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astovska
et al
.
:
J
ournal of
aoaC I
nternatIonal
v
ol
. 98, n
o
. 2, 2015
collecting 0.5 mL elution fractions in 20 evaporation tubes
or flasks. Add 0.5 mL isooctane to each elution fraction and
evaporate down to 0.5 mL using the optimized evaporation
conditions. Analyze each fraction by GC/MS.
(
c
) Determine PAH elution profile by plotting analyte
response (peak area or height) in a given fraction normalized
to the sum of analyte responses in all tested fractions vs the
elution volume.
See
Figure
2014.08B
for an example of a
PAH elution profile. It is recommended to add an additional
0.5 mL on top of the determined elution fraction (corresponding
to 100% recovery) as a safety margin ensuring good analyte
recoveries in routine practice. This would result in the optimum
elution volume of 7 mL for the silica cartridge tested in
Figure
2014.08B
.
(
d
) To check the effectiveness of fat removal, dissolve 100mg
pure fish oil (or any suitable fat) in 1 mL hexane and apply it
onto the silica gel cartridge, which was conditioned with 6 mL
hexane–dichloromethane (3 + 1, v/v) and 4 mL hexane. Elute
with the optimum elution volume of hexane–dichloromethane
(3 + 1, v/v), which was determined in the previous step (e.g.,
7 mL for the example in Figure
2014.08B
). Collect this
fraction in an evaporation tube or flask, which empty weight
(after heating in an oven to remove moisture) was recorded to
4 decimal places using an analytical balance. Elute the cartridge
with additional 3 × 1 mL hexane–dichloromethane (3 + 1, v/v)
and collect in three evaporation tubes/flasks of known empty
weight.
(
e
) Evaporate the four elution fractions to dryness and
gravimetrically determine the amount of fat eluting in each
fraction by subtracting the empty weights from newly recorded
weights after solvent evaporation. There should be no fat
eluting in the optimum elution fraction for PAHs (this can also
be observed visually in the tubes).
(
f
) If there is fat coeluting with PAHs, then the PAH and fat
elution profiles have to be reexamined to determine optimum
elution volume for PAH and fat separation (potentially
sacrificing up to 5% of late-eluting PAH amounts if necessary)
or a different silica gel cartridge has to be used.
G. GC/MS Analysis
(
a
)
GC conditions.
—Table
2014.08G
provides GC
conditions that were used by the collaborative study participants.
Other conditions (e.g., column, temperature and flow program,
and injection technique and volume) can be used as long as the
laboratory qualification criteria for separation, sensitivity, and
linearity are met. The injection temperature or program needs
to be optimized to enable quantitative transfer of less volatile
PAHs. If programmable temperature vaporizer (PTV) solvent
vent mode is used, solvent venting parameters (temperature,
time, flow, pressure) need to be carefully optimized to prevent
losses of the volatile PAHs, especially naphthalene. The
separation criteria (demonstrated in Figure
2014.08C
) include
(
1
) a baseline separation of benzo[
a
]pyrene and benzo[
e
]pyrene
(concentration ratio of 1:5), (
2
) at least 50% valley separation
of anthracene and phenanthrene (concentration ratio 1:2.5;
evaluated for the anthracene peak), and (
3
) at least 50% valley
Table 2014.08E. Analyte concentrations in the mixed
stock standard solution
Analyte
Concentration, µg/mL
Anthracene
10
Benz[
a
]anthracene
10
Benzo[
a
]pyrene
10
Benzo[
b
]fluoranthene
10
Benzo[
g,h,i
]perylene
10
Benzo[
k
]fluoranthene
10
Chrysene
25
Dibenz[
a,h
]anthracene
10
Fluoranthene
25
Fluorene
10
Indeno[1,2,3-
cd
]pyrene
10
Naphthalene
50
Phenanthrene
25
Pyrene
25
1-Methylnaphthalene
25
2,6-Dimethylnaphthalene
25
1-Methylphenanthrene
25
1,7-Dimethylphenanthrene
10
3-Methylchrysene
25
Table 2014.08F. Dilution scheme for preparation of the
calibration standard solutions
Calibration
level
Vol. of mixed
stock standard
solution, µL
Vol. of working
PAH solution
a
,
µL
Vol. of
working
PAH
solution B, µL
Vol. of
13
C-PAH
1 µg/mL
solution, µL
Final
vol.
a
, µL
1
—
—
50
50
1000
2
—
—
100
50
1000
3
—
—
200
50
1000
4
—
—
500
50
1000
5
—
100
—
50
1000
6
—
200
—
50
1000
7
—
500
—
50
1000
8
100
—
—
50
1000
a
Bring to volume using isooctane.
Figure 2014.08B. An example of elution profiles of PAHs on a silica gel SPE cartridge and
determination of the optimum elution volume.
Optimum elution volume
Figure 2014.08B. An example of elution profiles of PAHs on a
silica gel SPE cartridge and determination of the optimum elution
volume.
Candidates for 2016 Method of the Year
243