Application of tetra-isopalmitoyl ascorbic acid in cosmetic formulations: Stability studies and in vivo efficacy

Abstract

Liposoluble vitamin C derivatives, such as tetra-isopalmitoyl ascorbic acid (IPAA), are often used in dermocosmetic products due to their higher stability than vitamin C free form as well as its proposed effects in skin; however, there are no studies analyzing IPAA stability or its in vivo effects when present in dermocosmetic formulations. Thus, this study aimed to evaluate chemical stability and pre-clinical and clinical efficacy of dermocosmetic formulations containing IPAA in skin hydration and microrelief. Chemical stability of the formulations added with 1% IPAA was evaluated by heat stress during 35 days by HPLC. For pre-clinical evaluation, experimental formulations were topically applied on hairless skin mice during 5 days and animal skins were analyzed by non-invasive biophysic techniques (water content of stratum corneum, TEWL, viscoelasticity, and microrelief) and by histopathological studies. For clinical efficacy tests, the formulations were topically applied to the forearm and face of human volunteers, and 3 h and 15 days after applications, the skins were evaluated by the same non-invasive techniques mentioned before. Results showed that formulations containing IPAA had medium stability and had pronounced moisturizing effects on stratum corneum and on viable epidermis. These formulations also improved skin microrelief especially in relation to skin smoothness and roughness.

Introduction

The use of l-ascorbic acid (vitamin C) for topical application is not a new procedure. It has been used in pharmaceutical and cosmetic preparations for a long time on the basis of its many favorable effects on the skin [1]. However, free vitamin C is unstable, being easily oxidized and inactivated when exposed to air, and also it cannot penetrate into the skin because of its hydrophilicity [2].

Some marketed products containing unstable free vitamin forms need great investments in specific methods of packaging and containers and have short shelf-lives. Therefore, more stable and safer new delivery systems, such as microemulsions, microcapsules, nanospheres, liposomes [3] as well as the use of ester derivatives such as magnesium ascorbyl phosphate and sodium ascorbyl phosphate (water-soluble derivatives) and ascorbyl-6-palmitate and tetra-isopalmitoyl ascorbic acid (IPAA) (liposoluble derivatives) are an attempt to prolong formulation stability and are increasingly substituting the use of vitamin C free form [4], [5]. Moreover, liposoluble derivatives such as IPAA are able to better penetrate the skin [1] and have increased stability when compared to vitamin C free form [6].

However, formulations supplemented with vitamins, even in the ester form, may present low shelf-life [7]. It is well known that the stability of all vitamin C derivatives is influenced also by structural properties of the formulations [1], [8]. Thus, in the development of a cosmetic formulation, it is necessary to choose suitable vehicle raw materials because they must be compatible with the active substances selected to attend the indication for use of the product [9].

Cosmetic formulations containing IPAA have to be submitted to studies of chemical stability in order to evaluate their quality, since the integrity of the vitamin derivative must be kept constant during the formulation shelf-life.

Stability prediction is usually performed by accelerated storage conditions [7] such as temperature variation to induce rapid chemical alterations in formulations, which are usually detected by quantification of some components over time. Thus, in this study, a High Performance Liquid Chromatography (HPLC) method was developed to predict the chemical stability of formulations containing a liposoluble ester derivative of vitamin C [10].

Vitamin C, an essential skin component, has an important role in collagen production, being a co-factor in proline and lysine synthesis [11] and participates in connective tissue repair and cicatrization. The vitamin also captures free radicals in the skin and eliminates singlet oxygen species, superoxide, hydroxyl and peroxyl groups, and hypochlorous acid [2], [8], [12], thus preventing UV-induced damages and improving photo-aged skin [13]. However, on exposition to UV radiation, the levels of the antioxidant are sharply reduced [14].

Some studies are conducted to evaluate whether ascorbic acid properties may be extrapolated to its derivatives [12], [15]. Maia Campos et al. [16] observed that liposoluble tetra-isopalmitoyl ascorbic acid (IPAA) and hydrosoluble magnesium ascorbyl phosphate were efficient in neutralizing free radicals and that the latter could have a hydration effect in the deeper epidermis layers.

IPAA is the first lipoidic-liquiform vitamin C derivative by itself and has been one of the frequently used ascorbic acid derivatives in cosmetic c formulations [6]. Ochiai et al. [17] demonstrated 84% liberation of vitamin C by the derivative in reconstructed skin. Studying the effects of IPAA on keratinocyte cultures, the authors demonstrated reduction of oxidative stress and of interleucin-1 and prostaglandin E₂ production induced by UVB. Secretion of melanocyte proliferation factors was also suppressed, indicating that the hyperpigmentation effect induced by UVB may be alleviated.

Application of IPAA may also alleviate skin damage following physiologic doses of UVA irradiation and should be adopted in the preventative and protective strategies against UVA injuries [6].

IPAA, which has already been widely used as an additive in cosmetic formulations [6], has also been approved as “quasi drugs” by Japanese authorities. These “quasi drugs” are only approved based on equivalency evaluation and examination conducted by the Pharmaceutical and Medical Device Agency and thus they are products with proven efficacy in a specific claim category, in this case for skin whitening, recognized by the Ministry of Health and Welfare [18].

Considering that IPAA is a liposoluble vitamin C derivative more stable than the free form, it could have vitamin C skin effects and also provide other skin benefits such as hydration, smoothness, and reduction of wrinkles. However, it still needs to be analyzed by in vivo studies, mainly clinical studies, which are necessary to confirm the effects of formulations added with IPAA on human skin in actual conditions of use since most studies involving the derivative were performed in vitro.

Regarding in vivo efficacy testing, histopathological and histometrical studies determine epithelial thickness and the number of cell layers in the viable epidermis and are able to show alterations produced on the skin by a specific treatment and may have a role in the understanding of the effects of topical substances [19]. Since these are invasive techniques, hairless mouse could be an appropriate model for preliminary studies on anti-aging properties of topical formulations.

On the other hand, modern non-invasive skin biophysical techniques are often used since they allow evaluation of cosmetic products under actual conditions of use [16]. The skin biophysical methods typically measure selected properties that depend on the measuring principle. Hydration, skin viscoelasticity and microrelief, barrier function, and skin thickness are examples of parameters that can be quantified non-invasively using these techniques [16], [20].

Thus, the aim of this study was to evaluate chemical stability and pre-clinical and clinical efficacy of dermocosmetic formulations containing IPAA by investigating possible alterations caused by these formulations in the epidermis. There are not studies analyzing the stability of this liposoluble vitamin C derivative when vehiculated in topical formulations and their efficacy by pre-clinical and clinical parameters, mainly using biopsies and objective measurements, which are an important tools in efficacy studies.