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33 Antiaging Actives in Sunscreens Karl Lintner Sederma, Paris, France Introduction Strategies of Antiaging Actives in Sunscreens Prevention of Damage (“Slowing Down the Aging Process”) Vitamins Botanicals Enzymes Miscellaneous Treatment of UV-Induced Age Symptoms Barrier Repair Tissue Repair Conclusions References 673 677 677 677 680 681 683 684 685 686 689 689 INTRODUCTION The first and second editions of Sunscreens did not contain a chapter equivalent to the present one. Including “antiaging actives” in the present book reflects some of the changes occurring in cosmetic formulations and marketing strategies. As in so many other domains, we see a blurring of frontiers, a mixing of categories, and a (deliberate?) gradual disappearance of clear distinctions and definitions. 673 674 Lintner Although the title of Sunscreens has not changed and certainly suggests to most readers the general category of “cosmetic products used on the skin during extensive sun exposure in order to protect the skin against the deleterious effects of direct sunlight,” the apparent simplicity of this description is deceptive. Even more ambiguity is contained in the title of this chapter, with the two powerful, but vague, concepts of “antiaging” and “actives” (or “cosmeceuticals,” as these ingredients are sometimes, erroneously, called). It appears therefore necessary to start with a few definitions of our own, that we will use within the scope of this chapter, without prejudice to different meanings in other parts of the book. Let us call “sunscreens” the finished cosmetic consumer product that bears a clear message of “protection against solar radiation” such as the prevention of erythema, sunburns, sometimes even cancer. This would include in most cases “suntan lotions,” “sun care products,” “sunblocks,” and the like. Generally, it would not include “after-sun lotions” and “self-tanning products.” Although the word “sunscreen” is sometimes also used to designate the chemical entity that blocks the sunlight from reaching the skin, these chemicals contained in “sunscreens” should be called ultraviolet (UV) filters or UV reflectors. Difficulties in nomenclature also arise because of different legislations in different parts of the world (cf. the section on regulatory aspects) and because of technical and marketing considerations: a “sunscreen” of today contains, more and more often, specific skin and/or body care active ingredients, accompanied by a corresponding claim (this is the reason for this chapter); on the other hand, an increasing number of classic “skin care” (i.e., face care, lip care, makeup, body care, and even hair care) products boast sun protection factors (SPFs) in the 5– 15 range. These products have primary skin care claims (moisturizing, antiwrinkle, firming, . . .) and offer the sun protection as an additional benefit. So where is the borderline between the two? A “sunscreen” of SPF 15 with an additional antiwrinkle claim is a “sunscreen” (e.g., Biotherm’s recent launch with exactly that name: “Antiwrinkle Suncare”) because the marketeer positions the product as such (advertising, point of sale, timing of promotional activity), whereas Yves-Saint Laurent’s spring launch of Age Expert (“age-defying cream”) has an SPF15 but is clearly not positioned as a “sunscreen”. It claims to contain DHEA-like actives and lycopene as a free radical scavenger and is a “classical” face care product. In any case, the New Zealand Society of Dermatology proposes on its website that “sunscreens should be applied daily, more often when outdoors.” While this makes sense, it is certainly not the daily routine of the general population. How about “actives”? My opinion and arguments for it can be found in the proceedings of the PCIE conference (1). Briefly, any cosmetic ingredient that has (i) demonstrated cosmetic activity on human skin (or its appendages), (ii) a substantiated claim, and (iii) a plausible “story” to go with it can be considered an active: this encompasses then the wide field of ingredients (such as found in the CTFA/INCI dictionary) from botanicals (various types of plant extracts) to pure chemical entities that possess a function that is clearly different from the Antiaging Actives in Sunscreens 675 galenic purpose (emulsifiers, texturizers, thickeners, preservatives, fragrances etc.). Again, it may occur that an ingredient functions as an “active” in one product (positioned as such by the marketeer) and as a basic ingredient in another product (glycerin, lecithin, and lanolin come to mind). In the main body of this chapter, we shall concentrate on “actives” that are particularly suited or are already in use in the general category of “sunscreens,” “actives” that make sense in the context of sun care and skin protection/treatment. We can therefore now drop the quotation marks from the word active. Finally, “antiaging” (again with quotation marks): the term is catchy, seductive, but very vague. Some countries have regulations forbidding the use of this “claim” in cosmetic advertising or on the packaging. As usual, there are two aspects to the concept: prevention and treatment. Antiage prevention implies that a consumer product helps “reduce the speed of the appearance of the clinical signs of (cutaneous) aging,” based on the protective active contained in the product. Antiage treatment promises to reverse (some of ) the visible signs of skin aging (such as wrinkle reduction, firmness improvement, moisturization of dry skin, etc.), based on actives that “restore, regenerate, repair, . . .” skin items such as “barrier, extracellular matrix, collagen fibers, hydrolipid balance” and the like. If one really wants to get a more global perspective of the antiaging field in general, books like Pharmacological Intervention in Aging and Age-Associated Disorder (2) are a good place to start. Yu and Yang introduce the discussion there with a “critical evaluation of the Free Radical Theory of Aging” (see in following text, the impact of free radicals and antioxidant strategies). Many articles and references therein are useful for finding ideas in “antiaging”. In the following sections we consider both types of “antiaging” actives and their respective merits in sunscreens. I apologize for the lengthy introduction and the many quotation marks and hope the reader has thus a clear picture of where this chapter is headed and what the various terms are meant to convey. In view of the many detailed chapter headings before and after this one in the present book, it would be redundant to repeat much that is described about the dangers of the sun to our skin, about the photobiology, the physics of filters, the differences between chronological aging and photoaging. If modern UV filters are so well suited to protect us against the sun’s dangerous rays, for what reason (other than a marketing and/or label claim) should the formulator of a modern sunscreen add antiaging actives to his product? For one, and most importantly, UV filters are not absolute: even an SPF 60 (not allowed as a claim everywhere) will wear off with time, or may not be applied in sufficient amount from the beginning, or the exposure of the person wearing it continues beyond the period of protection. Any well-chosen additional active in the product will help decrease the damages that are not prevented by the UV filter. For instance, this seems to be particularly true for the combination of UV-A filters with antioxidant protective molecules. The presently available 676 Lintner UV-A filters are rarely (not?) able to block out all of the UVA radiation such as to prevent all free radical generation in the deeper layers of the skin. Second, the trend in all cosmetic formulas and products goes that way: makeup mascaras, foundations, lip sticks, powders, also cleansers, body care, and scalp care SKUs all contain actives for additional benefits. True skin care needs a global, and continuous, approach. We need the sunlight for the synthesis of vitamin D and for our psychic well-being (“healthy tan”), and we desire silky, youthful skin: for this we need the optimum combination of sunscreen and skin care actives. Before reviewing the traditional actives used and useful in sunscreens and presenting a few new ideas in the final section, Table 33.1 summarizes the rationale for the different types of actives that might make sense in sunscreens, Table 33.1 Rationale for Adding Different Types of Actives to Sunscreens Type of danger/ damage Dryness Dryness Skin sagging Wrinkles Skin thinning Roughness Inflammation (redness) Melasma, age spots Yellowing, elastosis Telangiectasis Blackheads, whiteheads Free radical damage Lipoperoxidation Enzyme damage DNA damage Apoptosis Type of active proposed “Cause” Moisturizer/humectant Barrier repair: ceramides Firming, elasticity enhancing Tissue repair: ECM, collagen stimulation, cell metabolism, skin tighteners Tissue repair: ECM, collagen stimulation, cell metabolism, skin tighteners Smoothing, emollients Anti-inflammatories, soothing actives Skin “whitening” IR, UV-A, UV-B UV-B UV-A, UV-B Elastase inhibitors UV-A, UV-B Anticouperose, veinotonic Antiacne UV-B UV-B Radical scavengers UV-A Antioxidants Enzymes, pseudoenzymes DNA repair Cell repair UV-A UV-A, UV-B UV-B UV-B Increased sophistication UV-A, UV-B UV-A, UV-B UV-B UV-A, UV-B UV-A P Antiaging Actives in Sunscreens 677 although some appear far-fetched. Very roughly, one can partition these actives along the following lines. UV-B rays generate actinic damages that need antiage treatment (long-term), UV-A radiation needs to be addressed with actives more immediately: to prevent oxidation, to scavenge the radicals, to reduce local inflammation and thus avoid long-term damages of molecular nature to accumulate (antiage Prevention). STRATEGIES OF ANTIAGING ACTIVES IN SUNSCREENS Prevention of Damage (“Slowing Down the Aging Process”) Vitamins One of the first and most widely used categories of actives formulated in sunscreens is the one comprising the vitamins C and E, sometimes A (retinoids) or a few of the B group. The literature on the effects of vitamins C and E as photoprotective agents in cell cultures (in vitro) and on animals is impressively large. Although general consensus is expressed that the protective effect afforded by these molecules “might be beneficial” to human skin, there is astonishingly little documentation of the benefits of vitamin uses in cosmetic finished products, especially sunscreens, on human skin (in vivo). Pehr and Forsey concluded in 1993, that “after 44 years of research there is still scant proof of vitamin E’s effectiveness [. . .]; it is of no use in [. . .] skin damage induced by ultraviolet light” (3). It is not the purpose of this chapter to present an exhaustive review of this topic, as antioxidants will also be discussed in a separate chapter. Such a review can be found, for instance, in Pinnell (4) in the form of a lecture, followed by a quiz. A rapid overview of the literature however shows that research into the effects of topical vitamin application continues, many papers focusing on combinations of vitamins, such as E and C, E and A. Vitamin E (a-tocopherol) is a ubiquitous, liposoluble molecule, the major activity of which is as an antioxidant. Although more powerful antioxidants can be found in nature, vitamin E is most accessible, by synthesis or extraction, is colorless and well documented as being toxicologically safe. Ritter et al. (5) show the beneficial effects of tocopherol application before UV irradiation on mice. They note an increase in epidermal thickness, which might contribute to the decrease in the number of sunburn cells. The concentration of tocopherol in the vehicle (50% in ethanol) is however quite unrealistic in cosmetic and sunscreen applications. Saral et al. (6) studied vitamin E acetate (the more stable ester form of vitamin E that is preferentially employed in sunscreens) by applying it topically for 3 weeks on guinea pigs before a single UV-B dose. Measuring lipid peroxide levels and enzyme scavenging activity (cf. also following text) these authors find that tocopherol acetate did prevent the UV-B-induced effects. Trevithick et al. (7) studied the application of tocopherol acetate immediately after UV-B irradiation on mice and found decreases in sunburn cell number, inflammatory infiltration, 678 Lintner and edematous swelling. Even delayed application (8 h after irradiation) afforded some protection, again however at high concentrations (5%). Another interesting study was carried out on the antioxidant activity of a-tocotrienol in topical application (8). Although it did not address UV irradiation, the oxidant damages were induced by benzylperoxide (10%). Application of a 5% w/v preparation of a-tocotrienol for 7 days reduced the BPO-induced lipid peroxidation significantly. For cosmetic purposes, 5% vitamin E again appears unrealistically high. Vitamin C has several properties, which make it attractive to the formulator of sunscreens. Not only does the molecule possess antioxidant (reducing) activity, but it also stimulates the synthesis of collagen (in vitro) and contributes to the hydroxylation and the correct lay-down of collagen fibers in skin tissue. Its cosmetic use is found particularly in “antiwrinkle” creams (based on the collagen stimulation claim) and in “skin whitening” products (based on the inhibition of melanogenesis). Darr et al. (9) investigated the topical use of vitamin C on pigskin and found protection against UV-B damages as measured by erythema and sunburn cell formation. As innate vitamin C concentrations decrease as a result of UV irradiation, topical supplementation is a potential strategy to counter this deleterious effect of sunlight. Follow-up studies later confirmed the protective effect of vitamin C, when formulated together with a UV filter (oxybenzone) (10). An even more broad-spectrum protection is achieved with the combination of vitamin C, vitamin E and the sun-filter, as vitamin C affords particular protection against the UV-A-mediated phototoxicity in this animal model. Some more recent studies suggest benefits to using vitamin C together with vitamin E in topical products such as sunscreens. Moison et al. (11) describe a synergistic effect between the two molecules in protecting the lipids, also on pigskin; moreover, the inherent vitamin C and E content of the skin is maintained at its levels, against depletion by UV-B radiation. Steenvorden and Beijersbergen (12) investigated vitamins C and E independently and conjointly: topical application on mice before UV-B irradiation led to reduced immunosuppression. In their model, no synergy was found, however. Both studies cited showed that vitamin C concentration needed to be about 500 – 1000 times higher than vitamin E to obtain comparable efficacy. Human in vivo studies were carried out by Dreher et al. (13) using three antioxidant molecules, alone or in combination: vitamin C, vitamin E, and melatonin. Slight synergistic results are obtained by combining the ingredients two by two or in a threesome and applying them in a vehicle 30 min before UV exposure. Skin color and skin blood flow were used as markers for UV-induced damage. The same authors then studied the effects of these combinations when applied 30 min, 1 h, and 2 h after UVR exposure, using the same end-points (14). As no protective effect whatsoever was noted, the authors concluded that UVR-induced skin damage is rapid; antioxidants can alleviate or prevent Antiaging Actives in Sunscreens 679 damages only when present before or during sun exposure. Melatonin is of course used in oral supplementation against jet lag, but Reiter et al. (15) describe in much detail its antioxidant, radical scavenging activity, and life span prolongation! Lin et al. (16) tried a further combination of 15% vitamin C and 1% vitamin E on pig skin and found that repeated application of this cocktail reduced erythema, sunburn cells, as well as thymidin dimers (DNA damage) generated by repeated UV irradiation with a solar simulator. While the protection against this latter aspect is of importance in the prevention of mutations and their consequences, the sunscreen formulator may again have difficulties in incorporating these levels of ingredients in the finished product. And what about vitamin A and its derivatives? Kligman (17) in 1987 recommended the use of retinoic acid to replenish the inherent pool of this molecule in the skin after its depletion by UV light. Together with Schwartz (18) he also demonstrated that post-UV irradiation treatment with 0.05% retinoic acid stimulated collagen synthesis in vivo in albino hairless mice. Ho et al. (19) showed in 1992 that retinoic acid augments UV-induced melanogenesis, an interesting side effect to all other activities known for retinoids. The study was carried out on a specific mouse strain and confirmed on two human volunteers. As retinoic acid is considered a prescription drug in most countries, the cosmetic industry became interested in retinol, retinol esters, and retinaldehyde. Thus, more recently, Boisnic et al. (20) published a study with a retinaldehyde cream, applied to an ex vivo human skin model. Eighteen days of regular UV-A exposure simulated photoaging; this was followed by application of the retinaldehyde cream for 2 weeks. The UV-A-induced alterations of collagen and elastic fibers were reversed by the retinaldehyde, and collagen synthesis rates were restored to the levels of unexposed skin. Sorg and colleagues (21,22) have interested themselves in the combination effects of retinoids and vitamin E; they find certain specific benefits, depending on the type of irradiation (UV-A, UV-B), the time of application (before or after exposure), and other parameters. More data on human volunteers in studies on retinoids in conjunction with UV radiation can be found in Kang et al. (23). One opposing opinion is, however, expressed in Ref. (24) under the aggressive title: “Tretinoin and cutaneous photoaging: new preparation. Guaranteed adverse effects!” Knowing that the skin contains various antioxidants all together, a more holistic approach was taken by Greul et al. (25) where a combination of b-carotene, lycopene, vitamin C, vitamin E, selenium, and proanthocyanidins was tested in a double-blind placebo-controlled human study involving UV irradiation and skin analysis. Findings concerned significant differences in MMP 1 and MMP 9 expression and a slowdown in the development and grade of UV-induced erythema. This reference is given tongue in cheek (excuse the pun), as the study did not use the antioxidant mixture topically, but orally. Somewhat similar results were obtained in the SUVIMAX study (26). In summary, “vitamins are good for you.” Their use in sunscreens is widespread; based on the numerous studies, even if most of them are animal or in vitro 680 Lintner studies, their claim to “antiaging” activity is not far-fetched. Vitamins C and E would be considered more of the “preventive” type (antioxidant, to be used before or during sun exposure), vitamin A and derivatives are more the “repair” type, undoing some of the UV-caused damages. Their main drawback is the difficulty in formulating stable vehicles, such that the right amount of efficacious vitamins can be guaranteed for sufficient shelf life. Botanicals Like the antioxidants, botanicals, also called plant extracts, are also discussed in another chapter in the present book. Nevertheless, they merit a short mention here, as an increasing number of ingredients of plant origin are offered and used that are tested and positioned as “antiaging” actives, and thus used or useful in sunscreens. Plant extracts cover the spectrum from hydroglycolic solutions of analytically ill-defined nature to pure, isolated, chemically identified molecules, and all products that present intermediate stages of purification. The reputation of the plant kingdom is one of almost unlimited source of potential healing activities, thousands of substances yet undiscovered. A closer look reveals, however, that with some notable exceptions, a few broad categories suffice to describe the benefits obtained from plant extracts for cosmetic claims: we find antioxidant activity (polyphenols, vitamins, flavonoids), anti-inflammatory properties (nonsteroidal enzyme inhibitors), tissue repair molecules (di- and triterpenes). All of these activities can be employed for “antiage” claims, all of them make sense in the context of sunscreen formulation. A few references gleaned from the peer-reviewed literature shall illustrate this concept. It is of course impossible to list here all the commercially available plant-derived cosmetic ingredients (cf. CTFA dictionary) that are claimed to be antiaging based on some or another in vitro, ex vivo, or even in vivo test with or without UV irradiation included in the test protocol. Green tea is a favorite among the botanicals with well-known reputation. In vitro scavenging of hydrogen peroxide and prevention of UV-induced oxidative damages on skin cells in culture by various fractions of green and black teas, including purified epigallocatechin gallate (EGCG) were described by Wei et al. (27). The pure molecule enhanced the observed activity and is considered the major active substance in these preparations. An in vivo study by Vayalil et al. (28) on hairless mice confirmed the prevention of UV-induced lipid peroxidation by green tea polyphenols, such as EGCG. Interestingly, the authors also quantified the amount of inherent antioxidant enzymes (catalase, glutathione peroxidase): whereas UV irradiation depleted these enzymes in the skin, EGCG application before single UV-B doses prevented this loss by 50 –90% (see also following text). Less well known botanical extracts such as those obtained from methanolic maceration of Capparis spinosa L. buds (29), crude ethanol extracts from Culcitium reflexum H.B.K. (30), or Chromolaena odorata (31), to cite just a few exotic Antiaging Actives in Sunscreens 681 ones, all contain flavonoids, phenolic acids, coumarins and the like. They all show in vitro antioxidant activity that can be used for antiage claims in sunscreens. Prunus persica Batsch extracts rich in kampferol glycoside derivatives (32) also showed inhibition of UV-induced edema on mouse ear and tumor prevention (33) in UV-B- and UV-C-irradiated mice. A further aspect, not yet widely recognized or mentioned in this context of sunscreen protection by antiaging products is discussed by Okano et al. (34). It is known that with advancing age, proteins and sugar molecules react, unspecifically, in a process called glycation to give what has been aptly termed, advanced glycation endproducts (AGEs). Okano et al. describe that AGEs are not only inherently a sign of aging skin (less elasticity of the glycated proteins) but also contribute actively to aging by reducing fibroblast proliferation, matrix synthesis, and by generating reactive oxygen species (ROS) during UV exposure! He then describes that unspecified extracts of Paenia suffruticosa and Sanguisorba officinalis inhibit AGE formation and scavenges hydrogen peroxide at the same time. A review of photochemoprevention by botanical antioxidants in view of their use in sunscreens is given in Ref. (35). A typical example of anti-inflammatory activity of a botanical extract useful in a suncare product is described by Hughes-Formella et al. (36). UV-B irradiation, provoking erythema on the back of 30 volunteers was followed by application of a Hamamelis virginiana lotion 7, 24, and 48 h after irradiation. Significant differences in erythema values (chromameter, visual scoring) were observed with respect to the vehicle lotion. This type of use is, however, better suited for after-sun products than for the sunscreen itself and we shall not dwell on these applications. Enzymes We have cited earlier two studies (6,28) that mentioned antioxidant enzymes of the skin. This aspect has received less attention in the sunscreen and protection field; two reasons may account for this. Technical difficulties in analyzing enzyme activities on human skin, and the inherent instability of enzymes which make them hard to formulate and stabilize in finished cosmetic sunscreens. Nevertheless, basic research into the innate enzyme defense system of the skin has progressed, and a number of in vitro, animal and human in vivo studies point to the delicate balance that is required between the various enzymes in the skin. We shall first review the salient facts about cutaneous defense enzymes and then discuss the possibility of using antiaging actives within this scope. Once again, the problem turns around the free radicals, lipoperoxidation, and other oxidative damages. ROS such as superoxide anion, hydroxyl radical, singlet oxygen, and hydrogen peroxide cause numerous deleterious effects on structural and functional (enzyme) proteins, lipid membranes, tissue polysaccharides, and genetic material (DNA). The molecules present in the skin that are 682 Lintner supposed to protect us against these damages are the vitamins (see preceding text), a few other antioxidants (melanin, urocanic acid, glutathione, and ubiquinone) and specific enzymes: essentially superoxide dismutase (SOD), glutathione peroxidase (GPO), and catalase. It now has become evident that these inherent antioxidant defense systems of the skin are rapidly overwhelmed by the amount of sun exposure we stress them with in today’s lifestyle. Not only are vitamins C and E depleted in the skin by UV irradiation, but the same also occurs with the enzymes. Miyachi et al. (37) describe the decrease of SOD activity in mice after a single dose of UV light, Pence and Naylor (38) confirm this observation in hairless mice and add that catalase activity also was significantly depressed. Punnonen et al. (39) extended this observation to human epidermis. A quantitative analysis of the localization of these enzymes (and nonenzymatic antioxidants) in murine skin and their decrease after UV exposure is presented by Shindo et al. (40). These acute effects are in opposition to long-term irradiation, as Okada et al. (41) show: after 36 weeks of regular UV exposure, SOD activity increased with UV-B, but not with UV-A; catalase activity however was strongly depressed by UV-A. Although catalase, which detoxifies hydrogen peroxide into water and molecular oxygen is the enzyme most frequently cited as being necessary in conjunction with SOD, which transforms the superoxide anion into hydrogen peroxide (itself a cytotoxic molecule), the two enzymes do not react in similar ways to long-term UV exposure. A few, more pointed investigations into the details can be found in Shindo and Hashimoto (42), Filipe et al. (43), Aricioglu (44), and Naderi-Hachtroudi et al. (45) and references therein. A thorough investigation on humans, carried out over winter and summer season, confirms this fact: catalase is easily destroyed by UV-A light in summer, more active in winter (oh, the logic of nature!), whereas SOD is much more resilient (46). This then leads to a potential buildup of hydrogen peroxide in the skin, not necessarily the best thing to occur. The need for a balanced antioxidant enzyme system thus becomes apparent. Two approaches are possible: (a) to stimulate and/or protect the innate enzyme system, so that even under UV exposure, it retains its efficacy, and (b) to supply the lacking enzymes by topical application, for instance, within a sunscreen, as well as presun or postsun products. Hoppe and colleagues (47), as well as Maes and coworkers (48) presented examples of the first strategy: they show that molecules such as salicin in skin fibroblasts (Hoppe) and vitamin D derivatives or betulinic acid in keratinocytes (Maes) are able to stimulate the synthesis of heat shock proteins which are able to protect the catalase against UV-induced degradation. These molecules could therefore be used advantageously in sunscreens as antiaging actives in as much as they induce protection of our own antiaging defense systems. Other molecules that induce heat shock could be worthwhile looking for. A more controversial proposal is put forward by Inal et al. (49) who show that treatment of rats with quercetin (a plant-derived molecule) reduces the UV induced damages Antiaging Actives in Sunscreens 683 to SOD, catalase, GPO significantly. Quercetin is often described as mutagenic (based on Ames tests) and its behavior under UV light (photostability) would need investigation. Once more it may be instructive to refer to orally administered substances such as deprenyl, a monoamine oxidase B inhibitor which upregulates SOD activity and has shown to prolong the “remaining life span of old rats” (50). Antiaging concepts may be found in many strange places. Strategy (b) has a few limitations. Usually available enzymes such as SOD and catalase (extracted from yeast or other biotech sources) are not easy to stabilize in cosmetic formulas, to say the least. Complicated packaging stratagems or encapsulation techniques may overcome the problem; it is, however, well known that enzymes—relatively large proteins—are inherently unstable in aqueous environments, and also heat and UV sensitive. Furthermore, SOD alone on the skin would lead, at least theoretically, to a buildup of hydrogen peroxide, already described by Maes and colleagues (46) as being the “natural” problem of seasonal variations of these enzyme activities. Adding the fragile catalase is not only difficult, it is also not possible for any formula sold in Europe because of an archaic prohibition of catalase use in cosmetic products (51). A neat solution to this problem is afforded by antioxidant enzymes originating from organisms that live and thrive under extreme conditions of heat: the “extremophiles.” Discovered a little more than a decade ago, these bacteria live close to the hydrothermal vents at the bottom of the ocean, at temperatures that can reach 80 –1008C. It is possible today to cultivate these organisms at sea level, in industrial fermenters, and to extract heat-stable antioxidant enzymes that mimic the skin’s SOD, catalase and GPO activity. Further, the enzymes are the more active, the hotter it gets, up to 1008C (which is unrealistic from a cosmetic point of view anyway). They are thus ideal for incorporation into sunscreens where the exposure to UV and to the sun’s heat will not only not destroy the defensive activity afforded by them, but also even increase it with increasing outside temperature of irradiation. An active ingredient based on this concept is described by Lintner et al. (52,53). Thermus thermophilus bacteria, harvested 6000 ft below the California coast, are fermented at 758C, then extracted and concentrated to yield a high potency solution containing superoxide anion dismutating (SOD), hydrogen peroxide converting, and GPO mimicking activity. In vitro tests carried out on this cosmetic ingredient include protection of human fibroblasts in culture, lipoperoxidation inhibition, protection of DNA against the formation of 8-oxo-guanidine, collagen contraction. Studies on human volunteers show the persistence of cutaneous catalase against UV-A irradiation and a decrease in in vivo lipoperoxidation of the stratum corneum. Miscellaneous A few more (nonexhaustive!) ingredients of diverse nature that might be of interest in photoprotection can be found in the literature. Pinnell and coworkers have reviewed the evidence supporting the antioxidant role of zinc in UV protection 684 Lintner (54), Mitani et al. (55), on the other hand, reminds us that iron is bad for the skin and that Kojic acid treatment prior to sun exposure may help reduce UV-induced wrinkling (in hairless mice). A complex but very promising concept is presented by Maes and coworkers (56): they found that creatine, the precursor molecule to phosphocreatine (PCr), protects cells from UV damage either by pretreatment or after UV irradiation. The story involves cellular energy, as creatine is neither a filter, nor an antioxidant, but a key molecule in the chemical energy management (ATP, PCr) of the cells. The additional energy reserves afforded by supplementation in the culture medium with creatine allow the repair mechanisms (thymidine dimer excision, for instance) to function more efficiently, thus protecting the cells against apoptosis and further damage. These authors confirm the beneficial effects of creatine in a clinical study where they show that the number of UVinduced sunburnt cells is diminished by topical application of creatine. An intriguing study from back in 1978 shows that caffeine and theophylline protect mice ears from UV-induced tumors (57). Knowing that these molecules stimulate the pool of cyclic AMP (an essential ingredient in the cellular processes of both melanogenesis and lipolysis), their use in sunscreens has been promoted in Sun Active Body Refiner (SPF 8) by Lancaster/Coty in a recent launch. Treatment of UV-Induced Age Symptoms As mentioned in the introduction, “reversing” some of the signs of aging is of course also considered “antiage” activity. Is it realistic? Can anything but retinoic acid reduce some of the wrinkles, the sagging skin, the dryness, and loss of tonus that comes with (photo)aging? And even if so, does it make sense to include these antiaging actives in sunscreens? Apart from price considerations in the highly competitive market, the relatively seasonal aspect of sunscreen use and the relatively short contact times (when compared with “standard” skin care products) would cast doubt on the proposition. Whatever the theoretical considerations say, the market has already acted and begun to introduce sunscreens that contain various actives with some type of antiage and repair claims. It is not for us to judge the scientific validity, but to describe possible concepts and ideas that may be useful to the marketeer, if sufficiently documented by experimental evidence. Once more, it is not possible to review here the enormous mass of antiage and wrinkle repair ingredients of synthetic, marine, botanical, or biotechnological origin proposed on the market, which all might be considered, based on their merit, for inclusion in sunscreens. We shall examine two major aspects—barrier repair of the skin surface and tissue repair in the deeper layers—and discuss some actives that appear to have clearly perceivable, demonstrated benefits. Contrary to the “prevention type” products discussed above, the interaction between the repair active and the Antiaging Actives in Sunscreens 685 sunscreen and/or the UV irradiation is not compulsory. We shall simply review the “repair” aspect as a possibility to boost sunscreen marketing appeal, an added, but logical, benefit to the use of these products. Barrier Repair Scanning the literature on the relationship between skin barrier and UV irradiation, one realizes quickly that the subject is more complex than expected. First: definitions. For our purpose here, we limit the terms barrier, barrier function, and barrier repair to the epidermis, essentially to the stratum corneum (SC) where ceramides, cholesterol, and corneocytes constitute the cutaneous barrier. Although this is purely arbitrary—and not necessarily consistent with my general view of barrier repair—it is convenient and simple for the purpose at hand. On one hand, UV-B irradiation stimulates barrier synthesis: the epidermis thickens, ceramide synthesis is increased, involucrine (a distinct marker protein of cell differentiation and cornification) increases (58 –60). On the other hand, this seems to be a transient effect, an immediate reaction of the skin to the danger of UV rays. Long-term effects of UV exposure clearly lead, especially in old age, to a diminished barrier function (61,62); all systems of the skin suffer through photoaging, and so does the capacity to repair the important structure that is called stratum corneum: enzymes necessary for the process are fewer in number and less active, lipids are peroxidized, the skin is thinner, and the normal desquamation process is altered. When should barrier enhancement actives be used in a sunscreen? Only for “mature” skin? Starting when? Or as a preventive (again?) measure, right from the start, even on young skin? Too few in vivo studies are available to form a clear prescription. A few ideas may help in making one’s own decision on the type of “barrier function antiage” active to use in sunscreens. Hydroxy acids: Lactic acid, one of the most widely used actives in skin care, is known to stimulate many processes in skin, in particular the proliferation of keratinocytes and barrier repair. A 4-week in vivo study by Rawlings et al. (63) showed that L -lactic acid increases ceramide synthesis by 38% over baseline. This is confirmed by a similar study using TEWL as a measure of barrier repair. Rendl et al. (64) investigated more immediate effects in a model of human skin (reconstituted epidermis) and found that lactic acid in a cream increased growth factors (VEGF), and decreased angiogenin secretion. They conclude that the regulation of keratinocyte growth factors and cytokines by AHA may explain some of the therapeutic effects observed in treating photoaged skin with lactic acid. Scott (65) reviewed a large number of AHA and BHA containing preparations and found only a few of them active on photoaged skin. Glycolic acid, found in fruit and milk sugars is described as a cosmetic ingredient with photoprotective activity. Hong et al. (66) describe its inhibition of UV-induced skin tumorigenesis in hairless mice and investigate some of the 686 Lintner complex mechanisms involved. However, a more recent study of 2003 by Kaidbey et al. (67) suggests that AHAs can increase the sensitivity of the skin to UV light. After pretreatment for 4 weeks (24 applications of a 10% glycolic acid product or placebo on the back of 29 Caucasian subjects) the skin was irradiated with 1.5 MED. They observed increased sunburn-cell induction and lowered MEDs and conclude that 10% glycolic acid sensitizes the skin to the damaging effects of UV light. Thus, clearly more systematic studies are needed to determine the benefits of hydroxy acids in sunscreens for antiaging purposes. Ceramides: The large family of complex lipids called ceramides needs no review here. They are the essential element in the cement of cell cohesion of the stratum corneum; long-chain lipids, highly insoluble, ceramides are not so much “biologically active” as structurally important. There is thus less possible controversy about their use in sunscreens. In view of the outdoor activities that go with the use of sunscreens, the abrasion, frequent bathing and the sun exposure, it seems reasonable to use ceramides or ceramide promoting actives in the formula. Any barrier repair contribution will be beneficial to the skin. The improvement of barrier function by ceramides in general has been described in numerous papers (68 and references therein), rarely though in conjunction with UV irradiation. Various studies report the effects of substances that stimulate keratinocyte differentiation and ceramide synthesis: niacinamide (vitamin B3) (69,70), avocadofuran (71), vitamin C (72), calcium (73), mevalonic acid (74), ursolic acid liposomes (75), and others. The T. thermophilus ferment described in the previous section (52,53) has one additional antiaging benefit. Not only does it contain the heat-stable anti-oxidant SOD and catalase-like enzymes to protect the skin against the heat and the free radicals, it also turns out to stimulate keratinocyte differentiation, involucrin synthesis, and barrier repair by increased ceramide and cholesterol production. In vivo, this translates to greater resistance of the skin barrier against aggression and to better moisture retention, both important antiage concepts (76). Tissue Repair The most important antiage activity, from a cosmetic point of view, is to reduce wrinkles. Wrinkles are of course, as we have said at the outset, a major, visible, consequence of the actinic damages sunlight, rather excessive sunlight, generates in the skin. Does it therefore make biological, physiological, scientific sense, to include antiwrinkle actives in sunscreens? Similar arguments as those used for barrier repair actives in sunscreens hold here, too. Sunscreens, if properly used, are in contact with the exposed skin for quite some time. During the outdoor activities that incite the consumer to use a sunscreen, she or he will hardly use other skin care products. But the benefits of truly active tissue repair, antiage molecules lie in extended use, regular exposure to their action, and constancy. Antiaging Actives in Sunscreens 687 In the way a good modern skin care (face care) product should offer at least some SPF—even if not positioned as a sunscreen (cf. Introduction), in the same way a sunscreen may offer tissue repair ingredients to bridge the periods between morning face preparation and the night cream. Two major categories of antiage ingredients are presently of great interest, that follow the wave of hydroxy acids and retinoids discussed earlier: the isoflavones (phytohormones), which are proposed as plant-derived “(pseudo)substitutes” of estrogen, for mature (i.e., postmenopausal) skin, and the matrikines, natural protein fragments with specific, tissue repairing activity which are wound healing research inspired new cosmetic ingredients. We shall limit our discussion mainly to those two fields. Isoflavones: Although there appears to be an impressive amount of literature on the benefits of isoflavones (genistein, daidzein, puerarin, biochanin A, and others), more of it is again concentrated on demonstrating the antioxidant (and thus protective) effects of these molecules (extracted most often from soy, sometimes from red clover or more exotic plants) than on their wound-healing and tissue repair activities. At least, this is true with respect to peer-reviewed published studies. But hundreds of references to the stimulating and repair activating properties of these molecules—in pure form or presented as enriched extracts— are nevertheless found on websites and in promotional documents. Two recent publications by Widyarini et al. (77) and Kang et al. (78) describe the protective effect of isoflavones against UV-induced inflammation and photoaging. More in the spirit of the present section and of significant interest is the study by Myazaki et al. (79), which shows that topical genistein and daidzein stimulate hyaluronic acid production in human keratinocyte culture and in hairless mice. This is tissue repair of a type that antiaging claims require. Schmid and Zülli (80) have gone further and measured the skin thickness increase by topical application of soy isoflavones in a human, placebo-controlled trial. The most recent document on soy isoflavone activity in human skin is by Kawai (81), demonstrating the ability of these extracts to stimulate collagen synthesis in the skin. Cosmetic research has most certainly produced many more examples and data about the skin repair benefits of isoflavone (“phytoestrogens”); for reasons of intellectual property and fierce competition, only a small portion of this research sees publications in peer-reviewed journals. It can thus only be surmised that the use of these ingredients in sunscreens helps keep the skin in better (i.e., more youthful) condition. Matrikines: A whole new concept in tissue repair, and thus in antiaging strategy, is offered by the discovery of matrikines. The term was coined by Macquart (82) to designate protein fragments (peptides) of small size, which are generated by the gradual hydrolysis of natural, structural proteins in the connective tissue. But not just any breakdown product of proteins will be a matrikine. During wound healing and/or inflammation, proteolytic enzymes break down collagen, elastin, fibronectin, and other structure proteins into smaller 688 Lintner pieces. Certain peptide sequences, thus released, possess mediator (“kinin”) or messenger (ormon1 ¼ “hormone”) function: they act on nearby cells (fibroblasts) to stimulate them into neosynthesis of tissue macromolecules, or to attract them to the damaged tissue site (chemotaxis). Like all mediators or signal molecules, these peptides of specific amino acid sequence, act at very low (nano- to micromolar) concentration, but achieve dramatic effects in the rapid regeneration of tissue. These peptides, derived from the natural sequence of the damaged proteins, are usually in the tri- to hexamer range. Some of these matrikines have found cosmetic use, for which it was necessary to attach a lipophilic, fatty acid chain in order to assure skin diffusion and bioavailability (83). It has thus been shown that a few parts per million of the tripeptide palmitoyl-Gly-His-Lys (a serum protein fragment) is able to stimulate collagen and GAG synthesis in vitro, which translates into skin thickening and antwrinkle effects in vivo. Equally low concentrations of palmitoyl-Val-GlyVal-Ala-Pro-Gly (a fragment of elastin) or palmitoyl-Gly-Gln-Arg-Pro (a fragment of Immunoglobulin IgG) have potent skin repair activities (84,85) that may be used in antiaging compositions. As a concrete example of cosmetic use of this concept, the matrikine peptide palmitoyl-Lys-Thr-Thr-Lys-Ser shall be presented in more detail in the following. Discovered by Katayama et al. (86) during wound healing related research on lung cells, it was investigated for skin applications (87) first in in vitro studies on normal human fibroblasts which demonstrate the extracellular matrix (ECM) stimulation by the palmitoylated peptide: collagen I and GAG are increased over baseline by amounts varying between 50% and 250%. This is confirmed on full thickness skin tissue where 2 ppm of Pal-KTTKS achieve the same result as 1000 ppm of vitamin C and in clinical, vehicle, or benchmark (retinol, moisturizer) controlled studies using the Pal-KTTKS peptide in topical antiwrinkle creams (87 – 89). The use of 3 – 5 ppm of Pal-KTTKS in a topical formula leads to significant, measurable, and consumer perceivable benefits in reducing wrinkle volume, depth, density, and to overall improvement of the facial skin when used for up to 6 months. Skin biopsies that were taken during one of the panel studies (88) on both the Pal-KTTKS treated group and the placebo group, at time points T ¼ 0, T ¼ 2 months, and T ¼ 4 months show that the treated skin has improved collagen IV and elastin fiber assemblies whereas the control group showed no notable changes in the skin samples. Matrikines such as Pal-KTTKS are thus ideal candidates for cosmetic tissue repair. Nothing in the nature, activity, or mechanism of action of these molecules with true antiaging properties prevents them from being used in sunscreens. They are compatible with any kind of formulation, they are stable, their mode of action is unperturbed by UV and outdoors. They are clearly an additional benefit to sunscreens, as they demonstrably can repair some of the actinic photodamage. Antiaging Actives in Sunscreens 689 CONCLUSIONS Some aspects of this chapter may appear somewhat polemic and/or tongue in cheek. Whereas UV absorbing molecules are clearly designated as sun filters and constitute a well-defined category of chemicals, the notion of “antiage” actives (cosmeceuticals?) is much less well characterized, whence the occasional asides. We have tried to demonstrate that the notion of antiaging actives in sunscreens opens many possibilities to the formulator to improve the basic sunscreen products, to add real benefits and to allow for variety in claims and marketing positioning. Prevention of sun damages on the skin can be reinforced by some of the antioxidant and photoprotective agents; treatment of sun damage during or immediately after sun exposure with repair actives is also justified. Teaching the consumer on how to “manage” the sunlight (prevention goes beyond using sunscreens and includes wearing adequate clothes, avoiding the hottest hours of the day, etc.) has become part of the marketeer’s obligation. Depending on the country, however, from the USA to Europe to East Asia, the legislations on sunscreens, claims, and formulations, are quite different and complex. Adding antiage actives to these sunscreens makes the legal situation even more complex with respect to advertised claims. Other chapters in this book address the regulatory aspects of sunscreens per se: the notion of which actives can legally be called “actives” varies even more. 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