PERFUME PHOTO [O\ ICI'I'• 699 F•AC •3A / COeW•ZN Figure 2. Phototoxic responses in one subject tested with 8 different materials Figure 8. Hyperpigmentation in a subject 23 days after positive response to bcrgamol

700 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS Table I Phototoxicity of Oil of Bcrgamot, Coumarin-psoralcn (C-P) Nonvolatile Fraction, TLC- Separated C-P Components, and Ethyl Ether-Precipitated Portion of C-P • Substance Components Fraction Concen- Showing tration Positive Skin (%) Response Oil of bergamot C-P fraction C-P, TLC band 1 C-P, TLC band 2 C-P, TLC band 3 C-P, TLC band 4 C-P (ethyl ether-pptd. portion recrystallized 3X with methanol), fraction 3A About 2% C-P Bergapten, limettin, bergamottin, bergaptol, methoxygeranoxycoumarin Mainly bergamottin Mainly 7-methoxy-5-geranoxycoumarin Limettin-bergapten Band 3 to origin (includes bergaptol) Mainly bergapten (ca. 90 %) 100 9/9 1 9/9 0.5 1/9 O. 5 3,/9 0.5 9/9 0.5 2/9 0.5 9/9 • Test materials (diluted with 95% alcohol) were applied to a 5 X 12-cm area of "stripped" forearm skin of nine subjects. The areas were irradiated for 40 minutes with UV light (prin- cipally 360-370 nm) emitted from an Inspectolite (Hanovia) at a distance of 8-10 cm. to the limettin-bergapten mixture obtained from the TLC band g. The results (Table II) indicate that pure bergapten is phototoxic down to 0.002% concentration pure limettin on the other hand, though strongly fluorescent under UV, is not phototoxic. Commercial Samples Fifteen commercial perfumes were purchased in the San Francisco area in 1968 for human photopatch tests by the methods described earlier. An old (1964) sample of "Shalimar" Perfume (el) was in- cluded as well as a new sample (½t2). Results show that "Shalimar" "Replique," and "Park 8c Tilford No. 3" produced phototoxicity in some test subjects (Table III). On chemical analysis, our sample of "Shall- mar" el was found to contain 0.1% bergapten. This is equivalent to 30% bergamot, using Cieri's recent analytical data, i.e., 0.33,% bergapten in bergamot (12). "Shalimar" •t2, on the other hand, contained no ber- gapten and produced no phototoxicity in four subjects tested. Samples o15 "Replique," subsequently procured in the Washington area in 1969 and tested on hairless mice, as described later, showed no phototoxic ef- fects. Furthermore, "Park and Tilford No. 3" is no longer manufac- tured.