Penzyme test

Other miscellaneous assays for penicillin or other 3-lactams in milk is the Penzyme Test which uses cell wall enzjrmes inhibited by 3-lactam drugs in a kinetic assay. This test system is purported to be able to detect 0.005 units penicillin/mL and requires approximately 30 min to complete. It, like many other assays, detects 3-lactam antibiotics only. 

The enzymatic colorimetric format is followed by the Penzyme test. This test is a qualitative enzymatic assay for rapid detection of -lactam residues in milk (28-30). The detection principle of the Penzyme test is based on measurement of the degree of inactivation of the enzyme oo-carboxypeptidase is involved in the synthesis of the bacterial cell wall by -lactam antibiotics. These residues bind specifically with the enzyme and inactivate it, thus interfering with bacterial cell wall formation. 

In the Penzyme test, freeze-dried Streptomyces oo-carboxypeptidase is placed in sealed vials to which the milk sample is added. A preliminary incubation is carried out for 5 min at 47 C to cause some inactivation of the enzyme molecules. The degree of inactivation is dependent on the amount of -lactams presumed to be present in the milk sample. Subsequent addition of a reagent tablet containing synthetic o-alanine oligopeptide and o-amino acid oxidase followed by reincubation at 47 C for 15 min results in the release of o-alanine, its amount... 

In 1989, an experimental study designed by the International Dairy Federation in 53 laboratories of 22 different countries to achieve deeper insight into state of proficiency of routinely applied tests showed that the most frequently used microbial inhibitor screening tests were the disc assays with Bacillus stearo-tliermopliilus, Delvotest-P, Brilliant black reduction test, acidification test, CHARM II test, and the Penzyme test (32), Currently available microbial inhibitor tests for screening of residual antibacterials in milk and milk products are presented in Table 27.1. 

In general, the Penzyme test is applicable for screening milk samples for the presence of f)-lactam compounds with detection limits close to the USFDA tolerances (Table 5.6) and is, therefore, applicable for the regulatory monitoring in accordance with the USA system. However, in all cases the detection limits quoted are greater than the current EU MRLs, and for that reason the test is not considered as a useful monitoring tool for the European market. 

An important consideration is the applicability of the rapid test to the range of sample matrices, and its susceptibility to naturally occurring interference. In a study conducted by Andrew et al., the performance of rapid antibiotic residue screening tests including the Penzyme test were evaluated in presence of different milk compositions and qualities. Metabolic changes that occur because of disease affect the composition and quality of the milk produced. Mastitis is the major disease for which antibiotic treatment is used and after which antibiotic residue screening assays are employed. 

The enzyme exploited in the Penzyme milk test (Neogen Corporation, Michigan, USA) for P-lactam residues is D,D-carboxypeptidase. The assay is based on the following two properties of d, D-carboxypeptidase ... 

The Delvo-X-PRESS (DSM Food Specialties, Delft, The Netherlands) is an enzyme-based rapid test, which operates in a similar mode to that of the Penzyme. The Delvo-X-PRESS is specially designed to control milk from bulk tank and silos within minutes for the presence of P-lactam residues. The Delvo-X-PRESS is a qualitative. 

TABLE 5.6 Manufacturer s Reported Detection Limits for the Penzyme Milk Test and the Delvo-X-PRESS for Various fS-Lactam Compounds in Spiked Bovine Milk Compared to USEDA Tolerances and EU MRLs... 

Gibbons-Burgener et al. " evaluated the performance of the rapid SNAPBL, DelvoTest SP, and Penzyme milk test-kits for testing raw milk samples from individual cows. 

---