Development of lamellar gel phase emulsion containing marigold oil (Calendula officinalis) as a potential modern wound dressing
Graphical abstract
Introduction
Chronic wounds to the skin present a serious public health risk and are painful, unsightly, and can require limb amputation if left unattended. Adequate treatments are needed to increase the overall patient quality of life. Products designed for the treatment of chronic wounds and/or skin ulcers require optimizing properties such as the ease of application, increased patient comfort, and the ability to retain adequate moisture in the bed of the wound (Popovich et al., 2010).
Calendula officinalis (Asteraceae) L., or marigold, is an herbal plant that has been used to treat wounds since the 13th century in Europe (Parente et al., 2012), and nowadays is used virtually worldwide. This plant possesses several biological properties such as antimicrobial (Faria et al., 2011), antimetastatic (Preethi et al., 2010), and antiparasitic activity (Szakiel et al., 2008). Its main activity as an anti-inflammatory constituent (Parente et al., 2012) which was observed in animal models (Preethi and Kuttan, 2009) and clinical tests, justifies its utilization as a cosmetic and personal care product for wound treatment (Grimme and Augustin, 1999). Secondary metabolites such as flavonoids, tannins, saponins, terpenoids, coumarins and others (Santos et al., 2006, Schmidt et al., 2009) alone or in association are directly associated with these effects. Also, marigold oil (C. officinalis) could be used to treat thermal burn injury, acute dermatits (Pommier et al., 2004) during irradiation for breast cancer and skin rashes (Parente et al., 2012). C. officinalis extract is present in almost 200 cosmetic formulations and significant experimental wound healing studies or treatments have been performed using only C. officinalis extract or cream containing the extract (Chandran and Kuttan, 2008).
The choice of formulation type for the treatment or care of the skin is important because it may affect the mode of active compounds distribution in its surface. Lamellar gel phase (LGP) or liquid crystals formulations (emulsions) can be obtained due to the bi-layer arrangement (separated by water layers) of the surfactant molecules in the interfacial film. The LGP emulsions provide benefits such as enhanced stability and incorporation of active components in the matrix of (LGP) phase, water retention and the controlled release of active ingredients (Kudla et al., 2010). LGP could thus be used for wound healing purposes due to its adequate viscosity and because of the probability that it might increase the residence time of the formulation on the wound surface.
In the present study, we optimized the preparation of LGP emulsions using the ternary and pseudo phase’s ternary diagram with different proportions of C. officinalis oil, distilled water, and a mixed emulsifier (cetyl alcohol 2 polyoxyethylene and stearyl alcohol 2 polyoxyethylene) whose HLB values were 6.0. This work is based on previous studies by Santos et al. (2006). The novel system was designed with the aim of achieving an efficient lamellar gel phase emulsion with wound healing activity. Santos et al. (2006) did not use the ternary and pseudo phase’s ternary diagram to select the best emulsion. In addition to this, the previous formulations using C. officinalis oil were not stable. Here, we have presented a detailed physicochemical characterization (preliminary tests) of samples to select the best LGP emulsion (i.e. good amount of anisotropic structure and stability). The selected LGP formulation was analyzed according to the rheological behavior as well as the intrinsic and accelerated physical stability (pH, apparent viscosity (η), and conductivity values) in different temperatures. In addition, in vitro tests for cytotoxic activity toward L929 cells and in vivo studies on experimental wound healing in rats were performed. Thus, the present study aimed to compare LGP emulsions to simple emulsions (both containing C. officinalis oil) in the wound healing profile.
Section snippets
Material
C. officinalis (calendula oil or marigold oil) from flowers were supplied by Beraca (Beraca Ingredients, Sao Paulo, Brazil). The information and specifications about the marigold oil can be found at the European Pharmacopeia (CAS No.: 84776-23-8, 70892-20-5; EINECS No.: 283-949-5). The lipophilic (cetyl alcohol 2 polyoxyethylen, named Ceteth-2, HLB = 5.3) and hydrophilic surfactants (stearyl alcohol 2 polyoxyethylene, named Steareth-20 HLB = 15.4) were kindly provided by Oxiteno (Sao Paulo, Brazil).
Pre-formulation of LGP emulsions and development of ternary and pseudo phase’s ternary diagram
A ternary phase diagram was constructed to represent the formulations with the percentage of C. officinalis oil, distilled water, and surfactants (Ceteth-2 and Steareth-20). The weight ratio of surfactants (Ceteth-2/Steareth-20 = 0.93/0.07) was chosen to achieve an HLB value of 6.0. These surfactants were chosen due to their low toxicity with the skin and low incompatibility regarding other excipients (Santos and da Rocha-Filho, 2007). In addition, it has been suggested that the anisotropic
Conclusions
The LGP emulsion (15.0% marigold oil; 10.0% of blend surfactants and 75.0% of purified water [w/w/w]) has demonstrated greater stability and its low spread ability suggests longer contact with wounds. The C. officinalis oil was not detrimental and also was non-cytotoxic as assayed in L929 cells. The LGP emulsion was relatively stable and promoted better quality wound healing in a rat skin wound model than the other groups. LGP emulsion seemed to modulate the inflammatory phase of wound healing.
Conflict of interest
The authors declare that they have no competing interests
Acknowledgments
This work was supported by CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – Brazil), CNPq and FAPESP (Fundação de Amparo à Pesquisa do Estado de São Paulo – Brazil).
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