Clinical Safety and Efficacy Evaluation of a Dissolving Microneedle Patch Having Dual Anti-Wrinkle Effects With Safe and Long-Term Activities

Abstract

Background

Anti-aging products are widely used, but the desire for safe and more efficient anti-aging products continues to increase. Dissolving microneedle patches (MNPs) have provided a more efficient transdermal drug delivery solution. MNP is a promising candidate for developing better anti-aging products.

Objective

To develop a more efficient anti-aging MNP product, we fabricated a dual anti-wrinkle microneedle patch (named DA-MNP) using droplet extension (DEN^®) technology and evaluated its skin puncture ability, safety, and efficacy through clinical studies.

Methods

A DA-MNP comprising hyaluronic acid (HA) polymer backbone, acetyl octapeptide-3, and L-ascorbic acid 2-glucoside and sodium cyclic lysophosphatidic acid was fabricated using DEN^® technology. Placebo MNPs comprising only HA were also fabricated. Twenty-four healthy subjects were enrolled in this comparative clinical study. The DA-MNP or placebo MNP was separately applied to the left and right eyes of subjects for overnight. Assessments, including wrinkle improvement, trans-epidermal water loss (TEWL), eye lifting and adverse effects were evaluated at each scheduled visit day for 28 days.

Results

The DA-MNP showed mechanical strength enough for puncturing the stratum corneum. Compared to placebo MNP group, the DA-MNP treated group showed an effective eye wrinkles improvement and better anti-aging of skin, with reduced TEWL, enhanced skin elasticity and lifting, and no adverse effects.

Conclusion

The present study demonstrated that the fabricated DA-MNP exhibited fast acting on deep wrinkles and enhanced anti-aging efficacy, with no skin safety concern. Thus, this DA-MNP may serve as a new transdermal delivery solution for skin wrinkling and aging.

INTRODUCTION

Recent transdermal drug delivery advances include various devices (employing processes, such as iontophoresis, sonophoresis, electroporation, and photomechanical waves delivery), and various skin-application formulations (such as, ointments, creams, gels, liposomes, and emulsions)¹. However, although many of these devices exhibit exceptional drug delivery efficiency, some of them are somewhat less user friendly and expensive devices. Moreover, many conventional skin-application formulations, such as creams, showed lower delivery efficiency due to the skin barrier. Thus, existing products fail to fully address the needs of users².

One of the primary obstacles to efficient drug absorption through the skin (transdermal drug delivery) is the stratum corneum, the outermost layer of the skin. It selectively permits only small molecules (<500 Da) and lipophilic materials to pass through while inhibiting the absorption of water soluble and high molecular weight substances. Thus, many active molecules, such as water soluble and high molecular weight substances, which are commonly utilized in the pharmaceutical industry are not well absorbed through the skin. Accordingly, developing more efficient and effective transdermal delivery technologies is critical to overcoming this challenge³.

Dissolving microneedle patches (MNPs) have emerged as a promising advancement in transdermal drug delivery. On application, the MNP physically penetrates the skin barrier and creates microchannels in the stratum corneum. Subsequently, the skin moisture gradually dissolves the microneedles, releasing the formulated drugs and promoting their absorption. This mechanism facilitates efficient delivery of high molecular weight peptide drugs and hydrophilic drugs by enabling skin barrier penetration⁴,⁵.

Skin aging is characterized by wrinkling, caused by loss of skin moisture, elasticity, and firmness. Various intrinsic (genetics, hormone balance, and cellular metabolism) and extrinsic (environmental) factors contribute to skin aging. Identifying efficient functional ingredients that can antagonize wrinkle generation is crucial to improving skin wrinkles.

However, inadequate absorption of functional ingredients, across the skin barrier, the anti-aging product may have insignificant wrinkle improvement effects. Further, despite being well absorbed, fundamental wrinkle improvement, by activating collagen or hyaluronic acid (HA) synthesis in the skin, requires long-term application of the anti-aging product⁶.

Here, to overcome these issues and achieve a more rapid wrinkle improving anti-aging product having long-term skin improvement effects, we developed a microneedle patch (named DA-MNP) having dual anti-wrinkle effects with safe and long-term activities. Firstly, to ensure efficient delivery of the selected anti-aging functional ingredients and minimize skin irritation, we selected the microneedle length that can create microchannels with minimal skin invasion. Secondly, we selected 3 face-specific anti-wrinkle and anti-aging functional ingredients. The first functional ingredient, acetyl octapeptide-3 (SNAP-8^TM), was a botulinum toxin analog which acts by blocking neurotransmitters and relaxing muscles to improve wrinkles. SNAP-8^TM is composed of 8 amino acids (Ac-Glu-Glu-Met-Gln-Arg-Arg-Ala-Asp-NH₂) and reportedly influences muscle contraction resulting in a more relaxed and line-free face, especially around the forehead and the eyes⁷. The second functional ingredient, L-ascorbic acid 2-glucoside (AA2G), was a stabilized ascorbic acid (Vitamin C) derivative, which is activated by D-glucosidase in the cell membrane. Basically, wrinkles are caused by lack of regeneration of connective tissues which fill the extracellular matrix (ECM). AA2G promotes the synthesis of collagen, which is one of the main components of ECM connective tissue, and has been reported to exhibit longer-term antioxidant efficacy in skin⁸,⁹,¹⁰. Therefore, AA2G was selected as a functional ingredient for wrinkle improvement. The third functional ingredient used in this study was sodium cyclic lysophosphatidic acid (NcPA) which is known to increase skin moisture and ECM-fillers via upregulating synthesis of HA (a major ECM connective tissue)¹¹,¹². Skin application of this fabricated MNP (DA-MNP) loaded with these 3 functional ingredients was expected to show a wrinkle improvement with long-term anti-aging effects because of high delivery efficiency of active ingredients by MNP’s penetration through the stratum corneum barrier.

To prove this assumption, we conducted skin safety clinical studies to determine the possibility for its regular use and a comparative clinical study for assessment of anti-wrinkle effects with safe and long-term activities using the fabricated DA-MNP and placebo MNP.

MATERIALS AND METHODS

Fabrication of MNPs by droplet extension (DEN^®) manufacturing method

HA polymer was dissolved in water and mixed using a planetary centrifugal mixer (ARV-310; THINKY Corporation, Tokyo, Japan). Subsequently, SNAP-8^TM (MW 105.16, water solubility 10 mg/ml; Incospharm Corp., Daejeon, Korea), AA2G (MW 338.26, water solubility 879 mg/ml; Hayashibara Co., Ltd., Okayama, Japan), and NcPA (MW 400.00, water solubility 50 mg/ml; Sansho Co., Ltd., Osaka, Japan) were added sequentially to the HA-polymer mixture and mixed using the planetary centrifugal mixer. The final blended mixture was dispensed on the hydrocolloid adhesive patch as precision-controlled droplets. The droplets on the hydrocolloid adhesive patch were contacted with another hydrocolloid adhesive patch and extended to target lengths (350, 500, and 800 μm) for the DA-MNP and placebo MNPs. The DA-MNP and placebo MNPs were air dried and separated by blowing room temperature air. The finalized DA-MNP and placebo MNP products were packed in aluminum and polyethylene terephthalate blister pouches to conserve humidity and protect them from physical damage. Fabricated microneedle characteristics (microneedle length, shape, and tip diameter) were microscopically observed (13.4×–40× magnification, SMZ 745T; Nikon, Tokyo, Japan).

Measurement of skin penetration depth

For this study, we purchased a minipig skin consisting of commercialized whole skin components (stratum corneum, epidermis, dermis layers) from Apures, Pyeongtaek, Korea (https://www.apures.com). The DA-MNP and placebo MNPs (350, 500, and 800 μm long microneedles) were applied to minipig skin (800 μm thickness) for few seconds and removed. The tissue was sectioned and stained with hematoxylin and eosin (H&E, BBC Biochemical, Mount Vernon, WA, USA) according vendor recommended method. The skin penetration depth of MNPs was microscopically measured (10×–20× magnification, ECLIPSE Ni-U; Nikon).

Comparative clinical study design

Comparative clinical study was approved and conducted at P&K Korean Skin Research Center (Seoul, Korea) with the approval by P&K Korean Skin Research Center Institutional Review Board (PNK-17807-K1R). Twenty-four subjects with eye wrinkles (aged 30–65) were chosen for the study. The DA-MNP or placebo MNP groups were applied to the left and right eyes, separately. MNPs were applied once a day at night for 14 days, and then applied once every 3 days for the next 14 days (until day 28). Observational visits were scheduled at various intervals after the application, including before (0 day), day 1 (with a 2-hour application), day 3, day 7, day 14, and day 28 (with overnight application). Any adverse events (ADEs) were observed on each visit. ADEs were evaluated on erythema, edema, and scaling by clinical researchers. Itching, stinging, burning, tightness, and prickling were surveyed by clinical participants.

Quantitative evaluation of wrinkle improvement

For skin roughness (wrinkle) assessment, wrinkle assessment 1: R1 (skin roughness), R2 (maximum roughness), R3 (average roughness), R4 (smoothness depth), and R5 (arithmetic average roughness), was quantitatively analyzed using Skin-Visiometer SV 600 (Courage-Khazaka Electronic GmbH, Köln, Germany). Wrinkle assessment 2: Ra (Average roughness), Rz (Roughness), Rp (Peak height), Rv (Valley depth), was also quantitatively analyzed using PRIMOS CR (Canfield Scientific Inc., Parsippany, NJ, USA).

Overall skin conditions

Trans-epidermal water loss (TEWL) was measured using VapoMeter (Delfin Technologies Ltd., Kuopio, Finland) for evaluating water loss of skin (g/h·m²), which makes it possible to predict skin water retention capacity. The average of triplicate measurements was reported. Skin elasticity of the eye rim was calculated through R2 value measured by Cutometer CM580 (Courage-Khazaka Electronic GmbH). The R2 value closer to 1 represents better elasticity.

For measuring eye lifting, the subjects were photographed by F-ray’s Topography which indicates contour on skin. It was analyzed by Image-pro^® plus (Media Cybernetics, Rockville, MD, USA). The same area was measured in “before” and “after use.” Eye lifting was evaluated by the angle between 2 lines: one was drawn as parallel line from eye-head and another was the curve at the end of eye-tail. The smaller degree between 2 lines indicates the skin lifting improvement.

Statistical analysis

The statistical analysis package SPSS 19.0 (IBM, Armonk, NY, USA) was used to evaluate the efficacy of DA-MNP in affecting skin changes. The significance level was set at 5% (p<0.05 was statistically significant). Probability was rounded to 3 decimal places. Continuous variables were summarized as average with standard deviation and categorical variables were summarized as frequency with percentage. For “before” and “after use” comparison, the parametric test, paired t-test and the non-parametric test, and Wilcoxon signed rank test were used according to the statistical treatment for analysis. For repeated measurement, the parametric test, repeated measures analysis of variance (post hoc: Bonferroni correction) and the non-parametric test, Friedman test (post hoc: Wilcoxon signed rank test with Bonferroni correction) were used according to the statistical treatment for analysis. For comparison between DA-MNP and placebo MNP, change rate was used as evaluation data. The statistical analysis was conducted using both parametric and non-parametric tests. The independent t-test, a parametric test, was used to compare the means of 2 independent groups, while the Mann-Whitney U test, a non-parametric test, was used to compare the medians of 2 independent groups.

RESULTS

Characteristics of the fabricated MNP (DA-MNP)

The DA-MNP was made using DEN^® method. The fabricated DA-MNP had a cone shape with a concave form from the lower end to the upper end. The results showed that the diameter of the lowest end of the fabricated DA-MNP was 800–900 μm, the middle was 250–300 μm, and the controlled tip diameter was <60 μm. The total number of microneedles on a patch was 140 (Fig. 1). Composition of the DA-MNP was determined to be 5% AA2G, 4% NcPA, 0.03% SNAP-8, and 90% HA. The DA-MNP length was approximately 350 μm. Placebo MNPs consisted of 100% HA with 3 different microneedle lengths (350, 500, and 800 μm).

Fig. 1. The fabricated microneedle patch (DA-MNP) product. (A) Schematic diagram of microneedle patch production by droplet extension. (B, C) Microscopic images (13.4× magnification). (D) Microscopic image (40× magnification). (E) Specification of DA-MNP: the base diameter, 800–900 μm; the middle, 250–300 μm; tip diameter, 60 μm; the length of microneedles, 350 μm. Each microneedle was cone shaped.

DA-MNP: dual anti-wrinkle microneedle patch.

Skin puncture ability and penetration depth of DA-MNP and MNPs

The mechanical strength of the DA-MNP was 0.25 N. It was reported that the key factors, namely the length, shape, and pitch of each microneedle, importantly determine the skin penetration ability of MNPs¹³. Since it was previously reported that a minimum mechanical strength of 0.1 N is required for skin puncture of dissolving microneedles¹⁴, the observed mechanical strength of our DA-MNP, 0.25 N, suggested that the fabricated DA-MNP exhibits enough mechanical strength to penetrate the skin.

On the other hand, microneedles may generally have different degrees of penetration depth according to the elasticity of the skin¹⁵,¹⁶. As shown in Fig. 2, the results of penetration test of MNPs in the present study showed most microneedles penetrated the micropig skin by about 60% of their lengths. Placebo MNPs with lengths 350 and 500 μm reached the epidermis and epidermis–dermis junction, respectively. The 800 μm placebo MNP reached far deeper into the dermis layer with confirmed penetrance of about 500 μm (Fig. 3). These results suggested that skin penetration depth can be controlled by varying microneedle lengths. Based on these results, we decided the length of DA-MNP fabricated by DEN^® to be approximately 350 μm.

Fig. 2. Skin penetration depth of MNPs depending on microneedle length. (A) After applying MNP to micropig skin, the puncture site of microneedle and skin was cut vertically, and the cross section was observed for penetration depth with a microscope (magnification:10×). (B) H&E staining image of the micropig skin after application of the 350 μm long MNP (magnifications: left 10×, right 20×). (C) H&E staining image of the micropig skin after application of the 500 μm long MNP (magnifications: left 10×, right 20×). (D) H&E staining image of the micropig skin after application of the 800 μm long MNP (magnifications: left 10×, right 20×), showing the MNP tip reached the dermis layer.

MNP: microneedle patch, H&E: hematoxylin and eosin.

Fig. 3. The results of eye wrinkle improvement assessment 1 after DA-MNP and placebo MNP application: R1, skin roughness; R2, maximum roughness; R3, average roughness; R4, smoothness depth; R5, arithmetic average roughness.

DA-MNP: dual anti-wrinkle microneedle patch, MNP: microneedle patch.

^*p<0.05 by repeated measure analysis of variance, post hoc Bonferroni correction; ^**p<0.05 by independent t-test.

^#p<0.025 (=5%/2) by post hoc Wilcoxon sign test with Bonferroni correction.

Skin safety is very important for cosmetic products because they are used daily. Thus, since our 350-μm MNP sufficiently penetrated the stratum corneum, we decided the appropriate DA-MNP length to be 350 μm, to confer our DA-MNP with improved capabilities for more efficient and safer transdermal drug delivery via formation of microchannels in stratum corneum.

DA-MNP application showed rapid improvement of wrinkle and overall skin condition

A total of 24 subjects were initially enrolled in the study, but 3 subjects dropped out, resulting in a final 21 subjects who completed the study. Out of the 21 subjects, 13 had dry skin, 2 had dry to normal skin, 5 had normal skin, and 1 had oily skin. Two different evaluations, namely eye wrinkle assessments 1 and 2, were used to assess eye wrinkle improvement. The results of eye wrinkle assessment 1 were comprehensively summarized in Fig. 3. The skin roughness (R1) and average skin roughness (R3) parameters in the DA-MNP treated group showed a significant decrease (p<0.025) on 14th and 28th day of DA-MNP application. On the other hand, maximum roughness (R2), smoothness depth (R4), and average arithmetic roughness (R5) parameters showed a significant decrease (p<0.025–0.05) on day 28 of DA-MNP application. All roughness evaluation parameters (R1–R5) were significantly different between DA-MNP and placebo groups (p<0.05) on 28th day of application. Although the R2 and R3 parameters in the placebo group showed a tendency to decrease, the decrease was not statistically significant (Fig. 3).

The results of eye wrinkle assessment 2 (Ra, Rz, Rp, and Rv parameters) were summarized in Fig. 4. The Ra parameter was significantly decreased after 28 days of DA-MNP application (p<0.01). The Ra parameter before and after placebo MNP application was not significantly different at any time-point during post-application. Compared to the placebo group, the DA-MNP group showed a faster and significant difference (p<0.05), on 14th and 28th day of product application. Meanwhile, the Rz parameter was significantly decreased (p<0.01) on 7th, 14th, and 28th day of DA-MNP application. However, Rz parameter remained constant with the placebo MNP application. The Rz parameter of the DA-MNP group and placebo group was significantly different (p<0.05) on 14th and 28th day of product application.

Fig. 4. The results of eye wrinkle improvement assessment 2 after the DA-MNP and placebo MNP application. (A) Ra, Average roughness. (B) Rz, Roughness. (C) Rp, Peak height. (D) Rv, Valley depth. The values were expressed as the percent changes following an equation below: $[\Delta Change\%=\left[\frac{After-Before}{Before}\right]\times 100]$.

DA-MNP: dual anti-wrinkle microneedle patch, MNP: microneedle patch.

^*p<0.01 (=5%/5) by post hoc Wilcoxon signed rank test with Bonferroni correction; ^**p<0.05 by independent t-test.

^##p<0.05 by Mann-Whitney U test.

The Rp parameter showed a significant decrease (p<0.01) at 2 hours and 28th day of DA-MNP application. But the Rp parameter of the placebo group did not change significantly with placebo MNP application. The Rp parameter DA-MNP group and placebo group was significantly different (p<0.05) on 28th day of product application. Lastly, the within-group Rv parameter of DA-MNP and placebo groups were similar at all observational time-points. However, the Rv parameter of DA-MNP group was significantly different from placebo group (p<0.05) on 28th day of product application. DA-MNP showed wrinkle improvement immediately after the first application, compared to the placebo MNP. Moreover, while significant improvement in Rp (measuring peak height of wrinkle) was seen after just one application of the DA-MNP, significant improvement in Rz (measuring roughness) was seen on the 7th day of application.

Based on the results of eye wrinkle assessment 1 and 2, we concluded that the fabricated DA-MNP has a faster anti-wrinkle effect (with only one or a few applications). The DA-MNP showed far more efficient anti-wrinkle effects than placebo MNP during all the varying durations of applications.

In addition, TEWL was significantly decreased (p<0.05) at 7 and 14 days of product application in the DA-MNP group (Fig. 5A). No change in TEWL was observed in the placebo group. Moreover, TEWL was significantly different (p<0.05) between the DA-MNP and placebo groups, with 7 and 14 days of product application. Thus, the TEWL decrease, allays concerns about skin damage due to skin puncture caused by DA-MNP.

Fig. 5. Comparative results of TEWL, effects of DA-MNP and placebo MNP application on skin elasticity and lifting. (A) DA-MNP group showed a significant decrease in TEWL at 7th and 14th day of product application. The placebo group did not show any significant time-course dependent changes with product application; (B) elasticity and lifting were significantly increased at 14th day of DA-MNP product application. Compared to the placebo group, the DA-MNP group showed a significant difference in the results of the eye-lifting test at 14th day of product application, but no changes were observed in the skin elasticity test results.

TEWL: trans-epidermal water loss, DA-MNP: dual anti-wrinkle microneedle patch, MNP: microneedle patch.

^*p<0.05 by paired t-test; ^**p<0.05 by Mann-Whitney U test.

^#p<0.05 by repeated measures analysis of variance, post hoc Bonferroni correction; ^##p<0.05 by Mann-Whitney U test.

Further, the DA-MNP group showed a significant increase (p<0.05) in skin elasticity and eye-lifting parameters at 14th day of product application (Fig. 5B). However, skin elasticity and eye lifting parameters in the placebo group were not significantly changed. The eye-lifting parameter in the DA-MNP group was also significantly different (p<0.05) compared to the placebo group. Meanwhile, the skin elasticity was similar between the 2 groups (DA-MNP and placebo group). According to the results of skin elasticity and eye lifting tests, the delivery of anti-aging ingredients through microchannels in skin using DA-MNP seems to have excellent facial skin improvement effects, especially, in eye lifting.

A representative image of the time course of eye wrinkles improvement in response to DA-MNP and placebo MNP application, is shown in Fig. 6. The DA-MNP group showed faster wrinkle improvement and continued improvement of overall skin conditions when compared to the placebo group.

Fig. 6. Representative time course of eye wrinkle improvement with application of DA-MNP and placebo MNP (visualized using PRIMOS) over the course of total 28 days: (A) the DA-MNP application group; (B) placebo MNP application group. Wrinkle improvement was confirmed in both groups. However, the DA-MNP group showed better wrinkle improvement compared to the placebo group.

DA-MNP: dual anti-wrinkle microneedle patch, MNP: microneedle patch.

DISCUSSION

Microneedles-based transdermal drug delivery is widely acknowledged as a highly beneficial innovation. Although the microneedles manufacturing methods are different, they have been clinically proven in improving skin wrinkles and skin tone¹⁷,¹⁸. Nevertheless, for daily use products such as cosmetics, concerns still persist regarding the damage caused in the skin due to the process of microchannels creation using microneedles.

Since the penetration depth of microneedles through the skin is dependent on their length, as well as their strength, due to the skin elasticity¹⁹, we first tried to determine the penetration efficiency using dissolving placebo MNPs with different lengths (350, 500, 800 μm). The results showed that while those placebo MNPs with lengths 350 and 500 μm reached the epidermis and epidermis–dermis junction, the 800 μm MNP reached far deeper into the dermis layer, still maintaining appropriate strength of each microneedle. Thus, with the results from the through-skin penetration tests using different microneedle lengths, we decided the length of our DA-MNP to be approximately 350 μm, to avoid possible unwanted damage to the deeper region of the dermis layer, considering the safety concern of the daily use of cosmetic products. Of course, some microneedles with longer length have been developed and reported as clinically safe. For example, a MNP-vaccine clinical trials using microneedles with the length of 600 μm were reported as safe²⁰. However, generally, these MNP-vaccines are not administered on a daily basis and do not need long-term use. Thus, safety report from this study must be interpreted differently for long-term daily use MNP, especially, for skincare MNP products such as our target product (DA-MNP) which must be used daily. In these contexts, we aimed to develop a DA-MNP with minimal skin invasive ability (with a length of 350 μm), thereby the microchannels created by DA-MNP microneedles could effectively deliver functional ingredients in a minimally invasive manner. In this clinical study, no adverse effects were observed despite repeated use in the same skin area for 28 days, indicating that the 350-μm length DA-MNP fabricated using DEN^® technique could be considered to be quite safe and useful for daily use of functional cosmetic products.

On the other hand, conventional cosmetic formulations generally contain SNAP-8^TM, AA2G, or NcPA, as efficient anti-wrinkling ingredients. However, reported conventional cosmetic formulations such as cream and topical serums verified to require to take time between 4 to 12 weeks for anti-wrinkle effectiveness²¹,²²,²³. Because conventional products generally require several weeks of use to improve wrinkles, consumers are still demanding not only faster wrinkle improvement, but also safer functional products. In this context, we took care to design a dual-acting MNP (DA-MNP) for an effective eye wrinkles’ improvement and better anti-aging of skin than conventional functional cosmetics. Thus, in the present study, we formulated our DA-MNP by combining these all 3 compounds together to increase the anti-wrinkle effectiveness of DA-MNP.

As confirmed by the present comparative clinical study, our DA-MNP showed relatively rapid wrinkle care results; the Rp (peak wrinkle height) value of the eye wrinkles decreased significantly after a single use. Furthermore, a significant decrease in Rz (roughness) value was observed within 7 days of use and decrease in Ra (average roughness) value was observed within 14 days of use. The deep wrinkle improvement observed after a single use appears to be due to the short-term effect of SNAP-8’s muscles relaxing mechanism, and subsequent long-term wrinkle improvement and anti-aging effects could be due to the complex actions of AA2G and NcPA which are involved in synthesis of several major ECM components, such as collagen and HA. Meanwhile, an improvement in wrinkle reduction, skin elasticity, and eye-lifting effect was observed within 28 days of use. Additionally, skin moisture loss typically caused by the physical skin puncture and invasiveness of microneedles was not observed in this comparative clinical study. Moreover, ADEs such as erythema, edema, and scaling on the skin site where the product was applied were not observed at all visits. Itching, stringing, burning, fitness, and pickling were not reported by all subjects. Although the skin safety concern generally needs to be confirmed through a larger clinical study, the clinical study results in the present study suggest that our new type of MNP product (DA-MNP) did not evoke any significant skin safety concerns.

In summary, the DA-MNP appeared to show an effective wrinkle improvement capability compared to previously reported conventional formulations in terms of an excellent percutaneous delivery efficiency of functional ingredients through minimally skin invasive microchannels. Additionally, the present clinical study demonstrated the skin safety of our DA-MNP. And the skin safety of active ingredients and the DA-MNP itself was secured. Thus, the DA-MNP loaded with anti-aging ingredients can be a new type of topical delivery solution to treat or prevent the skin wrinkling and aging.