Agar Test pH 6,0 for Inhibition Test

Antimicrobial inhibitors contained in the samples diffuse into the nutrient media and cause growth-free inhibition zones to develop on the otherwise thickly covered plates. Repeated tests with pH 6.0, pH 8.0 and pH 7.2 are necessary, as penicillin and streptomycin are optimally active at pH 6.0 and 8.0 respectively and the activity optimum of sulfonamide is found at pH 7.2. Addition of trimethoprim to Test Agar pH 7.2 considerably increases the sensitivity of the test system to sulfonamides.

Complete inhibition of growth with a zone of inhibition of at least 2 mm is considered as a positive result. An inhibitory zone of 1-2 mm must be regarded as questionable. This only applies if the controls, prepared at the same time, show inhibition zones measuring about 6 mm.

Typical formula in g/L, adjusted and/or supplemented as required to meet performance criteria.

Preparation

Suspend 25.5 grams of the medium in one liter of distilled water. Mix well and dissolve by heating with frequent agitation. Boil for one minute until complete dissolution. Sterilize in autoclave at 121 °C for 15 minutes. Cool to 45-50 °C and aseptically add a spore suspension of Bacillus subtilis (BGA) containing approximately 1 x 10⁷ CFU/mL (final concentration in medium 1 x 10⁴ CFU/mL). Pour into plates.

Instructions for use

1. Prepare meat samples by cutting 2 mm thick slices of tissue 8 mm in diameter. A total of 6 discs are required for the complete 3-plate test (Test Agar pH 6.0, pH 7.2 and pH 8.0 LC-2024).
2. Using sterile forceps, place two of the discs in diametrically opposite positions on each test plate.
3. Aseptically place an appropriate control disc on each of the three plates.
4. Incubate the plates at 30 °C for 18-24 hours.

• Arbeitsgruppe des Instituts für Lebensmitteltechnologie und Verpackung der Technischen Universität München: Merkblätter für die Prüfung von Packmitteln, Merkblatt 18 "Prüfung auf antimikrobielle Bestandteile in Packstoffen". - Verpackgs.-Rdsch., 25; Techn.-wiss. Beilagen; 5-8 (1974).