# Blue-light activated molybdenum cluster-based photosensitizers in NaDES – formulated creams for dermatological photodynamic therapy

Michaela Kubáňová, Kaplan Kirakci, Leslie Boudesocque-Delaye, Laura Divoux, Antonín Kaňa, Jaroslav Zelenka, Tomáš Ruml, Emilie Munnier, Kamil Lang, Alexis Verger

<figure><img src="image_1.png" /></figure>

## ► To cite this version:

Michaela Kubáňová, Kaplan Kirakci, Leslie Boudesocque-Delaye, Laura Divoux, Antonín Kaňa, et al. Blue-light activated molybdenum cluster-based photosensitizers in NaDES – formulated creams for dermatological photodynamic therapy. International Journal of Pharmaceutics, 2025, 689, pp.126491. 10.1016/j.ijpharm.2025.126491. hal-05510929

HAL Id: hal-05510929

https://hal.science/hal-05510929v1

Submitted on 14 Feb 2026

HAL is a multi-disciplinary open access
archive for the deposit and dissemination of sci-
entific research documents, whether they are pub-
lished or not. The documents may come from
teaching and research institutions in France or
abroad, or from public or private research centers.

L'archive ouverte pluridisciplinaire HAL, est
destinée au dépôt et à la diffusion de documents
scientifiques de niveau recherche, publiés ou non,
émanant des établissements d'enseignement et de
recherche français ou étrangers, des laboratoires
publics ou privés.

<figure><img src="image_2.png" /></figure>

Distributed under a Creative Commons CC BY 4.0 - Attribution - International License
# Blue-light activated molybdenum cluster-based photosensitizers in NaDES – formulated creams for dermatological photodynamic therapy

Michaela Kubáňová<sup>a,b ID</sup>, Kaplan Kirakci<sup>b</sup>, Leslie Boudesocque-Delaye<sup>c ID</sup>, Laura Divoux<sup>c,d</sup>, Antonín Kaňa<sup>e ID</sup>, Jaroslav Zelenka<sup>a</sup>, Tomáš Ruml<sup>a</sup>, Emilie Munnier<sup>d ID</sup>, Kamil Lang<sup>b</sup>, Alexis Verger<sup>d,* ID</sup>

Check for
updates

## ARTICLE INFO

Keywords:

Photodynamic therapy

Topical drug delivery

Molybdenum clusters

Natural deep eutectic solvents (NaDES)

Semi-solid cream formulation

Skin microbiome

Antimicrobial resistance

## ABSTRACT

Photodynamic therapy (PDT) emerges as an innovative strategy combining light and drug treatment. The most prevalent use of this therapy is for the elimination of cancerous and pre-cancerous cells. However, in recent years PDT has been shown to be a suitable approach to combat the rising threat of antimicrobial resistance through the photoinactivation of pathogenic bacteria. Photosensitizers predicated on octahedral hexa-molybdenum cluster (Mo₆) complexes activated by blue light were formulated into semi-solid topical pharmaceutics containing three distinct types of Mo₆ clusters as potential novel actives for cosmetics purposes. For the first time, semi-solid cosmetic formulations were developed utilizing natural deep eutectic solvents as carriers of these active compounds and tested for their stability, viscosity, skin penetration, and singlet oxygen formation. After that, negligible toxicity for human cells was confirmed using in vitro testing on HaCaT cell lines. Finally, photoinactivation of the most prevalent skin bacterial pathogens *Staphylococcus aureus* and *Cutibacterium acnes* was demonstrated under mild conditions that preserve viability of natural symbiotic skin microflora.

## 1. Introduction

The escalating threat of antimicrobial resistance (AMR) represents a significant global health concern. Conventional antibiotics, previously the mainstay of infectious disease management, are exhibiting diminishing efficacy against an expanding array of resistant pathogens. One of the contributing factors being the utilization of antibiotics for dermatological conditions caused by microbial pathogens such as *Cutibacterium acnes* (former *Propionibacterium acnes*). Topical and systematic antibiotics, as well as over-the-counter antibiotic formulations are commonly used as primary treatments for severe forms of acne vulgaris, consequently leading to the emergence of antibiotic-resistant strains. This phenomenon underscores the urgent need for the development of more non-antibiotic therapeutic alternatives (Dessinioti and Katsambas, 2022). In addition to acne, the opportunistic pathogen *Staphylococcus aureus* is commonly associated with inflammatory skin conditions such

as dermatitis and atopic dermatitis, impetigo or abscess resulting in severe cutaneous lesions (Alexander et al., 2020).

Resistant strains of this pathogen are also widely prevalent among patients, especially in hospital settings (Cavalcante et al., 2021).

Photodynamic therapy (PDT) demonstrates tremendous potential in eradicating skin pathogens. Its action mechanism relies on utilizing light-sensitive compounds known as photosensitizers which, when activated by specific wavelengths of light generate reactive oxygen species (ROS), that induce a cytotoxic response by causing lipid peroxidation, protein oxidation and DNA damage (Verger et al., 2021). Initially developed for treating certain types of cancers, its application has expanded beyond oncology, showing promise in combating microbial infections and addressing the global challenge of antimicrobial resistance. This versatile approach leverages the unique interactions between light, photosensitizers, and biological tissues, offering a non-invasive, targeted treatment option with minimal side effects and little

<footnote><sup>a</sup> Department of Biochemistry and Microbiology, University of Chemistry and Technology Prague 166 28 Prague, Czech Republic</footnote>

<footnote><sup>b</sup> Institute of Inorganic Chemistry of the Czech Academy of Sciences, 250 68 Husinec-Rež, Czech Republic</footnote>

<footnote><sup>c</sup> Université de Tours, UR 7502 SIMBA, Faculté de Pharmacie, 31 avenue Monge, 37200 Tours, France</footnote>

<footnote><sup>d</sup> Université de Tours, CNRS UPR 4301 CBM, Département NMNS, Tours, France</footnote>

<footnote><sup>e</sup> Department of Analytical Chemistry, University of Chemistry and Technology Prague, 166 28, Prague, Czech Republic</footnote>

<footnote>* Corresponding author at: Faculté de pharmacie de Tours - Laboratoire de Pharmacie Galénique, Département NanoMédicaments et NanoSondes, Centre de Biophysique Moléculaire UPR CNRS 4301, 31, avenue Monge, 37200 Tours, France.
E-mail address: alexis.verger@univ-tours.fr (A. Verger).</footnote>
to no proclivity to induce resistance within microbial populations (Dias et al., 2022; Maisch, 2015, 2009). Topical PDT has been previously established using 5-aminolevulinic acid or methyl aminolevulinate (Keyal et al., 2016; Wiegell and Wulf, 2006; Yang et al., 2021). Blue light has been demonstrated as an effective treatment for mild to moderate acne vulgaris, due to the excitation of bacterial porphyrins acting as photosensitizers (Diogo et al., 2021; Pei et al., 2015).

To improve the phototoxic efficiency of this treatment, octahedral molybdenum-halogen cluster complexes ($Mo_6$) are interesting alternative to traditional porphyrins. When excited by X-ray, UV and visible light up to 550 nm, $Mo_6$ complexes generate high levels of singlet oxygen, $O_2(^1\Delta_g)$, making them exemplary type II photosensitizers for the blue light PDT (Kirakci et al., 2013,2012). Their properties such as hydrophobicity, membrane targeting and yields of $^1O_2$ can be modulated by the choice of the six outer ligands. Phototoxic effects of these compounds have been previously demonstrated in both anti-cancer PDT (Pribyl et al., 2024; Verger et al., 2023,2022) as well as photo-inactivation of bacteria (Felip-León et al., 2017; Guégan et al., 2023; Kirakci et al., 2020a; Vorotnikova et al., 2019). Usually, these clusters are soluble only in organic solvents (Kirakci et al., 2019b), which are generally not appropriate to be used in cosmetic products (Boudesocque-Delaye et al., 2024). Thus, our aim was to incorporate these compounds as a part of an oil phase using natural deep eutectic solvents (NaDES), a green alternative to traditional solvents. NaDES in the final semi-solid formulation were also intended to protect $Mo_6$ clusters from hydrolysis and subsequent loss of biological activity.

NaDES are composed of naturally occurring metabolites such as organic acids, amino acids or sugars. Due to their strong hydrogen bonding and favorable sensory properties, NaDES represent a safe and viable alternative to traditional organic solvents (Aduloju et al., 2023). NaDES are created as a eutectic mixture of two constituent components – hydrogen bond donor and acceptor – that, when combined, exhibit a melting point lower than that of each individual component. Depending on the constituents, NaDES can be either hydrophilic or hydrophobic (Kua and Gan, 2019). Consequently, they have been explored as drug carriers and effective solvents, particularly suited for hydrophilic compounds (Jelinski et al., 2019). NaDES also exhibit skin-conditioning properties and enhance the delivery of active ingredients (Boudesocque-Delaye et al., 2024; Rente et al., 2022; Verger et al., 2024). The most used NaDES are based on choline-chloride, however, their use is restricted in cosmetics due to the EU regulations (UNION,

2009). Therefore, NaDES based on fatty acids and 1,3-propanediol were chosen for this study to solubilize the cluster compounds. Moreover, thanks to the free fatty acid content, they can contribute to the skin microbiome-modulating and antibiotic effects of the active component (Desbois and Smith, 2010).

Herein, we report for the first time the successful solubilization of $Mo_6$ clusters in natural deep eutectic solvents (NaDES), thereby unveiling their potential for topical cosmetic applications in photodynamic therapy (PDT) through NaDES-based formulations. Three distinct types of outer ligands were used: phosphinate (1) ($Na_2[Mo_6I_8(O-POPh_2)_6]$, phosphonium (2) [$Mo_6I_8(OCOC_4H_8PPh_3)_6Br_4$, and cholate (3) $Na_2[Mo_6I_8(olate)_6]$) (Fig. 1). Final formulations were characterized by their physical and photophysical properties and tested for biological safety and phototoxicity against skin pathogens in vitro.

## 2. Materials and methods

### 2.1. Cluster compounds preparation

$Mo_6$ cluster compounds with selected ligands were prepared similarly as previously reported by reaction of $Na_2[Mo_6I_8(OMe)_6]$ with the corresponding acids (Kirakci et al., 2020b,2019b,2019a).

### 2.2. Blank cream formulation

The formulation was initially developed for a 100 g batch size using the phase inversion temperature (PIT) method, employing a VMI rotor-stator homogenizer. The beaker containing oil phase (B1) consisted of 20.0 g *Helianthus annuus* seed oil, 2.5 g cetyl alcohol, and 2.5 g glyceryl stearate and the beaker with aqueous phase (B2) comprised 73.75 g deionized water and 0.25 g preservative were separately heated to 70 °C until complete melting of solid components was observed. The oil phase was transferred to the VMI rotor-stator homogenizer vessel, and initial homogenization was set at 1000 rpm. The aqueous phase was introduced gradually into the oil phase under continuous agitation. Upon complete addition of B2, the rotor speed was increased to 1500 rpm and maintained for 10 min to ensure thorough emulsification. Subsequently, 1.0 g of Amigum® was incorporated as a gelling agent. The rotor speed was further increased to 1800 rpm and maintained for additional 10 min to ensure uniform dispersion of the gelling agent. The final emulsion was transferred to a 100 mL plastic container for storage

<figure><img src="image_3.png" /><figcaption>**Fig. 1.** Schematic representation of molybdenum cluster complex [$Mo_6I_8L_6$]$^n$ and selected ligands (L).</figcaption></figure>
and stability testing.

## 2.3. SpeedMixer® scale-down

For small-scale prototyping, the preparation was adapted to a Hauschild SpeedMixer® dual asymmetric centrifugal mixing system with the use of magnetic stirring wheels. This dual asymmetric centrifugal mixing technique allows for rapid and efficient homogenization of multi-component systems without the use of blades.

Per one batch, 25 g of cream was formulated per individual sample. The oil phase (B1) consisted of 5 g sunflower oil, 0.625 g cetyl alcohol, and 0.625 g glyceryl stearate. The aqueous phase (B2) comprised 18.44 g distilled water and 0.06 g preservative. Both phases were separately heated to 60 °C in beakers until complete melting of solid components was observed. The heated phases were then transferred to a 100 mL mixing vessel equipped with two magnetic stirring rings. Emulsification was performed using a SpeedMixer® at 2500 rpm for 5 min. Following initial emulsification, 0.25 g of Amigum was incorporated to increase the viscosity and stability of the emulsion. The formulation was then subjected to a second mixing cycle under identical conditions (2500 rpm, 5 min) to ensure uniform distribution of the gelling agent.

For control and cluster sample groups, NaDES-cluster complexes were added after the heating and mixing of B1 and B2 phases, just before the first mixing cycle. All samples were prepared in triplets. To simulate accelerated aging conditions, the samples were stored in a climate-controlled chamber at 40 °C ± 1°C and 75 % ± 5 % relative humidity.

## 2.4. Solubility assay

A mass of 25 mg of the cluster compound was combined with 975 mg of the selected NaDES in a glass vial. A magnetic stir bar was introduced, and sample was subjected to thermal agitation at 50 °C (±0.5 °C) under continuous magnetic stirring for 24–48 h. In cases where complete solubilization was not achieved, an additional 1000 mg of NaDES was introduced to the system, and the stirring was prolonged for a further 24 h at the same temperature. For compound (2), due to its lower solubility, the initial concentration was reduced by a factor of 10 to 2.5 mg per 987.5 mg of NaDES.

The solubilization process was monitored visually. Temperature was maintained using a calibrated heating block, and stirring speed was kept constant at 600 rpm throughout the experiment. All mass measurements were performed using an analytical balance with a precision of ±0.1 mg. For clarity, the final eutectic system selected for all formulations and analyses is the caprylic acid: 1,3-propanediol (1:2 M ratio) deep eutectic solvent. Throughout the manuscript, this solvent is referred to by its full name or the abbreviation CA:PD (1:2) DES.

## 2.5. Standard stability analysis

Samples were measured on day 1 and day 30 after production, pH, conductivity and visible separation were evaluated. The color of samples was measured using Konica Minolta CR-400 colorimeter in L*a*b color space. All measurement were taken in technical triplets.

## 2.6. Turbiscan analysis

Samples were disposed in 2 ml glass vials, ensuring no air bubbles were created. Turbiscan measurement assay was performed using the protocol of 15 min run under 50 °C. Backscattering, Particle size and Total Stability Index were measured on day 1 and day 30.

## 2.7. Rheology

Rheological properties were characterized using a Kinexus pro + rheometer (Netzsch, Germany) with a standard cone-plate geometry (4° angle, 40 mm diameter). All measurements were performed in triplicate

(a fresh sample for each run), with a 10-minute relaxation period to reach equilibrium before each run. The analysis was conducted at 25 °C to mimic the application conditions of the cream formula. Samples were evaluated for properties such as viscosity, deformation, and frequency sweep. Apparent viscosity was measured at increasing shear rates from 0.1 to 1000 s⁻¹, with 10 points per decade over a 20-minute acquisition period. The deformation sweep experiment was performed at a frequency of 1 Hz, with deformation ranging from 0.1 % to 100 % and 50 points per decade during acquisition. The frequency sweep test was conducted at a fixed oscillation amplitude within the viscoelastic range, with the frequency varying from 0.1 Hz to 50 Hz, acquiring 25 points per decade.

## 2.8. Textural analysis

Texture properties of semi-solids were evaluated using the EZ-SX texture analyzer (Shimadzu) at the room temperature at the speed of 10 mm/min, ranging from 0 mm to 5 mm with a trigger threshold being 0.005 N. Properties like hardness, springiness, Texture properties of semi-solids were evaluated using the EZ-SX texture analyzer (Shimadzu) at the room temperature at the speed of 10 mm/min, ranging from 0 mm to 5 mm with a trigger threshold being 0.005 N. Properties like hardness, springiness, cohesiveness and stringiness were obtained using Trapezium-V software.

## 2.9. Spectral characterization

Luminescence properties were measured using FLS1000 spectrometer (Edinburgh Instruments, UK) with a cooled PMT-900 photon detection module (Edinburgh Instruments, UK). Mixed oil phase (16 % sunflower seed oil + 4 %DES) solutions were saturated with air or argon to assure different oxygen concentrations for phosphorescence analyses. Cream was measured in air atmosphere in the form of a film deposited on quartz substrate. The FLS1000 spectrometer was also used for time-resolved phosphorescence measurements ($\lambda_{exc} = 405$ nm, VPLED Series) and the recorded decay curves were fitted to exponential functions by the Fluoracle software (v. 2.13.2, Edinburgh Instruments, UK). Phosphorescence quantum yields of the samples were recorded using a Quantaurus QY C11347-1 spectrometer (Hamamatsu, Japan).

## 2.10. Franz cell permeation assay

Pre-dried and preheated Franz cells containing magnetic stirrers were filled with PBS as the receiving liquid, and a Strat-M® membrane was placed on top of the cells so that the membrane was in contact with the PBS. The system was incubated at 37 °C and left for 30 min to equilibrate. Silicone was applied to the edges of the donor compartments, which were then placed on top of the Strat-M® membranes and secured with clips. Approximately 1 g of the sample emulsions was applied on top of the membranes without any air bubbles, and the donor compartments were covered with parafilm. The samples were incubated at 37 °C for 24 h. After incubation, 200 µl of the receiving liquid was collected, and the residual emulsion was discarded. The Strat-M® membrane was removed, cleaned with PBS, dissolved in chloroform, filtered, and the chloroform was evaporated. The residue was resolubilized with 200 µl of DMSO. All collected samples, both from the receiving liquid and the membrane, were analyzed using ICP-MS to detect the presence of molybdenum.

## 2.11. Microbiology testing

Pathogenic strains *Staphylococcs aureus* (CCM 3953), *Cutibacterium acnes* (CCM 3343), as well as microbiota strains *Micrococcus luteus* (isolated from skin) and *Staphylococcus epidermidis* (isolated from skin) were cultivated at 37 °C and stored at 4 °C on LB (Luria-Bertani) and TSA (Tryptone Soy Agar) agar (in case of *C. acnes*). All experiments were
performed in biological triplicates. Stock inocula of the selected bacteria were prepared by diluting colonies in distilled water or phosphate buffer (20 mM phosphate, pH 7.4) and standardizing the suspension to 1 McFarland (McF). A 100 µL aliquot of the inoculum was mixed with 10 µL of various concentrations of NaDES and/or cluster complexes. Samples were incubated for 2 h in the dark at laboratory temperature. For photodynamic therapy (PDT) experiments, samples were subsequently irradiated using a 12 × 10 W LED source (Cameo) with blue light (460 nm, 18 mW·cm⁻²) for 15 min. The quantification of inactivated bacteria was performed using the Miles and Misra Pathogenic strains *Staphylococcs aureus* (CCM 3953), *Cutibacterium acnes* (CCM 3343), as well as microbiota strains *Micrococcus luteus* (isolated from skin) and *Staphylococcus epidermidis* (isolated from skin) were cultivated at 37 °C and stored at 4 °C on LB (Luria-Bertani) and TSA (Tryptone Soy Agar) agar (in case of *C. acnes*). All experiments were performed in biological triplicates. Stock inocula of the selected bacteria were prepared by diluting colonies in distilled water or phosphate buffer (20 mM phosphate, pH 7.4) and standardizing the suspension to 1 McFarland (McF). A 100 µL aliquot of the inoculum was mixed with 10 µL of various concentrations of NaDES and/or cluster complexes. Samples were incubated for 2 h in the dark at laboratory temperature. For photodynamic therapy (PDT) experiments, samples were subsequently irradiated using a 12 × 10 W LED source (Cameo) with blue light (460 nm, 18 mW·cm⁻²) for 15 min. The quantification of inactivated bacteria was performed using the Miles and Misra method on LB or TSA agar after 24-hour cultivation at 37 °C (72-hours for *C. acnes* in strict anaerobic conditions).

## 2.12. *In vitro cytotoxicity testing*

The immortalized keratinocyte cell line HaCaT was used for the potential toxicity evaluation. Cells were cultured in full medium composed of DMEM high-glucose medium (Sigma-Aldrich) supplemented with 10 % FBS at 37 °C under a 5 % CO₂ atmosphere. The toxicity of NaDES and the NaDES-phosphinate cluster complex (C1) was investigated 24 and 72 h after treatment, and phototoxicity was assessed 2 and 24 h after treatment followed by PDT. The cluster dissolved in NaDES was diluted in PBS with 0.1 % Tween 20 as a surfactant. A fixed volume of 10 µL with different concentrations was added to 100 µL of DMEM medium containing the cells. For phototoxicity experiments, the cluster-treated cells were illuminated using a 12 × 10 W LED source (Cameo) with blue light (460 nm) for 15 min (18 mW·cm⁻²). In the control experiments, the same volume of 0.1 % Tween 20 in PBS was used. Cell viability was analysed using the resazurin assay (Sigma-Aldrich).

# 3. Results and discussion

## 3.1. Cluster compounds solubility in NaDES

Since the solubilization of Mo₆ clusters in NaDES has not been previously reported, a selection of potentially suitable lipophilic NaDES was generated using in silico screening. This computational pre-screening was followed by empirical solubility assays to identify the optimal NaDES formulations. In addition to NaDES, pure sunflower oil was evaluated as a solvent; however, it failed to adequately dissolve the clusters. Based on the nature of the peripheral ligands, a NaDES composed of medium-chain fatty acids—specifically, caprylic acid (C8) and lauric acid (C12) in a 1:1 M ratio—was selected for phosphinate (1) and cholate-based cluster (3). Nonetheless, this system was not suitable for the cluster solubilization. For the phosphonium-substituted cluster (2), a urea:glycerol-based NaDES was initially identified as the most promising candidate.

Finally, a NaDES consisting of caprylic acid and 1,3-propanediol in a 1:2 M ratio demonstrated the most favorable solubilization capacity across all three cluster types. Concentrations of 2.5 % w/w for clusters

(1) and (3), and 0.25 % w/w for cluster (2) were achieved.

## 3.2. Blank cream formulation

A semi-solid base formulation was developed utilizing ingredients with low comedogenic potential, specifically targeting acne-prone skin. *Helianthus annuus* (sunflower) seed oil was selected as the primary lipid phase component due to its low comedogenicity index. The preservative system consisted of benzyl alcohol and dehydroacetic acid as a broad-spectrum antimicrobial agent with established efficacy in cosmetic formulations. For rheological modification, Amigum® (Sclerotium Gum) was incorporated as a natural polysaccharide gelling agent (Lapasin et al., 2017). This biopolymer combines the pseudoplastic properties of guar gum with the electrolyte stability of xanthan gum, providing enhanced texture and stability to the formulation.

Following the formula optimization, a dual emulsifier system comprising cetyl-alcohol and glyceryl-stearate was selected to enhance the stabilization of oil phase and incorporated cluster compounds, as well as improve the overall emulsion integrity. This combination of emulsifiers is known to form lamellar liquid crystalline phases at the oil-water interface, contributing to long-term stability (Ballmann and Mueller, 2008). The cream composition is presented in Table S1.

## 3.3. Cream with NaDES and Mo₆ compound formulation

For the final cream formulation, 4 % w/w of the sunflower oil component was substituted with the optimized NaDES composition (Table 1). Three experimental cream groups were established: a blank cream with the original formula, a control cream, where the CA:PD (1:2) DES was present without any cluster compounds, and cluster creams (C1-C3), containing one of the selected NaDES-cluster complexes at predetermined concentrations.

Three cream groups were created as shown in Table 2. Each sample group contained a quartet of cream samples.

## 3.4. Stability evaluation

The standard stability conducted over 30 days-period showed no visible destabilization of the cream with no phase separation observed and conductivity measurements confirmed the o/w cream. However, the average pH in all sample group has been lowered after 30 days from around 5 to 4, as visible in Table S2. Colorimetry used to determine average color of each sample group in Fig. S1, indicates a significant color shift over 30 days in the cream containing the cholate cluster (C3) and overall darkening of cream groups C1–C3. All in all, the inclusion of Mo₆ clusters led to a noticeable color change from white to a subtle yellow, in contrast to the Blank and Control sample groups.

Next, optical microscopy with image analysis of oil droplet diameters was performed (Fig. 2). This analysis revealed that the microemulsion predominantly comprised small droplets, accompanied by a smaller number of larger domains. The presence of larger domains suggests minor coalescence of the oil phase—an occurrence typical in cosmetic creams (Castel et al., 2017; McClements, 2012). Statistical analysis using paired t-tests demonstrated no significant differences in droplet diameters between the blank, control, and cluster cream groups. After 30 days, slight aggregation of droplets was observed, but the overall changes in diameter remained statistically insignificant, indicating good

<table><caption>**Table 1**
Control and cluster cream formula.</caption><thead><tr><th>Oil phase</th><th>(% w/w)</th><th>Water phase</th><th>(% w/w)</th></tr></thead><tbody><tr><td>Sunflower seed oil</td><td>16</td><td>Water</td><td>73.75</td></tr><tr><td>NaDES (+cluster)</td><td>4</td><td>Preservative</td><td>0.25</td></tr><tr><td>Cetyl-alcohol</td><td>2.5</td><td>Amigum</td><td>1</td></tr><tr><td>Glyceryl-stearate</td><td>2.5</td><td></td><td></td></tr></tbody></table>
<table><caption>**Table 2**

Cream sample groups.</caption><thead><tr><td>Sample group</td><td>NADES</td><td>Cluster</td></tr></thead><tbody><tr><td>Blank</td><td>no</td><td>No</td></tr><tr><td>Control</td><td>yes</td><td>No</td></tr><tr><td>C1</td><td>yes</td><td>(1) phosphinate</td></tr><tr><td>C2</td><td>yes</td><td>(2) phosphonium</td></tr><tr><td>C3</td><td>yes</td><td>(3) cholate</td></tr></tbody></table>

emulsion stability and no meaningful differences between sample groups throughout the storage period.

Secondly, Turbiscan analysis was measured to determine the stability by backscattering. On day 1, all samples, including the blank, control, and experimental formulations, showed relatively uniform backscattering profiles across the vial's height, indicating good initial dispersion without significant phase separation (Fig. 3). Some differences in C2 and C3 creams have emerged, showing a shift in 30 days and suggesting possible oil phase droplets aggregation, especially for the cream C3. Moreover, the baseline of backscattering for the cream C3 was higher than in previous samples. In control cream a slight creaming or phase shift was observed, however, overall, all samples maintained reasonable stability over the 30-day period.

It should be noted that the droplet size values reported in this study originate exclusively from Turbiscan backscattering analysis. Optical microscopy was used only to provide a qualitative visualization of the emulsion microstructure, as its resolution is insufficient to accurately measure droplets below ~1 µm.

### 3.5. Rheology

All tested creams exhibited non-Newtonian behaviour with slight shear thinning, as viscosity decreased with increased shear rate (Fig. S2). This is considered a common and desirable feature among

cosmetic semi-solid emulsions including creams. Under higher shear forces, such as during spreading on the skin, the viscosity decreased, enhancing user-friendly application characteristics (Gilbert et al., 2013; Martin et al., 2025).

Moreover, all the tested cream formulas appeared to be very stable overall, as the viscosity profiles showed minimal variations after a 30-day aging period, with no significant differences between samples.

Slight differences were observed in the spreadability area (viscosity at lower shear rates). The blank and control creams exhibited a slightly higher viscosity, suggesting a more cohesive structure and a thicker application feel. The most noticeable decrease in viscosity was observed for cholate cluster (C3), which may enhance the ease of application (Kim et al., 2024).

Additionally, all the creams containing clusters exhibited a decrease in viscosity after 30 days. Nevertheless, these differences remained within acceptable limits, indicating stable performance over time. To identify the linear viscoelastic region (LVR), strain sweep measurements were conducted across a deformation range from 0.1 to 100 % at a controlled temperature of 25 °C and a fixed frequency of 1 Hz (Fig. S3). These measurements provide more detailed understanding of the viscoelastic properties independent of the applied strain amplitude. The parameters measured were mainly the storage modulus (G') and loss modulus (G'') which indicate the elastic and viscous components of the formulations, respectively.

All creams exhibit stable rheological profiles with only minor decreases in viscoelastic properties, confirming the formulation robustness and suitability for cosmetic application. The blank formula consistently demonstrated the strongest viscoelastic behavior as it exhibited the highest G' values, indicating strong elasticity and relatively lower G'' confirming predominantly elastic response.

The addition of clusters generally decreased the G' and G'' values, with C3 showing the most pronounced effect, suggesting its potential for a softer and more spreadable product. After 30 days of aging, a slight decrease in both, G' and G'' was observed across all cream formulas,

<figure><img src="image_4.png" /><figcaption>**Fig. 2.** Optical microscopy of cream formulations on day 1 and day 30 after aging simulation. Image analysis of oil droplet diameters.</figcaption></figure>

<figure><img src="image_5.png" /><figcaption>**Fig. 3.** Turbiscan backscattering measurement on day 1 and day 30 of all freshly prepared sample groups.</figcaption></figure>
suggesting minor relaxation of the internal structure due to the storage. However, the changes were minimal, indicating good stability.

The frequency sweep analysis (Fig. S4) of emulsions evaluated G' and G'' across the range of frequencies, representing the elastic and viscous responses, respectively. All formulations exhibited stable profiles with gradual increases in both G' and G'' at higher frequencies. Vibrations in the range of 15 to 50 Hz were observed, corresponding to a subtle structural transition within the formula, where a shift between elastic (G') and viscous (G'') behavior begun. A crossover point (liquefaction) was observed around 50 Hz.

Overall, the results demonstrate that all cream samples possess stable viscoelastic properties. This balance between stability and user-friendly characteristics makes them suitable Overall, the results demonstrate that all cream samples possess stable viscoelastic properties. This balance between stability and user-friendly characteristics makes them suitable for cosmetic application. The inclusion of clusters introduced slight variations in texture, particularly softening the formulas, which could enhance spreadability and overall user experience (Shirata and Campos, 2016).

### 3.6. Textural analysis

All creams exhibited similar values in hardness with minor variations. The cream containing phosphinate cluster (C1) demonstrated a slightly higher value, suggesting a firmer texture compared to the others. More notable differences were observed in cohesiveness values, where samples without clusters showed slightly lower values, whereas cream containing the cholate cluster (C3) exhibited higher cohesiveness indicating better structural integrity. Adhesiveness was minimal across all the tested creams, with no significant differences observed. Springiness, which reflects recovery after deformation, was also relatively similar for all creams, although slightly higher in the sample C3. The stringiness, associated with the stretchiness and tackiness of the formulation, was highest in the blank formulation, followed by the control. The addition of clusters notably reduced this value. Overall, no significant differences in the texture of the formulations were observed (Fig. S5).

### 3.7. Spectral characterization

The photophysical properties of the cluster complexes dissolved in mixed oil phase (16 %sunflower seed oil + 4 %DES) were investigated and are summarized in Table 3. Upon excitation at 400 nm, the typical red phosphorescence of the cluster was observed with maxima at approximately 690 nm and quantum yield ranging from 0.20 to 0.50 in Ar-saturated NaDES (Fig. 4A). The lifetimes ranged from 142 to 198 µs and were reduced to 3.9–7.5 µs in air-saturated mixed oil phase (Fig. 4B). The efficient quenching of phosphorescence by oxygen suggested the production of singlet oxygen. Measuring the weak phosphorescence signal of singlet oxygen at 1274 nm confirmed that the singlet

<caption>**Table 3**</caption>

Photophysical properties of cholate, phosphinate, and phosphonium in mixed oil phase at room temperature.ª

<table><thead><tr><th>Sample</th><th>λ<sub>L</sub> (nm)</th><th>Φ<sub>L</sub></th><th>τ<sub>L</sub> (μs)</th><th>τ<sub>air</sub> (μs)</th></tr></thead><tbody><tr><td>(1) phosphinate</td><td>691</td><td>0.50</td><td>198</td><td>7.5<sup>b</sup></td></tr><tr><td>(2) phosphonium</td><td>693</td><td>0.25</td><td>176</td><td>6.6<sup>b</sup></td></tr><tr><td>(3) cholate</td><td>691</td><td>0.20</td><td>142</td><td>3.9<sup>b</sup></td></tr></tbody></table>

oxygen photosensitization was maintained in mixed oil phase (Fig. 4C).

The photophysical properties of the cluster complexes in the cream were also evaluated, evidencing comparable emission spectra than in mixed oil phase albeit red-shifted to approximately 710 nm (Fig. 4D). However, a much longer lifetime of emission of 1.2 ms in air atmosphere was evidenced for all complexes (Fig. 4E).

### 3.8. Franz cell penetration testing

The penetration of the clusters through a skin-like membrane into the receiving solution was assessed using a Franz cell device to estimate the potential absorption of the cluster from the emulsions in the skin. The amount of the cluster in the receptor media and the skin-like membrane was measured as the quantity of molybdenum and recalculated back to the original cluster complex. Overall, the final concentration in the receptor media ranged from 0.03 to 0.06 nM (Table 4). The penetration of molybdenum clusters through the Strat-M® synthetic membrane was found to be extremely low, with concentrations in the receptor medium on the order of a few nanomolars. The Strat-M® membrane is a well-established *in vitro* model of human skin, widely used for preliminary assessment of topical formulations due to its reproducibility, ease of handling, and good correlation with ex vivo human skin permeability for both hydrophilic and lipophilic compounds (Kichou et al., 2023). Although C2 showed an eight-fold higher concentration in the receptor medium compared to C1, the absolute values remain minimal, indicating negligible systemic absorption. These results support the potential safe use of these clusters in cosmetic applications, as the low penetration suggests minimal risk of systemic exposure in line with regulatory safety requirements (EU Cosmetic Regulation 1223/2009).

### 3.9. Antimicrobial activity

Antimicrobial activities of NaDES and clusters were measured using *Staphylococcus aureus*, a prototypical human skin pathogen with high risk of contracting antibiotic resistance. Initial experiments have demonstrated strong antimicrobial activity of NaDES itself, even at concentrations as low as 0.16 % w/w (Fig. S6A). Consequently, it was not possible to conclusively evaluate the phototoxicity of the cluster compounds within the final cream formula. The antibacterial effect of fatty acid-based NaDES has been previously documented, as fatty acids possess antibacterial properties and can modulate the skin microbiome (Wils et al., 2024).

It was found that the primary mechanism underlying the antimicrobial effect of NaDES used in this study was related to the low pH of C8:1,3 propanediol (1:2). Introducing this NaDES into an environment with buffering capacity significantly reduced its toxic effect on bacteria. Conducting experiments in buffered peptone water reduced its toxicity fourfold. Specifically, in the presence of high concentrations of phosphate salts (100 mM, phosphate-buffered water), the antimicrobial activity was substantially diminished (Fig. S6B,C).

Subsequently, the antimicrobial effects of pure Mo₆ cluster complexes were tested by performing a PDT experiment on the planktonic form of S. aureus (Fig. S7). Strong dark toxicity and complete photoinactivation were observed in the cluster 1, while no toxicity was detected for 2, and mild phototoxicity (a one-order-of-magnitude reduction in CFU) was found in 3. The cluster concentrations corresponded to those in the final creams, meaning that the concentration of the cluster 2 was one order of magnitude lower compared to the other clusters. This factor might have also contributed to the lack of bacterial inhibition observed. Based on these results, further experiments were conducted using only the cluster 1 dissolved in NaDES (1:NaDES) in the ratio identical to the cream C1.

Photoinactivation with 1:CA:PD (1:2) DES was demonstrated for S. aureus (Fig. 5A), with dark toxicity being two orders of magnitude lower. Efficient phototoxicity was observed even at lower concentrations, with

<footnote>ª λ<sub>L</sub> – phosphorescence maximum (λ<sub>exc</sub> = 400 nm); τ<sub>L</sub> and τ<sub>air</sub> – amplitude average lifetimes in argon- and air-saturated mixed oil phase (λ<sub>exc</sub> = 405 nm, λ<sub>em</sub> = 690 nm); Φ<sub>L</sub> – phosphorescence quantum yields in argon-saturated mixed oil phase, respectively (λ<sub>exc</sub> = 320–400 nm, experimental error of Φ<sub>L</sub> is ±0.01).</footnote>

<footnote>ᵇ Biexponential decay. Such long lifetime might result from an equilibrium between the excited states of the cluster complexes and a component of the cream, as already observed in a Mo₆ cluster complex with anthracene carboxylate ligands (Kirakci et al., 2014).</footnote>
<figure><img src="image_6.png" /><figcaption>**Fig. 4.** (A) Emission spectra in argon-saturated mixed oil phase, excited at 400 nm. (B) Phosphorescence decay kinetics at 700 nm in argon-, and air- saturated mixed oil phase, excited at 400 nm. (C) Emission spectra of produced singlet oxygen in oxygen-saturated mixed oil phase, excited at 400 nm. (D) Emission spectra in cream, excited at 300 nm. (E) Phosphorescence decay kinetics at 700 nm in air- saturated cream, excited at 300 nm.</figcaption></figure>

<caption>**Table 4**</caption>

Transfer of molybdenum clusters through the StratM® membrane. Percentage calculated from original amount in the cream on the top of the StratM® membrane.

<table><thead><tr><th>Samples</th><th>StratM membrane (%)</th><th>Receptor media (%)</th></tr></thead><tbody><tr><td>C1</td><td>0.00003 ± 0.00006</td><td>0.00015 ± 0.00002</td></tr><tr><td>C2</td><td>&lt;LOQ <sup>b</sup></td><td>0.00121 ± 0.00014</td></tr><tr><td>C3</td><td>&lt;LOQ <sup>b</sup></td><td>0.00024 ± 0.00021</td></tr></tbody></table>

a two-order-of-magnitude reduction in bacterial viability at 1 % of the 1: NaDES solution, which corresponds to 85 µM of pure cluster 1.

C. acnes was found to be more susceptible to PDT (Fig. 5B), showing complete photoinactivation at the same 1 % concentration of the 1:CA: PD (1:2) DES solution. Given that C. acnes is a primary target in acne treatment due to its direct role in skin inflammation (Dessinioti and Katsambas, 2010), this may be considered a highly significant outcome. The high susceptibility of C. acnes to PDT is likely due to its endogenous porphyrin production, which is well-documented in previous studies (Ashkenazi et al., 2003; Dréno et al., 2018).

Importantly, bacterial strains representing the normal skin microbiota (M. luteus and S. epidermidis) were more resistant to PDT treatment (Fig. 5C, D). Significant phototoxicity was observed only at the highest concentrations tested. These results align with previous research on topical antibacterial PDT using 5-aminolevulinic acid (ALA-PDT), where a marked reduction in C. acnes was reported, while commensal bacteria remained largely unaffected or even increased (Yang et al., 2021). The potential of antimicrobial PDT to modulate the skin microbiome and enhance microbiota composition has been demonstrated for porphyrins, showing the increase in the relative abundance of *Actinobacteria*, *Firmicutes*, and *Bacteroidetes* accompanied with a decrease in *Corynebacterium* and *Cutibacterium* (Guo et al., 2023). Notably, beneficial strains such as *Lactobacillus* spp. exhibit greater resistance to oxidative stress,

suggesting that they may be less adversely affected compared to pathogenic strains (Feng and Wang, 2020; Kong et al., 2020). Although skin conditions have typically multifactorial origins, evidence shows the critical role of normal microbial flora in mitigating skin inflammation. This highlights the importance of maintaining a healthy skin microbiome as a protective factor (Guo et al., 2023).

### 3.10. *In vitro* cytotoxicity

As of now, the most studied DES systems are based on choline chloride or phosphonium. Hydrophobic NaDES are less studied and there is still limited data regarding their toxicity on human cell lines (Wils et al., 2021). Generally, IC₅₀ values vary depending on their specific NaDES composition and components used. For example, NaDES comprising choline chloride and 1,2-propanediol found that increasing the ratio of 1,2-propanediol enhanced the toxicity (Lomba et al., 2021). For the CA:PD (1:2) DES employed in our study, data are scarce. Given the structural similarity between 1,2-propanediol and 1,3-propanediol, it is plausible that NADES containing higher concentrations of 1,3-propanediol may exhibit similar trends in cytotoxicity. According to *in vitro* experiments with human epidermis cell line HaCaT (Fig. 6, Table 5), NaDES showed minimal toxicity at concentrations under 1.25 % w/w. Therefore, lowering of the overall concentration of CA:PD (1:2) DES-cluster complex in the final cream to 1 % would eliminate the acute toxicity of the product. The phototoxicity of CA:PD (1:2) DES-cluster was significant from 0,16 % w/w. Dark toxicity of C1 was measured from 0.63 % and higher. IC₅₀ of NaDES was 3.01 ± 0.03 % w/w, which corresponds to 65 mM and for the NaDES-cluster complex only 0.36 ± 0.01 % w/w in 72 h, which corresponds to 7.7 mM of NaDES and 0.03 mM concentration of 1 cluster. However, considering a very low transport trough *stratum corneum*, even 4 % addition in formulation may be perceived as safe.

<footnote><sup>b</sup> LOQ – limit of quantification.</footnote>
<figure><img src="image_7.png" /><figcaption>**Fig. 5.** Planktonic test of dark toxicity and phototoxicity of (Na2[Mo6I8(OPOPh2)6]):CA:PD (1:2) DES solution in 0.1 % Tween 20 and phosphate buffered water (100 mM phosphate) on (A) staphylococcus aureus, (B) cutibacterium acnes, (C) micrococcus luteus and (D) staphylococcus epidermidis. Control group contained 0.1 % Tween 20 and 5 % CA:PD (1:2). ND – no detectable colonies.</figcaption></figure>

<figure><img src="image_8.png" /><figcaption>**Fig. 6.** Cytotoxicity testing. (A) Dark toxicity after 24 h and 72 h. (B) Phototoxicity after incubation for 2 h and 24 h. Samples contained CA:PD (1:2) DES or 1: CA:PD (1:2) DES solution using HaCaT keratinocytes cell line. All samples were containing 0.1 % Tween including control group.</figcaption></figure>

## 4. Conclusion

This application presents a trail-blazing, novel approach to combat skin infections without the use of antibiotics, thereby mitigating the contribution of skin treatments to antibiotic resistance development.

For the first time, it has been demonstrated that Mo6 cluster-based complexes can be solubilized in the innovative natural deep eutectic solvents while retaining their photosensitizing properties and maintaining efficient singlet oxygen production. Additionally, in this form,

they can be used as active cosmetic ingredients and incorporated into cosmetic emulsions, showing new possibilities for the treatment of skin conditions such as acne or dermatitis caused by bacteria. Although the final formula presented in this study requires further optimization, these findings suggest that CA:PD (1:2) DES is as a suitable solvent and carrier of Mo6 cluster complexes as active ingredients for topical PDT applications.

Moreover, the Mo6 clusters selectively target C. acnes while having a limited impact on beneficial microbiota. This further supports their
<caption>**Table 5**</caption>

Calculated IC₅₀ for CA:PD (1:2) DES or 1: CA:PD (1:2) DES solution in HaCaT cells incubated with the mixtures for the indicated time before metabolic activity determination.

<table><thead><tr><th>Dak toxicity</th><th>24 h (%)</th><th>72 h (%)</th></tr></thead><tbody><tr><td>NaDES</td><td>3.01 ± 0.03</td><td>3.01 ± 0.03</td></tr><tr><td>1:NaDES</td><td>0.45 ± 0.02</td><td>0.36 ± 0.01</td></tr><tr><td>Phototoxicity</td><td><strong>2 h (%)</strong></td><td><strong>24 h (%)</strong></td></tr><tr><td>NaDES</td><td>2.29 ± 0.12</td><td>2.72 ± 0.03</td></tr><tr><td>1:NaDES</td><td>0.15 ± 0.05</td><td>0.12 ± 0.01</td></tr></tbody></table>

potential as an effective and microbiome-friendly treatment for acne and other skin conditions. Our results reinforce previous observations that bacteria with thicker cell walls and lower porphyrin levels (e.g., *S. epidermidis* and *M. luteus*) exhibit greater PDT resistance compared to *C. acnes*.

However, concentration of cluster:NaDES originally used for the tested formulation seems unnecessary high and 4 % addition can be decreased under 1 % for safety purpose and to target only pathogenic bacterial strains.

### CRediT authorship contribution statement

**Michaela Kubáňová:** Writing – original draft, Methodology, Investigation, Conceptualization. **Kaplan Kirakci:** Writing – review & editing, Validation, Supervision, Methodology, Investigation, Conceptualization. **Leslie Boudesocque-Delaye:** Writing – review & editing, Validation, Supervision, Methodology, Investigation, Conceptualization. **Laura Divoux:** Writing – review & editing, Methodology, Investigation. **Antonín Kana:** Methodology, Investigation, Data curation. **Jaroslav Zelenka:** Writing – review & editing, Validation, Supervision, Project administration. **Tomáš Ruml:** Writing – review & editing, Validation, Supervision. **Emilie Munnier:** Writing – review & editing, Validation, Supervision. **Kamil Lang:** Writing – review & editing, Validation, Supervision. **Alexis Verger:** Writing – review & editing, Visualization, Validation, Supervision, Project administration, Methodology, Investigation, Data curation, Conceptualization.

### Declaration of competing interest

The authors declares that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

### Acknowledgement

The co-author thanks the EIG Concert Japan PHOTOMOS_H2O (No. 8I24001) for financial support.

### Appendix A. Supplementary data

Supplementary data to this article can be found online at https://doi.org/10.1016/j.ijpharm.2025.126491.

### Data availability

Data will be made available on request.

### References

Aduloju, E.I., Yahaya, N., Mohammad Zain, N., Anuar Kamaruddin, M., Hamid, A.A., M., 2023. An overview on the use of DEEP eutectic solvents for green extraction of some selected bioactive compounds from natural matrices. Adv. J. Chem. Sect. A 6, 253–300. https://doi.org/10.22034/ajca.2023.389403.1356.

Alexander, H., Paller, A.S., Traidl-Hoffmann, C., Beck, L.A., De Benedetto, A., Dhar, S., Girolomoni, G., Irvine, A.D., Spuls, P., Su, J., Thyssen, J.P., Vestergaard, C., Werfel, T., Wollenberg, A., Deleuran, M., Flohr, C., 2020. The role of bacterial skin

infections in atopic dermatitis: expert statement and review from the International Eczema Council Skin Infection Group. Br. J. Dermatol. 182, 1331–1342. https://doi.org/10.1111/jbd.18643.

Ashkenazi, H., Malik, Z., Harth, Y., Nitzan, Y., 2003. Eradication of Propionibacterium acnes by its endogenic porphyrins after illumination with high intensity blue light. FEMS Immunol. Med. Microbiol. 35, 17–24. https://doi.org/10.1111/j.1574-695X.2003.tb00644.x.

Ballmann, C., Mueller, B.W., 2008. Stabilizing effect of cetostearyl alcohol and glyceryl monostearate as co-emulsifiers on hydrocarbon-free O/W glyceride creams. Pharm. Dev. Technol. 13, 433–445. https://doi.org/10.1080/10837450802247952.

Boudesocque-Delaye, L., Ardeza, I.M., Verger, A., Grard, R., Théry-Koné, I., Perse, X., Munnier, E., 2024. Natural deep eutectic solvents as a novel bio-based matrix for ready-to-use natural antioxidants-enriched ingredients: extraction and formulation optimization. Cosmetics 11, 17. https://doi.org/10.3390/cosmetics11010017.

Castel, V., Rubiolo, A.C., Carrara, C.R., 2017. Droplet size distribution, rheological behavior and stability of corn oil emulsions stabilized by a novel hydrocolloid (Brea gum) compared with gum arabic. Food Hydrocoll. 63, 170–177. https://doi.org/10.1016/j.foodhyd.2016.08.039.

Cavalcante, F.S., Saintive, S., Carvalho Ferreira, D., Rocha Silva, A.B., Guimarães, L.C., Braga, B.S., Dios Abad, E., Ribeiro, M., Netto Dos Santos, K.R., 2021. Methicillin-resistant Staphylococcus aureus from infected skin lesions present several virulence genes and are associated with the CC30 in Brazilian children with atopic dermatitis. Virulence 12, 260–269. https://doi.org/10.1080/21505594.2020.1869484.

Desbois, A.P., Smith, V.J., 2010. Antibacterial free fatty acids: activities, mechanisms of action and biotechnological potential. Appl. Microbiol. Biotechnol. 85, 1629–1642. https://doi.org/10.1007/s00253-009-2355-3.

Dessinioti, C., Katsambas, A., 2022. Antibiotics and antimicrobial resistance in acne: epidemiological trends and clinical practice considerations. Yale J. Biol. Med. 95, 429–443.

Dessinioti, C., Katsambas, A.D., 2010. The role of Propionibacterium acnes in acne pathogenesis: facts and controversies. Clin. Dermatol. 28, 2–7. https://doi.org/10.1016/j.clindermatol.2009.03.012.

Dias, L.M., Ferrisse, T.M., Medeiros, K.S., Cilli, E.M., Pavarina, A.C., 2022. Use of photodynamic therapy associated with antimicrobial peptides for bacterial control: a systematic review and meta-analysis. Int. J. Mol. Sci. 23, 3226.

Diogo, M.L.G., Campos, T.M., Fonseca, E.S.R., Pavani, C., Horliana, A.C.R.T., Fernandes, K.P.S., Bussadori, S.K., Fantin, F.G.M.M., Leite, D.P.V., Yamamoto, Á.T. A., Navarro, R.S., Motta, L.J., 2021. Effect of blue light on acne vulgaris: a systematic review. Sensors 21, 6943. https://doi.org/10.3390/s210206943.

Dréno, B., Pécastaings, S., Corvec, S., Veraldi, S., Khammari, A., Roques, C., 2018. Cutibacterium acnes (Propionibacterium acnes) and acne vulgaris: a brief look at the latest updates. J. Eur. Acad. Dermatol. Venereol. 32, 5–14. https://doi.org/10.1111/jdv.15043.

Felip-León, C., Arnaud del Valle, C., Pérez-Laguna, V., Isabel Millán-Lou, M., Miravet, J.F., Mikhailov, M., Sokolov, M.N., Rezusta-López, A., Galindo, F., 2017. Superior performance of macroporous over gel type polystyrene as a support for the development of photo-bactericidal materials. J. Mater. Chem. B 5, 6058–6064. https://doi.org/10.1039/C7TB01478C.

Feng, T., Wang, J., 2020. Oxidative stress tolerance and antioxidant capacity of lactic acid bacteria as probiotic: a systematic review. Gut Microb. 12, 1801944. https://doi.org/10.1080/19490976.2020.1801944.

Gilbert, L., Picard, C., Savary, G., Grisel, M., 2013. Rheological and textural characterization of cosmetic emulsions containing natural and synthetic polymers: relationships between both data. Colloids Surf. Physicochem. Eng. Asp. 421, 150–163. https://doi.org/10.1016/j.colsurfa.2013.01.003.

Guégan, R., Cheng, X., Huang, X., Němečková, Z., Kubáňová, M., Zelenka, J., Ruml, T., Grasset, F., Sugahara, Y., Lang, K., Kirakci, K., 2023. Graphene oxide sheets decorated with octahedral molybdenum cluster complexes for enhanced photoinactivation of staphylococcus aureus. Inorg. Chem. 62, 14243–14251. https://doi.org/10.1021/acs.inorgchem.3c01502.

Guo, Y., Zeng, M., Yuan, Y., Yuan, M., Chen, Y., Yu, H., Liu, R., Ruan, Z., Xie, Q., Jiao, X., Lu, T., 2023. Photodynamic therapy treats acne by altering the composition of the skin microbiota. Skin Res. Technol. 29, e13269. https://doi.org/10.1111/srt.13269.

Jelinski, T., Przybylek, M., Cysewski, P., 2019. Natural deep eutectic solvents as agents for improving solubility, stability and delivery of curcumin. Pharm. Res. 36, 116. https://doi.org/10.1007/s11095-019-2643-2.

Keyal, U., Bhatta, A.K., Wang, X.L., 2016. Photodynamic therapy for the treatment of different severity of acne: a systematic review. Photodiagn. Photodyn. Ther. 14, 191–199. https://doi.org/10.1016/j.pdpdt.2016.04.005.

Kichou, H., Bonnier, F., Dancik, Y., Bakar, J., Michael-Jubeli, R., Caritá, A.C., Perse, X., Soucé, M., Rapetti, L., Tfayli, A., Chourpa, I., Munnier, E., 2023. Strat-M® positioning for skin permeation studies: a comparative study including EpiSkin® RHE, and human skin. Int. J. Pharm. 647, 123488. https://doi.org/10.1016/j.ijpharm.2023.123488.

Kim, J., Jeong, E.H., Baik, J.H., Park, J.D., 2024. The role of rheology in cosmetics research: a review. Korea-Aust. Rheol. J. 36, 271–282. https://doi.org/10.1007/s13367-024-00108-y.

Kirakci, K., Demel, J., Hynek, J., Zelenka, J., Rumlová, M., Ruml, T., Lang, K., 2019a. Phospinate apical ligands: a route to a water-stable octahedral molybdenum cluster complex. Inorg. Chem. 58, 16546–16552. https://doi.org/10.1021/acs.inorgchem.9b02569.

Kirakci, K., Fejfarová, K., Kučeráková, M., Lang, K., 2014. Hexamolybdenum cluster complexes with pyrene and anthracene carboxylates: ultrabright red emitters with the antenna effect. Eur. J. Inorg. Chem. 2014, 2331–2336. https://doi.org/10.1002/ejic.201402076.
Kirakci, K., Kubát, P., Dušek, M., Fejfarová, K., Sícha, V., Mosinger, J., Lang, K., 2012. A highly luminescent hexanuclear molybdenum cluster – a promising candidate toward photoactive materials. Eur. J. Inorg. Chem. 2012, 3107–3111. https://doi.org/10.1002/ejic.201200402.

Kirakci, K., Kubát, P., Langmaier, J., Polívka, T., Fuciman, M., Fejfarová, K., Lang, K., 2013. A comparative study of the redox and excited state properties of (nBu4N)2[Mo6X14] and (nBu4N)2[Mo6X8(CF3COO)6] (X = Cl, Br, or I). Dalton Trans. 42, 7224.

Kirakci, K., Nguyen, T.K.N., Grasset, F., Uchikoshi, T., Zelenka, J., Kubát, P., Ruml, T., Lang, K., 2020a. Electrophoretically deposited layers of octahedral molybdenum cluster complexes: a promising coating for mitigation of pathogenic bacterial biofilms under blue light. ACS Appl. Mater. Interf. 12, 52492–52499. https://doi.org/10.1021/acsami.0c19036.

Kirakci, K., Zelenka, J., Krížová, I., Ruml, T., Lang, K., 2020b. Octahedral molybdenum cluster complexes with optimized properties for photodynamic applications. Inorg. Chem. 59, 9287–9293. https://doi.org/10.1021/acs.inorgchem.0c01173.

Kirakci, K., Zelenka, J., Rumlová, M., Cvačka, J., Ruml, T., Lang, K., 2019b. Cationic octahedral molybdenum cluster complexes functionalized with mitochondria-targeting ligands: photodynamic anticancer and antibacterial activities. Biomater. Sci. 7, 1386–1392. https://doi.org/10.1039/C8BM01564C.

Kong, Y., Olejar, K.J., On, S.L.W., Chelikani, V., 2020. The potential of lactobacillus spp. for modulating oxidative stress in the gastrointestinal tract. Antioxidants 9, 610. https://doi.org/10.3390/antiox9070610.

Kua, Y.L., Gan, S., 2019. Natural deep eutectic solvent (NADES) as a greener alternative for the extraction of hydrophilic (polar) and lipophilic (non-polar) phytonutrients. Key Eng. Mater. 797, 20–28.

Lapasin, R., Abrami, M., Grassi, M., Šebenik, U., 2017. Rheology of Laponitescleroglucan hydrogels. Carbohydr. Polym. 168, 290–300. https://doi.org/10.1016/j.carbpol.2017.03.068.

Lomba, L., Ribate, M.P., Sangüesa, E., Concha, J., Garralaga, M.A.P., Errazquin, D., García, C.B., Giner, B., 2021. Deep Eutectic solvents: are they safe? Appl. Sci. 11, 10061.

Maisch, T., 2015. Resistance in antimicrobial photodynamic inactivation of bacteria. Photochem. Photobiol. Sci. 14, 1518–1526. https://doi.org/10.1039/c5 pp00037h.

Maisch, T., 2009. A new strategy to destroy antibiotic resistant microorganisms: antimicrobial photodynamic treatment. Mini Rev. Med. Chem. 9, 974–983. https://doi.org/10.2174/138955709788681582.

Martin, C., Vrignaud, E., Boudesocque-Delaye, L., Verger, A., Munnier, E., Grisel, M., Gore, E., 2025. Natural eutectic systems as emerging green multifunctional ingredients: a sensory-related perspective. J. Mol. Liq. 436, 128277. https://doi.org/10.1016/j.molliq.2025.128277.

McClements, D.J., 2012. Nanoemulsions versus microemulsions: terminology, differences, and similarities. Soft Matter 8, 1719–1729. https://doi.org/10.1039/C2SM06903B.

Pei, S., Inamadar, A.C., Adya, K.A., Tsoukas, M.M., 2015. Light-based therapies in acne treatment. Indian Dermatol. Online J. 6, 145–157. https://doi.org/10.4103/2229-5178.156379.

Přibyl, T., Rumlová, M., Mikysková, R., Reiniš, M., Kaňa, A., Škoch, K., Zelenka, J., Kirakci, K., Ruml, T., Lang, K., 2024. PEGylated Molybdenum-Iodine Nanocluster as

a Promising Radiodynamic Agent against Prostatic Adenocarcinoma. Inorg. Chem. 63, 4419–4428. https://doi.org/10.1021/acs.inorgchem.4c00084.

Rente, D., Cvjetko Kubalo, M., Panić, M., Paiva, A., Caprín, B., Radojić Redovniković, I., Duarte, A.R.C., 2022. Review of deep eutectic systems from laboratory to industry, taking the application in the cosmetics industry as an example. J. Clean. Prod. 380, 135147. https://doi.org/10.1016/j.jclepro.2022.135147.

Shirata, M.M.F., Campos, P.M.B.G.M., 2016. Importance of texture and sensorial profile in cosmetic formulations development. Surg. Cosmet. Dermatol. 8. https://doi.org/10.5935/scd1984-8773.201683861.

Union, p., 2009. Regulation (EC) No 1223/2009 of the european parliament and of the council. Off. J. Eur. Union L 342, 59.

Verger, A., Brandhonneur, N., Molard, Y., Cordier, S., Kowouvi, K., Amela-Cortes, M., Dollo, G., 2021. From molecules to nanovectors: current state of the art and applications of photosensitizers in photodynamic therapy. Int. J. Pharm. 604, 120763. https://doi.org/10.1016/j.ijpharm.2021.120763.

Verger, A., Dollo, G., Brandhonneur, N., Martinais, S., Cordier, S., Lang, K., Amela-Cortes, M., Kirakci, K., 2023. PEGylated poly(lactic-co-glycolic acid) nanoparticles doped with molybdenum-iodide nanoclusters as a promising photodynamic therapy agent against ovarian cancer. Adv. Mater. https://doi.org/10.1039/D3MA00206C.

Verger, A., Dollo, G., Martinais, S., Molard, Y., Cordier, S., Amela-Cortes, M., Brandhonneur, N., 2022. Molybdenum-iodine cluster loaded polymeric nanoparticles allowing a coupled therapeutic action with low side toxicity for treatment of ovarian cancer. J. Pharm. Sci. 111, 3377–3383. https://doi.org/10.1016/j.xphs.2022.09.010.

Verger, A., Kichou, H., Huang, N., Perse, X., Ardeza, I.M., Pradel, C., Conceicao, G.M.D. R., Atanasova, B., Legrand, F.X., Despres, A., Boudesocque-Delaye, L., Munnier, E., 2024. Effects of hydrophilic natural deep eutectic solvents on the rheological, textural, and sensory properties of carboxymethylcellulose-based cosmetic hydrogels. ACS Sust. Chem. Eng. 12, 7187–7199. https://doi.org/10.1021/acssuschemeng.4c01866.

Vorotnikova, N.A., Alekseev, A.Y., Vorotnikov, Y.A., Evtushok, D.V., Molard, Y., Amela-Cortes, M., Cordier, S., Smolentsev, A.I., Burton, C.G., Kozhin, P.M., Zhu, P., Topham, P.D., Mironov, Y.V., Bradley, M., Efremova, O.A., Shestopalov, M.A., 2019. Octahedral molybdenum cluster as a photoactive antimicrobial additive to a fluoroplastic. Mater. Sci. Eng. C 105, 110150. https://doi.org/10.1016/j.msec.2019.110150.

Wiegell, S.R., Wulf, H.C., 2006. Photodynamic therapy of acne vulgaris using 5-amino-levulinic acid versus methyl aminolevulinate. J. Am. Acad. Dermatol. 54, 647–651. https://doi.org/10.1016/j.jaad.2005.12.033.

Wils, L., Leman-Loubière, C., Bellin, N., Clément-Larosière, B., Pinault, M., Chevalier, S., Enguehard-Gueiffier, C., Bodet, C., Boudesocque-Delaye, L., 2021. Natural deep eutectic solvent formulations for spirulina: preparation, intensification, and skin impact. Algal Res. 56, 102317. https://doi.org/10.1016/j.algal.2021.102317.

Wils, L., Yagmur, M., Bellin, N., Phelippe, M., Chevalley, A., Bodet, C., Boudesocque-Delaye, L., 2024. Innovative alkanediol-based eutectic solvents for extracting/pre-formulating dermatologically valuable free fatty acids from spirulina and porphyridium cakes. Mar. Drugs 22, 281. https://doi.org/10.3390/md22060281.

Yang, Y., Tao, S., Zeng, R., Zheng, H., Ge, Y., 2021. Modulation of skin microbiome in acne patients by aminolevulinic acid-photodynamic therapy. Photodiagn. Photodyn. Ther. 36, 102556. https://doi.org/10.1016/j.pdpdt.2021.102556.