Analysis of pesticides in egg products will have higher priority after fipronil

The insecticide was found in Dutch and Belgian farms and has affected 15 European countries as well as Switzerland and Hong Kong.

Investigations are focused on Poultry Vision, a pest control firm from Belgium and Chickfriend, a Dutch poultry farm cleaning company.

Products were marketed under DEGA 16 and COOPER BOOST and used on poultry farms.

Use of amitraz, an insecticide used against insects and mites, is also being investigated as it cannot be used in poultry. It is a moderate toxic substance that can cause effects on the central nervous system.

TLR International Labs was approved by NVWA to do fipronil testing following confirmation of contamination. It performs such analysis using Sciex 6500+ Triple Quad LC-MS/MS Systems.

Eurofins recommended unannounced inspections along the supply chain to prevent such unexpected contamination cases.

Thermo Fisher said some labs are working 24/7 to cope with demand.

TLR International Laboratories using Sciex instruments

Dr Niels Martha, lab manager at TLR International Laboratories, Netherlands, said it assures results within eight hours for the most urgent class of samples.

“Fipronil analysis was always included in our method and was routinely tested as part of many monitoring programmes. Unfortunately, the products that were introduced into the chicken farms were not previously tested.”

Dr Ashley Sage, senior manager, Applied Markets Development, EMEAI and Phil Taylor, global marketing manager, Food, Environmental & Forensics of Sciex, said the most common techniques are LC-MS/MS (liquid chromatography coupled with tandem mass spectrometry) and GC-MS (gas chromatography coupled with mass spectrometry) for targeted screening. There is also growing interest in high resolution (HR)-MS.

Food samples have to be prepared using extraction methods such as QuEchERs, mini-luke or SweET. When using GC-MS additional sample preparation steps such as derivitisation are required.

Taylor said the contamination has sparked an increase in enquiries from testing firms who need mass spectrometry-based methods for testing fipronil.

“There’s growing interest in using SWATH data independent acquisition for non-targeted analysis of food; this approach is ideal for analysing all compounds in a sample, including unknown contaminants.”

Sage said most CROs are routinely testing customers’ products for more than the 240 pesticides required to be monitored in foodstuffs in the EU.

“To protect consumers from other substances that may be in use, these labs typically test upwards of 500 compounds per analysis, and as demand grows for greater vigilance, it’s likely that non-targeted screening with SWATH, for example, will complement these targeted approaches.” 

Agriculture Committee MEPs are to hear from the Commission on the issue this Thursday.

Meanwhile, the Joint Research Centre is developing a certified reference material in response to the crisis.

Achieving sensitivity in fat containing matrices

Marion Mandix and Johannes Jaschik, analytical service managers at Eurofins Food Testing Germany, said fipronil can be analysed with different chromatographic systems. The challenge is to achieve the necessary sensitivity for detection even in fat containing matrices such as meat and eggs.

“Therefore, Eurofins is primarily using LC-MS/MS-systems for detection of fipronil in such products. Sample preparation is important to guarantee the reproducibility for the different matrices. We are accredited for the matrices animal and vegetable fat, eggs and egg products, beverages.”

The Dutch site is approved by the NVWA and FAVV to do the original and contra analyses.

Eurofins is analysing fipronil and its metabolites within two days or faster if needed by clients. Analysis is performed with an LOQ (Limit of Quantification) of 0,003 mg/kg (LC-MS/MS).

Mandix and Jaschik said every food business operator must guarantee traceability of raw materials and products to ensure transparency in case of a crisis.

“A reliable long-term relationship between food producers and their suppliers and service providers is a key factor for brand protection.

“However, there is no analytical strategy available to prevent unexpected cases like this. Therefore, it is highly recommended to carry out unannounced inspections along the manufacturing and supply chain.”

Fipronil can be used legally as an insecticide for some vegetables, e.g. potatoes, onions, leeks, cabbages and is routinely checked as part of standard multi residue screening.

“Untargeted techniques would expand the scope of the screening, but are technically limited in sensitivity, reliability and are only applicable for specific groups of substances,” said Mandix and Jaschik.

“As there is no analytical method available that covers all possible risks, you would need to combine several methods, targeted and untargeted, leading to relatively high costs. Due to the current findings, the analysis of pesticides, including acaricides and insecticides, in egg products will have a higher priority in future.”

Thermo Fisher Scientific: Some labs working 24/7

Dr Khalil Divan, senior director, food and beverage at Thermo Fisher Scientific, said testing for Fipronil and the metabolite, fipronil sulphone, in eggs and meat is relatively straightforward.

It typically involves a generic extraction with enough clean-up to remove unwanted co-extracted lipids, followed by determination using LC-MS or GC-MS.

“The clean-up step is usually any combination of dispersive solid phase extraction using C18 material, back extraction with hexane, and/or a freezing out step.

“We know of one laboratory who is using GC- triple quadrupole MS as the primary quantitation method, LC-triple quadrupole MS as the confirmatory method, and because of political and legal sensitivities, also using GC-high resolution accurate mass (GC-Orbitrap) to provide increased confidence in the accurate identification and quantification of fipronil and fipronil sulphone in selected samples.”

Labs are typically offering a 24 hour turnaround service although it is possible, depending on calibration method, to analyse an individual sample in three to four hours from receipt.

Dr Divan said some labs are working 24/7 to cope with demand.

“It is difficult to protect the food chain against all eventualities, as is evident from the frequent food safety and food fraud issues, such as the misuse of isofenphos methyl on vegetables, sudan dyes in spices, chloramphenicol in acquaculture products, nitrofen in animal feed, dioxins in pork and many others.

“An increase of screening strategies using high resolution accurate mass (HRAM) mass spectrometry, combined with liquid chromatography and with gas chromatography to maximize the scope, may help to detect unwanted contaminants or food fraud more quickly, but it is not easy. While MS technology has advanced rapidly in the last few years, there are still limitations in extraction.”

British egg processors criticised the buying policies of UK supermarkets after several of them withdrew egg-containing products contaminated with fipronil.

Ian Jones, chairman of British Lion Egg Processors, said: “The major retailers are operating to double standards when it comes to eggs. All of them stock British Lion shell eggs but they use imported eggs in many of their other foods containing eggs.”

SGS, Agilent and Waters

SGS has three labs accredited to perform fipronil testing and analysis on eggs and egg products in Berlin (Germany), Varna (Bulgaria) and Mersin (Turkey).

In Berlin and Varna, the labs can determine fipronil content in eggs at 0.003 mg/kg.

Other SGS labs can perform testing to detect fipronil in food using GC/MS, GC/MS/MS and LC/MS/MS mass spectrometry techniques.

Chinese scientists at the National Reference Laboratory of Veterinary Drug Residues have developed a sensitive and reliable method to determine residual fipronil in chicken egg and muscle. 

It uses an Agilent high-performance liquid chromatography system and Zorbax column.

German scientists at the Saxon Sate Laboratory of Health and Veterinary Affairs have developed a sample preparation method to determine 97 GC and 81 LC amenable residues. 

It uses an Agilent HPLC system coupled to an Agilent triple-quadrupole mass spectrometer.

Fipronil and fipronil sulfone can be determined using LC-MS/MS with electrospray ionization (EI) in negative ion mode. However, matrix effects from co-eluting co-extractives can suppress the mass spectrometry signal, reducing sensitivity of the method, said Waters.

The firm offers an atmospheric pressure GC (APGC) ionization source that can be used with GC as an inlet to a triple quadrupole mass spectrometer for trace-level quantification.

APGC offers ‘soft’ ionization, meaning that less fragmentation is observed for many compounds compared with techniques such as EI. Reduced fragmentation can provide higher sensitivity and specificity and simplify precursor ion selection in MS/MS analyses.

Waters said labs such as Primoris Belgium have observed lower detection limits using APGC-MS/MS compared to LC-MS/MS. The lab is ISO17025 accredited for fipronil analysis in food of animal origin.

EU hearing Thursday

The hearing for Agriculture Committee MEPs this week is on the illegal use of substances containing fipronil at laying hen farms and consequences for the food chain.

Annie Schreijer-Pierik of the European People's Party (PPE) asked a written question about the financial support measures the European Commission will introduce for affected breeders in different Member States to avoid market disruption stemming from increasing bankruptcies.

Ivo Belet, of the same party, said there was an ‘inadequate exchange’ of food safety alerts.

“The exchange of information between national food safety agencies is clearly failing: food safety alerts should be automatically shared throughout Europe within three days via the Rapid Alert System for Food and Feed, but in the Fipronil affair there was no communication between agencies for several weeks.”

Sophie Montel (ENF) said France could be self-sufficient in eggs but because of the free-trade model it is forced to import items which can be of a lower quality and are sometimes dangerous.