BS ISO 8784-1-2014 Pulp paper and board Microbiological examination Enumeration of bacteria and bacterial spores based on disintegration《纸浆 纸和纸板 微生物检验 基于分解的细菌和细菌孢子的计数》.pdf

1、BSI Standards Publication BS ISO 8784-1:2014 Pulp, paper and board Microbiological examination Part 1: Enumeration of bacteria and bacterial spores based on disintegrationBS ISO 8784-1:2014 BRITISH STANDARD National foreword This British Standard is the UK implementation of ISO 8784-1:2014. It super

2、sedes BS ISO 8784-1:2005 which is withdrawn. The UK participation in its preparation was entrusted to Technical Committee PAI/11, Methods of test for paper, board and pulps. A list of organizations represented on this committee can be obtained on request to its secretary. This publication does not p

3、urport to include all the necessary provisions of a contract. Users are responsible for its correct application. The British Standards Institution 2014. Published by BSI Standards Limited 2014 ISBN 978 0 580 71019 3 ICS 07.100.99; 85.040; 85.060 Compliance with a British Standard cannot confer immun

4、ity from legal obligations. This British Standard was published under the authority of the Standards Policy and Strategy Committee on 31 December 2014. Amendments/corrigenda issued since publication Date T e x t a f f e c t e dBS ISO 8784-1:2014 ISO 2014 Pulp, paper and board Microbiological examina

5、tion Part 1: Enumeration of bacteria and bacterial spores based on disintegration Ptes, papiers et cartons Analyse microbienne Partie 1: Dnombrement des bactries et des spores bactriennes bas sur la dsintgration INTERNATIONAL STANDARD ISO 8784-1 Third edition 2014-12-01 Reference number ISO 8784-1:2

6、014(E)BS ISO 8784-1:2014ISO 8784-1:2014(E)ii ISO 2014 All rights reserved COPYRIGHT PROTECTED DOCUMENT ISO 2014 All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized otherwise in any form or by any means, electronic or mechanical, including photoc

7、opying, or posting on the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address below or ISOs member body in the country of the requester. ISO copyright office Case postale 56 CH-1211 Geneva 20 Tel. + 41 22 749 01 11 Fax + 41 22 749 09

8、47 E-mail copyrightiso.org Web www.iso.org Published in SwitzerlandBS ISO 8784-1:2014ISO 8784-1:2014(E)Contents Page Foreword iv Introduction v 1 Scope . 1 2 Normative references 1 3 T erms and definitions . 1 4 Principle 2 5 Culture media and diluents . 2 5.1 General . 2 5.2 Water . 2 5.3 Culture m

9、edia for total bacteria count and spore count . 2 5.4 Diluents . 2 6 Apparatus and equipment 3 6.1 General . 3 6.2 List of equipment 3 7 Sampling 3 8 Preparation of the test material . 4 8.1 General . 4 8.2 Determination of dry-matter content . 4 8.3 Weighing . 4 8.4 Disintegration 4 9 Determination

10、 of the total bacterial count and spore count 5 9.1 General . 5 9.2 Plating for total bacterial count 5 9.3 Plating for spore count . 6 9.4 Incubation . 6 10 Enumeration of the colonies 6 11 Calculation and report 6 11.1 Calculation 6 11.2 Interpretation 7 11.3 Report . 8 12 Test report . 8 Annex A

11、informative) Dilution fluid 9 Annex B (informative) Precision 10 Bibliography .14 ISO 2014 All rights reserved iiiBS ISO 8784-1:2014ISO 8784-1:2014(E) Foreword ISO (the International Organization for Standardization) is a worldwide federation of national standards bodies (ISO member bodies). The wo

12、rk of preparing International Standards is normally carried out through ISO technical committees. Each member body interested in a subject for which a technical committee has been established has the right to be represented on that committee. International organizations, governmental and non-governm

13、ental, in liaison with ISO, also take part in the work. ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization. The procedures used to develop this document and those intended for its further maintenance are described in t

14、he ISO/IEC Directives, Part 1. In particular the different approval criteria needed for the different types of ISO documents should be noted. This document was drafted in accordance with the editorial rules of the ISO/IEC Directives, Part 2 (see www.iso.org/directives). Attention is drawn to the pos

15、sibility that some of the elements of this document may be the subject of patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of any patent rights identified during the development of the document will be in the Introduction and/or on the ISO list

16、of patent declarations received (see www.iso.org/patents). Any trade name used in this document is information given for the convenience of users and does not constitute an endorsement. For an explanation on the meaning of ISO specific terms and expressions related to conformity assessment, as well

17、as information about ISOs adherence to the WTO principles in the Technical Barriers to Trade (TBT) see the following URL: Foreword - Supplementary information The committee responsible for this document is ISO/TC 6, Paper, board and pulps, Subcommittee SC 2, Test methods and quality specifications f

18、or paper and board. This third edition cancels and replaces the second edition (ISO 8784-1:2005), which has been technically revised. The second edition was applicable to yeast and mould, as well as bacteria. The following main changes have been made with respect to the previous edition: This third

19、edition is only applicable to bacteria and bacterial spores, and no longer applicable to yeast and mould; incubation temperature changed from (37 C 1 C) to (32 C 2 C) (9.4); 2 parallel determinations are to be made (Clause 8 and Clause 9); the result can be reported “as received” in addition to repo

20、rting on a dry-mass basis ( 8.2, 11.1, and Clause 12). ISO 8784 consists of the following parts, under the general title Pulp, paper and board Microbiological examination: Part 1: Enumeration of bacteria and bacterial spores based on disintegrationiv ISO 2014 All rights reservedBS ISO 8784-1:2014ISO

21、 8784-1:2014(E) Introduction This part of ISO 8784, which deals with the microbiological examination of dry market pulp, paper, and paperboard, is broadly based on ISO 4833 1although the conditions are not identical. However, it provides specific amplification where necessary. It is intended for the

22、 estimation of colony-forming units, CFU, aerobic bacteria, and bacterial spores. Because of the exacting techniques required in aseptic procedures, reproducible good quality results can only be ensured by skilled microbiological technicians. ISO 2014 All rights reserved vBS ISO 8784-1:2014BS ISO 87

23、84-1:2014Pulp, paper and board Microbiological examination Part 1: Enumeration of bacteria and bacterial spores based on disintegration 1 Scope This part of ISO 8784 specifies a method for determining the total number of colony-forming units of bacteria and bacterial spores in dry market pulp, paper

24、 and paperboard after disintegration. The enumeration relates to specific media. 2 Normative references The following documents, in whole or in part, are normatively referenced in this document and are indispensable for its application. For dated references, only the edition cited applies. For unda

25、ted references, the latest edition of the referenced document (including any amendments) applies. ISO 186:2002, Paper and board Sampling to determine average quality ISO 7213:1981, Pulps Sampling for testing ISO 638:2008, Paper, board and pulps Determination of dry matter content Oven-drying method

26、3 T erms a nd definiti ons For the purposes of this document, the following terms and definitions apply. 3.1 bacteria microscopic, single-celled organisms that possess a prokaryotic type of cell structure, which reproduce by fission and are able to grow under the test conditions specified in this pa

27、rt of ISO 8784 3.2 bacterial spores highly resistant, dormant structures EXAMPLE Endospores from certain genera of bacteria. 3.3 total bacterial count number of colony-forming units (CFU) of bacteria and bacterial spores formed after incubation in a standard culture medium, under the test conditions

28、 specified in this part of ISO 8784 3.4 spore count number of colony-forming units (CFU) of bacterial spores formed after incubation in a standard culture medium, under the test conditions specified in this part of ISO 8784 INTERNATIONAL ST ANDARD ISO 8784-1:2014(E) ISO 2014 All rights reserved 1BS

29、ISO 8784-1:2014ISO 8784-1:2014(E) 4 Principle This poured plate method involves enumeration of colonies in a standard culture medium. A fibre suspension, prepared from paper, paperboard, or pulp samples, is plated in agar. Two parallel determinations are made. For enumeration of bacterial spores, th

30、e fibre suspension is heated for 10 min at 80 C prior to plating. The plates are incubated at 32 C for 48 h. The total numbers of bacteria or bacterial spores are enumerated by counting the colonies formed in the agar. The mean value of 2 parallel determinations is calculated and the results are exp

31、ressed as the number of CFU per gram of sample. 5 Culture media and diluents 5.1 General All substrates and diluents shall be appropriately sterilized. When preparing the culture medium, make sure that the ingredients are completely dissolved by mixing while heating prior to dispensing into suitable

32、 containers for sterilization. See ISO 11133 2for quality assurance and guidelines on preparation and production of culture media. 5.2 Water When water is mentioned in a formula, use distilled water or purified water, see ISO 11133 2 . 5.3 Culture media for total bacteria count and spore count Cultu

33、re medium shall be prepared as follows, or from commercially available dehydrated culture media according to the manufacturers instructions. Ready-to-use medium may be used when its composition is comparable to that given in this part of ISO 8784. To test the performance of the medium, see ISO 11133

34、 2 . Plate count agar (PCA) composition per litre:Tryptone 5,0 gYeast Extract 2,5 gDextrose 1,0 gAgar 15,0 gWater 1 000 mlFinal pH 7,0 0,2 If PCA is not available, Tryptone glucose extract (TGE) agar may be used (see A.3). The use of TGE as an alternative culture medium is acceptable if it gives com

35、parable results as the standard culture medium. The culture medium used shall be stated in the test report (see Clause 12). 5.4 Diluents Ringers solution (see A.1) is preferred, although other isotonic solutions may be used. Ringers tablets are commercially available. To facilitate the release of ce

36、lls from the fibres, it is recommended to add 20 l of Tween 80 (see A.2) per litre to the Ringers solution prior to sterilization by autoclaving. The diluent used and if Tween 80 has been added, shall be stated in the test report (see Clause 12).2 ISO 2014 All rights reservedBS ISO 8784-1:2014ISO 87

37、84-1:2014(E) 6 Apparatus and equipment 6.1 General All laboratory equipment and parts of the equipment in direct contact with the sample and the diluent or the culture medium shall be sterilized. NOTE For advice on standard microbiological equipment, see ISO 7218 4 . 6.2 List of equipment 6.2.1 Use

38、ordinary microbiological laboratory equipment, and the following. 6.2.2 Suitable wrapping material, e.g. aluminium foil (non-coated and inert), ready-to-use envelopes of different sizes or self-closing plastic bags, all of which are commercially available. 6.2.3 Disintegrator, high speed electrical

39、blender with metal (preferably stainless steel) or glass cup that can be sterilized. NOTE Other homogenizing system with equivalent efficiency may be used. 6.2.4 Incubator, capable of maintaining a constant temperature of 32 C 2 C. 6.2.5 Petri dishes, having a diameter of 90 mm (standard) or 140 mm

40、to 150 mm (alternative). 6.2.6 Pipettes, of wide-mouth type suitable volume. The width of the mouth must be large enough so that a 1 % fibre suspension can easily be drawn into the pipette tip. NOTE A suitable volume is 10 ml or 50 ml. 6.2.7 Water bath, capable of maintaining a temperature of 80 C 2

41、 C. 6.2.8 Colony-counting equipment or magnifying device, with a magnification between 1,5 and 2,5 shall be used. NOTE The use of an additional lens might be necessary to increase the magnification, up to 10 , to facilitate the counting of pin-point bacterial colony-forming units and also to ensure

42、that no other particles except bacterial colonies are counted (see Clause 10). 6.2.9 Balance, with an accuracy of 0,01 g. 6.2.10 Sterilizing unit, an autoclave capable of sterilization at 121 C. 7 Sampling Make sure that the sampling procedure is performed using aseptic techniques. If the sample is

43、to represent a lot of paper or paperboard, the sampling shall be in accordance with ISO 186:2002. From each unit of paper or paperboard to be sampled, cut several top layers and discard them to eliminate surface contamination. Use a sterile knife to cut through several layers of the paper or board s

44、ample, producing a stack of sheets. Discard the top sheet. ISO 2014 All rights reserved 3BS ISO 8784-1:2014ISO 8784-1:2014(E) If the sample is to represent a lot of pulp, the sampling shall be in accordance with ISO 7213:1981. From each unit of dry market pulp to be sampled, discard several top shee

45、ts from each bale to eliminate surface contamination. In other cases, sample a sufficient number of units so that the test material is representative of the paper, the paperboard, or the dry market pulp to be tested. In all sampling and examination procedures, make sure that the test material taken

46、is representative of the sample received. Ide a l ly, a s a mple shou ld c ont a i n at le a s t fou r she e t s , e ac h of t hem h av i n g a m i n i mu m s i z e of 20 0 m m 250 m m of dry market pulp, paper, or paperboard (at least 2 sheets for testing and 2 protective sheets). NOTE For paperboa

47、rd or thicker material, it might be sufficient to use only 1 sheet for each parallel determination. For thinner paper, more than 2 sheets can be used for each parallel determination. After sampling, wrap the unexposed test material in suitable wrapping material (6.2.2). 8 Preparation of the test mat

48、erial 8.1 General Preferably, conduct the procedure under aseptic conditions. A laminar flow hood is recommended for plating. Unwrap the test material under aseptic conditions. Remove the protective sheets on the top and bottom of the sample stack without touching the test sheets in the centre of th

49、e sample stack. The procedure in 8.3 and 8.4 shall be repeated for the 2 parallel determinations. 8.2 Determination of dry-matter content If the result is to be reported on a dry-mass basis, determine the dry-matter content of the test material, X, in accordance with ISO 638:2008. If the result is to be reported on an “as received”-mass basis (not on a dry-mass basis), omit the determination of dry-matter content and report accordingly see 11.1 and Clause 12 j). 8.3 Weighi