Biopharmaceutical Sciences, Biomed Biopharm Res., 2022; 19(2):1-18
Evaluation of the protective and moisturizing effects of cosmetic formulations for the skin containing extracts with film-forming properties
Bruna Penachin1, Letícia Kakuda1, Patrícia Maria Berardo Gonçalves Maia Campos1*
1School of Pharmaceutical Sciences of Ribeirão Preto – University of São Paulo, Avenida do Café, S/N, Ribeirão Preto - SP, 14040-903, Brasil
The development of formulations that provide efficient film-forming properties is important since film formation can protect the skin and hair from the action of external aggressors, such as pollution. Thus, the aim of this study was to evaluate the film-forming effect of cosmetic formulations for the skin containing extracts of Kappaphycus alvarezii and Caesalpinia spinosa, as well as the use of hemisqualane as an alternative to synthetic silicones. The evaluation of efficacy was performed by biophysical techniques and skin imaging techniques, as well as by a sensorial analysis. Sixteen women, aged 18 to 30 years, participated in the study. The results obtained demonstrated that the presence of the extracts in the formulation enabled the formation of a film that helped maintain skin barrier integrity, reduced desquamation, and improved parameters related to hydration. Finally, the study also showed that there were no significant differences between the formulations containing silicones and hemisqualane, which supports the use of hemisqualane as a more sustainable alternative in cosmetic formulations.
Keywords: biopolymer, film-forming effect, hemisqualane, clinical efficacy, cosmetics
Received: 12/10/2022; Accepted: 1/12/2022
For more than a century, the cosmetics industry has been responsible for developing products that have an essential meaning for consumer society, as they are related to how people express themselves (1). This sector does not seem to suffer even in times of crisis and constantly continues searching for innovative projects (1).
During the pandemic, some changes were observed concerning the consumer profile by increasing the demands for cosmetic products with skin care and health care as their primary appeal (2). Therefore, among the proposals for innovative formulations are those that provide a film-forming effect to the skin or the hair, protecting them from the action of aggressive external agents, such as atmospheric pollution (3,4).
Thus, such formulations are interesting from the point of view of innovation and for the potential benefit, they can provide to the skin (5). These products act either by forming a polymeric film (synthetic or natural) on the skin surface or using a residual liquid film, resulting in a thin and transparent layer in the area (4,6).
The search for raw materials is a crucial step in the research and development process of cosmetic products, considering that this industry follows market demand. Consumers worldwide are also attentive to the search for innovative cosmetics formulations concerned with aspects related to product sustainability (7).
This way, some natural ingredients can be applied to provide a film-forming effect to the formulation. The red algae extract (Kappaphycus alvarezii) can be combined with tara extract (from the Caesalpinia spinosa fruit) through a controlled hydrolysis process that results in a natural biopolymer that has the potential to form a non-occlusive, flexible and resistant film on the skin surface (4). This association is rich in polysaccharides and, due to the characteristic of the polymer, presents an expressive potential for the promotion of the film-forming effect with clinical applicability to reduce the skin-pollutant interaction (4).
The cosmetic industry widely uses synthetic silicones due to their versatility in their possible applications. These substances, even in small concentrations, are used as emollient agents that provide important sensorial characteristics (such as improving the spreadability of the formulation and dry touch), and they are also used to facilitate the incorporation of some substances (for example, chemical and physical sunscreens) in cosmetic preparations (8,9). However, even though they provide pleasant sensory characteristics to formulations, silicones remain in the environment for many years and can cause harmful effects to the aquatic environment and human health (7,9,10).
Due to these concerns, alternatives containing natural and biodegradable substances that can replace the functions of synthetic silicones are becoming increasingly important in the cosmetology area (10). Therefore, in addition to polysaccharides, the C13-15 alkane (also known as hemisqualane), a saturated hydrocarbon from sugar cane, is another natural ingredient of interest. As this alkane is obtained from a renewable and biodegradable source, it can function as a natural alternative to synthetic silicones, acting as a light, innovative and multifunctional emollient that confers numerous advantages to improving the spreadability, texture, and sensory properties of formulations (9,10).
Biophysical techniques are clinically relevant to evaluate the film-forming effect of these formulations because they allow the evaluation of the skin noninvasively and in real-time (4). This equipment allows, for example, the evaluation of the stratum corneum integrity, the barrier function of the skin, and microrelief, which are damaged in exposure to pollution, since skin with cutaneous alterations presents high transepidermal water loss and lower hydration values (4,11,12).
Finally, the clinical evaluation of the immediate effect of cosmetics allows the assessment of the action of these products on the skin immediately following their application, such as the action in improving the barrier function or the ability to form a film on the skin surface, which are characteristics proposed by the biopolymer composed of the red algae and tara extracts.
In consideration of all the aspects mentioned above, the aim of the present study was to evaluate the film-forming effect of cosmetic formulations for the skin containing tara (Caesalpinia spinosa) and red algae (Kappaphycus alvarezii) extracts, using biophysical and skin imaging techniques and sensory analysis, as well as to evaluate the use of hemisqualane as an alternative to the synthetic silicones.
Material and methods
Development of formulations
The formulations were developed considering the specifications of the active substance under study, the sensory characteristics, and the interaction with the raw materials used as a vehicle in cosmetic formulations. As presented in Table 1, a gel-cream cosmetic formulation (named P2) was developed based on emulsifier, polymer, emollients, humectants, preservatives, and containing or not (the vehicles) the extracts of tara (Caesalpinia spinosa) and red algae (Kappaphycus alvarezii).
The extracts that were used were obtained commercially (SILAB, France) and have MSDS (Material Safety Data Sheets) as listed by the company responsible for the commercialization of these substances (SILAB, France). Our research group has also previously investigated these extracts (4). Regarding composition, the active substance is composed of a mixture of red algae (Kappaphycus alvarezii extract) and tara (Caesalpinia spinosa fruit extract).
Thus, the formulations received the following names for identification purposes: gel-cream with hemisqualane vehicle (P1), gel-cream with hemisqualane and the active substance (P2), gel-cream with silicones vehicle (PS1), and gel-cream with silicones and the active substance (PS2).
Stability study and visual evaluation of formulations
The tests regarding the stability of the formulations followed the guidelines described in ANVISA’s (National Health Surveillance Agency) Stability Guide for Cosmetic Products, in Brazil.
Preliminary tests of centrifugation and analysis of organoleptic characteristics were performed daily for 28 days. For this purpose, the formulations were stored in plastic and opaque recipients under controlled temperatures (37ºC and 45ºC) and humidity (70% RH) conditions. After the preliminary stability tests, the formulations were submitted to accelerated stability studies by rheological behavior, which was performed every 7 days during a 28-day period (13,14).
The pH value of the formulations was verified by a 10% (w/w) aqueous dispersion of each formulation in Digimed DM20 (15,16). The centrifugation test was performed with 3 grams of each formulation developed, centrifuging at 3000 rpm for 30 minutes at room temperature (25 °C) in a centrifuge (Excelsa® Baby II, model 206-R, power 0.0440, Fanem, Guarulhos, SP, Brazil) (17).
Samples were stored at room temperature (≈ 25 °C), 37 ºC, and 45 ºC for 28 days, the samples were visually observed daily for the following changes: color, smell, phase separation, and homogeneity (18).
Study of the rheological behavior
A Brookfield DV3T (Brookfield, Middleboro, MA USA) cone and plate type rheometer equipped with a CP-51 spindle coupled to the RHEOCALCT® software was used for the rheology study. For each analysis, 0.50 and the speed rotation was increased progressively from 0 to 10 rpm, with 2 seconds between each speed, resulting in a curve composed of “shear rate” versus “shear stress”. A second, descending, curve was then generated with the inverse decrease of speed (16).
To characterize rheological behavior, after preparation the formulations were kept at approximately 25ºC for 24 hours, and then the parameters of flow index, consistency, minimum apparent viscosity, and thixotropy were evaluated (14,16).
Analysis of the texture profile and spreadability
The texture profile analysis and spreadability were performed using the TA XT Plus® (Stable Microsystems, Surrey, United Kingdom), coupled with the Exponent software. The method is based on the insertion of an analytical probe into the sample, with a previously defined speed and deepness. To evaluate the spreadability, the shear work test was conducted using the TTC Spreadability Rig (HDP/SR) (16). Then to evaluate the other properties, the texture test was performed where the probe used was the Back Extrusion rig (A/BE) (16,20). Both analyses were done in triplicate.
From the resulting graphic of force (N) by time (t) were obtained the following parameters: cohesiveness, consistency, firmness, viscosity index, and spreadability (16,20,21).
The sensorial analysis was performed according to the protocol established by the Ethics Committee in Research Involving Human Being from the School of Pharmaceutical Sciences of Ribeirão Preto from the University of São Paulo (CEP: CAAE 58730816.5.0000.5403). To this end, 16 participants aged between 18 and 30 years were recruited. To each, 42 μL of the formulations was applied to a region area corresponding to 4x5 cm² and delimited on the forearm, making twenty circular movements during the application (23). The areas were randomly defined. The first region received the vehicle formulation containing hemisqualane.The vehicle with the silicones was applied to the second region. The third region received the formulation containing hemisqualane and the extracts. Finally, the formulation containing the silicones and the active substance was applied in the fourth region.
After application, formulations were evaluated by means of a questionnaire, where each participant evaluated the aspects related to spreadability, soft touch, and the touch sensation. After 5 minutes, the volunteers also evaluated the dry touch, tackiness, white residue, and hydration. The participants evaluated each statement by agreeing, disagreeing, or partially agreeing with the statements regarding the parameters mentioned before. Finally, the participants were asked about their preference for the formulation.
Short-term clinical efficacy study
To identify the immediate effects, such as film formation, the water content of the stratum corneum, the transepidermal water loss (TEWL), and the skin microrelief were evaluated using Corneometer® CM 82, Tewameter® TM 210 and Visioscan® VC 98 (all Courage & Khazaka, Cologne, Germany) equipment respectively. The parameters described above were measured prior to (with basal values) and two hours following the application of the formulations in their respective regions.
The Corneometer® CM 825 was used to determine the water content of the stratum corneum. This equipment is based on the principle of measuring electrical capacitance. The results were provided in arbitrary units (AU) where 1 AU is estimated to correspond to 0.2 - 0.9 mg of water per gram of stratum corneum (24).
To determine the transepidermal water loss from the skin, the Tewameter® TM 210 was used, coupled with software to measure the transepidermal water loss based on the diffusion principle described by Adolf Fick. The probe remained on the skin for two minutes in the region of the forearm that we evaluated. The average value of three measurements was used in the subsequent calculations (4). The values are given in g.m2.h-1.
The Visioscan® VC 98 was used to determine the cutaneous microrelief, providing qualitative and quantitative information on the skin surface in physiological conditions through optical profilometry techniques using a digitalization image process obtained by a video camera. Among the parameters related to the skin surface (SELS - Surface Evaluation of Living Skin), the Sesc (related to skin desquamation) was evaluated.
Statistical analysis and presentation of results
The experimental data obtained from the analysis were submitted to statistical analysis using Prism8 and Origin 8 software. To evaluate normality, the Shapiro-Wilk test was performed. When the curve obtained was normal, the one-way ANOVA test with Tukey's post-test was applied. The Kruskal-Wallis test with Dunn’s post-test was applied when the curve was not normal. For the results, p-values less than or equal to 0.05 were considered significant. Finally, the calculation of the average, standard deviation, and coefficient of variation was performed to present the results as tables, graphs, and figures with the discussion based on the literature data.
Stability study and visual evaluation of formulations
After preparation, the four formulations presented pH values compatible with the skin's pH (25,26,27), in a range where the pH presented variations between 4.5 and 5.5. After the centrifugation cycles, no phase separation was observed in the formulations under study, which remained homogeneous in appearance and were considered stable. Finally, in these conditions and after the 28-day period, no visual changes were observed in the formulations in relation to color, smell, phase separation, and homogeneity (18).
Study of the rheological behavior
The formulations containing the active substance (P2 and PS2) or not (P1 and PS1) presented a non-Newtonian profile, pseudoplastic (with flow index less than 1), and thixotropic behavior (28,29). Regarding the hysteresis area, P2 and PS2 presented significantly lower values compared to those obtained for their respective vehicles, P1, and PS1 (Figure 1).
In relation to the formulations containing the natural extracts, it was observed that for P2 and PS2 there is a significant increase in the consistency index and minimum apparent viscosity (VMA) compared to P1 and PS1. There was no significant difference in the flow index between the preparations (Figure 2).
Analysis of the texture profile and spreadability
For the formulations containing the active substance, P2, and PS2, it is observed that for both the consistency and firmness parameters, there is a significant decrease (p<0.05) in the values when compared to their vehicles (Figure 3). The same decrease can be observed when the cohesiveness and viscosity index parameters are compared between the preparations containing the studied extracts and the vehicles (Figure 3). Furthermore, there were no significant differences (p<0.05) in the shear work values of the formulations containing the active substances.
In relation to the sensory evaluation noticed immediately following the application of the formulations (Figure 4), P2 presented high acceptability in terms of the spreadability of the formulation and the other preparations. Even so, for P1 and PS1, all volunteers agreed on good spreadability.
As for the velvety/satin touch, more volunteers agreed that P2 conferred this characteristic compared to the results obtained by the other formulations. Regarding the softness, PS2 conferred a better result among the preparations.
In the sensory analysis performed 5 minutes following the application of the formulations, it was observed that for the parameters of hydration and stickiness, P1 and P2 showed similar results, and were better than those of the PS1 and PS2 formulations (Figure 5). The data regarding the dry feel of the formulation indicates that P2 shows a better result than the others and has slightly higher acceptability than P1. As for the formation of white residue, the volunteers agreed with the statement, "the formulation doesn’t leave a white residue on the skin" for all the formulations.
When asked about their preference for one of the formulations, 87.5% of the participants preferred the formulations containing hemisqualane, the natural alternative to the use of synthetic silicones. From these, 43.75% chose formulation P2.
Short-term clinical efficacy study
In relation to the aqueous content of the stratum corneum, all formulations showed a significant increase compared to the initial time and 2 hours after application (Figure 6).
Regarding the skin peeling results, the preparations containing the active substance (P2 and PS2) showed a significant (p<0.05) reduction of the parameter evaluated in relation to the baseline time when compared to the vehicle results (Figure 7).
For the results obtained regarding transepidermal water loss (Figure 8), the formulations containing the extracts (P2 and PS2) showed significantly (p<0.05) lower values when compared to the basal values.
The choice for this type of gel-cream formulation occurred based on studies previously developed within our research group (4). Thus, the formulation was based on carbomer (viscosity agent/rheological modifier), EDTA Disodium (chelating agent) and glycerin (wetting agent) and Nikkomulese 41® (Nikkol, Japan) was used as emulsifying base. In addition, Naticide® (Sinerga, Italy) was used as a natural preservative and butylene glycol as a wetting agent.
In analyzing the data related to rheological behavior, the lower values observed for P2 and PS2 are essential because, for thixotropic fluids, the lower these values, the less the fluids' rheological behavior will depend on time. With this, there is a faster viscosity recovery after applying a shear force (14,28,30). Therefore, it is crucial to observe the rheological behavior of a formulation where the film forming ability is evaluated because film formation significantly influences the preparation's flow properties (28,30).
Thus, the addition of the active substance resulted in a decrease in thixotropy, indicating a rapid recovery of the film structure after the application of a force because the result showed a significant but not extreme decrease, allowing there to be a quick and sufficient recovery of the film structure after the application of force, which may be, for example, interesting after the force applied during a topical administration of the formulation (14,30).
The texture profile analysis was used as an important tool to evaluate the mechanical properties of the formulations to predict their sensory characteristics and also used to correlate the data obtained with those collected during the evaluation performed by the volunteers; once the sensory is a question that directly influences the acceptability of cosmetic preparations (16,31,32). Thus, as there were no significant differences in the values of the shear work and the flow index of the formulations containing the active substances compared to the others, it may be an indication that the addition of the extracts did not compromise the spreadability of the formulation, since both parameters are closely related to this characteristic (16,32).
In relation to cohesiveness, a parameter related to the ability to form a polymeric structure due to the union of the formulation particles, we can relate it to the tackiness of the preparation (21). Therefore, it is observed that the more cohesive the formulation, the greater will be the tackiness (21). Thus, when these results are analyzed, it is observed that the addition of the active substance caused the cohesiveness to decrease significantly compared to their respective vehicles (P1 and PS1). These results support what was perceived in the sensory analysis for P2 but not for PS2, which was considered sticky after 5 minutes of application.
The sensory analysis was performed with an untrained group. This choice is important because it simulates the consumer's opinion and can evaluate the volunteers' perception of the sensory characteristics of the formulations after application (33). This step was essential to the study because consumers tend not to adhere to the use of cosmetic preparations that do not present a pleasant sensorial, even if they contain appeals considered attractive concerning the formulation (32). In the results observed immediately after application, the formulations showed similar results for the parameters spreadability and softness sensation, with a slightly better result for P2 concerning the smooth touch evaluation.
However, for the evaluation made 5 minutes after the application, it is possible to observe a significant difference in the volunteers' perception of the formulations through the collected results. In the volunteers' perception, P1 and P2 were considered more hydrating and showed better results about perceived stickiness. Thus, reflecting the results, there was a preference of 87.5% of the participants for the formulations containing the natural alternative to the use of synthetic silicones, and of these, 43.75% chose the P2 formulation.
The evaluation of the sensory properties regarding the hydration sensation promoted by the four formulations corroborates the results observed in the immediate clinical study, where all preparations showed an increase in the water content of the stratum corneum two hours after application, which may demonstrate that they all accomplished their hydration role, despite the presence of the extracts (4,5,34). As the sensory analysis performed was qualitative, the perceived hydration is possibly due to the presence of raw materials with hydration and wetting capacity in the vehicles, as is the case of glycerin and butylene glycol (5).
For the evaluation of skin desquamation (Sesc), the lower the values compared to the baseline results the more hydration is imparted to the skin (4). Thus, it is observed that there was a significant decrease in P2 and PS2 values demonstrating that the presence of the active substance resulted in formulations with the ability to provide hydration, improving desquamation (4). In this case, possibly with the addition of the active substance the desired effects about film formation are a consequence of the composition of the emulsion since this appears to interfere directly with the polymeric structure of the carbomer present in the formulation (4).
In relation to the transepidermal water loss, the formulations containing the active substance showed a significant decrease in the results, when compared to the basal values. These results show the ability of the biopolymer to form an efficient film, since the film formed decreases the evaporation of superficial water from the skin, reducing desquamation, protecting against external aggressors, and helping to protect the barrier function of the skin (4,34).
The extracts under study are a polymer obtained commercially through the controlled hydrolysis of sugars (without the use of catalysts or chemical additives), which allows the selection of polysaccharides with well-defined structures. In this process, galactomannans are derived from Caesalpinia spinosa and sulfated galactans are from Kappaphycus alvarezii. The interpolymerization of these sugars occurs, which will determine the resistance of the film formed. This way, obtaining the active substance presents the technology of forming a network of interpenetrated natural polymers, forming a dense and cohesive mesh with strong biomechanical properties, as described in the technical report from SILAB (France) (4). Thus, the presence of the extracts in the formulation, because of the formation of an efficient film due to the polymeric structure, helped maintain skin integrity as it was effective in reducing desquamation, and improving parameters related to hydration and skin barrier integrity (4,34).
The formulation added to the red algae extract (Kappaphycus alvarezii) and tara (Caesalpinia spinosa) showed a reduction in the thixotropy rheological parameter, which is directly related to the rapid recovery of viscosity after shear, a characteristic related to film-forming properties. Furthermore, the analysis of sensorial properties showed that there are no differences between synthetic silicones and hemisqualane, which allows the use of hemisqualane as a more sustainable alternative to provide adequate sensorial properties to cosmetic formulations.
Therefore, the addition of the extract to the formulation promoted increased hydration and decreased skin desquamation and transepidermal water loss within 2 hours of application. Finally, the tara and red algae extract showed properties that correspond to the film-forming, which added benefits to the formulation such as improving the barrier function of the skin and may promote a protective effect against external damage to the skin.
The author would like to thank the study group NEACTEC (Núcleo de Estudos Avançados em Tecnologia de Cosméticos), FCFRP (School of Pharmaceutical Sciences of Ribeirão Preto - University of São Paulo) and the participants of clinical study.
Authors Contribution Statement
BP was responsible for the experimental part, collection and analysis of data and writing the article; LK was responsible for the experimental part, collection and evaluation of results and review of the writing; PMC coordinated the study, including the conceptual design, supervision, and was responsible for the final revision of the article.
Conflicts of interest
The authors of the study declare that they have no financial and/or personal relationships that could represent any potential conflict of interest.
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