© 2009 PerkinElmer© 2009 PerkinElmer© 2009 PerkinElmer© 2012 PerkinElmer
The Determination of Benzene, Toluene, Ethyl
Benzene, Xylenes and Styrene in Olive Oil using HS Extraction and GC/MS
Andrew TiplerChromatography R&D Manager
PerkinElmer Inc., Shelton, CT
Olive Oil
Olive oil is produced by grinding olives and extracting the oil by mechanical (pressing) or chemical (solvent extraction) means.
Olive oil is composed mainly of the mixed triglyceride esters of oleic acid and palmitic acid and of other fatty acids, along with traces of squalene and sterols.
It is commonly used in cooking, cosmetics, pharmaceuticals, and soaps and as a fuel.
Olive Oil Production
Country Production in tons (2009)
Production % (2009)
Consumption (2005)
Annual per capita
consumption (kg)
World 2,907,985 100% 100% 0.4
Spain 1,199,200 41.2% 20% 13.6
Italy 587,700 20.2% 30% 12.4
Greece 332,600 11.4% 9% 23.7
Syria 168,163 5.8% 3% 7.0
Tunisia 150,000 5.2% 2% 11.1
Turkey 143,600 4.9% 2% 1.2
Morocco 95,300 3.3% 2% 1.8
Portugal 53,300 1.8% 2% 7.1
France 6,300 0.2% 4% 1.3
United States 2,700 0.1% 8% 0.6
Others 169,122 5.8% 18% 1.2
Source: Wikipedia.com
Effects of Air Pollution
Olive trees are good at absorbing air pollutants and passing them on to the olives and thus into the oil
“Concentrations in olive oil were found to be around several tens of µg/kg for benzene; a few hundred µg/kg for toluene and many hundreds of µg/kg for the sum of ethyl benzene and xylenes. Substantial concentrations of styrene were found which however was probably of natural origin. The situation was considered to be sufficiently serious that a definition of provisional limits for these components was proposed (benzene: 50µg/kg; toluene: 180µg/kg; ethyl benzene: 50µg/kg; xylenes: 280µg/kg). However, these were not imposed as a consequence of disagreement in the European Union” . Maurus Biedermann, Konrad Krob, Gianni Morchio, Z Lebensm Unters Forsch (1995) 200; 266-272.
Despite the strong concerns, even today, there still appears to be no agreement on the acceptable limits of BTEXS in olive oil.
On 10 June 2011, the US National Toxicology Program has described styrene as "reasonably anticipated to be a human carcinogen".
Acceptable limits for BTEXS in olive oil
5.05 5.55 6.05 6.55 7.05 7.55 8.05 8.55 9.05 9.55 10.05Time0
100
%
Scan EI+ TIC
1.17e89.92
8.85
7.94
6.63
5.24
5.004.92
4.62
4.72
6.49
8.07
Ben
zen
e
Tolu
ene
p-X
ylen
em
-Xyl
en
e
Eth
ylb
enze
ne
o-X
ylen
e
Sty
ren
e
BTEXS (~0.6ng) separation using GC/MS
30m x 0.25mm x 1.0µm Elite-Wax35(1)-10-130(0)
Sampling Olive Oil
Olive oil is a heavy sample matrix and is not very GC-friendly. Direct injection will cause problems with breakdown and contamination.
Some form of extraction technique is required to separate the BTEXS from the sample matrix prior to introduction into the GC.
Don’t do this in your lab!
Headspace Sampling
Headspace sampling provides a very easy way to extract the volatile BTEXS compounds from the olive oil matrix.
10g of sample are placed in a vial which is then sealed with a cap and maintained at 90°C for 20 minutes. The volatile components (including BTEXS) will partition into the vapor phase
0.15mL of the vapor phase is transferred to the GC column for analysis.
Analytical System
Clarus SQ8 GC/MS TurboMatrix HS Sampler
Pressure-Balanced Headspace Sampling
Pressurization Sampling
V1 V1
Equilibration
V1
PHS
V2
Valve closed for just a few seconds
Olive oil sample
5.05 5.55 6.05 6.55 7.05 7.55 8.05 8.55 9.05 9.55 10.05Time0
100
%
1: Scan EI+ TIC
1.17e8
4.62
4.914.63
6.57
5.70
6.295.91
6.06
7.21
7.97 8.34
8.21 10.43
8.91
Eth
ano
lB
enze
ne
Tolu
ene
p-X
ylen
em
-Xyl
en
e
Eth
ylb
enze
ne
o-X
ylen
e
Sty
ren
e
Total Ion Chromatogram from Spiked Olive Oil
10g olive oil spiked with 17ng/g BTEXS
30m x 0.25mm x 1.0µm Elite-Wax35(1)-10-130(0)
5.10 5.60 6.10 6.60 7.10 7.60 8.10 8.60 9.10 9.60 10.10Time0
100
%
0
100
%
0
100
%
0
100
%
2.61e49.93
9.38e38.17
8.077.94 8.86
4.80e46.62
6.95e45.24
Be
nze
ne
Tolu
en
e
p-X
yle
ne
m-X
yle
ne
Eth
ylb
en
zen
e
o-X
yle
ne
Sty
ren
e
SIM @ 78 Daltons
SIM @ 92 Daltons
SIM @ 106 Daltons
SIM @ 104 Daltons
Single Ion Monitoring Improves Selectivity and Detectability
SiFi Enables ‘Simultaneous’ Full and Single Ion Data collection
5.10 5.60 6.10 6.60 7.10 7.60 8.10 8.60 9.10 9.60 10.10Time0
100
%
OliveOilAStd_060
9.95
Ben
zen
e @
78
Da
Tolu
ene
@ 9
2 D
a
p-X
ylen
e @
106
Da
m-X
yle
ne
@ 1
06 D
a
Eth
ylb
enze
ne
@ 1
06 D
a
o-X
ylen
e @
106
Da
Sty
ren
e @
104
Da
SIM of Olive oil spiked with 17ng/g BTEXS
y = 178.38x - 60.006R² = 0.9998
0
2000
4000
6000
8000
10000
12000
14000
16000
18000
0 10 20 30 40 50 60 70 80 90 100
Peak
Are
a (C
ount
s)
Concentration (ppb w/w)
Extracted Calibration Plot for Benzene
y = 12.021x - 3.8872R² = 0.9997
0
200
400
600
800
1000
1200
0 10 20 30 40 50 60 70 80 90 100
Peak
Are
a (C
ount
s)
Concentration (ppb w/w)
Extracted Calibration Plot for Styrene
Quantitative Performance
Compound RSD (% at 45ng/g, n=10)
r2
(from 4.5 to 90ng/g)Limit of Detection
(ng/g)Benzene 1.7 0.9998 0.1
Toluene 3.8 0.9986 0.2
Ethylbenzene 2.3 0.9995 0.3
p-Xylene 3.5 0.9997 0.3
m-Xylene 3.7 0.9998 0.3
o-Xylene 3.3 0.9995 0.3
Styrene 3.5 0.9997 0.3
Standard mixtures prepared in 10g ‘clean’ olive oil
5.10 5.60 6.10 6.60 7.10 7.60 8.10 8.60 9.10 9.60 10.10Time0
100
%
0
100
%
0
100
%
0
100
%
TIC1.84e3
S/N:RMS=23.81
9.809.7110.03
10.17
TIC2.47e3
S/N:RMS=39.67
7.93
7.807.738.06
8.85
8.348.57 8.939.05
TIC1.10e4
S/N:RMS=71.15
TIC3.90e3
S/N:RMS=14.84
5.015.42
Benzene, 0.9ng/g
Toluene, 5.9ng/g
m-Xylene, 5.4ng/g
Styrene, 3.1ng/g
Low-Level Determination of BTEXS in Olive Oil
Background from ‘Clean’ Olive Oil
5.10 5.60 6.10 6.60 7.10 7.60 8.10 8.60 9.10 9.60 10.10 Time0
100
%
OliveOilA_061
5.10 5.60 6.10 6.60 7.10 7.60 8.10 8.60 9.10 9.60 10.10 Time0
100
%
OliveOilAStd_060
9.95
Ben
zen
e @
78
Da
Tolu
ene
@ 9
2 D
a
p-X
ylen
e @
106
Da
m-X
yle
ne
@ 1
06 D
a
Eth
ylb
enze
ne
@ 1
06 D
a
o-X
ylen
e @
106
Da
Sty
ren
e @
104
Da
‘Clean’ Olive Oil
‘Clean’ Olive Oil spiked with 17ng/g BTEXS
BTEXS in Olive Oil from Local Supermarket (ng/g)
Sample Source Benzene
Toluene
Ethylbenzene
p-Xylene
m-Xylene
o-Xylene
Styrene
California 0.89 5.86 1.66 1.45 5.24 3.77 3.07
Italy, Greece, Spain Tunisia 2.86 27.55 6.12 5.86 16.73 8.75 41.34
Italy, Spain, Greece, Tunisia 3.07 24.22 13.47 7.85 23.64 13.97 39.59
Italy, Spain, Tunisia, Turkey, Argentina 2.99 17.03 3.74 3.44 9.35 6.14 40.09
Spain, Argentina 2.43 34.99 7.22 7.42 18.97 10.65 126.11
Italy, Spain, Greece, Tunisia, Morocco, Syria, Turkey 4.09 35.71 19.13 17.10 59.31 28.10 61.05
Italy, Greece, Spain, Tunisia 1.25 2.79 ND 1.80 3.74 3.17 7.39
The sensitivity of the MS detector in SIM mode enables detection of low levels of BTEXS with standard headspace sampling. Detection limits of 0.1 to 0.3ng/g have been demonstrated which compare favorably against those in other published papers (~5ng/g).
The HS sampling system simplifies sample preparation and eliminates the effects from a heavy sample matrix entering the chromatographic column and the MS detector.
Good quantitative performance has been shown.
Significant levels of BTEXS were found in olive oils purchased at a local (Connecticut) supermarket.
Conclusions