References Materials for Chemical Analysis || Introduction

Reference Materials for Chemical Analysis Certification, Availability, and Proper Usage

Edited by Markus Stoeppler, Wayne R. WOK PeterJjenks

0 Wiley-VCH Verlag GmbH, 2001

I’ 1

Introduction Edited by Markus Stoeppler

1.1 Historical

Markus Stoeppler and HumphtyJM Bowen

This Chapter reviews some selected historical examples of the development, production and use of reference materials (RMs), from the past century up to the present. It is, of necessity, a rather short and incomplete review describing international efforts in this area. From the references given at the end of this Chapter and at the end of Chapter 3, the reader may investigate further into the past.

1.1.1 Early Developments

The history of reference materials is closely linked with the development of analytical chemistry. In the 19th Century all chemicals were, in comparison with those of today, of poor purity. Thus, for volumetric analysis suitable purified materials as primary standards had to be specified. One of the first examples was the recom- mendation of As(II1) oxide by Gay-Lussac (1824) for this purpose. Somewhat later, Sorensen (1887) proposed criteria for the selection of primary chemical standards. These were further elaborated by Wagner (1903) at the turn of the last century. It is worthwhile mentioning that their criteria were quite similar to those used today.

One of the first attempts to use a biological RM was for the analysis of the fat content of milk. This was carried out in London in the late 1880’s by a number of analpcal chemists who were trying to identify adulterated milk. At that time milk was sold unpackaged and at least 20 % of the milk sold in London was adulterated by dilution with water. This work appears to be the first empirical round-robin approach for characterization of a RM.

In medicine the need for standards was just as acute. Beal (1951) mentioned that the U.S. Pharmacopoeia VI, issued in 1880, took a big step forward by adding tests for purity and quality of the materials described in it, but the use of reference materials as an integral part of the pharmaceutical monographs for drugs did not start until the 1950’s. Another example of activities at the end of the 19th Century was associated with the introduction, by Ehrlich, of the first diphtheria antitoxin and his

2 1 7 Introduction

discovery of Salvarsan (arsphenamine), the first effective cure for syphilis. Whilst these were by no means the first such medicines, Ehrlich was the first to calibrate the purity of the preparations of these substances by bio-assay against an arbitrary standard kept at low temperature (Ehrlich et al. 1894; Ehrlich 1910). Others followed his lead, from 1921 the State Serum Institution of Copenhagen, Denmark, collected a set of international standards for therapeutic agents including diphteria antitoxin as well as arsphenamines. These standards were increased in number after the discovery of vitamins and sex hormones in the 1930’s (e.g. Dale 1939). A main source of these clinical standards was the National Institute of Medical Research in London. In 1948 a total of 35 International Standards were held at this institute (Miles 1948). In the 1960’s the need for reliable quality control of clinical chemistry determinations had been highlighted in the USA and England following the introduction of the first reliable automated analytical systems, the Technicon Auto-Analyzer Whiteheads work at the University of Birmingham led to the introduction of rou- tine inter-laboratory performance testing schemes and the regular use of thoroughly validated reference materials (Radin 1967; Meinke 1971; Booth et al. 1974; White- head 1976). Today the UK-EQAS scheme and the CAP scheme in the USA share a common philosophy of continuous quality improvement through repeated use of inter-laboratory studies - a concept far from the “pass-fail” mentality common in other disciplines, see also Sections 4.1 and 6.2.

Other scientific disciplines required standards. When the American Type Culture Collection (ATCC) was founded in 1925, one of its chief roles was to be a source of standards for the rapidly developing public health laboratory activity in the USA. In this context we mean standard organisms, rather than standard materials or chemicals, but their use was analogous, they helped produce better analpcal data.

In 1901, the U.S. National Bureau of Standards (NBS) - now the National Insti- tute of Standards and Technology (NIST) - was founded because of the increasing demand for various kinds of standards in the rapidly developing engineering industries. The early history of the NBS reference material program started in 1905 with a cooperative effort within the iron and steel industry whereby industrial analysts helped characterize the individual reference materials. Cooperation with NB S was recognized as a mark of achievement for the laboratory, so this effort served a dual purpose. It both helped the laboratory develop its measurement skills and also helped NI ST understand the measurement problems associated with a given matrix.

The analysis of irons and steels is mostly about the measurement of inorganic elements in an inorganic matrix. Hence a variety of separation and measurement techniques could be used, compared and evaluated to arrive at the best answer. Prob- lems were usually solved by looking for complete dissolution and understanding the interferences affecting the various methods that were employed. Since most methods used were related to gravirnetric quantities (i.e. weighed quantities of pure inorganic substances), traceability to the mole was not an issue and comparability and compatibility of measurements is what was sought.

NIST’s first four certified reference materials were steel samples, and these were followed by many others. The program supplied analytically well characterized homogeneous materials. This program included, from the beginning, homogeneity

7 . 7 Historical I 3

evaluation, cooperative multi-laboratory characterization and an evaluation of the measurement process to assure accurate values (Cochrane 1966; Uriano and Gra- vatt 1977). The success of this work initiated many requests from other industries for the development of appropriate materials and led to a rapid growth of this part of the NBS program and the adoption of the name “Standard Reference Material” or “SRM”; later these terms were registered as Trade Marks of NIST.

By 1951 NBS was advertizing 541 SRMs, of which 200 were alloys, ores or ceramics and 204 were hydrocarbons or oils (Bright 1951). Similar work for RMs in the fields of metallurgy and ceramics was performed by many U.S. commercial sources and in other countries as well. Examples of early suppliers of appropriate RMs include in the United Kingdom the Bureau of Analyzed Samples Ltd. (BAS), who issued RMs from 1916; in Germany the Federal Institute of Materials Research and Testing (BAM), founded 1904 as the “Royal Bureau of Materials Testing” and who issued RMs from 1912, and the Physikalisch-Technische Bundesanstalt (PTB), the Japanese Iron and Steel Institute, the French BNM (Bureau National de Mktrologie) and IRSID (Institut de Recherches de la Siderurgie FranGaise) and the Polish Com- mittee on Standardization and Measures. All of these organizations still carry out their work; and their RMs have developed and evolved as the demands of the metallurgy industry have increased.

In geochemistry, the introduction of RMs did not take place until 1951 but, once RM usage became a regular part of geochemical analysis, the consequences were not far short of amazing. For many years geochemical analysts had been concerned about the accuracy of their determinations of major elements in rocks, but it was the potential of emission spectrometry for the determination of trace elements which set off the production of the first rock Certified Reference Materials (CRMs), G-I and W-I by the U.S. geological Survey (USGS) (Ahrens 1951). Geochemical CRMs characterized by a number of different institutes, including NBS, were distributed in increasing numbers by the USGS. This led in the following years to remarkable improvements in resolving major disagreements between analysts using similar or distinct techniques in geochemical analysis. From about 1965 many other international organizations were supplying geochemical RMs, (e.g. Richardson 1995; Imai et al. 1996; Potts 1997). For details see Section 6.4.

Until the 1950’s the only “biological” reference materials available were a few commercial sera produced by Seronorm A/S, the Welcome Foundation and others, as a result of the U.K. and U.S. clinical chemists’ initiatives. As many elements are found in biological matrices at much lower levels than in industrial and geological samples, improvement of the quality of elemental analysis in agricultural app- lications was the aim of a committee convened at Michigan State University in November 1950. About g kg of leaves from four orchard trees, apple, cherry, peach, and orange were homogenized and distributed to 16 U.S. laboratories in a round-robin exercise. The results showed good agreement for the major elements Ca, K, Mg, N, and P, but were much less precise for essential trace elements (Kenworthy et al. 1956). A similar collaborative study using nine vegetable RMs sent out to 13 Canadian laboratories was reported somewhat later (Ward and Heeney 1960).

4 7 Introduction

Humphry Bowen did pioneering work for the development and use of appropriate biological matrix reference materials. He was at that time developing techniques for radiochemical neutron activation analysis (RNAA) and realized that there were no standards or RMs to check the accuracy of results in this work. He had some geochemical CRMs but these were unsuitable because biological materials have a different matrix and contain orders of magnitude lower concentrations of most trace elements. Thus he prepared in 1960 IOO kg of kale (Brassica oleracea) powder, a large amount for the time (Bowen 1965); for details of the preparation see Section 2.1. Indeed, Bowen’s kale served for more than two decades as a reference and valuable aid for many analysts (Wainerdi 1979; Bowen 1984). Thus his work and the wide demand for his kale RM was pivotal to the future direction of biological reference material and stimulated the planning, preparation, distribution and analysis of further materials of similar kind.

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1.1.2 Growth and Maturity

The production of biological matrix RMs for elemental analysis began in national and international institutions in the late 1g60’s. The National Bureau of Standards, now NIST, announced its intention to produce biological RMs in 1967. Orchard leaves, its first botanical reference material, was certified and distributed in 1971 (Meinke 1971). Workers at NIST were also the first to see the need to introduce sterilization of CRMs using y-radiation for longer shelf life. In 1911 NBS offered just 23 certified reference materials, by 1975 more than 1000 SRMs were available with approximately 700 of these intended for use in chemical analysis (Cali and Stanley 1975). At the end of the last century (~ggg) , about the same number of NIST SRMs were available, with many now including a considerable percentage certified for organic constituents and other analytes never dreamed of by Bowen (Trahey 1998). As we enter the zIst Century, attention turns to the measurement of DNA in plant material as the arbiter of genetic identity, and RMs will be required, see Section 5.4.

Another important part of the history of reference materials has been the contribution of the International Atomic Energy Agency (IAEA) in Vienna. The IAEA began an analytical quality control service (AQCS) aiming at assisting its Member states to maintain and improve the quality of analytical data obtained in their laboratories. Indeed in the 1960’s AQCS was concerned primarily with radioactive measurements. Later it became involved with the reliability of nuclear methods in elemental analysis and practically all IAEA AQCS RMs started out as intercomparison materials. The first five IAEA biological reference materials were issued in 1970. They included a marine RM (mussel shells) from the IAEA laboratory at Monaco, and by 1983 the IAEA had issued 28 CRMs, but most of them were quickly exhausted because of their popularity (Parr 1984). The 1998199 IAEA AQCS catalogue contains more than 90 CRMs of environmental and biological origin for a wide range of determinedness, encompassing radionuclides, trace elements, petroleum hydrocarbons, pesticides, and PCBs (International Atomic Energy Agency 1998).

1. I Historical I 5

In the 1970’s and 1980’s, a number of other organizations started programs de- signed to provide biological, environmental, and food RMs and CRMs. Projects for the development and certification of food matrices were initiated by the U.S. Depart- ment of Agriculture, Agriculture Canada and the U.S. Food and Drug Administra- tion (Wolf and Ihnat 1984; Ihnat and Wolf 1984; Tanner 1984) in co-operation with NIST. Examples are a total diet SRM (Wolf et al. 1990) and a series of agricultural/ food RMs (Ihnat and Wolynetz 1993).

Early in the 1970’s the then European Economic Community (EEC) established a community-wide CRM program in order to pull together many of the diverse and widespread RM activities under the direction of the Community Bureau of Ref- erence (BCR) (van der Eijk 1979). BCR made first a compilation that covered the major producers, both governmental and private, not only in Europe but on a world- wide basis (Commission of the European Communities 1973; Cali and Stanley 1975). The BCR of the EEC, Brussels, now the 5th framework program of the Eur- opean Comissions DG Research, initiated in 1970 a program to make available a broad range of CRMs (Griepink et al. 1991); the first biological CRMs were issued in 1983. A large number of candidate materials were initially prepared within that program at the Joint Research Centre Ispra (Rossi and Colombo 1979). It should be mentioned that the continuous efforts of Herbert Muntau at the Ispra Laboratories were the basis for many new environmental and biological CRMs (Muntau 1979, 1980, 1984). BCR started also the production and certification of food CRMs in co- operation with several qualified European laboratories (Wagstaffe 1984). In 1984 the Institute for Reference Materials and Measurements (IRMM), previously called Central Bureau for Nuclear Measurements (CBNM), initially purely nuclear, was given a major role in the storage and distribution of BCR RMs. At the same time a series of major investments were started at the IRMM to set up facilities for the preparation of highest quality candidate RMs in economically attractive conditions (Kramer et al. 1998); see also Section 2.2. From 1995 IRMM had complete responsibility for stock management, sales policy, and renewal of sold-out materials. The increase in production and certification is significant: in 1984 BCR issued about IOO individual CRMs, in 1999 this number had increased to 570, including nuclear and isotopic materials (IRMM 1999).

Valuable contributions were made by two Canadian agencies, particularly by the National Research Council Canada (NRCC) who, from about 1976, provided marine and marine biological CRMs certified for metals, metal species and organic constituents (Berman 1984; Willie 1997). More recently their Halifax laboratories have issued a highly respected range of CRMs for the determination of shellfish toxins. Another Canadian producer, the National Water Research Institute (NWRI) specia- lized in marine (water and sedimentary) CRMs, and from the late 1980’s their matrix materials certified also for organic compounds (Chau et al. 1979; Lee and Chau 1987).

In the United Kingdom chemical RMs were first produced some time in the late 1960’s at the National Physical Laboratory (NPL) Division of Chemical Standards at Teddington. This Division was transferred to the Laboratory of the Government Che- mist LGC in November 1978. Early work was based on the development of highly

6 I 1 Introduction

purified and certified pesticides, issued as CRMs. Further developments continued under LGC and the range expanded steadily. In the 1980’s the Validity of Analytical Measurement (VAM) programme started to consider the role CRMs play in the production of valid results. Following a number of consultation exercises a list of needed matrix CRMs was developed and during the 1990’s a wide range of matrix CRMs were produced and certified. At present it offers about 300 CRMs.

Further afield, in 1978 the Japanese National Institute for Environmental Studies (NIES) started the production of a series of biological and environmental matrix CRMs, certified for a number of trace elements (Okamoto and Fuwa 1985). Recently also the certification of metal species in some materials was reported (Okamoto and Yoshinaga 1999).

A remarkable level of activity can be seen in China. The National Research Center for CRM (NRCCRM) was founded in 1980 and the certification and accreditation program for “ G B W RMs started in 1983 by co-operation with many Chinese Insti- tutions. In 1993 around Go RMs and CRMs were available (Chai Chifang 1993) and in 1999 the availability of about 1000 CRMs was reported, around 30 of them clinical, IOO environmental, zoo geological, and 300 metallic matrix materials (Rong and Min 1999).

Increasing activities for the production and certification of biological, environmental and geological CRMs from the late 1980’s have been also reported from Poland (Dybczyhslti 1995) and the Czech Republic (ICutera et al. 1995,1998).

From 1983 to 1997, seven international symposia on biological and environmental reference materials have been alternately held in the USA and Europe. The pro- ceedings mirror the global developments and problems that were discussed during these meetings including also the needs of developing countries. These symposia are described together with other RM meetings in some detail in Chapter 8.

1.1.3 Milestones and The Future

Finally some additional milestones in organization, research and development of RMs need to be mentioned. The large increase in the number of reference materials being produced led in 1975 to the formation of an I S 0 Council Committee on Ref- erence Materials (ISO-REMCO) charged with the establishment of international guidelines on principles of certification, methods of use, needs, availability and nomenclature (Klich 1999). see also Sections 1.2 and 1.3.

Significant improvements in analytical methodology and better controlled sampling procedures for environmental and biological materials led in the 1980’s to the recognition that older data describing the analysis of trace metals in, for example, natural waters and biological fluids were erroneously high, sometimes by orders of magnitude. The causes have been found to be due to inadequate methods and contamination from sampling and analytical tools. This reality must be reflected in the preparation of a new class of, as far as possible, contamination-free reference materials prepared with utmost care and reflecting the state of the art at the end of the 20th Century. Examples for this are the procedures applied at NRCC for the prep-

1 7 7.2 The Theoretical Basis

aration of natural water CRMs (Berman et al. 1983) and the efforts to produce simi- larly contamination-free bovine (Veillon et al. 1984) and human serum CRMs. The latter were prepared under rigid quality control by a group of expert laboratories and called “second generation biological RMs” (Versieck et al. 1988); for more details of preparation of this material see Section 2.1.

Another step forward was the introduction of milling of solid, mainly biological, materials at liquid nitrogen (cryogenic) temperatures. These techniques allowed homogenization under “contamination minimized” conditions, previously thought impossible to achieve. Cryogenic techniques were used to prepare materials like hair. Another advantage of the cryogenic procedures is that smaller particle sizes can be obtained than with most conventional procedures (Zeisler et al. 1983; Schla- dot and Backhaus 1988; Kramer et al. 1993). The technique was successfully applied and tested in the preparation of Specimen Bank materials for long-term stored under cryogenic temperatures without any change of chemical composition. From the experience gained during these programs, cryogenic milling and long-term cryogenic storage offers unique possibilities to prepare RMs with practically indefinitely long shelf-lives (Stoeppler and Zeisler 1993; Emons 1997).

Two challenging, but very difficult tasks have been tackled mainly or increasingly during the last two decades: the certification of organometallic species and valency states of elements (see Section 3.3), and organic compounds (see Section 3.4). But doubtless this was just the beginning and a wealth of work waits in the future to serve all needs of the analytical community (Quevauviller and Maier 1999).

From the mid 1980’s the rise of Quality Standards, Total Quality Management and Accreditation schemes created a booming demand for RMs and CRMs. Thus, the use and production of matrix RMs rapidly increased the new IAEA database lists 56 producers from 22 countries and about 1640 RMs. The 1998 Comar database, which covers a much wider scope, lists more than 200 producers and around 10 ooo RMs; see Chapter 8 for more details.

The demand for RMs and CRMs continues to grow. As traditional chemical analysis moves into biochemistry and molecular biology the demand for RMs does not abate: the only question is “what is next?” Chapter g considers these, and other future issues critically.

1.2 The Theoretical Basis

Adriaan van der Veen

The basis for the preparation and use of reference materials (RMs) is given in I S 0 Guides 30-35. These documents deal with the following aspects of the preparation and use of RMs

30. Terms and definitions 31. Certificates, reports, and labels 32. Calibration using RMs 33. Other uses of RMs

8 I Introduction

34. Quality systems of RM producers 35. General and statistical principles of the preparation of RMs

The definition of a certified reference material (CRM) is given in I S 0 Guide 30

I Each of these documents will be reviewed briefly in this Section.

(1992) and it forms the root of all other I S 0 Guides:

Reference material, accompanied by a certificate, one or more of whose property values are certified by a procedure which establishes its traceability to a n accurate realization Ofthe unit in which the property values are expressed, andfou which each certij?ed value is accompanied by a n uncertainty at a stated level of con- jidence.

Additionally, there is also a definition for a RM:

Material or substance, one or more of whose property values are suflciently homogeneous and well established to be used for the calibration of a n appa- ratus, the assessment Ofa measurement method, or fo r assigning values to materials.

The key difference between a CRM and an RM is the traceability. In order to play any role at all in metrology, traceability is a key property. “Traceability” refers to a property value of the CRM, and thus to the underlying measurements. Insufficient traceability of these measurement results will eventually lead to a RM that cannot be certified, as the property value cannot be related to other standards. In the ideal case, traceability is realized up to the International System of Units, SI, but this is only feasible for a very small number of CRMs.

Most CRMs are so-called “matrix-CRMs”, identifying that they have been made from material sampled in nature. For these materials, it is impossible to come up with property values traceable to SI, as preparation steps cannot directly be related to that. At best, a comparison can be made among methods, from which usually one is the technically best established method, and the results of such a comparison may flow into the establishment of the property values and their respective uncertainties.

The most important document, accompanying a CRM is its certificate. I S 0 Guide 31 (1981) provides guidance for the establishment of certificates, labeling of CRMs, and certification reports. The certificate contains among other information the certified values and their respective uncertainties. As important as this information is the traceability statement, which defines to what references the CRM is traceable. Ideally, a CRM is traceable to a suitable (combination) of SI units. This is not always possible, so other “stated references” may appear here. Especially when certifying matrix reference materials, malting the measurements traceable to SI does not imply that the CRM is traceable to SI as well. The steps necessary to transform the sample into a state that can be measured may have a serious impact on the traceability of the values, and thus on the traceability statement.

Although labels and certificates are mandatory, certification reports are not. It also depend on the kind of RM, whether such a report is of any relevance. For instance, for the certification of gas mixtures, a certification report would not usually

1 9 7.2 The Theoretical Basis

contain more information than is already presented on the certificate. In other cases, a certification report might be of interest, but for other reasons (economical, political) is not feasible. For many matrix RMs, brief or extensive certification reports are made available, containing for instance the results of the homogeneity and stability studies, as well as the results from the characterization measurements. This allows the user to gain some extra insight in the properties of the RM, and possibly about problems in measuring the CRM with certain methods.

There are two main uses of a RM: calibration and method performance checking. I S 0 Guide 32 (1997) deals with the use of RMs for calibration purposes. RMs used for calibration purposes are usually RMs prepared by synthetic means. Commonly, the property values of these RMs are known from preparation, and verified by some kind of suitable measurement technique. This can be a technique directly providing a value for a property of interest, or a technique that allows the comparison of the new material against older measurement standards.

I S 0 Guide 32 provides guidance in two ways. Apart from the guidance on using RMs for calibration purposes, it also provides information on the preparation and use of calibrants in a laboratory, and checking them against other RMs or measurement standards.

I S 0 Guide 33 (1998) deals with other uses of RMs. It elaborates on various uses of RMs, excluding calibration, which is the subject of I S 0 Guide 32. In most cases, RMs are used as a quality control measure, i.e. to assess the performance of a measurement method. Most matrix RMs are produced with this purpose in mind. Other purposes of RMs are the maintenance of conventional scales, such as the octane number and the pH scale. I S 0 Guide 33 provides guidance on the proper use of RMs, and therefore it is together with I S 0 Guide 32 the most important document for users of CRMs.

The assessment of the performance of a method is commonly checked by means of a (C)RM. In those cases where there is no RM available, considerable effort is requested from the laboratory to assess the performance of their own methods. The aspect of traceability of the certified value(s) is also of great importance: whenever necessary, the laboratory will make modifications in its procedures if the result of a measurement using the RM appears to be unsatisfactory. If the traceability of the values to other references is not fully established, then this judgement may be clouded by doubts about the certified value(s).

Another question to be addressed by the laboratory is the portability of the measurement results on the RM to their common test samples. The behaviour of matrices that are named the same may still widely differ. There are also examples known where, for selected parameters, it is very well possible to transfer the results on the RM directly to the daily practice. As a rule, this is not possible. The conch- sions drawn from a measurement on a RM should be translated with care to the measurement practice.

The result from a measurement on a RM is commonly a difference between the observed value and the certified value. This difference is called measurement bias, and can, appreciating both the uncertainty on the RM as well as the uncertainty added during the measurement, be tested for (statistical) significance. I S 0 Guide 33

10 I Introduction

provides the uncertainty calculations necessary to carry out such an assessment. Commonly, if a measurement bias appears to be significant, the laboratory attempts to improve the measurement procedure, effectively reducing the measurement bias. It should be noted that a measurement bias smaller than the expanded combined uncertainty from the RM and measurement is not meaningful, unless a history record exists that shows trends. This kind of trend analysis may be important for the laboratory, but falls outside the scope of I S 0 Guide 33.

Another important use of RMs is the maintenance of conventional scales. The octane number of gasoline is an example of such a scale. The scale is defined through chemicals. This definition can be realized through RMs. Another example is the pH scale, which is defined by buffers with pH = 4, pH = 7, and pH = 10.

These buffers are defined as mixtures of salts, dissolved in water. These define the pH scale can be used by laboratories for the purpose of calibrating their pH meters.

By now, it should be clear what role RMs play in measurement science. This puts great responsibility on the producers of RMs, as they must see how to satisfy the requirements set implicitly or explicitly by the users regarding matrix, parameters, uncertainty, and traceability. Laboratories use RMs often as a quality control measure, but it this obviously only valid if the RM is produced under proper conditions.

In order to ensure the quality of RMs, it is highly recommended that producers work under a quality system. I S 0 Guide 34 (1996) provides guidance on how to set up a quality system for the production of RMs. It builds on I S 0 Guide 25 ( ~ g g o ) , setting apart from the same requirements a series of extra requirements related to material and sample management, homogeneity and stability testing, and the traceability of the property values. In the first edition, I S 0 Guide 34 was a document to be used on top of I S 0 Guide 25. In the voting draft of I S 0 Guide 34:1ggg, the document has become stand-alone, thus fully discussing the requirements for a quality system.

Demonstrating competence in this field is of particular interest. Field laboratories are required to demonstrate their traceability through using RMs where possible and appropriate. Obviously, the producers of these RMs must also be able to demonstrate quality and traceability, as otherwise the international measurement infra- structure becomes a set of isolated smaller networks, rather than one big network. So, providing traceability is one of the key issues in the production of RMs.

Whereas I S 0 Guide 34 sets requirements for the quality system of a CRM producer, I S 0 Guide 35 (1989) provides guidance on how to implement many of these requirements. Among these, the document also provides a general and statistical outline of the process that leads to CRMs. The current edition of I S 0 Guide 35 is a little outdated, but still most of the contents are valid.

The preparation of RMs is possibly the most complex subject dealt with in the I S 0 Guides 30-35. In I S 0 Guide 35, the general requirements as well as the statistical frame for setting up certifications of RMs are discussed. As RMs cover a very wide area, ranging from high-quality gas mixtures with very tight uncertainties, via soils certified for organic contaminants up to microbiological RMs, the exact imple- mentation of these requirements is as manifold as there are RMs.

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I” 7.3 Technical Requirements

I S 0 Guide 35 provides details on how to implement homogeneity testing, stability testing, and different ways of characterizing RMs. The document also provides a statistical framework on how to evaluate the results of these measurements and how to establish the certified value and its uncertainty. For the producer of a RM, this guide is probably the most important one, as it details a possible implementa- tion of the production of traceable RMs. The next Section gives an overview of the technical requirements when producing RMs, along the lines of I S 0 Guide 35.

1.3 Technical Requirements

Adriaan van der k e n

The preparation of a reference material requires a great deal of planning prior to undertaking any actual activity in the project. A substantial part of the planning deals with the amounts of material needed, as well as with the design of the homogeneity, stability, and characterization studies. The design also includes the choice of appropriate measurement methods for these studies. The number of samples to be produced is also a very important variable in the planning process. With a basic outline of the items mentioned, the amount of raw material to be sampled can be esti- mated.

The planning of a project starts with the definition of what reference material is to be produced. Usually, this definition just say is something like:

8 “preparation of a soil reference material containing a series of trace elements at relevant concentration levels for environmental analytical chemistry”

Although this definition might need some further specification, it is satisfactory to start the design of the project. The first task in such a project is to obtain a sufficient amount of raw material with the desired properties. The amount of material needed is dictated by the following parts of the projects:

8

the number of samples of reference material needed the need for a feasibility study the number of samples needed for the homogeneity study the number of samples needed for the stability study the number of samples needed for a characterization of the reference rnate- rial

Each of these aspects will be addressed briefly in this Section. The number of samples of reference material needed is a commercial issue in the

first place. An important variable is the number of samples likely to be sold during the lifetime (“shelf life”) of the reference material. As the lifetime is a function of the intrinsic stability of the material, this variable also affects the amount of raw material is needed. For instance, microbiological materials have limited intrinsic stability, and therefore their lifetime is expected to be shorter than for a dry sediment certified for trace elements. So, under the assumption of an equal number of sam-

12 I 7 Introduction

ples to be dispatched per year, the number of samples needed for the rnicrobiologi- cal material is greater than for the dry sediment.

In those cases where there are any doubts about the feasibility of producing a sufficiently homogeneous and stable reference material, a feasibility study might be needed. For this study, an extra amount of material is needed. Questions regarding the best way of preparing the sample, the stability of the material, or the fitness for purpose might justify the inclusion of a feasibility study in the project. In the BCR projects, it is common practice to have a feasibility study, which usually has as the sole purpose of assessing the performance of the laboratories in the collaborative study in relation to the certification of the reference material. The feasibility study allows the participants to fine-tune their equipment, their methods, and their procedures in view of the characterization measurements. In each of these cases, a considerable extra number of samples is needed.

The design of a project aiming to produce a new reference material may also include aspects of blending of materials. In several areas, like for instance in environmental chemistry, reference materials are needed with a very wide range of parameters at appropriate (concentration) levels. Often, it is impossible to find all parameters in one material. In those cases, blending two or a few similar matrices may lead to a batch of raw material suitable for the project. The problems associated with this practice are potentially greater regarding homogeneity and stability of the reference material. The extra problems with obtaining sufficient homogeneity are obvious. Extra problems regarding stability usually come from differences in the matrix, allowing for physical and/or chemical processes that would not take place otherwise. Adaptation of the preparation procedures may very well solve these (potential) problems.

Another issue in the preparation of reference material is the required shelf life. The shelf life of reference material is the time that it remains stable under proper storage conditions. Depending on the nature of the mechanisms affecting the stability of the material, various actions can be taken to improve the shelf life. Reduction of the moisture content is one of the first options to be considered. In many cases, moisture plays a key role in mechanisms leading to instability of the matrix and/or parameters. In other cases, sterilization or pasteurization of the material might be considered in order to stop bacterial activity. When preparing solutions, additives may increase the shelf life. Obviously, the shelf life of material is also a function of the storage conditions.

In the preparation of many solid state reference materials, reduction of the grain size plays an important role. Usually this reduction is required because of the measurement methods to be used both in the projects and later by the users of the reference material, as well as to come to an acceptable minimum sample intake. The minimum sample intake can be defined as the minimum amount of material needed, so that the heterogeneity of the material does not affect the repeatability of the measurement method. The reduction of the grain size is usually implemented by crushing and/or grinding techniques. The techniques employed and the equipment used must be suitable for the purpose of processing the material. Potential problems of contamination, loss of volatile components, and/or other physical and

1.3 Technical Requirements

chemical changes during grinding must be addressed in advance and evaluated during the project whenever necessary.

After reduction of the grain size, the material can be divided into portions appropriate for use as a reference material in a laboratory. The size of these portions depends on the expectations of the users, and what is technically possible. In some cases, these test portions are intended for use as such, but in most cases some sub- sampling is needed. Both approaches have their advantages and disadvantages. The need for sub-sampling can be both: on one hand if no sub-sampling by the user is needed, the producer will have less concern about possible complaints from custo- mers with respect to the homogeneity of the material. On the other hand however, many written standards and other procedures involve some kind of sub-sampling, and having a reference material that does not require sub-sampling implies that the user does not have the option of verifying this step with the reference material. In some cases, providing samples for single-use is a necessity: after opening, the material has so little stability that it is unlikely that it could be used again. In those cases, the desired size of the sample is a portion suitable for single-use only.

There is a wide choice of method and equipment for dividing the material into portions. Again, the choice of the method as well as the equipment depends on the nature of the material. A basic requirement however is that the method used guar- antees within certain limits that it may be expected that the first sample produced will have the same properties as the last one. The use of dynamic riffling techniques is only one option. Automated subdivision techniques and equipment is becoming increasingly popular, as these systems allow producing batches of a few thousands of samples; and they can also be linked to a labelling system. Especially in those projects, where portions for single use are produced the number of samples is usually large.

The whole process so far dealt with obtaining the raw material and the production of sub-samples from it. The issue of homogeneity and stability, as well as the characterization of the material under proper conditions now needs full attention. In the past, homogeneity and stability testing were primarily intended to see whether the sample preparation was completed successfully. This is perfectly reflected in I SO Guide 35 (1989). In recent years, complete new categories of reference materials have come to existence which cannot be dealt with that way. Despite all the effort put in the preparation and conservation of the material, there still may be some heterogeneity and instability left. Just testing for significance with respect to the repeatability of the test method is incorrect in two ways. It does not answer the question of how this remaining heterogeneity and instability affects the uncertainty of the reference material. Furthermore, it leaves open the option of selecting measurement method with a poor repeatability, so that the homogeneity study will not demonstrate any heterogeneity.

In cases where test pieces (or items) are prepared, the issue of obtaining a homogeneous batch of items is even more complex. Here the preparation procedure sets limits, in combination with the properties to be certified. The uncertainty of the property values should appreciate this fact, as otherwise the uncertainty of the reference material is only valid for the batch, not for a single item from the batch. This

14 7 Introduction

is an essential requirement, which obviously may also have an impact on the design of the project. In doubtful cases, a feasibility study might be needed to investigate whether a reference material can be obtained with an appropriate level of uncertainty.

For the issue of stability, the same reasoning holds. In heavy metals analysis, lim- itations of stability are only an issue for solutions and for some highly unstable matrices. In most matrices, heavy metals are fairly stable and a lifetime of these materials typically exceeds ten years. In organic analysis, as well as in microbiological analysis, materials usually have limited stability. These judgements can obviously only be made when comparing data from the stability study with the uncertainty from the characterization of the reference material. For most proper reference materials in these categories, some uncertainty from limited stability should be taken into consideration. Obviously, if a material shows a slow but steady degradation, the data should be analyzed critically in order to find out whether material could be certified. An additional problem is that usually the stability study is also accompanied by a considerable uncertainty, which should be taken into account somehow.

After the verification of homogeneity and stability, the characterization of material can take place. Frequently, this step is named certification rather than characterization, which is wrong in view of the discussion about homogeneity and stability and their impact on the reference material. The certification of our reference material is more than the characterization of the material. However, for most people working on the development of measurement methods, the characterization is the most interesting part of the project. This probably explains the huge amount of literature available.

The characterization of a reference material can take place in different ways. Depending on the source cited, there are three or four mainstream approaches; I S 0 Guide 35 (1989) distinguishes between three:

I

characterization by multiple methods characterization by a single method

characterization by means of an inter-laboratory study

The third approach is also known as a collaborative study or a collaborative trial. Both names underpin the joint effort of the coordinator and participants to characterize the reference material. In any case, the measurement methods used in the characterization should be traceable to what is called “stated references”, and prefer- ably to SI. The aspect of traceability of measurement results goes well beyond the actual measurements; it also includes the transformation of the sample from the state of the reference material to the state in which it can be measured. An example of such a transformation is the destruction of the sample.

Traceability of measurement results is essential in the establishment of a certified reference material. As stipulated in I S 0 Guides 30 and 35, a certified reference material can only be certified if there is an uncertainty statement with a traceability statement. Basically, traceability means anchoring. In classical analytical chemistry, that SI system is often the best choice as a reference (= “anchoring point”). However, there is a wide range of parameters either defined by a method or defined by the

1 1 5 7.3 Technical Requirements

conditions under which the measurements take place; and for these measurements another reference might be more appropriate. The choice of the references is primarily the responsibility of the producer. However, he has to look at the measurement practice in the particular area.

Another aspect of traceability of the results is the linkage of data from the homogeneity study, the stability study, and the characterization study of the reference material. In order to establish this link, the coordinator must be in the position to demonstrate that the results of these three studies have a common reference. Such a reference can be a calibrant, reference material, or possibly some realization by means of a suitable method. If such a common reference is not available, it is impossible to link the data sets, and therefore it is impossible to translate the results from the homogeneity and stability studies to the characterization of the material. This is also an aspect that should be addressed in the design of the project.

In addition to the requirements regarding traceability of measurement results, the measurement methods employed should represent “state-ofithe-arY‘ in the particular field. Failing to do so would lead to a reference material with an uncertainty that has become too large to serve as a quality control. The better the methods perform in terms of uncertainty and traceability, the better the reference material will serve the interests of the (potential) users.

The measurement method used for the homogeneity study should have a very good repeatability. For a stability study, where often samples are measured at different days, the reproducibility of the measurement method is of primary importance. So, the methods for homogeneity and stability studies are not necessarily the same. This is not a problem, as long as this common reference already mentioned is available. For the characterization of the reference material, especially in the case of matrix reference materials, it is often desirable to use multiple methods, and often also multiple laboratories. Under these conditions, it is easier to arrive at an uncertainty that represents the “state-of-the-art” of the laboratories.

In summary, the preparation of reference material involves the following steps:

Definition of the reference material, i.e. the matrix, the properties to be certified, and their desired levels Design of a sampling procedure Design of a sample preparation procedure Selection of method appropriate for homogeneity and stability testing Design of the characterization of the reference material Sampling Sample preparation Homogeneity testing Stability testing Characterization of the reference material Combination of the results from homogeneity testing, stability testing, and characterization and assembling an uncertainty statement Set-up of a certificate and, if appropriate, a certification report

16 1 Introduction

It should always be kept in mind that certified reference materials are used by laboratories, authorities, and regulating bodies for quality control purposes. The primary objective of a reference material is therefore the anchoring of measurements. This anchoring of the measurements of a laboratory using the reference material under ideal conditions i s as good as the anchoring of the measurements used for establishing the reference material. So, there i s a great responsibility for all persons involved in certification projects to work under proper “traceable” conditions, because otherwise the resulting reference material is useless. A reference material with a lack of traceability to stated and acceptable references cannot be used as such. Moreover, the users of reference materials expect to buy traceability, and their interests are only served if the producers paid sufficient attention to these aspects.

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References Materials for Chemical Analysis || Introduction