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1、J IS JAPANESE I N D USTR IAL STANDARD Translated and Published by Japanese Standards Association JIS K 0350-40-10 :2002 (JIWA/ JSA) Testing method for enumeration of total bacteria in industrial water and wastewater ICs 13.060.70 Reference number : JIS K 0350-40-10 : 2002 (E) PROTECTED BY COPYRIGHT
2、6 s Copyright Japanese Standards Association Provided by IHS under license with JSALicensee=IHS Employees/1111111001, User=Wing, Bernie Not for Resale, 03/12/2007 08:27:10 MDTNo reproduction or networking permitted without license from IHS -,-,- K 0350-40-10 : 2002 Foreword This translation has been
3、 made based on the original Japanese Industrial Standard established by the Minister of Economy, Trade and Industry through deliberations at the Japanese Industrial Standards Committee according to the proposal of establishing a Japanese Industrial Standard from the Japan Industrial Water Associatio
4、n (JIWA)/the Japanese Standards Association (JSA), with a draft of Industrial Standard based on the provision of Article 12 Clause 1 of the Industrial Standardization Law. JIS K 0350 consists of the following four parts. JIS K 0350-10-10 Testing methods for standard plate counts of hetero- trophic b
5、acteria in industrial water and wastewater JIS K 0350-20-10 Testing methods for detection and enumeration of coliform organisms in industrial water and waste- water JIS K 0350-30-10 Testing methods for detection and enumeration of heterotrophic bacteria in industrial water and waste- water JIS K 035
6、0-40-10 Testing method for enumeration of total bacteria in industrial water and wastewater Date of Establishment: 2002-03-20 Date of Public Notice in Official Gazette: 2002-03-20 Investigated by: Japanese Industrial Standards Committee Standards Board Technical Committee on Environment and Recyclin
7、g Policy JIS K 0350-40-10 : 2002, First English edition published in 2002-12 Translated and published by: Japanese Standards Association 4-1-24, Akasaka, Minato-ku, Tokyo, 107-8440 JAPAN In the event of any doubts arising as to the contents, the original JIS is to be the final authority. O JSA 2002
8、All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying and microfilm, without permission in writing from the publisher. Printed in Japan PROTECTED BY COPYRIGHT Copyright
9、Japanese Standards Association Provided by IHS under license with JSALicensee=IHS Employees/1111111001, User=Wing, Bernie Not for Resale, 03/12/2007 08:27:10 MDTNo reproduction or networking permitted without license from IHS -,-,- JAPANESE INDUSTRIAL STANDARD JIS K 0350-40-10 : 2002 Testing method
10、for enumeration of total bacteria in industrial water and wastewater 1 Scope This Japanese Industrial Standard specifies testing method for enumeration of total bacteria in industrial water and wastewater. 2 Normative references The standards given in Attached Table 1 contain pro- visions which, thr
11、ough reference in this Standard, constitute provisions of this Standard. The most recent editions of the standards (including amendments) shall be applied. 3 Definitions For the purposes of this Standard, the definitions shall be in ac- cordance with JIS K 0101, JIS K 0102, JIS K 0550 and JIS K 0211
12、, and the fol- lowing definition shall apply. 3.1 sample. total bacteria Total bacteria refer to all bacteria which are contained in the 4 Matters in common Matters in common shall be as follows: 4.1 accordance with JIS K 0050. General rules The general matters common to chemical analysis shall be i
13、n 4.2 Water Water used in this Standard shall be A2 water or A3 water(1) speci- fied in JIS K 0557. Carry out the filtration sterilization according to 4.4 d) before use. Note (1) The metal-made still shall not be used. Reagents When the name of reagent is specified, the most excellent grade specifi
14、ed in JIS shall be used, and when there is no reagent specified in SIS, the reagent giving no hindrance to the test shall be used. The mass concentration of the solution of reagents shall generally be expressed in the unit of g/L or mgL. Water to be used for the preparation of reagents shall be that
15、 of 4.2. The names of reagents shall be coordinated as much as possible with those men- tioned in the nomenclature of compounds specified by the Chemical Society of Japan and in JIS reagents, which are based on the nomenclature system for inorganic compounds and for organic compounds prescribed by I
16、nternational Union of Pure and Applied Chemistry (IUPAC). Care shall be sufficiently taken in accordance with the related laws and regula- tions for treating reagents and wastewater and the like. Sterilizing procedures of implement and the like Sterilizing procedures of implement and the like shall
17、be carried out as follows: PROTECTED BY COPYRIGHT Copyright Japanese Standards Association Provided by IHS under license with JSALicensee=IHS Employees/1111111001, User=Wing, Bernie Not for Resale, 03/12/2007 08:27:10 MDTNo reproduction or networking permitted without license from IHS -,-,- 2 K 0350
18、-40-10 : 2002 a) b) C d) 4.5 Dry heat sterilization This is used made implements. Sterilize them at 1 h. High-pressure steam sterilization for sterilization of glass-made and metal- approximately 170 “C for approximately This is used for sterilization of culture media, dilution water, a sample conta
19、iner, culture media that has been used, etc. Sterilize them at 121 “C for 15 min to 20 min. Flame sterilization This is used for the sterilization of a test tube and the mouth part of a flask. Sterilize them by keeping the mouth part aslant in flame and turning for a while, after and before cultivat
20、ing operation. Filtration sterilization This is used for the sterilization of water or solu- tion. Sterilize it by filtration with the filter apparatus as shown in the filter (see Fig. 2) in 6.2 d) using 0.2 pm in pore diameter and approximately 50 mm in diameter of the membrane filter(2). Note (2)
21、Wrap the membrane filter to be used in aluminium foil (or parchment paper, etc.) in advance, place it in a glass petri dish, and carry out the high-pressure steam sterilization in b) on it, Or commercial prod- ucts which have been sterilized may be used. For handling a mem- brane Filter, the pincett
22、e in 6.2 b) shall be used. Disinfection operations Disinfection operations shall be carried out as fol- lows: a) b) 4.6 Before and after test operation, disinfect fingers and a testing bench. Use cresol soap solution (10 g/L), cationic soap solution (1 g/L to 10 g/L), or alcohol for dis- infection (
23、which is specified in the Japanese Pharmacopoeia) or ethanol (80 vol%) ethanol (95) specified in JIS K 31021, for disinfection of Fingers. For the disinfection of a testing bench, either spray cationic soap solution (10 g/L), alcohol for disinfection, ethanol (80 vol%) or the like, or wipe with the
24、cloth moistened with these solutions. Employed implements such as a pipette, a sample container, a water sampler, and so on, shall be immersed in disinfection solution such as cresol soap solu- tion (30 g/L to 50 g/L) and the like for 1 day (or shall be sterilized by high-pres- sure steam), and wash
25、 with water sufficiently until disinfection solution is completely removed. Glass ware implements For glass ware implements, those which are speci- fied in JIS R 3503 and JIS R 3505 shall generally be used. However, when special implements are required, one example of them for each item shall be ill
26、ustrated or ex- plained. Moreover, when heating operations are involved, the borosilicate glass-1 specified in JIS R 3503 shall be used. 5 Sample 5.1 sampler. Sampling Sampling shall be carried out using a sample container or water 5 . 1 . 1 Reagents The reagents shall be as follows: PROTECTED BY CO
27、PYRIGHT Copyright Japanese Standards Association Provided by IHS under license with JSALicensee=IHS Employees/1111111001, User=Wing, Bernie Not for Resale, 03/12/2007 08:27:10 MDTNo reproduction or networking permitted without license from IHS -,-,- 3 K 0350-40-10 : 2002 a) Glutaraldehyde solution l
28、75-pentanedio1 (glutaraldehyde) solution for an elec- tron microscope. Information : A commercial product is available. Glutaraldehyde solution of approximately 20 % is easy to handle. 5.1.2 Implements Implements shall be as follows: Water sampler A Heyroth water sampler(3). The water sampler, put i
29、n a portable box, shall be sterilized by dry heat sterilization in 4.4 a ) . Fig. 1 shows an example. Note (3) For sampling from the depth at which sampling with a Heyroth water sampler is difficult, a Vandorn water sampler may be used. In this case, transfer the sample to the sample container in b)
30、. I I I I I I I I I I , I ! ,A- . A : Glass-made sample B : Stopper C : Chain D : Chain for stopper opening E : Chain for metal fittings of bottle holder plate F : Bottle holder plate G : Portable box H : Cover of portable box I : Weight container (100 ml) Fig. 1 An example of a Heyroth water sample
31、r Sample container 100 ml glass bottle with a ground stopper. Carry out dry heat sterilization in 4.4 a) or high-pressure steam sterilization in 4.4 b) on the sample ontainer() whose stopper part and neck part have been covered with aluminium foil (or parchment paper), etc. Alternatively, a steriliz
32、ed polyethyl- ene bottle for bacteria test may be used. Take care so that the sample con- tainer is not subjected to contamination until sampling begins. Note (4) When sampling the water which contains oxidizing material as residual chlorine, place 20 mg to 30 rng of sodium thiosulfate pentahydrate
33、(pow- dered) specified in JIS K 8637 into the sample container, and then Ster- ilize it by high-pressure steam method in 4.4 b) or by Oxirane (ethylene oxide) method. The commercially available sterilized polyethylene bottle for bacte- ria test filled with sodium thiosulfate beforehand may be used.
34、PROTECTED BY COPYRIGHT Copyright Japanese Standards Association Provided by IHS under license with JSALicensee=IHS Employees/1111111001, User=Wing, Bernie Not for Resale, 03/12/2007 08:27:10 MDTNo reproduction or networking permitted without license from IHS -,-,- 4 K 0350-40-10 : 2002 5.1.3 Operati
35、ons Sampling shall be carried out as follows: Sampling of surface water When the surface water in lake and marsh, river, waterway, overflow, water tank, etc. can be directly sampled, take the sample into a sample container. On this occasion, care shall be taken so that the wa- ter touched with hands
36、 and fingers is not sampled. When the surface water can not be sampled directly, take it with a water sampler(3). Sampling water at definite depth Take the water at a definite depth using a water sampler(3). Sampling from water tap When the material of water tap is proof against the flame sterilizat
37、ion, sterilize a water tap by the flame sterilization in 4.4 c) in advance of opening the tap, and after discharging the water in the pipe suf- ficiently, take water into a sample container. When the flame sterilization is not practical, remove the dirt on the periph- ery and inside of water tap, an
38、d disinfect with ethanol (80 vol%), etc. in ad- vance. Open the tap and after discharging the water in the pipe sufficiently, take water into a sample container. Sampling from piping and apparatus Similarly to c), take water into a sample container. Handling of sample Carry out the test immediately
39、after sampling. When the test is not carried out immediately, add glutaraldehyde solution so that its con- centration becomes 0.1 vol% to 1.0 vol% and preserve it in a dark place at O O C to 5 O C (do not freeze). 6 Testing method For test of total bacteria, apply the method of counting those which
40、produce the pale or orange color fluorescence at the excitation wavelength of 460 nm to 490 nm as total bacteria by staining DNA of bacteria with 3,6-bis (dimethylamino) acridine (acridine orange) or those which produce the pale fluores- cence at the excitation wavelength of 330 nm to 380 nm as tota
41、l bacteria by staining DNA of bacteria with 4 , 6-diamidino-2-phenylindole (DAPI). 6.1 Reagents Reagents shall be as follows: 6.1.1 Water In accordance with 4.2. 6.1.2 Staining reagents Staining reagents shall be as follows: a) Acridine orange solution (10 g/L) After dissolving 1 g of acridine orang
42、e(5) in 100 ml of water, carry out the filtration sterilization according to 4.4 d). DAPI solution (1 g/L) After dissolving 0.1 g of 4 , 6-diamidino-2-phenylindole di-hydrochloride (DAPII(5) in 100 ml of water, carry out the filtration steriliza- tion according to 4.4 d ) . Transfer this solution in
43、to a sterilized brown bottle and preserve it in a cold and dark place. Note ( 5 ) Since this is suspected of carcinogenicity, care shall be taken so that it does not stick to the skin, clothes, etc. b) PROTECTED BY COPYRIGHT Copyright Japanese Standards Association Provided by IHS under license with
44、 JSALicensee=IHS Employees/1111111001, User=Wing, Bernie Not for Resale, 03/12/2007 08:27:10 MDTNo reproduction or networking permitted without license from IHS -,-,- 6 . 1 . 3 scope. 5 K 0350-40-10 : 2002 Non-fluorescent oil immersion oil Non-fluorescent oil for fluorescence micro- Information : Co
45、mmercial products are available. Implements and apparatus Implements and apparatus &all be as follows: Measuring pipette 1 ml to 10 ml. Put it into a pipette sterilizer with its tip first, or wrap in aluminium foil or parchment paper, and carry out the dry heat sterilization according to 4.4 a ) . P
46、incette The tip shall be flat and smooth. Carry out the flame sterilization according to 4.4 c) immediately before using. Membrane filter (black) The black-colored membrane filter made of poly- carbonate of 0.2 pm in pore diameter, approximately 25 mm in diameter. Filter (separate type) An example s
47、hall be shown in Fig. 2. Wrap each part of the filter with aluminium foil (or parchment paper, etc.) and carry out the sterilization which is suitable for the materials. Filter in details Filter in details (in the case of filtration sterilization) collection of bacteria) (in the case of A : Upper fi
48、lter pipe Bi : Membrane filter Bi! : Membrane filter (black) C : Membrane filter support D : Lower filter pipe E : Rubber stopper (silicone) F : Metal clamp G : Suction bottle Fig. 2 An example of a filter (separate type) Slide glasses The standard type (76.0 mm X 26.0 mm) of quality class 2 specifi
49、ed in JIS R 3703. Cover glasses The standard type (24 mm x 24 mm or 32 mm x 24 mm) of quality class 2 specified in JIS R 3702. Incident-light fluorescence microscope Eyepiece grid micrometer Stage micrometer PROTECTED BY COPYRIGHT Copyright Japanese Standards Association Provided by IHS under license with JSALicensee=IHS Employees/11