Abstract Sunscreen is an important product for protecting the skin from the harmful effects of the sun's UV rays. Ethanol extract of watery rose apple leaf (WRAL) contains flavonoids, which acts as natural UV filters. Titanium dioxide (TD), an inorganic filter, provides broad spectrum protection. This study aims to develop a sunscreen cream by combining WRAL extract and TD, and to investigate the synergistic efficacy based on the SPF value. This study began with characterizing the total flavonoid content of the WRAL extract. Five formulations were prepared using either single or combined UV filters at varying ratios. These formulations results were evaluated for physical properties and SPF value determined in vitro. The optimum formula underwent accelerated stability, skin hydration and irritation tests. The WRAL extract contains a total flavonoids content of 46.33 ± 1.02 mg QE/g extract. Then, F3 cream, containing 10% WRAL extract and 6% TD, was found to be the optimum formula. The results showed that the cream met quality requirements, with a pH of 6.12 ± 0.01, spreadability of 5.5 ± 0.15 cm, viscosity of 60 ± 10 dPas, and SPF value of 40.22 ± 0. The cream was physically stable at 25°C and under heating-cooling conditions during storage, improved skin hydration, did not cause skin irritation and the most preferred formula. Thus, the cream preparation containing WRAL extract as a natural UV filter and titanium dioxide as an inorganic UV filter is a promising sunscreen formulation, offering new opportunities for the development of skin care products.

Keywords: SPF value, Sunscreen, Syzygium aqueum, Titanium dioxide, UV filter

Citation:  Setiowati, A.D., Sari, T.A., and Muadifah, A. 2026. The synergistic efficacy of natural and inorganic UV filter combinations on the SPF value of sunscreen cream preparations. Natural and Life Sciences Communications. 25(2): e2026029.

Graphical Abstract:

INTRODUCTION

Based on wavelength, there are three categories of solar ultraviolet (UV) radiation: UVA (315-400 nm), UVB (280-315 nm), and UVC (100-280 nm). UVC rays are scattered and absorbed by the ozone layer, preventing them from reaching the earth's surface, while about 90-99% of UVA rays and 1-10% of UVB rays reach the surface (Li et al., 2023). Repeated exposure to UV rays can lead to changes in skin structure. Short-term repeated exposure can cause erythema, or sunburn, while long-term exposure can lead to a loss of skin elasticity and may result in skin cancer (Geoffrey et al., 2019).

Sunscreen is essential for shielding the skin from the damaging effects of ultraviolet radiation. The effectiveness of sunscreen is measured by the sun protection factor (SPF), which indicates protection levels against UVB rays. The protection level increases as the SPF value rises (Rajasekar et al., 2024). UV filters are categorized as either organic or inorganic. Organic filters, such as oxybenzone, avobenzone, meradimate, and octocrylene, absorb UV radiation, whereas inorganic filters, such as titanium dioxide and zinc oxide, reflect and scatter it (Jesus et al., 2022).

Organic UV filters can trigger allergic reactions, and their combinations are limited due to photoinstability and the possibility of unfavorable synergistic interactions (Gilbert et al., 2013). In animal studies, these organic filters were shown to have detrimental effects on hormone levels (Schneider and Lim, 2019). In contrast, inorganic UV filters are widely used because they do not penetrate deeper into the skin, reducing the allergic reactions. However, their scattering effect often causes a whitening appearance on the skin (Chavda et al., 2023). Titanium dioxide, one of the most commonly used inorganic filters, offers several advantages such as broad-spectrum protection from both UVA and UVB rays, is photostable, and is safe for children and people with sensitive skin (Egambaram et al., 2020). Sunscreens combining organic and inorganic UV filters have been widely studied. Serpone et al. (2007) incorporated titanium dioxide into micro-fine sunscreens to avoid SPF value decreases due to the organic UV filters’ photoinstability.

Despite the advantages of inorganic filters, there is increasing interest in safer, natural-based UV filters to overcome cosmetic and safety limitations. The utilization of plant extracts as photoprotective agent is increasingly popular due to their minimal potential negative effects, and reducing the amount of synthetic ingredients required in sunscreen. In vitro and in vivo investigations have demonstrated that numerous phytochemicals possess photoprotective properties against UV radiation (Tangyuenyongwatana and Gritsanapan, 2022; Tran et al., 2024).

Syzygium aqueum (S. aqueum) is a medicinal plant commonly found in tropical countries such as Indonesia and Malaysia. This plant is also known as watery rose apple. Its leaf extract has been studied and proven to have cosmetic benefits (Itam et al., 2021). The ethanol extract of watery rose apple leaf (WRAL) has an EC₅₀ value of 35.60 ppm, which although relatively low, demonstrates strong antioxidant activity. The polyphenol content of Syzygium species—including flavonoids, phenolics, and tannins—contributes to this activity (Yumita et al., 2023). The UV absorption properties of polyphenolic compounds, particularly flavonoids, have been widely studied. With their antioxidant capacity, flavonoids offer significant photoprotective potential and can serve as natural sunscreen agents (Ghazi, 2022). Previous studies have developed sunscreen gels containing WRAL ethanol extract at various concentrations. At 10%, the extract produced an SPF of 6.19 ± 0.03 (Rusydi et al., 2023). Therefore, inorganic sunscreen agents such as titanium dioxide need to be incorporated to enhance the photoprotective activity of sunscreen creams containing plant extracts (Widiyati et al., 2021).

Based on the literature review, no previous studies have examined the effectiveness of sunscreen creams containing WRAL ethanol extract and titanium dioxide. This study aims to develop a sunscreen formulation combining natural and inorganic UV filters and investigate the synergistic efficacy based on the SPF value, offering an alternative sunscreen cream that is both safe and effective as a photoprotector.

MATERIALS AND METHODS

Plant material collection and identification

Two kilograms of fresh watery rose apple (Syzygium aqueum) leaves were obtained in March 2024 in Tulungagung, East Java, Indonesia at coordinates 8°11'12.4"S 111°44'29.3"E. The species was identified as Syzygium aqueum by a plant taxonomist at Laboratory of Herbal Materia Medica Batu, East Java, Indonesia under determination number 000.9.3/616/102.20/2024. The collected samples were taken to the laboratory, where the leaves were separated from the stems, rinsed with running water, and air-dried. The wilted leaves were chopped into small pieces and then dried in an oven at 50°C for 24 hours. Once completely dehydrated, the leaves were ground into a fine powder using a blender and stored at room temperature for further use.

Extract preparation

The preparation of WRAL extract was carried out by following the procedure of Srisuksomwong et al. (2023) with slight modifications. The powder was then extracted using the maceration method. An amber maceration vessel was used to soak 350 grams of powder in 1.75 liters of 96% ethanol for three days at room temperature under frequent manual shaking. Filtration was performed in two stages: first using cloth, followed by filter paper to remove fiber from the solution. Using a rotary evaporator (Buchi R-300, Switzerland), the filtrate was evaporated and thickened in a waterbath to produce a concentrated extract. The percentage yield of extract was then calculated. The extracts were stored at 4°C in dark glass bottles until further analysis.

Total flavonoid content (TFC) determination

The method described by Alara et al. (2018) was slightly modified and used to quantify TFC in the ethanol extract of WRAL. For sample preparation, the extracts were dissolved in methanol at a concentration of 50 mg/mL. Quercetin, used as a standard, was diluted to 1, 5, 10, 25, and 50 ppm to construct a calibration curve. An aliquot of 0.1 mL of the extract solution or quercetin was thoroughly mixed with 0.1 mL of 2% AlCl₃ solution and incubated at 25 ± 2°C for 1 hour. Deionized water was then added to adjust the volume to 1 mL. The absorbance was measured using a UV-Vis spectrophotometer (Shimadzu 1240, Japan) at 420 nm. The quercetin standard calibration curve with the linear equation of y = 0.0064x + 0.0137, R² = 0.9945, was used to calculate the TFC content in the extract. The results were presented as quercetin equivalents in milligrams per gram of extract (mg QE/g).

Total phenolic content (TPC) determination

Quantitative analysis of phenolic content in the extract was conducted using the Folin-Ciocalteau method, with gallic acid as the standard. The procedure followed Rosdianto et al. (2023), with slight modifications. The extract sample was dissolved in methanol at a concentration of 50 mg/mL. Gallic acid was diluted to concentrations of 2, 5, 10, 25, and 50 ppm to construct the calibration curve. The test was carried out by pipetting 50 μL of the extract or standard solution into a suitable container, adding 500 μL of Folin-Ciocalteau reagent (10% in water) and 400 μL of 1 M sodium carbonate, and incubating the mixture for 15 minutes at 25 ± 2°C. Absorbance was measured at 765 nm using a UV-Vis spectrophotometer. The gallic acid calibration curve (y = 0.0044x + 0.0093, R² = 0.9752) was used to calculate the total phenol content, expressed as milligrams of gallic acid equivalent per gram of extract (mg GAE/g).

Total tannin content (TTC) determination

Quantitative analysis of tannin content in the extract was performed using the Folin-Ciocalteau method, with tannic acid as the standard. The procedure followed Chandran and Kavitha (2016), with slight modifications. The extract was dissolved in methanol at a concentration of 50 mg/mL. Tannic acid was diluted to concentrations of 0.1, 0.5, 1, 2.5, and 5 ppm to construct the calibration curve. The test was carried out by pipetting 0.1 mL of the extract or standard solution into a 10 mL volumetric flask containing 7.5 mL of distilled water, 0.5 mL of Folin-Ciocalteau reagent, and 1 mL of 35% sodium carbonate. Distilled water was then added to a final volume of 10 mL, the mixture was shaken until homogeneous, and incubated for 30 minutes at 25 ± 2°C. Absorbance was measured at 700 nm using a UV-Vis spectrophotometer. The tannic acid calibration curve (y = 0.0801x + 0.0303, R² = 0.964) was used to calculate the total tannin content, expressed as milligrams of tannic acid equivalent per gram of extract (mg TAE/g).

Preparation of sunscreen creams

The preparation of the oil-in-water (o/w) cream followed the procedure outlined by Widiyati et al. (2021) with minor modifications. The aqueous phase consisted of glycerin, triethanolamine, methylparaben, ascorbic acid, and distilled water, which were placed in a 500 mL beaker and heated on a hotplate to 70°C. The oil phase consisted of stearic acid, lanolin, cetyl alcohol, propylparaben, and virgin coconut oil, which were placed in another 500 mL beaker and heated to 70°C. The aqueous phase was then gradually added to the oil phase while stirring with a homogenizer at 500 rpm until a semi-solid preparation formed. The water apple (Syzygium aqueum) leaf extract was subsequently added with continuous stirring, followed by the addition of titanium dioxide (particle size d10 ≥ 0.1 µm, d50 about 0.8 µm, d90 ≤ 2.5 µm; spherical shape; rutile crystalline form), which was stirred for 15 minutes until a cream formed. The concentrations of extract and titanium dioxide used in the formulations are presented in Table 1.

Table 1. Sunscreen cream preparation formulas with various concentrations of single and combination active ingredients.

Evaluation of sunscreen cream preparations

Organoleptic characteristics

The organoleptic characteristics in this research included color, odor, and homogeneity were observed visually using the method described by Mishra et al. (2014).

pH determination

A pH 7 buffer solution was used to calibrate the pH meter (AMT20, AMTAST, Florida, USA). The samples were placed in beakers, and their pH was measured using the pH meter at room temperature.

Spreadability measurement

The spreadability was measured by placing 1 gram of the sample on a glass plate. Another glass plate was placed on top of the first plate, with a 50-gram weight applied for 1 minute. The weight was added periodically until the diameter remained constant. The diameter of the spread sample was measured in centimeters using a caliper (Kusmita et al., 2024).

Viscosity measurement

The viscosity test was conducted following the procedure outlined by Zulkarnain et al. (2022). The sample was placed in a chamber, and rotor number 1 was installed until it was immersed in the sample. The viscometer (Rion VT-04F, Japan) was activated to allow the rotor to rotate. Once the needle on the viscometer stabilized at a viscosity scale, the reading was recorded in units of dPas (1 dPas = 100 cps).

Sensory evaluation

All procedures involving human participants in this study were approved by the Research Ethics Commission of the Faculty of Dentistry, Airlangga University (Approval no. 0353/HRECC.FODM/IV/2024). Written informed consent was obtained from all participants in accordance with ethical standards. Sensory evaluation in this study was based on the research by Balboa et al. (2017) with slight modifications. A total of 20 healthy female panelists aged 20–40 years participated as volunteers and completed a questionnaire on five cream samples. Each sample underwent a hedonic assessment covering five parameters: appearance, odor, texture, spreadability, and whitening effect. The preference levels for each parameter were measured using a 1–10 scale, where 1 indicated the lowest preference and 10 the highest. Each panelist evaluated five cream sample formulas across these five parameters.

SPF measurement of sunscreen cream

Following the methodology outlined by Widiyati (2017), the in vitro SPF value was calculated using UV-Vis spectrophotometry to assess the UVB photoprotection effectiveness of each cream formulation. A total of 50 mg of cream was placed into a 50 mL volumetric flask, and isopropanol was added as a solvent. The absorbance data of the sample solution was recorded in the wavelength range of 290-320 nm at 5 nm interval. Triplicate was measured at each point. The SPF value was calculated using the absorbance that was acquired by using the following Mansur equation.

Note: CF = correction factor (= 10), EE = erythema effect spectrum, I = solar intensity spectrum, Abs = absorbance of sunscreen cream.

Accelerated stability test

The sunscreen formula that meets physical quality requirements and achieves the highest SPF is considered the optimum formula. The optimum formula was subjected to accelerated stability studies at different storage temperatures. Six heating-cooling cycles were used to conduct the stability test. Each cycle of heating and cooling included a 24-hour period in the fridge at 4°C and a 24-hour period in the oven at 40°C (Jaikampan et al., 2025). The samples were examined at the end of each cycle. They were also tested for stability in a temperature-controlled room at 25°C over a 28-day storage period and were sampled on days 7, 14, 21, and 28. At each sampling interval, observations on their organoleptic, pH, viscosity and spreadability were conducted.

Skin hydration test

The optimum formula of sunscreen cream preparation was then evaluated for skin hydration and irritation. The skin hydration and irritation test on respondents was carried out with the permission of the Research Ethics Commission of the Faculty of Dentistry, Airlangga University No. 0353/HRECC.FODM/IV/2024. This test was carried out over one week involving 15 respondents. The inclusion criteria included being healthy women aged 18-30 years, having no previous skin problems, and consenting to participate in the study by filling out a willingness form. The exclusion criteria included individuals with skin diseases such as dermatitis, extensive wounds, or systemic diseases, those who were pregnant, and those who failed to participate in the research for three consecutive days. On the first day, a skin analyzer (model: CR-302, U-trak, China) was used to determine the respondent's skin hydration on the back of their hands. The cream preparation was applied nightly to a 5 × 5 cm area on the back of the left hand for a week. On the seventh day, the hydration condition of the back of the respondent's left hand was reassessed using the skin analyzer. The results of the skin hydration test before and after applying the cream were then compared (Rohmani et al., 2021). Skin hydration was assessed by measuring the water content of the epidermis. Hydration levels were categorized as follows: <33% for dehydrated skin, 34–37% for dry skin, 38–42% for normal skin, and 43–46% for very hydrated skin (Widiawaty et al., 2022).

Skin irritation test

The cream preparation was applied to the back of the left hand in a 1 x 1 cm area for 24 hours. Erythema and edema were assessed during the time and recorded (Mishra et al., 2014).

Statistical analysis

All data were obtained from triplicate, and the results were expressed as mean ± standard deviation (SD). Statistical analyses were performed using a paired t-test with SPSS version 16.0, with a P-value of < 0.05 considered statistically significant. A paired t-test was used to determine differences in stability test results at each sampling point compared to the baseline (day 0) throughout the storage period and to assess skin improvement in the hydration test (before and after application). Sensory evaluation scores were analyzed using the non-parametric Kruskal–Wallis test to determine significant differences among the five cream formulations.

RESULTS

Characterization of extract

The WRAL extraction produced extracts with a blackish green appearance, a thick semisolid texture, and a distinctive odor. The results of the percentage of extract produced and the total flavonoid, phenolic, and tannin contents are shown in Table 2.

Table 2. Percentage yields and phytochemical contents of Syzygium aqueum leave extract.

Preparation of sunscreen cream

The sunscreen cream formulation results are presented visually in Figure 1. The resulting sunscreen cream ranges in color from white to brownish green, depending on the type and concentration of active ingredients added. Cream F1 produces a white cream preparation that is difficult to apply and causes a white cast effect on the skin, resulting in lower cosmetic acceptance. Cream F2 has a brownish green appearance and leaves a green tint on the skin when applied. Furthermore, creams F3 and F5 have a similar color, both appearing yellowish green, providing a smooth finish, and not leaving color marks on the skin even though the resulting cream is tinted. In contrast, cream F4 has a green appearance and leaves a slight color mark on the skin when applied.

Figure 1. Visual appearance of sunscreen cream formula F1 (6% TD); F2 (10% extract); F3 (10% extract, 6% TD); F4 (15% extract, 6% TD); F5 (10% extract, 9% TD).

Evaluation of sunscreen cream

Organoleptic characteristics, pH, spreadability, and viscosity measurement

The organoleptic characteristics (color, odor, and homogeneity), pH, spreadability, and the viscosity of the sunscreen cream formulations are shown in Table 3. The pH values of the resulting cream formulas range from 6.09 to 6.46, which are in accordance with the pH requirements of topical preparations, set between 4 and 7 (Bora et al., 2019). Furthermore, all cream formulas meet the viscosity requirements for good semisolid preparation, which are between 50 and 1,000 dPas (Rohmani and Pangesti, 2024). A higher the viscosity value indicates greater resistance flow, making it more difficult to apply. Conversely, a lower viscosity in topical preparation results in a higher spreadability value. The five creams also met the spreadability requirements, ranging from 5 to 7 cm (Aisyah et al., 2024). A greater spreadability value facilitates easier application of the cream, thus increasing the contact area between the active ingredients and the skin. In addition, good spreadability can enhance the absorption rate of active ingredients by the skin (Rohmani and Pangesti, 2024).

Table 3. Physical evaluation of sunscreen cream.

Sensory evaluation

The sensory test on the cream samples measured appearance, odor, texture, spreadability, and whitening effect. The preference levels were assessed using a 10-point scale, with 1 indicating "very dislike" and 10 indicating "very like." The panelists' preferences for the cream formulas are presented in Figure 2. Formula F1 was the most preferred in terms of appearance and odor, while F3 was favored for texture, spreadability, and whitening effect. The overall panelist preferences for formulas F1 to F5 were 6.30 ± 2.84, 5.90 ± 0.55, 7.79 ± 0.88, 6.23 ± 0.57, and 6.76 ± 0.59, respectively, indicating that F3 was the most preferred and acceptable formulation. Sensory evaluation data were analyzed using the non-parametric Kruskal–Wallis test. The results indicated significant differences between cream formulations F1-F5 across five test parameters (P < 0.05). Variations in active ingredient concentrations among formulations significantly influenced sensory parameters.

Figure 2. Sensory evaluation of the sunscreen cream formula F1-F5 on various parameters.

Figure 3. In vitro SPF value of sunscreen cream F1-F5. F0 = cream base, F1-F5 = sunscreen cream formulation 1-5, control = commercial sunscreen.

Sun protection factor (SPF) value

Figure 3 shows the results of measuring the SPF value of the produced cream formula. F1 cream, with titanium dioxide as its single active ingredient, yielded an SPF value of 5.10 ± 0.15, while F2 cream, using WRAL extract alone, yielded an SPF value of 20.24 ± 0.19. F3 cream, a combination of the two active ingredients in a 1:1 ratio, resulted in an SPF value of 40.22 ± 0. F4 cream, with a 1.5:1 ratio of WRAL extract and titanium dioxide, had an SPF value of 12.31 ± 0.41. Finally, F5 cream, with a 1:1.5 ratio, produced an SPF value of 10.92 ± 0.49. The F0 formula (cream base) had an SPF value of 4.58 ± 0.23, as it contained virgin coconut oil (VCO), which provides protection againts UVB radiation (Varma et al., 2019). Commercial products containing both organic and inorganic UV filters showed an SPF value of 49.89 ± 2.58, consistent with the information on their product labels. These evaluation show that F3 is the optimum formula, SPF nearly eightfold compared to titanium dioxide alone and nearly twofold compared to WRAL extract alone.

Based on the results of the physical quality evaluation and the in vitro SPF value, cream F3 was chosen as the optimum formula. It demonstrated good organoleptics, met the physical requirements for semisolid preparations, and had the highest SPF value. The optimum formula of cream F3 was then subjected to accelerated stability test, skin hydration and irritation test.

Accelerated stability test

F3 sunscreen cream was subjected to accelerated stability test at different storage temperatures, including 6 heating-cooling cycles and 25°C for 28 days. For the heating-cooling, the sampling was conducted every cycle, while for the 25°C, the sampling was carried out every 7 days. At each sampling time, evaluations on its organoleptics, pH, spreadability and viscosity were conducted. The results of the accelerated stability test at each sampling time of F3 sunscreen cream are available in Table 4.

The organoleptic evaluation showed no change in color, odor, and homogeneity over time. The pH, spreadability and viscosity parameters remained stable throughout the storage period. The statistical analysis of each sampling point indicated a P-value of > 0.05, meaning that the values were not significantly different from those of day 0 for each parameter.

Table 4. F3 sunscreen cream stability test over time at various temperatures.

Figure 4. The hydration levels of the skin after applying F3 sunscreen cream for seven days on 15 respondents.

Skin hydration test

Figure 4 shows the results of the skin hydration level in 15 respondents before and after 7 days of the cream usage. The skin hydration test showed that the mean hydration level of respondents’ skin prior to treatment was 36.67%, classified as dry skin. After one week of applying the F3 cream, the average hydration level increased to 43.60%, classified as very hydrated skin. The F3 sunscreen cream significantly increased the skin hydration level, as indicated by the P-value of < 0.05 between the measurements taken before and after use for each respondent. These findings indicate that F3 cream has good efficacy in increasing skin hydration.

Skin irritation test

The respondents who participated in the irritation test were the same as those in the hydration test. After 24 hours of the irritation test, skin changes were evaluated. The results showed that none of the 15 respondents experienced redness or edema after 24 hours of exposure to the cream sample. The erythema and edema scores were both 0. F3 cream can be safely applied to the skin, as its ingredients are safe for use in sunscreen cosmetics.

DISCUSSION

Syzygium aqueum is a native plant of Indonesia. According to the research by Manaharan et al. (2012), WRAL extract possesses cosmeceutical properties. The ethanol extract of WRAL in this study exhibited a total flavonoid content of 46.33 ± 1.02 mg QE/g, measured using UV-Vis spectrophotometer. Flavonoids can absorb UV light in the range of 200 to 400 nm, exhibiting intensive absorption with broad bands due to their aromatic groups and conjugated bond structures. Flavonoids offer three distinct photoprotection effects: UV light absorption, direct and indirect antioxidant activity, and modulation of several signaling pathways. Given its large number of natural compounds that perform well as sunscreen active ingredients, natural UV filters are emerging as a promising alternative suitable for all skin types (Saewan and Jimtaisong, 2013; Carvalho et al., 2023; Fonseca et al., 2023).

The ethanol extract of WRAL in this study contained 31.87 ± 0.96 mg GAE/g of total phenols. Polyphenols in this extract react with specific redox reagents to form a blue complex that can be quantified using a UV-Vis spectrophotometer. This reaction produces a blue chromophore through a phosphotungstic–phosphomolybdenum complex. Phenolic compounds act as natural antioxidants due to their structure, which consists of a phenol ring and hydroxyl groups bound to an aromatic ring that facilitate oxidation by donating hydrogen atoms to free radicals (Blainski et al., 2013). The ethanol extract of WRAL in this study also contained 40.52 ± 0.72 mg TAE/g of total tannins, another class of phenolic compounds that contribute to antioxidant properties (Sharma and Argawal, 2015).

Based on previous studies, one of the key compounds in the WRAL extract is myricetin, a flavonol known to protect keratinocytes from UVB damage. This highlights WRAL extract as a highly suitable skincare ingredient due to its antioxidant activity, tyrosinase inhibition properties, and UVB-blocking capability. Myricetin has a protective effect against UVB by inhibiting lipid peroxidation and hydrogen peroxide production. It enhances the proliferation of normal keratinocytes while reducing oxidative damage from UVB exposure. Studies have demonstrated that myricetin possesses photoprotective activity, making it a suitable addition to sunscreen formulations for preventing UV-induced skin damage (Huang et al., 2010; Palanisamy et al., 2011).

Titanium dioxide (TiO₂) is the most widely used inorganic UV filter for photoprotection. It provides excellent protection against UV radiation and is chemically inert, reducing the risk of allergic sensitization (Nunes et al., 2018). TiO₂ is considered the most effective inorganic UV filter due to its high refractive index, which efficiently blocks both UVA and UVB rays. TiO₂ exists in two crystal forms, anatase and rutile, each with distinct photocatalytic properties. Since the anatase form has been shown to be more cytotoxic and photoreactive, the rutile form is more commonly used in sunscreens (Gilbert et al., 2013). According to Schneider and Lim (2019), TiO₂ poses a very low health risk to humans due to its minimal absorption in both damaged and normal skin. The FDA has also recognized TiO₂ as a safe and effective UV filter (Adler and DeLeo, 2020). TiO₂ reflects and scatters light, offering broad-spectrum protection. However, its scattering effect often leads to a whitening appearance on the skin, which can reduce aesthetic value (Ferreira et al., 2023). In this study, combinations between WRAL extract and titanium dioxide as active ingredients were used as natural and inorganic UV filters, respectively, and they were incorporated in the cream base formula, as shown in Table 1.

The all five resulting cream preparations met the physical quality requirements, including pH, spreadability and viscosity. However, F3 cream was the most preferred by panelists based on the highest hedonic score. The SPF value of the combination of WRAL extract and titanium dioxide in a ratio of 1:1 yields optimum results in cream F3. This is higher than the SPF value of each single active ingredient, which are lower, as seen in F1 (titanium dioxide) and F2 (extract). This indicates that flavonoids are insufficient to fully suppress photochemical reactions catalyzed by UV radiation. Combining flavonoids with UV filters enhances their synergistic efficacy and improves photoprotection (Hubner et al., 2019).

However, when the ratio includes a 1.5-fold increase of either the extract or titanium dioxide, the SPF values decrease, as seen in creams F4 and F5. This aligns with a study conducted by Dianursanti et al. (2020), beyond a certain flavonoid concentration, the SPF value begins to decline. Not all formulations exhibit a linear increase in SPF with higher concentrations of sunscreen actives. Previous research reported that a rutin nanocream containing 5% TiO₂ achieved the optimum SPF, whereas increasing the concentration to 7.5% resulted in a decrease. This finding demonstrated that an optimum concentration enables better arrangement and conformation of TiO₂ molecules within the formulation, thereby enhancing UV-blocking effectiveness (Kamel and Mostafa, 2015). This study also finds that the development of this combination is more protective, based on its SPF value, compared to other combinations of natural UV filters with titanium dioxide, as reported in previous studies (Rohmani and Pangesti, 2024).

Amnuaikit and Boonme (2013) found that the viscosity of sunscreen creams directly correlates with their SPF values, as optimal viscosity enhances adhesiveness efficacy. The incorporation of 10% flavonoids (rutin and quercetin) in formulations has been shown to produce an SPF value comparable to homosalate, a standard sunscreen ingredient. When combined with TiO₂, the resulting SPF value reaches 30, demonstrating the synergistic effect of flavonoids and inorganic UV filters in boosting SPF (Cefali et al., 2016). TiO₂ concentrations ranging from 2.5% to 10% provide the necessary efficiency when combined with other filters. At concentrations of 5%–10%, there is no significant difference in SPF and UVA protection, as the effectiveness depends on the additional filters used to enhance efficiency (Ghamarpoor et al., 2023).

Monsalve-Bustamante et al. (2020) reported that using plant extracts combined with physical sunscreens (inorganic UV filters) effectively protects the skin against UV rays while maintaining the skin matrix against oxidative stress and synergistically increasing SPF value compared to single filters. Plant extracts rich in flavonoids possess an aromatic ring structure that enables UV absorption, allowing them to act as natural UV filters with antioxidant and anti-inflammatory properties (Li et al., 2023). Flavonoids protect plants from UV radiation by absorbing sunlight and scavenging reactive oxygen species (ROS). The presence of phenolic hydroxyl groups in flavonoids facilitates ROS scavenging, further enhancing their protective effects (Milutinov et al., 2024). These combined activities reinforce flavonoids' role as SPF boosters. Sunscreen effectiveness is often enhanced by incorporating antioxidants, which improve photoprotective properties (Shabrina et al., 2025).

The UV absorption capacity of flavonoids is similar to the mechanism of organic UV filters or chemical sunscreens (Manaia et al., 2013). Flavonoids, through their UV-absorbing properties, can act synergistically when combined with inorganic UV filters in a single formulation. Inorganic UV filters such as TiO₂, reflect or scatter light at the skin surface, thereby increasing the optical path of photons, while flavonoids absorb the retained photons (Lademann et al., 2005). In addition, due to its intrinsic photocatalytic activity under UV irradiation, TiO₂ facilitates the generation of ROS (Morsella et al., 2016). At the molecular level, the elevated presence of hydroxyl groups in flavonoids augments their ability to donate hydrogen atoms, which promotes the stabilization of reactive free radicals (Niksic et al., 2025). This synergistic interaction results in higher UV absorption and reflection, which can increase the SPF value compared to when either component is used alone.

The reliability of the Mansur method for SPF measurement in accordance with USP guidelines was evaluated by Zaid et al. (2018). Their study, conducted on FDA-approved homosalate sunscreen products, demonstrated that the Mansur method is accurate for in vitro SPF assessment, producing results comparable to those of the FDA method. Owing to its accuracy and simplicity, the Mansur method remains the most widely used technique for in vitro SPF determination. In vitro SPF evaluation is commonly performed to estimate in vivo SPF values. Mansur et al. (2016) reported a strong correlation between in vitro and in vivo SPF values in human skin. Among the three tested formulations, the base formula had a lower in vivo SPF value, likely due to biological factors. Despite this correlation, only in vivo testing is accepted for sunscreen registration with healthcare regulatory agencies for commercial purposes. In vitro SPF testing is primarily used as a preliminary screening method to select the most promising formulation for subsequent in vivo testing.

In this study, we selected an oil-in-water (o/w) emulsion type for the cream due to its advantages over water-in-oil (w/o) emulsions, including better spreadability on the skin surface and a non-sticky feel, making it more cosmetically acceptable. The optimum formula F3 demonstrated physical stability both at 25°C and under extreme conditions, confirming that the formula meets the essential parameters for good cream preparation (Zulfaidah et al., 2023). Additionally, F3 sunscreen cream significantly increase skin hydration after one week of use, attributable to the inclusion of virgin coconut oil (VCO) and glycerin in the formula. Formulations containing glycerin not only benefit photoprotection but also enhance skin hydration (Ouyang et al., 2013). Clinical trials by Varma et al. (2019) demonstrated that VCO reduces skin inflammation, improves epidermal barrier function, and provides hydration benefits. Furthermore, the sunscreen cream preparation did not cause skin irritation because the ingredients contained were non-irritant.

Study limitations

A limitation of this study is the exclusive use of the in vitro method to assess the photoprotective potential of the cream formulation containing a combination of WRAL extract and TiO₂. While in vitro SPF assessment offers valuable preliminary insights, in vivo studies are essential to comprehensively validate the formulation's efficacy. Additionally, long-term stability and photostability of the formulation were not addressed, despite being critical factors for ensuring safe application in cosmetic products. Furthermore, to strengthen the robustness and enhance the generalizability of the findings, future studies should include a larger sample size and a more diverse demographic representation of the respondents.

CONCLUSION

WRAL ethanol extract contains a total flavonoid content of 46.33 ± 1.02 mg QE/g extract, demonstrating that flavonoids can function as natural UV filters. The combination of WRAL ethanol extract as a natural UV filter with titanium dioxide as an inorganic UV filter has been successfully formulated into a sunscreen cream preparation. Cream F3, containing 10% WRAL extract and 6% titanium dioxide, is the optimum formula, characterized by a yellowish-green color, a mild leaf scent, homogeneous texture, and overall good physical properties. Other evaluations of physical quality also suggest other requirement fulfillments, including a pH of 6.12 ± 0.01, a spreadability of 5.5 ± 0.15 cm, and a viscosity of 60 ± 10 dPas. The optimum formula F3 provides an SPF value of 40.22 ± 0, indicating the synergistic efficacy of the combination of natural and inorganic UV filters. It is physically stable both at 25°C and under heating-cooling conditions during the storage period. Additionally, it significantly increases skin hydration (P-value > 0.05) and did not cause skin irritation.

ACKNOWLEDGEMENTS

The authors would like to thank Karya Putra Bangsa Health Science Academy for providing instruments.  

AUTHOR CONTRIBUTIONS

Arfinda Diah Setiowati: Conceptualization (Lead), Investigation (Lead), Data Curation (Lead), Formal Analysis (Equal); Tri Anita Sari: Writing – Original Draft (Lead), Writing – Review & Editing (Lead), Validation (Equal), Visualization (Lead), Supervision (Equal), Formal Analysis (Equal), Conceptualization (Supporting); Afidatul Muadifah: Supervision (Equal), Validation (Equal), Conceptualization (Supporting).

CONFLICT OF INTEREST

The authors declare that they hold no competing interests.

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