Lipid-matrix effects on tyrosinase immobilization in Langmuir and Langmuir-Blodgett films.

The immobilization of the enzyme tyrosinase (Tyr) in lipid matrices can be explored to produce biosensors for detecting polyphenols, which is relevant for the food industry. Herein, we shall demonstrate the importance of the lipid composition to immobilize the enzyme tyrosinase in Langmuir-Blodgett (LB) films. Tyr could be incorporated into Langmuir monolayers of arachidic acid (AA), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1,2-dipalmitoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (sodium salt) (DPPG), having as the main effect an expansion in the monolayers. Results from polarization-modulated infrared reflection-absorption spectroscopy (PM-IRRAS) pointed to electrostatic interactions between the charged residues of Try and the lipid headgroups, in addition to changes in the order of lipid chains. The interaction between Tyr and DPPC in Langmuir monolayers can be correlated with the superior performance of DPPC/Tyr LB films used as biosensors to detect catechol by cyclic voltammetry. The molecular-level interactions assessed via PM-IRRAS are therefore believed to drive an immobilization process for Tyr in the lipid LB matrix and may serve as a general criterion to identify matrices that preserve enzyme activity.

Chain Cleavage of Bioinspired Bacterial Membranes Photoinduced by Eosin Decyl Ester.

Photodynamic therapy (PDT) is promising for bacterial inactivation since cellular internalization of photosensitizers (PS) is not crucial for the treatment effectiveness. Photoinduced damage in the lipid envelope may already induce microbial inactivation, which requires PS capable of easily penetrating into the membrane. Herein, we report on the insertion of the PS eosin decyl ester (EosDec) into Langmuir films of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), 1,2-dioleoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (DOPG), and cardiolipin (CLP) used as mimetic systems of bacterial membranes. Surface pressure isotherms and polarization-modulated infrared reflection absorption spectroscopy (PM-IRRAS) indicated that the hydrophobic nature of EosDec favored deeper penetration in all the phospholipid monolayers. The incorporation of EosDec led to monolayer expansion, especially in the anionic DOPG and CLP owing to repulsive electrostatic interactions, and induced disorder in the lipid chains. Light irradiation of DOPE, DOPG, and CLP monolayers containing EosDec increased the rate of material loss to the subphase, which is attributed to cleavage of lipid chains triggered by contact-dependent reactions between excited states of EosDec and lipid unsaturations. The latter is key for membrane permeabilization and efficiency in microbial inactivation.

Role of Toluidine Blue-O Binding Mechanism for Photooxidation in Bioinspired Bacterial Membranes.

The alarming increase in bacterial resistance to antibiotics has demanded new strategies for microbial inactivation, which include photodynamic therapy whose activity relies on the photoreaction damage to the microorganism membrane. Herein, the binding mechanisms of the photosensitizer toluidine blue-O (TBO) on simplified models of bacterial membrane with Langmuir monolayers of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) and 1,2-dioleoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (DOPG) were correlated to the effects of the photoinduced lipid oxidation. Langmuir monolayers of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) were also used as a reference of mammalian membranes. The surface pressure isotherms combined with polarization-modulated infrared reflection absorption spectroscopy revealed that TBO expands DOPC, DOPE, and DOPG monolayers owing to electrostatic interactions with the negatively charged groups in the phospholipids, with a stronger adsorption on DOPG, which has a net surface charge. Light irradiation made the TBO-containing DOPC and DOPE monolayers less unstable as a result of the singlet oxygen (1O2) reaction with the chain unsaturation and hydroperoxide formation. In contrast, the decreased stability of the irradiated TBO-containing DOPG monolayer suggests the cleavage of carbon chains. The anionic nature of DOPG allowed a deeper penetration of TBO into the chain region, favoring contact-dependent reactions between the excited triplet state of TBO and lipid unsaturations or/and hydroperoxide groups, which is the key for the cleavage reactions and further membrane permeabilization.

Breast cancer subtype specific biochemical responses to radiation.

External beam radiotherapy is a common form of treatment for breast cancer. Among patients and across different breast cancer subtypes, the response to radiation is heterogeneous. Radiation-induced biochemical changes were examined by Raman spectroscopy using cell lines that represent a spectrum of human breast cancer. Principal component analysis (PCA) and partial least squares discriminant analysis (PLSDA) revealed unique Raman spectral features in the HER2 and Ki67 subtype. The changes in Raman spectral profiles to different doses of radiation (0-50 Gy) included variations in the levels of proteins, lipids, nucleic acids and glycogen. Importantly, the differences in radiation-induced changes on the normal breast epithelial cell line MCF10A could be discriminated within and across the various breast tumor cell lines. These results demonstrate a novel approach to uncover differences between breast cancer cell subtypes and surrounding normal tissues by their biochemical variations in response to radiation.

Molecular-Level Modifications Induced by Photo-Oxidation of Lipid Monolayers Interacting with Erythrosin.

Incorporation into cell membranes is key for the action of photosensitizers in photomedicine treatments, with hydroperoxidation as the prominent pathway of lipid oxidation. In this paper, we use Langmuir monolayers of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) as cell membrane models to investigate adsorption of the photosensitizer erythrosin and its effect on photoinduced lipid oxidation. From surface pressure isotherms and polarization-modulated infrared reflection-absorption spectroscopy (PM-IRRAS) data, erythrosin was found to adsorb mainly via electrostatic interaction with the choline in the head groups of both DOPC and DPPC. It caused larger monolayer expansion in DOPC, with possible penetration into the hydrophobic unsaturated chains, while penetration into the DPPC saturated chains was insignificant. Easier penetration is due to the less packed DOPC monolayer, in comparison to the more compact DPPC according to the monolayer compressibility data. Most importantly, light irradiation at 530 nm made the erythrosin-containing DOPC monolayer become less unstable, with a relative surface area increase of ca. 19%, in agreement with previous findings for bioadhesive giant vesicles. The relative area increase is consistent with hydroperoxidation, supporting the erythrosin penetration into the lipid chains, which favors singlet oxygen generation close to double bonds, an important requirement for photodynamic efficiency.

Molecularly designed layer-by-layer (LbL) films to detect catechol using information visualization methods.

The control of molecular architectures has been exploited in layer-by-layer (LbL) films deposited on Au interdigitated electrodes, thus forming an electronic tongue (e-tongue) system that reached an unprecedented high sensitivity (down to 10(-12) M) in detecting catechol. Such high sensitivity was made possible upon using units containing the enzyme tyrosinase, which interacted specifically with catechol, and by processing impedance spectroscopy data with information visualization methods. These latter methods, including the parallel coordinates technique, were also useful for identifying the major contributors to the high distinguishing ability toward catechol. Among several film architectures tested, the most efficient had a tyrosinase layer deposited atop LbL films of alternating layers of dioctadecyldimethylammonium bromide (DODAB) and 1,2-dipalmitoyl-sn-3-glycero-fosfo-rac-(1-glycerol) (DPPG), viz., (DODAB/DPPG)5/DODAB/Tyr. The latter represents a more suitable medium for immobilizing tyrosinase when compared to conventional polyelectrolytes. Furthermore, the distinction was more effective at low frequencies where double-layer effects on the film/liquid sample dominate the electrical response. Because the optimization of film architectures based on information visualization is completely generic, the approach presented here may be extended to designing architectures for other types of applications in addition to sensing and biosensing.

Photochemical outcomes triggered by gold shell-isolated nanorods on bioinspired nanoarchitectonics for bacterial membranes.

Boosted by the indiscriminate use of antibiotics, multidrug-resistance (MDR) demands new strategies to combat bacterial infections, such as photothermal therapy (PTT) based on plasmonic nanostructures. PTT efficiency relies on photoinduced damage caused to the bacterial machinery, for which nanostructure incorporation into the cell envelope is key. Herein, we shall unveil the binding and photochemical mechanisms of gold shell-isolated nanorods (AuSHINRs) on bioinspired bacterial membranes assembled as Langmuir and Langmuir-Schaefer (LS) monolayers of DOPE, Lysyl-PG, DOPG and CL. AuSHINRs incorporation expanded the isotherms, with stronger effect on the anionic DOPG and CL. Indeed, FTIR of LS films revealed more modifications for DOPG and CL owing to stronger attractive electrostatic interactions between anionic phosphates and the positively charged AuSHINRs, while electrostatic repulsions with the cationic ethanolamine (DOPE) and lysyl (Lysyl-PG) polar groups might have weakened their interactions with AuSHINRs. No statistical difference was observed in the surface area of irradiated DOPE and Lysyl-PG monolayers on AuSHINRs, which is evidence of the restricted nanostructures insertion. In contrast, irradiated DOPG monolayer on AuSHINRs decreased 4.0 % in surface area, while irradiated CL monolayer increased 3.7 %. Such results agree with oxidative reactions prompted by ROS generated by AuSHINRs photoactivation. The deepest AuSHINRs insertion into DOPG may have favored chain cleavage while hydroperoxidation is the mostly like outcome in CL, where AuSHINRs are surrounding the polar groups. Furthermore, preliminary experiments on Escherichia coli culture demonstrated that the electrostatic interactions with AuSHINRs do not inhibit bacterial growth, but the photoinduced effects are highly toxic, resulting in microbial inactivation.

Toward the optimization of an e-tongue system using information visualization: a case study with perylene tetracarboxylic derivative films in the sensing units.

The wide variety of molecular architectures used in sensors and biosensors and the large amount of data generated with some principles of detection have motivated the use of computational methods, such as information visualization techniques, not only to handle the data but also to optimize sensing performance. In this study, we combine projection techniques with micro-Raman scattering and atomic force microscopy (AFM) to address critical issues related to practical applications of electronic tongues (e-tongues) based on impedance spectroscopy. Experimentally, we used sensing units made with thin films of a perylene derivative (AzoPTCD acronym), coating Pt interdigitated electrodes, to detect CuCl(2) (Cu(2+)), methylene blue (MB), and saccharose in aqueous solutions, which were selected due to their distinct molecular sizes and ionic character in solution. The AzoPTCD films were deposited from monolayers to 120 nm via Langmuir-Blodgett (LB) and physical vapor deposition (PVD) techniques. Because the main aspects investigated were how the interdigitated electrodes are coated by thin films (architecture on e-tongue) and the film thickness, we decided to employ the same material for all sensing units. The capacitance data were projected into a 2D plot using the force scheme method, from which we could infer that at low analyte concentrations the electrical response of the units was determined by the film thickness. Concentrations at 10 µM or higher could be distinguished with thinner films--tens of nanometers at most--which could withstand the impedance measurements, and without causing significant changes in the Raman signal for the AzoPTCD film-forming molecules. The sensitivity to the analytes appears to be related to adsorption on the film surface, as inferred from Raman spectroscopy data using MB as analyte and from the multidimensional projections. The analysis of the results presented may serve as a new route to select materials and molecular architectures for novel sensors and biosensors, in addition to suggesting ways to unravel the mechanisms behind the high sensitivity obtained in various sensors.

Mechanisms of hypericin incorporation to explain the photooxidation outcomes in phospholipid biomembrane models.

Cell membranes are the first barriers for drug binding and key for the action of photosensitizers (PS). Herein, we report on the incorporation of the PS hypericin into Langmuir monolayers of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and 1,2-dioleoyl-sn-glycero-3-phospho-L-serine (DOPS) to represent eukaryotic cell membranes, and 1,2-dioleoyl-sn-glycero-3-phospho(1'-rac-glycerol) (DOPG) to mimic bacterial membranes. Surface pressure (π) vs mean molecular area (Å) isotherms showed a high degree of interaction (binding, penetration and relative solubilization) of hypericin into DPPC and DOPC monolayers. On the other hand, electrostatic repulsions govern the interactions with DOPG and DOPS, favoring hypericin self-aggregation, as visualized by Brewster angle microscopy (BAM). Indeed, the larger domains in BAM were consistent with the greater expansion of DOPG monolayers with incorporated hypericin, owing to stronger electrostatic repulsions. In contrast to DPPC, light-irradiation of DOPC monolayers containing hypericin induced loss of material due to hydrocarbon chain cleavage triggered by contact-dependent reactions between triplet excited state of hypericin and chain unsaturations. The mild effects noted for both irradiated DOPS and DOPG monolayers are attributed to hypericin self-aggregation, which may have decreased the singlet oxygen quantum yield (Φ1O2) via self-quenching, despite the increased instability induced in the monolayers.

Immunoassay platform with surface-enhanced resonance Raman scattering for detecting trace levels of SARS-CoV-2 spike protein.

The early diagnosis of Coronavirus disease (COVID-19) requires either an accurate detection of genetic material or a sensitive detection of viral proteins. In this work, we designed an immunoassay platform for detecting trace levels of SARS-CoV-2 spike (S) protein. It is based on surface-enhanced resonance Raman scattering (SERRS) of methylene blue (MB) adsorbed onto spherical gold nanoparticles (AuNPs) and coated with a 6 nm silica shell. The latter shell in the SERRS nanoprobe prevented aggregation and permitted functionalization with SARS-CoV-2 antibodies. Specificity of the immunoassay was achieved by combining this functionalization with antibody immobilization on the cover slides that served as the platform support. Different concentrations of SARS-CoV-2 antigen could be distinguished and the lack of influence of interferents was confirmed by treating SERRS data with the multidimensional projection technique Sammon's mapping. With SERRS using a laser line at 633 nm, the lowest concentration of spike protein detected was 10 pg/mL, achieving a limit of detection (LOD) of 0.046 ng/mL (0.60 pM). This value is comparable to the lowest concentrations in the plasma of COVID-19 patients at the onset of symptoms, thus indicating that the SERRS immunoassay platform may be employed for early diagnosis.

The efficiency of photothermal action of gold shell-isolated nanoparticles against tumor cells depends on membrane interactions.

Photoinduced hyperthermia with nanomaterials has been proven effective in photothermal therapy (PTT) of tumor tissues, but a precise control in PTT requires determination of the molecular-level mechanisms. In this paper, we determined the mechanisms responsible for the action of photoexcited gold shell-isolated nanoparticles (AuSHINs) in reducing the viability of MCF7 (glandular breast cancer) and especially A549 (lung adenocarcinoma) cells in vitro experiments, while the photoinduced damage to healthy cells was much smaller. The photoinduced effects were more significant than using other nanomaterials, and could be explained by the different effects from incorporating AuSHINs on Langmuir monolayers from lipid extracts of tumoral (MCF7 and A549) and healthy cells. The incorporation of AuSHINs caused similar expansion of the Langmuir monolayers, but Fourier-transform infrared spectroscopy (FTIR) data of Langmuir-Schaefer films (LS) indicated distinct levels of penetration into the monolayers. AuSHINs penetrated deeper into the A549 extract monolayers, affecting the vibrational modes of polar groups and carbon chains, while in MCF7 monolayers penetration was limited to the surroundings of the polar groups. Even smaller insertion was observed for monolayers of the healthy cell extract. The photochemical reactions were modulated by AuSHINs penetration, since upon irradiation the surface area of A549 monolayer decreased owing to lipid chain cleavage by oxidative reactions. For MCF7 monolayers, hydroperoxidation under illumination led to a ca. 5% increase in surface area. The monolayers of healthy cell lipid extract were barely affected by irradiation, consistent with the lowest degree of AuSHINs insertion. In summary, efficient photothermal therapy may be devised by producing AuSHINs capable of penetrating the chain region of tumor cell membranes.

Modulating photochemical reactions in Langmuir monolayers of Escherichia coli lipid extract with the binding mechanisms of eosin decyl ester and toluidine blue-O photosensitizers.

Photodynamic damage to the cell envelope can inactivate microorganisms and may be applied to combat super-resistance phenomenon, empowered by the indiscriminate use of antibiotics. Efficiency in microbial inactivation is dependent on the incorporation of photosensitizers (PS) into the bacterial membranes to trigger oxidation reactions under illumination. Herein, Langmuir monolayers of Escherichia coli lipid extract were built to determine the binding mechanisms and oxidation outcomes induced by eosin decyl ester (EosDEC) and toluidine blue-O (TBO) PSs. Surface-pressure isotherms of the E. coli monolayers were expanded upon EosDEC and TBO, suggesting incorporation of both PSs. Fourier-transform infrared spectroscopy (FTIR) of Langmuir-Schaefer (LS) films reveled that the EosDEC and TBO binding mechanisms are dominated by electrostatic interactions with the anionic polar groups, with limited penetration into the chains. Light-irradiation reduced the relative area of E. coli monolayer on TBO, indicating an increased loss of material to the subphase owing to the chain cleavage, generated by contact-dependent reactions with excited states of TBO. In contrast, the increased relative area of E. coli monolayers containing EosDEC suggests lipid hydroperoxidation, which is PS contact-independent. Even considering a small chain penetration, the saturated EosDEC may have partitioned towards saturated reach domains, avoiding direct contact with membrane unsaturations.

Plasma membrane permeabilization to explain erythrosine B phototoxicity on in vitro breast cancer cell models.

Lipid oxidation is ubiquitous in cell life under oxygen and essential for photodynamic therapy (PDT) of carcinomas. However, the mechanisms underlying lipid oxidation in rather complex systems such as plasma membranes remain elusive. Herein, Langmuir monolayers were assembled with the lipid extract of glandular breast cancer (MCF7) cells and used to probe the molecular interactions allowing adsorption of the photosensitizer (PS) erythrosine B and subsequent photooxidation outcomes. Surface pressure (π) versus area (cm2/mL) isotherms of MCF7 lipid extract shifted to larger areas upon erythrosine incorporation, driven by secondary interactions that affected the orientation of the carbonyl groups and lipid chain organization. Light-irradiation increased the surface area of the MCF7 lipid extract monolayer containing erythrosine owing to the lipid hydroperoxidation, which may further undergo decomposition, resulting in the chain cleavage of phospholipids and membrane permeabilization. Incorporation of erythrosine by MCF7 cells induced slight toxic effects on in vitro assays, differently of the severe phototoxicity caused by light-irradiation, which significantly decreased cell viability by more than 75% at 2.5 × 10-6 mol/L of erythrosine incubated for 3 and 24 h, reaching nearly 90% at 48 h of incubation. The origin of the phototoxic effects is in the rupture of the plasma membrane shown by the frontal (FSC) and side (SSC) light scattering of flow cytometry. Consistent with hydroperoxide decomposition, membrane permeabilization was also confirmed by cleaved lipids detected in mass spectrometry and subsidizes the necrotic pathway of cell death.

Coupling surface-enhanced resonance Raman scattering and electronic tongue as characterization tools to investigate biological membrane mimetic systems.

The surface-enhanced Raman scattering (SERS) effect and sensor and biosensor analyses are widely applied to investigate drug-biomolecule interactions or to detect trace amount of analytes. In this work, surface-enhanced resonance Raman scattering (SERRS) and an electronic tongue system using impedance spectroscopy were brought together, combining sensitivity and structural level information. Taking advantage of the use of layer-by-layer (LbL) films of phospholipids as biological membrane mimetic systems, cardiolipin (CLP) and dipalmitoyl phosphatidyl glycerol (DPPG) were applied as transducers onto Pt interdigitated electrodes forming an array of sensing units. This e-tongue system was able to detect the phenothiazine methylene blue (MB) below nanomolar concentrations. SERRS was applied to investigate the MB molecular arrangement (monomers or aggregates) when in contact with the phospholipids at trace levels of concentration. The key point was the adsorption of Ag nanoparticles (AgNPs) within the phospholipid LbL films. This approach did not compromise the e-tongue performance and allowed the recording of in situ SERRS spectra for the LbL films after immersion into MB aqueous solutions. The detection of MB through SERRS gave similar results to those reported in the literature but now with an unprecedented sensitivity.

Photo-Induced Necrosis on Oropharyngeal Carcinoma (HEp-2) Cells Mediated by the Xanthene Erythrosine.

The photodynamic therapy (PDT) has been outstanding as a promising alternative for treating different carcinomas. However, the lack of detailed knowledge on the mechanisms of action prevents exploitation of the therapy full potential. Herein we shall evaluate not only the photodynamic efficiency but the mechanism of cell death triggered by the photoactivated erythrosine in oropharyngeal cancer cells (HEp-2). Cytotoxic assays were performed by MTT at distinct concentrations (10-3 to 10-6 mol/L) and incubation time (3, 24 and 48 h) of erythrosine in HEp-2 in vitro culture. In addition to the cytotoxic effect, the mechanisms of cell death were evaluated by flow cytometry following the annexin V/propidium iodide double staining protocol. Erythrosine was incorporated by HEp-2 cells in a dose- and time-dependent pathway. The incubation of erythrosine in dark has not shown any significant effect over the culture until 24 h and 1.25×10-6 mol/L concentration, from which a small portion (<25% and statistically significant) of the cell population have undergone apoptosis. On the other hand, 50% of cell viability is reduced mainly by necrosis when 10, 3.75 and 1.9×10-6 mol/L of erythrosine concentrations at 3, 24 and 48 h of incubation are photoactivated, respectively. Bioinspired models of tumor membrane based on Langmuir monolayers of 2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and 1,2-dioleoyl-sn-glycero-3-phospho-L-serine (DOPS) mixture reveled that electrostatic interactions with the lipid head groups are the main driving forces allowing the erythrosine adsorption. Furthermore, light-induced hydroperoxidation significantly increased the surface area of the monolayers, which might be the origin of the necrotic pathway triggered in HEp-2 cells.

Correlating Artepillin C cytotoxic activity on HEp-2 cells with bioinspired systems of plasma membranes.

Artepillin C is the main compound present in propolis from Baccharis dracunculifolia, whose antitumor activity has been the focus of many studies. Herein, we shall investigate the Artepillin C mechanisms of action against cells derived from the oropharyngeal carcinoma (HEp-2). Cytotoxicity tests revealed that the concentrations of Artepillin C required to reduce cell viability by 50% (CC50) are dependent on the incubation time, decreasing from 40.7 × 10-5 mol/L to 15.7 × 10-5 mol/L and 9.05 × 10-5 mol/L considering 12, 24 and 48 h, respectively. Hydrophobic interactions on neutral species of Artepillin C induce aggregation over the HEp-2 plasma membrane, given the acid conditions of the cellular culture. Indeed, Langmuir monolayers mimicking cellular membranes of tumor cells revealed Artepillin C affinity to interact with 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) containing 20 mol% of 1,2-dipalmitoyl-sn-glychero-3-phosphoserine (DPPS), leading aggregation on giant unilamellar vesicles (GUVs) at pH 3.2. Moreover, leakage experiments on GUVs have shown that the presence of DPPS enhances the efflux of the fluorescent probe signaling the membrane permeabilization, which is the origin of the necrotic pathway triggered in HEp-2 cells, as observed by flow cytometry assays.

Molecular-level effects on cell membrane models to explain the phototoxicity of gold shell-isolated nanoparticles to cancer cells.

Metallic nanoparticles are promising agents for photothermal cancer therapy (PTT) owing to their photostability and efficient light-to-heat conversion, but their possible aggregation remains an issue. In this paper, we report on the photoinduced heating of gold shell-isolated nanoparticles (AuSHINs) in in vitro experiments to kill human oropharyngeal (HEp-2) and breast (BT-474 and MCF-7) carcinoma cells, with cell viability reducing below 50 % with 2.2 × 1012 AuSHINs/mL and 6 h of incubation. This toxicity to cancer cells is significantly higher than in previous works with gold nanoparticles. Considering the AuSHINs dimensions we hypothesize that cell uptake is not straightforward, and the mechanism of action involves accumulation on phospholipid membranes as the PTT target for photoinduced heating and subsequent generation of reactive oxygen species (ROS). Using Langmuir monolayers as simplified membrane models, we confirmed that AuSHINs have a larger effect on 1,2-dioleoyl-sn-glycero-3-phospho-l-serine (DOPS), believed to represent cancer cell membranes, than on 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) taken as representative of healthy eukaryotic cells. In particular, data from polarization-modulated infrared reflection absorption spectroscopy (PM-IRRAS) revealed an increased conformational order of DOPS tails due to the stronger adsorption of AuSHINs. Furthermore, light irradiation reduced the stability of AuSHINs containing DOPC and DOPS monolayers owing to oxidative reactions triggered by ROS upon photoinduced heating. Compared to DOPC, DOPS lost nearly twice as much material to the subphase, which is consistent with a higher rate of ROS formation in the vicinity of the DOPS monolayer.

Taking advantage of electrostatic interactions to grow Langmuir-Blodgett films containing multilayers of the phospholipid dipalmitoylphosphatidylglycerol.

The use of phospholipids as mimetic systems for studies involving the cell membrane is a well-known approach. In this context, the Langmuir and Langmuir-Blodgett (LB) methods are among the main techniques used to produce ordered layers of phospholipids structured as mono- or bilayers on water subphase and solid substrates. However, the difficulties of producing multilayer LB films of phospholipids restrict the application of this technique depending on the sensitivity of the experimental analysis to be conducted. Here, an alternative approach is used to produce LB films containing multilayers of the negative phospholipid dipalmitoylphosphatidylglycerol (DPPG). Inspired by the electrostatic layer-by-layer (LbL) technique, DPPG multilayer LB films were produced by transferring the DPPG Langmuir monolayers from the water subphase containing low concentrations of the cationic polyelectrolyte poly(allylamine hydrochloride) (PAH) onto solid substrates. Fourier transform infrared (FTIR) absorption spectroscopy revealed that the interactions between the NH(3)(+) (PAH) and PO(4)(-) (DPPG) groups might be the main driving forces that allow growth of these LB films. Besides, ultraviolet-visible (UV-vis) absorption spectroscopy showed that the multilayer LB films can be grown in a controlled way in terms of thickness at nanometer scale. Cyclic voltammetry showed that DPPG and PAH are more packed in the LB than LbL films. The latter finding is related to the distinct molecular architecture of the films since DPPG is structured as monolayers in the LB films and multilamellar vesicles in the LbL films. Despite the interaction with PAH, cyclic voltammetry also showed that DPPG retains its biological activity in LB films, which is a key factor since this makes DPPG a suitable material in sensing applications. Therefore, multilayer LB films were deposited onto Pt interdigitated electrodes forming sensing units, which were applied in the detection of a phenothiazine compound [methylene blue (MB)] using impedance spectroscopy. The performance of DPPG in single-layer and multilayer LB films was compared to the performance of sensing unities composed of DPPG in single-layer and multilayer LbL films, showing the importance of both the thickness and the molecular architecture of the thin films. As found in a previous work for LbL films, the high sensitivity reached by these sensing units is intimately related to changes in the morphology of the film as evidenced by the micro-Raman technique. Finally, the interaction between MB and the (DPPG+PAH) LB films was complemented by pi-A isotherms and surface-enhanced resonance Raman scattering (SERRS).

Evidence of photoinduced lipid hydroperoxidation in Langmuir monolayers containing Eosin Y.

Photodynamic therapy (PDT) efficiency depends on many factors including the incorporation of the photosensitizer (PS) in cell membranes and possible lipid hydroperoxidation. In this study, we show that hydroperoxidation may be photoinduced when eosin Y is incorporated into Langmuir monolayers that serve as cell membrane models. This occurs for Langmuir monolayers of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), which have unsaturation in their hydrophobic chains. In contrast, light irradiation had no effect on monolayers of saturated 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). Evidence of hydroperoxidation was obtained from the area increase in eosin-containing DOPC and POPC monolayers upon irradiation, which was accompanied by a decrease in monolayer thickness according to grazing incidence X-ray off-specular scattering (GIXOS) data. Furthermore, the changes in polarization-modulated infrared reflection absorption spectroscopy (PM-IRRAS) induced by irradiation were consistent with hydroperoxide migration toward the lipid hydrophilic heads.. In summary, this combination of experimental methods allowed us to determine the effects of eosin Y interaction with cell membrane models under irradiation, which may be associated with the underlying mechanisms of eosin Y as photosensitizer in PDT.

Co-Deposition of Gold Nanoparticles and Metalloporphyrin Using the Langmuir-Blodgett (LB) Technique for Surface-Enhanced Raman Scattering (SERS).

The synergistic effect produced by metallic nanoparticles when incorporated into different systems empowers a research field that is growing rapidly. In addition, organometallic materials are at the center of intensive research with diverse applications such as light-emitting devices, transistors, solar cells, and sensors. The Langmuir-Blodgett (LB) technique has proven to be suitable to address challenges inherent to organic devices, since the film properties can be tuned at the molecular level. Here we report a strategy to incorporate gold nanoparticles (AuNPs) into the LB film by co-deposition in order to achieve surface-enhanced Raman scattering (SERS) of the zinc(II)-protoporphyrin (IX) dimethyl ester (ZnPPIX-DME). Prior to the LB co-deposition, the properties of the Langmuir monolayer of ZnPPIX-DME at the air-water interface, containing AuNPs in the subphase, are studied through the surface-pressure versus mean molecular area (π-A) isotherms. The ZnPPIX-DME+AuNPs π-A isotherm presented a significant shift to higher molecular area, suggesting an interaction between both ZnPPIX-DME molecules and AuNPs. Those interactions are a key factor allowing the co-deposition of both AuNPs and ZnPPIX-DME molecules onto a solid substrate, thus forming the LB film. SERS of ZnPPIX-DME was successfully attained, ensuring the spatial distribution of the AuNPs. Higher enhancement factors were found at AuNP aggregates, as a result of the intense local electromagnetic field found in the metal nanoparticle aggregates. The main vibrational bands observed in the SERS spectra suggest a physical adsorption of the ZnPPIX-DME onto the surface of AuNPs. The latter is not only in agreement with the interactions pointed out by the π-A isotherms but also suggests that this interaction is kept upon LB film co-deposition.