Robust experimental design for optimizing the microbial inhibitor test for penicillin detection in milk

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To use experimental design techniques and a multiple logistic regression model to optimize a microbiological inhibition test with dichotomous response for the detection of Penicillin G in milk.

Methods and Results:

A 23 × 22 robust experimental design with two replications was used. The effects of three control factors (V: culture medium volume, S: spore concentration of Geobacillus stearothermophilus, I: indicator concentration), two noise factors (Dt: diffusion time, Ip: incubation period) and their interactions were studied. The V, S, Dt, Ip factors and V × S, V × Ip, S × Ip interactions showed significant effects.


The use of 100 μl culture medium volume, 2 × 105 spores ml−1, 60 min diffusion time and 3 h incubation period is recommended. In these elaboration conditions, the penicillin detection limit was of 3·9 μg l−1, similar to the maximum residue limit (MRL). Of the two noise factors studied, the incubation period can be controlled by means of the culture medium volume and spore concentration.

Significance and Impact of the Study:

We were able to optimize bioassays of dichotomous response using an experimental design and logistic regression model for the detection of residues at the level of MRL, aiding in the avoidance of health problems in the consumer.

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