[Pattern of contact sensitization to paraphenylenediamine and its detection in hair dyes]. / Patrón de sensibilización por contacto a parafenilendiamina y su detección en tintes capilares.

BACKGROUND: One of the greatest challenges in occupational dermatology is the identification of chemical substances used by patients in their work in order to determine their allergenic potential. Numerous techniques have been described for the identification of allergenic compounds. These tests must be sensitive, specific, and safe. We describe a study to detect the presence of paraphenylenediamine (PPD) in hair dyes that are commercially available in Spain. MATERIAL AND METHODS: We undertook an experimental study involving qualitative and semiquantitative detection of PPD in hair dyes sold in Spain. The qualitative technique we used was a previously described colorimetric method involving dilution of the dye with isopropyl alcohol followed by addition of a reagent solution (1g of vanilla in 15 ml of isopropyl alcohol and 7.5 ml of hydrochloric acid). A quantitative study was then done in which the dye was extracted in 96% ethanol and subjected to 1-dimensional thin-layer chromatography. RESULTS: A total of 15 brown and 12 blonde dyes were analyzed. PPD was identified in all of the brown dyes analyzed, irrespective of whether it was indicated (n = 12) or not (n = 3) in the composition. PPD was found in 6 of the 9 blonde dyes that indicated it in the composition and 2 of the 3 in which it was not indicated. Semiquantitative analysis by thin-layer chromatography revealed that the concentration of PPD in brown hair dyes (mean, 3%) was higher than in blonde dyes (mean, 0.1-0.3%). CONCLUSIONS: The presence of PPD in hair dyes is related to the color of the dye. It is consistently present in darker dyes and at low levels in blonde dyes. This study highlights the clinical and epidemiological importance of identifying allergens in dermatology, particularly in occupational dermatology.

Simultaneous determination of dihydroxybenzene and phenylenediamine positional isomers using capillary zone electrophoresis coupled with amperometric detection.

In general capillary zone electrophoresis (CZE) separation models, o-, m-, and p-phenylenediamine isomers can be separated in a weak acidic running buffer for their pK(a) values being 4.52, 5.64, 6.04, respectively, while o-, m-, and p-dihydroxybenzene isomers can be separated in a weak basic buffer for their pK(a) values being 9.40, 9.40 and 10.04, respectively. So, it is hard to find a suitable running buffer at a fixed pH in normal CZE for simultaneous separation of these two groups of positional isomers. In this paper, a novel method based on alternately running two different pH buffers in CZE coupled with amperometric detection (CZE-AD) was designed to simultaneously determine these two groups of positional isomers. It is found that when two different pH running buffers were employed alternately under appropriate order and time, the six analytes could be separated perfectly in less than 20 min and the detection limits were as low as 10(-7) mol/L. Furthermore, the effects of working electrode potential, pH and concentration of running buffer, separation voltage and injection time on CZE-AD were investigated. Experimental results demonstrated that the introduced method was practical to simultaneously determine two groups of positional isomers with different pK(a) and had some advantages of high sensitivity, good repeatability and small sample requirement.

Microbial transformation of aniline derivatives: regioselective biotransformation and detoxification of 2-phenylenediamine by Bacillus cereus strain PDa-1.

A bacterial isolate, strain PDa-1, grew well on basal medium supplemented with 2-phenylenediamine, sucrose, and ammonium nitrate and completely transformed 2-phenylenediamine. The isolate was identified as Bacillus cereus. The product formed from 2-phenylenediamine was identified by EI-MS and NMR as 2-aminoacetanilide; whole cells converted 2-phenylenediamine to the product with a 76% molar yield. Whole cells also showed a broad substrate specificity toward 20 of 26 tested arylamines with substituent groups of various size and positions. Especially 2-aminobenzoic acid, 4-aminosalicylic acid, 5-aminosalicylic acid, and 2-aminofluorene were converted completely to the corresponding product with an aminoacetyl group. Cell extracts of strain PDa-1 had a high arylamine N-acetyltransferase activity. The partially purified enzyme converted 2-phenylenediamine to 2-aminoacetanilide. Strain PDa-1 constitutively expressed the enzyme in the absence of 2-phenylenediamine. Effects of 2-phenylenediamine and 2-aminoacetanilide on growth indicated that this enzyme probably plays a role in the detoxification of toxic arylamines in this strain.

Fabrication and performance of a three-dimensionally adjustable device for the amperometric detection of microchip capillary electrophoresis.

A microchip CE-amperometric detection (AD) system has been fabricated by integrating a two-dimensionally adjustable CE microchip and an AD cell containing a one-dimensionally adjustable disk detection electrode in a Plexiglas holder. It facilitates the precise 3-D alignment between the channel outlet and the detection electrode without a complicated 3-D manipulator. The performance of this unique system was demonstrated by separating five aromatic amines (1,4-phenyldiamine, aniline, 2-methylaniline, 4-chloroaniline, and 1-naphthylamine) of environmental concern. Factors influencing their separation and detection processes were examined and optimized. The five analytes have been well separated within 140 s in a 74 cm long separation channel at a separation voltage of +2500 V using a 10 mM phosphate buffer (pH 3.5). Highly linear response is obtained for the five analytes over the range 20-200 microM with the detection limits ranging from 0.46 to 1.44 microM, respectively. The present system demonstrated long-term stability and reproducibility with RSDs of less than 5% for the peak current (n = 9). The new approach for the microchannel-electrode alignment should find a wide range of applications in CE, flowing injection analysis, and other microfluidic analysis systems.

[Fast optimization of stepwise gradient conditions for ternary mobile phase in reversed-phase high performance liquid chromatography].

In high performance liquid chromatography, it is necessary to apply multi-composition gradient elution for the separation of complex samples such as environmental and biological samples. Multivariate stepwise gradient elution is one of the most efficient elution modes, because it combines the high selectivity of multi-composition mobile phase and shorter analysis time of gradient elution. In practical separations, the separation selectivity of samples can be effectively adjusted by using ternary mobile phase. For the optimization of these parameters, the retention equation of samples must be obtained at first. Traditionally, several isocratic experiments are used to get the retention equation of solute. However, it is time consuming especially for the separation of complex samples with a wide range of polarity. A new method for the fast optimization of ternary stepwise gradient elution was proposed based on the migration rule of solute in column. First, the coefficients of retention equation of solute are obtained by running several linear gradient experiments, then the optimal separation conditions are searched according to the hierarchical chromatography response function which acts as the optimization criterion. For each kind of organic modifier, two initial linear gradient experiments are used to obtain the primary coefficients of retention equation of each solute. For ternary mobile phase, only four linear gradient runs are needed to get the coefficients of retention equation. Then the retention times of solutes under arbitrary mobile phase composition can be predicted. The initial optimal mobile phase composition is obtained by resolution mapping for all of the solutes. A hierarchical chromatography response function is used to evaluate the separation efficiencies and search the optimal elution conditions. In subsequent optimization, the migrating distance of solute in the column is considered to decide the mobile phase composition and sustaining time of the latter steps until all the solutes are eluted out. Thus the first stepwise gradient elution conditions are predicted. If the resolution of samples under the predicted optimal separation conditions is satisfactory, the optimization procedure is stopped; otherwise, the coefficients of retention equation are adjusted according to the experimental results under the previously predicted elution conditions. Then the new stepwise gradient elution conditions are predicted repeatedly until satisfactory resolution is obtained. Normally, the satisfactory separation conditions can be found only after six experiments by using the proposed method. In comparison with the traditional optimization method, the time needed to finish the optimization procedure can be greatly reduced. The method has been validated by its application to the separation of several samples such as amino acid derivatives, aromatic amines, in which satisfactory separations were obtained with predicted resolution.

[Study on qualitative analysis method of high performance liquid chromatography by simultaneous multi-wavelength detection].

In this paper, we introduce an HPLC qualitative method, that is, multiwavelength detection qualitative analysis method. According to Beer's law, for a certain compounds the ratio of peak areas at different wavelengths is a constant. Comparing the ratio of peak areas of a component at different wavelengths with that of the standard by means of simultaneous multi-wavelength detection, we established a convenient qualitative method. This method has been applied successfully to qualitative analysis of 18 aromatic amines in the prohibited azo dyes.

Contact allergy to colour developing agents in the guinea pig.

Colour developing agents, derivatives of p-phenylenediamine, can cause contact allergy. Patch test reactions to more than one colour developer are sometimes seen in patients. To study whether this is due to simultaneous sensitization or cross-reactivity, guinea pig maximization tests (GPMT) with CD-2, CD-3 and CD-4 were carried out. 5 experiments were performed, using pet. or water as vehicles. When pet. was used, the challenge concentrations could be raised and cross-reactivity between the colour developers, but not with p-phenylenediamine-dihydrochloride, was revealed. When water was used as vehicle, the challenge concentrations were limited because of staining of the test sites and irritation. CD-2, CD-3 and CD-4 were found to be extreme sensitizers according to the classification by Magnusson and Kligman. The importance of using an appropriate vehicle to obtain optimal conditions for the GPMT is stressed. To study the purity and stability of the chemicals used, analysis by HPLC of the test substances at different stages of the GPMT procedure was performed. Aqueous solutions of the colour developers were found to be unstable, while pet. mixtures were stable.

Identification of a toxic impurity in commercial diphenylamine.

The impurity in commercial diphenylamine which induces polycystic kidney disease in rats has been identified as N,N,N'-triphenyl-p-phenylenediamine. The structure of the compound was confirmed by mass and proton magnetic resonance spectrometry and by comparison with a synthetic standard prepared by the Ullman coupling of iodobenzene and p-phenylenediamine or N,N-diphenyl-p-phenylenediamine. Laboratory studies with highly purified diphenylamine indicated that N,N,N'-triphenyl-p-phenylenediamine can be formed by heating diphenylamine.

Purpuric allergic contact dermatitis to paraphenylenediamine.

A 59-year-old saleswoman of black hats presented with a severe purpuric eruption of the exposed areas of the face, neck and arms. Patch testing to paraphenylenediamine produced a purpuric test reaction. Similar eruptions have been reported due to N-isopropyl-N-phenylparaphenylenediamine (IPPD), a rubber antioxidant. The patient had previous dermatitis underneath the elastic portions of her undergarments. It was determined that IPPD is added to elastic material used in some elastic trim on undergarments in the United States. This relationship is discussed.