AOAC Final Action Methods in 2018

AOAC Official Method 2014.08 Polycyclic Aromatic Hydrocarbons (PAHs) in Seafood Gas Chromatography-Mass Spectrometry First Action 2014

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,

[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.

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 2 3 4 5 6 7 8

5

12.5

25 50

50 50 50 50 50 50 50 50

0.5

1.25

2.5

5 5 5 5 5 5 5 5

10 20 50

25 50

1 2 5

2.5

5

100 250 500

5

10 25 50

125 250 500

12.5

100 200 500

10 20 50

25 50

1000 2500 5000

100 250 500

1250 2500

125 250

1000

100

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.

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