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Increase Removal of Fat and Pigment from Avocado Extracts Prior to GC-MS Analysis of Pesticide and Metabolite Residues

By: Katherine K. Stenerson and Jennifer Claus, Reporter US Volume 31.2

The health benefits of avocado are associated with its high content of healthy fat, fiber, vitamins, and minerals. Avocado consumption in the United States has steadily grown, with sources of the fruit being both domestic and imported1. The fat content of avocado is typically in the range of 10-15%, and this can pose a special analytical challenge when performing pesticide residue analysis. If not removed, fat compounds can interfere with analysis, resulting in elevated detection limits and contamination of LC and GC systems. The same issues can result from pigments; therefore, the effective removal of these compounds is also critical.

QuEChERS Cleanup Sorbents

QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe) has become a very popular method for the extraction and cleanup of fruit and vegetable samples prior to pesticide residue analysis2-4. The extraction step uses a salt (such as magnesium sulfate) to drive compounds into acetonitrile, added as the extraction solvent. Cleanup is then accomplished using dispersive SPE (dSPE) with sorbents that include:

  • PSA (primary-secondary amine) – for removal of polar pigments, sugars, and organic acids
  • C18 – for removal of lipids and non-polar components
  • Graphitized carbon black – for removal of chlorophyll and carotenoids

It is common to use blends of cleanup sorbents based on the interferences that need to be removed. Due to the fat content of avocado, a PSA/C18 blend is appropriate for extracts intended for pesticide analysis.

Recently, a new cleanup sorbent, Supel™ QuE Z-Sep+, was developed. This sorbent consists of both C18 and zirconia bonded to the same silica particles. The C18 binds fats through hydrophobic interaction, while the zirconia acts as a Lewis acid, attracting compounds with electron donating groups; such as the hydroxyl (-OH) groups in mono and diglycerides.


In this work, Z-Sep+ is compared to a PSA/C18 blend for effectiveness in the QuEChERS cleanup of avocado extracts prior to pesticide residue analysis by GC-MS. Extraction and cleanup procedures are summarized in Table 1. Multiple replicates of both unspiked and spiked avocado samples were processed. Spiked samples were prepared with a mixture of hydrophobic and polar pesticides, each at 20 ng/g. All extracts were analyzed by GC-MS using large volume injection (LVI). Quantitation was done using a calibration curve prepared with matrix-matched standards.

Table 1. Extraction and Cleanup Procedures

  1. Place 3 g of a homogenized avocado sample into a 50 mL centrifuge tube (Product No. 55248-U). Add spike solution if a spiked replicate.
  2. Add 25 mL of acetonitrile (Product No. 34481), and shake for one minute.
  3. Add the contents of an Acetate Extraction Tube (Product No. 55234-U), and shake for one minute.
  4. Centrifuge for five minutes.
  5. Transfer 3 mL of the supernatant into the appropriate cleanup tube, Z-Sep+ (Product No. 55296-U) or PSA/C18 (Product No. 55229-U).
  6. Shake for one minute, then centrifuge for three minutes.
  7. Transfer 1 mL of the supernatant into an autosampler vial for GC-MS analysis.

Matrix Removal

Gravimetric analysis was performed on avocado extracts, cleaned and uncleaned, to determine the level of matrix residue remaining in the supernatants. This data is displayed in Figure 1. The results clearly show that significantly less residue remains in cleaned extracts, and Z-Sep+ sorbent removes more matrix than PSA/C18.

Comparison of matrix residue remaining

Figure 1. Comparison of Matrix Residue Remaining

Pigment Removal

Extracts were visually compared for color remaining after. As shown in Figure 2, less color remained in the extract cleaned with Z-Sep+. This indicates Z-Sep+ has a greater capacity for pigment removal than PSA/C18.

Z-Sep+ vs. PSA/C18

Figure 2. Avocado Extracts after Cleanup


Figure 3 shows GC-MS total ion chromatograms (TICs) in full scan mode of uncleaned and Z-Sep+ cleaned avocado extracts. Much less background is observed in the TIC of the Z-Sep+ cleaned extract than the uncleaned extract. This indicates that Z-Sep+ can be effectively used to reduce unwanted matrix background in difficult samples such as avocado.

Z-sep+ cleaned avocado extract vs. uncleaned avocado extract

Figure 3. GC-MS Chromatograms (same y-axis)


column: SLB®-5ms, 20 m x 0.18 mm I.D., 0.36 µm (Product No. 28576-U); oven: 70 °C (2 min), 15 °C/min to 325 °C (6 min); inj. temp.: Programmed, 25 °C (0.89 min), 600 °C/min to 325 °C (5 min); detector: MS Scan mode; carrier gas: helium, 1 mL/min constant; injection: 10 µL LVI, PTV solvent vent, rapid injection speed, split vent; flow: 100 mL/min (5 psi) until 0.28 min, 60 mL/min at 2.78 min; liner: 4 mm I.D., split type, wool packed FocusLiner™ with single taper design

Pesticide Recovery

Use of a cleanup sorbent should not adversely affect the recoveries of the analytes of interest. This was studied for both Z-Sep+ and PSA/C18 by determining the average recovery and reproducibility of 3 replicate spiked avocado samples cleaned with each sorbent. The results are depicted in Figure 4, showing average % recovery and the recovery range obtained for each pesticide. Recovery was better for most pesticides using Z-Sep+. In the case of PSA/C18, matrix interference prevented analysis of cyfluthrin, cypermethrin, and deltametrin. Reproducibility obtained with Z-Sep + cleanup was also better than PSA/C18, as indicated by the tighter recovery ranges indicated in Figure 4.

Z-sep+ cleanup vs. PSA/C18 cleanup average recovery and range

Figure 4. Average Recovery and Range (n=3)


The data presented here indicates that Z-Sep+ sorbent can be effectively used to remove fat and pigment interferences from avocado, a high fat matrix. It was found to remove more matrix from avocado extracts than traditional PSA/C18 sorbent, and produce a lower background when analyzed using GC-MS. Recoveries of pesticides of polar and lipophilic character were good, indicating no adsorption problems for these analytes.

Legal Information

SLB is a registered trademark of Sigma-Aldrich Co. LLC
Supel is a trademark of Sigma-Aldrich Co. LLC
FocusLiner is a trademark of SGE Analytical Science Pty Ltd.




  1. Carman, Hoy F.; Li, Lan; Sexton, Richard J.; An Economic Evaluation of the Hass Avocado Promotion Order's First Five Years. Gianni Foundation Research Report 351, Univ. of California Agriculture and Natural Resources. December 2009.
  2. Lehotay, S. J. Quick, Easy, Cheap, Effective, Rugged and Safe (QuEChERS) Approach for the Determination of Pesticide Residues. Proceedings AOAC Annual Meeting, St. Louis, MO USA, 2004.
  3. AOAC Official Method 2007.01, Pesticide Residues in Foods by Acetonitrile Extraction and Partitioning with Magnesium Sulfate.
  4. EN15662:2008, Foods of plant origin – Determination of pesticide residues using GC-MS and/or LC-MS/MS following acetonitrile extraction/partitioning and cleanup by dispersive SPE – QuEChERS-method.