1、raising standards worldwideNO COPYING WITHOUT BSI PERMISSION EXCEPT AS PERMITTED BY COPYRIGHT LAWBSI Standards PublicationBS ISO 21338:2010Water quality Kineticdetermination of the inhibitoryeffects of sediment, othersolids and coloured sampleson the light emission of Vibriofischeri (kinetic lumines
2、centbacteria test)BS ISO 21338:2010 BRITISH STANDARDNational forewordThis British Standard is the UK implementation of ISO 21338:2010.The UK participation in its preparation was entrusted to TechnicalCommittee EH/3/5, Biological Methods.A list of organizations represented on this committee can beobt
3、ained on request to its secretary.This publication does not purport to include all the necessaryprovisions of a contract. Users are responsible for its correctapplication. BSI 2010ISBN 978 0 580 56634 9ICS 13.060.60Compliance with a British Standard cannot confer immunity fromlegal obligations.This
4、British Standard was published under the authority of theStandards Policy and Strategy Committee on 31 August 2010.Amendments issued since publicationDate Text affectedBS ISO 21338:2010Reference numberISO 21338:2010(E)ISO 2010INTERNATIONAL STANDARD ISO21338First edition2010-07-15Water quality Kineti
5、c determination of the inhibitory effects of sediment, other solids and coloured samples on the light emission of Vibrio fischeri (kinetic luminescent bacteria test) Qualit de leau Dtermination cintique des effets inhibiteurs des chantillons de sdiment, autres solides et des chantillons colors sur l
6、a luminescence de Vibrio fischeri (essai cintique de bactries luminescentes) BS ISO 21338:2010ISO 21338:2010(E) PDF disclaimer This PDF file may contain embedded typefaces. In accordance with Adobes licensing policy, this file may be printed or viewed but shall not be edited unless the typefaces whi
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11、org Published in Switzerland ii ISO 2010 All rights reservedBS ISO 21338:2010ISO 21338:2010(E) ISO 2010 All rights reserved iiiContents Page Foreword iv Introduction.v 1 Scope1 2 Normative references1 3 Terms and definitions .2 4 Principle2 5 Interferences 3 6 Reagents and materials 4 7 Apparatus.5
12、8 Sampling and sample pre-treatment .5 9 Procedure.6 10 Evaluation.7 11 Expression of results9 12 Criteria of validity 11 13 Test report11 Annex A (informative) Precision data .13 Annex B (informative) Typical kinetic curves from different samples 17 Annex C (informative) Dilution series .18 Bibliog
13、raphy20 BS ISO 21338:2010ISO 21338:2010(E) iv ISO 2010 All rights reservedForeword ISO (the International Organization for Standardization) is a worldwide federation of national standards bodies (ISO member bodies). The work of preparing International Standards is normally carried out through ISO te
14、chnical 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-governmental, in liaison with ISO, also take part in the work. ISO collaborates close
15、ly with the International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization. International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2. The main task of technical committees is to prepare International Standards. Draft
16、 International Standards adopted by the technical committees are circulated to the member bodies for voting. Publication as an International Standard requires approval by at least 75 % of the member bodies casting a vote. Attention is drawn to the possibility that some of the elements of this docume
17、nt may be the subject of patent rights. ISO shall not be held responsible for identifying any or all such patent rights. ISO 21338 was prepared by Technical Committee ISO/TC 147, Water quality, Subcommittee SC 5, Biological methods. BS ISO 21338:2010ISO 21338:2010(E) ISO 2010 All rights reserved vIn
18、troduction The method specified in this International Standard is a kinetic modification of the luminescent bacteria test specified in ISO 11348. The kinetic method overcomes the problems arising from intense colour and turbidity in the samples. There is no need for sedimentation or centrifugation o
19、f turbid samples, or for the correction of colour as described in ISO 11348. This kinetic method uses luminometers capable of dispensing luminescent bacteria to the samples and measuring the luminescent intensity over a period of time. In the method, the bacterial suspension is dispensed and mixed w
20、ith the sample in the measurement chamber of the luminometer. Several suitable instruments are commercially available, but only a few of them are capable of cooling the measurement chamber to (15 1) C as specified in ISO 11348. However, if the bacterial suspension and test samples are kept at (15 1)
21、 C in a thermo-block before the measurement and during the whole incubation, the actual temperature during the contact time is (15 1) C. The measurements specified in this International Standard can be carried out using freshly prepared bacteria, as well as freeze- or liquid-dried bacterial preparat
22、ions. The various bacterial preparations can deliver different results, especially in the presence of heavy metals (see ISO 11348). The laboratories responsible for the results have the opportunity to select the most suitable bacterial preparation based on expert judgement and information about the
23、samples to be tested. BS ISO 21338:2010BS ISO 21338:2010INTERNATIONAL STANDARD ISO 21338:2010(E) ISO 2010 All rights reserved 1Water quality Kinetic determination of the inhibitory effects of sediment, other solids and coloured samples on the light emission of Vibrio fischeri (kinetic luminescent ba
24、cteria test) WARNING Persons using this International Standard should be familiar with normal laboratory practice. This International Standard does not purport to address all of the safety problems, if any, associated with its use. It is the responsibility of the user to establish appropriate safety
25、 and health practices and to ensure compliance with any national regulatory conditions. IMPORTANT It is absolutely essential that tests conducted in accordance with this International Standard be carried out by suitably trained staff. 1 Scope This International Standard specifies the kinetic direct-
26、contact method for determining the inhibitory effect of suspensions of sediment and other solid samples, and also for problematic turbid or coloured aqueous samples on the light emission of the marine bacterium Vibrio fischeri (NRRL B-11177). This method is applicable to: a) sediment samples and wat
27、er suspensions of sediments (fresh water, brackish, and seawater sediments); b) effluents (especially turbid and coloured); c) aqueous extracts (e.g. leachates, eluates, elutriates) of soil, solid waste, and other solid material (especially turbid and coloured). 2 Normative references The following
28、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. ISO 5667-16:1998, Water quality Sampling Part 16: Guidance on b
29、iotesting of samples ISO 5814, Water quality Determination of dissolved oxygen Electrochemical probe method ISO 11348-1, Water quality Determination of the inhibitory effect of water samples on the light emission of Vibrio fischeri (Luminescent bacteria test) Part 1: Method using freshly prepared ba
30、cteria ISO 11348-2, Water quality Determination of the inhibitory effect of water samples on the light emission of Vibrio fischeri (Luminescent bacteria test) Part 2: Method using liquid-dried bacteria ISO 11348-3:2007, Water quality Determination of the inhibitory effect of water samples on the lig
31、ht emission of Vibrio fischeri (Luminescent bacteria test) Part 3: Method using freeze-dried bacteria BS ISO 21338:2010ISO 21338:2010(E) 2 ISO 2010 All rights reserved3 Terms and definitions For the purpose of this document, the following terms and definitions apply. 3.1 contact time duration of con
32、tact between one object or substance and another NOTE In the test, the contact time is the time available to control or sample for contact with the test bacteria. 3.2 control sample sample used in a laboratory in order to check or monitor the instrument or measurement performance or to monitor chang
33、es in a sample under investigation 3.3 correction factor dimensionless multiplier to correct data for known influences affecting their values as measured NOTE In the test, the correction factor, fkt, serves to correct the initial luminescence intensity of the sample. 3.4 peak value maximum signal re
34、corded in response to a stimulus NOTE In the test, the peak value is the maximum signal which is recorded immediately after all the bacteria are in contact with the sample. 3.5 reference sample when the effect or behaviour of a substance is known from previous tests (reference substance) and when th
35、is substance is examined within the framework of a test series as test sample, this is called the reference sample NOTE Adapted from ISO 5667-16:1998. 3.6 test sample test sample is made from the sample by means of various preparatory steps specific to the sample and the test, e.g. by dissolving, ho
36、mogenizing, sedimenting, filtering, neutralizing or aeration ISO 5667-16:1998 4 Principle The inhibition of light emission by cultures of Vibrio fischeri is measured kinetically by following the light emission of cultures from the very beginning of the assay. This is accomplished by dispensing the l
37、uminescent bacteria suspension into the sample in a cuvette or other suitable vessel (e.g. microtiter plate) already in the measuring position in the luminometer. The light emission is measured and recorded from the moment of dispensing of the bacterial suspension to the sample until the maximum val
38、ue has been reached and not only at the maximum value of intensity (peak value) which usually occurs within 5 s of mixing, and after a contact time of 15 min and 30 min or optionally 5 min (Figure 1). BS ISO 21338:2010ISO 21338:2010(E) ISO 2010 All rights reserved 3Key t time y relative light units
39、1 start measurement 2 inject bacteria 3 record peak value from 0 s to 5 s 4 mix the sample before recording signal at 30 min Figure 1 Principal schematic protocol for the kinetic luminescent bacteria test Vibrio fischeri suspension is dispensed and mixed into the sample in the measurement chamber of
40、 the luminometer. The test criterion is the decrease of the luminescence at each endpoint compared to the peak value, taking into account a correction factor, fkt , which is measured from intensity changes of control samples during the exposure time. The inhibitory effect of the sample can be determ
41、ined as the lowest ineffective dilution (LID) value, or as effective concentration (EC20or EC50) values by means of dilution series (e.g. as described in Annex C). The LID value is the most concentrated test batch tested at which the inhibition of light emission is 3 mg/l is required for the test. I
42、f the oxygen concentration of the undiluted sample is less than 3 mg/l, use appropriate methods to oxygenate the sample, e.g. aeration or stirring. Measure the pH of all samples and sample water suspensions. If the pH is between 6,0 and 8,5, no adjustment is necessary. The adjustment of the pH value
43、, however, may alter the nature of the sample. On the other hand, the pH of the sample and the pH of the test batch can differ because of the buffer capacity of the test medium. It may be necessary to carry out tests on both the pH-adjusted and the non-pH-adjusted samples. If necessary, adjust the p
44、H of the samples by adding either hydrochloric acid (6.4) or sodium hydroxide solution (6.3). Depending on the purpose of the test, the pH may be adjusted to 7,0 0,2 or to the upper (8,5 0,2) or lower (6,0 0,2) limits. Choose the concentration of the hydrochloric acid or the sodium hydroxide solutio
45、n to restrict the volume added to not more than 5 % of total volume. Add 20 g of sodium chloride per litre to the sample or to the neutralized sample. For salt water samples, ISO 11348-3:2007, Annex D gives further information. 9 Procedure 9.1 Initial preparations 9.1.1 Preparation of test solutions
46、 Prepare the reference samples according to 6.6. Test each batch of bacteria after delivery with all three reference substances. Test at least one of the three reference substances with each vial of stock suspension reconstituted. Prepare the samples according to 8.2. Prepare, in the first set of te
47、st tubes (7.5), the sample dilution series, the reference sample (6.6) and the control (6.2) required. Serially dilute the sample. Make the dilutions in separate tubes and transfer them to the measurement cuvettes or plates. Normally, in this test, equal volumes of test suspension and sample (or dil
48、uted sample) are mixed, giving tested dilution levels within a series of D W 2. The minimum dilution which can be tested is 1,00 in 1,25, prepared by mixing 4 volumes of sample with 1 volume of test suspension (e.g. 800 l sample plus 200 l test suspension). The corresponding D value is 1. For this D
49、 value, an extra control batch is needed which is made up by adding 200 l of the test suspension to 800 l sodium chloride solution (see Annex C). Maintain the test tubes containing the sodium chloride solution (6.2) for controls, the reference samples (6.6), the samples (8.2) and the samples of the dilution series (Annex C) at (15 1) C. Choose test conditions which ensure that the maximum temperature deviation in the thermo-block within one test is no more than 0,3 C. BS ISO 21338:2010I
