Discovery of piperonal-converting oxidase involved in the metabolism of a botanical aromatic aldehyde.

Piperonal-catabolizing microorganisms were isolated from soil, the one (strain CT39-3) exhibiting the highest activity being identified as Burkholderia sp. The piperonal-converting enzyme involved in the initial step of piperonal metabolism was purified from strain CT39-3. Gene cloning of the enzyme and a homology search revealed that the enzyme belongs to the xanthine oxidase family, which comprises molybdoenzymes containing a molybdopterin cytosine dinucleotide cofactor. We found that the piperonal-converting enzyme acts on piperonal in the presence of O2, leading to formation of piperonylic acid and H2O2. The growth of strain CT39-3 was inhibited by higher concentrations of piperonal in the culture medium. Together with this finding, the broad substrate specificity of this enzyme for various aldehydes suggests that it would play an important role in the defense mechanism against antimicrobial compounds derived from plant species.

Optimal evaluation time point for patch testing to predict skin irritation of commercial topical drugs.

To develop patch testing (PT) that better reflects the skin irritation resulting from repeated applications, we methodically classified PT results and examined correlations between these results with results from repeated application tests. A total of 115 commercial topical drugs were used for patch testing. In two groups of 30 healthy individuals with normal back skin, 55 or 60 commercial topical drugs were applied to the back for 24 h. Skin reactions were scored for each group at 2 and 24 h after patch removal. Based on time-dependent changes in skin reaction at each evaluation time, PT results were classified into six different types. For repeated application testing, a total of 17 products with different types of PT results were selected. A total of 104 healthy individuals divided into three groups (n = 22-52 each) received application of 6-7 test materials, differing for each group, for 3 weeks on both arms. Incidences of positive symptoms induced by repeated drug application excluding transient erythema showed a greater correlation with reaction levels at 24 h after patch removal than 2 h after patch removal. Many test materials that triggered a high incidence of positive symptoms after repeated application also exhibited erythema or greater reaction up to 24 h after patch removal. PT offers a useful test system for predicting skin irritation that develops after repeated application of topical drug. Moreover, assessment at 24 h after patch removal is suggested as the optimal evaluation time point to reflect the results of repeated application tests.