Tiêu chuẩn iso 06887 4 2017

© ISO 2017 Microbiology of the food chain — Preparation of test samples, initial suspension and decimal dilutions for microbiological examination — Part 4 Specific rules for the preparation of miscell[.]

Microbiolog y ofthe food chain —

Part 4:

Specific rules for the pr eparation of

Micro iolo ie de la ch îne alimentaire — Pré aration de s

éch ntillo s,de la sus pension mè e et de s d utions décimale s en v e

de l’e xamen micro iolo ique —

Partie 4 : Règle s spécifique s p ur la pré aration de produits v riés

COPYRIGHT PROTECTED DOCUMENT

© ISO 2017, P blshed in Sw itz rlan

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written permis ion Permis ion c n be req esed from either ISO at the ad r es below or ISO’s member bod y in the c u try of

F reword v

1 Sc ope 1

2 Nor mati ve r eferenc es 2

3 Terms and definitions 2

4 Principle 2

5 Diuents 2

5.1 Basic mate ials 2

5.2 Diuents for g eneral use 2

5.2.1 Peptone salt olution 2

5.2.2 Bufer ed peptone wate 2

5.3 Diuents for spe ial purp ses 3

5.3.1 Double-s r eng th bufer ed peptone wate 3

5.3.2 P osphate bufer ed diuent 3

5.4 Dis ribution an s e i zation of the diuent 3

5.5 Pe formanc tes ing of diuents 3

5.6 Enzyme solutions 3

5.6.1 A lpha-am ylase solution 3

5.6.2 Celulase solution 4

5.6.3 Pa ain solution 4

6 A pparatus 4

7 S mpl n and sample types 5

8 Preparatio of samples 5

8.1 General 5

8.2 A cidic pr od ct 5

8.3 High-fat fo ds, exclu ing mar garines an spreads (e.g o e 20 % of total mas as fat) 6

8.4 Har d an dry pr od ct 6

9 Specific pr oc edures 6

9.1 Dehydrated an low a w prod ct 6

9.1.1 General 6

9.1.2 A ppar atus 6

9.1.3 Pr ep ration of samples .7

9.1.4 Pr ep ration of initial suspension 7

9.1.5 Resuscitation 8

9.1.6 Wate activity 8

9.2 Flours, c r eal g rains an b -prod ct an animal fe ds 9

9.3 Gelatine (powder ed and leaf)

1 9.3.1 Pr ep ration of samples 1

9.3.2 Pr ep ration of initial suspension 1

9.4 Mar garine an spr eads 1

9.4.1 Samplng 1

9.4.2 Pr ep ration of tes sample 1

9.5 Eg g s an eg g pr oduct 1

9.5.1 Fr esh whole eg g s 1

9.5.2 Microflor a of whole eg g shel 1

9.5.3 Inte nal microflor a 1

9.5.4 Bulk whole lq id eg g , eg g white an eg g y lk 1

9.5.5 Dehydrated whole eg g and dried eg g white 1

9.5.6 Whole eg g micr oflora ( hel plusy lk plus white) 1

9.6 Bake y g oods, p stry and cakes 1

9.6.1 General 1

9.6.2 Pr ep ration of samples 1

9.7 Fr esh fruit an veg eta les(pr e-p cked) 1

9.7.1 Sampleprep r ation of multi-comp nent pr od ct 1

9.7.2 Pr e-packed prod ct of one type of fruit or veg eta le .1

9.8 Fermented prod ct or othe pr od cts containing via le micr oor ganisms 1

9.8.1 General 1

9.8.2 Diuent 14

9.9 Beverag es (alcoholc an non-alcoholc drin s an b tled wate s, stil or car bonated) 14 9.9.1 General 14

9.9.2 De-gas ing b inve sion an mix ing 14

9.9.3 De-gas ing using ultrasou d 14

9.1 A lte native protein pr od ct (co ked inse ts, tex tur ed veg eta le pr otein or myco r otein) 14

9.1 1 General 14

9.1 2 C oked inse t 14

9.1 3 Tex tur ed veg eta le pr otein an myco r otein 1

10 F rthe di utio s 15

Biblog raph y 16

ISO (he Int ernational Org nization for Stan ardization) is a worldwide fede ation of national s an ards

b dies (ISO membe b dies) The work of pr p ring Int ernational Standards is normaly car ied out

through ISO t ech ical committ ees Each membe b dy int er st ed in a subje t for w hich a t ech ical

committ ee has be n es a l shed has the right t o be r pr sent ed on that committ ee Int ernational

org nizations, g ove nmental an non-g ove nmental, in laison with ISO, also take part in the work

ISO cola orat es closely with the Int ernational Ele trot ech ical C mmis ion (IEC) on al matt ers of

ele trot ech ical s an ardization

The proc d r s used t o develo this document an those int en ed for it furthe maint enanc ar

desc ibed in the ISO/IEC Dir ctives, Part 1 In p rticular the dife ent a pro al c it eria ne ded for the

dife ent ty es of ISO document should be not ed This document was draft ed in ac ordanc with the

edit orial rules of the ISO/IEC Dir ctives, Part 2 ( e www iso org dir ctives)

A tt ention is drawn t o the p s ibi ity that ome of the element of this document ma be the subje t of

p t ent right ISO shal not be held r sponsible for identifying any or al such pat ent right Detais of

any p t ent right identif ied d ring the develo ment of the document wi be in the Introd ction an / r

on the ISO ls of p t ent de larations r c ived ( e www iso org p t ent )

Any trade name used in this document is information given for the convenienc of use s an does not

cons itut e an en orsement

For an ex lanation on the meaning of ISO spe ific terms an ex r s ions r lated to conformity as es ment,

as wel as information a out ISO’s adhe enc to the Wor ld Trade Org nization (WTO) principles in the

Te h ical Bar ie s to Trade (TBT) se the folowing URL: www iso org / iso/ for word html

The committ ee r sp nsible for this document is ISO/TC 34, Fo d produc ts , Subcommitt ee SC 9,

Microbiolog y ofthe food chain — Pr eparation oftest

Part 4:

WARNING — The use of this document may inv lve hazardous materials, o eratio s and

equipment It is the respo sibiity of the use of this document to establ sh ap ro riate safety

and he lth practices an to determine the ap l cabi ity of re ulatory l mitatio s before use

This document spe if ies rules for the pr p ration of samples an diutions for the mic o iolo ical

ex mination of spe if ic fo d prod ct not co e ed in othe p rt of ISO 6 8 , w hich deal with mor

g ene al cat eg ories This document co e s a wide rang e of misc laneous prod ct , but does not inclu e

new prod ct brought on t o the market aft er publcation

ISO 6 8 -1 defines the g ene al rules for the pr paration of the initial suspension an diutions for

mic o iolo ical ex mination

This document ex clu espr p ration of samples for b th en me ation an det ection t es methods w hen

pr paration detais ar spe ified in the r lev nt Int ernational Stan ards

This document is a plca le t o the folowing prod ct :

— acidic (low pH) prod ct ;

— hard an dry prod ct ;

— dehydrat ed, fr e e-dried an othe low a

wprod ct ( inclu ing those with inhibit ory pro e ties);

— flours, w hole c r al grains, c r al b -prod ct ;

— animal fe d, cat le cake, kib les and pet chew s;

— g elatine (p wde ed an leaf);

— marg rines, spr ads an non-dairy prod ct with ad ed wat er;

— eg s an eg prod ct ;

— bakery g oods, p s ries an cakes;

— fr sh fruit an veg eta les;

— fe ment ed prod ct an othe prod ct containing via le mic o rg nisms;

— alcoholc an non-alcoholc beve ag es;

— alt ernative prot ein prod ct

2 Normati ve referenc es

The folowing document ar r fe r d t o in the t ext in such a wa that some or al of their cont ent

cons itut es r q ir ment of this document F or dat ed r fe enc s, only the edition cit ed a ples F or

un at ed r fe enc s, the lat es edition of the r fe enc d document ( inclu ing any amendment ) a ples

ISO 68 7-1, Mic ro iolo y o the fod chain — Prep ration o tes t s amples , initial s us pens io a d dec imal

diutio s fr mic ro iolo ic al ex min tio — Part 1: G ene al rules fr the pr p ratio o the initial

s us pens io a d dec imal diutio s

ISO 7 1 , Mic ro iolo y o f od a d a imal fe din s tufs — G ene al r q ir ments a d g ida ce fr

mic ro iolo ic al ex min tio s

3 Terms and definitions

F or the purposes of this document, the t erms an definitions given in ISO 6 87-1 a ply

ISO an IEC maintain t erminolo ical data ases for use in s an ardization at the folowing ad r s es:

— IECEle tro edia: a aia le at ht p:/ www ele tro edia org

— ISO Onlnebrow sing plat orm:a ai a le at ht ps:/ www iso org o p /

4 Principle

The g ene al principles for sample pr p ration an subseq ent st eps ar detai ed in ISO 68 7-1 This

document desc ibes spe if ic measur s for prod ct not co e ed in othe part of ISO 6 8

An initial suspension is pr par d t o o tain as uniform a dis ribution as p s ible of the mic o rg nisms

contained in the t es p rtion

F or det ection methods, the pr -enrichment or enrichment suspension is pr p r d in the same wa ,

using the medium r commen ed in the method of ex mination conc rned F or en me ation methods

used for al fo d ty es co e ed in this document, g ene al diuent ar document ed in the r lev nt

clauses of ISO 6 8 -1 an spe if ic diuent ar inclu ed in Cla use 5

If ne es ary, furthe diutions ar pr p r d in orde t o r d c the n mbe of mic o rg nisms pe u it

v lume t o alow, aft er incub tion, o se v tion ofany growth ( in the case of lq id media) or colonies ( in

the case of a ar plat es or a ar tubes) ass at ed in each spe ific s an ard

5.1 Basic materials

R efe t o ISO 68 7-1

5.2 Di uents for g eneral use

5.2.1 P epto e salt solutio

R efe t o ISO 68 7-1

5.2.2 Bufered pepto e water

R efe t o ISO 68 7-1

5.3 Di uents for special pur poses

5.3.1 Double-streng th bufered peptone water

5.3.1.1 Compositio and pr eparation

R efe t o ISO 68 7-1

5.3.1.2 A pplcatio

Use this di uent for highly acidic prod ct of pH ≥ 3,5 t o pH ˂ 4,5 t o o tain an initial suspension of pH

7,0 ± 0,5

NOTE B fered pept one water (5.2.2)is suff icient or prod cts of pH 4,5an a ove

5.3.2 Phosp ate bufer ed diuent

9,0 g

Potas ium dihydrog en phosphat e (KH

2PO

4

5.3.2.2 Pr eparatio

Dis olve the comp nent in thewat er, b heating if ne es ary

If ne es ary, adjus the pH so that, aft er st eri zation, it is pH 7,0 ± 0,2 at 2 °C

5.3.2.3 A pplcatio

Phosphat e bufe ed solution is used as a diuent or g elatine (9.3)

5.4 Distribution and ster il zation of the di uent

5.6.1.2 Preparatio

Dis olve the α-amy lase in the wat er an st eri iz the solution b p s ing through a 0,2 µm membrane

filt er The enzyme solution can be st or d for up t o 1 month at 5°C ±3 °Cor up t o 3 months at ≤− 20 °C

Final composition of the α-amy lase solution ma ne d t o be adjust ed depen ing on the enzymatic

activity of the comme cial α-amy lase used and thethickening pro e ties of the t es sample

5.6.1.3 A pplcatio

This enzyme solution is ad ed at the rat e of 1 ml t o 1 0 0 ml of diuent (1 % v lume fraction) t o

improve solubi ity of swel ng s ar h prod ct , c r als an c r al-containing prod ct (9.1.4.3)

5.6.2 Celulase solutio

5.6.2.1 Compositio

5.6.2.2 Preparatio

Dis olve the c lulase in the wat er an st eri z the solution b p s ing through a 0,2 µm membrane

filt er The enzyme solution can be st or d for up t o two we ks at 5 °C ± 3°C or upt o 1 month at ≤− 20 °C

NOTE Use of fresh solution wil ensure ma imum enzyme activity

5.6.2.3 A pplcatio

This enzyme solution is ad ed at the rat e of 1 ml t o 1 0 0 ml of diuent (1 % v lume fraction) t o

improve solubi ity of carb x methy l c lulose, locus beans, caro , guar an cas ia gums(9.1.4.3)

Dis olve the p p in in the wat er an st eri iz the solution b p s ing through a 0,2 µm membrane filt er

The enzyme solution can be st or d for up t o 1 month at 5 °C ± 3 °C

NOTE Use of fresh solution wil ensure ma imum enzyme activity

5.6.3.3 A pplcatio

This enzyme solution is ad ed at the rat e of 2 ml t o 1 0 0 ml of diuent (2 % v lume fraction) t o

improve solubi ity of g elatine (9.1.4.3 and 9.3)

6 A pparatus

Usual mic o iolo ical la orat ory eq ipment for g ene al use ( e ISO 7 1 an ISO 6 8 -1) an , in

p rticular, the folowing

6.1 Homog enizer

6.1.1 Rotar y homog enizer (blen e )

R efe t o ISO 7 1 If a larg e sample is t o be homog eniz d, the eq ipment should inclu e a st erie 1 l b w l

6.1.2 P eristaltic homog enizer

R efe t o ISO 7 1 W ith st erie plas ic b gs or f ilt er b gs t o r tain p rticulat e mat erial w he e ne es ary

6.2 Domestic g rater , s e ie

6.3 Hammer or other heavy implement, ca a leof c ushing hard mate ials

6.4 Wate baths, ca a le of being maintained at 44 °C to 47°Cor as s ated for spe ific purp ses

6.5 Ster ile s is or s, kni ves, s alpels an forc eps

6.6 Ster ile spatulas, spoo s or sc oo s

6.7 Ster ile c or er s, for taking samples at depth

6.8 Stir r er, ca a le of o erating with a horizontal motion

6.9 Ster ile w ide-necked flasks or othe c ontainer , of 50 ml ca acity

6.10 Ultraso ic bath, with o erating fr eq ency of 3 MHz to 45 MHz

7 Sampl ng and sample types

Car y out samplng in ac ordanc with the spe ific s an ard a pro riat e t o the prod ct conc rned or

se ISO/TS 1 7 8 Some guidanc on samplng c rtain prod ct is inclu ed in Cla use 9 for clarity If a

spe if ic s an ard is not a ai a le, it is r commen ed that a r ement be r ached on this subje t b the

p rties conc rned

8 Preparation ofsamples

Al pr p rations an manipulations shal be car ied out using aseptic t ech iq es an st eri e eq ipment

( e ISO 7 1 ) Gene al sample pr paration proc d r s ar given in ISO 68 7-1, but ad itional detai for

some cat eg ories is given in 8.2 t o 8.4

8.2 Acidic pr oducts

It isimp rtant t o conside the en use of the prod ct w hen t es ing acidic samples

If the prod ct is t o be used as an ingr dient in a f inal prod ct of highe pH, then the pH of the initial

suspension of the t es portion shal be adjust ed t o pH 7,0 ± 0,5 with the diuent spe if ied at 5.2.2 or

5.3.1 or othe s with eq iv lent bufe ing ca acity

F or pH adjus ment of mode at ely acidic samples (pH ≥ 3,5 t o pH < 4,5), use double-s r ngth bufe ed

pept one wat er (5.3.1) S e ISO 6 87-1

If highly acidic (pH < 3,5) samples (e.g low pH fruit an juic s or vineg rs an pickles) ar t est ed for

acid-t ole ant an acido hi c sp iag e org nisms using a pro riat e media, the pH of such samples shal

not be adjust ed

8.3 High-fat foods, e xc luding margar ines and spreads ( e.g over 20 % of total mas as fat)

A diuent with betwe n 1 g l an 1 g l of p ly sorb t e 8 [p lyox ethy lene (2 ) sorbitan mono leat e ]

ac ording t o the es imat ed fat cont ent shal be used t o impro e emulsification d ring suspension (e.g

for a fat cont ent of 40 %, ad 4 g l)

Alt ernative surfactant an emulsif ie s ar a aia le u de v rious trade names, but hepro ortions t o

use should be det ermined b the la orat ory

8.4 Hard and dry pr oducts

Do not homog eniz hard or dry prod ct in a rotary homog eniz r (6.1.1)for mor than 2,5 min at a time

t o a oid ex ces ive heating that ma damag e the mic o rg nisms pr sent

Homog eniz dry an hard het erog eneousprod ct b mincing or grin ing the la orat ory sample Av id

ex ces ive heating d ring this proc s b homog enizing for pe iods of no mor than 1 min at a time,

with suita ler s int erv lsa pled, depen ing on the prod ct being proc s ed Minc or grin u ti the

sample is visibly homog eneous

R esuscitation at la orat ory ambient t empe atur (1 °Ct o 2 °C)for up t o 1 h is r commen ed t o as is

in the r co e y of s r s ed org nisms from al hard an dry prod ct

9 Specific pr ocedures

9.1 Dehydrated and low a

wproducts

9.1.1 General

The folowing ar r g rded as dehydrat ed prod ct :

— dehydrat ed meat an veg eta les;

— dried soups, b ui on cubes an gra y mix es;

— powde ed beve ag es (t ea, coco an coco -b sed prod ct , cofe , dehydrat ed fruit juic );

— raw c lulose, soluble c lulose, dextrin, sorbit ol, sug rs, glucose, glutamat e;

— he bs, spic s, fla ourings an colourings;

— poly sac haride g el ing ag ent , alginat es, gums, et c

— cocon t, p rtialy dehydrat ed veg eta le /yeas /meat/f ish extract ;

— chocolat e an confe tione y (b rs or sweet );

— dehydrat ed w hole eg an dried eg w hit e;

— c r als, flours, animal fe ds;

— powde ed or peletiz d via le mic o rg nisms(e.g yeas s for b ke y)

9.1.2 A pparatus

Plas ic b gs with f ilt er inse t (6.1.2) ar r commen ed t o as is in the pipet ing of prod ct with

subs antial insoluble matt er in suspension