EN 13623-2010 pdf download.Chemical disinfectants and antiseptics – Quantitative suspension test for the evaluation of bactericidal activity against Legionella of chemical disinfectants for aqueous systems – Test method and requirements (phase 2, step 1).
b) Note for each plate the exact number of colonies but record -> 330” for any counts higher than 330 and determine the Vt-values according to 5.6.2.2.
c) Calculate the numbers of cfu/ml in the test suspension N and in the validation suspension N, using the methods given in 5.6.2.3 and 5.6.2.5. Verify according to 5,7.
5.4.2 Product test solution
The concentration of a product test solution shall be ten times the desired test concentration because it is diluted to 10 % during the test and the method validation (5.5.2 or 5.5.3). Product test solutions shall be prepared in hard water at minimum three different concentrations to include one concentration In the active range and one concentration in the non-active range (5.8.2). The type of hard water is selected according to the intended use: BFHW (5.2.2.10) for products intended to be used for treatment of cooling water and HWGP (5.2.2.7) for products intended to be used for treatment of water for general purposes. The product as received may be used as one of the product test solutions, in this case the highest tested concentration is
10%.
For solid products, dissolve the product as received by weighing at least 1.0 g ± 10 mg of the product in a volumetric flask and filling up with hard water (5.2.2.7 or 5.2.2.10). Subsequent dilutions (lower concentrations) shall be prepared in volumetric flasks (5.3.2.12) on a volume/volume basis in hard water (5.2.2.7 or 5.2.2.10).
For liquid products, dilutions of the product shall be prepared with hard water on a volume/volume basis using volumetric flasks (5.3.2.12).
The product test solutions shall be prepared freshly and used in the test within 2 h. They shall give a physically homogeneous preparation that is stable during the whole procedure. If during the procedure a visible inhomogeneity appears due to the formation of a precipitate or flocculant (for example, through the addition of the interfering substance), it shall be recorded in the test report.
NOTE Counhng micro-organisms embedded in a precipitate or flocculant is difficult and unreliable.
The concentration of the product stated in the test report shall be the desired test concentration. Record the test concentration in terms of mass per volume or volume per volume and details of the product sample as received.
5.5 Procedure for assessing the bactericidal activity of the product
5.5.1 General
5.5.1.1 Experimental conditions (obligatory and additional)
1) the obligatory test organism is Legionella pneumophila (5.2.1);
2) additional test organisms (other strains of Legoriei(a pneumophiia or species of Legionella) may be
used.
5.5.1.2 Choice of test method
The method of choice Is the dilution-neutralization method. To determine a suitable neutralizer carry Out the validation of the dilution neutralization method (5.5.2.3, 5.5.2.4 and 5.5.2.5 in connection with 5.5.2.6) using a neutralizer, chosen according to laboratory experience and published data.
If this neutralizer Is not valid, repeat the validation test using an alternative neutralizer taking into account the information given in Annex B.
If both neutralizers are found to be invalid, the membrane filtration method (5.5.3) may be used.
5.5.1.3 General Instructions for validation and control procedures
The neutralization and/or removal of the bactericidal or bacteriostatic activity of the product shall be controlled and validated — only for the highest product test concentration — for each of the used test organisms and for each experimental condition (contact time). These procedures (experimental condition control, neutralizer or filtration control and method validation) shall be performed at the same time with the test and with the same neutralizer — or rinsing liquid — used in the test.
The same hard water (5.2.2.7 or 5.2.2.10) used in the test (5.5.2.2) shall be used in the validation and controls. The same agar (5.2.2.3) used in the test (5.5.2.2) shall be used in the validation and controls.
5.5.1.4 Equilibration of temperature
Prior to testing, equilibrate all reagents (product test solutions (5.4.2). test suspension (5.4.1.4), validation suspension (5.4.1.5) hard water (5.2.2.7 or 5.2.2.10) and interfering substance (5.2.2.8) to the test temperature of either 20 °C for products intended for non cooling water applications or 30 °C for cooling water products (5.5.1.1, a)) using the water bath (5.3.2.2) controlled at (20 ± 1) °C or (30 ± 1) °C.
Check that the temperature of the reagents is stabilized at the chosen temperature.
The neutralizer (5.2.2.5) or the rinsing liquid (5.2.2.6) and water (5.2.2.2) shall be equilibrated at a
temperature of (20 ± 1) °C.
5.5.1.5 Precautions for manipulation of test organisms
Do not touch the upper part of the test tube sides when adding the test or the validation suspensions (5.4.1).EN 13623-2010 pdf download.