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α-Lipoic Acid Is Ineffective as a Topical Antioxidant for Photoprotection of Skin11This work was done in Durham, North Carolina, USA.

2004; Elsevier BV; Volume: 123; Issue: 5 Linguagem: Inglês

10.1111/j.0022-202x.2004.23446.x

1523-1747

Jing-Yi Lin, Fu-hsiung Lin, James A. Burch, M. Angélica Selim, Nancy A. Monteiro‐Riviere, James M. Grichnik, Sheldon R. Pinnell,

Vitamin C and Antioxidants Research

To the Editor: α-lipoic acid is synthesized in mitochondria of cells (Morikawa et al., 2001Morikawa T. Yasuno R. Wada H. Do mammalian cells synthesize lipoic acid? Identification of a mouse cDNA encoding a lipoic acid synthase located in mitochondria.FEBS Lett. 2001; 498: 16-21https://doi.org/10.1016/S0014-5793(01)02469-3Abstract Full Text Full Text PDF PubMed Scopus (107) Google Scholar) and covalently attached through lysyl residues to α-keto acid dehydrogenase complexes (Biewenga et al., 1997Biewenga G.P. Haenen G.R. Bast A. The pharmacology of the antioxidant lipoic acid.Gen Pharmacol. 1997; 29: 315-331Crossref PubMed Scopus (728) Google Scholar). The attached α-lipoic acid is a cofactor for the enzyme and together with dihydrolipoic acid, its reduction derivative, serve as a redox couple carrying electrons from the substrate of the enzyme to NAD+. Despite the fact that there is little or no free α-lipoic acid normally in tissues (Hermann et al., 1996Hermann R. Niebch G. Borbe H.O. et al.Enantioselective pharmacokinetics and bioavailability of different racemic alpha-lipoic acid formulations in healthy volunteers.Eur J Pharm Sci. 1996; 4: 167-174https://doi.org/10.1016/0928-0987(95)00045-3Crossref Scopus (99) Google Scholar), α-lipoic acid has been proposed as an antioxidant for topical application to skin to protect the skin against ultraviolet photodamage including skin cancer and photoaging changes (Podda et al., 2001Podda M. Zollner T.M. Grundmann-Kollmann M. Thiele J.J. Packer L. Kaufmann R. Activity of alpha-lipoic acid in the protection against oxidative stress in skin.Curr Problems Dermatol. 2001; 29: 43-51Crossref PubMed Google Scholar). We have previously studied topical combination vitamin C (L-ascorbic acid) and vitamin E (α-tocopherol) in skin and demonstrated synergistic photoprotection (Lin et al., 2003Lin J.Y. Selim M.A. Shea C.R. Grichnik J.M. Omar M.M. Monteiro-Riviere N.A. Pinnell S.R. UV photoprotection by combination topical antioxidants vitamin C and vitamin E.J Am Acad Dermatol. 2003; 48: 866-874https://doi.org/10.1067/mjd.2003.425Abstract Full Text Full Text PDF PubMed Scopus (267) Google Scholar). In this study, we use the same model to evaluate topical α-lipoic acid formulations. The experimental design has previously been published in detail (Lin et al., 2003Lin J.Y. Selim M.A. Shea C.R. Grichnik J.M. Omar M.M. Monteiro-Riviere N.A. Pinnell S.R. UV photoprotection by combination topical antioxidants vitamin C and vitamin E.J Am Acad Dermatol. 2003; 48: 866-874https://doi.org/10.1067/mjd.2003.425Abstract Full Text Full Text PDF PubMed Scopus (267) Google Scholar). Experiments were performed on weanling white Yorkshire pigs in accord with the guidelines prepared by the Committee on Care and Use of Laboratory Animals of the Institute of Laboratory Resources, National Research Council (National Institutes of Health, publication no. 86-23, revised 1996). Five percent DL-α-lipoic acid (Pharmline, Florida, New York) was dissolved in 50% ethanol. A commercial formulation containing 15%L-ascorbic acid (vitamin C) and 1%DL-α-tocopherol (vitamin E) (Primacy C+E, SkinCeuticals, Garland, Texas) and a commercial formulation of á-lipoic acid (Face Firming Activator with DMAE, N.V. Perricone, Meriden, Connecticut) were purchased and opened immediately prior to use. Antioxidant solutions (500 μL) were applied to patches of shaved back skin (7.5 × 10 cm) daily for 4 days. A 1000-W solar simulator (Lightning Cure 200, Hamamatsu, Japan) fitted with a WG295 Schott filter to eliminate wavelengths less than 295 nm delivered ultraviolet irradiation to the skin's surface through a liquid light guide at an intensity of 5 mW per cm2 of UVB and about 40 mW per cm2 of UVA as measured by a radiometer (IL1700, International Light, Newburyport, Mississippi). MED was determined as the lowest dose resulting in erythema with perceptible borders (40 mJ per cm2 of UVB). Each patch was given solar-simulated irradiation in triplicate from 0.5 × to 2.5 × MED at 0.5 × MED intervals or 1 × to 5 × MED at 1 × MED intervals. Evaluation was carried out 24 hours later. Erythema was measured by colorimeter evaluation in the “a” mode (ColorMouse Too, Color Savvy Systems, Springboro, Ohio) of 8 × 12-in photographic enlargements of skin. Each spot, together with adjacent unirradiated skin, was measured in triplicate. The difference between irradiated and unirradiated skin determined the erythema measurement. Sunburn cells were determined in formalin-fixed 8 mm punch biopsy sections stained with hematoxylin and eosin. When irradiation damage was extensive, the figure 35 sunburn cells per mm was used as an upper limit. Results are expressed as mean±SD (n=3). The p-values were calculated by Student's t test. No significant protection could be detected either by reduction of erythema (Figure 1a) or by sunburn cell (Figure 1b) analysis when untreated skin was compared to skin treated by 5%α-lipoic acid or a commercial formulation of α-lipoic acid. As previously reported, a formulation combining 15%L-ascorbic acid and 1%α-tocopherol provided meaningful photoprotection against solar-simulated irradiation as revealed by erythema (Figure 2a) or sunburn cells (Figure 2b). Adding 5%α-lipoic acid to the vitamin C and E solution provided no additional photoprotection (Figure 2a and b).Figure 2Topical α-lipoic acid does not supplement photoprotection provided by topica vitamins C+E. Colorimeter (a) and sunburn cell (b) data from untreated skin, skin treated with 5% vitamin C and 1% vitamin E, or skin treated with 5% vitamin C and 1% vitamin E and 5%α lipoic acid.View Large Image Figure ViewerDownload (PPT) Although α-lipoic acid has antioxidant properties, its potential as an antioxidant has been thought to depend on reduction to the more potent dihydrolipoic acid. Although small amounts of dihydrolipoic acid have been measured in murine skin following topical application of α-lipoic acid (Podda et al., 1996Podda M. Rallis M. Traber M.G. Packer L. Maibach H.I. Kinetic study of cutaneous and subcutaneous distribution following topical application of [7,8-14C]rac-alpha-lipoic acid onto hairless mice.Biochem Pharmacol. 1996; 52: 627-633https://doi.org/10.1016/0006-2952(96)00337-1Crossref PubMed Scopus (40) Google Scholar), this reaction is very inefficient. Reduction of α-lipoic acid to dihydrolipoic acid can be achieved with the mitochondrial enzyme dihydrolipoamide dehydrogenase, but it is not clear whether free α-lipoic acid has ready access to the mitochondrial compartment (Biewenga et al., 1996Biewenga G.P. Dorstijn M.A. Verhagen J.V. Haenen G.R. Bast A. Reduction of lipoic acid by lipoamide dehydrogenase.Biochem Pharmacol. 1996; 51: 233-238Crossref PubMed Scopus (44) Google Scholar). Glutathione reductase also can carry out the reduction, but the rate of the reaction is slow (Pick et al., 1995Pick U. Haramaki N. Constantinescu A. Handelman G.J. Tritschler H.J. Packer L. Glutathione reductase and lipoamide dehydrogenase have opposite stereospecificities for alpha-lipoic acid enantiomers.Biochem Biophys Res Commun. 1995; 206: 724-730https://doi.org/10.1006/bbrc.1995.1102Crossref PubMed Scopus (84) Google Scholar). Several studies have shown that the reaction rate constant of dihydrolipoic acid with reactive oxygen species is slower than that of α-lipoic acid (Bisby and Parker, 1998Bisby R.H. Parker A.W. Antioxidant reactions of dihydrolipoic acid and lipoamide with triplet duroquinone.Biochem Biophys Res Commun. 1998; 244 ([erratum appears in Biochem Biophys Res Commun 1998 Aug 10;249(1):297].): 263-267Crossref PubMed Scopus (18) Google Scholar;Lu and Liu, 2002Lu C. Liu Y. Interactions of lipoic acid radical cations with vitamins C and E analogue and hydroxycinnamic acid derivatives.Arch Biochem Biophys. 2002; 406: 78-84https://doi.org/10.1016/S0003-9861(02)00411-3Crossref PubMed Scopus (39) Google Scholar;Trujillo and Radi, 2002Trujillo M. Radi R. Peroxynitrite reaction with the reduced and the oxidized forms of lipoic acid: New insights into the reaction of peroxynitrite with thiols.Arch Biochem Biophys. 2002; 397: 91-98https://doi.org/10.1006/abbi.2001.2619Crossref PubMed Scopus (166) Google Scholar). Moreover, dihydrolipoic acid may be pro-oxidant (Scott et al., 1994Scott B.C. Aruoma O.I. Evans P.J. et al.Lipoic and dihydrolipoic acids as antioxidants. A critical evaluation.Free Radical Res. 1994; 20: 119-133Crossref PubMed Scopus (282) Google Scholar). Once α-lipoic acid is oxidized after reaction with a free radical, its one-electron oxidation product is strongly pro-oxidant potentially causing free radical damage to cellular and tissue components(Lu and Liu, 2002Lu C. Liu Y. Interactions of lipoic acid radical cations with vitamins C and E analogue and hydroxycinnamic acid derivatives.Arch Biochem Biophys. 2002; 406: 78-84https://doi.org/10.1016/S0003-9861(02)00411-3Crossref PubMed Scopus (39) Google Scholar). In contrast, for example, ascorbyl radical has a relatively low pro-oxidant potential (De Tullio and Arrigoni, 2004De Tullio M.C. Arrigoni O. Hopes, disillusions and more hopes from vitamin C. [Review] [103 refs].Cell Mol Life Sci. 2004; 61: 209-219https://doi.org/10.1007/s00018-003-3203-8Crossref PubMed Scopus (67) Google Scholar). In fact, it prefers to react with itself and disproportionate rather than attacking other substrates. It is not clear whether free α-lipoic acid provides much physiologic antioxidant activity. In mitochondria it is covalently attached to mitochondrial enzymes through lysyl residues. Although available in the diet in tissues with high metabolic activity (e.g., heart), proteolytic enzymes do not effectively cleave the linkage between α-lipoic acid and lysine (Biewenga et al., 1997Biewenga G.P. Haenen G.R. Bast A. The pharmacology of the antioxidant lipoic acid.Gen Pharmacol. 1997; 29: 315-331Crossref PubMed Scopus (728) Google Scholar). Free α-lipoic acid is not normally detected in plasma (Hermann et al., 1996Hermann R. Niebch G. Borbe H.O. et al.Enantioselective pharmacokinetics and bioavailability of different racemic alpha-lipoic acid formulations in healthy volunteers.Eur J Pharm Sci. 1996; 4: 167-174https://doi.org/10.1016/0928-0987(95)00045-3Crossref Scopus (99) Google Scholar) or in skin (Podda et al., 1994Podda M. Tritschler H.J. Ulrich H. Packer L. Alpha-lipoic acid supplementation prevents symptoms of vitamin E deficiency.Bioch Biophys Res Commun. 1994; 204: 98-104https://doi.org/10.1006/bbrc.1994.2431Crossref PubMed Scopus (100) Google Scholar). Following oral or parenteral infusion, it remains in plasma only about 30 minutes and is cleared by liver during first pass (Hermann et al., 1996Hermann R. Niebch G. Borbe H.O. et al.Enantioselective pharmacokinetics and bioavailability of different racemic alpha-lipoic acid formulations in healthy volunteers.Eur J Pharm Sci. 1996; 4: 167-174https://doi.org/10.1016/0928-0987(95)00045-3Crossref Scopus (99) Google Scholar;Teichert et al., 2003Teichert J. Hermann R. Ruus P. Preiss R. Plasma kinetics, metabolism, and urinary excretion of alpha-lipoic acid following oral administration in healthy volunteers.J Clin Pharmacol. 2003; 43: 1257-1267https://doi.org/10.1177/0091270003258654Crossref PubMed Scopus (132) Google Scholar). Similar rapid clearance may also occur after percutaneous absorption. In summary, we have tested α-lipoic acid at high doses topically to skin to determine if it provides photoprotection against solar-simulated radiation. We were unable to detect protection using α-lipoic acid alone or together with vitamins C and E. In addition, a commercial formulation of α-lipoic acid provided no protection. Although we are unable to explain this lack of effect, it seems reasonable that α-lipoic acid absorbs ultraviolet light in the UVA spectrum—it has an absorption spectrum at 330 nm (Matsugo et al., 1996Matsugo S. Han D. Tritschler H.J. Packer L. Decomposition of alpha-lipoic acid derivatives by photoirradiation-formation of dihydrolipoic acid from alpha-lipoic acid.Biochem Mol Biol Int. 1996; 38: 51-59PubMed Google Scholar)—and is oxidized to its free radical and subsequently destroyed (Podda et al., 1997Podda M. Traber M.G. Packer L. Alpha-lipoate: Antioxidant properties and effects on skin.in: Fuchs J. Packer L. Zimmer G. Lipoic Acid in Health and Disease. Dekker, New York1997: 163-180Google Scholar). Conversion to dihydrolipoic acid is inefficient resulting in little interaction with vitamins C and E to augment their photoprotection. This research was supported in part by NIH grant R43CA83538. Solutions were prepared by Mostafa M. Omar, PhD of Phytoceuticals, Elmwood Park, New Jersey. Thanks to Doren Madey, PhD for help in preparation of the manuscript. Sheldon Pinnell is a consultant for SkinCeuticals, Garland, Texas.