Improved sample preparation and HPLC/MS determination of pesticides in fat-rich vegetables
Frank Michel, Emily Barrey, Katherine Stenerson, Olga Shimelis, Michael Ye
[email protected] Food Science Asia 2015, 27.10.2015, Singapore
1
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
QuEChERS Method: dispersive SPE
Quick Easy Cheap Effective Rugged Safe
• M. Anastassiades, S. Lehotay et al., J. AOAC Int. 86 (2) 412-431, 2003 • EN 15662:2008 • AOAC Official Method 2007.01 2
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
QuEChERS Method: dispersive SPE
LCGC North America, Jul 1, 2010 © 2015 Sigma-Aldrich Co. LLC. All rights reserved.
3
Literature values for extraction of problematic pesticides from fatty matrices (more then 5% fat) Matrix
Cleanup used
Avocado
QuEChERS, PSA/C18
Avocado
GPC, extraction using ethyl acetatecyclohexane
Dough Flaxseeds
• •
QuEChERS, PSA/C18
Hexachlorobenzene
Endosulfan Reference
27%
90%
S. Lehotay & K. Mastovska 2005
-
78%
J.L.F. Moreno et al, 2006
52%
112%
25%
73%
U. Koesukwiwat et al., 2010
Fat from matrix interferes with analysis Non-polar pesticides are compromised when removing fat
4
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
QuEChERS Method: Removing fat out of fat-rich samples • Fat-containing samples: – Freezing out of the fat – C18 SPE adsorbent removing the fat – Zirconia-coated silica: Supel QuE Z-Sep/C18 (Discovery® DSC-18 + Z-Sep) for samples containing 15% fat
5
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
New adsorbents for fat removal Zirconiacoated silica
Phospholipids
6
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
Proposed Retention Mechanism O O O O O O
Triglyceride H3C H3C
O
H3C
O
H3C
(CH2)n
CH3
(CH2)n (CH2)n
R2 R3
CH3 CH3
R1 O HO Zr
H3C
H3C
C18
O
O Zr
Z-Sep
H3C
R1
O
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
Monoglyceride HO
7
Supel QuE Z-Sep/C18 and Z-Sep+ adsorbents • Supel QuE Z-Sep/C18 • Mixture of Discovery DSC-18 and Z-Sep (zirconia-coated silica)
Zr Zr Zr Zr + Zr Zr Zr Zr
C18 C18 C18 C18 C18 C18 C18 C18
• Supel QuE Z-Sep+ • Dual Bonded C18 and zirconia coated on silica
Zr C18 Zr C18 Zr C18 Zr C18 8
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
Comparison of sorbents for retention of model fatty compounds
25 mg of sorbent mixed with 1 mL of olein solution in acetonitrile (400mg/mL total concentration).
100% 90% 80% 70% 60% 50% 40% 30%
The remaining oleins were quantified by LCELSD and retention was calculated.
20% 10% 0% Z-Sep+
Z-Sep/C18
C18
PSA
NH2
alumina
LRA
Silica
9
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
Comparison of different adsorbents: Removal of oleins monoolein dioleins triolein 100% 80% 60% 40% 20% 0% Z-Sep+
Z-Sep
C-18
Z-Sep/C18 mix 10
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
Pesticides in oranges by LC/MS • Similar recoveries for most pesticides • Better recoveries for some pesticides using Z-Sep/C18
Z-Sep/C18 PSA/C18 PSA
% recovery
– Anilazine, Naled, Edifenphos
120 100 80 60 40 20 0
11
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
Pesticides in Apples by LC/MS Pesticides from green apples at 50 ppb
Comparison between Z-Sep/C18 cleanup and PSA cleanup
PSA
recovery
2 sets of compounds: organophosphates, carbamates, benzoylurea classes
Z-Sep/C18 150%
Z-Sep/C18
PSA
100% 50% 0% 1 2 3 4 5 6 7 8 9 10 33 11 34 12 35 13 14 15
compound number
Green Apple
Pesticides from green apples at 10 ppb
• Conclusion • Better overall green and yellow pigment removal by Z-Sep/C18 than PSA • Comparable recovery in most cases.
reco very
Z-Sep/C18 140% 120% 100% 80% 60% 40% 20% 0%
PSA
17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 compound number 12
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
Pesticides in olives by LC/MS • 38 pesticides tested in canned olives • Comparison of PSA, C18/PSA and Z-Sep/C18 • Recoveries for most pesticides are similar between 2 methods • Better recoveries for some pesticides observed using ZSep/C18 • Fenhexamide (no ion suppression for Z-Sep/C18)
140.0
C18/PSA 120.0
Z-Sep/C18
100.0
PSA
80.0 60.0 40.0
• Anilazine (no ion suppression for Z-Sep/C18) • Sethoxydim (no ion suppression for Z-Sep/C18)
20.0 0.0 FENHEXAMID
ANILAZINE
SETHOXYDIM
SPIROMESIFEN
PHOSMET
13
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
QuEChERS: Pesticides in avocado by GC-MS • Pesticide mix included hydrophobic compounds (e.g.organochlorines, hexachlorbenzene) and some other more polar classes all GC-MS amenable • QuEChERS extraction was performed – Ratio 3 g sample to 25 mL acetonitrile extraction solvent improved recoveries – Tested cleanup used no adsorbent, PSA/C18 and Z-Sep+
Extraction and Cleanup Procedures
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
QuEChERS: Pesticides in avocado by GC-MS Results: Matrix Removal Amount of Matrix Residue Remaining
•
Gravimetric Analysis: level of matrix residue in supernatants
•
Z-Sep+ removes more matrix than PSA/C18
15
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
QuEChERS: Pesticides in avocado by GC-MS Results: Pigment Removal •
Less color remained in the extract cleaned with Z-Sep+.
•
Z-Sep+ has a greater capacity for pigment removal than PSA/C18.
16
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
QuEChERS: Pesticides in avocado by GC-MS Results: Chromatography •
Much less background is observed in the TIC of the Z-Sep+ cleaned extract than the uncleaned extract.
•
Indicates that Z-Sep+ can be effectively used to reduce unwanted matrix background in difficult samples such as avocado.
GC-MS TICs in full scan mode of uncleaned and Z-Sep+ cleaned avocado extracts
17
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
QuEChERS: Pesticides in avocado by GC-MS Results: Pesticide Recovery
•
Z-Sep+ showed better recovery overall.
•
PSA/C18: matrix interference prevented analysis of cyfluthrin, cypermethrin and deltametrin.
•
Z-Sep+ showed better reproducibility than PSA/C18
18
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
Summary • Fat is challenging for the analysis of non-polar pesticides using QuEChERS approach • Usual approaches to remove fat such as freezing out or adding C18 SPE adsorbent are insufficient for non-polar pesticides • Zirconia-coated silica (Z-Sep/C18 and Z-Sep+) is a targeted approach for fat removal not compromizing non-polar pesticides – Z-Sep/C18 and Z-Sep+ cleanup more effective for removing fatty matrix interference (80 % of oleins retained by Z-Sep/C18, nearly 100% retained by Z-Sep+) than C18 – Similar recovery for most pesticides, higher for certain ones – Less retention of hydrophobic pesticides on Z-Sep/Z-Sep+ – Better removal of matrix components
• Viable alternative to PSA/C18 adsorbents or freezing out in current methods 19
© 2015 Sigma-Aldrich Co. LLC. All rights reserved.
Thank you for your attention!
20 © 2015 Sigma-Aldrich Co. LLC. All rights reserved.
