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Evaluation of SARs for the prediction

of skin irritation/corrosion
potential–structural inclusion rules in
the BfR decision support system
a a a a
A. Gallegos Saliner , I. Tsakovska , M. Pavan , G. Patlewicz
a
& A. P. Worth
a
European Chemicals Bureau, Institute for Health and Consumer
Protection, Joint Research Centre, European Commission, 21020
Ispra (VA), Italy

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To cite this article: A. Gallegos Saliner, I. Tsakovska, M. Pavan, G. Patlewicz & A. P. Worth (2007):
Evaluation of SARs for the prediction of skin irritation/corrosion potential–structural inclusion rules
in the BfR decision support system , SAR and QSAR in Environmental Research, 18:3-4, 331-342

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 SAR and QSAR in Environmental Research,
Vol. 18, Nos. 3–4, May–June 2007, 331–342

Evaluation of SARs for the prediction of skin

irritation/corrosion potential – structural inclusion rules
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in the BfR decision support systemy

A. GALLEGOS SALINER*, I. TSAKOVSKA, M. PAVAN,
G. PATLEWICZ and A. P. WORTH
European Chemicals Bureau, Institute for Health and Consumer Protection,
Joint Research Centre, European Commission, 21020 Ispra (VA), Italy

(Received 11 May 2006; in final form 27 September 2006)

The German Federal Institute for Risk Assessment (BfR) has developed a Decision Support
System (DSS) to assess certain hazardous properties of pure chemicals, including skin and eye
irritation/corrosion. The BfR – DSS is a rule-based system that could be used for the regulatory
classification of chemicals in the European Union. The system is based on the combined use of
two predictive approaches: exclusion rules based on physicochemical cut-off values to identify
chemicals that do not exhibit a certain hazard (e.g., skin irritation/corrosion), and inclusion
rules based on structural alerts to identify chemicals that do show a particular toxic potential.
The aim of the present study was to evaluate the structural inclusion rules implemented in the
BfR–DSS for the prediction of skin irritation and corrosion. The following assessments were
performed: (a) a confirmation of the structural rules by rederiving them from the original
training set (1358 substances), and (b) an external validation by using a test set of 200 chemicals
not used in the derivation of the rules. It was found as a result that the test data set did not
match the training set relative to the inclusion of structural alerts associated with skin
irritation/corrosion, albeit some skin irritants were in the test set.

Keywords: Structure–Activity Relationship (SAR); Structural alert; Skin irritation/corrosion;

BfR–Decision Support System (BfR–DSS); Structural inclusion rule

1. Introduction

1.1 Assessment of skin irritation/corrosion

The skin can be exposed to products and their ingredients either through use of
cosmetic and pharmaceutical products intended for use on the skin or through
accidental exposure. Therefore, the potential of a particular product or ingredient to
cause skin irritation or corrosion needs to be evaluated. Skin irritancy and corrosivity

*Corresponding author. Email: ana.gallegos@jrc.it

yPresented at the 12th International Workshop on Quantitative Structure-Activity Relationships in
Environmental Toxicology (QSAR2006), 8–12 May 2006, Lyon, France.

SAR and QSAR in Environmental Research

ISSN 1062-936X print/ISSN 1029-046X online ß 2007 Taylor & Francis
DOI: 10.1080/10629360701304014
 332 A. Gallegos Saliner et al.

are assessed in the hazard classification of chemicals for regulatory purposes, and in the
safety assessment of ingredients and mixtures used in industrial, pharmaceutical and
consumer products. The standard test method for assessing the potential of chemicals to
cause skin irritation and corrosion is the Draize rabbit skin test, which is included in
international regulatory requirements for the testing of chemicals. However, for
scientific, economic and animal welfare reasons, there is the need to use alternative
(non-animal) methods for the determination of local irritation and corrosion.
In the hazard classification of chemicals, the purpose is to assess the irritation/
corrosion potential according to classification schemes defined by regulatory authorities.
In the European Union (EU), the proposed REACH legislation (Registration,
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Evaluation, and Authorisation of Chemicals) [1–3] envisages an extensive system for

assessing the physicochemical, (eco)toxicological and environmental properties of New
and Existing Chemicals based on their tonnage [4]. At the international level, a unified
classification system for hazard endpoints is provided by the United Nations Globally
Harmonised System (GHS) for Classification and Labelling [5].
The safety assessments of cosmetic ingredients are aimed at demonstrating that
products will not cause adverse effects under the exposure conditions of intended use.
The EU Cosmetics Directive [6] imposes the legal obligation that cosmetics sold to
consumers are safe; thus, the main concern is the generation of false negatives.

1.2 BfR–Decision Support System

The German Federal Institute for Risk Assessment (BfR) has developed a Decision
Support System (DSS) for predicting local lesions [7–9]. It is used to assess chemicals for
skin/eye irritation/corrosion potential in support of regulatory decisions on classifica-
tion and labelling.
The DSS provides categorical predictions of hazard. The predictions are generated
from: (a) physicochemical data of the query chemical, by applying exclusion rules based
on physicochemical cut-off values [10]; and (b) from the structural formula of the
chemical, by applying inclusion rules based on structural alerts (SARs) [11]. The system
predicts the irritation and corrosion potential caused by a single contact of a chemical
with skin and/or eyes. In particular, the DSS applies the appropriate EU risk phrase
according to EU regulations, providing suitable data for legal classification and
labelling.
The current version of the DSS [12, 13] is based on a proprietary confidential training
set of 1358 chemicals with experimental data for skin and eye irritation and corrosion.
The training set was compiled from confidential data submitted under the EU New
Chemicals Notification procedure, and contained within the New Chemicals Database
(NCD) [14].
Recently, evaluations of the DSS physicochemical exclusion rules for eye irritation
and corrosion [15] and for skin irritation and corrosion [16] have been reported. The
aim of the present study was to evaluate the structural inclusion rules implemented in
the BfR–DSS for predicting skin irritation and corrosion. The following aspects were
addressed: an assessment of how the structural rules were derived from the rulebase
training set of 1358 substances, and an external validation using a test set of 200
chemicals not used in rule derivation. A related activity, the evaluation of the structural
inclusion rules for predicting eye irritation, has been undertaken in parallel by
Tsakovska et al. [17].
 Skin irritation/corrosion SAR 333

Table 1. R-phrases for skin irritation/corrosion.

Risk phrase Effect Description

R34 Causes burns Full thickness destruction of the skin

occurs as a result of up to 4 h exposure.
R35 Causes severe burns Full thickness destruction of the skin occurs as
a result of up to 3 min exposure.
R38 Irritating to skin Defined reversible inflammation of the skin as
a result of up to 4 h of exposure
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2. Definition of the endpoint

The classification and labelling of substances within the EU is based on the use of Risk
phrases (R-phrases). The definitions and the grounds upon which chemicals are
assigned R-phrases for skin irritation/corrosion are presented in table 1. The GHS
system adopts separate categories for skin irritation and corrosion [18].
The protocols recommended for the different required tests are described in the
Dangerous Substances Directive (DSD) and in the OECD Testing Guidelines for
chemicals. Test methods B.4 [19], and B.40 [20] in Annex V to the DSD deal separately
with skin irritation and skin corrosion, respectively. In the OECD test guidelines, the
corresponding test methods are included in Test Guideline 404 [21]. Recently, two
in vitro skin corrosion tests have been validated [22, 23]. The definitions according to
the OECD guidelines and to the GHS are given in the next sections.

2.1 Skin irritation

Dermal irritation is defined in OECD TG 404 as ‘the production of reversible damage
of the skin following the application of a test substance for up to 4 hours’ [21]. In the
EU this effect is assigned ‘Xi’ and the R-phrase R38. For the grading of dermal
responses, there is a consensus among all the authorities to use the Draize grading scale
for dermal lesions.

2.2 Skin corrosion

Dermal corrosion is defined in OECD TG 404 as ‘the production of irreversible damage
to skin’; namely, visible necrosis through the epidermis and into the dermis, following
the application of a test substance for up to four hours [21]. In the EU this effect is
assigned ‘C’ and either R34 or R35.

3. Method

3.1 Definition of the BfR structural inclusion rules

In the BfR–DSS, structural inclusion rules have been derived from empirical knowledge
relating specific local effects and structural formulae [8]. The rules are based on the
presence of structural alerts (SARs) and take the general form:
IF ðsubstructureÞ A THEN Effect B
 334 A. Gallegos Saliner et al.

The most recently updated lists of structural alerts for skin corrosion and for skin
irritation [11] are given in the three first columns of tables 2 and 3, respectively.

3.2 Evaluation of the BfR structural inclusion rules

The rulebase training set and the external test set were assessed by using Leadscope [24],
a data management and support system tool. Leadscope possesses a unique chemical
hierarchy containing over 27,000 chemical fingerprints. The hierarchy can be exploited
to perform specific structural searches.
In order to evaluate the structural rules, the compounds underpinning the skin
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corrosion and irritation structural alerts were evaluated by using Leadscope in different
phases. In the first run, only structures exactly matching the substructure (including
pre-defined constraints) were retrieved. In the second run, a less restricted search was
performed to see if a larger number of structures would be retrieved. In this case, any
restrictions defined in the BfR alerts were relaxed. For example, in the case of the alkyl
side chain constraint, closed aliphatic chains were allowed, but aromatic cycles,
heteroatoms and different functional groups were excluded. Finally, in the third run, for
those structural rules representing a low number of compounds, all compounds
containing the structural alert irrespective of any pre-defined limitations were
considered. The third run was performed to explore a possible extension to the existing
structural alert limits.

4. Results

4.1 Composition of skin irritant and corrosive compounds

The original rulebase training set comprised 1358 substances. Out of these substances,
17 non-irritant/corrosive polymers were excluded, leaving 1341 substances, including
1169 non-classified compounds (87.2%) and 172 skin irritants/corrosives (12.8%).
Among the 172 classified compounds, 95 substances (7.1%) were labelled R38, and
77 substances (5.7%) were labelled R34/R35.
The test set was made up of 200 substances, including 179 non-classified compounds
(89.5%) and 21 skin irritants/corrosives (10.5%). Among the classified compounds,
14 substances (7%) were labelled R38 and 7 substances (3.5%) were labelled R34/R35.
The composition of irritant, corrosive and non-classified substances for both datasets
is outlined in table 4.

4.2 Evaluation of the BfR structural inclusion rules

Table 2 details the number of structures which underpin each of the structural alerts for
skin corrosion in the training set (both first and second runs). In the test set, no
substances contained any of the skin corrosion alerts.
The number of structures containing structural alerts for skin irritation in the training
set is displayed in table 3. In the third run, since predominantly non-labelled
compounds were retrieved, investigations of potential structural rule extensions were
not feasible. No substance in the test set contained any of the skin irritation alerts.
 Downloaded by [University of Saskatchewan Library] at 20:12 14 October 2012

Table 2. BfR structural alerts for the prediction of skin corrosion (EU Risk phrases: R34 ¼ Causes burns; R35 ¼ Causes severe burns), and number of structures
containing the skin corrosion structural alerts in the training set (numbers in brackets indicate the number of misclassifications).

Limit/ First run Second run

ID Alert Compound class remarks restricted search relaxed search Total Coverage

1 Substituted benzoic Hal ¼ F, Cl R1–4 ¼ Any 5 (1) – 5 (1) 31.59%

O acid halogenides
C Hal
R1

R4 R2
R3

2a R–CH2–N¼C¼O Aliphatic isocyanates R ¼ Alkyl (aliphatic chain) – 1 1 5.26%

2b R–CH2–N¼C¼S Aliphatic iso(thio)cyanates R ¼ Alkyl (aliphatic chain) – – – 0%
3 Chlorosilanes R1–3 ¼ Any (e.g. further halogen) 3 – 3 15.79%
R2
R1 Si R3
Cl

4 Mixed oxy-carboxysilanes R1,2 ¼ Any 1 – 1 5.26%

CH3 O
R2 O Si O C R1
Skin irritation/corrosion SAR

CH3

5 Aliphatic amines R1 ¼ Alkyl (aliphatic chain which may contain – 7 7 36.84%

R1 ether functions) R2,3 ¼ H or alkyl (aliphatic chain)
N
R3 R2

6 Alkali salts of aliphatic R1 ¼ H or alkyl (aliphatic chain) 1 – 1 5.26%

R1 alcohols R2 ¼ Alkyl (aliphatic chain)
+
R2 CH O Alkali
335

1–6 ALL 10 (1) 8 18 (1) 100%

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336

Table 3. BfR structural alerts for the prediction of skin irritation (EU Risk phrase: R38 ¼ Irritating to skin), and number of structures containing skin irritation
structural alerts in the training set (numbers in brackets indicate false positives).

First run Second run

ID Alert Compound class Limit/remarks restricted search relaxed search Total Coverage

1 Substituted di-halogen- Hal ¼ F, Cl, Br, I R1–3 ¼ Any 2 – 2 9.09%

O benzoic acids
C OH
Hal R1
A. Gallegos Saliner et al.

Hal R2
R3

2a R–Si (OCH3)3 Tri-alkyloxysilanes R ¼ Alkyl (aliphatic chain) 1 – 1 4.55%

2b R–Si (OC2H5)3 Tri-alkyloxysilanes R ¼ Alkyl (aliphatic chain) 1 – 1 4.55%
2c Di-alkyloxysilanes R1,2 ¼ Alkyl (aliphatic chain) 3 (1) 1 (1) 4(2) 27.27%
R2

R1 Si(OCH3)2
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2d Di-alkyloxysilanes R1,2 ¼ Alkyl (aliphatic chain) – – – 0%

R2

R1 Si(OC2H5)

3 Aliphatic alcohols R1 ¼ Alkyl (aliphatic chain, saturated or – 1 (1) 1(1) 9.09%

R1 with C10–C20 chains containing double bonds) R2,3 ¼ H or
R2 C OH alkyl (aliphatic chain)
R3

4 Aliphatic aldehydes R ¼ Alkyl (aliphatic chain, saturated or 1 (1) (2) 1(3) 18.18%
O containing double bonds)
R C H

5 Aliphatic alpha-halogen esters Hal ¼ F, Cl, Br, I R1–2 ¼ Any, not 1 (1) – 1(1) 9.09%
R2 O containing nitrogen
Hal CH C O R1

6 Tri- and tetrahalogene benzenes Hal ¼ F, Cl, or Br 4 – 4 18.18%

Hal3–4
Skin irritation/corrosion SAR

1–6 ALL 13 (3) 2 (4) 15(7) 100%

337
 338 A. Gallegos Saliner et al.

Table 4. Composition of substances in the training and test set.

Non-labelled R38 R34/R35

N. % N. % N. %

Training set 1169 87.17 95 7.09 77 5.74

Test set 179 89.5 14 7.00 7 3.50

Table 5. Composition and number of skin corrosive (R34 and R35), and irritant substances (R38) predicted
in the training and test set using the inclusion rules coded as structural alerts in Leadscope. The cumulative
percentage of coverage of alerts for the first run, and the second run is included for the training set.
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R34/R35 R38

Pred Pred

Exp 1st 2nd Exp 1st 2nd

Training set 77 10 (1) 8 95 13 (3) 2 (4)

14.29% 24.68% 16.84% 23.16%
Test set 7 0 0 14 0 0

Exp: Number of classified chemicals.

Pred: Number of chemicals predicted with Leadscope. Numbers in brackets account for the number of misclassified
compounds.

4.3 Coverage of rules

A comparison of the composition of the training and test sets and the numbers of hits
obtained by applying the structural rules is given in table 5. The number of hits in the
first and second run using Leadscope is highlighted in the second column of each
category. The first subcolumn includes chemicals identified in the first run, and the
second subcolumn includes the additional chemicals identified in the second run. Since
no compounds were found in the test set which contained the structural alerts, only the
results for the training set will be discussed. The coverage of skin corrosives and
irritants was calculated by dividing the number of hits (which account for true and false
positives) by the composition for each class.
It should be noted that the relaxed search strategy (including mainly aliphatic rings)
increased the coverage quite dramatically: an increase of 73% for skin corrosives and an
increase of 38% for skin irritants. The coverages of the rulebase for irritant and
corrosive compounds, adjusted for the number of correctly and incorrectly predicted
compounds, are shown in figure 1 and table 6.
The coverage of alerts is very low: for the corrosion alerts, the total coverage is 24.7%
and the ‘true’ coverage, correcting for the single misclassified chemical, is 23.4%. For
irritation alerts, the total coverage of 23.2% decreases to a ‘true’ coverage of 15.8%
when a correction is made for the seven misclassified compounds. It is worth noting that
only misclassifications by one class were found: one compound predicted to be
corrosive was labelled R38 and seven compounds containing irritation alerts were
not labelled. There are no cases of corrosive compounds labelled as non irritants or
vice-versa. Since the structural rules do not predict the absence of effects, it is not
possible to make an assessment of false negatives by considering only the alerts.
 Skin irritation/corrosion SAR 339

100 95
90
80 77

70
Composition
60
‘True’ cover.
50
40 ‘False’ cover.
30
Total cover.
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22
18 19
20 15
10 7
1
0
R34/R35 R38

Figure 1. Coverage of structural alerts in the training set.

Table 6. Percentage of true coverage, false coverage, and total coverage of alerts
in the training set.

R34/R35 R38

‘True’ coverage 23.38% 15.79%

‘False’ coverage 1.30% 7.37%
Total coverage 24.68% 23.16%

4.4 Goodness-of-fit and predictivity

The goodness-of-fit and predictivity were assessed in terms of Cooper statistics [25].
Since BfR rules are only intended to identify active (corrosive/irritating) compounds,
and since there was a large prevalence of non-labelled substances which could influence
the goodness-of-fit statistics, non-labelled compounds were disregarded. The definitions
and values of goodness-of-fit statistics are detailed in table 7.
Given the large difference between the prior probability of a class and the prior
proportional probability of classes, it makes sense to eliminate the non-labelled
compounds from the statistics to assess sensitivity and classification rate. Indeed, the
no-model condition is the situation according to which all the objects are assigned
to the class most represented among the ones compared. Goodness-of-fit values close
to the ones of the no-model condition give evidence of a poor result of the classification
method. Thus, the exclusion of non-active compounds is also intended to avoid a
no-model scenario much better than the global goodness-of-fit (32 correctly classified
compounds out of 1341 compounds yields a sensitivity of 2.39%).
The true classification rate (sensitivity) is the ability of the DSS to detect known active
compounds for each class. The false classification rate is generally calculated as the
inverse of the sensitivity. In contrast, the positive predictive value focuses more on
the effect of individual chemicals, since it acts as a conditional probability, and it is the
probability that a chemical classified as active is really active. Note that Cooper
statistics values do vary according to the prevalence or proportion of active chemicals
in a population.
 Downloaded by [University of Saskatchewan Library] at 20:12 14 October 2012

340

Table 7. Definitions and values of the goodness-of-fit parameters (performance).

Statistic Formula Value Definition

Prior probability of a class
Prior proportional prob-
ability of a class
No-Model Error Rate,
NOMER%
True Classification Rate for
a class (Sensitivity for a
class)*
False Classification rate for
a class*
Positive Predictive Value
for a class*
P ¼ 1/G 33.33% (C, I, non-labelled)
Pg ¼ ng/n PR34/R35 ¼ 5.74% PR38 ¼ 7.08%
Pnon-labelled ¼ 87.17%
NOMER% ¼ (n
nM)/n  100 12.83%
Sg ¼ cgg0 /ng  100 SR34/R35 ¼ 23.38% SR38 ¼ 15.79%
1
Sg SR34/R35 ¼ 76.62% SR38 ¼ 84.21%
cgg0 /ng0  100 SR34/R35 ¼ 94.74% SR38 ¼ 68.18%
Probability that an object belongs to a class
supposing that every class has the same
probability.
Probability that an object belongs to a class taking
into account the number of objects in the class
Error provided in absence of model.
Number of objects correctly assigned to a class as a
percentage of the total number of objects
belonging to that class.
Percentage of objects that are falsely assigned to a
class.
Number of objects correctly assigned to a class as a
percentage of the total number of objects assigned
A. Gallegos Saliner et al.

to that class.

cgg0 ¼ Number of objects correctly classified in class g.

ng ¼ Total number of objects belonging to class g.
ng0 ¼ Total number of objects assigned to class g.
n ¼ Total number of objects.
nM ¼ Number of objects of the most represented class.
G ¼ Number of classes.
*Non-labelled compounds have been eliminated for the calculation of statistics.
 Skin irritation/corrosion SAR 341

As a consequence, the difference between the sensitivity and the positive predicted
value is very large. This difference can be explained by the fact that the coverage of the
alerts was very low: the implicit results would be 59 false negatives for skin corrosion
and 80 false negatives for skin irritation.

5. Conclusions

The application of the structural alerts for skin irritation and skin corrosion to the
training set of the BfR rulebase resulted in a low number of hits, indicating that the set
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of structural alerts has a limited coverage of the chemical space. The coverage could be
improved by slightly relaxing the pre-defined restrictions imposed on the rules. In the
relaxed search for skin irritant compounds, the false coverage increases, while the ‘true’
coverage does not significantly improve. The relaxed search for skin corrosive
compounds detects a significant number of corrosives missed in the first run.
However, the search for structural alerts irrespective of the constraints (third run)
did not allow the scope of the structural alerts to be extended.
The low coverage of the BfR structural rulebase for skin irritation and corrosion has
to be considered in the context in which the rulebase is intended to be applied. The rules
were designed to identify true positives without too many overpredictions. As a
consequence, if the rules predict a substance to be irritant or corrosive, then there is
a very high probability that the predicted classification is correct. This is reflected in the
high positive predictivity values. Furthermore, the structural inclusion rules should be
applied in a tiered strategy, following the application of physicochemical exclusion
rules. A previous analysis has shown that nearly a quarter of the labelled chemicals
in the same training set are covered by more than one exclusion rule. Thus, the coverage
of the structural rulebase, when corrected for the non-classified chemicals correctly
identified by the physicochemical rulebase, would be greater.
Since this study represents a partial assessment of the BfR rulebase (only the
structural rulebase was examined), further work should assess the combined application
of both the physicochemical and structural rulebases. Furthermore, it is recommended
that research is performed to identify new alerts, with a view to extending the coverage
of the DSS.

Acknowledgements

The authors thank Dr Matthias Herzler (BfR; Federal Institute for Risk Assessment,
Berlin, Germany) for his contributions to this study.

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