Determination of Aromatic Amines from Azo Colorants According to DIN EN ISO 17234-1

Importance of the topic

Aromatic amines released from azo dye cleavage pose significant health risks due to their potential carcinogenicity. Regulatory standards such as DIN EN ISO 17234-1 mandate sensitive analytical methods for detecting trace levels of these compounds in consumer goods. Robust, automated workflows enhance safety monitoring and reduce manual effort in routine laboratory operations.

Objectives and study overview

This application note evaluates an automated sample preparation protocol combined with two analytical detection strategies—HPLC-MS/MS and HPLC-UV—for the determination of 23 targeted aromatic amines generated by reductive cleavage of azo dyes. The study compares two solid-phase extraction phases: CHROMABOND® XTR (alkaline activated) and conventional Kieselguhr.

Methodology and instrumentation

Leather and textile samples were degreased with n-hexane, extracted with citrate buffer at 70 °C, and reductively cleaved using sodium dithionite. Extracts underwent liquid-liquid partitioning with methyl tert-butyl ether and SPE cleanup. Automated processing was achieved on the FREESTYLE system equipped with SPE and EVAporation modules, coupled to direct HPLC injection.

Instrumentation

• FREESTYLE BASIC robot with SPE and EVAporation modules
• Agilent 1290 Infinity II HPLC system with API 5500 Triple Quad MS/MS (ESI, MRM)
• Thermo Ultimate 3000 HPLC-UV system (detection at 240, 280, 305 nm)
• CHROMABOND® XTR, alkaline activated cartridges (15 mL, 3 g) and Kieselguhr phase cartridges

Main results and discussion

HPLC-MS/MS with CHROMABOND® XTR yielded average recoveries between 78 % and 132 % for all 23 amines, exceeding ISO requirements and showing standard deviations < 10 % for all but one analyte. The Kieselguhr phase produced lower and more variable recoveries, with several compounds falling below the 70 % threshold. Chromatograms demonstrated clear separation of all targets within a 14 min gradient run.

Benefits and practical applications

The automated SPE-EVAporation workflow minimizes hands-on time, reduces manual errors, and enables unattended overnight operation. The CHROMABOND® XTR cartridge simplifies sample cleanup by eliminating manual activation steps. Integration with direct HPLC injection streamlines the entire analytical pipeline, making the method suitable for QA/QC laboratories in textile, leather, and consumer goods testing.

Future trends and opportunities

Advancements may include coupling to high-resolution mass spectrometry for broader compound screening, miniaturized SPE formats for lower solvent consumption, and integration of machine-learning algorithms for automated data interpretation. Expanding the method to cover additional degradation products could further strengthen consumer safety assessments.

Conclusion

The combination of automated SPE-EVAporation on the FREESTYLE platform with CHROMABOND® XTR cartridges and HPLC-MS/MS detection delivers a high-throughput, reliable method for quantifying aromatic amines from azo dyes. The approach surpasses DIN EN ISO 17234-1 performance criteria and offers significant operational efficiencies for routine analytical laboratories.

Reference

DIN EN ISO 17234-1:2015. Determination of aromatic amines derived from azo colorants.