BRITISH STANDARD Chemical disinfectants and antiseptics — Quantitative suspension test for the evaluation of fungicidal or yeasticidal activity of chemical disinfectants and antiseptics used in food, industrial, domestic and institutional areas — Test method and requirements (phase 2, step 1) ICS 71.100.35 BS EN 1650:2008 +A1:2013 BS EN 1650:2008+A1:2013 National foreword This British Standard is the UK implementation of EN 1650:2008+A1:2013 It supersedes BS EN 1650:2008 which is withdrawn The start and finish of text introduced or altered by amendment is indicated in the text by tags Tags indicating changes to CEN text carry the number of the CEN amendment For example, text altered by CEN amendment A1 is indicated by !" The UK participation in its preparation was entrusted to Technical Committee CH/216, Chemical disinfectants and antiseptics A list of organizations represented on this committee can be obtained on request to its secretary This publication does not purport to include all the necessary provisions of a contract Users are responsible for its correct application Compliance with a British Standard cannot confer immunity from legal obligations This British Standard was published under the authority of the Standards Policy and Strategy Committee on 30 June 2008 © The British Standards Institution 2013 Published by BSI Standards Limited 2013 ISBN 978 580 80140 Amendments/corrigenda issued since publication Date Comments 30 June 2013 Implementation of CEN amendment A1:2013 EUROPEAN STANDARD EN 1650:2008+A1 NORME EUROPÉENNE EUROPÄISCHE NORM May 2013 ICS 71.100.35 Supersedes EN 1650:2008 English Version Chemical disinfectants and antiseptics - Quantitative suspension test for the evaluation of fungicidal or yeasticidal activity of chemical disinfectants and antiseptics used in food, industrial, domestic and institutional areas - Test method and requirements (phase 2, step 1) Antiseptiques et désinfectants chimiques - Essai quantitatif de suspension pour l'évaluation de l'activité fongicide ou levuricide des antiseptiques et des désinfectants chimiques utilisés dans le domaine de l'agro-alimentaire, dans l'industrie, dans les domaines domestiques et en collectivité - Méthode d'essai et prescriptions (phase 2, étape 1) Chemische Desinfektionsmittel und Antiseptika Quantitativer Suspensionsversuch zur Bestimmung der fungiziden oder levuroziden Wirkung chemischer Desinfektionsmittel und Antiseptika in den Bereichen Lebensmittel, Industrie, Haushalt und öffentliche Einrichtungen - Prüfverfahren und Anforderungen (Phase 2, Stufe 1) This European Standard was approved by CEN on April 2008 and includes Amendment approved by CEN on 28 March 2013 CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this European Standard the status of a national standard without any alteration Up-to-date lists and bibliographical references concerning such national standards may be obtained on application to the CEN-CENELEC Management Centre or to any CEN member This European Standard exists in three official versions (English, French, German) A version in any other language made by translation under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC Management Centre has the same status as the official versions CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and United Kingdom EUROPEAN COMMITTEE FOR STANDARDIZATION COMITÉ EUROPÉEN DE NORMALISATION EUROPÄISCHES KOMITEE FÜR NORMUNG Management Centre: Avenue Marnix 17, B-1000 Brussels © 2013 CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Members Ref No EN 1650:2008+A1:2013: E BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Contents Page Foreword Introduction Scope Normative references Terms and definitions Requirements Test method 5.1 Principle 5.2 Materials and reagents 5.3 Apparatus and glassware 11 5.4 Preparation of test organism suspensions and product test solutions 13 5.5 Procedure for assessing the fungicidal or yeasticidal activity of the product 18 5.6 Experimental data and calculation 24 5.7 Verification of methodology 27 5.8 Expression of results and precision 28 5.9 Interpretation of results - conclusion 29 5.10 Test report 29 Annex A (informative) Referenced strains in national collections 31 Annex B (informative) Examples of neutralizers of the residual antimicrobial activity of chemical disinfectants and antiseptics and rinsing liquids 32 Annex C (informative) Graphical representations of dilution-neutralization method and membrane filtration method 34 Annex D (informative) Example of a typical test report 38 Annex E (informative) Precision of the test result 44 Bibliography 47 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Foreword This document (EN 1650:2008+A1:2013) has been prepared by Technical Committee CEN/TC 216 “Chemical disinfectants and antiseptics”, the secretariat of which is held by AFNOR This European Standard shall be given the status of a national standard, either by publication of an identical text or by endorsement, at the latest by November 2013, and conflicting national standards shall be withdrawn at the latest by November 2013 Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights CEN [and/or CENELEC] shall not be held responsible for identifying any or all such patent rights This document includes Amendment 1, approved by CEN on 2013-03-28 This document supersedes !EN 1650:2008" The start and finish of text introduced or altered by amendment is indicated in the text by tags !" This European Standard was revised to include the results of a collaborative trial (ANDISTAND), to correct obvious errors and ambiguities, to harmonize the structure and wording with other quantitative suspension tests of CEN/TC 216 (existing or in preparation), and to improve the readability of the standard and thereby make it more understandable According to the CEN/CENELEC Internal Regulations, the national standards organisations of the following countries are bound to implement this European Standard: Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and the United Kingdom BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Introduction This European Standard specifies a suspension test for establishing whether a chemical disinfectant or antiseptic has or does not have a fungicidal or yeasticidal activity in the fields described in the scope This laboratory test takes into account practical conditions of application of the product including contact time, temperature, test organisms and interfering substance, i.e conditions which may influence its action in practical situations The conditions are intended to cover general purposes and to allow reference between laboratories and product types Each utilization concentration of the chemical disinfectant or antiseptic found by this test corresponds to defined experimental conditions However, for some applications the recommendations of use of a product may differ and therefore additional test conditions should be used BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Scope This document specifies a test method and the minimum requirements for fungicidal or yeasticidal activity of chemical disinfectant and antiseptic products that form a homogeneous, physically stable preparation when diluted with hard water or - in the case of ready-to-use-products - with water Products can only be tested at a concentration of 80 % or less as some dilution is always produced by adding the test organisms and interfering substance This document applies to products that are used in food, industrial, domestic and institutional areas excluding areas and situations where disinfection is medically indicated and excluding products used on living tissues except those for hand hygiene in the above considered areas The following areas are at least included: a) processing, distribution and retailing of: 1) b) food of animal origin: 2) food of vegetable origin: milk and milk products; beverages; meat and meat products; fruits, vegetables and derivatives (including sugar, distillery ); fish, seafood, and related products; flour, milling and baking; eggs and egg products; animal feeds; animal feeds; etc etc institutional and domestic areas: catering establishments; public areas; public transports; schools; nurseries; shops; sports rooms; waste containers (bins ); hotels; dwellings; clinically non-sensitive areas of hospitals; offices; etc BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) c) other industrial areas: packaging material; biotechnology (yeast, proteins, enzymes, ); pharmaceutical; cosmetics and toiletries; textiles; space industry, computer industry; etc EN 14885 specifies in detail the relationship of the various tests to one another and to “use recommendations” NOTE The method described is intended to determine the activity of commercial formulations or active substances under the conditions in which they are used NOTE 2 This method corresponds to a phase step test Normative references The following referenced documents are indispensable for the application of this document For dated references, only the edition cited applies For undated references, the latest edition of the referenced document (including any amendments) applies EN 12353, Chemical disinfectants and antiseptics – Preservation of test organisms used for the determination of bactericidal, mycobactericidal, sporicidal and fungicidal activity EN 14885, Chemical disinfectants and antiseptics – Application of European Standards for chemical disinfectants and antiseptics ISO 4793, Laboratory sintered (fritted) filters – Porosity grading, classification and designation Terms and definitions For the purposes of this document, the terms and definitions given in EN 14885 apply Requirements The product shall demonstrate a reduction of at least a decimal log (lg) when diluted with hard water (5.2.2.7) or - in the case of ready-to-use products - with water (5.2.2.2) and tested in accordance with Clause under simulated clean conditions (0,3 g/l bovine albumin solution - 5.2.2.8.2) or simulated dirty conditions (3 g/l bovine albumin solution - 5.2.2.8.3) according to its practical applications and under the other obligatory test conditions (one or two selected test organisms, 20 °C, 15 min) The fungicidal activity shall be evaluated using the following two test organisms: Candida albicans (vegetative cells); BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Aspergillus niger (spores) The yeasticidal activity shall be evaluated using the following test organism: Candida albicans (vegetative cells); Where indicated, additional specific fungicidal or yeasticidal activity shall be determined applying other contact times, temperatures, interfering substances and test organisms (in accordance with 5.2.1, 5.2.2.8 and 5.5.1.1) in order to take into account intended specific use conditions NOTE For these additional conditions, the concentration defined as a result can be lower than the one obtained under the obligatory test conditions Test method 5.1 Principle 5.1.1 A sample of the product as delivered and/or diluted with hard water (or water for ready to use products) is added to a test suspension of fungi (yeast cells or mould spores) in a solution of an interfering substance The mixture is maintained at (20 ± 1) °C for 15 ± 10 s (obligatory test conditions) At the end of this contact time, an aliquot is taken, and the fungicidal and/or the fungistatic activity in this portion is immediately neutralized or suppressed by a validated method The method of choice is dilution-neutralization If a suitable neutralizer cannot be found, membrane filtration is used The numbers of surviving fungi in each sample are determined and the reduction is calculated 5.1.2 The test is performed using the vegetative cells of Candida albicans and the spores of Aspergillus niger (fungicidal activity) or only the vegetative cells of Candida albicans (yeasticidal activity) as test organisms (obligatory test conditions) 5.1.3 Additional and optional contact times and temperatures are specified Additional test organisms can be used 5.2 Materials and reagents 5.2.1 Test organisms The fungicidal activity shall be evaluated using the following strains as test organisms: 1) Candida albicans ATCC 10231 Aspergillus niger ATCC 16404 The yeasticidal activity shall be evaluated using only Candida albicans NOTE See annex A for strain references in some other culture collections The required incubation temperature for these test organisms is (30 ± 1) °C (5.3.2.3) If required for specific applications, additional strains may be chosen from, e.g for breweries: 1) The ATCC numbers are the collection numbers of strains supplied by the American Type Culture Collection (ATCC) This information is given for the convenience of users of this standard and does not constitute an endorsement by CEN of the product named BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Saccharomyces cerevisiae DSM 1333 Saccharomyces cerevisiae var diastaticus DSM 70487 If additional test organisms are used, they shall be incubated under optimum growth conditions (temperature, time, atmosphere, media) noted in the test report If the additional test organisms selected not correspond to the specified strains, their suitability for supplying the required inocula shall be verified If these additional test organisms are not classified at a reference centre, their identification characteristics shall be stated In addition, they shall be held by the testing laboratory or national culture collection under a reference for five years 5.2.2 5.2.2.1 Culture media and reagents General All weights of chemical substances given in this standard refer to the anhydrous salts Hydrated forms may be used as an alternative, but the weights required shall be adjusted to allow for consequent molecular weight differences The reagents shall be of analytical grade and/or appropriate for microbiological purposes They shall be free from substances that are toxic or inhibitory to the test organisms NOTE To improve reproducibility, it is recommended that commercially available dehydrated material is used for the preparation of culture media The manufacturer's instructions relating to the preparation of these products should be rigorously followed NOTE For each culture medium and reagent, a limitation for use should be fixed BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Annex C (informative) Graphical representations of dilution-neutralization method and membrane filtration method C.1 Dilution – neutralization method 34 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) 35 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) C.2 Membrane filtration method 36 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) 37 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Annex D (informative) Example of a typical test report NOTE All names and significations in Annex D are imaginary apart from those used in this European Standard NOTE Test reports for yeasticidal activity should be entitled “EN 1650, YEASTICIDAL ACTIVITY”, and be presented in the same format NOTE Only the test results of one replicate for Aspergillus niger and for Candida albicans are given as an example -HHQ Laboratories Antiseptville/Euroland Tel ++011.57 83 62-0 Fax ++011-57 83 62-19 e-mail:h.h.Q.lab@net.com TEST REPORT EN 1650, FUNGICIDAL ACTIVITY (obligatory and additional conditions) Client: Centipede Formulations Inc., Markkleeberg / Euroland Disinfectant-sample Name of the product: Z Batch number: 91-71-51 Manufacturer or - if not known - supplier: Centipede Formulations Inc (manufacturer) Storage conditions (temp and other): Room temperature, darkness Appearance of the product: Liquid, clear, yellowish Active substance(s) and their concentration(s): Not indicated Product diluent recommended by the manufacturer for use: Potable water Period of testing Date of delivery of the product: 2006-03-16 38 Dates of tests: see “Test results” (attached) BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Experimental conditions Product diluent: hard water Concentrations of the product tested: see “Test results” (attached) Obligatory conditions: test-organisms: Candida albicans ATCC 10231 and Aspergillus niger ATCC 16404; test temperature: 20 °C; contact time: 15 min; interfering substance: 3,0 g/l bovine albumin = dirty conditions; Incubation temperature: 30 °C Additional conditions: test organism: Saccharomyces yyy; Test temperature: 10 °C; contact time: 60 min; interfering substance: 10,0 g/l reconstituted milk; Incubation temperature: 30 °C Special remarks regarding the results: All controls and validation were within the basic limits At least one concentration of the product demonstrated a lg reduction of less than lg No precipitate during the test procedure (test mixtures were homogeneous) Test results: see attached sheets Conclusion: For the product Z (batch 91-71-51), the fungicidal concentration for general purposes determined according to the EN 1650 standard (obligatory conditions) under dirty conditions is: 1% (v/v) (the mean reduction of six replicates with the limiting test organism Aspergillus niger was 1,2 x 10 Candida albicans was tested once and showed a reduction of lg or more at a lower concentration than Aspergillus niger) For the product Z (batch 91-71-51), the yeasticidal concentration for specific purposes determined according to the EN 1650 standard at 10°C, with 60 contact time, with milk as interfering substance using Saccharomyces yyy as test organism is: 0,5% (v/v) Antiseptville, 2006-10-10 Alexandra May, MD, PhD, Scientific Director 39 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) !Test results (fungicidal suspension test) EN 1650 .(Phase 2, step 1) Product-name: Z Batch No.: 91-71-51.…….….…………… Remarks: Dilution neutralization method Pour plate Spread plate Number of plates / ml Neutralizer: .Lecithin 3,0 g/l in diluent Membrane filtration method Rinsing liquid: …………………… Test temperature: 20 °C Interfering substances: bovine albumin 3,0 g/l …………………… ……… Test organism: Aspergillus niger ATCC 16404 Incubation temperature:.30 °C…….…… Internal lab no: QS 58/00 Date of test:.2006-08-05 Responsible person: Fang Signature: Fang Diluent used for product test solutions: hard water.….Appearance of the product test solutions: clear…………… Validation and controls Presence of a high concentration of A brasiliensis spiny spores in the spore suspension Validation (Nv0) suspension Experimental Conditions control (A) Vc1 84 x= Vc1 85 Vc2 87 85,5 Vc2 83 30 ≤ x of Nv0 yes ≤ 160 ? no x Test suspension and Test x= of A is ≥ 0,5x yes Neutralizer control (B) no x 81 84 Vc2 85 of Nv0? x 40 83 no Vc1 79 Vc2 86 x x= 82,5 of C is ≥ 0,5x x of Nv0? yes no N Vc1 Vc2 x wm = 147,27 x 105 ; lg N = 7,17 10 -5 139 154 N0 = N/10 ; lg N0 = 6,17 10 -6 14 17 6,17 ≤ lg N0 ≤ 6,70? Test-suspension (N and N0): x= of B is ≥ 0,5x x of Nv0? yes no filtration Method validation (C) Product conc.: 10 ml/l or Vc1 yes yes no Conc of the product % Vc1 Vc2 Na = x x10 lg Na lg R Contact time (min) 0,50 > 165 > 165 > 650 > 3,22 < 2,95 15 0,75 57 65 610 2,79 3,38 15 1,00 < 140 < 2,15 > 4,02 15 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) -5 -6 Remarks: Counting per plate for (N): 10 = 60 + 79, 72+ 82, 10 = + 10, + 15 Explanations: Vc = count per ml (one plate or more) x wm = x R = reduction (lg R = lg N0 – lg Na) = average of Vc1 and Vc2 (1 + duplicate) weighted mean of x " 41 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Test results (fungicidal suspension test) EN 1650 .(Phase 2, step 1) Product-name: Z Batch No.: 91-71-51.…….…………………… Remarks: Dilution neutralization method Pour plate Spread plate Number of plates / ml Neutralizer: .Lecithin 3,0 g/l in diluent …………………………… Membrane filtration method Rinsing liquid: ………………………………… Test temperature: 10°C Interfering substances: bovine albumin 3,0 g/l ……………………………… Test organism: Candida albicans ATCC 10231 ………… Incubation temperature:.30°C Date of test:.2006-08-27 Internal lab no: QS 58/08 Responsible person: Fang Signature: Fang Diluent used for product test solutions: hard water Appearance of the product test solutions: clear……………………… Validation and controls Validation (Nv0) Vc1 76 x= Vc1 82 Vc2 72 74 Vc2 81 30 ≤ x of Nv0 ≤ 160 ? yes x no Test suspension and Test x= 81,5 of A is ≥ 0,5x x of Nv0? yes Vc1 81 Vc2 72 x no the Vc1 70 x= 85,5 Vc2 76 73 of Nv0? x x= of B is ≥ 0,5x yes no x of C is ≥ 0,5x x of Nv0? yes no N Vc1 Vc2 x wm = 259,54 x 105 ; 10 -5 241 283 N0 = N/10 ; lg N0 = 6,41 10 -6 20 27 6,17 ≤ lg N0 ≤ 6,70? Test-suspension (N and N0): Conc of product % 42 Neutralizer or filtration control Method validation (C) Product conc.: 7,5 ml/l (B) suspension Experimental Conditions control (A) Vc1 Vc2 Na = x x10 lg Na lg R lg N = 7,41 yes no contacttime (min) 0,25 > 660 600 > 300 > 3,80 < 2,61 15 0,50 21 28 245 2,39 4,02 15 0,75 < 140 < 2,15 > 4,26 15 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Remarks: Counts per spread plate (2 / ml): Nv0 38+38, 34+38 A 60+22, 52+29 B 32+49, 32+40 -5 -6 36, 25+51 N 10 120+121, 130+153 N 10 10+10, 15+1 Na 0,25% >330+>330, 320+280 13+8, 7+21 Na 0,75% 1+7 C 34 + Na 0,5% Explanations: Vc = count per ml (one plate or more) x = average of Vc1 and Vc2 (1 + duplicate) x wm = weighted mean of x R = reduction (lg R = lg N0 – lg Na) 43 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Annex E (informative) Precision of the test result A collaborative study (ANDISTAND 1997 to 1999) was carried out to determine the precision of the test method within and between different laboratories The study involved 12 laboratories from different European countries Each laboratory replicated the test three times Although the interfering substance was different (EN 1657-high soiling) the results can be extrapolated to this European Standard (EN 1650) The tests were performed using the dilution neutralization or membrane filtration methods with sodium dichlorisocyanurate and phenol on Aspergillus niger and Candida albicans (as test organisms) The complete results and the statistical evaluation of this study are described in documents CEN/TC 216 HWG N 121 + N 121 Annexes + N 121 Corrigendum The agreement between laboratories, expressed in terms of fungicidal effect (reduction R = lg), is very good at low and high concentrations, but less good at intermediate levels Uncountable data were replaced by theoretical fake values of reduction factor When counts were below the lower counting limit (15 viable colonies), the reduction factor could vary between 3,33 × 10 (the maximum value when data were countable) and 5,00 × 10 (no surviving colonies), corresponding respectively to 4,52 and 6,70 log reduction In order to simulate a possible variability, three theoretical reduction factors (one for each replicate) were chosen in this range: 4,5; 5,6; and 6,7 (in logarithmic terms) When counts were above the upper counting limit (150 viable colonies), the reduction factor could vary between 1,00 × 10 (no reduction) and 1,00 × 10 (the minimum value when data were countable), giving the log reduction values of, respectively, 0,00 and 3,00 In this case, the three chosen fake theoretical reduction factors were 0,0; 1,5; and 3,0 All these theoretical values were collected and are shown in Table E.1 Table E.1 — Theoretical reduction factors used for fungi (in logarithmic terms) Below the lower limit (15) Above the upper limit (150) All three replicates are 4,5 – 5,6 – 6,7 0,0 – 1,5 – 3,0 Only two replicates are 4,5 – 6,7 0,0 – 3,0 5,6 1,5 Only one replicate is Variance analyses were performed on the reduction factors in order to point out the tested conditions with significant obtained results and to estimate the two types of variability: "within" and "between" laboratories The differences in reduction factors across the laboratories were essentially obtained with medium and high dilutions of the tested products Some of those significant results were due to outliers, certain laboratories giving completely different reduction factor from most of the others In other cases, it was more difficult to point out the outliers, but the significant differences were the consequences of the important "within lab" variability In most cases, the "between" estimated standard deviation, which varied (in logarithmic terms) from 0,89 to 2,42 for sodium dichlorisocyanurate and from 0,47 to 1,89 for phenol, was higher than the "within" value, which oscillated between 0,63 and 1,80 Nevertheless, in some cases, this ratio was inverted ("between" dispersion lower than "within" one), due probably to the techniques used for replacing uncountable data, which overestimated the "within" variability and reduced the "between" variation 44 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) The "inside" variability was also used to estimate the precision of the obtained reduction factors Two different hypotheses were taken into account: the first, which could be qualified as the "worst" case, used the maximum of the calculated "inside" variability (σ = 1,66), while the second one was based on an average "inside" dispersion (σ = 1,23) The estimated precision depended also on the sample size (number of replicates) and the confidence level (90% probability used in the calculation) With three replicates, the "worst" case led to a reduction factor precision of ± 2,80, while the "mean" case was about ± 2,07 In other words, if the precision target is fixed to ± lg reduction, ten replicates were needed in the "worst" case and seven in the "mean" one (Figure E.1 or Table E.2) Key A B X Y Worst case Mean case Sample size (number of replicates) Precision (in log terms) Figure E.1— Precision of the reduction factor obtained with fungi (in logarithmic terms) 45 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Table E.2 — Precision of the reduction factors obtained for fungi (in logarithmic terms) Replicates Standard deviation (σ) 1,0 1,1 1,2 1,3 1,4 1,5 1,6 1,7 1,8 ± 4,46 ± 4,91 ± 5,36 ± 5,80 ± 6,25 ± 6,70 ± 7,14 ± 7,59 ± 8,04 ± 1,69 ± 1,85 ± 2,02 ± 2,19 ± 2,36 ± 2,53 ± 2,70 ± 2,87 ± 3,03 ± 1,18 ± 1,29 ± 1,41 ± 1,53 ± 1,65 ± 1,77 ± 1,88 ± 2,00 ± 2,12 ± 0,95 ± 1,05 ± 1,14 ± 1,24 ± 1,33 ± 1,43 ± 1,53 ± 1,62 ± 1,72 ± 0,82 ± 0,90 ± 0,99 ± 1,07 ± 1,15 ± 1,23 ± 1,32 ± 1,40 ± 1,48 ± 0,73 ± 0,81 ± 0,88 ± 0,95 ± 1,03 ± 1,10 ± 1,18 ± 1,25 ± 1,32 ± 0,67 ± 0,74 ± 0,80 ± 0,87 ± 0,94 ± 1,00 ± 1,07 ± 1,14 ± 1,21 ± 0,62 ± 0,68 ± 0,74 ± 0,81 ± 0,87 ± 0,93 ± 0,99 ± 1,05 ± 1,12 10 ± 0,58 ± 0,64 ± 0,70 ± 0,75 ± 0,81 ± 0,87 ± 0,93 ± 0,99 ± 1,04 11 ± 0,55 ± 0,60 ± 0,66 ± 0,71 ± 0,77 ± 0,82 ± 0,87 ± 0,93 ± 0,98 12 ± 0,52 ± 0,57 ± 0,62 ± 0,67 ± 0,73 ± 0,78 ± 0,83 ± 0,88 ± 0,93 13 ± 0,49 ± 0,54 ± 0,59 ± 0,64 ± 0,69 ± 0,74 ± 0,79 ± 0,84 ± 0,89 14 ± 0,47 ± 0,52 ± 0,57 ± 0,62 ± 0,66 ± 0,71 ± 0,76 ± 0,80 ± 0,85 15 ± 0,45 ± 0,50 ± 0,55 ± 0,59 ± 0,64 ± 0,68 ± 0,73 ± 0,77 ± 0,82 NOTE It cannot be excluded that the precision may be better or worse when other test organisms, products, and/or interfering substances are tested However, it is likely that the precision in these cases will be in the same range 46 BS EN 1650:2008+A1:2013 EN 1650:2008+A1:2013 (E) Bibliography [1] European Pharmacopoeia (EP), Edition 1997 supplement 2000, Water for injections 47 This page deliberately left blank