**1. Introduction**

It is well known that the body barrier to external pathogenic attacks is represented by the skin, which prevents microbial invasion, so every damage or wound can provide an environment for microbial growth, leading to infection and prolonged wound healing [1–6]. The e fficacy of antibiotics is superior to that of other drugs; thus, antibiotics are widely used [7]. However, the antibiotic resistance is currently a health emergency, so drug-resistant bacterial infections are becoming more common with a consequent increase in public spending [8,9]. About 60–70% of the existing antibiotics are not

active against intracellular infections due to their low intracellular retention as a result of their poor permeability. Nanomaterials represent an attractive solution for the hydrophilicity barrier, because they can habitually penetrate cells, and increase their intracellular activity [10,11]. Small-size nanoparticles have the advantage of being characterized by a larger contact surface, which is able to enhance their penetration and therapeutic effects [12–15].

Among others nanomaterials, a good candidate is represented by silver nanoparticles (AgNPs) that can be easy functionalized [16,17], inducing hydrophilic behavior [18,19], antibacterial properties [13,20], and reduced inflammatory response [21], with low resistance phenomena in Gram-positive bacteria and Gram-negative bacteria [22–24]. The dimension and shape of AgNPs have a strong influence on antibacterial activity. In fact, a higher surface/volume ratio produces the higher rate of silver ions release [25]. AgNPs' effects have been studied against the multidrug-resistant bacteria such as *P. aeruginosa, E. coli, Streptococcus pyogenes, S. aureus, Klebsiella pneumoniae, Salmonella* species, and *Enterococcus species* [26,27]. In these papers, the bactericidal action is mainly due to the inhibition of cell wall synthesis, nucleic acid synthesis, and protein synthesis mediated by the 30S ribosomal subunit. The strong bactericidal effect of AgNPs against the multidrug-resistant bacteria is mostly due to their multiple mechanisms to disrupt microbial cells [28]. Moreover, AgNPs can improve the antibiotic effects against *S. aureus* and *E. coli* [29].

Today, in medicinal practice, there are wound dressings, contraceptive devices, surgical instruments, bone prostheses, and dental implants that are coated or embedded with nanosilver [30–33]. Moreover, in the last decade, the research field of AgNPs has moved to the possibility of their use as an anticancer drug, due to their inherent cytotoxic effect on cancer cells [34].

However, the instability of silver nanoparticles limits their industrial application in several cases, and most of the methods to prepare AgNPs cause environmental pollution and low production efficiency. To overcome this problem, silver nanoparticles are usually loaded onto carriers [35].

Moreover, despite AgNPs having multiple mechanisms for antibacterial effects, recent studies showed bacterial resistance to them: the resistance evolves without any genetic changes; only phenotypic change is needed to reduce the nanoparticles' colloidal stability and thus eliminate their antibacterial activity [36,37].

As a response to this problem, hybrids/composites with AgNPs dispersed on carriers or supports have been studied to enhance antibacterial activity compared with sole AgNPs: it is of significance to seek the optimal choice of carriers to combine with AgNPs in order to construct ideal antibacterial agents. Various AgNPs-based nanocomposites with different structures and morphologies have been developed up to now, such as an amorphous silica matrix dispersed with AgNPs [38], AgNPs core@silica shell [39], mesoporous silicas loaded with AgNPs [40], hollow mesoporous silica spheres with AgNPs in the cavity [41,42], fibers coated with AgNPs [43], etc. Although extensive efforts have been devoted to fabricating a lot of these AgNPs-based nanocomposites involving different carriers' structures, there are still few systematic investigations on the effects of structures on antibacterial performance [44]. The use of drug delivery systems (DDS) has been proposed to overcome important issues in the release of active pharmaceutical molecules, such as unfavorable pharmacokinetics and biodistribution with a consequent decrease of side effects.

Nanocarriers represent an innovative approach to overcome these issues [45,46]. Among other nanocarriers, such as liposomes, polymersomes, micelles, and polymer-based vesicles, the niosomes systems, non-ionic surfactant vesicles [46,47], have attracted attention from researchers because of their ability to encapsulate different kinds of drugs for the purpose of increasing their stability and efficacy. In fact, niosomes enable modulating the drug concentration loading in a range of interest for the biological applications (0.3–5.0 μg/mL for AgNPs) and to consent to drug-release control [48].

In this research study, AgNPs were synthetized using 3-mercapto-1-sodium propanesulfonate (3MPS) to induce hydrophilic behavior, improving the niosomal entrapment efficiency and reducing the bilayer destabilization. AgNPs were loaded in two different niosomes, Tween 20 and Span 20 ones, producing two different systems, namely NioTw20 + AgNPs and NioSp20 + AgNPs. A deep

physical chemical characterization was carried out to obtain information on hydrophilic AgNPs and their influence on the preparation and characterization of Nio + AgNPs. Moreover, stability studies were performed in water, bovine serum, and human serum to assess their use in biological compartments. Hydrophilic and lipophilic probe release profiles were obtained in HEPES and in human serum. Both systems proved to be able to entrap AgNPs, are stable, and maintain the ability to entrap also hydrophilic or lipophilic model molecules, and so are promising systems for biotechnological applications.
