ISO/DIS 10993-16:2025(en)

ISO/TC 194/WG 13

Secretariat: DIN

Date: 2025-10-13

Biological evaluation of medical devices — Part 16: Toxicokinetic evaluation for degradation products and leachables

Évaluation biologique des dispositifs médicaux — Partie 16: Évaluation de la toxicocinétique des produits de dégradation et des substances relargables

© ISO 2025

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Contents Page

Foreword 3

Introduction 5

1 Scope 1

2 Normative references 1

3 Terms and definitions 1

4 Guidance on toxicokinetic evaluation 3

5 Circumstances in which toxicokinetic evaluation shall be considered 3

6 Mathematical modelling for the evaluation of toxicokinetics 4

6.1 General considerations 4

6.2 Rationale for the use of mathematical modelling 4

6.3 Data requirements 5

7 Principles for design of toxicokinetic animal studies 5

8 Output of toxicokinetic evaluation 6

Annex A (informative) Mathematical modelling 7

A.1 General 7

A.2 Classical compartment open models 7

A.2.1 General 7

A.2.2 Zero-order kinetics 7

A.2.3 First order kinetics 7

A.2.4 Two compartments model 7

A.2.5 Michaelis-Menten kinetics 8

A.3 Physiologically based toxicokinetic (PBTK) models 8

Annex B (informative) Information on animal toxicokinetic testing 9

B.1 General considerations 9

B.2 Guidance on specific types of tests 10

B.2.1 General 10

B.2.2 Absorption 11

B.2.3 Distribution 11

B.2.4 Metabolism and excretion 11

Annex ZA (informative) Relationship between this European Standard the General Safety and Performance Requirements of Regulation (EU) 2017/745 aimed to be covered 13

Foreword

ISO (the International Organization for Standardization) is a worldwide federation of national standards bodies (ISO member bodies). The work of preparing International Standards is normally carried out through ISO technical committees. Each member body interested in a subject for which a technical committee has been established has the right to be represented on that committee. International organizations, governmental and non-governmental, in liaison with ISO, also take part in the work. ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization.

The procedures used to develop this document and those intended for its further maintenance are described in the ISO/IEC Directives, Part 1. In particular the different approval criteria needed for the different types of ISO documents should be noted. This document was drafted in accordance with the editorial rules of the ISO/IEC Directives, Part 2 (see www.iso.org/directives).

Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of any patent rights identified during the development of the document will be in the Introduction and/or on the ISO list of patent declarations received (see www.iso.org/patents).

Any trade name used in this document is information given for the convenience of users and does not constitute an endorsement.

For an explanation on the voluntary nature of standards, the meaning of ISO specific terms and expressions related to conformity assessment, as well as information about ISO’s adherence to the World Trade Organization (WTO) principles in the Technical Barriers to Trade (TBT) see the following URL: www.iso.org/iso/foreword.html.

This document was prepared by Technical Committee ISO/TC 194, Biological and clinical evaluation of medical devices, in collaboration with the European Committee for Standardization (CEN) Technical Committee CEN/TC 206, Biological and clinical evaluation of medical devices, in accordance with the Agreement on technical cooperation between ISO and CEN (Vienna Agreement).

This fourth edition cancels and replaces the third edition (ISO 10993‑16:2017), which has been technically revised with the following changes:

a) the title is changed to accommodate methods alternative to animal studies;

b) the introduction is modified;

c) the scope is modified;

d) a new clause 4 was added to include guidance on toxicokinetic evaluation;

e) normative Annex A has been moved to clause 5 and the title is changed to accommodate methods alternative to animal studies;

f) a new clause 6 was added to include mathematical modelling for the evaluation of toxicokinetics;

g) a new clause 8 was added to address possible outcome of toxicokinetic evaluation;

h) a new informative Annex A was added to include additional information on mathematical modelling;

i) Clause 5 guidance on test methods, has been moved to new informative Annex B.

j) the bibliography has been updated.

A list of all the parts in the ISO 10993 series can be found on the ISO website.

Introduction

Medical devices can release leachables (e.g., residual catalysts, processing aids, residual monomers, fillers, antioxidants, plasticizers, etc.) or degradation products which migrate from the material and have the potential to cause adverse effects in the body. Toxicokinetic evaluation describes the fate of a foreign chemical in the body with time by assessing absorption, distribution, metabolism, and excretion of the chemical and can be of value in assessing the safety of medical devices. Toxicokinetic evaluation can also be applicable to medical devices containing active ingredients, in which case, pharmaceutical legislation is to be considered.

Traditionally, animal studies have been used for toxicokinetic evaluation. The need for and extent of toxicokinetic animal study should be carefully considered; it is preferably to replace animal studies with mathematical modelling or evaluation of existing toxicological and toxicokinetic data. The need for consulting toxicokinetic experts for these considerations is emphasized.

A considerable body of published literature exists on the use of toxicokinetic methods to study the fate of chemicals in the body (see Bibliography).

Biological evaluation of medical devices — Part 16: Toxicokinetic evaluation for degradation products and leachables

This document describes the considerations for inclusion of toxicokinetic evaluation in the biological evaluation of medical devices and provides principles on designing and performing toxicokinetic evaluation relevant to medical devices.

The following documents are referred to in the text in such a way that some of or all their content constitutes requirements of this document. For dated references, only the edition cited applies. For undated references, the latest edition of the referenced document (including any amendments) applies.

ISO 10993‑1, Biological evaluation of medical devices — Part 1: Requirements and general principles for the evaluation of biological safety within a risk management process

ISO 10993‑2, Biological evaluation of medical devices — Part 2: Animal welfare requirements

ISO 10993‑17, Biological evaluation of medical devices — Part 17: Toxicological risk assessment of medical device constituents

For the purposes of this document, the terms and definitions given in ISO 10993‑1 and the following apply.

ISO and IEC maintain terminological databases for use in standardization at the following addresses:

— IEC Electropedia: available at https://www.electropedia.org/

— ISO Online browsing platform: available at http://www.iso.org/obp

3.1

absorbable device

medical device designed to biodegrade and to be absorbed by the body over time; typically serving a temporary function

3.2

absorption

action of a material or substance, or its decomposition products passing through or being assimilated either by cells or tissue or both over time

3.3

bioavailability

extent of systemic absorption (3.1) of specified substance

3.4

biodegradation

degradation due to the biological environment

Note 1 to entry: Biodegradation might be modelled by in vitro tests.

3.5

constituent

chemical that is present in or on the finished medical device or its material of construction

3.6

clearance

rate of removal of a specified substance from the body or parts of the body by metabolism (3.15) or excretion (3.10)

3.7

c_max

maximum concentration of a specified substance in plasma

Note 1 to entry: When the maximum concentration in fluid or tissue is being referred too, it should have an appropriate identifier, e.g., c_max, liver, and be expressed in mass per unit volume or mass.

3.8

degradation product

product which is derived from breakdown of a material by physical, chemical or biological means

3.9

distribution

process by which an absorbed substance or its metabolites circulate and partition within the body

3.10

excretion

process by which an absorbed substance or its metabolites are removed from the body

3.11

extract

liquid that results from extraction of the test substance (3.16) or control

3.12

half-life

t₁/₂

time for the concentration of a specified substance to decrease to 50 % of its initial value in the same body fluid or tissue

3.13

leachable

constituent (3.5) released from a medical device under clinical use conditions

Note 1 to entry: A leachable (e.g., additives, monomeric or oligomeric constituent of polymeric material) can be extracted under laboratory conditions that simulate normal conditions of exposure.

3.14

mean residence time

statistical moment related to half-life (3.12) which provides a quantitative estimate of the persistence of a specified substance in the body

3.15

metabolism

process by which an absorbed substance is structurally changed within the body by enzymatic and/or non-enzymatic reactions

Note 1 to entry: The products of the initial reaction can subsequently be modified by either enzymatic or non-enzymatic reactions prior to excretion (3.10).

3.16

test substance

degradation product (3.8) or leachable (3.13) used for toxicokinetic study

3.17

t_max

time at which c_max (3.6) is observed

3.18

volume of distribution

V_d

parameter for a single-compartment model describing the apparent volume which would contain the amount of test substance (3.15) in the body if it were uniformly distributed

4.1 Potential hazards exist in the use of most medical devices. Chemical characterization identifies chemical hazards and shall precede toxicokinetic considerations. However, it is neither necessary nor practical to conduct toxicokinetic evaluation for all identifiable intended and unintended leachables and degradation products, nor for all medical devices.

NOTE More information on potential risks can be found in ISO 10993‑17 and ISO 14971.

4.2 The need for toxicokinetic evaluation as part of the biological evaluation of a medical device shall be considered taking into account the finished device and its constituent chemicals, intended and unintended leachables and degradation products in combination with the intended use of the device, e.g. nature and duration of contact (Figure 1).

Possible toxicokinetic interaction between active ingredients and leachables or degradation products should also be considered.

4.3 Mathematic modelling shall be considered when sufficient relevant background data exist.

4.4 In vitro methods, which are appropriately validated using positive or negative controls or both, reasonable and practically available, reliable and reproducible, shall be considered for use in preference to in vivo tests. This shall be done in accordance with ISO 10993-1. Where appropriate, in vitro experiments (e.g., tissue, homogenates or cells) may be conducted to investigate probable rather than possible degradation products.

Figure 1 — Process of assessing the need for toxicokinetic evaluation

5.1 Toxicokinetic evaluation shall be considered if the following conditions are met:

a) the device is designed to be absorbable;

b) the device is a permanent contact implant, and significant corrosion or biodegradation is known or likely, and/or migration of leachables from the device occurs;

c) substantial quantities of potentially toxic or reactive degradation products or constituents released into the body during clinical use;

d) substantial quantities of pharmacologically active constituents are likely or known to be released from a medical device;

e) substantial quantities of nano-objects are likely or known to be released from a medical device into the body during clinical use.

NOTE 1 The meaning of the term “substantial quantities” is dependent on the properties of the chemicals or nano-objects in question and is based on expert judgements.

NOTE 2 See ISO/TR 10993‑22 for information on nano-objects.

5.2 Toxicokinetic evaluation or in vivo studies are not required if:

a) sufficient toxicological data or toxicokinetic data relevant to the degradation products and leachables already exist;

b) sufficient toxicological data or toxicokinetic data relevant to the active pharmacologically ingredients already exist;

c) achieved or expected rates of release of degradation products and leachables from a particular device have been judged to demonstrate safe levels of clinical exposure. This shall be done in accordance with ISO 10993-17;

d) clinical exposure of degradation products and leachables is documented as safe with reference to historical experience.

5.3 Where materials are complex and contain products which are either endogenous or they are so similar to endogenous products that they cannot be analytically distinguished, a toxicokinetic evaluation is usually not feasible.

NOTE 1 Other studies, such as implantation studies that include histological evaluation of the implant and implant site, can yield useful information on the toxicokinetic behaviour of the device.

Mathematical functions can be used as an alternative to animal studies to describe the time- and dose-dependent processes of absorption, distribution, metabolism, and elimination of a chemical substance and metabolites thereof in animals and humans. For this purpose, classical compartment open models and physiologically based toxicokinetic models are used. Both are applied to fit concentration-time data and to predict concentration-time courses of a xenobiotic and its metabolite(s) in organs, tissue, blood, plasma, and/or other secrete such as urine after repeated or continuous exposures.

NOTE Validated commercial products for toxicokinetic modelling exist.

Toxicokinetic data are normally derived from animal studies. A major concern is that animal derived toxicokinetic data are not always reliable for extrapolation to humans due to differences in physiology, biochemical and metabolic pathways. For this reason and for a general attitude and regional legislation in reducing animal testing there is an increasing pressure to develop and use alternative (non-animal) toxicokinetic methods to reliably determine the necessary toxicokinetic parameters. See Annex A for different mathematical models.

Such modelling requires the collection of physical and chemical properties, and physiological parameters relating to tissue uptake and clearance. Physical and chemical information may include molecular weight, octanol-water partition coefficient and pKa. If insufficient literature data exists, in vitro studies such as metabolism or tissue uptake may be addressed via organ homogenates or tissue slices. The data shall be created using appropriate quality assurance controls, e.g. positive or negative controls.

7.1 If a toxicokinetic evaluation is needed (see 5.1 and 5.2) and cannot be completed by mathematic modelling (see clause 6) animal tests shall be considered. Toxicokinetic studies shall be designed on a case-by-case basis.

NOTE Annex B provides information on animal toxicokinetic testing.

7.2 A study protocol shall be written prior to commencement of the study. The study design, including methods, shall be defined in this protocol. The study design shall state the physiological fluid, tissue or excreta in which analyte levels will be determined. Animal tests shall be done in accordance with ISO 10993-2.

7.3 The results of extraction studies, chemical characterization, and toxicological risk assessment should be considered to determine the methods to be used for toxicokinetic studies. Information on the chemical and physicochemical properties, surface morphology of the material and biochemical properties of any leachable should also be considered.

NOTE More information on extraction studies can be found in ISO 10993-12, on chemical characterization in ISO 10993-18, and on toxicological risk assessment of device constituents in ISO 10993-17.

NOTE The extent and rate of release of leachables depend on the concentration at the surface, migration to the surface within the material, solubility and flow rate in the physiological milieu.

7.4 It is recommended to undertake toxicokinetic studies with a characterized leachable or degradation product that has the potential of being toxic. However, the performance of toxicokinetic studies on mixtures is possible under certain conditions. An extract liquid, or a ground or powdered form of the material or device may be used in exceptional circumstances and shall be justified in the study design. In either case, the dose (both chemical/chemicals and amount) administered shall be documented.

NOTE More information on extraction studies can be found in ISO 10993-12.

7.5 Analytical methods shall be able to detect and characterize degradation products, leachables and metabolites in biological fluids and tissues. Quantitative analytical methods shall be specific, sensitive and reproducible. Limit of detection/quantification shall be defined and justified. Validation/qualification of the method shall be performed.

NOTE Use of radiolabelled compounds can be useful in detecting unknown metabolites.

7.6 Analyte recovery from the matrix shall be documented.

NOTE Blood is convenient to sample and thus is often the fluid of choice for kinetic parameter and absorption studies. It is necessary to specify whether analysis is on whole blood, serum or plasma and to provide validation of this choice. Binding to circulating proteins or red cells can be determined in vitro.

7.7 There shall be sufficient data points with adequate time intervals to allow determination of kinetic parameters. In theory, this should cover several terminal half-lives; in practice, the constraints of the analytical method may necessitate a compromise.

Toxicokinetic evaluation of characterized leachables and degradation products can serve as an input to various parts of the biological evaluation for a medical device. Use of toxicokinetic data in a biological evaluation may include, but is not limited to:

a) estimating a clinically relevant systemic exposure dose from mathematical modelling for the toxicological risk assessment of hazardous constituents known to be released from a device;

b) identification of potential target organs of toxicity that may require further investigation in a systemic toxicity study;

c) determining the systemic absorption and tissue concentration of a pharmacologically active ingredient at its target site and off target sites;

d) providing data on the presence or absence of device constituents in specific tissues and organs to inform on bioaccumulation and potential health risks from long term exposure;

e) detection of metabolites;

f) understanding exposure route differences in bioavailability for route-to-route extrapolation.

Where toxicokinetic data has been generated via mathematical modelling or by an animal study, its use as part of the overall biological evaluation for a device shall be justified and documented.

1. 
   (informative)
   
   Mathematical modelling
   1. General

This annex provides an overview of mathematical models that may be used to determine toxicokinetic parameters relevant to degradation products and leachables from medical devices. It is advisable to consult toxicologist(s) or pharmacokineticist(s) with experience in toxicokinetics in the selection of method(s). The models are usually categorized as classical compartment open models and physiologically based toxicokinetic (PBTK) models.

• 1. Classical compartment open models
     1. General

Classical compartment open models are used to estimate concentration-time course of a chemical and its metabolite(s), usually in blood or plasma, or urine or tissue or organs for repeated or continuous exposures. Several models are in use varying in complexity and information obtained.

• • 1. Zero-order kinetics

The zero-order kinetic describes a situation where the elimination rate is constant without being dependant on the concentration of the chemical. This occurs often with chemicals that saturate metabolic enzymes or membrane transport systems increasing the risk of toxic reactions.

NOTE 1 Ethanol exhibits zero-order kinetics.

NOTE 2 The formula for calculating the rate (k) at time t where [A]₀ is the initial concentration and [A]_t is the concentration at time t is:

[A.1]

• • 1. First order kinetics

First order kinetics describes a situation where the rate of elimination is proportional to the concentration of the chemical. This implies that the elimination is more efficient when the concentration is high.

NOTE 1 Caffeine exhibits first-order kinetics

NOTE 2 The formula for calculating the rate (k) at time t where [A]₀ is the initial concentration and [A]_t is the concentration at time t is:

[A.2]

• • 1. Two compartments model

The two-compartment model describes a situation involving a central and a peripheral compartment. The chemical is introduced in the central compartment (i.e. blood stream, kidney, liver) and distributed to the peripheral compartment. The chemical is assumed to be eliminated from the central compartment only and redistributed from the peripheral compartment as the concentration in the central compartment decreases. For simplification, it is assumed that elimination from the central compartment and movement between the compartments follow first order kinetics.

• • 1. Michaelis-Menten kinetics

The Michaelis-Menten kinetics describe elimination that is dependent on rate limiting endogenous substances and how the elimination rate of a chemical is dependent on concentration of the endogenous substance(s). There are two distinct different situations, saturation of the endogenous substance i.e., carrier molecules or cell receptors and a surplus of these. In the first case, the elimination rate is constant and related to the rates of binding and release between the chemical and the endogenous substance.

• 1. Physiologically based toxicokinetic (PBTK) models

Physiologically based toxicokinetic (PBTK) models are mathematical descriptions of how a chemical enters the body, absorbs into the blood, moves between body tissues and the blood, and how the body metabolizes and eliminates the chemical. PBTK are useful tools for gaining rich insight into the kinetics of toxicants beyond what classic toxicokinetic models can provide. Such models make possible to describe the time course of distribution of toxicants to any organ or tissue, and to estimate the effects of changing physiologic parameters on tissue concentrations. Physiologically based toxicokinetic (PBTK) modelling is based on a priori knowledge of the physicochemical properties of a chemical coupled with biochemistry and realistic anatomy and physiology of relevant organ or tissue. PBTK models integrate a chemical and organ and tissue physiology into a mathematical modelling framework. An advantage of the PBTK is that the model can be adapted to different exposure situations, physiological conditions, and species by using model parameter values relevant to the system(s) in question. The availability and quality of the a priori data can be a limitation for the outcome of PBTK modelling.

PBTK models can be simplistic, with a small number of parameters that are important for describing the movement of a chemical in the body, or complex with many parameters that dictate chemical fate and movement. The choice between simplistic or complex PBTK model depends on what health question is being answered and the amount and type of information that is available to develop the model. Simplistic PBTK models are often used to get a quick snapshot of a chemical’s fate and movement in the body, typically if the target organ is known. In other instances, more complex PBTK models are used if the health question being answered requires precise details on how the chemical distributes to multiple tissues and organs.

1. 
   (informative)
   
   Information on animal toxicokinetic testing
   1. General considerations

B.1.1 The study should be performed on an appropriate sex and species; consider utilizing the same species used for the systemic toxicity studies. The animal welfare conditions should be as recommended in guidelines for the care and use of animals according to ISO 10993‑2.

The total number of animals employed, number of samples harvested from each animal as well as sufficient sample replicates generated per evaluation interval should be critically evaluated to obtain meaningful data for toxicokinetic evaluation. For instance, no less than n=2 replicates per sex per timepoint would be acceptable to generate a half-life from blood or plasma samples. Dose balance studies typically use 3 to 5 animals per sex per group to obtain reliable data.

B.1.2 A non-radiolabelled test substance may be utilized provided that suitable validated assay procedures for the test substance in the relevant samples exist and the metabolism of the test substance is well characterized.

B.1.3 If necessary, the test substance should be radiolabelled in a metabolically stable position, preferably with ¹⁴C or ³H, and of suitable radiochemical purity (>97 %). When using ³H, the possibility of tritium exchange should be considered. The specific activity and radiochemical purity of the test substance shall be known and reported.

B.1.4 The test substance should be administered by an appropriate route. This route should be relevant to the use of the medical device. The test substance should be prepared in a suitable vehicle taking into account the physicochemical properties of the test substance (leachable or degradation product) using appropriate route and dose of administration. The stability of the test substance in the vehicle shall be known and reported.

NOTE The study design can require the inclusion of other route(s) for comparison of percent absorption.

B.1.5 In dose balance studies, animals should be housed only in metabolism cages. for up to 7 d. See B.2.4.1 NOTE.’

B.1.6 Urine and faeces should be collected in low temperature vessels (or in vessels containing preservative that does not interfere with the analysis) to prevent post-elimination microbial or spontaneous modification. Blood for whole-blood or plasma analysis should be collected in the presence of a suitable anticoagulant.

B.1.7 Controls should, wherever possible, be collected prior to dosing. In some studies, collection of controls (e.g., tissues) is not possible from the test animals and these should be obtained from a control group.

B.1.8 Collection times should be appropriate to the type of study being performed, and may be carried out, as necessary, over periods of minutes, hours, days, weeks or even months. For studies involving excreta, this is usually a 24 h period over at least 96 h. Recovery of test article from excreta should target > 90 % of administered dose.

Where blood sampling is required, blood is collected according to a specified schedule to cover several terminal half-lives of leachable elimination. Blood sampling shall be limited to no more than 10 % of an animal's total circulating blood volume (CBV) for a single draw, and a total of 15 % over a 2-week period; fluid replacement is often recommended for sampling exceeding 10 % of the CBV, and multiple draws should be spaced out appropriately.

Larger animals may be needed as test systems to obtain sufficient sample volume to detect low concentrations of chemical(s) of interest. Alternatively, terminal collection (e.g. 2 mice/timepoint for x timepoints) to define a toxicokinetic profile may be necessary when employing rodent species.

B.1.9 Toxicokinetic animal studies should be performed in accordance with good laboratory practice or laboratory quality assurance controls.

B.1.10 The study report shall include the following information, where relevant:

a) strain and source of animals, age, sex (if females indicate reproductive state), environmental conditions, diet;

b) test substance and sample, purity, stability, formulation, amount administered;

c) test conditions, including route of administration;

d) assay methods, extraction, detection, validation/qualification;

e) overall recovery of material;

f) tabulation of individual results at each time point;

g) quality standard or good laboratory practice compliance statement;

h) presentation and discussion of results;

i) interpretation of results.

• 1. Guidance on specific types of tests
     1. General

B.2.1.1 The study should be designed to provide the necessary information for risk assessment, and therefore it is usually not necessary to examine all aspects.

B.2.1.2 Absorption, distribution, metabolism and excretion studies are a range of studies capable of being performed either individually, examining one of these aspects, or collectively, examining several aspects in one study.

B.2.1.3 Depending on the design of the study, a number of kinetic parameters may be determined including absorption rate, area under the plasma concentration versus time curve, area under the first moment plasma concentration versus time curve, volume of distribution, c_max, t_max, half-life, mean residence time, elimination rate and clearance.

B.2.1.4 Kinetic parameters can only be determined for a particular molecular species and hence the assay needs to be specific and sensitive to this molecular species. True kinetic parameters of a relevant compound can only be determined following intravenous administration. It may therefore be necessary to include a limited intravenous administration study in the design of the kinetic parameter studies. This allows the fraction of the dose absorbed to be calculated and this serves as a correction in estimating parameters in other studies.

In some instances, intra-arterial administration should be considered as some compounds are known to be cleared through the pulmonary system.

B.2.1.5 The appropriate kinetic model should be used in determining the kinetic parameters. A number of computer programs exist for estimating kinetic parameters. The software should be validated prior to use and this validation should be documented. The assumptions entered into the program and the choices in modelling should be documented.

• • 1. Absorption

Absorption depends on the route of administration, the physicochemical form of the test substance and the vehicle. It can be estimated from blood, serum, excreta and tissue concentrations. Bioavailability studies may be considered. The choice of the appropriate type of study depends on the other information required, availability of radiolabelled material and assay method. The absorption rate constant can be estimated reliably only if sufficient samples are taken in the absorption phase.

NOTE In vitro methods exist which can give important information on gastrointestinal and dermal absorption of chemicals.

• • 1. Distribution

B.2.3.1 Distribution studies generally require radiolabelled compounds.

Studies may be

— quantitative, determining levels in dissected tissues,

— qualitative, using whole-body autoradiography (WBA), or

— semiquantitative, using graded WBA reference doses.

B.2.3.2 In general, sampling times in distribution studies may be based on kinetic data and will depend on test sample elimination. Multiple sampling times may be used. Sampling is normally more frequent in the early phase of absorption and elimination; however, samples need to be obtained over as much of the elimination phase as possible. The major determinant is often assay sensitive.

• • 1. Metabolism and excretion

B.2.4.1 Metabolism cages should permit a separate collection of urine and faeces throughout the study. The use of metabolic cages designed for the collection of CO₂ and volatile metabolites should be considered if relevant for the excretion. For studies of up to 14 d, the urine and faeces should be individually collected over 24 h intervals until the end of the experiment. In some study designs, animals may be euthanized at intermediate times for evaluation of tissue distribution. Samples may be collected prior to 24 h when it is probable that the test substance or its metabolites will be rapidly excreted. For studies of longer duration, sampling over the initial period should occur as for the short-term studies. Thereafter, samples should be obtained for a continuous 24 h period per assessment period.

NOTE The use of metabolism cages for prolonged periods might be detrimental to animal welfare. Therefore, at the longer times, representative discontinuous samples can be collected and these results extrapolated to continuous sampling.

B.2.4.2 The carcasses and/or target organs of the individual animals should be retained for analysis, and blood collected for analysis of plasma and whole-blood concentrations. After collection of the samples from the metabolism cages at the sacrifice time, the cages and their traps should be washed with an appropriate solvent. The resulting washes can be pooled and a representative fraction retained for analysis.

B.2.4.3 The recovery or calculated recovery of a test substance should ideally be (100 ± 10) % when a radiolabelled compound is used. The recovery range specified might not be achievable in all cases, and reasons for any deviation should be stated and discussed in the report. The amount of test substance in each fraction should be analysed by suitably robust procedures (e.g. inclusion of appropriate standards) for either a radiolabelled or non-radiolabelled compound in the appropriate milieu. Where a radiolabelled compound is used, both parent compound and metabolites are assessed unless a specific assay is used.

B.2.4.4 Levels of radioactivity in the biological milieu should be determined, for example, by liquid scintillation counting; however, it should be stressed that this represents a mixed concentration of compound and metabolites, and no kinetic parameters can be derived from it. Where isolation of metabolites is considered necessary, this may involve a number of extractions and chromatographic procedures (e.g., high-pressure liquid chromatography, thin layer chromatography, gas-liquid chromatography), and the resulting material should be characterized by chemical methods and a variety of physical chemistry techniques (e.g., mass spectrometry, nuclear magnetic resonance spectroscopy).

B.2.4.5 The use of tissues, cells, homogenates and isolated enzymes for the study of metabolism in vitro is well documented. These methods identify potential metabolism which may not occur in vivo unless the compound is available at the appropriate site. The extents and rates of metabolism in vitro compared to in vivo will often differ.

Annex ZA
(informative)

Relationship between this European Standard the General Safety and Performance Requirements of Regulation (EU) 2017/745 aimed to be covered

This European standard has been prepared under M/575 to provide one voluntary means of conforming to the General Safety and Performance Requirements of Regulation (EU) 2017/745 of 5 April 2017 concerning medical devices [OJ L 117] and to system or process requirements including those relating to quality management systems, risk management, post-market surveillance systems, clinical investigations, clinical evaluation or post-market clinical follow-up.

Once this standard is cited in the Official Journal of the European Union under that Regulation, compliance with the normative clauses of this standard given in Table ZA.1 and application of the edition of the normatively referenced standards as given in Table ZA.2 confers, within the limits of the scope of this standard, a presumption of conformity with the corresponding General Safety and Performance Requirements of that Regulation, and associated EFTA Regulations.

Where a definition in this harmonised standard differs from a definition of the same term set out in Regulation (EU) 2017/745, the differences shall be indicated in the Annex Z. For the purpose of using this standard in support of the requirements set out in Regulation (EU) 2017/745, the definitions set out in this Regulation prevail.

Where the European standard is an adoption of an International Standard, the scope of this document can differ from the scope of the European Regulation that it supports. As the scope of the applicable regulatory requirements differ from nation to nation and region to region the standard can only support European regulatory requirements to the extent of the scope of the European Regulation for medical devices ((EU) 2017/745).

NOTE 1 Where a reference from a clause of this standard to the risk management process is made, the risk management process needs to be in compliance with Regulation (EU) 2017/745. This means that risks have to be ‘reduced as far as possible’, ‘reduced to the lowest possible level’, ‘reduced as far as possible and appropriate’, ‘removed or reduced as far as possible’, ‘eliminated or reduced as far as possible’, ’removed or minimized as far as possible’, or ‘minimized’, according to the wording of the corresponding General Safety and Performance Requirement.

NOTE 2 The manufacturer’s policy for determining acceptable risk must be in compliance with General Safety and Performance Requirements 1, 2, 3, 4, 5, 8, 9, 10, 11, 14, 16, 17, 18, 19, 20, 21 and 22 of the Regulation.

NOTE 3 When a General Safety and Performance Requirement does not appear in Table ZA.1, it means that it is not addressed by this European Standard.

NOTE 4 General Safety and Performance Requirements listed in Table ZA.1 are specific to biological safety for patients only.

Table ZA.1 — Correspondence between this European Standard and Annex I
of Regulation (EU) 2017/745 [OJ L 117] and to system or process requirements including those relating to quality management systems, risk management, post-market surveillance systems, clinical investigations, clinical evaluation or post-market clinical follow-up

Table ZA.2 — Applicable Standards to confer presumption of conformity as described in this Annex ZA

The documents listed in the Column 1 of table ZA.2, in whole or in part, are normatively referenced in this document, i.e. are indispensable for its application. The achievement of the presumption of conformity is subject to the application of the edition of Standards as listed in Column 4 or, if no European Standard Edition exists, the International Standard Edition given in Column 2 of table ZA.2.

Table ZA.3 — Correspondence between this European Standard and Annex II
of Regulation (EU) 2017/745 [OJ L 117]

WARNING 1 — Presumption of conformity stays valid only as long as a reference to this European standard is maintained in the list published in the Official Journal of the European Union. Users of this standard should consult frequently the latest list published in the Official Journal of the European Union.

WARNING 2 — Other Union legislation may be applicable to the product(s) falling within the scope of this standard.

Bibliography

[1] ISO 10993‑2, Biological evaluation of medical devices — Part 2: Animal welfare requirements

[2] ISO 10993‑9, Biological evaluation of medical devices — Part 9: Framework for identification and quantification of potential degradation products

[3] ISO 10993‑17, Biological evaluation of medical devices — Part 17: Toxicological risk assessment of medical device constituents

[4] ISO/TR 10993‑22, Biological evaluation of medical devices — Part 22: Guidance on nanomaterials

[5] ISO 14971, Medical devices — Application of risk management to medical devices

[6] EURL ECKVAM Toxicokinetics. https://eurl-ecvam.jrc.ec.europa.eu/validation-regulatory-acceptance/toxicokinetics/toxicokinetics (2015-09-14)

[7] FDA guideline for industry. Pharmacokinetics: Guidance for repeated dose tissue distribution studies. Fed. Regist. 1995, 60 pp. 11274–11275

[8] Harmonised Tripartite Guideline — Note for Guidance on Toxicokinetics: The Assessment of Systemic Exposure in Toxicity Studies, ICH S3A, 1994

[9] Harmonised Tripartite Guideline — Pharmacokinetics: Guidance for Repeated Dose Tissue Distribution Studies, ICH S3B, 1994

[10] ICH S3A Toxicokinetic guidance reference — Note for Guidance on Toxicokinetics: A Guidance for Assessing Systemic Exposure in Toxicity Studies (CPMP/ICH/384/95.)

[11] International Programme on Chemical Safety. Environmental Health Criteria 57: Principles of toxicokinetic studies. World Health Organization, Geneva, 1986

[12] OECD Guideline for Testing of Chemicals, 417: Toxicokinetics, Adopted: 22 July 2010

[13] U.S. Department of Health and Human Services, Food and Drug Administration. Physiologically Based Pharmacokinetic Analyses — Format and Content. Guidance for Industry. 2018.

[14] U.S. Environmental Protection Agency, Office of Research and Development. Physiologically-Based Pharmacokinetic (PBPK) Models. 2018 (www.epa.gov/research)

[15] AFZELIUS L. et al. State-of-the-art tools for computational site of metabolism predictions: Comparative analysis, mechanistic insights and future applications. Drug Metab. Rev. 2007, 39 pp. 61–86

[16] BAILLIE T.A. et al. Contemporary Issues in Toxicology: Drug Metabolites in Safety Testing, Toxicol. Appl. Pharmacol. 2002, 182 pp. 188–196

[17] BEATTY D.A., PIEGORSCH W.W. Optimal statistical design for toxicokinetic studies. Stat. Methods Med. Res. 1997, 6 pp. 359–376

[18] BOKKERS B.G., SLOB W. Deriving a data-based interspecies assessment factor using the NOAEL and the benchmark dose approach. Crit. Rev. Toxicol. 2007, 37 pp. 355–373

[19] BOOBIS A.R. et al. Interlaboratory comparison of the asessment of P450 activities in human hepatic microsomal samples. Xenobiotica. 1998, 28 pp. 493–506

[20] BOROUJERDI M. Pharmacokinetics and toxicokinetics. CRC Press 2015

[21] BOUVIER D’YVOIRE M., PRIETO P., BLAAUBOER B.J., BOIS F.Y., BOOBIS A., BROCHOT C. et al. Physiologically-based Kinetic Modelling (PBK Modelling): meeting the 3Rs agenda. The report and recommendations of ECVAM Workshop 63. Altern. Lab. Anim. 2007, 35 pp. 661–671

[22] CASARETT & DOULL’S Essentials of Toxicology, 2e. KLAASSEN C.D. & WATKINS J.B. Eds. Chapter 7. Toxicokinetics

[23] CHEN, C.-Y. LIN Z. Exploring the potential and challenges of developing physiologically-based toxicokinetic models to support human health risk assessment of microplastic and nanoplastic particles. Environ Inter 186: 2024 (https://doi.org/10.1016/j.envint.2024.108617)

[24] CLARK D.E. (Theme ed), Computational methods for the prediction of ADME and toxicity. Adv. Drug Deliv. Rev. 2002, 4 pp. 253–451

[25] COECKE S., BLAAUBOER B.J., ELAUT G., FREEMAN S., FREIDIG A., GENSMANTEL N. et al. Toxicokinetics and metabolism. Altern. Lab. Anim. 2005, 33 pp. 147–175

[26] COECKE S., PELKONEN O., LEITE S.B., BERNAUER U., BESSEMS J.G., BOIS F.Y. et al. Toxicokinetics as key to the integrated toxicity risk assessment based primarily on non-animal approaches. Toxicol. In Vitro. 2012, 27 pp. 1570–1577

[27] COSSON V.F., FUSEAU E., EFTHYMIOPOULOS C., BYE A., Mixed Effect Modeling of Sumatriptan Pharmacokinetics During Drug Development. I, Interspecies Allometric Scaling. J. Pharmacokinet. Pharmacodyn. 1997, 25 pp. 149–167

[28] CROSS D.M., BAYLISS M.K. A commentary on the use of hepatocytes in drug metabolism studies during drug discovery and drug development. Drug Metab. Rev. 2000, 32 pp. 219–240

[29] DIXIT R. Toxicokinetics: Fundamentals and applications in drug development and safety assessment. In: Biological Concepts and Techniques in Toxicology. Marcel Dekker, 2006, pp. 117–60

[30] DORNE J.L. Human variability in hepatic and renal elimination: implications for risk assessment. J. Appl. Toxicol. 2007, 27 pp. 411–420

[31] DORNE J.L. Impact of inter-individual differences in drug metabolism and pharmacokinetics on safety evaluation. Fundam. Clin. Pharmacol. 2004, 18 pp. 609–620

[32] DORNE J.L., WALTON K., RENWICK A.G. Human variability in xenobiotic metabolism and pathway-related uncertainty factors for chemical risk assessment: a review. Food Chem. Toxicol. 2005, 43 pp. 203–216

[33] DORNE J.L., WALTON K., SLOB W., RENWICK A.G. Human variability in polymorphic CYP2D6 metabolism, is the kinetic default uncertainty factor adequate? Food Chem. Toxicol. 2002, 40 pp. 1633–1656

[34] FILSER J.G. Toxicokinetic Models. In: Reichl, FX. & Schwenk, M. eds. Regulatory Toxicology. Springer 2021, Berlin, Heidelberg. pp. 1-11. https://doi.org/10.1007/978-3-642-36206-4_49-2

[35] FORKERT P.-G. Mechanisms of 1,1-dichloroethylene-induced cytotoxicity in lung and liver. Drug Metab. Rev. 2001, 33 pp. 49–80

[36] FRANTZ S.W. et al., Pharmacokinetics of ethylene glycol II. Tissue distribution, dose dependent elimination, and identification of urinary metabolites following single intravenous, peroral or percutaneous doses in female Sprague-Dawley rats and CD-1® mice, Xenobiotica, 26, pp. 1195-1220, 1996

[37] FUHR U. Induction of Drug Metabolising Enzymes: Pharmacokinetic and Toxicological Consequences in Humans. Clin. Pharmacokinet. 2000, 38 pp. 493–504

[38] GEHRING R., VAN DER MERWE D. Toxicokinetic-toxicodynamic modelling. In: GUPTA R.C. ed. Biomarkers in Toxicology. Academic Press 2014, pp. 149-153, ISBN 978-0-12-404630-6, DOI https://doi.org/10.1016/C2012-0-01373-7

[39] GUENGERICH F.P., RENDIC S., eds. Human cytochrome P450 enzymes, a status report summarizing their reactions, substrates, inducers and inhibitors – 1st update. Special issue on Human cytochromes P450 (Human CYPs), Drug Metab. Revs., 34, pp. 1-450, 2002

[40] GUNDERT-REMY U., SONICH-MULLIN C., IPCS Uncertainty and Variability Planning Workgroup and Drafting Group. (International Program on Chemical Safety). The use of toxicokinetic and toxicodynamic data in risk assessment: an international perspective. Sci. Total Environ. 2002, 288 pp. 3–11

[41] HANSCH C., MEKAPATI S.B., KURUP A., VERMA R.P. QSAR of Cytochrome P450. Drug Metab. Rev. 2004, 36 pp. 105–156

[42] HEDAYA M.A. Basic pharmacokinetics. CRC Press pharmacy education series. CRC Press, Boca Raton, FL, 2007

[43] HEINONEN M., OILA O., NORDSTRÖM K. Current issues in the regulation of human tissue-engineering products in the European Union. Tissue Eng. 2005, 11 pp. 1905–1911

[44] HENGSTLER J.G. et al., Cryopreserved primary hepatocytes as a constantly available in vitro model for the evaluation of human and animal drug metabolism and enzyme induction. Drug Metab. Revs., 32, pp. 81-118, 2000

[45] HOUSTON J.B., GALETIN A. Progress towards prediction of human pharmacokinetic parameters from in vitro technologies. Drug Metab. Rev. 2003, 35 pp. 393–415

[46] JOHANSON G. Toxicokinetics and Modeling, In: McQueen C.A. ed. Comprehensive Toxicology (Third Edition), Elsevier, 2018, pp. 165-187, ISBN 9780081006016, https://doi.org/10.1016/B978-0-12-801238-3.01889-4.

[47] KEEGAN G.M., LEARMONTH I.D., CASE C.P. Orthopaedic metals and their potential toxicity in the arthroplasty patient. A review of current knowledge and future strategies. J. Bone Joint Surg. Br. 2007, 89-B pp. 567–573

[48] KIRMAN C.R., SWEENEY L.M., CORLEY R., GARGAS M.L. Using physiologically-based pharmacokinetic modeling to address nonlinear kinetics and changes in rodent physiology and metabolism due to aging and adaptation in deriving reference values for propylene glycol methyl ether and propylene glycol methyl ether acetate. Risk Anal. 2005, 25 pp. 271–284

[49] KRISHNAN K. & ANDERSEN M.E. Physiologically based pharmacokinetic modelling in toxicology. In: Hayes A.W. ed. Principles and Methods of Toxicology, 3rd edn, Raven Press, New York, 1994, pp.149–188.

[50] KRISHNAN K., JOHANSON G. Physiologically-based pharmacokinetic and toxicokinetic models in cancer risk assessment, J. Environ. Sci. Health, Part C. Environ. Carcinogen. Ecotox. Rev. 2005, 23 pp. 31–53

[51] KWON Y., ed. Handbook of essential pharmacokinetics, pharmacodynamics and drug metabolism for industrial scientists. Springer Verlag, 2001

[52] LEWIS D.F.V., DICKINS M. Baseline lipophilicity relationships in human cytochromes P450 associated with drug metabolism. Drug Metab. Rev. 2003, 35 pp. 1–18

[53] LINHART I. Stereochemistry of styrene biotransformation. Drug Metab. Rev. 2001, 33 pp. 353–368

[54] LIPSCOMB J.C., OHANIAN E.V., eds. Toxicokinetics and risk assessment. Informa, 2006

[55] MAHMOOD I., GREEN M.D., FISHER J.E. Selection of the first-time dose in humans: comparison of different approaches based on interspecies scaling of clearance. J. Clin. Pharmacol. 2003, 43 pp. 692–697

[56] MCLANAHAN E.D., EL-MASRI H.A., SWEENEY L.M., KOPYLEV L.Y., CLEWELL H.J., JOHN F. WAMBAUGH, J.F. AND SCHLOSSER P.M., Physiologically Based Pharmacokinetic Model Use in Risk Assessment—Why Being Published Is Not Enough. Toxicol. Sci. 2012, 126 pp. 5–15

[57] MEEK B., RENWICK A., SONICH-MULLIN C. International Programme on Chemical Safety: Practical application of kinetic data in risk assessment — an IPCS initiative. Toxicol. Lett. 2003, 138 pp. 151–160

[58] MIZUTANI T. PM frequencies of major CYPs in Asians and Caucasians. Drug Metab. Rev. 2003, 35 pp. 99–107

[59] NESTOROV I. Whole Body Pharmacokinetic Models: Review Article. Clin. Pharmacokinet. 2003, 42 pp. 883–908

[60] POULIN P., THEIL F.-P. Prediction of pharmacokinetics prior to In Vivo studies. II. Generic physiologically based pharmacokinetic models of drug disposition. J. Pharm. Sci. 2002, 91 pp. 1358–1370

[61] RENWICK, A.G. Toxicokinetics. In: DUFFUS J.H., WORTH H.G.J. (eds), Fundamental Toxicology, Royal Society of Chemistry, 2007, pp 25-42.

[62] Roffey S.J., Obach R.S., Gedge J.I., Smith D.A. What is the objective of the mass balance study? A retrospective analysis of data in animal and human excretion studies employing radiolabelled drugs. Drug Metab. Rev. 2007, 39 pp. 17–44

[63] ROSSO F., MARINO G., GIORDANO A., BARBARISI M., PARMEGGIANI D., BARBARISI A. Smart materials as scaffolds for tissue engineering. J. Cell. Physiol. 2005, 203 pp. 465–470

[64] SCARFE G.B. et al., 19F-NMR and directly coupled HPLC-NMR-MS investigations into the metabolism of 2-bromo-4-trifluoromethylaniline in rat: a urinary excretion balance study without the use of radiolabelling, Xenobiotica, 28, pp. 373-388, 1998

[65] SCHROEDER K., BREMM K.D., ALÉPÉE N. BESSEMS J.G.M., BLAAUBOER B. Report from the EPAA workshop: In vitro ADME in safety testing used by EPAA industry sectors. Toxicol. In Vitro. 2011, 25 pp. 589–604

[66] SHEINER L.B., STEIMER J.-L. Pharmacokinetic/pharmacodynamic modelling in drug development. Annu. Rev. Pharmacol. Toxicol. 2000, 40 pp. 67–95

[67] SHEPARD T., SCOTT G., COLE S., NORDMARK A., BOUZOM F. Physiologically Based Models in Regulatory Submissions: Output From the ABPI/MHRA Forum on Physiologically Based Modeling and Simulation. CPT Pharmacometrics Syst. Pharmacol. 2015, 4 pp. 221–225

[68] SKORDI E., WILSON I.D., LINDON J.C., NICKOLSON J.K. Characterization and quantification of metabolites of racemic ketoprofen excreted in urine following oral administration to man by 1H-NMR spectroscopy, directly coupled HPLC-MS and HPLC-NMR, and circular dichroism. Xenobiotica. 2004, 34 pp. 1075–1089

[69] SLATTER J.G. et al., Pharmacokinetics, toxicokinetics, distribution, metabolism and excretion of linezolid in mouse, rat and dog, Xenobiotica, 32, pp. 907-924, 2002

[70] SMITH D.A., WALKER D.K., VAN DE WATERBEEMD H. Pharmacokinetics and metabolism in drug design. Methods and principles in medicinal chemistry, 31. Wiley-VCH, Weinheim, 2006

[71] SUN H.F., MEI L., CUNXIAN S., CUI X., WANG P. The in vivo degradation, absorption and excretion of PCL-based-implant. Biomaterials. 2006, 27 pp. 1735–1740

[72] TONNELIER A., COECKE S., ZALDÍVAR J.M. Screening of chemicals for human bioaccumulative potential with a physiologically based toxicokinetic model. Arch. Toxicol. 2012, 86 pp. 393–403

[73] TOZER T.N., ROWLAND M. Introduction to Pharmacokinetics and Pharmacodynamics — The Quantitative Basis of Drug Therapy. Lippincott, Williams & Wilkins, 2006

[74] VENKATAKRISHNAN K., VON MOLTE L.L., GREENBLATT D.J. Human drug metabolism and cytochromes P450: Application and relevance of in vitro models. J. Clin. Pharmacol. 2001, 41 pp. 1149–1179

[75] VOGEL H.G. et al., eds. Drug discovery and evaluation: safety and pharmacokinetic assays. Springer Verlag, Berlin, 2006

[76] WAGNER C., ZHAO P., PAN Y., HSU V., GRILLO J., HUANG S.M. et al. Application of Physiologically Based Pharmacokinetic (PBPK) Modeling to Support Dose Selection: Report of an FDA Public Workshop on PBPK. CPT Pharmacometrics Syst. Pharmacol. 2015, 4 pp. 226–230

[77] WALKER D.K. The use of pharmacokinetic and pharmacodynamic data in the assessment of drug safety in early drug development. Br. J. Clin. Pharmacol. 2004, 58 pp. 601–608

[78] WINTER M.E., ed. Basic clinical pharmacokinetics. Lippincott, Williams & Wilkins, Fifth Edition, 2009

[79] YACOBI A., SKELLY J.P., BATRA V.K. Toxicokinetics and new drug development. Pergamon Press, 1989

[80] YAN Z., CALDWELL G.W. Metabolism profiling, and cytochrome P450 inhibition & induction in drug discovery. Curr. Top. Med. Chem. 2001, 1 pp. 403–425

[81] YANNAS I.V. Synthesis of Tissues and Organs. ChemBioChem. 2004, 5 pp. 26–39

[82] ZHOU S. et al., Drug bioactivation, covalent binding to target proteins and toxicity relevance, Drug Metab. Rev., 37, pp. 41-214, 2005