EN ISO 5667-3:2003

SLOVENSKI STANDARD
01-februar-2004
1DGRPHãþD
SIST EN ISO 5667-3:1996
.DNRYRVWYRGH±9]RUþHQMH±GHO1DYRGLOR]DVKUDQMHYDQMHLQUDYQDQMH]Y]RUFL
YRGH,62
Water quality - Sampling - Part 3: Guidance on the preservation and handling of water
samples (ISO 5667-3:2003)
Wasserbeschaffenheit - Probenahme - Teil 3: Anleitung zur Konservierung und
Handhabung von Proben (ISO 5667-3:2003)
Qualité de l'eau - Echantillonnage - Partie 3: Lignes directrices pour la conservation et la
manipulation des échantillons d'eau (ISO 5667-3:2003)
Ta slovenski standard je istoveten z: EN ISO 5667-3:2003
ICS:
13.060.45 Preiskava vode na splošno Examination of water in
general
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

EUROPEAN STANDARD
EN ISO 5667-3
NORME EUROPÉENNE
EUROPÄISCHE NORM
December 2003
ICS 13.060.45 Supersedes EN ISO 5667-3:1995
English version
Water quality - Sampling - Part 3: Guidance on the preservation
and handling of water samples (ISO 5667-3:2003)
Qualité de l'eau - Echantillonnage - Partie 3: Lignes Wasserbeschaffenheit - Probenahme - Teil 3: Anleitung zur
directrices pour la conservation et la manipulation des Konservierung und Handhabung von Wasserproben (ISO
échantillons d'eau (ISO 5667-3:2003) 5667-3:2003)
This European Standard was approved by CEN on 21 November 2003.
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 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 Management Centre has the same status as the official
versions.
CEN members are the national standards bodies of Austria, Belgium, Czech Republic, Denmark, Finland, France, Germany, Greece,
Hungary, Iceland, Ireland, Italy, Luxembourg, Malta, Netherlands, Norway, Portugal, Slovakia, Spain, Sweden, Switzerland and United
Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION
EUROPÄISCHES KOMITEE FÜR NORMUNG
Management Centre: rue de Stassart, 36  B-1050 Brussels
© 2003 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 5667-3:2003 E
worldwide for CEN national Members.

CORRECTED  2004-03-03
Foreword
This document (EN ISO 5667-3:2003) has been prepared by Technical Committee ISO/TC 147
"Water quality" in collaboration with Technical Committee CEN/TC 230 "Water analysis", the
secretariat of which is held by DIN.
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 June 2004, and conflicting national
standards shall be withdrawn at the latest by June 2004.
This document supersedes EN ISO 5667-3:1995.
According to the CEN/CENELEC Internal Regulations, the national standards organizations of
the following countries are bound to implement this European Standard: Austria, Belgium, Czech
Republic, Denmark, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy,
Luxembourg, Malta, Netherlands, Norway, Portugal, Slovakia, Spain, Sweden, Switzerland and
the United Kingdom.
Endorsement notice
The text of ISO 5667-3:2003 has been approved by CEN as EN ISO 5667-3:2003 without any
modifications.
NOTE Normative references to International Standards are listed in annex ZA (normative).
Annex ZA
(normative)
Normative references to international publications
with their relevant European publications
This European Standard incorporates by dated or undated reference, provisions from other
publications. These normative references are cited at the appropriate places in the text and the
publications are listed hereafter. For dated references, subsequent amendments to or revisions of
any of these publications apply to this European Standard only when incorporated in it by
amendment or revision. For undated references the latest edition of the publication referred to
applies (including amendments).
NOTE Where an International Publication has been modified by common modifications, indicated
by (mod.), the relevant EN/HD applies.
Publication Year Title EN Year
ISO 3696 1987 Water for analytical laboratory use - EN ISO 3696 1995
Specification and test methods
ISO 5667-1 1980 Water quality - Sampling - Part 1: Guidance EN 25667-1 1993
on the design of sampling programmes
ISO 5667-2 1991 Water quality - Sampling - Part 2: Guidance EN 25667-2 1993
on sampling techniques
ISO 5667-16 1998 Water quality - Sampling - Part 16: EN ISO 5667-16 1998
Guidance on biotesting of samples
INTERNATIONAL ISO
STANDARD 5667-3
Third edition
2003-12-15
Water quality — Sampling —
Part 3:
Guidance on the preservation and
handling of water samples
Qualité de l'eau — Échantillonnage —
Partie 3: Lignes directrices pour la conservation et la manipulation des
échantillons d'eau
Reference number
ISO 5667-3:2003(E)
©
ISO 2003
ISO 5667-3:2003(E)
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ii © ISO 2003 — All rights reserved

ISO 5667-3:2003(E)
Contents Page
Foreword. iv
Introduction . vi
1 Scope. 1
2 Normative references . 1
3 Preservation of samples. 1
4 Recommendations . 8
5 Identification of samples. 9
6 Transport of samples. 9
7 Reception of samples . 9
Annex A (informative) Dutch investigation on prolonged preservation times. 28
Bibliography . 31

ISO 5667-3:2003(E)
Foreword
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
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-governmental, 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.
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 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 document may be the subject of patent
rights. ISO shall not be held responsible for identifying any or all such patent rights.
ISO 5667-3 was prepared by Technical Committee ISO/TC 147, Water quality Subcommittee SC 6, Sampling
(general methods).
This third edition cancels and replaces the second edition (ISO 5667-3:1994), which has been technically
revised.
ISO 5667 consists of the following parts, under the general title Water quality — Sampling:
 Part 1: Guidance on the design of sampling programmes
 Part 2: Guidance on sampling techniques
 Part 3: Guidance on the preservation and handling of water samples
 Part 4: Guidance on sampling from lakes, natural and man-made
 Part 5: Guidance on sampling of drinking water and water used for food and beverage processing
 Part 6: Guidance on sampling of rivers and streams
 Part 7: Guidance on sampling of water and steam in boiler plants
 Part 8: Guidance on the sampling of wet deposition
 Part 9: Guidance on sampling from marine waters
 Part 10: Guidance on sampling of waste waters
 Part 11: Guidance on sampling of groundwaters
 Part 12: Guidance on sampling of bottom sediments
 Part 13: Guidance on sampling of sludges from sewage and water-treatment works
 Part 14: Guidance on quality assurance of environmental water-sampling and handling
iv © ISO 2003 — All rights reserved

ISO 5667-3:2003(E)
 Part 15: Guidance on preservation and handling of sludge and sediment samples
 Part 16: Guidance on biotesting of samples
 Part 17: Guidance on sampling of suspended sediments
 Part 18: Guidance on sampling of groundwater at contaminated sites
 Part 19: Guidance on sediment sampling in marine areas
ISO 5667-3:2003(E)
Introduction
This part of ISO 5667 is intended to be used in conjunction with ISO 5667-1 and ISO 5667-2, which deal with
the design of sampling programmes and sampling techniques respectively.
vi © ISO 2003 — All rights reserved

INTERNATIONAL STANDARD ISO 5667-3:2003(E)

Water quality — Sampling —
Part 3:
Guidance on the preservation and handling of water samples
1 Scope
This part of ISO 5667 gives general guidelines on the precautions to be taken to preserve and transport all
water samples including those for biological analyses but not those intended for microbiological analysis.
These guidelines are particularly appropriate when spot or composite samples cannot be analysed on-site and
have to be transported to a laboratory for analysis.
2 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.
ISO 3696:1987, Water for analytical laboratory use — Specification and test methods
ISO 5667-1:1980, Water quality — Sampling — Part 1: Guidance on the design of sampling programmes
ISO 5667-2:1991, Water quality — Sampling — Part 2: Guidance on sampling techniques
ISO 5667-14:1998, Water quality — Sampling — Part 14: Guidance on quality assurance of environmental
water sampling and handling
ISO 5667-16:1998, Water quality — Sampling — Part 16: Guidance on biotesting of samples
ISO Guide 34:2000, General requirements for the competence of reference material procedures
3 Preservation of samples
3.1 General considerations
Waters, particularly fresh waters, waste waters and groundwaters, are susceptible to changes as a result of
physical, chemical or biological reactions which may take place between the time of sampling and the
commencement of analysis. The nature and rate of these reactions are often such that, if precautions are not
taken during sampling, transport and storage (for specific determinands), the concentrations determined may
be different to those existing at the time of sampling.
The extent of these changes is dependent on the chemical and biological nature of the sample, its
temperature, its exposure to light, the nature of the container in which it is placed, the time between sampling
and analysis, and the conditions to which it is subjected, for example agitation during transport. Further
specific causes of variation are as follows.
ISO 5667-3:2003(E)
a) The presence of bacteria, algae and other organisms can consume certain constituents of the samples.
These organisms can also modify the nature of the constituents to produce new constituents. This
biological activity affects, for example, the concentrations of dissolved oxygen, carbon dioxide and
compounds, of nitrogen, phosphorus and sometimes silicon.
b) Certain compounds can be oxidized by the dissolved oxygen present in the samples or by atmospheric
oxygen (e.g. organic compounds, Fe (II) and sulfides).
c) Certain substances can precipitate out of solution [for example calcium carbonate, metals and metallic
compounds such as Al(OH) ] or be lost to the vapour phase (for example oxygen, cyanides and mercury).
d) The pH and conductivity can be modified and the dissolved carbon dioxide changed by the absorption of
carbon dioxide from air.
e) Dissolved metals or metals in a colloidal state, as well as certain organic compounds can be irreversibly
adsorbed onto the surface of the containers or solid materials in the samples.
f) Polymerized products can depolymerize and conversely, simple compounds can polymerize.
Changes to particular constituents vary both in degree and rate, not only as a function of the type of water, but
also, for the same water type, as a function of seasonal conditions.
lt should be emphasized that these changes are often sufficiently rapid to modify the sample considerably in a
short time. In all cases, it is essential to take precautions to minimize these reactions and, in the case of many
determinands, to analyse the sample with a minimum of delay.
Preservation of water samples is necessary for a number of reasons, therefore it is generally necessary to
choose, from the various possible methods of preservation, a method that does not introduce contamination.
Fresh waters and groundwaters can be stored more successfully. In the case of potable waters, storage can
be solved easily by cooling, because these waters are less susceptible to biological and chemical reactions.
In many cases, if samples are analysed within 24 h, the preservation technique of cooling to between 1 °C to
5 °C is sufficient. Municipal or industrial sewage plant effluents should be preserved immediately after
sampling, because of the high biological activities in these samples.
This part of ISO 5667 describes the most commonly used preservation techniques and storage times.
[4]
In spite of investigations which have been carried out in order to recommend methods that enable water
samples to be stored without changes occurring to their composition, no guidance has been reported that
covers all situations. Users of particular test methods and analytical techniques described in International
Standards prepared by ISO/TC 147 are encouraged to take into account any relevant guidance offered in this
part of ISO 5667 when making decisions in relation to sample preservation and handling for such methods
and techniques.
3.2 Precautions to be taken
3.2.1 Container selection
The choice of sample container is of major importance and ISO 5667-2 provides some guidance on this
subject. Details of the type of container used for the collection and storage of samples are given in Tables 1
to 4. The same considerations given to this selection of suitable container material should also be given to the
selection of cap-liner materials. The guidance given here is to help in the selection of containers for general
use.
The containers used to collect and store the samples should be selected after taking into account the following
predominant criteria (especially when the analytes are present in trace quantities).
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ISO 5667-3:2003(E)
a) Minimizing sample contamination by the container or cap material, for example leaching of inorganic
constituents from glass (especially soda glass) and organic compounds and metals from plastics. Some
coloured caps may contain significant levels of heavy metals.
b) Ability to clean and treat the walls of the container to reduce surface contamination by trace constituents
such as heavy metals or radionuclides.
c) Chemical and biological inertness of the container or cap material in order to prevent or minimize reaction
between sample constituents and the container.
d) Containers may also cause changes to constituent concentrations by adsorption or absorption of analytes.
Trace metals are particularly susceptible to these effects but other analytes (for example detergents,
pesticides, phosphates) may also be affected.
Guidance should be sought from laboratory staff on the selection of sample containers and sampling
equipment.
Other factors should also be considered, e.g. resistance to temperature extremes, resistance to breakage,
ease of sealing and reopening, size, shape, mass, availability, cost, potential for cleaning and re-use.
Container blanks should always be taken, preserved and analysed as a check on the suitability of the
container and preservation procedures (see ISO 5667-14).
3.2.2 Container preparation
3.2.2.1 General
All preparation procedures should be validated to ensure positive or negative interferences do not occur. As a
minimum, this should include the analysis of:
a) blanks;
b) samples containing known levels of relevant analytes.
If disposable or single-use containers cannot be used, it is preferable to reserve a set of containers for a
particular determinand, thereby minimizing risks of cross-contamination. Care should be taken to prevent a
container, formerly holding a sample with a high concentration of a determinand, from contaminating a
subsequent sample containing a low concentration of the same determinand.
It may be necessary to wash new containers with water containing a detergent, in order to remove dust and
residues of packing materials, followed by thorough rinsing with water of an appropriate quality. The use of
cleansing reagents and solvents may cause interferences, e.g. residual contamination by phosphate-
containing detergents when undertaking nutrient analyses. If used, all cleaning reagents and solvents should
be of an appropriate quality. For the determination of silicon, boron and surfactants, detergents should not be
used for cleaning purposes.
3.2.2.2 Detergent-washed plastic or glass containers
The procedure should be as follows.
a) Wash the container and cap with a dilute solution of detergent and water.
b) Rinse thoroughly with tap water.
c) Successively rinse twice with water of an appropriate quality.
d) Drain thoroughly and replace cap.
Automatic dish washing machines may be used for this procedure.
ISO 5667-3:2003(E)
3.2.2.3 Solvent-washed glass containers
WARNING — Organic solvents may be hazardous. Provide suitable handling facilities and handle with
care.
The procedure should be as follows.
a) Wash the container and cap with a dilute solution of detergent and tap water.
b) Rinse thoroughly with tap water.
c) Successively rinse twice with water of an appropriate quality and dry.
d) Rinse with acetone of an appropriate quality and drain.
e) Rinse with a suitable solvent of an appropriate quality, dry and immediately replace cap.
The solvent should be compatible with the analytes of interest and the analytical method to be used.
3.2.2.4 Acid-washed containers in plastic or glass
The procedure should be as follows.
a) Wash the container and cap with a dilute solution of detergent and tap water.
b) Rinse thoroughly with tap water.
c) Rinse with an aqueous 10 % nitric acid solution.
d) Drain and completely fill with an aqueous 10 % nitric acid solution.
e) Cap and store for at least 24 h.
f) Empty the container, rinse with water of an appropriate quality and immediately replace cap.
Some manufactures will supply containers with a certificate of cleanliness. Such containers may not need
further cleaning or rinsing, provided the manufacturer supplies the containers with caps attached.
Automatic hot acid washers may be used for this procedure.
3.2.3 Filling the container
For samples requiring the determination of physico-chemical determinands, fill the container completely and
stopper it in such a way that there is no air space above the sample. This reduces interaction with the gas
phase, and minimizes agitation of the sample during transport.
Where samples are frozen as part of their preservation, sample containers should not be completely filled
(see 3.2.6).
3.2.4 Handling and preservation of samples for biological examination
The handling of samples for biological examination is different to that for samples requiring chemical analysis.
The addition of chemicals to the sample for biological examination can be used for either fixation or
preservation of the sample. The term “fixation” is used to describe the protection of morphological structures,
while the term “preservation” is used for the protection of organic matter from biochemical or chemical
degradation. Preservatives, by definition, are toxic and the addition of preservatives may lead to the death of
living organisms. Prior to death, irritation may cause the most delicate organisms, which do not have strong
cell walls, to collapse before fixation is complete. To minimize this effect, it is important that the fixation agent
4 © ISO 2003 — All rights reserved

ISO 5667-3:2003(E)
enters the cell quickly. Some preservatives, for instance acid solutions of Lugol, may lead to the loss of some
taxonomical groups of organisms, which can be a problem during certain parts of the year in certain areas.
This can be addressed by using an additional preservative, such as alkaline solutions of Lugol, during, for
example, the summer period when the appearance of silico-flagellates may be frequently observed.
The preservation of samples for biological examination should meet the following criteria:
a) the effect of the preservative on the loss of the organism should be known beforehand;
b) the preservative should effectively prevent the biological degradation of organic matter at least during the
storage period of the samples;
c) the preservative should enable the taxonomical groups of organisms to be adequately studied during the
storage period of the samples.
3.2.5 Handling and preservation of samples for radiochemical analysis
WARNING — Safety precautions and shielding depend on the activity of the sample.
There is little difference between the handling of samples for radiochemical analysis and the handling of
samples for physico-chemical analysis. Safety precautions depend on the nature of the radioactivity of the
sample. The preservation techniques for these samples depend on the type of emitter and the half-life of the
radionuclide of interest.
3.2.6 Cooling or freezing of samples
The cooling or freezing of samples is only effective if the process is applied immediately after the collection of
the samples. This necessitates the use of cool-boxes or refrigerators at the sampling location. Wherever a
temperature is given for cooling, the temperature of the sample environment is meant (not the temperature of
sample itself).
Simple cooling of the sample (in melting ice or in a refrigerator at a temperature between 1 °C and 5 °C) and
storage of the sample in the dark is, in most cases, sufficient to preserve the sample during transport to the
laboratory. Cooling cannot be considered as a means of long-term storage, particularly in the case of
wastewater samples (see Table 1). The sample should be kept and stored at a temperature lower than that
observed during the process of collection or filling of the container.
A small volume of ice does not have much cooling effect upon a large volume of warm water. Where a sample
contains determinands that are likely to be affected by biological activity, and where preservation on-site is not
possible, the temperature of the sample should be taken immediately on arrival at the laboratory. This is
particularly important when samples require transporting for several hours. Samples should be analysed or
cooled immediately at receipt in the laboratory. During transport, the temperature of the cooling system should
be monitored.
In general, storage of samples at temperatures below − 20 °C allows the samples to be stored for longer
periods of time. If samples are to be frozen, the container should be made of plastic and not be filled
completely. This reduces the risk to the sample container from being damaged. For some analytes, such as
nutrient determinands, freezing of the sample is the preferred method of preservation. In these cases, quick-
freezing with dry ice is a satisfactory procedure. The freezing of samples is not an appropriate procedure for
samples requiring analysis of volatile substances or if samples contain cells or bacteria or microalgae, which
can fracture and lose cell constituents during the freezing process. Nevertheless, it is necessary to control the
freezing and thawing technique in order to return the sample to its initial equilibrium after thawing. In this case,
the use of plastic containers (for example polyvinyl chloride or polyethene) is strongly recommended. For
thawing of samples, see ISO 5667-16.
3.2.7 Filtration or centrifugation of samples
Suspended matter, sediment, algae and other micro-organisms may be removed, either at the time of taking
the sample or immediately afterwards, by filtering the sample through membrane filter material (e.g. paper,
ISO 5667-3:2003(E)
polytetrafluoroethylene, glass) or by centrifuging. Filtration is, of course, not applicable if the membrane filter is
likely to retain one or more of the constituents to be analysed. It is equally essential that the membrane filter
assembly system not be a cause of contamination and be carefully washed before use, but in a manner
consistent with the final method of analysis.
Alternatively, the reason for filtering the sample may be to enable the proportion of soluble and insoluble forms
of an analyte to be determined (e.g. soluble and insoluble metal fractions).
Decanting the sample is not recommended as an alternative to filtration.
Membrane filters should be used with caution as various heavy metal compounds and organic material may
be adsorbed on the membrane filter surface, and soluble compounds (e.g. surfactants) within the membrane
filter can be leached out into the sample.
3.2.8 Addition of preservatives
Certain physical and chemical constituents can be stabilized by the addition of selective chemical compounds,
either directly to the sample after it has been taken, or beforehand, to the empty container.
Particular reagents, necessary for the specific preservation of certain constituents (e.g. the determination of
oxygen, total cyanides and sulfides) require the sample to be preserved on-site.
It is essential that the preservatives used do not interfere with the analysis; tests intended to check their
compatibility are necessary in case of doubt. Any dilution of the sample with added preservative solutions
should be taken into account during the analysis and calculation of results. It is preferable that the addition of
preservatives to samples be made using concentrated solutions so that only small volumes are used. In most
cases, this enables the corresponding dilution to be disregarded. The use of solid preservatives, for example
sodium hydroxide, is to be avoided as local heating may occur, adversely affecting the sample.
The fact that the addition of these agents can modify or change the chemical or physical nature of the
constituents means that these changes are not incompatible with the purpose of later determinations. For
example, acidification can solubilize colloidal constituents or solids, and should therefore be used with caution
if the aim of the analysis is the determination of dissolved constituents and then only for that purpose.
Filtration of the sample prior to the addition of preservative is essential for dissolved ions. Similarly, caution
should be applied if the aim of the analysis is to determine the toxicity of the sample to aquatic animals, as
certain components, particularly heavy metal compounds, are more toxic in the ionic form. Samples should
therefore be analysed as soon as possible.
It is essential to carry out a blank test, particularly in determinations for trace elements, to take into account
the possible introduction of an additional quantity of the determinand (for example acids can introduce a
significant amount of arsenic, lead and mercury) by the preservatives. In such cases, samples of the
preservatives used for the treatment of the water samples should be retained for use in the preparation of
blank tests.
3.3 Reagents
WARNING — Certain preservatives (e.g. acids, alkalis, formaldehyde) need to be used with caution.
Sampling personnel should be warned of potential dangers and that appropriate safety procedures
should be followed.
The following reagents are used for the preservation of samples and shall only be prepared according to
individual sampling requirements. Unless otherwise specified, all reagents used should be of at least
analytical reagent grade and water should be of at least ISO 3696:1987 Grade 2 purity. Acids referred to in
this part of ISO 5667 are the commercially available “concentrated” acids.
All reagents should be labelled with a “shelf life” which should not be exceeded. The “shelf life” represents the
period for which the reagent is suitable for use, if stored correctly. Any reagents that are not completely used
by the expiry of the “shelf life” date should be discarded.
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ISO 5667-3:2003(E)
Check the reagent dispensers periodically and discard any reagent where dispensers are shown to be
unsuitable.
Between field trips, reagents should be stored in clean, secure cabinets in order to prevent contamination.
It is essential that all samples requiring the same determinand determination be preserved together.
Each sample should be labelled accordingly after the addition of the preservative, since usually there may be
no visible indication as to which samples have been preserved and which have not.
3.3.1 Solids
.
3.3.1.1 Sodium thiosulfate pentahydrate, Na S O 5H O.
2 2 3 2
3.3.1.2 Ascorbic acid, C H O .
6 8 6
3.3.1.3 Sodium hydroxide, NaOH.
3.3.1.4 Potassium dichromate, K Cr O
2 2 7.
3.3.1.5 Copper sulfate, CuSO
4.
3.3.1.6 Sodium tetraborate, Na B O ⋅10H O.
2 4 7 2
3.3.1.7 Hexamethylenetetramine (hexamine, urotropine), C H N
6 12 4.
3.3.2 Solutions
3.3.2.1 Zinc acetate solution (ρ = 0,10 g/ml), C H O Zn.
4 6 4
3.3.2.2 Orthophosphoric acid (ρ = 1,7 g/ml), H PO
3 4.
3.3.2.3 Hydrochloric acid (ρ = 1,16 g/ml), HCl.
3.3.2.4 Nitric acid (ρ = 1,42 g/ml), HNO .
3.3.2.5 Sulfuric acid (8 mol/l), H SO .
2 4
3.3.2.6 Sodium hydroxide solution (ρ = 0,40 g/ml), NaOH.
3.3.2.7 Formaldehyde solution (volume fraction of 37 %) (Formalin), CH O.
WARNING — Beware of formaldehyde vapours. Do not store large numbers of samples in small work
areas.
3.3.2.8 Aqueous solution of disodium salt of ethylenediaminetetraacetic (EDTA) (ρ = 0,025 g/ml),
C H N Na O ⋅2H O.
10 14 2 2 8 2
3.3.2.9 Ethanol (volume fraction of 96 %).
3.3.2.10 Alkaline Lugol solution, with sodium acetate.
3.3.2.11 Acid Lugol solution, with acetic acid.
ISO 5667-3:2003(E)
3.4 Prolonged storage of samples
The timing of the maximum recommended preservation time before commencement of analysis begins
immediately after the sample has been taken.
For certain regulatory situations, there is a requirement for samples to be retained for a prescribed period of
time. Notwithstanding the analytical implications of this, this legal requirement takes precedence over the
guidance given in this part of ISO 5667.
Where samples are analysed after the maximum recommended preservation time, it is essential that the
results be accompanied with a statement to the effect that the analytical results may not be reflective of the
concentration present at the time of sampling.
In addition, where laboratories report data analysed after the maximum recommended preservation time, it is
essential that the analytical result be accompanied with a statement to the effect that the maximum
recommended preservation time has been exceeded.
Prolonged storage of samples may be appropriate if the laboratory can demonstrate that there is no difference
between the test result obtained after extended preservation time and the test result obtained within the
preservation time given in this part of ISO 5667. Procedures to be used for establishing homogeneity and
stability are outlined in ISO Guide 34.
3.5 General guidance
Staff should not be allowed to smoke near samples; in addition, samples should not be placed near any
source of engine exhaust.
Open samples should neither be placed (e.g. while samples are being filtered or preserved) near a fan or air
conditioner, nor near food and beverages.
Decontamination and cleaning is appropriate if reusable equipment (such as sample scoops) is used between
and during use.
The inner surfaces of bottles or caps should not be touched with fingers or other objects.
It is essential that empty bottles be stored and transported with caps tightly in place.
Extraneous matter should be kept out of sample bottles. If a measurement (such as temperature or pH) needs
to be made outside of a bottle, then a specific container should be used for that purpose, and the sample used
in the measurement discarded. Under no circumstances should a field measurement be made and the sample
returned to the sample container that is then subsequently forwarded to a laboratory for analysis.
Samples should be scrutinized for large particles, such as leaves or detritus and if these are observed, the
sample should be discarded and a new sample taken.
Preservation reagents should be scrutinized as contamination can sometimes be indicated by, for example, a
change in colour. If contamination is suspected, the reagent should be discarded.
4 Recommendations
For samples requiring analysis for certain organic determinands, an initial on-site extraction may be
advantageous. Alternative procedures such as on-site adsorption techniques or on-site headspace collection
may also be employed where appropriate.
As stated in 3.1, it is impossible to give guidance for storage times or the nature of the sample containers for
all preservation techniques. The efficiency of the preservation process depends not only on the constituents
that require analysis and their concentration levels, but also on the nature of the sample. In all cases, it is
essential that the method of storage be compatible with the analytical technique used. One objective of Tables
1 to 4 is to describe the most common preservation techniques.
8 © ISO 2003 — All rights reserved

ISO 5667-3:2003(E)
Further guidance is given in Table 2 on suitable preservation techniques used in conjunction with several
determinands. However, it is not reasonable or logical to combine organic and inorganic determinands,
because of the manner these determinations are treated within the laboratory.
The biological determinands are generally numerous and sometimes vary from one biological species to
another. For this reason, it is impossible to draw up an exhaustive checklist of all the precautions that should
be taken to preserve samples for biological analysis. The information given in Table 3 therefore relates only to
certain determinands generally studied for various animal or plant groups.
It should be noted that before carrying out any detailed study, it is essential to choose the determinands of
interest.
Table 4 gives techniques generally suitable for the preservation of radioactive samples.
There should be no significant statistical difference between the results of samples analysed immediately
following collection and those analysed after preservation. Results should be verified taking into account the
method of analysis and the guidance provided in this part of ISO 5667.
The sample volumes listed in Table 1 represent typical volumes required to perform a single determination on
the sample. Where more than one method is available for a particular determinant, the sample volumes
pertain to the method that requires the maximum sample volume. In some cases, it may be possible to take a
smaller volume of sample; however, this should only be undertaken after consultation with laboratory staff.
For a sample that requires the determination of more than one determinand, sometimes it is necessary to take
several sub-samples in order to meet the requirements of sample preservation. It is essential that extreme
care be taken to avoid cross-contamination which may occur. For example, nitric acid preservation used for a
metal sub-sample will contaminate the sub-sample taken for nitrate analysis.
5 Identification of samples
Sample containers should be labelled in a clear and unambiguous manner that is durable.
Additionally, it may be necessary to note, at the time of sampling, details which will enable a correct
interpretation of the information provided (for example, date and hour of sampling, name of person sampling,
nature and amount of preservatives added). The use of pre-printed labels, forms, etc. can facilitate the
practical attainment of these objectives.
Special samples of anomalous material should be clearly marked and accompanied by a description of the
observed anomaly. It is essential that samples containing hazardous or potentially hazardous materials, for
example acids, be clearly identified as such.
6 Transport of samples
Containers holding samples should be protected and sealed in such a way that samples do not deteriorate
and do not lose any of their constituents during transport. Packaging material should protect the containers
from possible external contamination and breakage, particularly near the opening of the container, and should
not be a source of contamination. During transportation, the samples should be stored according to the
guidance given in Tables 1 to 4. In cases where the storage and transportation time exceeds the maximum
recommended preservation time before commencement of analysis, whether or not the samples should be
analysed should be checked with the client, and if it is decided to proceed with the analysis, the time between
sampling and analysis should be reported.
7 Reception of samples
Laboratory staff should establish whether samples underwent cooling during transportation and if possible
whether a sample environmental temperature between 1 °C to 5 °C was maintained.
In all cases, and especially when a “chain of custody” process needs to be established, the count of sample
containers received in the laboratory should be verified against the number of sample bottles provided for
each sample.
ISO 5667-3:2003(E)
Table 1 — Techniques generally suitable for the preservation of samples —
Physico-chemical and chemical analysis
Maximum
recommended
Typical volume
Determinand to Preservation preservation
a
Type of container (ml) and filling Comments
be studied technique time before
b
technique
analysis after
preservation
c
14 days
Samples should
preferably be
analysed on-site
Cool to
(particularly for
Acidity and
Fill container
between 1 °C
P or G 24 h samples high in
alkalinity
completely to
and 5 °C.
dissolved gases).
exclude air.
Reduction and
oxidation during
storage can change
the sample
Extract sample
1 000
container as part of
Do not pre-rinse
the sample extraction
the empty Acidify to
procedure.
container with between pH 1
If the sample is
G with PTFE cap sample; analytes to 2 with HCl
Acidic herbicides 2 weeks
chlorinated, for each
liner or septum adhere to the wall and cool to
1 000 ml of sample,
of the bottle. between 1 °C
add 80 mg of
and 5 °C.
Do not completely
Na S O ⋅5H O to the
2 2 3 2
fill sample
container prior to
container.
collection.
Acidify to
between pH 1
to 2 with
1 000
HNO , cool to
Fill container
P or G between 1 °C 5 days
Adsorbable
completely to
and 5 °C, keep
organic halides
exclude air.
samples
(AOX)
stored in the
dark.
Freeze to
P 1 000 1 month
- 20 °C.
P acid-washed
Acidify to
Aluminium
100 between pH 1 1 month
G or BG acid-
to 2 with HNO
washed 3
Acidify to
between pH 1
to 2 with
P or G 500 21 days
H SO , cool to
Ammonia, free and 2 4 Filter on-site before
between 1 °C
ionized
preservation
and 5 °C.
Freeze to
P 500 1 month
− 20 °C.
10 © ISO 2003 — All rights reserved

ISO 5667-3:2003(E)
Table 1 (continued)
Maximum
recommended
Typical volume
Determinand to Preservation preservation
a
(ml) and filling Comments
Type of container
be studied technique time before
b
technique
analysis after
preservation
Cool to
Filter on-site before
P or G 500 between 1 °C 24 h
Anions (Br, F, Cl,
preservation.
and 5 °C.
NO , NO , SO and
2 3 4
See also
PO )
Freeze to
ISO 10304-1.
P 500 1 month
– 20 °C
Acidify to HCl should be used if
P acid-washed
between pH 1 the hydride
Antimony
100 1 month
to 2 with HCl technique is used for
G acid-washed
or HNO analysis.
3 .
HCI should be used
Acidify to pH 1
P acid-washed
if the hydride
Arsenic 500 to 2 with HCl 1
...