter
Analytical Approach for Determination
of Polycyclic Aromatic Hydrocarbons (PAHs)
Extraction Soxhlet extraction
Pressurized Fluid Extraction (PFE)
Use of different solvents
Clean Up/Isolation SPE (silica, amino)
LC (normal-phase)
total PAH fraction
isomer fraction
Separation and Analysis LC-fluorescence (total fraction)
LC-fluorescence (Isomer fraction)
GC/MS (different columns)
5% phenyl phase
50% phenyl phase
nonpolar phase (DB-XLB)
liquid crystal phase
Add 13C and/or deuterated
PAHs as internal standards
Recent SRM Certifications for
PAHs (µg/g)
SRM 1597a Coal Tar SRM 1649b Urban Dust
Phenanthrene 454 ± 7 (1.5%) 3.941 ± 0.047 (1.2%)
Fluoranthene 327 ± 7 (2.1%) 6.14 ± 0.12 (2.0%)
Pyrene 240 ± 7 (2.9%) 4.784 ± 0.029 (0.6%)
Benz[a]anthracene 98.1 ± 2.3 (2.3%) 2.092 ± 0.048 (2.3%)
Chrysene 66.2 ± 5.3 (8.0%) 3.008 ± 0.044 (1.5%)
Benzo[k]fluoranthene 41.2 ± 0.4 (1.0%) 1.748 ± 0.083 (4.7%)
Benzo[b]fluoranthene 66.1 ± 4.4 (6.7%) 5.99 ± 0.20 (1.6%)
Benzo[a]pyrene 93.5 ± 1.4 (1.5%) 2.47 ± 0.17 (6.7%)
Benzo[e]pyrene 50.4 ± 1.0 (2.0%) 2.970 ± 0.043 (1.5%)
Benzo[ghi]perylene 50.5 ± 0.6 (1.2%) 3.937 ± 0.052 (1.3%)
Indeno[1,2,3-cd]pyrene 55.5 ± 0.8 (1.4%) 2.96 ± 0.17 (5.7%)
Dibenzo[a,e]pyrene 9.08 ± 0.39 (4.3%) 0.538 ± 0.024 (4.5%)
Natural Environmental Matrix SRMs
for Organic Contaminants
Fossil Fuels/Combustion Products petroleum crude oil* shale oil* coal tar extract* coal tar/petroleum solution diesel particulate matter*
Marine and River Sediments**
Biological Tissues mussel tissue (frozen and freeze-dried)*** fish oil and fish tissue** whale blubber** human serum** human milk** human urine**
Air Particulate Matter air particulate matter (TSP)*** fine particulate matter (<10 µm)*** house dust***
* Values for PAHs
* Values for PCBs/pesticides
* Values for PBDEs
Recent environmental matrix SRMs
typically have values assigned as
appropriate for:
• 30 - 50 PAHs
• 10 - 15 Nitro-PAHs
• 40 - 50 PCB congeners
• 10 - 15 Chlorinated pesticides
• 10 - 15 PBDEs Wise et al., Anal. Bioanal. Chem.
386:1153-1190 (2006)
SRM 1640 Trace Elements
in Natural Water
SRM 2694a Simulated
Rainwater
SRM 1646a Estuarine Sediment
SRM 1944 NY/NJ Waterway Sediment
SRM 2702 Inorganics in Marine Sediment
SRM 8704 Buffalo River Sediment
SRM 1947 Lake Superior Fish Tissue
SRM 1566b Oyster Tissue
SRM 2976, 2977 Mussel Tissue
SRM 1648 Urban Particulate
SRM 2783 Air Particulate on Filter Media
SRM 2583 Trace Elements in Indoor Dust
The Biosphere of Environmental Matrix SRMs for Trace Elements
SRM 2781 Domestic Sludge
Not a comprehensive list of environmental NIST Standard Reference Materials; see www.nist.gov/srm
SRM 1515 Apple Leaves
SRM 1547 Peach Leaves
SRM 1570a Spinach Leaves
SRM 1573a Tomato Leaves
SRM 1548a Total Diet
SRM 1549 Milk Powder
SRM 1577b Bovine Liver
SRM 1598a Animal Serum
SRM 966 Toxic Metals in
Bovine Blood
SRM 2670a Toxic Metals in
Freeze-dried Urine
SRM 2709 San Joaquin Soil
SRM 2710 Montana Soil Highly Elevated
SRM 2711 Montana Soil Moderately Elevated
SRM 2780 Hard Rock Mine Waste
SRM 1632c Bituminous Coa,l etc.
SRM 1634c Trace Elements in Fuel Oil, etc.
Fish Tissue SRMs
Food vs. Environmental Matrix Two frozen fish tissue homogenate meets the
need for both environmental and food matrix CRMs
SRM 1946 Lake Superior Fish Tissue
SRM 1947 Lake Michigan Fish Tissue
Certified and Reference Concentrations for ~100 constituents including:
42 – 45 PCB congeners (13 congeners not value assigned in previous SRMs including 3 non-ortho-substituted PCB congeners)
17 Chlorinated pesticides
9 PBDE congeners
16 – 25 Fatty acids (largest number in any SRM including first values of omega-3 fatty acids)
Total Hg, methyl-Hg, and 11 elements
Proximates (fat, protein, carbohydrate)
Emerging Contaminants in Human
Monitoring
SRM 1589a Contaminants in
Human Serum issued
Serum collected in 1996
Certified for PCBs, pesticides, and
PBDEs
SRM 1589a used extensively by
CDC for human monitoring studies
including NHANES
Need for more contemporary
human serum to reflect decrease
in legacy contaminants and
increase in emerging contaminants
0
20
40
60
80
100
120
1984 1986 1988 1990 1992 1994 1996 1998 2000 2002
0
20
40
60
80
100
120
1984 1986 1988 1990 1992 1994 1996 1998 2000 2002
ng
/g lip
id
PCB 153
PBDE 47
Sjödin et al., Retrospective Time-Trend Study of Polybrominated Diphenyl Ether and Polybrominated and
Polychlorinated Biphenyl Levels in Human Serum from the United States, Environ. Health Perspect. 112, 654-658,
2004.
SRMs for Contaminants in Human
Serum and Milk
Collaboration with CDC to develop contemporary serum and milk SRMs for contaminant measurements
Two materials for both serum and milk: natural level and fortified (≈5 to10 x natural)
Target list of over 170 organic contaminants including:
PCBs and hydroxylated PCBs
Chlorinated pesticides
Chlorinated and brominated dioxins/furans
Brominated flame retardants (PBDEs)
PCNs
PFCs
Toxaphene
Halogenated phenols
Measurements by multiple methods at NIST and CDC
SRM 1957 Non-fortified Serum –
Method Comparison
GC/MS
SRM 1958 Fortified Serum –
Method Comparison
GC/MS
Per- and Poly-Fluorinated Alkyl
Compounds (PFCs)
F F F
F
F F F F
F F F F F F F F
F
S
O
O
O-
PFOA (C8)
PFOS
Perfluorocarboxylates PFCAs
Perfluorobutanoic acid PFBuA
Perfluoropentanoic acid PFPeA
Perfluorohexanoic acid PFHxA
Perfluoroheptanoic acid PFHpA
Perfluorooctanoic acid PFOA
Perfluorononanoic acid PFNA
Perfluorodecanoic acid PFDA
Perfluoroundecanoic acid PFUnDA
Perfluorododecanoic acid PFDoDA
Perfluoroalkanesulfonates/amides PFSs
Perfluorohexanesulfonate PFHxS
Perfluorooctanesulfonate PFOS
Perfluorooctanesulfonamide PFOSA
Perfluorobutanesulfonate* PFBS
F F
F
F
F F F F
F F F F F F F
O
O-
F F F
F
F F F F
F F F F F F F F
F
S
O
O
NH2
PFOSA
DuPont
3M Co.
PFC use began in 1956 as:
Stain protection for carpet, textile, leather, paper and board (fast food boxes)
Fire-fighting foams
Specialty surfactants (cosmetics, electronics, etching, medical uses, plastics)
Polymerization aid for Teflon (clothing, bedding, non-stick cookware, automobiles)
Determination of PFAAs by using
LC-MS/MS
PF
NA
and P
FO
S
PF
BA
PF
BS
PF
PeA
PF
HxS
PF
HpA
PF
HxA
PF
OA
PF
DA
PF
Un
A
PF
DoA
PF
TriA
PF
TA
PF
OS
15 PFAA standards Pentafluorophenyl column
Method of J. Reiner et al.
Anal. Bioanal. Chem. 2012
Determination of PFAAs in House
Dust using LC-MS/MS
PF
NA
and P
FO
S
PF
Pe
A
PF
HxS
P
FH
pA
PF
HxA
PF
OA
PF
DA
PF
OS
A
SRM 2585 Organic Contaminants in House Dust
PF
BS
Jessica Reiner, NIST
SRMs Analyzed for Determination
of PFAAs
SRM 2585 Organic Contaminants in House Dust
SRM 2781 Domestic Sludge
SRM PFOS PFOSA PFBA PFHxA PFHpA PFOA
2585 1255 ± 125 2218 ± 274 241 ± 33 243 ± 11 248 ± 29 528 ± 139
2781 215 ± 69 5.9 ± 1.0 23.4 ± 0.1 10.7 ± 1.0 6.9 ± 1.0 30.6 ± 0.3
Results are ng/g ± SD (n = 2)
Results reported for nine additional PFAAs in both SRMs
Jessica Reiner, NIST
Elemental Speciation Measurements in SRMs
Why is elemental speciation important?
• Great importance in biological and
environmental processes
• Toxic/beneficial effects often dependent
on species NOT the total concentration
• The specific chemical forms of an element
should be considered individually
Simultaneous measurements of mercury and tin species by
GC/ICP-MS: SRM 1974b Organics in Mussel Tissue (Mytilus
edulis)
µg L-1 as Arsenic
As (III) = 5.03 0.31
As (V) = 6.16 0.95
MMA = 7.18 0.56
DMA = 25.3 0.7
TMAO = 1.94 0.27
AB = 1.43 0.08
AC = 3.74 0.35
Total As = 52.8 3.5
SRM 2669, As Species in Frozen Human Urine: As speciation
by coupled IC/ICP-MS
Current Areas of Interest
• Biological Tissues
– Marine SRMs
– Fish, Oysters, Mussels
– Marine Mammals
• Clinical
– Blood, Urine
– Nutritional Supplements,
– Pharmaceutical/Nutraceutical Products
– Metalloproteins
• Environmental
– Soils, sediments
Arsenic Species in Frozen
Human Urine (SRM 2669)
Prolonged exposure to arsenic can lead to skin, pulmonary, and bladder cancer.
EPA limit on drinking water <10 ppb.
Urine is the best matrix for measurement of arsenic exposure.
Concentrations of total As and 7 As species were certified in the urine: arsonite (As III), arsonate (As V), monomethylarsonate (MMA), dimethylarsinate (DMA), trimethylarsine oxide (TMAO), arsenobetaine (DMA), and arsenocholine (AC).
SRM consists of two concentration levels: normal and elevated levels based on the 95 percentile as determined by CDC population studies.
0
1000
2000
3000
4000
5000
6000
7000
8000
9000
10000
0 5 10 15 20 25 30
Time (min)
cp
s @
m/z
75 As (III)
DMA
MMA
As (V)
AB
TMAO AC Development of SRM 2669 is a collaboration
between NIST and the Centers for Disease Control
and Prevention (CDC) with measurements using
different methods contributed by NIST, CDC, and
three additional Laboratories.
Produced in N2,
packaged with O2
absorber in gas
impermeable bags,
and stored at -80 C
to prevent species
conversion
Mutual Recognition Arrangement (MRA)
developed by the CIPM
MRA signed by 38 NMI Directors in 1999 and by 27 since then
Provides: Open, transparent, and comprehensive scheme to give users reliable
quantitative information on the comparability of metrology services worldwide needed for mutual recognition of national measurement standards and measurement certificates issued by national metrology institutes
Technical basis for wider agreements negotiated for international commerce and regulatory affairs
Requires: Declaring and documenting calibration and measurement capabilities
Evidence of successful participation in formal, relevant international comparisons
Demonstration of system for assuring quality of each NMI’s measurement services
Traceability to stated references and global confidence in this realization are the basis for mutual
recognition and confidence in data used to facilitate and underpin international trade and decisions
regarding health, safety, commerce, and scientific studies
27
6 pH
7 Electrolytic Conductivity
8 Metals and Metal Alloys 8.1 Ferrous Metals
8.2 Non-Ferrous Metals
8.3 Precious Metals
8.4 Other
9 Advanced Materials 9.1 Semiconductors
9.2 Superconductors
9.3 Polymers and Plastics
9.4 Ceramics
9.5 Other
10 Biological Fluids and
Materials 10.1 Blood, Plasma, Serum
10.2 Urine Fluids
10.3 Hair
10.4 Tissues
10.5 Bone
10.6 Botanical Materials
10.7 Other
11 Food 11.1 Nutritional Constituents
11.2 Contaminants
11.3 GMOs
11.4 Other
1 High Purity Chemicals 1.1 Inorganic Compounds
1.2 Organic Compounds
1.3 Metals
1.4 Isotopics
1.5 Other
2 Inorganic Solutions 2.1 Elemental
2.2 Anionic
2.3 Other
3 Organic Solutions 3.1 PAHs
3.2 PCBs
3.3 Pesticides
3.4 Other
4 Gases 4.1 High Purity
4.2 Environmental
4.3 Fuel
4.4 Forensic
4.5 Medical
4.6 Other
5 Water 5.1 Fresh Water
5.2 Contaminated Water
5.3 Sea Water
5.4 Other
Current CCQM Measurement Service Category Numbers and Categories
12 Fuels 12.1 Coal and Coke
12.2 Petroleum Products
12.3 Bio-mass
12.4 Other
13 Sediments, Soils, Ores,
and Particulates 13.1 Sediments
13.2 Soils
13.3 Ores
13.4 Particulates
13.5 Other
14 Other Materials 14.1 Cements
14.2 Paints
14.3 Textiles
14.4 Glasses
14.5 Thin Films
14.6 Coatings
14.7 Insulating Materials
14.8 Rubber
14.9 Adhesives
14.10 Other
15 Optical Properties
Recent Participation in CCQM Pilot
Comparisons
CCQM-P28 Ozone, Ambient Level
CCQM-P46 Preparation of Elemental Solution Standards
CCQM-P57 PCB Congeners in Tissue Extract
CCQM-P61 Volatile Organic Compounds in Solution
CCQM-P62 High-Purity Nickel
CCQM-P64 Study of Nonfat Soybean Powder Ca, Cu, Fe, and Zn
CCQM-P66 Determination of Cu and Cd in Multi-Nutrient Fertilizer
CCQM-P67 PCB Congeners in Tissue
CCQM-P68 19-Norandrosterone in Human Urine
CCQM-P69 PAHs in Soil
CCQM-P73 50 µmol/mol to 70 µmol/mol Nitric Oxide Preparative
CCQM-P75 Measurement of d34S in Methionine
CCQM-P77a Cortisol in Serum
CCQM-P77b Progesterone in Serum
CCQM-P78 Vitamins in Infant/Adult Formula
CCQM-P86 Analysis of Total Se and Selenomethionine in Pharmaceutical
Supplements
bold = NIST was the Coordinating or Co-coordinating Laboratory
Recent Participation in CCQM Key
Comparisons
CCQM-K15 SF6 and CFCs in Nitrogen
CCQM-K18 pH (Carbonate Buffer)
CCQM- K19 pH (Borate Buffer)
CCQM-K22 Volatile Organic Compounds in Air
CCQM-K26a Reactive Gases: Nitrogen Monoxide (NO) in Nitrogen (N2)
CCQM-K27.2 Ethanol in Water
CCQM- K36 Electrolytic Conductivity at 0.5 S/m and 5 mS/m
CCQM-K38 PAHs in Solution (also P31.a.1)
CCQM-K39 Chlorinated Pesticides in Solution (also P31.c.1)
CCQM-K40 PCB Congeners in Solution (also P31.b.1)
CCQM-K41 H2S in Air/Nitrogen
CCQM-K42 Minor Elements in Aluminum Alloy 5182
CCQM-K43 Measurement of Total Mercury in Salmon
CCQM- K47 Volatile Organic Compounds in Methanol
CCQM-K49 Essential and Toxic Elements in Bovine Liver
CCQM-K52 CO2 in Air
CCQM-K53 O2 in Nitrogen Preparative
CCQM-K54 n-Hexane in Nitrogen
bold = NIST was the Coordinating or Co-coordinating Laboratory
CCQM- K24 Cd in rice
CCQM-K5 p,p’-DDE in fish oil
NOMINAL VALUE: 7.5 mmol/mol
CCQM-K14: Calcium in Serum CCQM-16: Gas mixtures - Natural Gas Type IV – ethane (1 of 12 measurands) (low calorific mixture)
CCQM-K49 – Bovine Liver Fe Results
Key Comparison and Pilot Study of Fe, Zn, Se, Cd, Pb, Cr, As in bovine liver
All NMIs with inorganic analytical capabilities were requested to participate at least for Fe and Zn as part of a program to compare the capabilities of most NMIs on a regular basis
Results received from 21 NMIs and 4 non-NMIs
Material will be certified as SRM 1577c Trace Elements in Bovine Liver
U95% = ± 2.1%
CCQM-P39 and IMEP-20:
Pb in tuna fish IA
EA
_A
AS
BN
M-L
NE
LG
C
IRM
M
NM
IJ
EN
EA
IAE
A_IC
P-M
S
Fro
ntier
Geoscie
nces,
Inc.
50
40
30
20
10
0-1
0-2
0-3
0-4
0-5
0
Devia
tion f
rom
MM
-media
n in % GF-AAS
ICP-IDMS
ICP-MSext.
calibration
Types of Standard Reference Materials for
Chemical Composition Analyses
Pure compounds
Solutions
Natural matrices
Other (e.g., extracts,
metals)
http://www.nist.gov/srm
Use of Pure-Compound SRMs
Calibration of an instrument
Use as an internal standard
Method development/quality control in purity
assessment
Provide traceability
Use of Solution SRMs
Calibration solutions
Solutions for use as internal standards
Method development/quality assurance
Provide traceability
Uses of Natural Matrix SRMs
Analytical Method Validation
Control materials for quality assurance
of routine analytical measurements
Research applications – New method
development
Uses of Natural Matrix SRMs
Validate the complete analytical procedure including extraction, isolation, cleanup, separation, and quantification
Validate new analytical methods
For contaminants with values assigned PAHs, nitro-PAHs
PCBs, Pesticides, PBDEs
For contaminants not value assigned (emerging pollutants)
Homogeneous materials
Readily available to other laboratories world-wide for comparison of results
Analytical Method Validation
In general, natural-matrix materials
should not be used as calibrants
Relative uncertainties on calibrant SRMs range from less than a percent to a few parts per 100,000 (i.e., 0.001%). Certified value for copper in SRM 3114 Copper
Standard Solution – 0.16% relative expanded uncertainty
Relative uncertainties on natural-matrix materials are on the percent level. Certified value for copper in SRM 3244 Ephedra-
Containing Protein Powder – 9.8% relative expanded uncertainty
Plus, using a natural-matrix material for
calibration assumes that your material is
perfectly matched to it.
Q. If I analyze the SRM one time, how do I know
whether my result agrees with the assigned value?
To calculate the uncertainty on your single
value, you’ll need to know the reproducibility
standard deviation (sR) of your analysis.
If you don’t know this, you’ll need to do more
than one analysis.
22 )2/(/ Unsu R
Comparison of “your” single value to
the assigned value
a-Tocopherol in SRM 2384
0
1
2
3
4
5
6
7
8
9
10
1 2 3 4 5
Ma
ss F
rac
tio
n (
mg
/kg
)
Reference value
+U
-U
Uses of Natural Matrix SRMs
Analytical Method Validation
Control materials for quality assurance
of routine analytical measurements
Research applications – New method
development
Selection of a QC Material
Similar matrix Same sample preparation requirements
Likelihood of similar analyte recovery
Similar analyte levels Results fall within the same general range of the
calibration curve
Values assigned for analyte(s) of interest Check that the uncertainties on assigned values
make the material suitable for your purpose
Uses of Natural Matrix SRMs
Validate the complete analytical procedure including extraction, isolation, cleanup, separation, and quantification
Validate new analytical methods
For contaminants with values assigned PAHs, nitro-PAHs
PCBs, Pesticides, PBDEs
For contaminants not value assigned (emerging pollutants)
Homogeneous materials
Readily available to other laboratories world-wide for comparison of results
Analytical Method Validation
Method Validation: Comparison of
Results for New Constituents
L.Y. Zhu and R.A. Hites, Anal. Chem. 75:6696-6700 (2003)
Analyzed fish (2), mussel (3), and whale blubber tissue SRMs
SRM 1947 Lake Michigan Fish Tissue
0
10
20
30
40
50
60
70
80
90
BDE 47 BDE 99 BDE 100 BDE 153 BDE 154
Co
nc (
ng
/kg
) w
et
mass
NIST I NIST II NIST III Zhu and Hites