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© 2015 AOAC INTERNATIONAL
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) and 4 mL hexane, followed by application of the 1 mL extract
in hexane. The analytes are eluted with hexane–dichloromethane
(3 + 1, v/v) using volume determined for the given silica gel SPE
cartridges from the elution profiles of target analytes and fat,
which are dependent on the silica deactivation. The clean extract
is carefully evaporated, reconstituted in 0.5 mL isooctane, and
analyzed by GC/MS.
See
Figure
2014.08A
for the method flow
chart.
B. Apparatus
(
a
)
Homogenizer
.—WARING blender Model 38BL40 (Conair
Corp., Stamford, CT) or equivalent.
(
b
)
Solvent evaporator
.—Any suitable solvent evaporator,
such as a rotary vacuum evaporator, Kuderna-Danish evaporator,
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.
Candidates for 2016 Method of the Year
220