JOURNAL OF COSMETIC SCIENCE 136 CONCLUSION We have developed an HPLC–fl uorescence detection analysis for determination of OPP in paraben-free skin lotions by using NBD-COCl as a pre-column fl uorescence labeling reagent. This method is rapid, simple, convenient, inexpensive, and should be suitable for routine quality assessment of OPP in cosmetics. Two tested skin lotions were found to contain OPP at mean concentrations of 1.07±0.06 mg/g and 18.8±1.3 μg/g using the present system. The OPP level in the other lotion was below the limit of detection. Previous reports have dealt with determination of OPP levels in various sample matrixes, including fruits, serum, and environmental water (6,7,9,10,19–21). Our present method should be suitable for routine assessment of OPP levels, not only in cosmetics but also in those samples. REFERENCES (1) M. Davoren and A. M. Fogarty, In vitro cytotoxicity assessment of the biocidal agents sodium o-phenylphenol, sodium o-benzyl-p-chlorophenol, and sodium p-tertiary amylphenol using established fi sh cell lines, Toxicol In Vitro., 20, 1190–1201 (2006). (2) N. Kolbe and J.T. Andersson, Simple and sensitive determination of o-phenylphenol in citrus fruits us- ing gas chromatography with atomic emission or mass spectrometric detection, J. Agric. Food Chem., 54, 5736–5741 (2006). (3) E. M. Bomhard, S. Y. Brendler-Schwaab, A. Freyberger, B. A. Herbold, K. H. Leser, and M. Richter, o-Phenylphenol and its sodium and potassium salts: A toxicological assessment, Crit. Rev. Toxicol., 32, 551–626 (2002). (4) M. Takahashi, H. Sato, K. Toyoda, F. Furukawa, K. Imaida, R. Hasegawa, and Y. Hayashi, Sodium o-phenylphenate (OPP-Na) promotes skin carcinogenesis in CD-1 female mice initiated with 7,12-dimethylbenz[a]anthracene, Carcinogenesis, 10, 1163–1167 (1989). (5) Ministry of Health, Labour and Welfare. http://www.mhlw.go.jp/fi le/06-Seisakujouhou-11120000- Iyakushokuhinkyoku/keshouhin-standard.pdf. (2000), pp. 1–11. (6) R. D. Thompson, Determination of phenolic disinfectant agents in commercial formulation by liquid chromatography, J. AOAC Int., 84, 815–822 (2001). (7) L. Yang, A. Kotani, H. Hakamata, and F. Kusu, Determination of ortho-phenylphenol residues in lemon rind by high-performance liquid chromatography with electrochemical detection using a microbore column, Anal. Sci., 20, 199–203 (2004). (8) World Health Organization. Guidelines for drinking-water quality incorporating fi rst addendum. Recommen- dations, 3rd Ed. (World Health Organization, 2006), Vol. 1, pp. 427–428. (9) Y. Higashi and Y. Fujii, Determination of three phenylphenols in grapefruit juice by HPLC after pre-column derivatization with 4-fl uoro-7-nitro-2,1,3-benzoxadiazole, J. Anal. Chem. (2014) In press. (10) C. Blasco, Y. Picó, J. Mañes, and G. Font, Determination of fungicide residues in fruits and vegetables by liquid chromatography-atmospheric pressure chemical ionization mass spectrometry, J. Chromatogr. A, 947, 227–235 (2002). Table IV Levels of OPP in skin lotions and relative recovery values Skin lotion Concentration Relative recovecy (mean ± S.D., n=5) (%,mean ± S.D., n=5) A 1.07 ± 0.06 mg/g 87.0 ± 3.6 B 18.8 ± 1.3 μg/g 88.7 ± 5.4 C N.D. 88.9 ± 3.5 N.D., not determined (below the lower limit of quantifi cation).

DETERMINATION OF O-PHENYLPHENOL IN SKIN LOTION 137 (11) K. Imai, Analytical chemical studies on high-performance recognition and detection of bio-molecules in life, Yakugaku Zasshi, 123, 901–917 (2003). (12) T. Fukushima, J. Kawai, K. Imai, and T. Toyo’oka, Simultaneous determination of D- and L-serine in rat brain microdialysis sample using a column-switching HPLC with fl uorimetric detection, Biomed. Chromatogr., 18, 813–819 (2004). (13) Y. Higashi, S. Nakamura, H. Matsumura, and Y. Fujii, Simultaneous liquid chromatographic assay of amantadine and its four related compounds in phosphate-buffered saline using 4-fl uoro-7-nitro-2,1,3- benzoxadiazole as a fl uorescent derivatization reagent, Biomed. Chromatogr., 20, 423–428 (2006). (14) Y. Higashi, M. Sakata, and Y. Fujii, Simultaneous determination of the N-dealkylated metabolites of four butyrophenone-type agents in rat plasma by HPLC with fl uorescence detection after precolumn derivatization with 4-fl uoro-7-nitro-2,1,3-benzoxadiazole, J. Liq. Chromatogr. Rel. Technol., 30, 2747– 2754 (2007). (15) Y. Higashi, R. Gao, and Y. Fujii, Determination of fl uoxetine and norfl uoxetine in human serum and urine by HPLC using a Cholester column with fl uorescence detection, J. Liq. Chromatogr. Rel. Technol., 32, 1141–1151 (2009). (16) T. Toyo’oka, T. Mantani, and M. Kato, Characterization of labelling and de-labelling reagents for detec- tion and recovery of tyrosine residue in peptide, Biomed. Chromatogr., 17, 133–142 (2003). (17) Y. Higashi and Y. Fujii, HPLC-UV analysis of phenol and chlorophenols in water after pre-column de- rivatization with 4-fl uoro-7-nitro-2,1,3-benzoxadiazole, J. Liq. Chromatogr. Rel. Technol., 32, 2372– 2383 (2009). (18) Y. Higashi and Y. Fujii, HPLC-UV analysis of eugenol in clove and cinnamon oils after pre-column derivatization with 4-fl uoro-7-nitro-2,1,3-benzoxadiazole, J. Liq. Chromatogr. Rel. Technol., 34, 18–25 (2011). (19) Y. Yamazaki and T. Ninomiya, Determination of benomyl, diphenyl, o-phenylphenol, thiabendazole, chlorpyrifos, methidathion, and methyl parathion in oranges by solid-phase extraction, liquid chroma- tography, and gas chromatography, J. AOAC Int., 82, 1474–1478 (1999). (20) L. Yu, R. Schoen, A. Dunkin, M. Firman, H. Cushman, and A. Fontanilla, Determination of o-phenylphenol, diphenylamine, and propargite pesticide residues in selected fruits and vegetables by gas chromatography/mass spectrometry, J. AOAC Int., 80, 651–656 (1997). (21) X. Ye, L. J. Tao, L. L. Needham, and A. M. Calafat, Automated on-line column-switching HPLC-MS/ MS method for measuring environmental phenols and parabens in serum, Talanta, 76, 865–871 (2008). (22) J. Q. Huang, C. C. Hu, and T. C. Chiu, Determination of seven preservatives in cosmetic products by micellar electrokinetic chromatography, Int J Cosmet Sci., 35, 346–353 (2013). (23) C. Charnock and T. Finsrud, Combining esters of para-hydroxy benzoic acid (parabens) to achieve in- creased antimicrobial activity, J. Clin. Pharm. Ther., 32, 567–572 (2007). (24) P. D. Darbre and P. W. Harvey, Paraben esters: Review of recent studies of endocrine toxicity, absorp- tion, esterase and human exposure, and discussion of potential human health risks, J Appl Toxicol., 28, 561–578 (2008).