Novel Chemically Modified Curcumin (CMC) Derivatives Inhibit Tyrosinase Activity and Melanin Synthesis in B16F10 Mouse Melanoma Cells

Abstract

Skin hyperpigmentation disorders arise due to excessive production of the macromolecular pigment melanin catalyzed by the enzyme tyrosinase. Recently, the therapeutic use of curcumin for inhibiting tyrosinase activity and production of melanin have been recognized, but poor stability and solubility have limited its use, which has inspired synthesis of curcumin analogs. Here, we investigated four novel chemically modified curcumin (CMC) derivatives (CMC2.14, CMC2.5, CMC2.23 and CMC2.24) and compared them to the parent compound curcumin (PC) for inhibition of in vitro tyrosinase activity using two substrates for monophenolase and diphenolase activities of the enzyme and for diminution of cellular melanogenesis. Enzyme kinetics were analyzed using Lineweaver-Burk and Dixon plots and nonlinear curve-fitting to determine the mechanism for tyrosinase inhibition. Copper chelating activity, using pyrocatechol violet dye indicator assay, and antioxidant activity, using a DPPH radical scavenging assay, were also conducted. Next, the capacity of these derivatives to inhibit tyrosinase-catalyzed melanogenesis was studied in B16F10 mouse melanoma cells and the mechanisms of inhibition were elucidated. Inhibition mechanisms were studied by measuring intracellular tyrosinase activity, cell-free and intracellular α-glucosidase enzyme activity, and effects on MITF protein level and cAMP maturation factor. Our results showed that CMC2.24 showed the greatest efficacy as a tyrosinase inhibitor of all the CMCs and was better than PC as well as a popular tyrosinase inhibitor-kojic acid. Both CMC2.24 and CMC2.23 inhibited tyrosinase enzyme activity by a mixed mode of inhibition with a predominant competitive mode. In addition, CMC2.24 as well as CMC2.23 showed a comparable robust efficacy in inhibiting melanogenesis in cultured melanocytes. Furthermore, after removal of CMC2.24 or CMC2.23 from the medium, we could demonstrate a partial recovery of the suppressed intracellular tyrosinase activity in the melanocytes. Our results provide a proof-of-principle for the novel use of the CMCs that shows them to be far superior to the parent compound, curcumin, for skin depigmentation.

Novel Chemically Modified Curcumin (CMC) Analogs Exhibit Anti-Melanogenic Activity in Primary Human Melanocytes

“…To the best of our knowledge, this is the first study to evaluate the recovery of tyrosinase activity by HEMn-DP cells treated with curcumin analogs. Encouragingly, our results of partial recovery of tyrosinase activity of HEMn-DP cells treated with CMC2.24 and CMC2.23 in this study are in agreement with our results in our previous study [ 41 ].…”

“…Both CMC2.24 and CMC2.23 showed similar potency at inhibiting melanin production in human melanocytes and as CMC2.24 has already been reported to exhibit better stability and solubility than the parent compound curcumin, we anticipated that CMC2.23, a close structural analog of CMC2.24, might also demonstrate a similar activity profile. CMC2.5, on the other hand, is another potent candidate which might be effective as a skin-lightening agent, working through a distinct mechanism of action (suppressing tyrosinase protein levels without affecting tyrosinase activity) in addition to its significant antioxidant activity that was reported in our previous study [ 41 ]. The various distinct steps at which these CMCs act in melanogenesis have been summarized in a schematic ( Figure 4 ).…”

“…A previous study documented the recovery of inhibited tyrosinase activity by novel deoxyarbutin analogs in HEMn-DP cells, establishing their safety [ 45 ]. Currently, there are no reports on the recovery by cells after tyrosinase inhibition in cells treated with curcumin or its derivatives, except in our previous study where we evaluated the recovery capacity of B16F10 mouse melanoma cells after treatment with PC and CMCs [ 41 ]. To the best of our knowledge, this is the first study to evaluate the recovery of tyrosinase activity by HEMn-DP cells treated with curcumin analogs.…”

“…Despite this efficacy, the practical use of curcumin in commercial skin whiteners and clinical pharmaceuticals has been met with limited success due to its poor solubility, stability and bioavailability [ 40 ], in addition to the bright yellow coloration that is aesthetically unpleasing for personal care applications. In our earlier study [ 41 ], we reported on the capacity of four novel CMC analogs to inhibit tyrosinase enzyme activity in an acellular system as well as melanogenesis in B16F10 mouse melanoma cells; we have further begun to elucidate the mechanisms by which they inhibit melanogenesis. In the current study, we have extended our work and evaluated the anti-melanogenic activities of CMC2.14, CMC2.24, CMC2.23 and CMC2.5 in primary human melanocytes derived from darkly pigmented skin to validate the application of these compounds for treatment of hyperpigmentation disorders.…”

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“…So far, dermatologists and other scientists have developed some methods and materials to fulfill these aims. [11] Many skin whitening substances such as kojic acid and arbutin can inhibit the activity of tyrosinase [12] and reduce melanin synthesis. [13] Nicotinamide, the amide form of vitamin B 3 (niacin), [14] can effectively inhibit melanin transfer from melanocytes to keratinocytes.…”