Chemomphere, Vol. 10, No.

2, pp 15~ - 207 P r i n t e d i n Great B r i t a i n

O045-65~5/8Z/O201.~-..pPJJ02,00/0
~)1981 Pergamon Press Ltd.

OECD Chemicals Testing ProRramme Physico-Chemical Tests A.W. Klein, H. Harnisch, H.J. Poremski, and F. Schmidt-Bleek Umweltbundesamt Berlin (West)

S~mm ar,~ : The OECD-Expert Group on Physical Chemistry within the Chemicals Testing Programme began its work in February 1978 to prepare state-of-the-art reports on physico-chemical test methods and guidelines that can be used to predict the effects of chemicals on man and in the environment. In principle it adopted the following course of action: Selection of the most important properties of chemical substances within its respective area of testing in relation to health and environmental hazard assessment. Assembling appropriate information from the 0ECD member countries, national and international organizations and institutions, in particular, information on: - officially codified standard methods
-

consensus methods, and

- methods from scientific literature; Analysis of the available material with respect to its:
-

applicability to chemical substances of all kinds, practicability ( ease of performance ),

- reproducibility - cost ( labour and equipment )

Design of test guidelines according to a harmonized OECD format.

Revised version of a lecture given by Dr. A.W. Klein, Umweltbundesamt, Bismarckplatz I, D-tO00 Berlin 33, at the Swedish Information Meeting on the O E C D C h e m i c a l s Testing Programme, Solna, April 1980

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Improvement of the drafted Test Guidelines - by circulating an commenting them among the scientific public of the different OECD member countries, and
-

where necessary and possible in the time-frame, by conducting international laboratory intercomparison studies.

Final drafting within the OECD-Erpert Group and recommending of well-founded Test Guidelines to the OECD.

An extensive consultative process has been followed in the development of the Test Guidelines, leading to con~ibutions from other international institutions, academia, industry and other interested parties. Inspire of this not all problems could be solved, however, nor could all araising questions be answered satisfactorily within the short period of two years. Therefore, further international efforts will be necessary The OECD-Expert Group on Physical Chemistry reported early in 1980 to the OECD Chemicals Group, after generating 17 proposals for Test Guidelines and 8 preliminary proposals for Test Guidelines. The format for Test Guidelines, developed by the Expert Group on Physical Chemistry has since been adopted for all OECD Test Guidelines and forms the basis for the Methods to be adopted for Annex V of the EEC Directive 79/83~/EEC. The OEC -Expert Group on Physical Chemistry operated under Lead Country approach (Germany). Its Chairman was F. SchmidtBleak of the Umweltbundesamt (~nvironmental Agency). A total of about 70 e=~perts contributed toward its success and about the same number of laboratories participated in world-wide ring tests

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I.

Introduction During the last 10 y e a r s many c o u n t r i e s h a v e become aware

of the problem that all chemical products can be a potential risk to man and environment (1,2). Today, some 50 0OO commercial chemicals are being marketed. Most of these are produced and traded worldwide. It is a simple fact that we know very little about the environmental fate and about a wide spectrum of possible effects of most of these chemicals in the environment and upon man. While there is no clear-cut evidence that a large number of these chemicals is dangerous, it cannot be excluded that a considerable number may exert long-range, chronic effects upon man and his environment. At the same time, however, industrialised and developing countries alike c-nnot dissociate themselves from using chemicals. Too much depends upon their application in food production, health care, hygiene, comm~n~cation, transportation, convenience, and most other sectors of daily life. With i n c r e a s i n g c o n c e r n o v e r p o t e n t i a l and a c t u a l h a z a r d s t o man and h i s e n v i r o n m e n t t h r o u g h t h e h i t h e r t o l a r g e l y uncontrolled use of the commercial chemicals, a number o f k e y c h e m i c a l - p r o d u c i n g c o u n t r i e s r e c e n t l y p a s s e d or are in the legislative process of enacting general substance control legislation. J a p a n moved i n 197~ a l r e a d y , t h e U n i t e d S t a t e s e n a c t e d t h e T o x i c S ~ h s t a n c e s C o n t r o l Act i n 1979, and t h e Member S t a t e s o f E u r o p e a n Comm,,ntty a ~ e e d u p o n a common d i r e c t i v e i n 1979 ( 3 ) . On J u n e 25, 1980 t h e German B u n d e s t a g p a s s e d t h e German C h e m i c a l s Act ( ~ ) . T a k i n g a l s o a c c o u n t o f C a n a d a , S w i t z e r l a n d , Sweden, Norway and some o t h e r c o u n t r i e s , c l o s e t o 65 % o f t h e w~ed c h e m i c a l s p r o d u c t i o n i s now - o r w i l l s h o r t l y b e - u n d e r g e n e r a l l e g i s l a t i v e control. It is worth noticing that with this "new generation" of legal instruments, a significant shift from "curative" to "preventive" control is being instituted.

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2. International efforts on harmonizin~ test methods Non-harmonized t e s t i n g ( a n d e v a l u a t i o n ) o r o c e d u r e s , h o w e v e r , result in barriers to free international trade, as well as considerable c~ts to industry and administration alike (5). ~herefore, a broad international effort was started within the OECD in 1977, resulting in the "Chemica~Testing Programme". Six expert groups were established (FIE. I): Expert Group A "Physical Chemistry" (Lead Country: Germany) Erpert Group B "Ecotoxicology" (The Netherlands) Expert Group C "Degradation/Accumulation" (GermanY/ Japan) Expert Group D "Lone Term Health Effects" (United

Kingdom)

Expert Group E "Short Term Health Effects" (United States of America) Expert Group F "Step System Group" (Sweden) ( Figure I )

In p a r a g r a p h 1 of w o r k i n g mandate ( F ~ V / c h e m / 7 8 - ~ - R e v i s e d ) it was stated that: "The E x p e r t Groups w i l l b e convened b y d i f f e r e n t Lead Contries to prepare "State-of-the-art" -reports on t e s t methods t h a t can b e u s e d i n p r a c t i c e t o p r o v i d e i n f o r m a t i o n , f o r p r e d i c t i v e p u r p o s e s , on t h e p h y s i c a l chemical properties, e f f e c t s from c h e m i c a l s u b s t a n c e on biotic systems other than man, ... f!

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The international, mutual acceptability of data on chemicals for recognizing potential hazards to man and his environment depends upon the availability of and adherence to test methods (Test Guidelines, TG's) - laboratory quality assurance (GLP)
-

It is now anticipated that the OECD Council will pass the proposed concept of the inter~a~ional acceptability of data, generated on the basis of OECD TG's and GLP, in form of a decision early in 1981. This would mean that data thus produced become internationally convertible.

15e

3. Prediction of the potential hazard of chemicals on man and environment(6.? ~

The p o t e n t i a l hazard (H) arising to the environment by the release of a chemical substance can be regarded as a function of its environmental concentration (c), the exposure time (t), and t h e c r o s s s e c t i o n ~ , i . e . t h e p r o b a b i l i t y of chemical or biological effects: of the occurence

Consequently in order to perform a scientific risk assessment, the following information about the chemical is necessary: - production quantity use patterns environmental fate: compartmentalization desTadation (abiotic and biotic) accumulation (abiotic and biotic) - toxicity and ecotoxicity
-

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4. P~ysico-Chemical Properties in the Hazard Assessmerit o f S u b s t a n c e s (8, 9, 10). 4.1 General Sip~ificance of P~ysical Chemistry in T e s t i ~ Chemicals As a first step of its environmental hazard assessment the chemical has to be characterized, for the purpose of unequivocal identification of the substance and production related impurities. Basic information like structural data can be obtained by suitable analytical techuiques (e.g. by absorption spectra). In certain cases, by this way, a first estimate about the chemical and biological reactivity can be reached by structure/activity relationships, i.e. a comparison of structural resemblance to chemicals which are wellknown with respect to their environmental behaviour (11,12). Easily determined properties like melting point and boiling point give indication of impurities. Physico-chemical properties, however, do not only characterize substances, they also allow estimates with respect to - the mobility of chemicals (13-16) (e.g. by water solubility, adsorption, volatility) - their degradation behaviour (e.g. by h~drolysis or thermal and light stability) - their biological reac@ivity (14,17,18)(e.g. by partition coefficient, dissociation constant, permeability) noxious effects to man and the environment (19)(e.g. by complex formation ability, particle size, surface tension, and, including human well-being, corrosiveness) and other characteristics.
-

16o

The o b j e c t i v e o f t h e E x p e r t Group A on P h y s i c a l C h e m i s t r y was t o e l a b o r a t e - w i t h i n t ~ o y e a r s a f t e r i t s f i r s t m e e t i n g i n W a s h i n g t o n , F e b r u a r y 1978 - t h e s c i e n t i f i c and t e c h n i c a l d e t a i l s of the t e s t methods f o r t h e d e t e r m i n a t i o n of phys i c o - c h e m i c a l p r o p e r t i e s and t h e i r r e l a t i o n t o d e g r a d a t i o n , m o b i l i t y , a c c u m u l a t i o n and n o x i o u s e f f e c t s . As a first step towards harmonization through mutual international recognition of test results, those physicochemical properties were selected, which were thought to be most relevant for the assessment of the environmental behaviour of chemical substances.

@.2 Selection of Relevant P~ysico-Chemical Properties The selection of the most important physico-chemical properties of chemical substances in relation to health and environmental hazard assessment has been performed as follows: A first approach (20) was established by the 0ECD Expert Group A "Physical Chemistry" through intensive roundtable discussions during its first meeting in Washington, Febr u a r y 1978 ( T a b l e 1 ) .

Table

).

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In the course of following activities a systematic compilation and selection of relevant items shown schematically in Figure 2 has been performed by Hashimoto (21) on the basis of screening and systematizing glossaries on environmental chemistry as well as relevant hand- and textbooks. The resulting "Thesaurus of Environmental Sciences" was classified to four main categories: materials (e.g. soil, sulfate, waste water etc.) physical factors in the environment (e.g. static electricity, atmospheric pressure, radiation etc.) resulted phenomena, i.e. properties emerged from interactions of materials or caused by physical factors (e.g. oxidation, acidification, photochemical reaction etc.~ and characteristic properties (e.g. solubility, acidity, boiling point etc.)

(Fi~e2)
Materi&s, environmental factors and characteristic properties were ranked by their importance and the most important of the first two ones together with respective phenomena were re-arranged in a section A "Materials and Related Phenomena" and a section B "Environmental Factors and Related Phenomena". Then corresponding physico-chemical properties were assigned to the phenomena involved in the two sections. Those properties which could readily be measured and were regarded as mainly important at the same time were extracted and finally selected together with other porperties which seemed usefully for better understanding of the isolated phenomena (Table 2).

( Table

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I t i s remarkable t h a t t h e two approaches y i e l d e d v i r t u a l l y overlapping lists of environmentally important physicochemical p r o p e r t i e s .

Taking into consideration the significance of physicalchemical properties for forseeing effects upon the media air, water and soil - predicting the distribution between the compartments air, water, soil ("Compartmentalization") i.e. water solubility, adsorption/desorgtion, vapor-pressure - describing a part of abiotic degradation behaviour i.e. hydrolysis as a function of pH, thermal and air stability, oxidation - giving support in evaluating the biological reactivity, i.e. partition coefficient, dissociation constant, permea~nlity - eluciading possible noxious effects of certain classes of chemicals, i.e. complex formation ability, surface tension of a chemical in aqueous solution
-

5 o t h r e s u l t s were d i s c u s s e d by Export Group and combined i n a f i n a l s e l e c t i o n o f 20 p h y s i c o - c h e m i c a l p r o p e r t i e s (shown i n Table 3) d u r i n g i t s 3rd meeting i n B e r l i n , June 1979.
( Table 3 ).

4.3 Significance of Properties

The s i g n i f i c a n c e o f t h e s e l e c t e d p h y s i c o - c h e m i c a l p r o p e r t i e s in various areas of testing is s-mmarized in Fi~e 3. Depending on the test being considered, properties can be

required parts of testing the corresponding e n v i r o n m e n t a l b e h a v i o u r , i n d i c a t e d b y t h e s i g n "&", " p r e - r e q u i s i t e s " t o be known p r i o r t o a p p l y i n g t h e Test Guideline in the laboratory properly, indic a t e d by the sign "+", or useful background information ("guidance information") which aids in the selecting the relevant test, in performing a test in the best possible way and interpretating its results ("o").
( Figure 3 )

~.3-~ Environmental Fate The environmental fate of a substance includes the distribution in the environment (compartmentalization), its elimination through degradation / metabolization processes, and its accumulation in specific subsectors of the environment.

4.3.1.1 Environmental Compartmentalization The "Environmental Compartmentalization" of a substance is defined as the distribution (mass percentage on relative concentration) of a chemical among the environmental media under these simplifying assumptions:

the environment is a closed system c o n s i s t i n g of the environmental compartments: a i r , water, s e d i m e n t / s o i l , biota - the environment is at thermodynamic equilibrium, no degradation or accumulation processes occur during the distribution.
-

X64

Depending on the kind of transition being considered, different physico-chemical properties may be direct inputs for the hazard assessment. They are listed together with the corresponding environmental behaviour description at the top of Figure 3 and, by this way, it becomes obvious that, for example, the transition air/water can be described by the four phy_ sico-chemical parameters - Volatility fro m Aqueous Solution, owing to which many chemicals of low water solubility can enter the atmosphere although the vapor pressure of the pure substance may be quite small; - Density of Liquids and Solids, a determining factor of the settling behaviour of substances, especially in the aquatic environment; - Particle Size Distribution , as a factor which determines the distribution and mass transport of insoluble and non-volatile compounds in the media water and air; and Surface Tension of Aqueous Soluti0ns , affecting interracial properties.
-

Further parameters (in this case "Water Solubility", "Hydrolysis as a Function of pH", and "Dissociation Constants") have to be known before tests can be properly performed (+). On the other hand, testing of these and other properties will yield information which can be applied to justify omitting or substituting certain tests (o).

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~,.3.1.2 Degradation / Accumulation

In addition to the estimated compartmentalization, both the disappearance (Degradation / Hineralization) and locality specific accumulation of the substance need to be considered in order to obtain an estimate of environmental exposure. Within the OECD-Chemicals Testing Progr-mme, the Expert Group on Degradation / Accumulation was dealing with this subject. As indicated in Figure 3 it is agreed, however, that some of the physicochemical tests of a substance are well suited as "screening" tests for both degradation and, in particular, accumulation. It seems likely, for example, that a knowledge of the n-octanol/water partition coefficient could be used to predict the accumulation behaviour of a substance, whereas the hydrolysis behaviour is an important facet of degradation when in contact with water.

4.3.2 Toxicity / Ecotoxicity The physico-chemical properties which should be determined before toxicity and ecotoxicity testing, are s,,mmarized in the last two col~mns of Figure 3. Those marks indexed reflect the opinions expressed by the OECD-Expert Group B, D and E in their Final Reports. In our opinion Particle Size Distribution, Complex Formation Ability and Permeability are additionally to be used to predict toxic behaviour of a substance:

~6

The shape of a particle can in itself be indicative of its carcinogenic nature (e.g. asbestos fibre), and the complex formation ability may effectively mobilize heavy metals present in soil and sediments and make them accessible to food chains. Since prior to biological action substances have to be transported into biological systems, (penetration through plant cell membranes, skin resorption, stomach or intestinal mucous membranes), on the other hand the testing of a substance for permeability through cell membranes or through standardized skin-tissue culture gives an estimate of the possibility of biological action (screening for toxicology tests).

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5-

Ph~sico-chemical Properties as "Decision Points" in Test Flow Schemes As we have seen above, physico-chemical properties as "pre-requisites" are to be known before tests on other properties can be properly performed. As "guidance information" they assist in deciding if, when, to what extend and with what consequences tests should be applied. Low water solubility, for example, generally suggests lipophilicity and the possible need to check a tendency to bioaccumulate via partitioning. On the other hand high water solubility should lead automatically to studies on complex formation ability, adsorption/desorption, and partition coefficient, if the chemical under test has been found to be stable (22). With the pre-requisites for the individual tests known and some quantification of the guidance information test flow schemes can be developed providing a useful method for visualizing a systematic flow of the scientific, logical and necessary steps in the testing of chemical substances. The "Possible Flow Scheme" shown in FiE. ~ is based on an approach proposed by ElSpffer et al. (23) and contains some valuable revisions made by members of the OECD Expert Group A, especially by J.D. Bell, I. Asher, W.H.J. Strachan and R.L. Bohon. It discusses questions relating to which compartments can be reached by a substance. If anyone can be reached, information on degradation or accumulation, characteristic for this compartment, is required.

Fig. ~

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For illustration h e x a c h l o r o b e n z e n e (HCB) i s t a k e n as a model ( F I E . @, d a s h e d l i n e s ) . Because o f t h e h i g h volatility i n s p i r e o f t h e low v a p o r p r e s s u r e p h o t o d e E r a d a b i l i t y i n g a s - p h a s e and i n a d s o r b e d s t a t e h a s t o be t e s t e d . In a d d i t i o n , according to the h i g h p a r t i t i o n c o e f f i c i e n t ( n - o c t a n o l / w a t e r ) b i o c o n c e n t r a t i o n i n an a p p r o p r i a t e r e c e p t o r s h o u l d be d e t e r m i n e d . F u r t h e r m o r e anaerobic degradation tests (biotic and abiotic) are required due to the h/gh adsorption and low desorption behaviour. At t h e p r e s e n t t i m e t h e " P o s s i b l e Flow Scheme" r e p r e s e n t s only an initial approach which will need to be continually updated and revised to reflect actual experience in the laboratory, field and administrative sector. In cases where, for example, large quantities of a new substance are marketed, or dangerous de~Tadation products are detected further testing may be quite necessary.

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Elaboratin~ of Pormulated Test Guidelines for the Selected Physico-Chemical Properties

According to the working programme of the OECD Expert Groups, the "Test Guidelines" are worked out in the following steps: I. Collection and evaluation of existin~

- international
-

and national standards

consensus methods

- methods derived from scientific literature It was the aim to elaborate Test Guidelines which can be generally used for the determination of physico-chemical unknown, properties of new, and hitherto chemicals.

Further criteria for the selection of suitable methods were, - reliability - cost-efficiency - standardizability of methods. In order to avoid "double-efforts", international national and e.g.

standards were checked for suitability

first. It turned out that only few suitable standards existed inter alia because many tend to be too specific, e.g. designed for a certain class of chemical or only for certain measuring ranges. As a second step, the consensus methods, as they

are used in "routine laboratories" were screened and evaluated.

Finally, methods described in scientific literature were checked for potential use as a basis for Test Guidelines.

2. Elaboratin~ of "new" formulated Test Guidelines

The Drafts of the Test Guidelines were elaborated in cooperation of the participating member countries and were circulated and commented before coming to the plenary of the OECD-Expert Group.

3. _I~_provement of the drafted Test Guidelines After a further revision, the Test Guidelines were partly sent for "final" round of commenting to and within the member countries. Another part of the Test Guidelines (see Table 3) were tested on validity and quality in OECD/EEC-lutercomparison Testing Programmes. +) Essential improvemen~ were given by the participating laboratories and these revised drafts were finally discussed within the 0ECD-Expert Group on Physical Chemistry.

4. Final draftin~ within the OECD-Expert Grou~_and recommendinE ~ of well-founded Test Guidelines to the OECD

Nearly all of the finally revised Test Guidelines were recommended by the OECD Expert Group for adoption (see Table 3).

+)More information about the OECD/EEC-Laboratory Intercomparison Testing is given in the -nnex.

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Figure 5 shows this process of harmonization from the view of the Federal Republic of Germany as lead country of the Expert Group A and shall be demonstrated in the following section by the development of the Test Guideline "Vapor Pressure Curve".
(Figure 5)

Examwle: Vapor Pressure Curve I. Selection q f _ ~ h s _ ~ h ~ s i c o - c h e m i c a l _ ~ e ~ :

This property was selected as enviror~mentally relev~ut, because:

-

The vapor pressure of a substance gives an indication cf the probability of the phase transitions: liquid, solid ~ gaseous >gaseous

Vapor pressure is thus a significant factor for predicting the losding of the atmosphere. The vapor pressure of a substance can be -~ufficient as a basis for deciding a~ to whether or not a photochemically induced degradation test (in homogeneous gas phase or in an adsorbed phase) is necessary (23). The vapor pressure, apart from solubility in water,is the main parameter for calculating the volatility of a substance from aqueous solution (2~). 2. S u ~ e ~ of available test-methods: For the determination of the vapor pressure, there is no single vapor pressure curve measurement pro-

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cedure applicable to the entire range of vapor pressure. Therefore several methods are recommended to be used for the measurement of the vapor pressures from lo -5 to lo 5 Pa. The selection of the appTopriate test methods was done in the mentioned sequence: 4. The survey on the national and international standards (25) has shown that only one method (ASTM D-2879): "Isoteniscope") is applicable for the range of medium and higher pressures (lo up to 3 lo 5 Pa). 2. As a consequence of step 1, it was necessary to find out whether there are appropriate "consensus methods". These should be well-known methods like those used in GLP laboratories of the industry or research laboratories. As a result, t~hree methods could be found: - dynamic method - static method : range 5 " 4o 2 to lo 5 Pa : range lo to 3 4o 5 Pa .

- vapor pressure balance: range 4o -3 to 1 P a

3. In step 3 "i~ethods of the Scientific Literature" an additional method for the low pressure could be selected, the gas saturation method based on an EPA instruction (26).

3. Selection o f a~wropriate test methods and elaboratin~ of a formul~ted Test Guideline

B~sed on the survey of available test methods, the above mentioned five test methods were selected and elaborated as Test Guideline. All methods were d~scribed in detail and in a harmonized OECD-format (C 45/78).

17~

This "Preliminary Draft" was discussed and improved during the meetings of the OECD Expert Group, leading to ccntributions academia, from international institutions, industry and other interested parties.

4. Imwrovement of the Draft Test Guideline

During these meetings it became obvious that further experimental experience was quite necessary to yield essential improvements. Therefore, it was decided to start a "Leboratory Intercomparison Testing Prograz2e" in order to give an answer to questions which ha~ been raised on - comparability of the different cribed in the Test Guideline, - improvements of these methods by practical rience of the participating laboratories, e:~eand test methods des-

- screening for other and possibly better methods to determine the physico-chemical tested. For that purpose, the participatir~ laboratories properties bei~nG

were asked to perform not only - tests strictly according to the test procedures as submitted by the scientific
-

coordinator,

but also

"own" preferred methods for the determination of the corresponding physico-chemical property. In this case, the psrticipants were asked to describe this method as precisely as possible.

All test results were analyzed statistically.

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5.

Elaboratln~

of a harmonized

Test Guideline

Test results of the Intercomparison gramme as well as appropriate were discussed pating laboratories.

Testing

Pro-

comments

by experts

on a working party of the particiWeak points of the Test Guideof and for most of t h e m suggestions comparability. of the by the Test

line were detected

were made in order to improve the repeatability results and the interlaboratory Final revision under careful

consideration

suggested modifications OECD-Expert-Group Guideline A 80/5 (27).

then was performed

A leading to the harmonized

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7. Current Sta~e of E:roerience

Until now the Expert Group A discussed and finalized 16 Test Guidelines (cp. Table 3):
-

b~/-VIS Absorption Soectra Heltin~ Point / Meltin~ Rankle Boilin~ Point / Boilin~ Range Vapor Pressure Curve Water Solubility Adsorption / Desorotion Partition Coefficient (n-octanol-water) Complex Formation Ability Density of Licuids and Solids Particle Size Distribution Hydrolysis as a Function of pH Dissociation Constants Screening Test for Thermal Stability and Stability in air Viscosity of Liquids
Surface Tension of Aqueous Solutions

Fat Solubility of Solid and Liouid Substances

As it was felt that the first three among these could be applied without hesitance by every laboratory, they were not subjected to any Laboratory Intercomparison Testing Programme. Due to the very close time schedule of the OECD Chemicals Testing Programme, 8nother four Test Guidelines (Particle Size Distribution, Complex Formation Ability, Dissociation Constants, Screer~ng Test for Thermal Stability and Stability in Air) had to be developed without the benefit of a laboratory testing progr-mme at this time. They should, however, be subjected to future laboratory intercomparison work.

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Although having been in the Laboratory lntercomparison Testing Progrsmme, the Test Guideline - Volatility from Aoueous Solution could not reach the finalized state. The Expert Group A agreed that this guideline will require further revision. This task should be dealt with by the OECD up-dating mechanism (see below). The physico-chemical properties: - Relative molecular mass - Permeability - Corrosiveness are not described in form of a Test Guideline. The Expert Group A, however, considers these properties as important and recommends that Test Guidelines should be developed. Beyond the Test Guidelines mentioned above, eight further physico-chemical properties were also considered. These properties are relevant in assessing the potential immediate environmental hazard of substances in case of accidents. They are included in some legislation under the general heading of "Quality evaluation of substances". However, because the Expert Group did not necessarily posses~ first-hand in-depth expertise, and also due to time constraints, the guide lines for these properties were o~ly developed in a preliminary state: - Explosive properties of m~bst~uce.s - Oxidisin~'effects of ~ases yielding, oxsA~en - ~ T o p h o r i c behaviour of solids and licuids

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- ~,=~/roohoric behaviour of Eases and licuids - High infl-mmability of powdered, and pasty substances - High infl~mmability resp. inflammability of licuids - High infl-mmability of Eases - Substances which give off highly inflammable Eases in dangerous amounts on contact with water In the beginning of 198o a "Review ~rocedure" was started by the OECD-Secretariat and the Lead-Countries. Each OECD-Hember Country received copies of the Final Reports and in a final round they were asked for comments. the OECD-Council. In the meantime, the European Commission has iniThe "Review Panel" now has for finished work preparing the recommendations granular

tiated procedures to adapt the OECD Test Guidelines into Annex V of the Directive 79/831/EEC, Article 5 in that Directive requires application of test methods as described in Annex V. There is some indication that the discussions in Brtuxelles and Luxembourg might result in test methods in Annex V different from the OECD recommendation. inexcusable. The OECD Test Guidelines will become outdated when significant changes have occured in the state-of-theart. When such changes occur existing Guidelines will require revision and new Guidelines may need to be developed. Therefore an CECD_uD-dating mechanism will start in 1981 to deal with these tasks. For reasons inand dicated above this would be truly regrettable

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8. Conclusions I. The Test Guidelines have to be reviewed at regular time intervals for the purpose of improving to the state-of-the-art knowledge and further standardization ("Up-Dating" of TGs). 2. If there is more thsn one method for testing a physico-chemical property or a distinct range of this property (see: vapor pressure), the Test Guidelines should be differentiated into one reference method (e.g. gas saturation method) - additional methods (e.g. vapor pressure balance).
-

3. The reference method chosen should be est~olished with reference substances. These reference substances ought to be scaled uD in a "reference scale" and the additional or further methods have to be validated in reference to this scale before they are admitted to testing. a. The boundary conditions of testing have to be precised: 4.1. Testing of"purest grade" and "commercial grade" substances Until now, most of the Test Guidelines were developed for testing substances with highest degree of purity. Virtually, however, all substances of co~--mercial relevance are placed on the market with production related and other impurities (e.g. from raw materials). In some cases the impurities even exceed the substance itself by weight or by volume. In

179

addition, some products of the chemical industry can only be placed on the market after addition of stabilizers. It can be assumed that many of the Test Guidelines may be applied to "commercial grade" subst~uces, but several impurity related problems must be considered: - Identification of substance: Since spectra, structural formula, relative molecular mass, melting point, and boiling point are all part of the "identity-card" of a substance, they must be determined for the pure substance, otherwise literature will be cluttered with useless information. Sensitivity of tests in relation to impurities: Certain physico-chemical tests are inherently sensitive to the presence of particular types of impurities. Even a I percent impurity level was estimated to have a major i~oact upon test results in a najority of cases. Potential impact of impurities upon the environment: ~ e n e v e r a purified substance has been used in a physico-chemical test, the danger exists that enviror~oentally relevant results of impurities may be overlooked(e.g, an impurity may be n-octancl soluble as opposed to the West substance in the partition coefficient deternination). 4.2. Special needs for Test Guidelines applied in testing areas other than physical chemistry Some of the physico-chemical properties, e.g. water solubility, s~ability (hydrolysis,

180

oxidation, storage stability) are important pre-requisites before testing biodegradation, accumulation and exotoxicity. For this purpose properties have to be determined under certain boundary conditions which have not been considered until now. E~c~nple: Water Solubility The OECD-Expert Group B "EcotoxicoloGy" requested information on Water Solubilit[ State of Work/Answer by F~xpert Group A
-

at temperatures ranging Io - 3oC

TG developed for 2oOC not considered not considered, but TG could be extended not considered~ but TG could be extended

- at pH 6 and pH 8 - at high (35 g/1 NaC1) salt concentration - at high (4o0 mg/1CaC03) and low (20 hardness m~/1CaCO 5)

Also undecided is the question ~.Fnether water solubility should be determined in those media in which d e g r a d a t i o n / a c c ~ a l a t i o n or emotoxicity tests are to be perfoz~_ed. Therefore, it is still undetermined whether natural sea water or fresh water or simulated water should be used. The partitioning between water and p!~ncton (organic phase) c~_u effect the water solubility of a compound which can for exemple be reduced or e~_.h~anced. In simulated waters such effects do not occur.

181

Hashimoto (Xeio University, Yokohama, Hember of the OECD-V-~xp_ert Group A) checked this problem experimentally and found that the water solubility of hexachlorobenzene was reduced from 7.74 ppb in distilled water at 25C to 4.52 - 4.68 ppb in sea water at 25C. Both, the "salting out"-effect and the above mentioned partitioning phenomena may be responsible for these results. We hope that further international efforts will maintain and develop the h~urmonized state of the TGs and that both, harmonized TGs and GLP will lead to a mutual acceptance of data and on the base of that, in c o ~ e c t i o n with running activities as the OECD-Hazard Assessment Pro~ect to an equivalent eva!uation of chemicals.

182

Literature I. F. Schmidt-Bleek, P. Wagenknecht, U. Boehringer, K.-O. GGnther, W. Haberland, D. Kayser, H.-J. Poremski, S. Schultze-Wolfhagen, and P.-C. Storm: "Das Pmoblem der Umweltchemikalien vor der Verabschiedung eines Chemikaliengesetzes in der Bundesrepublik Deutschland." Chemospere 8, 581 - 722 (1979) 2. F. Schmidt-Bleek, U. Boehringer, W. Gierke, W. Haberland, D. Kayser, S. Lohse, P. Hubs, H.-D. Schenke, P.-C. Storm, P. Wagenknecht, "Pr~fung und Bewertung yon Stoffen auf ihre keit im Sinne des neuen Chemikaliengesetzes" Umweltbundesamt, Berlin (West) 1980 K.-O. GGnther, H.-J. Poremski, and B.-O. Wagner: Umweltgef~hrlich-

3. F. Schmidt-Bleek: "The Establishment of European Community Legislation" in: F. Coulston (Editor): "Regulatory Aspects of Carcinogesis and Food Additives: The Delaney Clause", PP. 339 - 342, Academic Press, New York, N.Y., 1979 4. F. Schmidt-Bleek: "Das neue Chemikaliengesetz" in: "Der Bundesminister des Innern informiert" Contributions from the "Zentrum des Wissens" apropos of the ENVITEC '80 Technology within Environmental Protection, 3 rd International Technical Fair, February 11-15, 1980; PP 7 - 18 5. F. Schmidt-Bleek and W. Bonberg: "Potential risks for nontariff barriers to trade with chemiclas"; Lecture at the International Conference on Chemicals Control, Stockholm, April 13, 1978

6. Y. Schmidt-Bleek: "Toxic Substances: Testing and Evaluating Chemicals", Prepared f o r Scope, May 12, 1980 7. F. Schmidt-Bleek: "Testing and Evaluating Methods for Environmental Chemicals" 10 th Annual Symposium on the Analytical Chemistry of Pollutants, Dortmund, Hay 28 - 30, 1980 8. OECD Chemicals Testing Programme, Expert Group "Physical Chemistry", Final Report, Umweltbundesamt, Berlin, Dec. 1979 9- A.W. Klein, H.J. Poremski, F. Schmidt-Bleek: "Physikalisch-Chemische Laborpr~ifung neuer Stoffe nach dem Chemikaliensesetz"; Sicherheit in Chemie und Umwelt, in press 10. H.J. Poremski, A.W. Klein, and F. Schmidt-Bleek: "The predictive value of physical chemical properties", International Symposium on "Principles for the Interpretation of the Results of Testing Procedures in Ecotoxicology", Sophia-Antipolis, Valbonne (Alpes-Maritimes), France, September 30 - October 2, 1980; Proceedings, in press

184

11. R.L. Dehn and C.T. Helmes: "An automatic procedure for assessing possible carcinogenic activity of chemicals prior to testing", Stanford Research Institute, June 197@, published by NTIS 12. C.F. Wilkinson: "Correlation of Biological Activity with Chemical Structure and Physical Properties", published in: Van Valkenburg: "Pesticide Formulations", Marcel Dekker, Inc. New York, q973 13. W. Klepffer, R. Frische, G. Esser, and H. Puchinger: "Physico-chemical properties and environmental behaviour of chemicals" Battelle Institute, Frankfurt q~. A.M. Stern and C.R. Walker: "Hazard Assessment of Toxic Substances: Environmental Fate Testing of Organic Chemicals and Ecological Effects Testing", in: "Estimating the Hazard of Chemical Substances to Aquatic Life", ASTM-STP 657 15. P.H. Howard, J. Saxena, and H. Sikka: "Determining the fate of chemicals", Env. Sci. Techn. q2, 398 - ~07 (q978) 16. S. B. Redding, D.H. Lin, H.L. Johnson, and Th. Mill: "Review of the environmental fate of selected chemicals", Standford Research Institute, May 1977, published by NTIS 17. B. Neely, D.R. Branson, and G.E. Blau: "Partition Coefficient to Measure Bioconcentration Potential of Organic Chemicals in Fish" Env. Sci. Techn. ~, 1113 - ~115 (197@)

185

18. 0ECD-document: C 29/78 J "Working Paper on Bioaccumulation",

Berlin, Nov. 1978

19. I. Borneff, Archiv f. Hygiene und Bakteriologie, 28, 1~7, (1963) 2o. OECD-document: A-78.4 "Environmental Relev~uce of the 19 Physico-Chemical Properties", Washington, Febr. 1978 21. OECD-document: A-78.25 "An Example of a Method of Selection of PhysicoChemical Properties for Environmental Studies", Berlin, Sept. 1978 22. 3 M, Enviror~ental ~ugineering & Pollution Control, St. Paul, Hi~u., Hay 5, 198o Contribution to: S~mmary of United States' Public Comments on the Reports of Expert Groups of the OECD Chemicals Testing Program Office of Pesticides and Toxic Substances, Environmental Protection Agency, Washington, D.C., July 19So 23. 0ECD-document: A 79-63 "The significance of physico-chemical properties in test flow schemes", Berlin, June 1979 24. D. N~ckay and P.J. Leinonen: "Rate of Evaporation of Low-Solubility Containa::ws from water bodies to atmosphere", ~uv. Sci. and Techn. Vol. 9, Dec. 1975, 1178-1!81 25. OECD-document: A 80.32 "S;~optica! Review abcut the ~ t i o n a l and international Standards for the Determination of Physico-Chemical Properties", Umweltbundesamt, Berlin, Dec. 1979 28. ~A-document "Preliminary Draft Guidance for Premar.ufacture fication", September 8, 1978 Noti-

186

27. OECD-document: A 80.5 "Test Guideline for Vapor Pressure Curve", Umweltbundesamt, Berlin, Dec. 1979 28. Report of the EEC-Laboratory Intercomparison Testing Programme on physice-chemical properties, Umweltbundesamt, Berlin, July 1979 29. OECD-document: A 79.82 "Summary of the OECD Laboratory Intercomparison Programme Part I - ", Umweltbundesamt, Berlin, September !979 30. OECD-document: A 80.30 "Summary of the OECD/EEC Laboratory Intercom~arison Testing Programme - Part II - ", Umweltbundesamt, Berlin, ~ y 1980 34 . G. Gottschalk and R.E. Kaiser: "Einft~hrung in die Varianzanalyse und Ringversuche", BIHochschultaschenbGcher (Bd. 775), Bibliographisches Institut Mannheim/Wien/ZGrich, 1976 32. ISO/DIS 5725 (1977), Precision of test methods Determination of repeatability and reproducibility 33. J. Wernicke: Personal communication ~4. A. Lindner: "Statistisohe Methoden f~r Natur~issenschaftler, Mediziner und Ingenieure", 3- Aufl., Basel 1960

187

Table 1: Relevant physico-chemical properties selected by the OECD-E:~ert Group A at its 1st meeting, Washington 1978 (2o)

1. Particle size distribution 2. Vapor pressure curve and boiling point 3. Melting point or melting point range 4. Density 5- Volatility of a substance from the adsorbed state 6. Volatility of a substance from an aqueous medium 7. Solubility in water 3. Solubility in fat (synthetic fat) 9. Partition coefficient (n-octanol/water) lo. Viscosity 11. Surface tension of an aqueous solution (test substance dissolved) 12. Adsorption/Desorpticn 13. Complex formation ability 14. Permeation 15. Absorption spectrun (IR/UV) 16. Dissociation constants 17. ~nermal stability (in air) 18. Hydrolysis Lu dependence on pH 19. Corrosiveness (including redoz potential)

188

Table 2: Finally selected physico-chemical properties according to Hashimoto (21) absorption of water/humidity acidity/alkalinity boiling point/melting point color complex formation ability decomposition temperature density dissociation constant expansion coefficient explosion temperature flash point grain size ignition temperature ionization potential light absorption curve odor permeability purity redox potential oxidation/reduction product partition coefficient permeability pH pH stability pulverization solubility into fat solubility into water solubility of colloid specific gravity specific gravity against air sublimation tempera~are surface tension thermal conductivity thermal stability turbidity vapor pressure vapor pressure from sulution viscosity volatility

189

Table Physico-chemical properties selected for their relevance hazard assessment of substances Selected physical chemical properties I. Relat. Molecular Mass/Struct~ral Formula 2. UV-VIS Absorption Spectra 3- Melting Point/Melting Range 4. Boiling Point/Boiling Range 5- Vapor Pressure Curve 6. Water Solubility ?. Adsorption/Desorption 8. Partition Coefficient (n-octanol/water) 9- Volatility f. Aqu. Solution 10. Complex Formation Ability 11. Density of Liquids and Solids 12. Particle Size/Fibre Shaoe 13. Hydrolysis as Fct. of pH x x x x x x x x x x x x x x x x x x x x x x x x x x TG reccmm, to CECD for adoption in the

Tested in Laboratories

14. Dissociation Const. in Water 15. Screen.-Test on Thermal Stab. and Stability in Air 16. Viscosity of Liquids !7. Surface Tension of Aqu. Solution 18. Fat Solubility 19. Permeability 20. Corrosiveness (Redox-Behaviour)

190

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191

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OECD/EEC-Laboratory Intercom~arison Testing I. General Remarks

Until now 1o Test Guidelines have been checked by three consecutive Testing Progrsm~es: I. In July 1978, the Commission of the ~-hropean Communities decided to organize a Lsboratory Intercomparison Testing Progr ~amme to prove the Test Guidelines of the four physico-chemical properties: Vapor Pressure Curve - Water Solubility
-

- Fat Solubility - Partition Coefficient (n-octanol/water). ,During this prcgra~me, Io substances were tested by 18 laboratories in ~ European countries (28). !i. in September 1978 the OECD-Expert Group A "Physical Chemistry" decided to organize an Intercomparison Testing of the Test Guidelines on the properties:
-

V<por Pressure Curve

- Water Solubility Fat Solubility - Partition Coefficient (n-oct./water) Density of Liquids and Solids - Viscosity of Liquids
-

Surface Tension of Aqueous Solutions

35 laboratories in 50ECD-member countries tester 17 test substances (29).

197

ili. On its 3rd meeting, ry Intercomparison following: - Hydrolysis

June 1979, the E:rpert Group A dea jount L~borato(Part II) on the Testing Programme

cided to start together with the ~ C

as a .~hnction of pH

- Volatility from Aqueous Solution - idsorptiorJDesorption 12 substances were tested by 22 laboratories cotuntries ~ o ) . in 8

In order to dete~nnine the quality of the test methods numerical results were subjected to statistical evaluation.

2. Statistical Evaluation of the CECD/~_V~C-Lsborator[ Interssmparison Testing Results

Test results usually are dependent on many factors, li~'e

-

test su~ostances

- laboratory equipment - staff (practice, e:,.-perience)

-

quality of the test methods etc.

- analD~ics,

in order to determine the quality of "test methods" by ~imple statistical methods, all other factors h~ve to by considered constantl , i.e. - the scientific coordinator has to supp!v identical compounds - the laborato~: equipment as well as the practice and the e:rp_erience of the staff have to be above ~::e "min~__mum level" to perfo-?~m the tests, and the analytical methods have to be identical.

19e

Although the OECD/EEC-Laboratory Intercomparison Testing could not always be strictly performed under the above mentioned conditions, all test results were analyzed statistically. As an illustration results of Fat Solubility and Vapor Pressure Curve testing are shown in the following: 2.1 Fat Solubi!i~y of Solid and Liguid Substances The following 2 substances were recommended by the Expert Group A as suitable for the purpose of checking the accuracy of the Test Guideline on Fat Solubility:
I. Hexachlorobenzene

2. Mercury (II) chloride From the seven laboratories" that took part in the laboratory comparison testing four ones determined the prooerty under consideration of only one substance. As furthermore no analyticel technique was prescribed results of different analytical methods had to be treated together in order to facilitate statistical evaluation in this investigation.

(Table A-I)
In the first step of evaluation the average for each substance and laboratcryand the standard deviations a n d r a n E e s for each substance as well as the average of all determinations in all laboratories - i.e., the total mean - have been calculated. The corresponding coefficient of variation has been computed according to Gottschalk & Kayser (31). Values obtained are shown in the Tables A-I and A-2. (Table A-2) The next step was to carry out an analysis of the repeatability and reproducibility of the test method according to ISO/DIS 5725 (32) on each data set presented. The puroose " Throughout this report the participating laboratories are referred to by arbitrary numbers.

199

of this evaluation is not only to show the discrepancies between the laboratories but also to give a numerical expression to the variations within the laboratories. The following equations are used for the determination of the measuring precision within and between laboratories: Standard Deviation of Reoeatability: i er ; si 2 P : : within laboratory variation number of laboratories

Standard Deviation of Reoroducibility: sI = s sr ; s~ : between-laboratory variation

Reweatability (si~nif. level 9~ %): r - t (95) . ~ . 2.83 . s r sr ; t : Student-factor according to ISO/DIS 5725)

; (t(95) ~ 2 . o

Reoroducibility (signif. level 9~ %): 2 1 R = t ( 9 5 ) " ~ . 2 V , s12 + s r ~ 2 . 8 3 . ~s2"+ s r

Conditions for the overall assessment of the measurin~ orecision ~;tx~ R/r ~.5 I. 5 w R / r , 2.0 2.0 ~ R/r ~ 2.8 2.8 R/r @ @ @ ~ fairly homogeneous sufficiently homogeneous inhomogeneous strongly inhomogeneous

The results obtained are summarized in the Tables A-3 and A-@.

2o0

(Table A-3) (Table A-@) From Table A-@. 3 it could be assumed that excessive systematic measuring errors or methodic difficulties turn the measurements worthless. But it should be kept in mind that the OECDlaboratory intercomparison testing was not performed exactly under ring test conditions as the number of participating laboratories was too small. Furthermore, the number of single measurements was not sufficient and there was no conformity in the number of measurements done. The results still yielded valuable indications of weak points of the Test Guideline and modifications were made in order to improve the repeatability of results and the between-laboratory reproducibility. 2.2 Vaoor Pressure Curve As suitable for the purpose of checking the accuracy of the different test methods (cp. chapter 4.1) of the Test Guideline on Vapor Pressure Curve the following substances were recommended by the Expert Group A: Reference Method dynamic method Range medium to high v.p. Test Substance I. Trichloroethylene 2. Di (2-ethyl-hexyl)phthalate

3. Phenol
4. Aniline 5. Toluene

static and isoteniscope method vapor pressure balance and gas saturation method

medium to high v.p. low v.p.

I. Phenol 2. Aniline 3. Toluene 4. 2. 34. Hexachlorobenzene Di (2-ethyl-hexyl)phthalate Pentachlorophenol Benzoic acid

201

Host of the participating eight laboratories pressure curves by only one method.

determined vapor some of the analysis be-

Furthermore,

results could not be included in the statistical ported.

cause only mean values or diagrams without raw data were reStatistical results therefore are only of limited on each value because of the small number of measurements substance.

For the evaluation the applicable results for each substance were transformed: p ~a] > log10 p

and summarized in different groups according to the method applied. The data of each group were subjected to a straight line fit by the method of least squares: y with y and x = lIT = log!o P A + B x

The goodness of curve fitting was determined by the r-square values (34).

The regression coefficients A and B as well as the corresponding r-square values are presented in Table A-5. The higher the calculated r-squares the smaller is the scattering of the measured values. (Table A-5) In Table A-6 the vapor pressure values (in Pa) calculated from the

regression curves are listed for the temperatures 10 C, 20 C and 30 C together with the range of calculated p at .C C. (Table A-6)

202

Table A-I: Fat Solubility (g/lO00 g fat) data of hexachloro. benzene

Lab. i 1002 1015 1032 lO25

1026

Mean ~i 11.2 11.1 11.4 12.1 11.3 3.0

Standard deviation si 0.632 0.451 0.139 0.497 0.360 0.179

No of values

1062

10 20 20 14 10 5

Average (total mean), ~ Standard deviation, s~ Range Coefficient of variation (31):

10.0 (11 . ~ ) 1 ) 3.5 (0.@) 1) 3.0 - 12.1 (11.1 - 12.1) !) o.3~ (o.o4) ~)

1)values in brackets without outlier, see DIXON's Test (Table A-3)

2o~

Table A-2: Fat solubility (g/1000 g fat) data of mercury (II) chloride

Lab. i

Mean xi

Standard defiation si

No of values

1015 1025 1047 1062

21.3 14.7 24.3 0.8

0.68 0.27 0-75 0.14

20 3 20 5

Average (total mean), ~ Standard deviation, s~ Range Coefficient of variation (31) :

15.3 10.5
0.8 - 24.3 0.68

204

Table A-~: Precision of the test method according to ISO/DIS 5725 (32) on fat solubility data of hexachlorobenzene A-3.~ Search and elimination of outliers

A-3.1 .I COCHRAN's maximum variance test The Cochran-test is not applicable since not all labs delivered equal numbers of measurement values.
A-3.1.2 DIXON's test of mean outliers Due to remote mean values the laboratory 1062 was found as outlier (sigaif. level 99 %) and eliminated: Lab Mean Dixon Test Val. Dixon Thresholds i 1062 ~i 3.0 D 0.89 95 % 0.63 99 % 0.74

No Dixon stragglers (signif. level 95 %) were found. A-3.2 Repeatability and Reoroducibility

STANDARD ~VIA~ION OF ~ V ~ T ~ Z ~ _ !
absolute s r = 0.45 3.9 %

relative s r .I00/~ =

STANDARD DEVIATION OF REPRODUCIBILITY absolute s I = relative s I. 100/~ = FURTh~EVALUATION With 95 % probability are the REPEATABILITY the REPRODUCIBILITY Furthermore the TOTAL HEWN is A-3.30VEPATI. ABS~SSMENT The overall assessment of the measuring precision within and between the laboratories is according to the quotient
R/r = q.1

0.25 2.2 %

r R

= =

0.13 0.14

= 11.4

FAIRLY HOMOGENOUS

205

Table A-A: Precision of the test method according to ISO/DIS 5725 (32) on fat solubility data of mercury (II) chloride A-~.I Search and elimination of outliers A-4.1.~ COCHRAN's Maximum variance test The Cochran-test is not applicable since not all labs delivered equal numbers of measurement values. A-@.I.2 DIXON's test of mean outliers No Dixon outliers (signif. level 99 %)/stragglers (signif. level 95 %) were found

A-~.2 Repeatability and reDroducibility STANDARD DEVIATION OF REPEATABILITY absolute s r relative Sr.100/~ = = 0.53 3.@ %

STANDARD DEVIATION OF REPRODUCIBILITY mabsolute s I relative Sl-100/~ FURTHER EVALUATION With 95 % probability are the REPEATABILITY r ~he REPRODUCIBILITY R Furthermore the TOTAL MEA~N is A-4.3 OVERALL ASSESSMENT The overall assessment of the measuring precision within and between the laboratories is according to the quotient R / r = 19.8 STRONGLY I N H O M O G E N O U S x = 15.3 = = 10.@ 68.3

0.15 2.96

206

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