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astovska
et al
.:
J
ournal of
AOAC I
nternational
V
ol
.
98, N
o
. 2, 2015
481
calculations reviewed by a peer, laboratory supervisor, or
manager.
Data Analysis
The Study Directors reviewed and compiled all the
data submitted by the participants. Statistical analysis was
conducted using the AOAC spreadsheet for blind duplicates (4)
to determine mean analyte concentrations, SD (S
r
) and RSD
(RSD
r
) for repeatability (for blind duplicate data), SD (S
R
) and
RSD (RSD
R
) for reproducibility, number of valid data points,
HorRat value (RSD
R
/predicted RSD
R
), and percentage recovery
for all data after removal of outliers (3). The following tests were
used in the AOAC spreadsheet (4) to determine outliers: (
a
)
the
Cochran test for removal of laboratories showing significantly
greater variability among replicate (within-laboratory) analyses
than the other laboratories for a given material, and (
b
)
the
Grubbs’ tests for removal of laboratories with extreme averages.
AOAC Official Method 2014.08
Polycyclic Aromatic Hydrocarbons (PAHs)
in Seafood
Gas Chromatography-Mass Spectrometry
First Action 2014
[Applicable for the determination of the following PAHs
in mussel, oyster, and shrimp: 1,7-dimethylphenanthrene,
1-methylnaphthalene, 1-methylphenanthrene, 2,6-dimethyl-
naphthalene, 3-methylchrysene, anthracene, benz[
a
]anthracene,
benzo[
a
]pyrene, benzo[
b
]fluoranthene, benzo[
g,h,i
]perylene,
benzo[
k
]fluoranthene,
chrysene,
dibenz[
a,h
]anthracene,
fluoranthene, fluorene, indeno[1,2,3-
cd
]pyrene, naphthalene,
phenanthrene, and pyrene. These were representative PAH
analytes selected for the collaborative study. The method
has been single-laboratory validated for 32 PAHs in fish and
shrimp (1), and, therefore, is expected to be applicable to other
GC-amenable PAHs and seafood matrices. The concentration
ranges evaluated within the collaborative study are given in
Table
2014.08A
.]
Caution
:
See Appendix B: Laboratory Safety
. Use appropriate
personal protective equipment such as laboratory coat, safety
glasses or goggles, appropriate chemical-resistant gloves, and
a fume hood. Dispose of solvents and solutions according
to federal, state, and local regulations. Always handle open
containers of solvents inside the fume hood, including the
pouring, mixing, evaporating, and preparing standard solution.
Keep containers covered or closed when not in use.
Hexane and isooctane
.—Highly flammable, liquid irritants.
Harmful if inhaled, swallowed, or absorbed through the skin.
May also cause skin and eye irritation.
Ethyl acetate
.—Highly flammable, liquid irritants. Harmful
if swallowed in quantity. Vapors may cause drowsiness.
Toluene
.—Highly flammable, liquid irritant. Harmful if
inhaled, swallowed, or absorbed through the skin. May also
cause skin and eye irritation. May cause drowsiness. Possible
teratogen.
Dichloromethane
.—Noncombustible,
liquid
irritant.
Harmful if inhaled, swallowed, or absorbed through the skin.
May also cause skin and eye irritation. Asphyxiant. Causes
central nervous system (CNS) depression. Possible carcinogen
and mutagen.
PAHs
.—Carcinogens, respiratory sensitizers, teratogens,
reproductive hazard, mutagens. Harmful if inhaled, swallowed,
or absorbed through the skin. May also cause skin and eye
irritation.
See
Tables
2014.08B
–
D
for results of the interlaboratory
study supporting acceptance of the method.
A. Principle
Homogenized seafood samples (10 g sample with a 5 µg/kg
addition of
13
C-PAH surrogate mixture) are mixed with 5 mL
water (or 10 mL water in the case of shrimp and other more
viscous samples) and shaken vigorously by hand with 10 mL
ethyl acetate in a 50 mL polypropylene centrifuge tube for
1 min. Subsequently, 4 g anhydrous magnesium sulfate and
2 g sodium chloride are added to the mixture to induce phase
separation and force the analytes into the ethyl acetate layer.
The tube is again shaken by hand for 1 min and then centrifuged
for 10 min at >1500 rcf. A 5 mL aliquot of the ethyl acetate
layer is evaporated, reconstituted in 1 mL hexane, and cleaned
on an SPE column with 1 g silica gel and approximately
0.2 g anhydrous sodium sulfate on the top. The column is
conditioned with 6 mL hexane–dichloromethane (3 + 1, v/v)
Table 2014.08A. PAH and
13
C-PAH concentrations in the calibration standard solutions
Concentration, µg/L
Equivalent concentration, µg/kg
Calibration level
BaP and others
a
Chr and others
b
Naph
c
13
C-PAHs BaP and others Chr and others Naph
13
C-PAHs
1
5
12.5
25
50
0.5
1.25
2.5
5
2
10
25
50
50
1
2.5
5
5
3
20
50
100
50
2
5
10
5
4
50
125
250
50
5
12.5
25
5
5
100
250
500
50
10
25
50
5
6
200
500
1000
50
20
50
100
5
7
500
1250
2500
50
50
125
250
5
8
1000
2500
5000
50
100
250
500
5
a
Analytes at 10 µg/mL in the mixed stock standard solution.
b
Analytes at 25 µg/mL in the mixed stock standard solution.
c
Analytes at 50 µg/mL in the mixed stock standard solution.