ISO 7124:2023

Eggs and egg products - Determination of fipronil and metabolites residues - Liquid chromatography tandem mass spectrometry method

This document specifies a liquid chromatography tandem mass spectrometry method (LC-MS/MS) for the determination of fipronil and metabolites (including fipronil-desulfinyl, fipronil-sulfide and fipronil-sulfone) residues in eggs and egg products.

Œufs et ovoproduits — Dosage des résidus de fipronil et de ses métabolites — Méthode de chromatographie en phase liquide couplée à la spectrométrie de masse en tandem

General Information

• Status: Published
• Publication Date: 12-Apr-2023
• ICS: 67.120.20 - Poultry and eggs
• Technical Committee: ISO/TC 34/SC 6 - Meat, poultry, fish, eggs and their products
• Drafting Committee: ISO/TC 34/SC 6 - Meat, poultry, fish, eggs and their products
• Current Stage: 6060 - International Standard published
• Start Date: 13-Apr-2023
• Due Date: 04-Jun-2024
• Completion Date: 13-Apr-2023

Overview

ISO 7124:2023 specifies a validated liquid chromatography–tandem mass spectrometry (LC‑MS/MS) method for the determination of fipronil and its metabolites in eggs and egg products. The method covers target analytes including fipronil, fipronil‑desulfinyl, fipronil‑sulfide and fipronil‑sulfone, and defines sample preparation, cleanup, chromatographic and mass spectrometric conditions, identification criteria and quantification using an isotope internal standard (fipronil‑C6).

Key Topics and Technical Requirements

• Scope: Determination of fipronil and metabolites residues in eggs and egg products using LC‑MS/MS.
• Sample handling: Homogenization of representative samples (start from ≥1 kg), preparation instructions for fresh eggs and egg products, and storage at −20 °C to −16 °C.
• Extraction: Acetonitrile extraction with addition of salts (sodium chloride, anhydrous sodium sulfate) and internal standard.
• Cleanup: Dispersive solid‑phase extraction (d‑SPE) using PSA, C18 and anhydrous magnesium sulfate prior to filtration (0.22 µm).
• Chromatography: Phenyl column (2.1 × 100 mm, 1.7 µm or equivalent), gradient elution between ammonium acetate/formic acid aqueous phase and methanol, 35 °C column temperature, 0.4 ml/min flow, 2 µl injection.
• Mass spectrometry: Electrospray ionization in negative polarity, multiple reaction monitoring (MRM) transitions specified for each analyte, with MS source and gas settings provided.
• Quantification & ID criteria: Internal standard calibration (fipronil‑C6), retention time match tolerance ±2.5%, confirmation by relative ion transition intensities, reporting in µg/kg.
• Standards & reagents: Reference to analytical grade solvents and reagents; standard materials with purity ≥95%; preparation and stability timelines for stock and working solutions.
• Performance: Procedures for blank tests, determination, calculation, limit of quantification and precision (annexes include chromatograms and precision data).

Applications

• Regulatory monitoring of pesticide residues in eggs and egg products for food safety and compliance with maximum residue limits (MRLs).
• Routine quality control testing by food laboratories, contract testing services and egg producers’ QC departments.
• Surveillance and investigative testing after suspected contamination incidents involving fipronil.
• Method transfer, validation and interlaboratory studies in analytical chemistry and food safety research.

Who Should Use This Standard

• Food testing and accredited analytical laboratories performing LC‑MS/MS residue analysis.
• National food safety and regulatory authorities enforcing residue limits.
• Egg and egg‑product manufacturers and processors for internal monitoring.
• Instrument vendors and method developers implementing validated analytical workflows.

Related Standards

• ISO 3696 - Water for analytical laboratory use (referenced for grade‑1 water).
• CAC/GL 50‑2004 - Recommended sampling guidance for foods (sampling recommendations referenced; sampling itself is not part of ISO 7124:2023).

For implementation, obtain the full ISO 7124:2023 standard from ISO or your national standards body to access complete procedural details, tables (MRM transitions, gradients), annex figures and validation data.

Standards Content (Sample)

INTERNATIONAL ISO
STANDARD 7124
First edition
2023-04
Eggs and egg products —
Determination of fipronil and
metabolites residues — Liquid
chromatography tandem mass
spectrometry method
Œufs et ovoproduits — Dosage des résidus de fipronil et de ses
métabolites — Méthode de chromatographie en phase liquide couplée
à la spectrométrie de masse en tandem
Reference number
© ISO 2023
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
be reproduced or utilized otherwise in any form or by any means, electronic or mechanical, including photocopying, or posting on
the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address below
or ISO’s member body in the country of the requester.
ISO copyright office
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CH-1214 Vernier, Geneva
Phone: [phone]
Email: [e-mail]
Website: www.iso.org
Published in Switzerland
ii
Contents Page
Foreword .iv
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Principle . 1
5 Sampling . 1
6 Preparing and storing the test sample . 2
6.1 Preparing . 2
6.1.1 Fresh eggs and egg products with shell. 2
6.1.2 Other egg products . 2
6.2 Storing . 2
7 Liquid chromatography tandem mass spectrometry method . 2
7.1 Reagents . 2
7.2 Preparation of solutions . 2
7.3 Standard materials . 3
7.4 Preparation of standard solutions . 3
7.5 Materials . 3
7.6 Apparatus . 3
7.7 Procedure . 4
7.7.1 General . 4
7.7.2 Extraction . 4
7.7.3 Purification . 4
7.7.4 LC-MS/MS analysis . 4
7.8 Identification criteria . 5
7.9 Determination . 6
7.10 Blank test . 6
7.11 Calculation . 6
7.12 Limit of quantification . 6
7.13 Precision . . . 7
8 Test report . 7
Annex A (informative) Information of fipronil and metabolites . 8
Annex B (informative) LC-MS/MS chromatograms for fipronil, fipronil-desulfinyl, fipronil-
sulfide, fipronil-sulfone and fipronil- C6 mixed standard solutions .10
Annex C (informative) Precision of the method .12
Bibliography .13
iii
Foreword
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This document was prepared by Technical Committee ISO/TC 34, Food products, Subcommittee SC 6,
Meat, poultry, fish, eggs and their products.
Any feedback or questions on this document should be directed to the user’s national standards body. A
complete listing of these bodies can be found at www.iso.org/members.html.
iv
INTERNATIONAL STANDARD ISO 7124:2023(E)
Eggs and egg products — Determination of fipronil and
metabolites residues — Liquid chromatography tandem
mass spectrometry method
1 Scope
This document specifies a liquid chromatography tandem mass spectrometry method (LC-MS/MS)
for the determination of fipronil and metabolites (including fipronil-desulfinyl, fipronil-sulfide and
fipronil-sulfone) residues in eggs and egg products.
2 Normative references
The following documents are referred to in the text in such a way that some or all of their content
constitutes requirements of this document. For dated references, only the edition cited applies. For
undated references, the latest edition of the referenced document (including any amendments) applies.
ISO 3696, Water for analytical laboratory use — Specification and test methods
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
ISO and IEC maintain terminology databases for use in standardization at the following addresses:
— ISO Online browsing platform: available at https:// www .iso .org/ obp
— IEC Electropedia: available at https:// www .electropedia .org/
3.1
fipronil and metabolites content
ω
mass fraction of fipronil and metabolites residues in egg and egg products
Note 1 to entry: The fipronil and metabolites content is determined according to the procedure specified in this
document.
Note 2 to entry: The fipronil and metabolites content is expressed in micrograms per kilogram.
4 Principle
The test portion is extracted with acetonitrile. The extract is cleaned up with dispersive solid phase
extraction. The fipronil and metabolites are determined and confirmed by LC-MS/MS, and are
quantified with an internal standard method.
5 Sampling
Sampling is not part of the method specified in this document. A recommended sampling method is
given in CAC/GL 50-2004.
It is important that the laboratory receive a sample which is truly representative and has not been
damaged or changed during transport or storage.
Start from a representative sample of at least 1 kg.
Store the sample in such a way that deterioration and change in composition are prevented.
6 Preparing and storing the test sample
6.1 Preparing
6.1.1 Fresh eggs and egg products with shell
Wash approximately 15 to 20 fresh eggs (with a mass of about 1 kg) and remove the shells. Smash and
homogenize the egg samples with the appropriate equipment (7.6.6). Place the sample in a polyethylene
bottle.
Weigh about 1 kg of egg products (with shells) and then remove the shells. Smash and homogenize the
egg samples with the appropriate equipment (7.6.6). Place the sample in a polyethylene bottle.
6.1.2 Other egg products
Homogenize the egg product samples. Place each sample in a polyethylene bottle.
6.2 Storing
Store the test sample and a standby sample at a temperature of −20 °C to −16 °C, respectively.
7 Liquid chromatography tandem mass spectrometry method
7.1 Reagents
Use only reagents of recognized analytical grade, unless otherwise specified.
7.1.1 Water, conforming to grade 1 of ISO 3696.
7.1.2 Acetonitrile, HPLC grade.
7.1.3 Methanol, HPLC grade.
7.1.4 Formic acid, HPLC grade.
7.1.5 Ammonium acetate, HPLC grade.
7.1.6 Anhydrous magnesium sulfate.
7.1.7 Sodium chloride.
7.1.8 Anhydrous sodium sulfate.
7.2 Preparation of solutions
7.2.1 Formic ac
...