Extraction of Planar Pesticides From Spinach and Fresh Oregano Using QuEChERS Methodology With a Novel Carbon Adsorbent

Although vital to plant life, the presence of chlorophyll in GC/MS analysis is highly problematic. A pigment of high molecular weight, it quickly causes fouling of injection liners and GC columns due to buildup at the injection port and GC column head. To preserve column lifetime, chlorophyll must be removed prior to GC/MS analysis.

Graphitized carbon black (GCB) has a planar, graphitic surface, allowing for removal of nearly 100% of chlorophyll from green samples. Unfortunately, when analyzing planar aromatic compounds such as hexachlorobenzene and chlorothalonil in green samples, poor analyte recovery results. Interaction between the planar structure of the GCB and these planar pesticides causes unwanted retention on the carbon sorbent.

A QuEChERS (quick, easy, cheap, effective, rugged and safe) cleanup method has been developed that uses Supel QuE Verde (MilliporeSigma, the life science business of Merck KGaA, Darmstadt, Germany, operating as MilliporeSigma in the U.S. and Canada), a sorbent combination containing Z-Sep+, primary-secondary amine (PSA) and an improved GCB. It provides improved recovery of planar pesticides while maintaining sufficient color removal in high-chlorophyll matrices. This article describes GC/MS analysis of select planar pesticides from spinach and oregano extracts.

Experimental

QuEChERS extraction and cleanup was performed in a manner similar to that outlined in the AOAC International Official Method 2007.01.1 Organic spinach and freshly picked oregano were extracted with acetonitrile. Figure 1 and Table 1 list the extraction and cleanup procedure for use with 2-mL tubes. When using 15-mL tubes, 8 mL of acetonitrile extract was transferred to the cleanup tubes. Acetonitrile extract was spiked with a mixture composed primarily of planar pesticides in acetonitrile to a concentration of 50 ng/mL. Samples were spiked after extraction to study the performance of the cleanup sorbents in relation to the planar pesticides. Extractions were performed in triplicate. Analysis of the final extracts was done by GC/MS using the conditions listed in Table 1.

Figure 1 – Chromatograms of planar pesticides in a) spinach and b) fresh oregano after cleanup with Supel QuE Verde. Peaks: 1) 2,6-dichlorobenzonitrile, 2) diphenylamine, 3) hexachlorobenzene, 4) pentachloronitrobenzene, 5) terbuthylazine, 6) chlorothalonil, 7) metribuzin, 8) dacthal, 9) folpet, 10) gamma-chlordane, 11) alpha-chlordane, 12) coumaphos. See Table 1 for conditions.
Table 1 – Planar pesticides cleanup conditions using Supel QuE Verde

Results and discussion

Background was evaluated by analyzing the final extracts using GC/MS in selected ion mode (SIM); chromatograms of the spiked extracts are shown in Figure 1. In the analysis of the planar pesticides in SIM, Supel QuE Verde cleanup removed enough matrix interferences to easily identify and quantitate all 12 residues in a mixture containing planar pesticides (Figure 1).

Figure 2 shows the spinach extract before cleanup, and the spinach and oregano extracts after cleanup with Supel QuE Verde. Total chlorophyll removal was evaluated using a spectrophotometer measuring absorbance at 664, 647 and 630 nm; the processed extracts were compared to the initial acetonitrile extract of each plant material (Figure 3). In all cases, chlorophyll removal was 95% or greater.

Figure 2 – QuEChERS extracts of a) spinach extract before cleanup, b) spinach extract after cleanup with Supel QuE Verde and c) oregano extract after cleanup with Supel QuE Verde.
Figure 3 – Percent total chlorophyll removal from spinach and oregano extracts by Supel QuE Verde.

Average analyte recoveries obtained from 50 ng/mL spiked spinach and oregano extracts using 2-mL Supel QuE Verde cleanup tubes are given in Figure 4. Recoveries were in the range 70–120% for all the planar pesticides tested in both matrices. Reproducibility, determined as %RSD for n = 3 spiked replicates, was very good for both matrices. Each pesticide exhibited a %RSD value less than 5%, with the exception of chlorothalonil, which had a %RSD value of 20%.

Figure 4 – Average percent recoveries from spinach and oregano extracts spiked at 50 ng/mL after cleanup with the 2-mL Supel QuE Verde tube (n = 3). Note: Recoveries with traditional carbon sorbents are typically below 70%.

Conclusion

Most food analysts agree that pesticide residue recovery should be between 70% and 120%, with chlorophyll removal 95% or greater. Because of the strong interaction between the planar pesticides and planar surfaces of GCB, a compromise is made between color removal and analyte recovery, especially regarding the most planar pesticides such as hexachlorobenzene and chlorothalonil. Supel QuE Verde combines a novel carbon with zirconia-coated silica (Z-Sep+) to provide an optimal balance between planar pesticide recovery and color removal. QuEChERS cleanup using Supel QuE Verde can be used prior to GC/MS analysis to provide sufficient chlorophyll removal while maintaining excellent recovery of planar pesticides from spinach and oregano matrices.

Reference

  1. AOAC Official Method 2007.01. Pesticide residues in foods by acetonitrile extraction and partitioning with magnesium sulfate. J. AOAC Int. 2007, 90, 485.

Michael Ye is R&D sample preparation manager, MilliporeSigma. Jennifer Claus is SPE product manager, MilliporeSigma, 595 N. Harrison Rd., Bellefonte, Penn. 16823, U.S.A.; tel.: 814-359-5407; e-mail: [email protected]www.emdgroup.com

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