Determination of Carbendazim and Benomyl Residues in Oranges and Orange Juice by Automated Online Sample Preparation Using TLX-LC-MS/MS

Significance of the Topic

The determination of carbendazim and its precursor benomyl in citrus products is critical for food safety monitoring and compliance with regulatory limits. These fungicides, widely used in agriculture, can pose health risks if residues exceed maximum residue limits. An efficient, automated analytical method ensures reliable detection at sub-ppm levels, supports regulatory controls, and addresses growing demands for high-throughput food quality testing.

Objectives and Study Overview

This study presents a rapid, automated online sample preparation method coupled with turbulent flow LC-MS/MS for the quantification of carbendazim and benomyl residues in oranges and orange juice. The primary goals were to achieve a limit of quantification below 0.01 mg/kg, demonstrate method performance suitable for routine regulatory analysis, and compare the approach with conventional QuEChERS workflows.

Methodology

Sample Preparation and Cleanup

• Sample homogenization or direct juice use followed by pH adjustment to 7 with ammonium hydroxide.
• Addition of internal standard (d4-imidacloprid) and methanol extraction.
• Centrifugation and transfer of supernatant into vials for automated cleanup.

Chromatographic Separation

• Online coupling of Thermo Scientific Transcend TLX turbulent flow column for matrix cleanup.
• Transfer of trapped analytes to Hypersil GOLD analytical column for gradient elution within a 13-minute cycle.

Mass Spectrometric Detection

• TSQ Quantum Access MAX triple quadrupole operated in positive SRM mode.
• Quantification based on internal standard calibration and characteristic ion transitions.

Used Instrumentation

• Thermo Scientific Transcend TLX-1 system
• TSQ Quantum Access MAX triple quadrupole mass spectrometer
• Turboflow Cyclone MCX-2 turbulent flow column
• Hypersil GOLD analytical column with guard

Main Results and Discussion

Linearity and Calibration

• Matrix-matched calibration from 0.005 to 0.1 mg/kg with R² > 0.99.
• Average response factor determined as 3.2.

Accuracy and Precision

• Recoveries ranged 96–115% at three spike levels (0.005, 0.01 and 0.05 mg/kg).
• Repeatability and intermediate precision below 10% RSD.

Detection Limits and Matrix Effects

• Method LOD and LOQ of 0.00015 and 0.0005 mg/kg, respectively.
• Minimal matrix effects (< 20% deviation in solvent vs matrix slopes).

Survey Analysis

• No residues exceeded 0.01 mg/kg in six commercial orange and juice samples.

Benefits and Practical Applications

The automated TLX-LC-MS/MS workflow offers high-throughput capacity of over 100 samples per day, reduced manual labor, and robust cleanup of complex citrus matrices. Its low detection limits and validated performance make it suitable for routine food safety monitoring by regulatory and industrial laboratories.

Future Trends and Opportunities

Expanding this approach to a broader range of pesticides and complex food matrices can further streamline multi-residue analysis. Advances in online sample preparation and high-resolution mass spectrometry may enhance sensitivity and selectivity, meeting future regulatory demands and large-scale monitoring programs.

Conclusion

The developed automated method using Transcend TLX and turbulent flow LC-MS/MS provides a rapid, sensitive, and reliable solution for quantifying carbendazim and benomyl in orange products. Its validated performance supports compliance with international MRLs and offers a compelling alternative to traditional sample preparation techniques.

References

1. Community Rapid Alert System for Food and Feed (RASFF) database.
2. FDA Compliance Policy for Carbendazim in Orange Juice.
3. Thermo Scientific Application Note Method 522133.
4. EU Commission Directive 2002/63/EC.
5. AOAC Single-Laboratory Validation Guidelines.
6. Harmonized Guidelines for Single-Laboratory Validation of Methods.
7. European Commission Pesticide Residue Public Database.
8. Mallat E., Barcelo D., Tauler R., Chromatographia 46 (1997) 342-350.