Rapid Analysis of Sudan and Other Prohibited Dyes in Chili Powder Using the ACQUITY UPLC H-Class System with Xevo TQD

Importance of the Topic

Sudan and related azo dyes are unauthorized colorants with carcinogenic potential. Their illicit use in spices such as chili powder poses a serious public health risk. Regulatory bodies including the European Union require routine monitoring. A robust analytical workflow that balances high throughput sensitivity and selectivity is essential to safeguard consumer safety and enforce legislation.

Goals and Overview of the Study

This study aims to develop and demonstrate a rapid method for simultaneous extraction separation and quantification of eleven prohibited dyes in chili and related spice matrices. The objectives include streamlining sample preparation using QuEChERS achieving chromatographic separation within twelve minutes and implementing tandem mass spectrometry for reliable detection at low micrograms per kilogram levels. The method is validated across a set of real commercial spice powders to assess performance under practical conditions.

Methodology and Instrumentation Used

Sample Preparation

• QuEChERS extraction based on CEN method 15662 combining water acetonitrile and buffered salts for dispersive cleanup
• Centrifugation and direct transfer of supernatant for analysis

UPLC Conditions

• ACQUITY UPLC H-Class System with BEH C18 2.1 x 100 mm column at 45 C
• Binary gradient of water methanol and acetonitrile each containing 0.1 percent formic acid
• Flow rate 0.4 mL per minute injection volume 5 microL total run time twelve minutes

MS/MS Detection

• Xevo TQD mass spectrometer in positive electrospray ionization
• Multiple reaction monitoring with optimized cone voltages and collision energies for each analyte
• Data acquisition and automated quantification with TargetLynx Application Manager

Main Results and Discussion

The optimized UPLC MS/MS method achieved baseline separation of all eleven dye compounds in under twelve minutes. Calibration curves in solvent and matrix matched standards exhibited excellent linearity with correlation coefficients above 0.99 across the range equivalent to 2.5 to 640 micrograms per kilogram of spice. Recoveries in a blank chili powder matrix at 10 micrograms per kilogram (50 micrograms per kilogram for Para red) ranged from 60 to 95 percent requiring potential use of internal standards to correct for matrix losses. Matrix effect evaluation revealed both ion suppression and enhancement across compounds highlighting the importance of matrix matching or isotopic internal standards. Analysis of fourteen commercial spice samples identified six positive instances of prohibited dyes with concentrations up to 276 micrograms per kilogram in curry powder. Sudan I Sudan II Rhodamine B and Sudan orange G were among the most frequently detected analytes. Automated flagging in TargetLynx provided rapid screening of non compliant samples by comparing ion ratios and action limits in accordance with EU Commission Decision 2002/657/EC criteria.

Benefits and Practical Applications

• High throughput analysis suitable for routine quality control in food testing laboratories
• Sensitivity down to low micrograms per kilogram levels to meet regulatory requirements
• Comprehensive coverage of multiple prohibited dyes in a single run
• Automated data processing and reporting reduces manual review and potential errors

Future Trends and Potential Applications

The approach can be extended to other spice matrices and fortified food products. Incorporation of isotopically labeled standards will further improve quantification accuracy. Integration with high resolution mass spectrometry platforms and ambient ionization techniques may enable even faster screening. Advances in miniaturized sample preparation and online cleanup could reduce solvent consumption and analysis time while maintaining performance.

Conclusion

A rapid robust UPLC MS/MS workflow combining QuEChERS extraction and tandem mass spectrometry has been developed for the determination of eleven Sudan and azo dyes in chili powder and related spices. The method demonstrates high selectivity sensitivity and throughput necessary for routine surveillance. Automated data processing ensures efficient identification of non compliant samples in line with regulatory standards.

Reference

1. European Commission Rapid Alert System for Food and Feed RASFF cases on Sudan dyes detected in food products 2011 and 2012
2. Waters DisQuE QuEChERS sample preparation method CEN method 15662 application note ID 720003048EN