contaminants in oils and fats: analysis and...
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AOCS-CCOA Joint Symposium on Functional Lipids Shanghai, China on 19-20 November 2014
Florence LACOSTE ITERG (Pessac, France)
f.lacoste@iterg.com
Contaminants in oils and fats: analysis and regulations
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Guillaume
CHANTRE
Technical
Industrial
Center
French
Ministry
of Industry
80
Staff
6 M€
(turnover)
ISO 9001
ISO 17025
Presentation of ITERG
2
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Polycyclic aromatic hydrocarbons
Mineral oil
Phthalates
3-MCPD esters & glycidol esters
Contents
3
Origins ?
Regulation ?
Recognized analytical methods ?
Risks ?
Risks Methods Regulation
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Polycyclic aromatic
hydrocarbons
4
Environmental contamination
Production process
Regulation
(EC) n°1881/2006 – contaminants in foodstuffs
Risks Methods Regulation
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
PAHs origins in vegetable oils
PAHs
Extraction
v
Oilseeds Crude oil
+
HAP
Active carbon
Heavy PAHs
Light PAHs
Deodorisation
Drying
Refined oil
(+ e HAP)
5
* Grapeseed oil * Coconut oil * Pomace olive oil * Sunflower oil
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014 6
benzo(b)fluoranthene benzo(k)fluoranthene benzo(a)pyrene indeno(1,2,3-cd)pyrene
dibenz(ah)anthracene benzo(ghi)perylene
HEAVY PAHs
naphthalene
acenaphthene acenaphthylene fluorene phenanthrene anthracene
fluoranthene pyrene benz(a)anthracene chrysene
LIGHT PAHs
Structure of PAHs
GT
GT
GT GT GT
GT : genotoxic
GT GT
GT
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
PAHs regulation: (EC) n°1881/2006
contaminants in foodstuffs
PAHs Maximum
levels oils and fats (µg/kg)
Maximum levels coconut
oil (µg/kg)
Maximum levels cocoa
butter (µg/kg)
Sum of 4 PAHs: - benzo[a]pyrene - benz[a]anthracene - benzo[b]fluoranthene - chrysene
10,0 20,0
35,0 from 1.4.2013 until 31.3.2015
30,0 from 1.4.2015
benzo[a]pyrene 2,0 2,0 5,0
from 1.4.2013
7
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
PAHs: determination methods
8
Method ISO 15302 ISO 15753 ISO 22959
BaP LOQ (µg/kg) 0,1 0,2 0,1
Reproducibility (CVR%) 27 % (2,1µg/kg) 41% (3,21µg/kg) 10 % (2,6µg/kg)
Principle
PAH isolation: liquid chromatography (alumina, silica gel,C18-silica gel) or HPLC (donor-acceptor complex chromatography, size-exclusion chromatography)
Analysis: HPLC/fluorescence or GC/MS
ISO 15302 : benzo[a]pyrene in oils (LC + HPLC/FLD)
ISO 15753 : 12 PAHs in oils (2 SPE + HPLC/FLD)
ISO 22959 : 17 PAHs in oils (DACC on-line + HPLC/FLD)
EN 16619 : 4 PAHs in foodstuffs (SEC + SPE + GC/MS)
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
0
10
20
30
40
50
60
70
80
90
100
<0,5 0,5 - 1 1 - 2 2 - 5 5 - 20 20 - 50 50 - 100 >100
% s
amp
les
Benzo[a]pyrene (µg/kg)
Virgin olive oil
Olive oil
Pomace olive oil
Grapeseed oil
Sunflower oil
EU limit
2 µg/kg
BaP : occurrence in vegetable oils
9
EU Scoop Task (Oct 2004)
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014 10
Edible oils: PAH4 Levels
Screening 2012 : 21 vegetable oils
PAH4 en µg/kg
benzo(a)pyrene chrysene benz(a)anthracene benzo(b)fluoranthene
ITERG data (2012)
0
2
4
6
8
10
12EU limit PAH4 : 10 µg/kg
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Mineral oil
11
Environmental contamination (air, soil)
Crop protection
Transport & storage
Production process
Regulation
(EC) n°1151/2009 - import of sunflower oil from Ukraine
EFSA Scientific Opinion on Mineral Oil Hydrocarbons in food (2012)
Risks Methods Regulation
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
EFSA Scientific Opinion, 2012
Mineral oil: complex mixture of hydrocarbons
MOSH: straight or branched alkanes & alkylated cycloalkanes
MOAH: aromatic hydrocarbons including alkyl-substituted
Hydrocarbon compound number in mineral oil > 100 000 for those with less than 20 carbon atoms !
Different products & composition: diesel fuel, white oil, lubricant …
Technical grade mineral oil contain 15-35 % MOAH, which is minimised in food grade MOSH (white oils)
Mineral oil composition
12
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Minutes
0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30
Millivo
lts
50
100
150
200
250
300
350
400
TRACE GC-Channel 1
sample 7
Name
13
Minutes
0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30
Millivo
lts
50
100
150
200
250
300
350
400
TRACE GC-Channel 1
sample 7
Name
Natural hydrocarbons
Mineral oil : unresolved complex mixture
EI
Mineral oil detected in vegetable oils
2008 contamination of sunflower oil from Ukraine with a mineral oil from unknown origin
2009 contamination of walnut oil with a food grade lubricant oil during refining process
2010 identification of compounds eluted as mineral oil in grapeseed oils
2011 contamination of milk fat with a food grade lubricant oil during production
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Fractionation of the sample by liquid chromatography on
silica gel impregnated with AgNO3
Quantification with an internal standard (C18 or C20)
GC/FID analysis on an short apolar column
MOSH determination: pr ISO17780
14
18
,5 g
Sili
ca g
el
Elution with hexane Solvent evaporation
Vide
40°C
GC analysis
On-column injection (2 µl) DB5 HT (15 m-0,25 mm-0,1 µm)
1 g oil
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
pr ISO 17780 validation
15
Mineral oil – 50 mg/kg spiked olive oil sample 2-2013 sample 4-2014
Spiking level (mg/kg) 50 50
Mean (m) 52 52
Repeatability limit (r) 19,3 15,3
Reproducibility limit (R) 51,7 28,1
Horrat value 4,1 2,2
0
20
40
60
80
100
120
140
160
26 3 8 23 21 34 15 31 1 19 4 33 36 9 28 17 30 27 29 24 11 10 35 5 32 12 6 22 2 18 20 14
Lab
me
an v
alu
e (
mg/
kg)
Laboratories
Sample 2: 50 mg/kg spiked olive oil
0
20
40
60
80
100
120
140
160
20 12 14 10 7 9 13 16 15 1 11 19 4 8 5 2 6 17
Lab
me
an v
alu
e (
mg/
kg)
Laboratories
Sample 4: 50 mg/kg spiked virgin olive oil
Method applicable from 50 to 1000 mg/kg
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Phthalates
16
Transport & storage
Production process
Regulation
EFSA Scientific Opinion on food additives, flavourings, processing aids and materials in contact with food (2005)
(EC) n°10/2011 - plastic materials and articles intended to come into contact with food
Risks Methods Regulation
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Chemical structures of phthalates
Di-methyl PHT DMP R1=R2=methyl
Di-ethyl PHT DEP R1=R2=ethyl
Di-isobutyl PHT DIBP R1=R2=isobutyl
Di-butyl PHT DBP R1=R2=butyl
Di-hexyl PHT DHexP R1=R2=hexyl
Benzyl butyl PHT BBP R1=benzyl R1=butyl
Di-n-heptyl PHT DHepP R1=R2=heptyl
Di-(2-ethyl hexyl) PHT DEHP R1=R2=ethyl-2 hexyl
Di-n-octyl PHT DNOP R1=R2=octyl
Di-isononyl PHT DINP R1=R2= isononyl
Di-isodecyl PHT DIDP R1=R2=isodecyl
17
C
C
O
O
O
OR1
R2
Oily visquous liquid
MW: 390,6 g/mol
BP: 385°C
Water solubility: 3 µg/l
High affinity for fat (log Kow : 7,5)
DEHP
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Flooring-Roofing- Wall covering Adhesives-Sealant-Rubber
Paints-Shower curtains Wires & cables-Fresheners
Cosmetics : Perfume Hairspray-Deodorant
Skin emollient-nail polish fingernail elongators
Car undercoating Dashboard-Door
panels-Safety glass
Shoes-Boots-Gloves Out-door & rainwear
Toys Child-care articles
Pharmaceuticals Medical devices :
Catheters-Blood bag
Phthalates are everywhere
18
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Specific migration limit in food
To be used as
BBP 30 mg/kg Plasticizer in single-use material containing non-fatty foods except infant formulae
DINP ∑ = 9 mg/kg
DIDP
DEHP 1,5 mg/kg Plasticizer in repeated use materials containing non-fatty foods DBP 0,3 mg/kg
(EC) n°10/2011 - Plastic materials &
articles into contact with food
19
Material containing these phthalates cannot be used for oils & fats
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014 20
Phthalates: ITERG’s procedure
Advantages of the method :
No contamination, no solvent
Rapid analysis (45 min)
Sensitive LOQ < 0,1 mg/kg (excepted DINP & DIDP)
HS SPME
1 g oil
Analysis
DB5ms (30 m; 0,25 mm; 0,25 µm)
PDMS
m/z 111 (adipates)
m/z 149 (phthalates) GC/MS
DEHP DHexP
DNPP DBP
DEP
DMP
7.5 10.0 12.5 15.0 17.5 20.0 22.5 25.0 minutes
0
1
2
3
4
5 50:500
DNOP
25 26 27 28 29 30 minute
s
0
50
100
150
200 kCounts
DINP
DIDP
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
0
1
2
3
4
543
DEHP in oils & fats
ITERG data, 2009
mg/kg
21
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
3-MCPD esters & glycidol esters
22
Production process (deodorisation)
Regulation for free 3-MCPD
(EC) n°1881/2006 – contaminants in foodstuffs (20 µg/kg in soya sauce)
Regulation for 3-MCPD esters
EFSA statement – related to 3-MCPD esters (2008)
EFSA Scientific Report on 3MCPD occurrence in food in Europe (2013)
Risks Methods Regulation
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
H2C
H2C
HC
O
AG
H2C
H2C
HC
O
OH
H2C
H2C
HC
OH
OH
Cl
H2C
H2C
CH
AG
AG
Cl
H2C
H2C
CH HO
Cl
AG
« free » 3-MCPD
Glycidol
3-MCPD monoesters « bound »
Glycidol esters (GEs)
3-MCPD diesters « bound »
49 7 14
3-MCPD esters & glycidol esters Structure
23
Hrncirik, OVID meeting, 2011
7 fatty acids
C12 - C14 C16 - C18
C18:1 - C18:2 C18:3
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
3-MCPD esters & glycidol esters Analysis strategy
24
Direct methods Indirect methods
Pros No chemical reaction Individual quantification of species
Standards available Good sensitivity
Cons Standards not available Expensive tools: HPLC/MS/MS Purification step needed
Reactivity of the species Global quantification of species
Standards AOCS-JOCS Cd 28-10 for GEs From Masukawa et al., JAOCS, 2010
AOCS Cd 29a-13 for 2- and 3-MCPD & GEs From Ermacora & Hrnirick, JAOCS, 2012
AOCS Cd 29b-13 for 2- and 3-MCPD & GEs From Kuhlmann, EJLST, 2011
AOCS Cd 29c-13 for 3-MCPD & GEs From DGF C-VI 18, 2010 & Weisshar, 2008
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014 25
Esters de 3-MCPD & glycidol
• Comparable results for the 3 indirect methods • In general, direct methods agreed with indirect methods • Methods did not give reliable results for concentrations < 1 mg/kg
2013 AOCS Collaborative Study
• Palm oil sample, not spiked. • All indirect methods were acceptable
• RBD Canola sample, not spiked. • Variable results between methods
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Occurrence of 3-MCPD & glycidol esters in
vegetable oils
26
Screening ITERG 2013
0.0
1.0
2.0
3.0
4.0
5.0
6.0
7.0
8.0
9.0
10.0
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. t
ou
rnes
ol
H. t
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H. t
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H. t
ou
rnes
ol
H. t
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H. t
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H. t
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ol
H. t
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ol
H. t
ou
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ol b
rute
H. t
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ol r
affi
née
H. p
épin
s d
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H. p
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H. p
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isin
H. p
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s d
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isin
H. p
épin
s d
e ra
isin
3MCPD (mg/kg)
sunflower
grapeseed
0.0
0.2
0.4
0.6
0.8
1.0
1.2
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. c
olz
a
H. t
ou
rnes
ol
H. t
ou
rnes
ol
H. t
ou
rnes
ol
H. t
ou
rnes
ol
H. t
ou
rnes
ol
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rnes
ol
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ou
rnes
ol
H. t
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rnes
ol
H. t
ou
rnes
ol
H. t
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H. t
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rnes
ol
H. t
ou
rnes
ol
H. t
ou
rnes
ol b
rute
H. t
ou
rnes
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affi
née
H. p
épin
s d
e ra
isin
H. p
épin
s d
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isin
H. p
épin
s d
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isin
H. p
épin
s d
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isin
H. p
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isin
H. p
épin
s d
e ra
isin
H. p
épin
s d
e ra
isin
H. p
épin
s d
e ra
isin
GLYCIDOL (mg/kg)
sunflower
rapeseed
grapeseed
rapeseed
Industrial specidifcation 3MCPD = 2 mg/kg Industrial specification glycidol = as low as possible
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Regulation, Methods, Risks
27
PAHs
Mineral oil
Phthalates
3-MCPD esters & glycidyl esters Risks Methods Regulation
Risks Methods Regulation
Risks Methods Regulation
Risks Methods Regulation
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014
Conclusions
• Research of contaminants is part of multiple controls conducted by fat and oil industry to fulfill the EC regulation n°1881/2006.
• In the absence of regulation, the detection of contaminants must be addressed in partnership with authorities according to the toxicity of molecules.
• Risks are rather limited due to the efficient elimination during oil-refining steps
• However some contaminants can be formed during the production process of vegetable oils such as 3-MCPD & glycidyl esters.
28
AOCS-CCOA Joint Symposium on Functional Lipids, Shanghai, Nov 2014 29
A special thanks to….
ITERG Analysis Department team
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