2.1 le blanc
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HOW CONFIDENT ARE WE?
Alain LeBlancSpecial projects’ manager,
support services and external quality assessment
Pierre DumasResearch & Development division
Alain LeBlancSpecial projects’ manager,
support services and external quality assessment
Pierre DumasResearch & Development division
Centre de toxicologie du Québec (CTQ) INSPQ, Québec, CanadaCentre de toxicologie du Québec (CTQ) INSPQ, Québec, Canada
ISBM, Manchester UK, 9th ‐ 11th September 2013
www.inspq.qc.ca/ctq
Among its expertise, the Centre de toxicologie du Québec (CTQ) offers analytical support in clinical, environmental and occupational health
The INSPQ is a provincial government body created to improve the coordination, development and use of expertise in public health
CENTRE DE TOXICOLOGIE DU QUÉBEC (CTQ)Claude Thellen
QUEBEC MINISTRY OF HEALTH AND WEALTHFARE (MSSS)INSTITUT NATIONAL DE SANTÉ PUBLIQUE DU QUÉBEC (INSPQ)
ENVIRONMENTAL HEALTH AND TOXICOLOGY DEPT. (DSET)
TOXICOLOGYLABORATORY BRANCH
Normand FleurySPECIAL PROJECTS, SUPPORT SERVICES and EQAS SECTOR
Alain LeBlanc
RESEARCHPierre Ayotte
METHOD DEVELOPMENTPatrick Bélanger
CLINICALMichel Lefebvre
ORGANICSEric Gaudreau
METALSCiprian‐Mihai
Cirtiu
PROJECT MANAGEMENTand SUPPORT
SERVICES
EXTERNAL QUALITY
ASSESSMENT SCHEMES
QUALITY MANAGEMENTSergine Lapointe
To make scientists, researchers and epidemiologists aware of day‐to‐day issues that
laboratories encounter when confrontedwith testing new environmental contaminants
Triclosan
• To develop reliable, robust and accurate analytical methods
• To assure tracability of the metrology
• To assure tracability of the data
• To assure the comparability of the analytical results
• To develop and maintain competence of staff
Sample collection, integrity and storage• Time of collection (spot/24 hrs; day‐to‐day variability)
• Handling (freeze/thaw)
• Stability (+ > 5 yrs)
• Homogeneity: free/conjugated species
• Contamination (collection process, container, …)
• Sampling container (adsorption onto plastics) / Tubes
• Are we sure we are measuring the right biomarker? in the right matrix?
• Shipping considerations (overnight, deep freeze)
Analytical innovation• Keep costs down; compromise with multi‐analyte methods
• Tend to less invasive matrices, but are they the right ones?
Analytical standards • Availability of conjugated analytes
• Accuracy (certificates, multiple sources)
• Impact on biomonitoring data (cycle to cycle)• Availability of appropriate internal standards
13C labeled and unlabeled analogs are usually not available at the same time• Custom synthesis (TRC, CANSYN, CHIRON, CIL, etc)
High cost, time to delivery, lot‐to‐lot, purity (UV, NMR, IR and MS spectra)
Analytical methods• Classical versus experimental protocols
• Sensitivity (LODs) and specificity
• Reportable limits (no international consensus; study comparisons)
• Laboratory contamination (ex: BPA, Triclosan, Parabens, BDEs, …)
Analytical instrumentation• Comparability
Reference materials• Availability (certified?)
• Appropriateness (same matrix?) commutability
• Internal RM 2nd source of standard!!
External quality assessment schemes• How to compare study results?
• Collaboration among reference labs or research centers
• To help reduce vulnerability
SAMPLE INTEGRITY AND STORAGE
-30
-20
-10
0
10
20
30Te
mpe
ratu
re°C
Sample handling: Chronological events(worst case scenario)
Sample Temperature Tracking Chart« Freeze/thaw »
Freeze/Thaw ‐ Stability of total BPA in urine (14 samples)
80
85
90
95
100
105
110
115
120
125
cycle 0 cycle 1 cycle 2 cycle 3 cycle 4
% B
PA
-10%
-5%
+5%
+10%
88
90
92
94
96
98
100
cycle 0 cycle 1 cycle 2 cycle 3 cycle 4
%C
onju
gate
d B
PA
Freeze/Thaw ‐ Stability of conjugated BPA in urine (14 samples)
• Free Triclosan can be rapidly adsorbed onto plastic tips according to type of material and % of acetonitrile in spike solution.
• Time residence in the pipette tips can increase the degree of adsorption.
Tip Y 0% ACN
Tip Y 5% ACN
Tip Y 50% ACN
Tip Y 0% ACN 1 Min
Tip Y 5% ACN 1 Min
Tip Y 50% ACN 1 Min
Tip B 0% ACN
Tip B 5% ACN
Tip B 50% ACN
Tip R 0% ACN
Tip R 5% ACN
Tip R 50% ACN
86
93
103
75
82
98
90
91
101
95
97
100
% Recovery in urine
11.7
12.7
13.7
14.7
15.7
16.7
17.7
18.7
4391
0
4449
5
4508
1
4566
6
4625
1
4683
7
4742
2
4800
7
4859
2
4917
8
4976
3
# séquence
Con
cent
ratio
n (u
g/L)
1.250
2.250
3.250
4.250
5.250
6.250
7.250
4391
0
4449
5
4508
1
4566
6
4625
1
4683
7
4742
2
4800
7
4859
2
4917
8
4976
3
# séquence
Con
cent
ratio
n (u
g/L)
Internal QC from unspiked urine. Endogenous conjugated form of trichlorophenol
External QC made from spiked urine using free trichlorophenol
Good stability
Poor stability
ANALYTICAL STANDARDS
Supplier MMP MEP MiBP MnBP MCHP MBzP MEHP MEHHP MEOHP MECPP MOP MCPP MNP
Certifiedsolutions
CIL‐2006 ‐25% ‐2% ‐ ‐47% ‐1% ‐63% ‐29% ‐11% ‐7% ‐ 12% ‐22% ‐39%
CIL‐2009 ‐9% ‐11% ‐5% ‐13% ‐12% ‐76% ‐14% ‐21% 4% 3% 2% ‐70% ‐16%
CIL‐2010 ‐3% ‐9% 0% ‐11% ‐7% 3% ‐11% ‐19% ‐9% ‐2% 3% ‐78% ‐10%
Chiron ‐ ‐ ‐3% ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐
Accustandard ‐26% ‐31% ‐ ‐19% ‐ ‐4% ‐27% ‐ ‐ ‐ ‐ ‐ ‐
Neatstandards
Accustandard 8% ‐7% ‐ ‐5% ‐ ‐2% ‐27% ‐ ‐ ‐ ‐ ‐ ‐
Aldrich 9% ‐ ‐ ‐6% ‐ ‐5% ‐ ‐ ‐ ‐ ‐ ‐ ‐
Chiron ‐ ‐21% ‐ 0% ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐
Dr Ehrenstorfer 13% ‐ ‐ ‐2% ‐ ‐ ‐29% ‐ ‐ ‐ ‐ ‐ ‐
CanSyn 9% ‐5% 4% ‐1% ‐6% ‐1% ‐8% ‐6% ‐4% ‐1% 10% ‐7% ‐3%
CIL 0% 0% 0% 0% 0% 0% 0% 0% 0% 0% 0% 0% 0%
TRC 7% ‐3% 4% ‐6% ‐10% ‐2% ‐11% 9% 9% ‐70% 12% ‐2% ‐6%
Langlois E., LeBlanc A., Simard Y., Thellen C. Accuracy investigation of phthalate metabolite standardsJournal of Analytical Toxicology, 2012 ; 36 :270-279
compounds synthesized at CHULBPA‐13C12 and TCS‐13C12 were obtained from Cambridge Isotope Laboratories
BPA
Triclosan
ANALYTICAL METHODS
• Cadmium in urine by ICP MS
• Mo‐O interferencecorrected VS uncorrected
• Mean concentration can be biased by 30 % with individual error greater than 200%
• Comparability between studies is strongly related to laboratory methodologies 130
78 9.523
32
21
0
5
10
15
20
25
30
35
0 50 100 150 200 250 300
Labo
ratorie
sCV
%
Ratio conc. Mo/Cd
Cd nmol/L
‐Mo in sample‐Mo in calibration curve
Cadmium Urine (nmol/L)QMEQAS 2011-2
Lab A
Lab B
Comparing United States and Canadian population exposures from National Biomonitoring Surveys: Bisphenol A intake as a case studyJudy S. LaKind, Johanne Levesque, Pierre Dumas, Shirley Bryan, Janine Clarke and Daniel Q. Naiman
Journal of Exposure Science and Environmental Epidemiology (2012) 22, 219‐226
X 1.6
X 1.6
QC Low QC High
Target value 2.90 ±0.23 10.17 ±0.58
CTQ 1 2.98 10.6
CTQ 2 2.82 10.2
PCB 153 (1µg/L) + PCB 132 (0.3 µg/L)Complete overlap of both congeners
NCI EI
0.000.200.400.600.801.001.201.401.601.802.00
35800 36800 37800 38800 39800
Blan
k level µg
/L
Sequence #
None Specific MetaboliteDiethyl thiophosphate (DETP)
Specific Metabolite3,5,6‐trichloro‐2‐pyridinol
Free BPA
G‐BPA
SO3‐BPA
(SO3)2‐BPA
y = 0.9149x - 0.0207R² = 0.9946
0
5
10
15
20
25
30
35
40
0 5 10 15 20 25 30 35 40
BP
A to
tal
µg
/L (G
C)
BPA free + BPA-glucuronide + BPA-sulfate + BPA-disulfate in µg BPA eq./L (LC)
LC‐MS/MS VS GC‐MS/MS
BPA
y = 1.0229x + 0.1818R² = 0.9954
0
100
200
300
400
500
600
700
800
900
0 100 200 300 400 500 600 700 800 900
TC
S t
ota
lµg
/L (
GC
)
TCS free + TCS-glucuronide + TCS-sulfate µg in TCS eq./L (LC)
LC‐MS/MS VS GC‐MS/MS
Triclosan
67.6
77.6
87.6
97.6
107.6
117.6
127.6
137.6
Con
cent
ratio
n (u
g/L)
# séquence
Setting up Single analyst + single instrument “Rush” multi analysts and instruments New standard batch
External Quality Assessment Schemesand Reference Materials
EQASBlood lead (17) AAFP CAP DigitalPt G‐EQUAS LAMP NEQAS NYSDH OELM PCI Penn QMEQAS RCPA SEKK SKZL TEQAS WIV‐ISP WSLHUrine BPA (1) G‐EQUASUrine Triclosan (0)Urine Triclocarban (0)
RMBlood lead (6) IAEA IRMM NIST RECIPE SERONORM UTAKUrine BPA (1) RECIPE [NIST]Urine Triclosan (0) [NIST]Urine Triclocarban (0)
More than 30 countries
250 laboratories
External Quality Assessment SchemesEQAS
Since 1979
• 3 continuing programs in blood, urine, serum and hairMetals: PCI, QMEQAS POPs: AMAP
• Material from human origin; unexposed volunteers
• 2 developing programs: MDA, Organic compounds in urine
• Voluntary participation
• Funded by the participants
• Reference material virtual store
36
REFERENCE MATERIALS
ISO/CEI 17043
www.inspq.qc.ca/ctq/sales
• Laboratories involved in HBM face challenges at all levels of operation
• Competence of staff must be developped and maintained
• Method validation must be brought to a higher level(stability, matrix effect, comparability of standards, …)
• Importance of External quality assessment schemes and referencematerials is obvious adapted QA strategy (work to do)
• A laboratory facility with a scientific and administratrive infrastructure ismandatory
• The term « new contaminant » alone shouldn’t set the rationale for biomonitoring
• A reference laboratory must have the capabilities and experience to tackle these recurring issues (not necessarily compatible with everylaboratory structure)
QUESTIONS?
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