EVALUATION OF NOVEL DISPERSIVE SPE AND PASS-THROUGH SPE OPTIONS FOR CLEANUP OF HIGH CHLOROPHYLL SAMPLES AFTER QUECHERS EXTRACTION

Importance of the Topic

Leafy vegetables like spinach often contain high levels of chlorophyll and related pigments that co-extract during QuEChERS pesticide sample preparation. These pigments can interfere with chromatography and mass spectrometry, leading to instrument contamination and inaccurate quantitation. Effective cleanup strategies are essential for reliable pesticide analysis in food safety laboratories.

Objectives and Study Overview

This study evaluated and compared three cleanup approaches for QuEChERS spinach extracts:

• Dispersive-SPE (dSPE) with 10 mg graphitized carbon black (GCB) per mL extract
• dSPE with 50 mg GCB per mL extract
• Pass-through SPE using Oasis PRiME HLB cartridges

Key goals were to assess pigment removal efficiency (chlorophylls, lutein, carotenes) via UPLC-PDA and to measure recoveries of three planar pesticides (cyprodinil, chlorothalonil, thiabendazole) by APGC-MS/MS and UPLC-MS/MS.

Methodology and Instrumentation

Sample preparation began with a standard AOAC QuEChERS extraction of homogenized spinach spiked at 50 µg/kg. After centrifugation, aliquots underwent either dSPE cleanup or pass-through SPE:

• dSPE: 2 mL extract treated with 150 mg Na₂SO₄, 50 mg C18, 50 mg PSA, plus either 10 mg or 50 mg GCB; shaken and centrifuged.
• Oasis PRiME HLB: 0.8 mL extract discarded, then 1.5 mL passed through 150 mg HLB cartridge under low vacuum; no conditioning required.

Instrumental analysis:

• UPLC-PDA on Waters Acquity I-Class with Cortecs UPLC T3 column at 450 nm for pigment profiling.
• APGC-MS/MS on Waters Xevo TQ-S for GC-amenable pesticides (Agilent 7890 GC with Restek Rxi-5ms column).
• UPLC-MS/MS on Waters Xevo TQ-S micro with BEH C18 column for LC-amenable pesticides.

Main Results and Discussion

All three cleanup protocols removed the majority of chlorophylls and carotenes. Specific findings:

• Oasis PRiME HLB removed >99 % of chlorophyll and >95 % of lutein with minimal pesticide losses.
• dSPE with 10 mg GCB achieved moderate pigment removal and negligible loss of planar pesticides.
• dSPE with 50 mg GCB completely eliminated pigments but caused significant losses of cyprodinil, chlorothalonil, and thiabendazole.

Pass-through cleanup generated cleaner extracts without conditioning steps, and different cartridge formats (Plus Short, Vac, Plus Light) showed equivalent performance.

Benefits and Practical Applications

• Oasis PRiME HLB cartridges streamline cleanup workflows, reduce solvent usage, and protect analytical systems.
• dSPE with reduced GCB load balances pigment removal and analyte retention for laboratories seeking minimal losses.
• The evaluated methods support high-throughput pesticide screening in QA/QC and research settings.

Future Trends and Applications

Advances may include:

• Broader adoption of pass-through SPE in automated and on-line cleanup platforms.
• Extension of PRiME HLB cleanup to other challenging matrices (e.g., herbs, teas, leafy greens).
• Development of mixed-mode sorbents tailored to remove diverse interferences.
• Miniaturized and high-throughput cartridge formats compatible with robotic liquid handlers.

Conclusion

Pass-through SPE using Oasis PRiME HLB cartridges offers superior pigment removal and preserves recoveries of sensitive planar pesticides in spinach QuEChERS extracts. While high GCB loads in dSPE can eliminate pigments, they risk significant analyte losses. Laboratories should select cleanup strategies that balance extract cleanliness and target compound integrity.