Accelerated solvent extraction of animal feedingstuffs for microbial growth inhibition screening for the presence of antimicrobial feed additives.

Three plate systems (combinations of indicator organism and growth medium) were evaluated for the detection of analytical standards of the banned feed additives avoparcin, bacitracin zinc, spiramycin, tylosin and virginiamycin. When authorized in the EU, the previously recommended minimum inclusion rate (MIR) for each compound was 5 mg kg(-1). One of the plate systems (Micrococcus luteus ATCC 10240, nutrient agar) detected all five additives. This plate was used in a further study that evaluated the suitability of accelerated solvent extraction (ASE) as a first step in the development of a rapid single-plate screening assay. A drug-free (negative control) feedingstuff was fortified with the compounds (0-50 mg kg(-1)), extracted by ASE and the extracts applied to the plate at each of three pH ranges - unadjusted extract (pH 5.7-5.9), pH 6.5 and 8.0. At pH 6.5, sub-MIR concentrations of virginiamycin and tylosin were detectable. Avoparcin was detectable at 6.3 mg kg(-1). The detection of zinc bacitracin was#10; pH-independent (10 mg kg(-1)). At pH 8.0, spiramycin was detectable at 5.4 mg kg(-1). Mean +/- SD analytical recoveries from fortified feedingstuffs (n = 10) ranged from 57 +/- 1.5% for avoparcin to 96 +/- 4% for virginiamycin. The five additives were also detectable following ASE extraction from a range of different feedingstuffs fortified with each of the drugs. A further 24 compounds permitted for use in animal feeds were tested. Of these, nine were detectable at their recommended MIR. It is concluded that ASE is a versatile technique suitable for the automated extraction of a range of antimicrobials from animal feedingstuffs. Employing ASE with this single-plate detection system permits the rapid antimicrobial screening of animal feedingstuffs and allows the detection of the banned additives. Whilst the method is applicable as a screening test, more specific postscreening methods would be necessary for subsequent identification (and quantification) of antimicrobials in screening-positive samples.