Entrapment of Amphipathic Drugs in Core-Shell Polymeric Nanoparticles under Batch Conditions─The Role of Control and Solubility Parameters.

The amphipathic bioactive compounds curcumin, resveratrol, and mitomycin C, which have similar solubility parameter component distributions, have been studied for encapsulation under batch conditions into core-shell nanocarriers composed of external hydrophobically functionalized polyelectrolytes and an inner matrix of polyesters or polyester blends: poly(l-lactide), poly(lactide-co-glycolide), and/or poly(ethylene succinate). Our contribution comprises determining the influence of process parameters on the properties and quality of the final products, namely core-shell nanoparticles loaded with appropriate drugs, according to process analysis technologymanagement. The crucial roles of the organic phase dosing rates and process temperatures were carefully investigated. Moreover, a technically feasible method of removing organic solvents from aqueous dispersions─stripping with inert gas─was employed and evaluated via FT-IR studies. The experiments were supported by the calculation and analysis of solubility parameters (δ) and dispersion (δd), polar (δp), and hydrogen bond (δh) components utilizing HSPiP software. The payload locus and sample morphology were studied via atomic force microscopy and X-ray photoelectron spectroscopy analyses with Ar+ sputtering. It was demonstrated that dosing rates of organic phases not exceeding ca. 0.5 mL/min per 1 L of aqueous dispersion of hydrophobically functionalized polyelectrolytes made it possible to obtain core-shell nanoparticles of ca. 100-150 nm with a very narrow polydispersity (PdI < 0.2). The locus of amphipathic payloads in nanocarriers, mostly within the core polymeric structure, was in good agreement with the results of solubility parameter component studies: water-insoluble polyesters with both polar and nonpolar interactions between chains serve as good host materials for amphipathic drugs.

Anti-adhesive, anti-biofilm and fungicidal action of newly synthesized gemini quaternary ammonium salts.

Newly synthesized gemini quaternary ammonium salts (QAS) with different counterions (bromide, hydrogen chloride, methylcarbonate, acetate, lactate), chain lengths (C12, C14, C16) and methylene linker (3xCH2) were tested. Dihydrochlorides and dibromides with 12 carbon atoms in hydrophobic chains were characterized by the highest biological activity against planktonic forms of yeast and yeast-like fungi. The tested gemini surfactants also inhibited the production of filaments by C. albicans. Moreover, they reduced the adhesion of C. albicans cells to the surfaces of stainless steel, silicone and glass, and slightly to polystyrene. In particular, the gemini compounds with 16-carbon alkyl chains were most effective against biofilms. It was also found that the tested surfactants were not cytotoxic to yeast cells. Moreover, dimethylcarbonate (2xC12MeCO3G3) did not cause hemolysis of sheep erythrocytes. Dihydrochlorides, dilactate and diacetate showed no mutagenic potential.

Newly synthesized surfactants as antimicrobial and anti-adhesion agents.

Quaternary ammonium salts (QAS) are widely used in medicine, industry and agriculture as disinfectants, biocides, and fungicides. QAS have the ability to coat various surfaces, prevent adhesion of microorganisms to them and inhibit the formation of biofilm. A group of surfactants derived from benzoic acid with different chemical structures was tested: monomeric QAS with different alkyl chain lengths (C12, C14, C16), gemini QAS containing 12-carbon alkyl chains and linkers of various lengths (3,4,6 methylene groups), as well as multifunctional QAS. Among the tested surfactants, monomeric QAS showed the highest bactericidal and fungicidal activity. All three groups of tested compounds inhibited the filamentation of C. albicans. The best antimicrobial activity was demonstrated by the monomeric surfactant C12AA, while the multifunctional equivalent (2xC12AA) was characterized by good anti-adhesive activity. All tested compounds are non-mutagenic and cause low hemolysis of sheep erythrocytes. Multifunctional and gemini surfactants are also non-toxic.

A Combination of ß-Aescin and Newly Synthesized Alkylamidobetaines as Modern Components Eradicating the Biofilms of Multidrug-Resistant Clinical Strains of Candida glabrata.

The current trend in microbiological research aimed at limiting the development of biofilms of multidrug-resistant microorganisms is increasingly towards the search for possible synergistic effects between various compounds. This work presents a combination of a naturally occurring compound, ß-aescin, newly synthesized alkylamidobetaines (AABs) with a general structure-CnTMDAB, and antifungal drugs. The research we conducted consists of several stages. The first stage concerns determining biological activity (antifungal) against selected multidrug-resistant strains of Candida glabrata (C. glabrata) with the highest ability to form biofilms. The second stage of this study determined the activity of ß-aescin combinations with antifungal compounds and alkylamidobetaines. In the next stage of this study, the ability to eradicate a biofilm on the polystyrene surface of the combination of ß-aescin with alkylamidobetaines was examined. It has been shown that the combination of ß-aescin and alkylamidobetaine can firmly remove biofilms and reduce their viability. The last stage of this research was to determine the safety regarding the cytotoxicity of both ß-aescin and alkylamidobetaines. Previous studies on the fibroblast cell line have shown that C9 alkylamidobetaine can be safely used as a component of anti-biofilm compounds. This research increases the level of knowledge about the practical possibilities of using anti-biofilm compounds in combined therapies against C. glabrata.

Multiheaded Cationic Surfactants with Dedicated Functionalities: Design, Synthetic Strategies, Self-Assembly and Performance.

Contemporary research concerning surfactant science and technology comprises a variety of requirements relating to the design of surfactant structures with widely varying architectures to achieve physicochemical properties and dedicated functionality. Such approaches are necessary to make them applicable to modern technologies, such as nanostructure engineering, surface structurization or fine chemicals, e.g., magnetic surfactants, biocidal agents, capping and stabilizing reagents or reactive agents at interfaces. Even slight modifications of a surfactant's molecular structure with respect to the conventional single-head-single-tail design allow for various custom-designed products. Among them, multicharge structures are the most intriguing. Their preparation requires specific synthetic routes that enable both main amphiphilic compound synthesis using appropriate step-by-step reaction strategies or coupling approaches as well as further derivatization toward specific features such as magnetic properties. Some of the most challenging aspects of multicharge cationic surfactants relate to their use at different interfaces for stable nanostructures formation, applying capping effects or complexation with polyelectrolytes. Multiheaded cationic surfactants exhibit strong antimicrobial and antiviral activity, allowing them to be implemented in various biomedical fields, especially biofilm prevention and eradication. Therefore, recent advances in synthetic strategies for multiheaded cationic surfactants, their self-aggregation and performance are scrutinized in this up-to-date review, emphasizing their applications in different fields such as building blocks in nanostructure engineering and their use as fine chemicals.

The effectiveness of newly synthesized quaternary ammonium salts differing in chain length and type of counterion against priority human pathogens.

Quaternary ammonium salts (QAS) commonly occur as active substances in disinfectants. QAS have the important property of coating abiotic surfaces, which prevents adhesion of microorganisms, thus inhibiting biofilm formation. In this study, a group of nine monomeric QAS, differing in the structure and length of the aliphatic chain (C12, C14, C16) and the counterion (methylcarbonate, acetate, bromide), were investigated. The study included an analysis of their action against planktonic forms as well as bacterial biofilms. The compounds were tested for their anti-adhesion properties on stainless steel, polystyrene, silicone and glass surfaces. Moreover, mutagenicity analysis and evaluation of hemolytic properties were performed. It was found that compounds with 16-carbon hydrophobic chains were the most promising against both planktonic forms and biofilms. Tested surfactants (C12, C14, C16) showed anti-adhesion activity but it was dependent on the type of the surface and strain used. The tested compounds at MIC concentrations did not cause hemolysis of sheep blood cells. The type of counterion was not as significant for the activity of the compound as the length of the hydrophobic aliphatic chain.

Rational Mitomycin Nanocarriers Based on Hydrophobically Functionalized Polyelectrolytes and Poly(lactide-co-glycolide).

Encapsulation of hydrophilic and amphiphilic drugs in appropriate colloidal carrier systems for sustained release is an emerging problem. In general, hydrophobic bioactive substances tend to accumulate in water-immiscible polymeric domains, and the release process is controlled by their low aqueous solubility and limited diffusion from the nanocarrier matrix. Conversely, hydrophilic/amphiphilic drugs are typically water-soluble and insoluble in numerous polymers. Therefore, a core-shell approach─nanocarriers comprising an internal core and external shell microenvironments of different properties─can be exploited for hydrophilic/amphiphilic drugs. To produce colloidally stable poly(lactic-co-glycolic) (PLGA) nanoparticles for mitomycin C (MMC) delivery and controlled release, a unique class of amphiphilic polymers─hydrophobically functionalized polyelectrolytes─were utilized as shell-forming materials, comprising both stabilization via electrostatic repulsive forces and anchoring to the core via hydrophobic interactions. Undoubtedly, the use of these polymeric building blocks for the core-shell approach contributes to the enhancement of the payload chemical stability and sustained release profiles. The studied nanoparticles were prepared via nanoprecipitation of the PLGA polymer and were dissolved in acetone as a good solvent and in an aqueous solution containing hydrophobically functionalized poly(4-styrenesulfonic-co-maleic acid) and poly(acrylic acid) of differing hydrophilic-lipophilic balance values. The type of the hydrophobically functionalized polyelectrolyte (HF-PE) was crucial for the chemical stability of the payload─derivatives of poly(acrylic acid) were found to cause very rapid degradation (hydrolysis) of MMC, in contrast to poly(4-styrenesulfonic-co-maleic acid). The present contribution allowed us to gain crucial information about novel colloidal nanocarrier systems for MMC delivery, especially in the fields of optimal HF-PE concentrations, appropriate core and shell building materials, and the colloidal and chemical stability of the system.

Biocompatible poly(ethylene succinate) polyester with molecular weight dependent drug release properties.

In the present study, we demonstrate that well-known molecular weight-dependent solubility properties of a polymer can also be used in the field of controlled drug delivery. To prove this, poly(ethylene succinate) (PES) polyesters with polycondensation time regulated molecular weights were synthesized via catalyst-free direct polymerization in an equimolar ratio of ethylene glycol and succinic acid monomers at 185 °C. DSC and contact angle measurements revealed that increasing the molecular weight (Mw, 4.3-5.05 kDa) through the polymerization time (40-80 min) increased the thermal stability (Tm= ∼61-80 °C) and slightly the hydrophobicity (Θw= ∼27-41°) of the obtained aliphatic polyester. Next, this biodegradable polymer was used for the encapsulation of Ca2+ channel blocker Nimodipine (NIMO) to overcome the poor water solubility and enhance the bioavailability of the drug. The drug/ polymer compatibility was proved by the means of solubility (δ) and Flory-Huggins interaction (miscibility) parameters (χ). The nanoprecipitation encapsulation of NIMO into PES with increasing Mw resulted in the formation of spherical 270 ± 103 nm NIMO-loaded PES nanoparticles (NPs). Furthermore, based on the XRD measurements, the encapsulated form of NIMO-loaded PES NPs showed lower drug crystallinity, which enhanced not only the water solubility but even the water stability of the NIMO in an aqueous medium. The in-vitro drug release experiments demonstrated that the release of NIMO drug could be accelerated or even prolonged by the molecular weights of PES as well. Due to the low crystallinity of PES polyester and low particle size of the encapsulated NIMO drug led to enhance solubility and releasing process of NIMO from PES with lower Mw (4.3 kDa and 4.5 kDa) compared to pure crystalline NIMO. However, further increasing the molecular weight (5.05 kDa) was already reduced the amount of drug release that provides the prolonged therapeutic effect and enhances the bioavailability of the NIMO drug.

Multilayered Curcumin-Loaded Hydrogel Microcarriers with Antimicrobial Function.

The design of multifunctional microcarriers has attracted significant attention because they combine various functions within a single system. In this study, we developed a set of multilayered hydrogel microcarriers, which were first loaded with chemotherapeutic curcumin (CUR), then, using the layer-by-layer (LbL) technique, coated through a polyelectrolyte shell consisting of chitosan (CHIT) or poly(allylamine hydrochloride) (PAH). As an outer layer with antimicrobial function, newly synthesised alkylene quaternary ammonium salt functionalised polyelectrolytes (A-QAS-PEs) were applied. For this purpose, poly(acrylic acid) (PAA) was decorated with different hydrophobic side chains (n-hexane and n-dodecane side entities) and different degrees of substitution (m) of quaternary ammonium groups (abbreviated as PAA-C(O)O-(CH2)n-N+(CH3)3(m); n = 6, 12; m = 8-14%). The grafting approach of PAA with the alkylene quaternary ammonium salt moiety was performed under mild reaction conditions using Steglich esterification followed by quaternisation. The structure of antimicrobial decorated PAA was confirmed by 1H NMR and FTIR, and the mean diameter of all multifunctional microparticles was characterised by SEM. The viscoelastic properties of the functional layers were studied using quartz crystal microbalance with a dissipation (QCM-D). The release of CUR from the microcarriers was described using a hybrid model, i.e., a combination of first-order kinetics and the Korsmeyer-Peppas model. The antimicrobial activity of functionalised PAA and multilayered CUR-loaded hydrogel microcarriers with quaternary ammonium function was assessed against Staphylococcus aureus and Serratia marcescens by the agar diffusion assay method. Only a limited inhibition zone of PAA was observed, but in the case of both antimicrobial decorated PAA and the corresponding multilayered nanocarriers, the inhibitory activity increase was achieved against both strains of bacteria.

The Effect of Molecular Weight on the Solubility Properties of Biocompatible Poly(ethylene succinate) Polyester.

Poly(ethylene succinate) (PES) is one of the most promising biodegradable and biocompatible polyesters and is widely used in different biomedical applications. However, little information is available on its solubility and precipitation properties, despite that these solution behavior properties affect its applicability. In order to systematically study these effects, biodegradable and biocompatible poly(ethylene succinate) (PES) was synthesized using ethylene glycol and succinic acid monomers with an equimolar ratio. Despite the optimized reaction temperature (T = 185 °C) of the direct condensation polymerization, relatively low molecular mass values were achieved without using a catalyst, and the Mn was adjustable with the reaction time (40-100 min) in the range of ~850 and ~1300 Da. The obtained crude products were purified by precipitation from THF ("good" solvent) with excess of methanol ("bad" solvent). The solvents for PES oligomers purification were chosen according to the calculated values of solubility parameters by different approaches (Fedors, Hoy and Hoftyzer-van Krevelen). The theta-solvent composition of the PES solution was 0.3 v/v% water and 0.7 v/v% DMSO in this binary mixture. These measurements were also allowed to determine important parameters such as the coefficients A (=0.67) and B (=3.69 × 104) from the Schulz equation, or the Kη (=8.22 × 10-2) and α (=0.52) constants from the Kuhn-Mark-Houwink equation. Hopefully, the prepared PES with different molecular weights is a promising candidate for biomedical applications and the reported data and constants are useful for other researchers who work with this promising polyester.

Biofilm eradication and antifungal mechanism of action against Candida albicans of cationic dicephalic surfactants with a labile linker.

Our research aims to expand the knowledge on relationships between the structure of cationic dicephalic surfactants-N,N-bis[3,3_-(dimethylamine)propyl]alkylamide dihydrochlorides and N,N-bis[3,3_-(trimethylammonio)propyl]alkylamide dibromides (alkyl: n-C9H19, n-C11H23, n-C13H27, n-C15H31)-and their antifungal mechanism of action on Candida albicans. The mentioned groups of amphiphilic substances are characterized by the presence of a weak, hydrochloride cationic center readily undergoing deprotonation, as well as a stable, strong quaternary ammonium group and alkyl chains capable of strong interactions with fungal cells. Strong fungicidal properties and the role in creation and eradication of biofilm of those compounds were discussed in our earlier works, yet their mechanism of action remained unclear. It was shown that investigated surfactants induce strong oxidative stress and cause increase in cell membrane permeability without compromising its continuity, as indicated by increased potassium ion (K+) leakage. Thus experiments carried out on the investigated opportunistic pathogen indicate that the mechanism of action of the researched surfactants is different than in the case of the majority of known surfactants. Results presented in this paper significantly broaden the understanding on multifunctional cationic surfactants and their mechanism of action, as well as suggest their possible future applications as surface coating antiadhesives, fungicides and antibiofilm agents in medicine or industry.

Studying the "Rigid-Flexible" Properties of Polymeric Micelle Core-Forming Segments with a Hydrophobic Phthalocyanine Probe Using NMR and UV Spectroscopy.

The aim of the performed studies was to thoroughly examine the internal structure of self-assembled nanocarriers (i.e., polymeric micelles-PMs) by means of a hydrophobic phthalocyanine probe in order to identify the crucial features that are required to enhance the photoactive probe stability and reactivity. PMs of hydrophilic poly(ethylene glycol) and hydrophobic poly(ε-caprolactone) (PCL) or poly(d,l-lactide) (PDLLA) were fabricated and loaded with tetra tert-butyl zinc(II) phthalocyanine (ZnPc-t-but4), a multifunctional spectroscopic probe with a profound ability to generate singlet oxygen upon irradiation. The presence of subdomains, comprising "rigid" and "flexible" regions, in the studied block copolymers' micelles as well as their interactions with the probe molecules, were assessed by various high-resolution NMR measurements [e.g., through-space magnetic interactions by the 1D NOE effect, pulsed field gradient spin-echo, and spin-lattice relaxation time (T1) techniques]. The studies of the impact of the core-type microenvironment on the ZnPc-t-but4 photochemical performance also included photobleaching and reactive oxygen species measurements. ZnPc-t-but4 molecules were found to exhibit spatial proximity effects with both (PCL and PDLLA) hydrophobic polymer chains and interact with both subdomains, which are characterized by different rigidities. It was deduced that the interfaces between particular subdomains constitute an optimal host space for probe molecules, especially in the context of photochemical stability, photoactivity (i.e., for significant enhancement of singlet oxygen generation rates), and aggregation prevention. The present contribution proves that the combination of an appropriate probe, high-resolution NMR techniques, and UV-vis spectroscopy enables one to gain complex information about the subtle structure of PMs essential for their application as nanocarriers for photoactive compounds, for example, in photodynamic therapy, nanotheranostics, combination therapy, or photocatalysis, where the micelles constitute the optimal microenvironment for the desired photoreactions.

Benefits of pH-responsive polyelectrolyte coatings for carboxymethyl cellulose-based microparticles in the controlled release of esculin.

Moderate and prolonged payload release in response to a particular factor is highly demanded for efficient carriers of low-molecular-weight, chemically unstable phytopharmaceuticals. Thus, the objective of our contribution was to establish the effect of pH-responsive polyelectrolyte coatings on the release properties of carboxymethyl cellulose-based microparticles designed to deliver phytopharmaceuticals through the gastrointestinal tract. Microparticles were fabricated via extrusion coupled with external gelation and further coated with polyelectrolytes (PEs) (chitosan, gelatin, or PAH and PSS) involving electrostatic interactions. Successful deposition of PEs was confirmed by FTIR, and their thickness and viscosity were characterized in terms of QCM-D and ellipsometric techniques. The encapsulation efficiency of esculin, used as a model phytopharmaceutical, as proven by UV-Vis studies, was over 57%. SEM and fluorescence microscopy revealed a micrometric size, a mostly spherical shape and an altered topography of the investigated microcapsules. The physical stability of the microcapsules in media of various pH values was confirmed with CLSM and gravimetric studies. Studies on human gingival fibroblasts in vitro revealed that the obtained microparticles did not induce any cytotoxic effects. Payload release was monitored in situ by means of CLSM and ex situ under gastrointestinal conditions in vitro. Mathematical evaluation of the microparticle release profiles using classical models led to the establishment of a new hybrid model that revealed the mechanism behind esculin release. We demonstrated that the application of a polyelectrolyte shell onto CMC-based microspheres may provide controlled delivery of the payload, with its release triggered by the pH and ionic strength of the medium. These observations suggest that the release manner of small-molecule glycosides under gastrointestinal conditions can be tailored by careful selection of suitable materials to obtain biocompatible and functional hydrogel microparticles.

Co-delivery of IR-768 and daunorubicin using mPEG-b-PLGA micelles for synergistic enhancement of combination therapy of melanoma.

Malignant melanoma is an emerging problem worldwide due to the high degree of lethalness. Its aggressiveness and the ability to metastasize along with the heterogeneity at the molecular and cellular levels, limit the overall therapeutic efficacy. Despite significant advances in melanoma treatment over the last decade, there is still a need for improved therapeutic modalities. Thus, we demonstrate here a combinatorial approach that targets multiple independent therapeutic pathways, in which polymeric micelles (PMs) were used as efficacious colloidal nanocarriers loaded with both daunorubicin (DRB) as a cytotoxic drug and IR-768 as a photosensitizer. This afforded the dual drug loaded delivery system IR-768 + DRB in PMs. The fabricated mPEG-b-PLGA micelles (hydrodynamic diameters ≈ 25 nm) had a relatively narrow size distribution (PdI > ca. 0.3) with uniform spherical shapes. CLSM study showed that mPEG-b-PLGA micelles were uptaken by mitochondria, which further contributed to excellent singlet oxygen generation capacity for PDT in A375 melanoma cells. Furthermore, the PMs were efficiently internalized by tested cells through endocytosis, resulting in much higher cellular uptake comparing to the free drug. As a result of these properties, IR-768 + DRB in PMs exhibited very potent and synergistically enhanced anticancer activity against A375 cells. Additionally, this combination approach allowed to reduce drug doses and provided low side effects towards normal HaCaT. This study indicates excellent properties of mPEG-b-PLGA micelles resulting in great therapeutic potential possessed by the developed nanoscale drug delivery system for combined chemo-photodynamic therapy of melanoma.

Hydrophobically Functionalized Poly(Acrylic Acid) Comprising the Ester-Type Labile Spacer: Synthesis and Self-Organization in Water.

One of the most important properties of hydrophobically functionalized polyelectrolytes (HF-PEs) and their assemblies is their ability to encapsulate hydrophobic/amphiphilic agents and provide release on demand of the entrapped payload. The aim of the present work was to synthesize and study self-organization behavior in aqueous solution of hydrophobically functionalized poly(acrylic acid) (PAA) comprising the ester-type pH labile moiety with various degrees of hydrophobization and side-chain lengths in the absence and presence of appropriate mono- and polyvalent electrolytes (i.e., NaCl or CaCl2). The synthesis and purification of hydrophobically functionalized PAA were performed under mild conditions in order to avoid chemical degradation of the polymers. The modified polyelectrolytes self-assembly in aqueous systems was monitored using diffusion-ordered nuclear magnetic resonance (DOSY NMR). The performed studies, supported by the all-atoms molecular dynamics simulations, revealed a strong dependence of polyelectrolyte self-assembled state on concentration-specific concentration regions with the coexistence of both smaller and larger aggregates were observed (values of hydrodynamic diameter DH around one nanometer and between two to six nanometers, respectively). Our investigations enabled us to gain crucial information about the self-assembly of the hydrophobically functionalized poly(acrylic acid) and opened the possibility of understanding and predicting its performance under various conditions.

New mild amphoteric sulfohydroxybetaine-type surfactants containing different labile spacers: Synthesis, surface properties and performance.

HYPOTHESIS: The excellent performance of zwitterionic alkylamide-type surfactants may be tuned by slight modifications of their structures, especially concerning linking groups between hydrophilic and hydrophobic moieties and the moderation of strong NH hydrogen bonds via the introduction of a methyl group attached to the nitrogen. EXPERIMENTS: The influence of the structure of alkylamidehydroxysulfobetaine-type surfactants on their adsorption and micellization, lime soap dispersing ability, wettability, as well as antimicrobial and hemolytic activities was studied in this work. We synthesized a series of novel surfactants with labile CON(Me)CH2 or CH2N(Me) CO spacers that were adjacent to the hydrophobic tail CnH2n+1 and separated from the zwitterionic hydroxysulfobetaine headgroup by C3H6 or C2H4 linkers, i.e., [(3-alkanoyilomethyoamine)propyl] dimethylammonium 2-hydroxypropanesulfonates (CnTMDAS) and [3-(alkylmetyloamino)-3-oxopropyl] dimethylammonium 2-hydroxypropanesulfonates (CnDMPAS), respectively. FINDINGS: The CnTMDAS surfactants showed slightly higher surface activity and lower CMC than the respective CnDMPAS with the same number of carbon atoms. Moreover, the CnTMDAS series exhibited significantly higher lime soap dispersing abilities and softer interactions with biological systems than CnDMPAS. The molecular modeling computations revealed that the difference in surface activity originated from lower dipole moment of CnTMDAS, hence, lower polarity, whereas higher free energy of dimerization of those surfactants with oleic acid may account for the favorable formation of mixed micelles.

Folate-directed zinc (II) phthalocyanine loaded polymeric micelles engineered to generate reactive oxygen species for efficacious photodynamic therapy of cancer.

Targeted and effective drug transport is becoming an attractive option in cancer therapy since it can improve drug efficacy and reduce drugs' side effects in normal tissues. In addition to using specific surface ligand molecules, the selective drug delivery can be accomplished via enhanced permeability and retention effect. Therefore, in our studies, we entrapped zinc (II) phthalocyanine (ZnPc) - a second generation photosensitizer - in folate-functionalized micelles of the biocompatible, FDA-approved for biomedical application diblock copolymer methoxypoly(ethylene oxide)-b-poly(L-lactide) (mPEG-b-PLLA) and its derivative with folate (FA) attached to the end of PEG chain (FA-PEG-b-PLLA). Dynamic light scattering (DLS) measurements confirmed the micellar size to be <150 nm in diameter, a low polydispersity index, and good colloid stability of the studied nanocarriers, while atomic force microscopy (AFM) was used to study their morphology. The application potential of the resulting micelles was evaluated in cyto- and photocytotoxicity studies in conjunction with intracellular distribution and accumulation imaging of the photosensitizer delivered to ovarian carcinoma (SKOV-3) and metastatic melanoma (Me45) cell lines. Reactive oxygen species generation study was performed after photodynamic reaction, and cellular cytoskeleton reorganization was visualized after undergoing a photodynamic reaction. The results demonstrated that the functionalized polymeric micelles are promising nanocarriers for photodynamic therapy procedures and can be used in anticancer drug delivery.

Förster Resonance Energy Transfer-Activated Processes in Smart Nanotheranostics Fabricated in a Sustainable Manner.

Multilayer nanocarriers loaded with optically activated payloads are gaining increasing attention due to their anticipated crucial role for providing new mechanisms of energy transfers in the health-oriented applications, as well as for energy storage and environmental protection. The combination of careful selection of optical components for efficient Förster resonance energy transfer, and surface engineering of the nanocarriers, allowed us to synthesize and characterize novel theranostic nanosystems for diagnosis and therapy of deep-seated tumors. The cargo, constrained within the oil core of the nanocapsules, composed of NaYF4 :Tm+3 , Yb+3 up-converting nanoparticles together with a second-generation porphyrin-based photosensitizing agent-Verteporfin, assured requisite diagnostic and therapeutic functions under near-IR laser excitation. The outer polyaminoacid shell of the nanocapsules was functionalized with a ligand-poly(l-glutamic acid) functionalized by PEG-ylated folic acid-to ensure both a "stealth" effect and active targeting towards human breast cancer cells. The preparation criteria of all nanocarrier building blocks meet the requirements for sustainable and green chemistry practices. The multifunctionality of the proposed nanocarriers is a consequence of both the surface-functionalized organic exterior part, which was accessible for selective accumulation in cancer cells, and the hydrophobic optically active interior, which shows phototoxicity upon irradiation within the first biological window.

Recent progress in the engineering of multifunctional colloidal nanoparticles for enhanced photodynamic therapy and bioimaging.

This up-to-date review summarizes the design and current fabrication strategies that have been employed in the area of mono- and multifunctional colloidal nanoparticles - nanocarriers well suited for photodynamic therapy (PDT) and diagnostic purposes. Rationally engineered photosensitizer (PS)-loaded nanoparticles may be achieved via either noncovalent (i.e., self-aggregation, interfacial deposition, interfacial polymerization, or core-shell entrapment along with physical adsorption) or covalent (chemical immobilization or conjugation) processes. These PS loading approaches should provide chemical and physical stability to PS payloads. Their hydrophilic surfaces, capable of appreciable surface interactions with biological systems, can be further modified using functional groups (stealth effect) to achieve prolonged circulation in the body after administration and/or grafted by targeting agents (such as ligands, which bind to specific receptors uniquely expressed on the cell surface) or stimuli (e.g., pH, temperature, and light)-responsive moieties to improve their action and targeting efficiency. These attempts may in principle permit efficacious PDT, combination therapies, molecular diagnosis, and - in the case of nanotheranostics - simultaneous monitoring and treatment. Nanophotosensitizers (nano-PSs) should possess appropriate morphologies, sizes, unimodal distributions and surface processes to be successfully delivered to the place of action after systemic administration and should be accumulated in certain tumors by passive and/or active targeting. Additionally, physically facilitating drug delivery systems emerge as a promising approach to enhancing drug delivery, especially for the non-invasive treatment of deep-seated malignant tissues. Recent advances in nano-PSs are scrutinized, with an emphasis on design principles, via the promising use of colloid chemistry and nanotechnology.

Lab-on-a-chip systems for photodynamic therapy investigations.

In recent years photodynamic therapy (PDT) has received widespread attention in cancer treatment due to its smaller surgical trauma, better selectivity towards tumor cells, reduced side effects and possibility of repeatable treatment. Since cancer is the second cause of death worldwide, scientists constantly seek for new potential therapeutic agents including nanotechnology-based photosensitizers used in PDT. The new-designed nanostructures must be carefully studied and well characterized what require analytically useful and powerful tools that enable real progress in nanoscience development. This review describes the current status of PDT investigations using microfluidic Lab-on-a-Chip systems, including recent developments of nanoparticle-based PDT agents, their combinations with different drugs, designs and examples of in vitro applications. This review mainly lays emphasis on biological evaluation of FDA approved photosensitizing agents as well as newly designed nanophotosensitizers. It also highlights the analytical performances of various microfluidic Lab-on-a-chip systems for PDT efficacy analysis on 3D culture and discusses microsystems designs in detail.