Detection Methods for Ethylene Oxide (EtO)

Ethylene Oxide (EtO) is a highly volatile and carcinogenic compound used in sterilization and fumigation processes. Due to its health risks, detecting its presence in food and environmental samples is crucial. The Food Safety and Standards Authority of India (FSSAI) has approved a method for detecting EtO and its reaction product, 2-Chloroethanol (2-CE), using advanced analytical techniques.

1. Gas Chromatography Tandem Mass Spectrometry (GC-MS/MS)

Principle:

• Gas Chromatography (GC) separates EtO from other sample components.

• Tandem Mass Spectrometry (MS/MS) detects and quantifies EtO and 2-CE using characteristic fragmentation patterns.

Variations in GC-MS/MS Techniques:

• Headspace (HS)-GC-MS/MS: Detects volatile compounds by analyzing gas phase above the sample.

• PTV or MMI-GC-MS/MS: Uses specialized injection techniques to enhance sensitivity.

• HS-Trap GC-MS/MS: Combines headspace analysis with a trapping system to increase detection limits.

Advantages:

✔ High sensitivity and specificity✔ Capable of analyzing complex food matrices✔ Simultaneous detection of both EtO and 2-CE

2. Headspace Gas Chromatography (HS-GC-MS)

Principle:

• Sample is incubated at a controlled temperature to allow EtO to evaporate.

• The gaseous phase is injected into the GC-MS system for separation and analysis.

• Mass spectrometry detects the ions of EtO and 2-CE and quantifies their concentrations.

Advantages:

✔ Minimizes sample handling and contamination✔ Ideal for highly volatile compounds✔ No need for extensive sample preparation

3. Multi-Step Enrichment Headspace-Trap GC-MS

Principle:

• Uses syringe-based headspace sampling combined with cryogen-free trapping to concentrate EtO.

• Multiple headspace injections ensure higher sensitivity.

• The enriched sample is analyzed via GC-MS.

Advantages:

✔ Extremely high sensitivity (detects very low concentrations)✔ Suitable for various food matrices, including spices and cereals✔ Avoids solvent interferences

4. QuEChERS Extraction with GC-MS/MS

Principle:

• Quick, Easy, Cheap, Effective, Rugged, and Safe (QuEChERS) extraction isolates EtO and 2-CE from food samples.

• The extract is analyzed using GC-MS/MS.

Advantages:

✔ Effective for diverse food matrices✔ Provides clean extracts with minimal sample degradation✔ Validated according to international quality standards

Key Considerations in EtO Analysis

• Storage Conditions: Samples should be kept at ≤ 10°C to prevent volatilization.

• Use of Internal Standards: 1,2-Dichloroethane-d4 is used as a reference for precise quantification.

• Calibration Standards: EtO concentrations are measured against matrix-matched standards to ensure accuracy.

• Quality Control: Methods follow SANTE/11312/2021 guidelines to ensure reproducibility.