ARTICLE
Paraphenylenediamine
(PPD) is an important primary intermediate with
a long history of extensive use in oxidation hair colouring products. The
European Cosmetic Directive 76/768 allows its use in oxidation hair colouring
products at a maximal concentration of 6%. In practice, the initial concentration
of PPD in the hair colourants prior to their mixing with a developer rarely
exceeds 4% [1].
PPD is a known allergen and occlusive patch testing has been used for
many years in dermatological practice to establish a diagnosis of PPD
contact allergy. In order to avoid the manifestation of an allergic reaction
by the hair-dyeing procedure, a "skin sensitivity test" or "dab test"
(open test), either self-applied or applied by the hairdresser, is recommended
before hair colouring [2]. This open test consists in the application
of the hair colourant formulation (i.e. colourant alone, without
the developer) to the retroauricular area 48 hours without washing prior
to hair dyeing.
Hair colouring should not be carried out when itching or reddening of
the skin is observed during the course of the open test.
Recently the validity of the open test or "skin sensitivity test" was
questioned by the Cosmetic, Toiletry and Perfumery Association (UK) on
the grounds that the testing procedure had not been standardised and that
the proportion of false negative reactions had not been established [3].
The aim of the present study was to confirm, under standardised conditions
and medical supervision, the effectiveness of the "skin sensitivity test"
(open testing) as a practical method of detection of type IV allergy to
PPD-containing hair dyes and, consequently, as an important factor in
the secondary prevention [4] of contact dermatitis.
Materials and methods
Three dermatological departments in three different countries (Italy,
Great Britain and Poland) participated in the study. The study protocol
was approved by the respective Hospital Ethics Committees. Informed consent
was obtained from all subjects.
Experimental group
Thirty contact dermatitis patients (2 males and 28 females, 18-74 year-old,
mean age 40 years) with positive patch tests to PPD during routine investigation
and with established clinical relevance of the positive patch test, were
included in the study. Clinical relevance was determined by self-declared
past exposure to hair colouring products and clinical manifestations compatible
with allergy to hair dyes. Sensitisation to PPD was occupationally-related
in 5 of the subjects (hairdressers). Allergy to other para-substituted
derivatives (not present in the tested hair dye formulation) was established
in a number of the patients: p-toluenediamine (n = 12), disperse orange
3 (n = 3) and o-nitro-p-phenylenediamine (n = 2).
Control group
Thirty sex- and age-matched PPD-negative subjects with no history of
adverse reactions to hair colouring products were included in the control
group.
Test material
The test material was a marketed hair colourant representative of the
current L'Oréal hair colouring technology, containing 1.8% of PPD
(black shade). The hair colourant contained also 6 other hair dye molecules,
at a total concentration inferior to 2% (2,4-Diaminophenoxyethanol HCL,
resorcinol, m-Aminophenol, o-Aminophenol, Hydroxybenzomorpholine and p-Aminophenol).
The hair dye matrix was composed of water, surfactants (< 30%), conditioning
agents (< 5%), alkalizing agents (< 3%), antioxidants and stabilizers
(< 2%) and a perfume (0.5%).
The hair colouring product was applied without mixing with a developer.
Experimental protocol
The retro-auricular areas were wiped with alcohol on Day 0. Circular
adhesive devices (3M) were applied, one behind each ear. One contained
the experimental product and the other, as a negative control, was left
empty. 0.1 ml of experimental test material was applied in the centre
of the adhesive device (open application) by means of a micropipette (Multipette
4780, Eppendorf) and combitips (Eppendorf). The hair colouring product
was spread manually over the surface delineated by the adhesive device
(1.75 cm2), the finger being protected by a vinyl glove. The
effective volume of experimental material deposited was approximately
0.085 ml.
Adhesive devices were removed 1 hour later and test sites were marked
with a Chemotechnique Diagnostics skin marker. The hair colourant was
left for 48 hours without washing. The reactions were recorded on Day
0 (1 hour post-application), Day 2 and Day 4.
In the absence of a universally accepted method for grading of open
tests, the scoring method used in the study was based on that proposed
by Johansen et al. [5].
The method comprised: i) the dermatologist's clinical evaluation
of the severity of the reaction, "overall clinical impression". The overall
clinical impression was recorded using a five-point grading scale: negative,
doubtful, weakly positive, moderately positive and strongly positive ;
ii) an objective evaluation using a set of objective parameters,
graded individually (erythema, infiltration, vesicles). Sensory manifestations
reported by the patients were also taken into account. The maximal score
for each reading was 19 (Table
I).
Statistical analysis
Mantel-Haenszel's test for ordinal variables and nonparametric Mann-Whitney's
test were used to compare the severity of the reactions evaluated as "overall
clinical impression" and as numerical scores respectively, between PPD-positive
and PPD-negative subjects at all reading times. To enable statistical
calculations, values were attributed to the dermatologist's "overall clinical
impression" as follows: negative = 0; doubtful = 1; weakly positive =
2; moderately positive = 3 and strongly positive = 4.
Kendall's tau-b was used to evaluate the correlation between the severity
of the reactions evaluated as "overall clinical impression" and the corresponding
objective numerical scores. Probability values of p < 5% were considered
to be significant.
Results
Day 0 (1 hour)
16/30 PPD-positive subjects and 13/30 PPD-negative subjects reacted
on Day 0 (1 hour post-application) with mild irritant reactions, which
were essentially low grade erythema and/or sensory manifestations (Fig.
1). Two patients in the experimental group and three patients in the
control group, who had erythematous reactions at the site of the experimental
test material, reacted with erythema of similar intensity to the adhesive
device applied on the contralateral side as a negative control. There
was no statistically significant difference between PPD-positive and PPD-negative
subjects either in the severity of the reactions assessed as "overall
clinical impression", nor as objective numerical scores (p > 5%).
Day 2
All 30 PPD-positive subjects reacted on Day 2 with erythema and infiltration
(individual papules or homogeneous infiltration). In 25/30 subjects vesicles
could also be observed. Sensory manifestations were recorded in most of
the subjects (27/30). Maximal intensity of the reactions was recorded
on Day 2 in all but two subjects. The reactions in 4/30 subjects were
evaluated with the maximal score (19/19). The reactions in all PPD-positive
subjects were evaluated as allergic in nature. Increased severity of reactions,
from weakly to strongly positive was correlated strongly with increasing
numerical scores (Table II).
Only 3 subjects in the control group had mild erythematous reactions
on Day 2 and in one subject a single isolated papule without erythema
was observed. All the reactions in the control subjects were evaluated
as negative, except in one subject, in whom the reaction (erythema) was
evaluated as doubtful. Severity of the reactions was statistically higher
in the PPD-positive subjects whether assessed as "overall clinical impression"
or as objective numerical scores (p < 10/00)
(Fig. 1).
Kendall's tau-b, used to evaluate the correlation between increasing
severity of the reactions assessed by "overall clinical impression" and
increasing numerical scores in all subjects (PPD-positive and PPD-negative),
was 0.88 (high degree of dependence), significant at 1% (Fig.
2).
Day 4
The severity of the reactions was reduced in most PPD-positive subjects
after Day 2, but on Day 4 there was still a clear-cut difference in the
nature and severity of the reactions between PPD-positive and PPD-negative
subjects (Fig. 1).
Discussion
The effectiveness of the open hair dye skin sensitivity test has been
investigated using a commercial hair colouring product containing 1.8%
PPD. Test subjects were 30 dermatological patients with contact dermatitis
to hair dyes; all were sensitised to PPD, by far the most frequent allergen
in hair colouring products [6-8].
Open testing is one of the oldest methods for clinical investigation
of allergy. Sabouraud was the first to use open testing in the retroauricular
area [2]. One reason for his choice of this particular test site was that
unsightly positive reactions will be hidden behind the ears. Further scientific
and/or practical arguments for the hair dye sensitivity test being performed
in that location are the follow-ing: i) a reaction to a contact
allergen is more easily elicited at a site of pre-existing dermatitis
to that particular allergen (hapten-specific local immunological memory)
[9-11]; ii) the site is protected from sunlight; iii) it
is easy to reach by the consumer and by the hairdresser.
Nowadays open testing as a diagnostic method in allergy investigation
is usually performed on the antecubital folds, upper back, flexor aspects
of the forearms or upper outer arms. This nonocclusive technique minimises
the risk of severe necrotic or false positive irritant reactions. The
Repeated Open Application Test (ROAT) [12] is a variant of the open application
test. It simulates the everyday use of dermatological and cosmetic products
and is used to define the clinical relevance of doubtful or positive diagnostic
patch tests. A single patch test procedure is more appropriate for testing
hair dyes, which are usually only applied once a month.
There is no generally accepted grading scale for open tests although
the ICDRG grading scale [13] may be used as a guideline. An eczematous
appearance with at least redness and infiltration (oedema) and/or scaling
is required by some authors for grading the reaction as positive, while
any degree of reaction, including redness is sufficient for others [5,
14]. Johansen et al. proposed a detailed grading scale and studied
the correlation between the numerical scores and the clinical evaluation
of the severity of the reactions in a ROAT to deodorants in 14 eczema
patients [5]. This study demonstrated a difference between the morphological
assessment of open test and patch test reactions. Thus, contrary to the
patch test evaluation by the ICDRG grading scale, homogeneous infiltration
was not a prerequisite for grading an open reaction as moderately or strongly
positive; likewise vesicles, which are a decisive feature of moderate
patch test reactions (++), were present only in a minority of ROAT reactions
graded as strongly positive [5]. In other words, less severe reactions
were graded positive in the open test than in the occluded patch test.
In our study skin reactions with morphological features strongly suggestive
of allergy were elicited in all PPD-positive subjects, with a maximal
intensity on Day 2. The morphology of the reactions on Day 2 was in most
cases that of a moderate or strongly positive reaction (++ or +++) even
by the ICDRG grading scale (vesicles present in 25/30 subjects), despite
the lack of occlusion or of repeated applications. There were no false
negative reactions either as far as clinical observation or the subjects'
perception was concerned; palpable infiltration accompanied by exudation
and/or pruritus could in no way pass unnoticed by the subjects. Conversely,
no reactions suggestive of allergy were elicited in PPD-negative subjects;
in only one case was there a doubtful, probably irritant reaction (erythema
only) on Day 2.
Previous open test studies have shown it is possible to elicit positive
reactions to PPD-containing hair dye formulations [15-17]. The methods
and results are unfortunately poorly described. To our knowledge the present
multicenter study is the first report on open testing of a hair dye formulation
carried out under standardised conditions and using a numerical score
in addition to the traditional clinical evaluation.
Good statistical correlation between the "overall clinical impression"
(qualitative evaluation) and the quantitative numerical score suggests
that the numerical method of scoring the results of an open test constitutes
an additional useful tool for quantifying and recording this type of reaction.
Abbreviations
ICDRG International Contact Dermatitis Research
Group
PPD Paraphenylenediamine
ROAT Repeated open application test
Acknowledgement
We thank Anne-Lise Garcel and Frank Rimek from L'Oréal
Recherche, Clichy, France for the statistical evaluation of data.
CONCLUSION
Our results indicate that the open test ("skin sensitivity test") as
described above, can detect individuals who would react to a PPD-containing
hair dye, thus preventing cases of contact dermatitis that might otherwise
involve the scalp and possibly the face. The "skin sensitivity test" is
a practical and easy to perform test that may be undertaken by consumers
and hairdressers; it can be considered as a very effective method for
detecting type IV hair dye allergy and as such, as an important factor
in the secondary prevention of allergic reactions.
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