As the number of U.S. states allowing the adult use of cannabis and cannabis products increases, so does the need for product testing before retail sale. States that have legalized recreational use have specified testing requirements for pesticide residues in cannabis flower and cannabis products. Because the specific pesticides and action levels vary from state to state, a comprehensive approach to residue analysis can meet the requirements of multiple U.S. state regulations with a single analysis. The challenge of quantifying pesticide residues in cannabis is complex because of the high concentration of cannabinoids and terpenes relative to the levels of pesticides that may be present. Here we present a straightforward acetonitrile extraction using a solid-phase extraction (SPE) cartridge and targeted dispersive solid-phase extraction (dSPE) cleanup. The final dilute extract is analyzed with both gas chromatography–tandem mass spectrometry (GC–MS/MS) and liquid chromatography–tandem mass spectrometry (LC–MS/MS) for a comprehensive target list (200+ compounds) that encompasses those identified on individual U.S. state lists. Limits of quantitation meet or exceed individual U.S. state requirements.
The LC–MS/MS extracts were prepared for analysis using only a final dilution step. This final dilution of 50 µL of extract into 950 µL of acetonitrile results in a factor of 20x. Combined with the initial extraction, this dilution leads to an overall dilution factor of 500x. Again, the background matrix is reduced (Figure 1) and the recoveries for target pesticides are within acceptance limits of 70–120%.
Each cannabis strain will have its own unique profile of cannabinoids and terpenes, which may present different background interferences affecting pesticide determinations. If dilution alone does not adequately address matrix effects or interferences, there is an optional dSPE cleanup that can be applied to extracts for LC–MS/MS analysis. This optional step uses PSA as a sorbent. PSA is a powerful cleanup sorbent for challenging samples, but may result in low recoveries for some compounds. Recoveries for spinotoram, spinosad, spirotetramat, and spiroxamine can be reduced to <50% when subjected to a dispersive technique containing PSA, and daminozide is unrecoverable.
A concept developed by Schenck and Wong (9) to address the loss of planar pesticides when using graphitized carbon as a cleanup sorbent can be applied to compounds strongly retained by PSA. To improve recoveries of planar pesticides when using graphitized carbon, toluene (25–30%) is added to sample extract to push planar pesticides off the sorbent. When using a dSPE designed for fatty samples (50 mg of PSA, 50 mg of C18, 150 mg of magnesium sulfate), a similar approach can be used with methanol. Adding methanol at the dSPE step will improve the recoveries for compounds retained by PSA. The data indicate that 10–20% methanol addition will increase recoveries for the problematic compounds (Figure 2). (See upper right for Figure 2, click to enlage; Figure 2: To increase the recovery for compounds retained by PSA using a dSPE cleanup methanol can be added to sample extract in the cleanup tube. The chart shows the percent recovery as a function of methanol added.) This approach will improve recoveries to acceptable levels with minimal matrix returning to the extract. Keep in mind that no amount of methanol will recover daminozide from PSA, and typical reversed-phase chromatography is not optimal for measuring daminozide residues. A more appropriate technique would be an extraction optimized for polar compounds and using hydrophilic-interaction chromatography (HILIC) chromatographic techniques (10).
Results and Discussion
A solvent extraction with a pass through solid-phase cleanup gives an extract with less coextracted materials when compared to extracts from a QuEChERS technique. Having a single extraction procedure that can be split to both MS/MS platforms streamlines the workflow, resulting in higher sample throughput. Dispersive cleanup and dilution techniques can be optimized for each analytical technique and compound list to give the necessary precision and accuracy. The cleanup and dilution procedure for GC–MS/MS analysis allows the laboratory to place the sample extract into a solvent that is more amenable to GC (hexane–acetone). An optional dispersive cleanup procedure for LC–MS/MS analysis can be used if adverse matrix effects or interferences are encountered.
This procedure was validated for 215 pesticides (Table I) split between the two MS/MS platforms. (See upper right for Table I, click to enlage; Table I: Comprehensive pesticide target list: GC–MS/MS [bold] LC–MS/MS [italic].) A set of five replicates was prepared at the limit of quantitation (LOQ) for each compound. The signal-to-noise ratio (S/N) criterion of 10:1 (quantitation ion) was met for each compound at the LOQ. For the GC–MS/MS-amenable compounds, 72 of the 74 pesticides had recoveries of 70–120%, and the percent relative standard deviation (%RSD) was less than 15% for all 74 compounds. For compounds analyzed by LC–MS/MS, all 141 pesticides had recoveries of 70–120%, and the %RSD was below 15% for 138 of 141. The LOQ was validated at 0.1 mg/kg for all compounds except those listed in Table II. (See upper right for Table II, click to enlage; Table II: Compounds with LOQs above 0.1 mg/kg.)
- Oregon Administrative Rules 333-007-0400.
- California Code of Regulations, Title 16, Division 42. Bureau of Cannabis Control, Chapter 11, § 5719.
- Washington Administrative Code 246-70-050.
- D.G. Farrer, “Technical report: Oregon Health Authority’s Process to Decide Which Types of Contaminants to Test for in Cannabis” (Oregon Health Authority, 2015).
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- CSN EN 15662, Foods of Plant Origin - Determination of Pesticide Residues Using GC-MS and/or LC-MS/MS Following Acetonitrile Extraction/Partitioning and Clean-Up by Dispersive SPE - QuEChERS-Method.
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- F.J. Schenck and J.W. Wong in Analysis of Pesticides in Food and Environmental Samples, J.L. Tadeo, Ed. (CRC Press Inc., Boca Raton, Florida, 2008), Chapter 6.
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Rick Jordan is the Laboratory Manager at Pacific Agricultural Laboratory in Sherwood, Oregon. Daniel Miller is the Technical Director at Pacific Agricultural Laboratory. Lilly Asanuma is a chemist at Pacific Agricultural Laboratory. Anthony Macherone is a Senior Scientist with Agilent Technologies and a visiting professor at The Johns Hopkins University School of Medicine. Direct correspondence to: [email protected]
How to Cite This Article
R. Jordan, L. Asanuma, D. Miller, and A. Macherone, Cannabis Science and Technology 1(2), 26-31 (2018).