Clinical and Instrumental Evaluation of the Anti-Aging Effectiveness of a Cream Based on Hyaluronic Acid and a Cream Based on Hyaluronic Acid and Vitamin C: A Prospective, Multicenter, 8-Week, Parallel-Group Randomized Study on 91 Subjects

Abstract

Introduction: Skin aging is a multifaceted process influenced by both intrinsic and extrinsic factors, resulting in visible changes such as wrinkles, loss of elasticity, uneven skin tone, and hyperpigmentation. Hyaluronic acid (HA) is widely recognized for its hydrating and structural support properties, while Vitamin C is known for its antioxidant and depigmenting effects. This study investigated the anti-aging efficacy of two topical formulations containing Jalubalance^® technology—HA delivered in Opuntia oil—with or without 1% Vitamin C. Background/Objectives: We conducted an 8-week, multicenter, randomized trial involving 91 women aged 30–50 years with mild-to-moderate photoaging. Participants were assigned to apply either HA-only cream (Group A) or a HA + Vitamin C cream (Group B) twice daily. The primary outcome was the percentage of subjects who achieved an improvement of at least one point in the hyperpigmentation score from baseline to week 8. Additionally, the study aimed to evaluate and compare the clinical and instrumental effects of both treatments, with a particular focus on improvements in wrinkles, elasticity, hydration, and pigmentation. Results: Both groups showed significant improvements across all measured parameters, including Glogau scores, wrinkle reduction, and skin elasticity. Instrumental analysis confirmed increased hydration and elasticity. Group B showed a significantly greater reduction in hyperpigmentation (−45%) compared to Group A (−31%, p < 0.05). At week 8, a ≥1-point reduction in hyperpigmentation score was observed in 56% of subjects in Group B and 30% in Group A (absolute difference: 26%; 95% CI: 5–43%; p < 0.05), highlighting the added benefit of Vitamin C on this parameter. Participant satisfaction was high, especially for the moisturization and brightening effects of both products. Conclusions: The topical application of Jalubalance-based creams effectively reduced signs of aging. The inclusion of Vitamin C provided enhanced benefits in reducing hyperpigmentation, suggesting its utility in personalized dermatological approaches for patients with pigmentation concerns.

1. Introduction

Skin aging is a complex multifactorial biological process, influenced by several factors, both intrinsic and extrinsic. Intrinsic aging, also known as chrono aging, is primarily driven by genetical factors that lead to physiological changes in cells, such as hormonal fluctuations and cellular senescence, which lead to a gradual structural deterioration of the skin, including skin thinning and the development of fine wrinkles [1,2,3,4].

Extrinsic aging, commonly referred to as photoaging, is induced by the exposition to environmental factors such as ultraviolet (UV) radiation, air pollution, smoking, and diet-induced factors (nutritional deficiencies). These stressors accelerate skin aging manifestations, resulting in deeper wrinkles, premature fine lines, uneven pigmentation, and a change in the skin texture, particularly in sun-exposed areas [5,6,7,8,9]

At the molecular level, skin aging is driven by several mechanisms, in particular oxidative stress, cellular senescence, degradation of the extracellular matrix (ECM) and mitochondrial dysfunction. Oxidative stress is the mechanism induced by an excessive generation of reactive oxygen species (ROS), that causes damage to DNA, proteins, and cellular membranes, leading to an impairing in tissue repair and degeneration of the tissue [9,10,11]. The accumulation of senescent cells contributes to chronic inflammation and tissue dysfunction (representing one of the hallmarks of aged skin) [12,13,14,15].

In addition, as age progresses, the components of ECM, especially collagen and elastin, undergo toward a process of degradation because of the increased action of metalloproteinases (MMPs), leading to the loss of elasticity and structural integrity [2,4,9,16].

Another key contributor is mitochondrial dysfunction, which exacerbates ROS production and cellular aging phenotypes [10,12].

The main molecule used in cosmetic formulation to counteract skin aging is hyaluronic acid (HA). This is a natural component of the skin extracellular matrix, and it demonstrated during several years of research that it is crucial in maintaining moisture, firmness, and elasticity of the skin. As the organism ages, the level of HA constantly decreases, as a direct consequence of this phenomenon, skin tends to be drier and less elastic, leading to the formation of fine lines and wrinkles [17]. Supplementation of topical HA helps to counteract chrono and photoaging effects, protecting skin from UV-induced damage, reducing ROS, inflammation, and apoptosis of skin cells [18,19,20], and maintaining skin hydration and elasticity, thus counteracting the natural decline in skin moisturization and firmness, as well as supporting the structure of extracellular matrix and, as a consequence, reducing wrinkles and fine line formation [17,21].

Also known as Ascorbic acid, Vitamin C acquired more relevance in different sectors, such as medicine, food, and now, cosmetics. Its potential as an ingredient in cosmetic formulations has been widely studied during this time, because of its protective role against oxidative damage and in boosting collagen production [22,23]. Vitamin C is also a well-established ingredient for skin brightening and reducing pigmentation. In fact, molecularly, Vitamin C interferes with the activity of the enzyme tyrosinase, which is essential for melanin production [24]. By inhibiting tyrosinase, Vitamin C reduces the formation of new pigments, helping to prevent hyperpigmentation [25,26]; it also helps to fade already existing dark spots by reducing melanin oxidation and supporting skin cell turnover [27,28,29].

Based on the literature results, we wanted to evaluate the clinical efficacy between two different face creams both containing hyaluronic acid-based technology (Jalubalance^®, Difa Cooper SpA, Caronno Pertusella, Italy), where hyaluronic acid is delivered in prickly pear oil (Opuntia oil), with one of them containing 1% of Vitamin C.

2. Materials and Methods

2.1. Population and Study Design

Between February 2025 and June 2025, a total of 91 participants (median age 43 ± 6 years), were enrolled in fifteen dermatology clinics affiliated with the Donne Dermatologhe Italia network, randomized in a 1:1 allocation ratio, and instructed to apply a cream formulated with red algae and Lactobacillus, enriched with Jalubalance^® technology (high-concentration hyaluronic acid delivered through Opuntia oil) (HA-only cream, Group A), or to apply a cream that combines the Jalubalance^® technology with 1% vitamin C (HA–Vitamin C cream, Group B). The products were applied to the entire face twice daily (morning and evening) for eight weeks, using 1.5 g of product per application (3 FTUs) on cleansed skin. A dedicated computer program was used to generate the randomization list. Both products were commercially available (Jaluronius Cream and Jaluronius Cream C, Cantabria Labs Difa Cooper, Caronno Pertusella, Italy).

The rationale for selecting these two formulations was to directly compare the anti-aging efficacy of a hydrating and elasticity-restoring active (HA) alone versus the same base formulation enriched with an antioxidant and depigmenting agent (Vitamin C), which has additional collagen-stimulating and melanin-inhibiting effects. The prescription-like regimen (twice daily, fixed amount, standardized cleansing before application) was intended to maximize adherence and reproducibility across study centers.

The main inclusion criteria were women with mild-to-moderate photoaging (Glogau score 1 or 2) and age between 30 and 50 years. The main exclusion criteria were pregnancy or breastfeeding, allergy to components present in the products, acute facial skin diseases, and esthetic procedures performed within the past six months (e.g., botulinum toxin injections, dermal fillers, laser treatments, chemical peels, topical retinoids). The study was conducted over 8 weeks, and participants were evaluated at the baseline (T0) and after 8 weeks (T1). The trial took place in fifteen dermatology clinics affiliated with the Italian Association “Donne Dermatologhe Italia”. The trial was conducted according to Good Clinical Practice Guidelines and Helsinki Declaration and consistent with the Good Clinical Practice (GCP) regulatory requirements. All subjects provided a signed informed consent. Study protocol was approved by the Internal Review Board of Donne Dermatologhe Italia (DDI-Pn01/24, 6 December 2024).

2.2. Study Outcomes

The primary efficacy outcome was the percentage of subjects who achieved an improvement of at least one point in the hyperpigmentation score from baseline to week 8.

Additional main efficacy outcomes included the evaluation of the Glogau score and a clinical score that assessed four parameters (wrinkles, elasticity, skin uniformity, and hyperpigmentation) with a 4-point score for each item (ranging from 0 = no issue to 3 = severe issue). The secondary efficacy outcome was the instrumental evaluation of treatment efficacy using the Digital Skin Moisture Meter, which uses Bioelectric Impedance Analysis (BIA) to measure skin hydration, sebum content, and skin elasticity. This device works by passing a very low, safe electrical current through the skin and measuring impedance. Hydration increases electrical conductance due to water’s conductive properties; sebum alters the skin’s surface capacitance, affecting the impedance pattern; elasticity is inferred from changes in the dielectric properties and mechanical resistance detected during measurement. Physiological reference ranges were 35–60% for hydration and 16–30% for oil content. Additionally, a VISIA^® (Canfield Scientific, Parsippany-Troy Hills, NJ, USA) objective face sculptor analysis was performed in a subgroup of 10 subjects.

At the end of the study (T1), the overall efficacy of the treatments was evaluated by each participant through a questionnaire evaluating four specific parameters: moisturizing effect, smoothing effect, skin tone uniformity, and brightening effect. Skin uniformity was assessed clinically within the four-item grading system (0–3 scale) and, in a subset of 10 participants, by VISIA^® imaging for pigmentation distribution and tone irregularities. This dual method was chosen because skin uniformity is a core component of photoaging assessment, with Vitamin C potentially improving melanin distribution and HA enhancing optical smoothness through hydration. For each parameter, subjects were asked to rate the perceived effectiveness using a 4-point scale (from excellent to none). This subjective evaluation provided insights into participant satisfaction and perceived benefits of the treatment, complementing the clinical and instrumental data.

2.3. Statistical Analysis

Sample size calculation was performed with the hypothesis that the cream with Vitamin C (HA + Vitamin C cream) could have a greater efficacy in reducing the hyperpigmentation score, with a high percentage of subjects achieving at least a 1-point reduction after treatment compared to baseline. The alternative hypothesis (H1) assumed that the Vitamin C cream would result in at least 58% of subjects showing a ≥1-point reduction in hyperpigmentation score, versus 30% in the control group (cream without Vitamin C). With an alpha value of 0.05 and a power of 80%, a total of 88 subjects should be enrolled in the trial. The sample size was calculated using G*Power statistical software (G*Power Version 6, Heinrich Heine University, Kiel, Germany).

Statistical analyses were conducted using GraphPad statistical software version 9.0 (GraphPad Software Inc., La Jolla, CA, USA). A two-way ANOVA test, a non-parametric t-test, and a chi-squared test were used to compare data at baseline and after 8 weeks of treatment. Data are expressed as mean ± standard deviation (SD), and a p-value < 0.05 was considered significant.

3. Results

This was a multicenter, randomized, prospective assessor-blinded trial that involved fifteen dermatology clinics affiliated with the Italian association “Donne Dermatologhe Italia”. The study was conducted according to the Declaration of Helsinki and GCP directive. The study protocol was approved by the Investigational Review Board of the Italian association “Donne Dermatologhe Italia” (dated 06 December 2024). The trial was carried out in Italy, between February 2025 and June 2025. A total of 91 women with mild-to-moderate photoaging were enrolled, and all the subjects completed the treatment period. Subjects were randomized into two groups: Group A (HA-only cream, n = 47, mean age 42 ± 6 years, mean Glogau score at baseline 1.7 ± 0.7) and Group B (HA + Vitamin C cream, n = 44, mean age 44 ± 6 years, mean Glogau score at baseline 1.7 ± 0.7) in a 1:1 ratio. The flow diagram of the study is illustrated in Figure 1.

After two months of treatment, the Glogau score was significantly reduced in both groups (Group A −34% and Group B −33%, p < 0.0001, two-way ANOVA test).

No significant differences were observed between the two groups (Figure 2), confirming the effectiveness of both treatments in reducing the signs of skin aging.

Each parameter was evaluated using a clinical scoring system ranging from 0 (no issue) to 3 (severe issue). A decrease in the score corresponds to an improvement in the assessed parameter.

Table 1. Score to evaluate the clinical efficacy of treatments. Group A: HA-only cream, Group B: HA + Vitamin C cream.

Parameter	Group A		Group B
	T0	T1	T0	T1
Wrinkles	1.3 ± 0.7	0.9 ± 0.6 ***	1.4 ± 0.7	1.0 ± 0.6 **
Elasticity	1.4 ± 0.6	0.6 ± 0.7 ****	1.4 ± 0.7	0.6 ± 0.5 ****
Skin uniformity	1.4 ± 0.8	0.7 ± 0.7 ****	1.6 ± 0.6	0.7 ± 0.6 ****
Hyperpigmentation	1.0 ± 0.7	0.7 ± 0.6 *	1.4 ± 0.8	0.8 ± 0.5 ****

* p < 0.05, ** p < 0.001, *** p < 0.0005, **** p < 0.0001. Two-way ANOVA test.

Table 1. Score to evaluate the clinical efficacy of treatments. Group A: HA-only cream, Group B: HA + Vitamin C cream.

Parameter	Group A		Group B
	T0	T1	T0	T1
Wrinkles	1.3 ± 0.7	0.9 ± 0.6 ***	1.4 ± 0.7	1.0 ± 0.6 **
Elasticity	1.4 ± 0.6	0.6 ± 0.7 ****	1.4 ± 0.7	0.6 ± 0.5 ****
Skin uniformity	1.4 ± 0.8	0.7 ± 0.7 ****	1.6 ± 0.6	0.7 ± 0.6 ****
Hyperpigmentation	1.0 ± 0.7	0.7 ± 0.6 *	1.4 ± 0.8	0.8 ± 0.5 ****

* p < 0.05, ** p < 0.001, *** p < 0.0005, **** p < 0.0001. Two-way ANOVA test.

The score related to wrinkles was significantly reduced in both groups (−34% Group A, p < 0.0005; −30% Group B, p < 0.001). No significant difference was observed between the two groups, confirming the efficacy of both treatments in reducing wrinkle-related signs of aging.

The elasticity score (−57% Group A and −55% Group B, p < 0.0001) and the skin uniformity (−49% Group A and −59% Group B, p < 0.0001) were significantly reduced in both groups, indicating an improvement in skin elasticity and skin tone. No significant differences were observed between the two groups.

The hyperpigmentation score was significantly reduced in both groups (−31% Group A, p < 0.05; −45% Group B, p < 0.0001), indicating an improvement in skin discoloration.

The reduction in the hyperpigmentation score in Group B (HA + Vitamin C cream) was statistically greater than in Group A (HA-only cream) when expressed as a percentage (Figure 3, p < 0.05, non-parametric t-test). This effect may be explained by the presence of Vitamin C in the cream formulation, confirming its role as inhibitor of melanin production through the suppression of the enzyme tyrosinase [30,31].

Regarding the primary outcome of the trial, 56% of subjects in Group B (HA + Vitamin C cream) showed a reduction of at least 1 point in the hyperpigmentation score at week 8 compared to baseline. In Group A, this percentage was 30% (p < 0.05, chi-squared test), resulting in an absolute difference of 26% (95% CI 5–43%).

Treatment efficacy was also instrumentally assessed using the Digital Skin Moisture Meter, which measures skin hydration (normal moisture value 35–60%), sebum content (normal oil value 16–30%), and elasticity. The results on skin hydration and sebum content are illustrated in

Table 2. Percentage of subjects with physiological and altered skin hydration at baseline (T0) and after 8 weeks of treatment (T1). Group A: HA-only cream, Group B: HA + Vitamin C cream.

	Group A		Group B
	T0	T1	T0	T1
Altered skin hydration	43%	16%	51%	9%
Physiological skin hydration	57%	84%	49%	91%

and

Table 3. Percentage of subjects with physiological and altered skin sebum content at baseline (T0) and after 8 weeks of treatment (T1). Group A: HA-only cream, Group B: HA + Vitamin C cream.

	Group A		Group B
	T0	T1	T0	T1
Altered sebum content	40%	25%	35%	23%
Physiological sebum content	56%	71%	65%	77%

, respectively.

In both groups, the percentage of subjects with physiological skin hydration significantly increased after two months of treatment (T1) compared to baseline (p < 0.05, chi-square test, Table 2). Similarly, an increase in the percentage of subjects with physiological sebum content was observed in both groups at T1, although this difference was not statistically significant (p > 0.05, chi-square test, Table 3).

After 2 months of treatment the skin elasticity was instrumentally measured (Figure 4). We could observe an increase in the elasticity score of both groups (Group A and Group B), indicating an improvement in this parameter, although no statistical difference was observed at T1 between Group A and Group B.

The overall efficacy of the two treatments was assessed at the end of the study through a questionnaire completed by the participants, evaluating their subjective perception of four parameters: moisturizing effect, smoothing effect, skin tone uniformity, and brightening effect.

The subjective evaluation confirmed the overall efficacy of both treatments across all assessed parameters. A high percentage of participants rated the effects as ‘excellent’ or ‘good’ in both groups. In particular, the moisturizing effect received the highest satisfaction, with 100% of Group A and 100% of Group B reporting it as excellent or good, demonstrating the high efficacy of both treatments in improving skin hydration results that highlight the efficacy of Jalubalance^® technology in moisturizing the skin.

The smoothing effect was also positively evaluated, with 59% (Group A) and 58% (Group B) rating it as excellent, and an additional 35% and 40% as good, respectively.

The skin tone uniformity evaluation showed slightly lower ratings but remained positive overall (94% Group A and 95% Group B reported the treatments as excellent or good), while the brightening effect was well perceived in both groups, with 98% (Group A) and 97% (Group B) reporting excellent or good results. Reports of ‘poor’ or ‘no effect’ were minimal, indicating a high level of participant satisfaction within both groups.

Figure 5 reports, as an example, the VISIA^® results for different skin parameters (wrinkles and brown spots) evaluated using both standardized lighting and cross-polarized lighting at baseline (T0) and after 8 weeks of treatment (T1) in a subject from Group B (HA + Vitamin C cream). The results are expressed as percentile scores using, as a comparison, a database of skin features of a group of people of the same age and skin type. The 50% percentile represents the average of the reference group; higher scores are associated with an improvement in terms of skin condition. Notably, both parameters showed visible improvement, including a reduction in brown spots, which can be attributed to the brightening and depigmenting action of Vitamin C in the formulation.

Correlation and Linear Regression Analysis

At baseline, Spearman’s rank correlation revealed a significant negative association between Glogau score and skin hydration (ρ = −0.326; p = 0.0025; N = 84) as well as skin elasticity (ρ = −0.423; p < 0.0001; N = 83). No significant correlation was observed with baseline sebum levels (ρ = −0.158; p = 0.151). When evaluating changes from baseline to week 8, significant negative correlations were found between ΔGlogau and Δhydration (ρ = −0.534; p = 3.3 × 10⁻⁷; N = 80), Δsebum (ρ = −0.367; p = 0.00075; N = 81), and Δelasticity (ρ = −0.374; p = 0.00059; N = 81). In all cases, increases in hydration, elasticity, or sebum content were associated with reductions in Glogau score, indicating clinical improvement in photoaging severity.

In a multivariate linear regression model with ΔGlogau as the dependent variable (R² = 0.446; overall p < 0.000001; N = 79), changes in hydration (β = −0.0258; 95% CI [−0.040; −0.012]; p = 0.001) and changes in sebum (β = −0.0328; 95% CI [−0.046; −0.020]; p < 0.0001) emerged as independent predictors of greater Glogau score reduction. No significant associations were observed for changes in elasticity (β = −0.0727; p = 0.250), age (β = −0.0105; p = 0.219), or treatment group (β = 0.062; p = 0.522) after adjusting for the other covariates in the model.

4. Discussion

Skin hydration plays a crucial role in maintaining healthy skin, skin’s physiological functions, and appearance [32,33]. Cosmetic ingredients are widely used in the population to restore physiological hydration and reduce trans epidermal water loss, contributing to an improved skin appearance and tactile properties [34].

Hyaluronic acid (HA) is widely recognized for its anti-aging effects on the skin. Its efficacy is based on its hydrating properties and its ability to improve skin elasticity, reduce wrinkles, and promote tissue regeneration. HA retains large amounts of water at the cellular level, keeping the skin hydrated. It also supports the production of collagen and elastin, and exerts antioxidant and anti-inflammatory properties, protecting the skin from damage caused by environmental factors (exposome) [17,21,35,36].

The efficacy of HA-containing creams and serums in improving skin hydration, reducing wrinkle depth, and enhancing skin smoothness has been well established in clinical research trials. A previous clinical trial investigated the moisturizing and skin barrier effects of a hydrating fluid containing hyaluronic acid (1%), glycerin (5%), and stem cell extract from Centella asiatica. The study, a randomized intra-subject design with blinded assessments, was conducted on 20 healthy women with a mean age of 40 years. The effects of the fluid were measured by using a Corneometer and a Vapometer to evaluate skin hydration and trans epidermal water loss (TEWL), respectively [37]. The results showed a significant increase in skin hydration lasting up to 24 h after a single application of fluid, with a 29% increase compared to baseline at 24 h. Additionally, TEWL was significantly reduced at the treated site compared to the control site, indicating an improvement in skin barrier function. A single application of the fluid demonstrated a long-lasting moisturizing and skin barrier-protective effect [37].

Jalubalance^® is an advanced and exclusive technology based on high-concentration hyaluronic acid (1%) delivered in Opuntia oil.

This clinical study demonstrated that Jalubalance^® technology, present in both the creams used for the treatments, significantly improves several signs of skin aging, including Glogau score, wrinkle depth, skin elasticity, skin tone uniformity, and hyperpigmentation. Instrumental and clinical findings confirmed that both products offer comparable efficacy in counteracting the visible signs of skin aging.

However, a significantly greater reduction in the hyperpigmentation score was observed in Group B (HA + Vitamin C cream) compared to Group A (HA-only cream, p < 0.05). This result may be related to the presence of Vitamin C in the formulation of the cream used for Group B. Vitamin C is a scientifically supported, multi-functional ingredient in anti-aging products. It supports collagen synthesis, protects against oxidative stress [22] and improved brightness, and lightening of skin [22,38]. Vitamin C inhibits the enzyme tyrosinase, reducing melanin production and helping to lighten hyperpigmentation [30,31].

In the present analysis, correlation data demonstrated that improvements in Glogau score were strongly and independently associated with changes in hydration and sebum levels, while baseline and changes in elasticity, although significantly correlated in bivariate analyses, did not retain significance in multivariate models. Interestingly, treatment group was not an independent predictor of ΔGlogau once physiological parameters were included, suggesting that the clinical benefits of the HA + Vitamin C formulation may be largely mediated through its effects on skin hydration and sebum regulation rather than a direct standalone effect of the active combination. The independent role of sebum changes as a predictor of clinical improvement—although less commonly reported—may reflect the contribution of an optimized hydrolipidic balance to skin smoothness and the visual perception of aging.

Some limitations should be considered in evaluating our study results. The main limitation was that this study is an open trial. However, to increase the internal validity of our results, we employed objective instruments to assess patients’ skin, which enhances the reliability of the findings by reducing potential operator error or subjective bias. Another limitation was the relatively short duration of the experiment, which may have limited our ability to observe long-term outcomes of the intervention. However, previous research on hydrating fluid containing 1% hyaluronic acid and 5% glycerin demonstrated a significant increase in skin hydration lasting up to 24 h after a single application [37]. To ensure the integrity of the results, measures were taken to minimize potential sponsor-related bias, such as independent statistical analysis. The role of the sponsor was limited to funding support and study design and did not influence data collection and analysis. These considerations provide confidence in the validity of our results, despite the constraints mentioned above.

5. Conclusions

The clinical and instrumental results of this study confirmed the moisturizing effectiveness of Jalubalance^® technology and the clinical efficacy of both products (Jaluronius cream and Jaluronius cream C) in reducing signs of skin aging, including wrinkle reduction, decreased hyperpigmentation, and improved firmness and elasticity, in subjects with mild-to-moderate skin aging.

Furthermore, the presence of Vitamin C in Jaluronius cream C contributed to the product’s brightening effect, as evidenced by a statistically greater reduction in hyperpigmented spots compared to the control group.

The findings of this study confirm that both creams represent highly effective anti-aging treatments. These results support the potential for personalized therapeutic approaches, allowing dermatologists to select the most appropriate formulation based on the specific clinical characteristics and pattern of skin aging in each patient.