Maximum allowable residue limits and analytical techniques of nitrofuran metabolites-pretreatment methods (1)

7.
1.
3 Maximum allowable residue limit

Studies have shown that NFs can induce mutations and have carcinogenicity and chronic toxicity.
In order to protect the health and safety of consumers, most countries have banned the use of such drugs in the production and breeding of food animals
.

Australia revoked the maximum residue limits of NFs in 1992 and banned their use; in 1993, the EU stipulated in Annex IV of 2377/90/EEC that nitrofurazone, nitrofurantoin and furazolidone were banned from being used in food animals, and furazolidone was listed in 1995.

7.
1.
4 Residue analysis technology

NFs are metabolized quickly in animals and have a short half-life.
The detection of the original drug is not enough to reflect the true residual level and drug use
.

In the study of NFs detection methods, the development process from the detection of prototype drugs to the detection of metabolites has been experienced, and the analysis of NFs metabolite residues has become an important technical means to determine whether NFs is abused or not

7.
1.
4.
1 Pre-processing method

NFs are photosensitive substances and are particularly sensitive to sunlight.
Therefore, sample pretreatment operations must be avoided under strong light
.

(1) Sample washing

In recent years, studies have found that NFs metabolites, especially semicarbazide (SEM), have complex mechanisms and diverse sources.
The abuse of nitrofurazone is not the only reason for the detection of SEM
.

In order to accurately reflect the actual use of NFs and reduce the interference of exogenous substances on the test results, only bound metabolites are measured in some animal-derived foods, and free metabolites are washed away, which can reduce the risk of misjudgment

(2) Hydrolysis and derivatization

NFs metabolites mainly exist in organism tissues in the form of protein conjugates, and can only be released under appropriate acidic conditions.
Usually, dilute hydrochloric acid is used to hydrolyze protein-bound metabolites in tissues
.

At the same time, NFs metabolites are all small molecule compounds.

Ortho-nitrobenzaldehyde (2-NBA) is usually used as a derivatization reagent, but a longer reaction time is required, generally requiring derivatization at 37°C for 16 hours
.

In order to shorten the derivatization time, Alexander and others hope to use other aromatic aldehydes (such as pyridine-3-carboxyformaldehyde, 2,2-dinitrobenzaldehyde, 2-hydroxy-5-nitrobenzaldehyde) to improve detection sensitivity or Shorten the reaction time

Currently, 2-NBA is a commonly used derivatization reagent
.

The mechanism of the derivatization reaction can be summarized as: under acidic conditions, the ketone-aldehyde group (-CHO) of the derivatizing agent and the nitrogen-containing nucleophilic group amino group (-NH ₂ ) of different metabolites undergo nucleophilic addition reaction of aldehyde and amine

Figure 7-2 Simultaneous hydrolysis and derivatization process of protein-bound NFs metabolites

Some researchers also use o-chlorobenzaldehyde (2-CBA) or 2-naphthaldehyde (NTA) as derivatization reagents
.

Zhu Jian established an LC-MS method for the determination of NFs metabolites AOZ and AMOZ residues in chicken and aquatic shrimp

Related Links: Metabolism and Toxicology of Nitrofuran Metabolites in the Body