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High-Quality Analysis of Pesticides in Cannabis Using QuEChERS, Cartridge SPE Cleanup, and GCxGC-TOFMS

    • Quickly and effectively extract medical marijuana samples for pesticide analysis.
    • Cartridge SPE cleanups of dirty extracts improve GC inlet and column lifetimes.
    • Selective GC columns increase accuracy of pesticide determinations for complex samples.

Over 20 states in the U.S. have legalized the use of recreational or medical cannabis because of therapeutic benefits for ailments such as cancer, multiple sclerosis, and ALS. Dosing methods include smoking or vaporizing and baked goods. Unlike other prescribed medicines regulated by U.S. FDA, marijuana is a Schedule 1 drug and is illegal on the federal level. As a result, medical cannabis patients have no safety assurances for their medication, which could contain harmful levels of pesticide residues. Currently, medical marijuana pesticide residue analysis methods are poorly defined and challenging to develop due to matrix complexity and a long list of potential target analytes.

In order to address matrix complexity, we combined a simple QuEChERS extraction approach with cartridge SPE (cSPE) cleanup, followed by GCxGC-TOFMS. Acceptable recoveries were obtained for most pesticides, and incurred pesticide residues were detected in some of the illicit marijuana samples used for method development.

QuEChERS Extraction Saves Time and Reduces Hazardous Solvent Use
Trace residue extraction procedures from dry materials like medical cannabis typically involve large amounts of solvent, long extraction times, and tedious concentration steps similar to the Soxhlet procedure or multiresidue methods from the Pesticide Analytical Manual. QuEChERS, with its simple 10 mL acetonitrile shake extraction and extract partitioning with salts and centrifugation, offers time savings, glassware use reduction, and lower solvent consumption.

Water was added to finely ground, dry cannabis samples to increase QuEChERS extraction efficiency, especially for more polar pesticides. A vortex mixer was used to shake the solvent and sample for at least 30 minutes prior to extract partitioning. When finished, it was easy to transfer the supernatant from the QuEChERS extraction tube for subsequent cSPE cleanup prior to analysis with GC or LC (Figure 1).

Figure 1: A quick and easy QuEChERS extraction, combined with cSPE, effectively prepared extracts for pesticide residue analysis from highly complex marijuana samples.

Post-centrifugation QuEChERS extracts

A. Post-centrifugation QuEChERS extracts

QuEChERS extracts loaded on SPE cartridge

B. QuEChERS extracts loaded on SPE cartridge

Final extract

C. Final extract


Cartridge SPE Cleanup Improves GC Inlet Uptime
Injecting chlorophyll-laden extracts into a GC gives reduced recoveries for less volatile pesticides, and results in degradation of sensitive pesticides like DDT and Dicofol (Table I). SPE cleanup with a 500 mg graphitized carbon black/500 mg PSA cartridge removes chlorophyll and traps fatty acids that interfere with qualitative pesticide identification and bias quantification. cSPE has increased sorbent capacity over dispersive SPE for thorough cleanup of complex extracts.

Table I:  Pesticide recoveries for a QuEChERS extract of cannabis give higher results when cSPE is used for cleanup. Dicofol and DDT are degraded in the inlet for the dirtier extract, yielding high DDD results.

Pesticide Classification With cSPE
Cleanup (%)
Without cSPE
Cleanup (%)
4,4´-DDD Organochlorine 83 230
4,4´-DDT Organochlorine 77 9
Bifenthrin Pyrethroid 86 89
Dicofol Organochlorine 84 ND
Azinphos methyl Organophosphorus 79 53
trans-Permethrin Organochlorine 68 17
Pyraclostrobin Strobilurin 73 19
Fluvalinate Pyrethroid 72 23
Difenoconazole Triazole 67 21
Deltamethrin Pyrethroid 68 20
Azoxystrobin Strobilurin 72 27

Orthogonal GC Columns Greatly Increase Separation Power for More Accurate Pesticide Results
GCxGC is a powerful multidimensional approach that gives two independent separations in one instrumental analysis. An Rxi®-5Sil MS and Rtx®-200 column combination distributes pesticides broadly in both dimensions, providing a highly orthogonal GCxGC system. More important though is separating pesticides from potential isobaric matrix interferences, as seen in the surface plot for the insecticide cypermethrin (Figure 2). Cypermethrin gas chromatographs as four isomers, and all would have experienced qualitative interference and quantitative bias from peaks in the foreground of the surface plot had only 1-dimensional GC been used. With GCxGC-TOFMS, cypermethrin was unequivocally identified in a marijuana sample at a low ppm level (Figure 3).

Figure 2:  GCxGC-TOFMS and orthogonal Rxi®-5Sil MS and Rtx®-200 columns allow incurred cypermethrins in a marijuana extract to be separated from interferences (m/z 163 quantification ion). (View larger)

PeakstR 1 (sec)tR 2 (sec)
1.Cypermethrin 12,2921.50
2.Cypermethrin 22,3041.54
3.Cypermethrin 32,3101.53
4.Cypermethrin 42,3131.58
Incurred Cypermethrins in a QuEChERS Extract of Marijuana by GCxGC on Rxi®-5Sil MS and Rtx®-200
GC_FF1204
ColumnRxi®-5Sil MS 30 m, 0.25 mm ID, 0.25 µm (cat.# 13623)
Rtx®-200 1.3 m, 0.25 mm ID, 0.25 µm (cat.# 15020)
Sample
Diluent:Toluene
Injection
Inj. Vol.:1 µL splitless (hold 1 min)
Liner:Premium 4 mm single taper w/wool (cat.# 23303.1)
Inj. Temp.:250 °C
Purge Flow:40 mL/min
Oven
Oven Temp.:Rxi®-5Sil MS: 80 °C (hold 1 min) to 310 °C at 5 °C/min
Rtx®-200: 85 °C (hold 1 min) to 315 °C at 5 °C/min
Carrier GasHe, corrected constant flow (2 mL/min)
Modulation
Modulator Temp. Offset:20 °C
Second Dimension Separation Time:3 sec
Hot Pulse Time:0.9 sec
Cool Time between Stages:0.6 sec
DetectorTOFMS
Transfer Line Temp.:290 °C
Analyzer Type:TOF
Source Temp.:225 °C
Electron Energy:70 eV
Mass Defect:-20 mu/100 u
Solvent Delay Time:5 min
Tune Type:PFTBA
Ionization Mode:EI
Acquisition Range:45-550 amu
Spectral Acquisition Rate:100 spectra/sec
InstrumentLECO Pegasus® 4D GCxGC-TOFMS
NotesRtx®-200 (cat.# 15020) is a 15 m column. A 1.3 m section was used here as the second dimension column.

Figure 3:  Positive mass spectral identification of incurred cypermethrin in illicit marijuana. (View larger)

PeakstR 1 (sec)tR 2 (sec)
1.Cypermethrin 22,3041.54
Sample and Reference Mass Spectra for Incurred Cypermethrin in a QuEChERS Extract of Marijuana by GCxGC on Rxi®-5Sil MS and Rtx®-200
GC_FF1206

See Figure 2 for instrument conditions.

Summary
QuEChERS and cSPE produced usable extracts from highly complex cannabis samples for high-quality pesticide residue analysis. The multidimensional separation power of GCxGC-TOFMS was then used to correctly identify and quantify pesticides in these complex extracts.

For more on medical cannabis analysis, visit www.restek.com/medical-cannabis

Acknowledgment
Randy Hoffman, a Police Evidence Technician at The Pennsylvania State University (PSU), supplied the seized marijuana samples while overseeing their handling. Frank Dorman at PSU assisted with QuEChERS extractions.

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