S. HIROSHIMENSIS STRAIN WITH ANTI-TYROSINASE ACTIVITY 39 analysis described above, we also identified the strain by using physiological character­ izations. Table I lists the comparison of the physiological characteristics between TI-C3 and BCRC 12423. All the criteria except esculin hydrolysis and melanin production gave identical results. Based on these results, the strain TI-C3 was identified as a strain of S. hiroshimensis. FERMENTATION OF S. HIROSHIMENSIS STRAIN TI-C3 S. hiroshimensis strain TI-C3 was cultivated using different carbon and nitrogen sources and analyzed for anti-tyrosinase activity. As shown in Figure 3, the strain TI-C3 ex­ hibited maximal anti-tyrosinsae activity (5 72 U/ml) when glucose was used as the 700 i 600 - e, 500 � ] 400 300 � ;§ 200 :a 100 = j,,,,,I a b 0 1000 900 800 700 600 500 400 300 200 100 0 Glycerol Glucose Com steep Malt extract liquor Maltose Sodium propionate Carbon Source ( 4 g/1) Soya Tryptone peptone Nitrogen Source (14 g/1) Yeast extract Sorbitol YMG control Figure 3. Inhibitory activity of the strain TI-C3 cultivated using different carbon (a) and nitrogen (b) sources. The fermentations and assays of anti-tyrosinase activity were done as described in Materials and Methods.

40 JOURNAL OF COSMETIC SCIENCE carbon source (Figure 3a). On the other hand, TI-C3 grew poorly and showed little anti-tyrosinase activity when cultivated using ammonia sulfate, casein, sodium nitrate, or urea as the nitrogen source (not shown). In contrast, TI-C3 displayed potent and mildly different anti-tyrosinase activity when cultivated using more complex and di­ gested nitrogen sources (Figure 36). TI-C3 showed 905 U/ml of anti-tyrosinase activity when malt extract was used as the nitrogen source. CONCLUSION The isolated strain TI-C3 displayed 905 U/ml of the anti-tyrosinase act1v1ty under cultivation using glucose and malt extract as the sole carbon and nitrogen sources, respectively. The strain was identified as a strain of S. hiroshimensis by morphological, physiological, and chemical analysis, gyr B sequences, and DNA-DNA hybridization analysis. The purification of the active compound from the strain TI-C3 is under research in our laboratory. REFERENCES (1) S. Y. Seo, V. K. Sharma, and N. Sharma, Mushroom tyrosinase: Recent prospects,]. Agr. Food Chem., 51, 2837-2853 (2003). (2) S. Briganti, E. Camera, and M. Picardo, Chemical and instrumental approaches to treat hyperpig­ mentation, Pig. Cell Res., 16, 101-110 (2003). (3) N. Baurin, E. Arnoult, T. Scior, Q.T. Do, and P. Bernard, Preliminary screening of some tropical plants for anti-tyrosinase activity,]. Ethnopharmacol., 82, 155-158 (2002). (4) Q. X. Chen and I. Kubo, Kinetics of mushroom tyrosinase inhibition by quercetin,j. Agr. Food Chem., 50, 4108-4112 (2002). (5) Y. M. Kim, J. Yun, C. K. Lee, H. Lee, K. R. Min, and Y. Kim, Oxyresveratrol and hydroxystilbene compounds: Inhibitory effect on tyrosinase and mechanism of action, ]. Biol. Chem., 277, 16340- 16344 (2002). (6) M. Shiino, Y. Watanabe, and K. Umezawa, Synthesis and tyrosinase inhibitory activity of novel N-hydroxybenzyl-N-nitrosohydroxylamines, Bioorg. Chem., 31, 129-135 (2003). (7) X. L. Piao, S. H. Baek, M. K. Park, and J. H. Park, Tyrosinase-inhibitor furanocoumarin from Angelica dahurica, Biol. Pharm. Bull., 27, 1144-1146 (2004). (8) T. S. Chang, C. F. Chiou, S. J. Chu, and M. Wu, Development of an efficiency high throughout screening method by using microplates, Fifty-First Chinese Chemical Engineering Conferences, 1096-1099 (2004). (9) E. B. Shirling and D. Gottlieb, Methods for characterization of Strepotmyces species, Int.]. Sys. Bacterial., 16, 313-340 (1966). (10) D. Berd, Laboratory identification of clinically important aerobic actinomycetes,J. Appl. Microbial., 24, 665-681 (1973). (11) T. Hasegawa, M. Takizawa, and S. Tanida, A rapid analysis for chemical grouping of aerobic actino­ mycetes,J. Gen. Appl. Microbial., 29, 319-322 (1983). (12) K. Hatano, T. Nishii, and H. Kasai, Taxonomic re-evaluation of whorl-forming Streptomyces (formerly Streptoverticillium) species by using phenotypes, DNA-DNA hybridization and sequences of gyrB, and proposal of Streptomyces luteireticuli (ex Katoh and Arai, 1957) corrig., sp. nov., nom. rev., Int.]. Sys. Evol. Microbial., 53, 1519-1529 (2003). (13) T. Ezaki, S. M. Saidi, S. L. Liu, Y. Hashimoto, H. Yamamoto, and E. Yabuuchi, Rapid procedure to determine the DNA base composition from small amounts of gram-positive bacteria, FEMS Microbial. Lett., 55, 127-130 (1990). (14) T. S. Chang and M. Tseng, Preliminary screening of soil actinomycetes for anti-tyrosinase activity,]. Mar. Sci. Technol., 14, 190-193 (2006). (15) M. P. Lechevalier and H. Lechevalier, Chemical composition as a classification of aerobic actinomy­ cetes, Int.]. Sys. Bacterial., 20, 435-443 (1970).