Evaluation of PTeCA (1-H-pyrrole-2,3,4,5-tetracarboxylic acid) in in vivo and in vitro hair matrix as a marker of oxidative cosmetic treatment

Alteration of hair matrix by cosmetic products presents a challenge for forensic hair analysis. In particular, oxidative treatments can lead to drugs depletion and, thus, false-negative results. Currently, the degradation products of eumelanin, 1H-pyrrole-2,3,5-tricarboxylic acid (PTCA) and its isomer isoPTCA, are being investigated as markers for oxidative hair treatment; however, they require the definition of threshold values since their formation naturally occurs from eumelanin degradation. Recently, the 1H-pyrrole-2,3,4,5-tetracarboxylic acid (PTeCA) concentration was significantly increased also after in vitro oxidative hair treatments with low percentages of H2O2 (1.9%).[1] Our previously published high-performance liquid chromatography tandem mass spectrometry (LC-MS/MS) method for PTCA analysis (LOQ 0.01 ng/mg) in hair matrix has also been fully validated for the simultaneous quantification of PTeCA (linearity range 0.01-2.5 ng/mg; LOQ 0.003 ng/mg) in real hair samples. [2] Then, the method was applied to N=3378 self-reported, treated (T; N= 503) and untreated (UT; N=2875), hair samples (<6 cm proximal) collected from the Laboratory of Forensic Toxicology of the University of Milan. In addition, the in vitro formation of PTeCA was assessed in N=240 untreated hair samples by N=9 different professional cosmetic treatments with and without oxidative agents (H2O2). In the UT group, PTCA was determined in 40% of the samples (N=1144) with a median PTCA concentration of 0.04 ng/mg (range 0.01-14.9 ng/mg), among which <2% of the samples (N=53) presented PTeCA concentration >LOQ (median 0.30 ng/mg; range 0.01-4.8 ng/mg). In the T group, PTCA was determined in 84% of the samples (N=425) with a median concentration of 0.75 ng/mg (range 0.01-58.1 ng/mg); while the median PTeCA concentration was 0.40 ng/mg (N=243; range 0.02-31.2 ng/mg), according to median PTeCA concentration measured in the N=53 samples with PTeCA>LOQ belonging to the UT group, suggesting a false self-declaration of untreatment. Finally, the in vitro cosmetic treatment confirmed the PTeCA formation only in oxidative conditions. Our data suggest that PTeCA could be a reliable marker for detecting oxidative cosmetic treatments in the hair matrix.