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Article

Influence of Silver Nanoparticles on Liposomal Membrane Properties Relevant in Photothermal Therapy

by
Maria Lyudmilova
1,*,
Lyubomir Stoychev
1,
Denitsa Yancheva
2,3,
Stoyanka Nikolova
4,
Mina Todorova
4,
Charilaos Xenodochidis
5,
Kamelia Hristova-Panusheva
5,
Natalia Krasteva
5 and
Julia Genova
1,*
1
Institute of Solid State Physics, Bulgarian Academy of Sciences, Tzarigradsko Chaussee 72, 1784 Sofia, Bulgaria
2
Institute of Organic Chemistry with Centre of Phytochemistry, Bulgarian Academy of Sciences, “Acad. Georgi Bonchev” Str., Build. 9, 1113 Sofia, Bulgaria
3
Department of Organic Chemistry, University of Chemical Technology and Metallurgy, 8 Kliment Ohridski Blvd., 1756 Sofia, Bulgaria
4
Department of Organic Chemistry, Faculty of Chemistry, University of Plovdiv, 4000 Plovdiv, Bulgaria
5
Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, “Acad. Georgi Bonchev” Str., Bl. 21, 1113 Sofia, Bulgaria
*
Authors to whom correspondence should be addressed.
Appl. Sci. 2026, 16(1), 220; https://doi.org/10.3390/app16010220
Submission received: 26 November 2025 / Revised: 20 December 2025 / Accepted: 23 December 2025 / Published: 24 December 2025
(This article belongs to the Section Nanotechnology and Applied Nanosciences)
Browse Figures
Review Reports Versions Notes

Abstract

Silver nanoparticles (AgNPs) are promising agents for nanomedicine but their interactions with lipid membranes, which are a key interfaces for drug delivery, require a deeper understanding. This study investigates the influence of fructose-capped AgNPs on the physicochemical properties of SOPC-based liposomal bilayers, with potential implications for drug delivery and photothermal therapy. We employed a multitechnique approach, including infrared (IR) spectroscopy, differential scanning calorimetry (DSC), thermally induced shape fluctuation analysis, and laser irradiation at 343, 515, and 1030 nm. Our results show that AgNPs incorporated into the bilayer cause measurable perturbations: DSC reveals a decrease in the main phase transition enthalpy (from 0.280 to 0.234 J/g) and temperature (from 2.80 to 3.41 °C), while shape fluctuation analysis indicates a reduction in bending modulus (from 1.18 × 10−19 J to 0.93 × 10−19 J), confirming increased membrane fluidity. FTIR confirms interactions of fructose-capped nanoparticles and the lipid’s carbonyl and phosphate groups. Furthermore, the AgNPs-liposomes exhibit a strong, wavelength-dependent photothermal response with a temperature increase of ≈22 °C under 515 nm laser irradiation, compared to only 3–5 °C at 1030 nm. We conclude that fructose-capped AgNPs moderately fluidify lipid bilayers while enabling efficient, controllable photothermal capability, making them excellent candidates for the eventual design of advanced liposomal systems for combined therapy and diagnostics.

1. Introduction

Silver has a long-standing history in biomedicine because of its broad-spectrum antimicrobial and antifungal properties. Even before the nanoscale era, bulk silver was employed to treat infections, burns, and wounds. At the nano scale, silver nanoparticles (AgNPs) inherit and greatly amplify these attributes: AgNPs typically measure less than 100 nm and contain thousands of silver atoms, giving them extremely high surface area-to-volume ratios and distinct size-dependent physicochemical characteristics, including high electrical and thermal conductivity, chemical stability and catalytic activity. These properties enable AgNPs to inhibit the growth of hundreds of bacterial, fungal, and algal species through controlled silver-ion release, making them highly relevant for medical, healthcare, and environmental applications. Recent advances in green nanotechnology, particularly biologically assisted synthesis routes, further enhance the relevance of AgNPs by enabling eco-friendly production with tunable surface chemistries and improved biosafety profiles, expanding their applicability across medicine, agriculture, electronics, and water-treatment technologies [1,2,3]. Peptides and proteins can be used as templates or to regulate AgNP nucleation, mirroring broader trends in green NP syntheses [4]. These unique properties (optical, thermal, and chemical) differ markedly from those of bulk silver and make AgNPs especially attractive in nanomedicine. However, AgNPs generate reactive species that disrupt microbial membranes and metabolic processes, producing their potent antibacterial/antifungal actions. The current focus on the nature of AgNPs is in cancer diagnostics and therapy: they can serve as cytotoxic or drug carrier agents against tumors. Due to their high surface-area-to-volume ratio, ease of functionalization, and excellent stability, silver NPs have gained significant interest as drug delivery systems. They have demonstrated versatility in delivering a wide range of therapeutic agents. Moreover, AgNPs enhance drug solubility, protect sensitive compounds from degradation, and provide controlled and sustained release at targeted sites [5]. Crucially, these nanoparticles exhibit surface plasmon resonance in the visible–near-IR range with tunable size and shape, enabling efficient absorption of near-infrared (NIR) light and converting it to heat [6]. This plasmonic heating under NIR irradiation is the physical basis of photothermal therapy (PTT) with metallic NPs. Thus, the combination of inherent bioactivity, high surface reactivity, and strong tunable optical absorption makes AgNPs highly beneficial for advanced drug delivery and photothermal applications in cancer treatment.
Parallel to the development of metallic NPs, liposomes have emerged as cornerstones in nanomedicine for drug delivery [7]. These biocompatible, self-assembled phospholipid bilayers mimic the structural characteristics of biological membranes and can encapsulate a wide range of therapeutic agents—both hydrophilic and hydrophobic. Liposomal formulations offer critical advantages, such as improved drug solubility, reduced systemic toxicity, extended systemic circulation, and the ability to be functionalized for controlled or triggered release [8,9]. For fundamental studies, well-defined phospholipids like SOPC (1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine) are ideal as they form fluid, stable bilayers at physiological temperature that resemble mammalian cell membrane. The integration of AgNPs with liposomal carriers present a compelling strategy to create multifunctional platforms that combine the diagnostic and photothermal capabilities of AgNPs with the superior drug delivery properties of liposomes. However, while both liposomes and AgNPs have been widely studied independently, the fundamental interactions between AgNPs and lipid bilayers remains poorly understood. Critical questions regarding how AgNPs influence membrane fluidity, structural integrity, and phase behavior must be answered to enable rational design.
This study therefore investigates the physicochemical effects of fructose-capped Ag-NPs on SOPC-based liposomal membranes. We focus specifically on elucidating the changes in bilayer elasticity, structure, and phase behavior. By probing these fundamental nanoparticle–membrane interactions in a controlled model system, this work provides the necessary foundation for the rational development of advanced, combined-modality nanotheraupetics that simultaneously leverage drug delivery and photothermal intervention.

2. Materials and Methods

2.1. Materials Sample Preparation

We purchased 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine (SOPC) from Avanti Polar Lipids, Inc., (Alabaster, AL, USA). Chloroform (99% purity) was obtained from Sigma-Aldrich and Merck (Merck KGaA, Darmstadt, Germany). All chemicals were used without further purification. The structural formula of the lipid used in the studies is given in Figure 1.
Fructose-capped silver nanoparticles (AgNPs) were synthesized as previously de- scribed, with slight modifications [10]; their spherical morphology and a thin fructose capping layer (≈2–15 nm) were confirmed by transmission electron microscopy (TEM, Figure 2).

2.2. Sample Preparation

2.2.1. Liposome Preparation for DSC and FTIR Spectroscopy

For both DSC and FTIR spectroscopy, samples were prepared using the same protocol, as follows: SOPC was dissolved in chloroform to a concentration of 10 mg/mL. For each sample, an aliquot containing 1 mg of SOPC was poured into a vial, and the solvent was evaporated under vacuum for 4 h to form an uniform lipid film. The dried film was hydrated with 100 µL of an aqueous solution containing double-distilled water and AgNPs to achieve final AgNP concentrations of 0, 0.5, 1.0, and 2.5 wt%. Finally, the samples were incubated in an ultrasonic bath at 35 °C for 8 h to form multilamellar vesicles.

2.2.2. Preparation of Giant Unilamellar Vesicles (GUVs) for Shape Fluctuation Analysis

Giant unilamellar vesicles (GUVs, diameter approx. 20–40 µm) were prepared using a modified electroformation method to ensure consistency, using the same lipid/AgNP solutions as for the spectroscopic and calorimetric studies. The electroforming chamber consisted of two indium tin oxide (ITO)-coated glass slides (layer thickness of 100 ± 20 nm, resistivity of approx. 100 Ω/square), separated by a PDMS spacer. For each AgNP concentration, freshly prepared solutions were deposited as multiple small droplets on the conducting surfaces of the slides and dried under vacuum for about 4 h to allow complete solvent removal. The chamber was then filled with double-distilled water and silver nanoparticles at the following concentrations, 1% and 2.5%, and connected overnight to a generator supplying a sinusoidal alternating voltage of 1.5 Vpp at 10 Hz. This procedure ensured the successful formation of GUVs with AgNPs embedded in the hydrophobic region of the lipid bilayer.

2.3. Infrared (IR) Spectroscopy

Infrared spectroscopy is a powerful analytical method that provides valuable molecular and structural information for a wide range of inorganic compounds, organic molecules, and biomolecular systems. Vibrational spectroscopy can identify the components in a mixture and reveal the structural changes resulting from molecular interactions or chemical transformations. These capabilities have made it especially useful for investigating conformational changes in lipid bilayers and membranes [11,12,13].
The attenuated total reflectance (ATR) IR spectra were acquired on a Bruker Invenio R spectrometer equipped with a diamond crystal Pike Technology ATR accessory. The samples were studied in liquid state by directly depositing several drops on the diamond surface. The spectra were recorded in the frequency region of 4000–600 cm−1 with 200 scans at a resolution of 2 cm−1, with water as the background. The spectra were baseline-corrected and smoothed at 17 smoothing points.

2.4. Differential Scanning Calorimetry (DSC)

DSC is a thermoanalytical technique that measures the difference in heat flow between a sample and a reference as their temperatures increase at a controlled rate [14,15]. When the sample undergoes a phase transition, additional heat is either absorbed (endothermic) or released (exothermic), which is recorded as a peak in the DSC thermogram. This method allows for the determination of key thermodynamic properties such as the changes in enthalpy, entropy, and heat capacity associated with phase transitions.
In this study, DSC measurements were performed using a DSC Discovery 250 instrument (TA Instruments, New Castle, DE, USA). Precisely weighed sample portions (typically between 15 and 30 mg) were placed in standard aluminum pans, hermetically sealed, and measured against an empty reference pan under identical conditions. The temperature protocol included an initial heating, followed by cooling and a final heating (further details in the following section) from −10 °C to 70 °C, with a scan rate of 5 °C/min.
For DSC data processing and analysis, a custom C++ program was developed as part of a course thesis and was used to automatically calculate the heating and cooling enthalpies of the AgNP-containing and control SOPC samples.

2.5. Thermally Induced Shape Fluctuation Method

One common and effective approach for quantifying the bending elasticity of lipid bilayers is thermally induced shape fluctuation analysis [16,17,18]. This technique is used to estimate how the mechanical properties of SOPC lipid bilayers are influenced by NPs. The spontaneous and thermally driven undulations of giant unilamellar vesicles are recorded via phase-contrast microscopy. Their contour fluctuations are then analyzed using spherical harmonic decomposition, and the bending modulus is extracted by fitting fluctuation spectra to the Milner–Safran theoretical model. Stroboscopic illumination is provided by a xenon flash lamp to capture clear pictures of the vesicles. To prevent eye irritation caused by direct exposure, sample positioning and stability were monitored indirectly via a video display. The xenon flash lamp’s pulsed light ensures that flicker is eliminated, allowing for stable, high-quality frames for subsequent quantitative analysis [19,20].
For the analysis, equatorial cross-sections of GUVs are recorded, usually capturing 400 images per second, and the mean diameter is measured in 256 directions along the vesicle’s radius. These images are then decomposed into harmonic functions. This non-invasive technique is highly sensitive to chemical or structural perturbations, including the presence of nanoparticles or bilayer compositions, allowing researchers to assess changes in mechanical properties due to such interactions [21,22,23].
Fluctuating giant vesicles were imaged using a Zeiss Axiovert (Carl Zeiss, Oberkochen, Germany) 100 phase-contrast microscope (LD Ph2 100×, NA 0.75) equipped with a custom stroboscopic illumination system. The setup employed a xenon flash lamp L6604 (external main discharge capacitor E7289-01, power supply C6096) synchronized with vertical sync pulses from a Hamamatsu, Japan (manufacturer of all mentioned equipment in the current paragraph) CCD camera (C2400-60, Hamamatsu Photonics, Hamamatsu City, Japan), producing light pulses of less than 3–4 µs (FWHM) at 2 J input energy, as specified in the manufacturer’s data sheet.

2.6. Laser Irradiation

The effect of electromagnetic radiation on AgNPs was studied to determine their ability to change temperature when exposed to a laser beam. All experiments were performed using a Pharos PH2-10W femtosecond laser (Light Conversion, Vilniaus, Lithuania) equipped with an automated harmonic generator emitting at wavelengths of 1030 nm, 515 nm, and 343 nm. The system delivers up to 10 W at 1030 nm, with corresponding maximum powers of 5.9 W and 2.8 W for the second and third harmonics. Pulse durations range from 170 fs for the fundamental wavelength to 130 fs for the higher harmonics, with repetition rates adjustable from a single pulse up to 200 kHz.

3. Results

3.1. Transmission Electron Microscopy (TEM)

Transmission electron microscopy (TEM) provides valuable insight into the morphology and surface chemistry of AgNPs, particularly in relation to their synthesis and stabilization mechanisms. The TEM image (Figure 2a) confirms the synthesis of smaller spherical particles. Many compounds are used as capping agents, such as polysaccharides, proteins, mannose, galactose, glucose, chitosan, and sodium alginate [24].
The TEM images, shown in Figure 2a,b, confirmed that fructose participated as a capping and protecting agent. A thin coat of fructose with an average thickness of 2–15 nm can be observed in the aforementioned TEM image. Many groups, such as hydroxyl, carboxyl, phenol, and carbonyl groups, are linked to oxygen and nitrogen with covalent bonds for the complex formation of silver, so they are probably absorbed on its surface [25,26]. This concept of small reducing sugar molecules acting as protecting agents is not strictly limited to NPs: as table sugar [27,28,29] is composed of the glucose and fructose, we hypothesize, based on prior studies, that sucrose serves as protector of cells against drought, both at low and high temperatures [30,31]. Similarly, we assume that fructose can act as a stabilizing and protecting agent for silver nanoparticles in experimental systems: supporting its ability to preserve the structural integrity of biological membranes under stress. Analogously, table sugar can protect various nanoparticles during synthesis and storage.

3.2. Dynamic Light Scattering (DLS)

The median size of the obtained particles was confirmed by DLS (Figure 3). The histogram of the sample displayed a trimodal particle size distribution, suggesting the presence of aggregates. Based on the DLS, the median size of the obtained particles was identified in the range of 10 to 40 nm.

3.3. Zeta Potential

The zeta potential of the sample was −17.94 mV. The negative charge of the zeta potential reflects the carboxylic groups of gluconic acid obtained when Ag+ is reduced to Ag0. Carboxylic acids, obtained in the oxidation of sugars, provide a negative surface charge density to counteract the van der Waals forces responsible for particle coalescence. Self-assembled carboxylic acids ensure dense coating on the metal surfaces and stabilize them [32].

3.4. UV–Vis Spectroscopy

An essential method for examining AgNPs in suspension is UV–Vis absorption spectroscopy. The absorption spectra in the 190–600 nm wavelength range were recorded in order to track the AgNP production in an aqueous solution (Figure 4). Different optical characteristics related to surface plasmon resonance are displayed by each kind of AgNP [33]. Figure 4 shows the time-dependent UV–Vis absorption spectra of AgNPs. Fructose displays an absorption peak at 189 nm and 280 nm.
The characteristic SPR band at 415 nm is an indicator of the formation of AgNPs. The spherical shape of the particles is supported by the band’s location between 350 and 420 nm [34]. The symmetrical peak of AgNPs showed a modest level of NP aggregation [35,36].

3.5. Infrared (IR) Spectroscopy

The ATR-FTIR spectra of the pure SOPC and the liposomes with varying amounts of NPs are depicted in the region of 3000–900 cm−1 in Figure 5.
In the FTIR-ATR spectrum of the pure SOPC, the IR band for the asymmetric C-H vibrations of the methylene groups was found at 2924 cm−1. As it can be seen from the spectra in Figure 5, the incorporation of AgNPs containing fructose in their cover layer in the liposomes did not notably influence the position of the band—it remained almost unchanged, varying between 2924 and 2923 cm−1. The frequency of the IR band for the symmetric stretching vibrations of the methylene groups also did not show a substantial change upon the addition of the NPs—it was found in all samples at ca. 2853–2854 cm−1. These wavenumbers correspond to a partially disordered state of the hydrocarbon chains as the bands shifted more than 5 cm−1 from the typical positions for lipids in the ordered phase [11,37,38]. However, between the samples, no significant change was observed, suggesting that the NPs had little effect on the packing of the hydrocarbon tails.
The characteristic IR band of the ester carbonyl (C=O) stretching vibration of pure SOPC was found at 1743 cm−1 and corresponded to partially hydrated carbonyl groups (Figure 5). In the liposomes containing silver nanoparticles with a fructose capping agent, the carbonyl band low-frequency shifted by approximately 10 cm−1, 1736–1732 cm−1, which evidenced the increased hydrogen bonding of the carbonyl groups. The observed changes in the C=O stretching band position was related to the interactions of the studied NPs that contained fructose in their cover layer with the ester groups of the SOPC membrane; obviously, the carbonyl groups of the SOPC molecules were involved in stronger and more hydrogen bonding interactions in the presence of the NPs. This effect could be attributed to the strong hydrogen bonding capacity of the polar fructose molecules, which are rich in hydroxy groups.
The IR bands for the phosphate stretching vibrations shifted from 1232 cm−1 in the pure SOPC (characteristic of partially hydrated phosphate groups) to 1228 cm−1 in the liposomes containing 0.5 and 1% NPs (Figure 5). This shift indicated that the hydrogen bonding interactions of the phosphate groups also increased in the presence of AgNPs containing fructose in their cover surface. Considering that the stretching vibrations of both the carbonyl and the phosphate groups were affected, the NPs may have incorporated within the polar region of the SOPC membrane.
In the IR spectrum of the sample containing 2.5% NPs, the position of the latter band was influenced by the emerging absorptions for C-O and C-C stretching vibrations of the fructose molecules in this region and could not be used for the assessment of the hydrogen bonding of the SOPC phosphate groups. In the region below 1250 cm−1, several bands arising from the fructose C–O and C–C stretching vibrations were observed, with the strongest bands appearing at 1160, 1101, 1085, 1064, and 1016 cm−1. The bands were the most intense in the spectrum of the sample containing 2.5% NPs, in accordance with it containing the highest amounts of fructose.

3.6. Differential Scanning Calorimetry (DSC)

Previous reports have shown that a water content of 10–20 wt% yields optimal thermotropic profiles for SOPC liposomes [39]. To ensure consistency and comparability, all SOPC samples with varying concentrations of AgNPs were prepared with 10 wt% water. At higher heating rates (e.g., from 2 to 5 °C/min), it was observed that both the transition temperature and calorimetric enthalpy increase [39]. A heating rate of 5 °C/minute was chosen as optimal for the DSC equipment, simultaneously balancing sensitivity and accuracy. For thermal analysis, each sample was equilibrated at room temperature before being placed in the DSC oven. The samples were first heated to 50 °C, followed by controlled cooling at a rate of 5 °C per minute down to −10 °C, based on the aforementioned experimental details. Subsequently, the samples were reheated from −10 °C to 70 °C at the same rate. This protocol ensured a consistent thermal history for all samples regardless of the AgNP concentration in the SOPC liposomes. Figure 6a,b show the differential scanning calorimetry (DSC) thermograms of SOPC liposomes, both in the absence and in the presence of various concentrations of AgNPs during heating and cooling cycles. The DSC results indicate that the incorporation of AgNPs into SOPC liposomes slightly alters the phase transition behavior. As the concentration of AgNPs increases, the main phase transition from the gel to the liquid crystalline state shifts to lower temperatures. This shift is evident in both the heating and cooling profiles, where the main endothermic and exothermic peaks move leftward on the temperature axis with increasing NP content.
For all studied concentrations of AgNPs, the transition peak became lightly broader and less defined compared to that of pure SOPC, suggesting a more difficult or more gradual phase change [40,41]. This effect can be observed in both the heating (Figure 6a) and cooling (Figure 6b) thermograms. The SOPC samples with the highest AgNP content of 2.5% show the most diffuse peak among the samples tested, indicating a subtle alteration in membrane organization. The enthalpy associated with the transition also decreases as more Ag-based nanocarriers are introduced (Table 1), reflecting a reduction in the energy required for the phase transformation. These changes are consistent with a membrane that undergoes a less ordered transition due to the incorporation of NPs. The quantified DSC results, summarized in Table 1, support the observations of the graphs.
For pure SOPC liposomes, the main phase transition temperature (Tm) during cooling is 3.41 °C. As the concentration of AgNPs increases, Tm steadily decreases, reaching 2.80 °C at 2.5 wt% AgNPs. The transition enthalpy (ΔH) also decreases with increasing nanoparticle concentration, from 0.280 J/g [Appendix A, Appendix B] in pure SOPC to 0.234 J/g at the maximal NPs content, reflecting a reduction in the energetic benefits of the phase transformation. A similar trend is observed for the heating cycle, where Tm slightly increases with increasing AgNPs concentration, but the heating enthalpy (ΔH) monotonically decreases. Together, these results suggest that the AgNPs induce subtle changes in the SOPC bilayer, resulting in broader and less-defined phase transitions. This effect facilitates membrane fluidization at reduced temperatures, supporting the theory of the phase transitions of systems with diluted defects [42].

3.7. Thermally Induced Shape Fluctuation Method

To ensure reliable and comparable results, we used thermally induced fluctuation to investigate the effect of AgNPs on SOPC lipid membranes, incorporating AgNP concentrations up to 2.5 wt%. The outcomes obtained with AgNPs were compared with those from pure SOPC and SOPC membranes in the presence of glucose [43,44,45].
Following a well-established methodology, we systematically investigated how AgNPs affect the elastic properties of SOPC lipid membranes. Similar to the bilayer softening with sugar, we anticipated that the adsorption and capping chemistry of AgNPs—particularly with fructose cap layers—would alter bending rigidity through surface pressure effects and interactions with lipid-head–group interactions. While higher concentrations sometimes led to membrane imperfections that complicated the formation of the large spherical vesicles needed for accurate analysis, we were still able to successfully measure the moduli of bending elasticity (kc) for the SOPC giant unilamellar vesicles (GUV) containing 1.0% and 2.5% AgNPs. These results are summarized in Table 2, presented as the weighted average from at least five separate vesicles. The data suggest that, within the margin of error, the AgNPs at the tested concentrations did not significantly alter the elastic behavior of the SOPC bilayer. For a statistical outlook, the experimental values of kc are presented in a histogram in Figure 7, thus giving a direct comparison of the bending elasticity modulus in the presence of fructose. Such a comparison is not accidental since sugars—like NPs—modulate bilayer mechanics through hydration and head–group interactions.
Sugars, well-known cryoprotectants under extreme conditions, displace water from the hydrophilic part of the molecule by forming multiple hydrogen bonds with the phospholipid head groups. This alters the tendency of lipid molecules to bind to water. However, as highlighted by Cacela and Hincha [46] and further discussed in disaccharide-membrane studies [47], once a critical concentration is reached, additional sugar molecules no longer displace water, and the system reaches equilibrium, with no further significant changes.
The progressive decrease in kc with the increase in the concentration of AgNPs suggests that NPs alter the lipid packing, reducing the bending rigidity of the bilayer. This observation is consistent with the DSC results, which show shifting and broadening of the main transition, supporting the interpretation that incorporating NPs induced subtle changes in membrane properties. This could be reflected in minor membrane destabilization or enhanced fluidity as a result of nanoparticle incorporation. The bending elastic modulus in the presence of fructose is included as a reference, which, as shown, has a much lower kc. This underlines that even with reduced rigidity, SOPC membranes with AgNPs are still significantly stiffer than lipid membranes in the presence of fructose (which has a bending elasticity modulus of almost half of that of pure SOPC). From a biomedical perspective, the observed concentration-dependent softening of SOPC membranes suggests that high AgNP loadings lead to subtle alterations in membrane integrity and thereby contribute to antimicrobial effects, although in vivo behavior will additionally depend on protein coronas, ionic environment, and cellular repair mechanisms.

3.8. Temperature Changes Under the Effect of Laser Irradiation in the Presence of Silver Nanoparticles

The experimental environment for cell cultures is suitable for the investigation of temperature changes under laser irradiation. In the presence of AgNPs, laser irradiation leads to a significant increase in the temperature, with the temperature dynamics depending on both the wavelength and power density of the radiation. The temperature response of the medium was investigated under laser irradiation at three wavelengths (1030 nm, 515 nm, and 343 nm) and power densities of 0.1 and 0.2 W/cm2 in the presence of the AgNPs. Dulbecco’s Modified Eagle Medium (DMEM) served as the reference medium for all measurements. All the experiments in this study were performed at 100 kHz repetition rate. To assess the temperature variation induced by laser exposure, AgNP suspensions were introduced into the medium at a concentration of 50 µL/mL.
A FOTRIC 320F thermal imaging camera (FOTRIC Inc., San Diego, California (CA); USA) was used to measure temperature changes in the studied systems during their laser irradiation. The results obtained for all studied cases are shown in Figure 8. The y-axis corresponds to the temperature differences in the silver nanoparticles throughout a 10 min irradiation process, with time indicated on the x-axis. The most pronounced thermal effect is registered at λ = 515 nm, where, at a higher power density (0.2 W/cm2), the temperature increase reaches over 30 °C within 10 min. This highlights effective absorption and conversion of optical energy into heat.
At a wavelength of 343 nm, a distinct temperature increase was also observed, al- though lower compared to that at 515 nm. In contrast, at λ = 1030 nm, the thermal effect was significantly weaker, regardless of the power density used. These results clearly demonstrate the spectral dependence of the photothermal response in the system and emphasize the key role of the plasmon resonance of AgNPs in the green range of the spectrum. This spectral behavior highlights their potential for applications in photothermal therapy, where selective heating in the presence of NPs can be used to target pathological cells with a minimal impact on healthy tissues.

4. Conclusions

In conclusion, our study demonstrates that the functional properties of lipid vesicles are profoundly and concentration-dependently altered by the incorporation of silver nanoparticles (AgNPs). We elucidated a multifaceted mechanism of interaction: FTIR-ATR spectroscopy revealed that AgNPs preferentially interact with carbonyl and phosphate groups in the lipid head region [48], while DSC thermograms indicated a perturbation of the hydrophobic core, leading to a broadened and less-energetically beneficial phase transition. This disruption in lipid packing was directly linked to a concentration-dependent softening of the membrane, as measured by a reduction in bending rigidity.
These findings have two critical implications. On the one hand, the nanoparticle-induced membrane destabilization provides a plausible mechanism for the known antimicrobial activity of AgNPs. On the other hand, the strong, wavelength-specific photothermal response of AgNPs confirms their potential for applications in controlled hyperthermia therapies. These two features underscore a central tenet of nanomedicine: the same property that confers therapeutic benefit (membrane interaction) can also pose a risk to membrane integrity.
Therefore, the successful implementation of AgNP–liposome hybrids in biotechnology and medicine hinges on a precise, quantitative understanding of these interactions. Future work will focus on translating these findings from biomimetic models to real cellular systems to validate their biological relevance and therapeutic efficacy.

Author Contributions

Conceptualization, J.G. and L.S.; methodology, J.G., L.S., M.L. and D.Y.; validation, J.G., L.S., M.L. and D.Y.; formal analysis, J.G., L.S., M.L. and D.Y.; investigation, J.G., L.S., M.L. and D.Y.; resources, J.G., L.S., M.L., S.N., M.T. and D.Y.; data curation, J.G., L.S., M.L. and D.Y.; writing—original draft preparation, M.L.; writing—review and editing, J.G., L.S., M.L., N.K., K.H.-P., C.X. and D.Y.; visualization, J.G., L.S., M.L., S.N., M.T. and D.Y.; supervision, J.G. and L.S.; project administration, L.S.; funding acquisition, J.G., L.S. and N.K. All authors have read and agreed to the published version of the manuscript.

Funding

This research received funding in 2023 via the Bulgarian National Science Fund (KP-06-N78/8 from 14 December 2023).

Data Availability Statement

Additional data are available upon request.

Acknowledgments

D.Y. is grateful for the financial support from the European Union NextGenerationEU, through the National Recovery and Resilience Plan of the Republic of Bulgaria, Project No. BG-RRP-2.004-0002, “BiOrgaMCT”. Equipment from INFRAMAT (Research Infrastructure from National roadmap of Bulgaria), supported by the Bulgarian Ministry of Education and Science, was used in a part of the present investigation.

Conflicts of Interest

The authors declare no conflicts of interest.

Abbreviations

The following abbreviations are used in this manuscript:
AgNPsSilver nanoparticles
DSCDifferential scanning calorimetry
SOPC1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine
FTIR-ATRFourier-transform infrared spectroscopy with attenuated total reflection
IR spectroscopyInfrared spectroscopy
PTTPhotothermal therapy
NIR lightNear-infrared light
EPR effectsEnhanced permeability and retention
TEMTransmission electron microscopy

Appendix A. Calculation of Heating Enthalpies of a Set of Samples by DSC (Differential Scanning Calorimetry)

The calculation of enthalpy values from the DSC thermograms in this study was performed using a dedicated C++ console program developed in accordance with the protocol established by Maria Lyudmilova. This protocol served as the reference standard for the experimental and computational methodology employed.
The console application processes raw thermogram data (temperature and heat flow) for each sample and calculates the enthalpy associated with the phase transitions observed during heating or cooling. Central features of the program include
  • File-based data input: the user specifies the name of a text file containing tabulated temperature and normalized heat flow measurements from the DSC experiment.
  • Trapezoidal numerical integration: the enthalpy (ΔH) for the relevant transition is computed using the trapezoidal rule, applying the general formula.
  • User-defined integration range: for each measurement, the user selects the temperature interval for integration to fully capture the enthalpic peak (as determined by the protocol guidance for each sample type).
  • Calculation of error due to protocol: the program permits entry of a protocol reference value for enthalpy, against which the calculated result is compared, providing absolute and relative errors.
  • Output: all results (file name, calculated ΔH, reference value, error) are displayed and saved as a C.SV file.

Appendix B. Protocol Adherence and Accuracy

The analysis intervals for each sample and nanoparticle concentrations are based on the peak positions and widths specified in the established protocol. This ensures methodological consistency and enables reliable comparison to experimentally validated reference values. The program’s logic and calculation steps mirror those performed in spreadsheet analyses and yield results within accepted experimental error margins (typically <5%). This software (Version 1.0) tool provides a reproducible and automated means for enthalpy calculation in SOPC/AgNP systems and can be adapted for other lipid/nanoparticle combinations as required.

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Applsci 16 00220 g001
Figure 1. Schematic representation of SOPC’s lipid structure.
Figure 1. Schematic representation of SOPC’s lipid structure.
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Figure 2. TEM characterization of AgNPs showing morphology and surface capping. (a) TEM image of biosynthesized AgNPs showing predominantly spherical morphology. (b) Higher-magnification TEM image showing a thin fructose layer (≈2–15 nm) around AgNPs, indicating successful capping and stabilization.
Figure 2. TEM characterization of AgNPs showing morphology and surface capping. (a) TEM image of biosynthesized AgNPs showing predominantly spherical morphology. (b) Higher-magnification TEM image showing a thin fructose layer (≈2–15 nm) around AgNPs, indicating successful capping and stabilization.
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Figure 3. Dynamic light scattering histograms of AgNPs.
Figure 3. Dynamic light scattering histograms of AgNPs.
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Figure 4. Time-dependent UV–Vis absorption spectra of fructose-capped AgNPs recorded in the 190–790 nm range, showing the emergence of the surface plasmon resonance band at ≈415 nm during nanoparticle formation.
Figure 4. Time-dependent UV–Vis absorption spectra of fructose-capped AgNPs recorded in the 190–790 nm range, showing the emergence of the surface plasmon resonance band at ≈415 nm during nanoparticle formation.
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Figure 5. ATR-FTIR spectra in the region of 3000 to 900 cm−1 of pure SOPC and three SOPC samples with varying amounts of AgNPs measured by referencing water as a background.
Figure 5. ATR-FTIR spectra in the region of 3000 to 900 cm−1 of pure SOPC and three SOPC samples with varying amounts of AgNPs measured by referencing water as a background.
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Figure 6. Thermal behavior of lipid systems containing 0–2.5%wt AgNPs. (a) Heating; (b) cooling. (a) DSC thermograms of the SOPC lipid system containing 0.5, 1.0, and 2.5 wt% silver nanoparticles during heating. (b) DSC thermograms of the SOPC lipid system containing 0.5, 1.0, and 2.5 wt% silver nanoparticles during cooling.
Figure 6. Thermal behavior of lipid systems containing 0–2.5%wt AgNPs. (a) Heating; (b) cooling. (a) DSC thermograms of the SOPC lipid system containing 0.5, 1.0, and 2.5 wt% silver nanoparticles during heating. (b) DSC thermograms of the SOPC lipid system containing 0.5, 1.0, and 2.5 wt% silver nanoparticles during cooling.
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Figure 7. Bending elasticity moduli of pure SOPC and SOPC containing AgNPs.
Figure 7. Bending elasticity moduli of pure SOPC and SOPC containing AgNPs.
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Figure 8. Thermal changes in DMEM with AgNPs during irradiation at three wavelengths.
Figure 8. Thermal changes in DMEM with AgNPs during irradiation at three wavelengths.
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Table 1. Thermal properties of SOPC liposomes (10% ddH2O) with various AgNP concentrations during cooling and heating.
Table 1. Thermal properties of SOPC liposomes (10% ddH2O) with various AgNP concentrations during cooling and heating.
SampleEXO Tm (°C)EXO ΔH (J/g)ENDO Tm (°C)ENDO ΔH
(J/g)
Pure3.410.2805.800.263
+0.5% AgNPs3.200.2795.800.306
+1.0% AgNPs3.330.2615.920.264
+2.5% AgNPs2.800.2346.300.258
Table 2. Bending elasticity modulus (kc) of the lipid membrane containing various concentrations of AgNPs, acquired through thermally induced shape fluctuation analysis. Bending modulus values are presented as weighted averages of five vesicles per concentration.
Table 2. Bending elasticity modulus (kc) of the lipid membrane containing various concentrations of AgNPs, acquired through thermally induced shape fluctuation analysis. Bending modulus values are presented as weighted averages of five vesicles per concentration.
Experimental SystemPure SOPCSOPC + 1.0
wt% Ag		SOPC + 2.5
wt% Ag		200 mM
Fructose
kc × 10−19 [J]1.18 ± 0.041.02 ± 0.100.93 ± 0.080.57 ± 0.04
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Lyudmilova, M.; Stoychev, L.; Yancheva, D.; Nikolova, S.; Todorova, M.; Xenodochidis, C.; Hristova-Panusheva, K.; Krasteva, N.; Genova, J. Influence of Silver Nanoparticles on Liposomal Membrane Properties Relevant in Photothermal Therapy. Appl. Sci. 2026, 16, 220. https://doi.org/10.3390/app16010220

AMA Style

Lyudmilova M, Stoychev L, Yancheva D, Nikolova S, Todorova M, Xenodochidis C, Hristova-Panusheva K, Krasteva N, Genova J. Influence of Silver Nanoparticles on Liposomal Membrane Properties Relevant in Photothermal Therapy. Applied Sciences. 2026; 16(1):220. https://doi.org/10.3390/app16010220

Chicago/Turabian Style

Lyudmilova, Maria, Lyubomir Stoychev, Denitsa Yancheva, Stoyanka Nikolova, Mina Todorova, Charilaos Xenodochidis, Kamelia Hristova-Panusheva, Natalia Krasteva, and Julia Genova. 2026. "Influence of Silver Nanoparticles on Liposomal Membrane Properties Relevant in Photothermal Therapy" Applied Sciences 16, no. 1: 220. https://doi.org/10.3390/app16010220

APA Style

Lyudmilova, M., Stoychev, L., Yancheva, D., Nikolova, S., Todorova, M., Xenodochidis, C., Hristova-Panusheva, K., Krasteva, N., & Genova, J. (2026). Influence of Silver Nanoparticles on Liposomal Membrane Properties Relevant in Photothermal Therapy. Applied Sciences, 16(1), 220. https://doi.org/10.3390/app16010220

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