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Encrypted peptides have been recently found in the human proteome and represent a potential class of antibiotics. Here we report three peptides derived from the human apolipoprotein B (residues 887-922) that exhibited potent antimicrobial activity against drug-resistant Klebsiella pneumoniae, Acinetobacter baumannii, and Staphylococci both in vitro and in an animal model. The peptides had excellent cytotoxicity profiles, targeted bacteria by depolarizing and permeabilizing their cytoplasmic membrane, inhibited biofilms, and displayed anti-inflammatory properties. Importantly, the peptides, when used in combination, potentiated the activity of conventional antibiotics against bacteria and did not select for bacterial resistance. To ensure translatability of these molecules, a protease resistant retro-inverso variant of the lead encrypted peptide was synthesized and demonstrated anti-infective activity in a preclinical mouse model. Our results provide a link between human plasma and innate immunity and point to the blood as a source of much-needed antimicrobials.

Therapeutic solutions to counter Burkholderia cepacia complex (Bcc) bacteria are challenging due to their intrinsically high level of antibiotic resistance. Bcc organisms display a variety of potential virulence factors, have a distinct lipopolysaccharide naturally implicated in antimicrobial resistance. and are able to form biofilms, which may further protect them from both host defence peptides (HDPs) and antibiotics. Here, we report the promising anti-biofilm and immunomodulatory activities of human HDP GVF27 on two of the most clinically relevant Bcc members, Burkholderia multivorans and Burkholderia cenocepacia. The effects of synthetic and labelled GVF27 were tested on B. cenocepacia and B. multivorans biofilms, at three different stages of formation, by confocal laser scanning microscopy (CLSM). Assays on bacterial cultures and on human monocytes challenged with B. cenocepacia LPS were also performed. GVF27 exerts, at different stages of formation, anti-biofilm effects towards both Bcc strains, a significant propensity to function in combination with ciprofloxacin, a relevant affinity for LPSs isolated from B. cenocepacia as well as a good propensity to mitigate the release of pro-inflammatory cytokines in human cells pre-treated with the same endotoxin. Overall, all these findings contribute to the elucidation of the main features that a good therapeutic agent directed against these extremely leathery biofilm-forming bacteria should possess.

Cationic Host Defense Peptides (HDPs) are endowed with a broad variety of activities, including direct antimicrobial properties and modulatory roles in the innate immune response. Even if it has been widely demonstrated that bacterial membrane represents the main target of peptide antimicrobial activity, the molecular mechanisms underlying membrane perturbation by HDPs have not been fully clarified yet. Recently, two cryptic HDPs have been identified in human apolipoprotein B and found to be endowed with a broad-spectrum antimicrobial activity, and with anti-biofilm, wound healing and immunomodulatory properties. Moreover, ApoB derived HDPs are able to synergistically act in combination with conventional antibiotics, while being not toxic for eukaryotic cells. Here, by using a multidisciplinary approach, including time killing curves, Zeta potential measurements, membrane permeabilization assays, electron microscopy analyses, and isothermal titration calorimetry studies, the antimicrobial effects of ApoB cryptides have been analysed on bacterial strains either susceptible or resistant to peptide toxicity. Intriguingly, it emerged that even if electrostatic interactions between negatively charged bacterial membranes and positively charged HDPs play a key role in mediating peptide toxicity, they are strongly influenced by the composition of negatively charged bacterial surfaces and by defined extracellular microenvironments.

Objective: This study investigates microcapsulated aqueous extracts of Garcinia kola (GK) and Hunteria umbellata (HU) seeds. Method: Extracts obtained after maceration of dried powdered seeds were prepared as microcapsules with chitosan-alginate by counterion coacervation method. Microcapsules were characterized using differential scanning calorimetry (DSC), x-ray diffractometry (XRD) and fourier transform infrared (FTIR) spectroscopy. In vitro release studies were carried out at pH 1.2 for 2 h and 6.8 for a further 10 h. Results: Between 20 and 50% extract release occurred from microcapsules after 2 h while conventional tablets released 100% after 1 h at simulated gastric pH. At pH 6.8, >80% of extract was released from microcapsules after 6 h. DSC revealed the presence of complex materials. XRD and FTIR showed stable character of the plant extracts within the microcapsules. Conclusion: Controlled release of aqueous extracts derived from these plants was achieved by microencapsulation and therefore can be developed as suitable delivery devices.

Objective: This study investigates the effect of selected excipients on microcapsules properties of seeds extracts of Garcinia kola (GK) and Hunteria umbellata (HU). Method: Extracts from macerated dried powdered seeds were microencapsulated with chitosan-alginate including either talc, HPMCAS, Eudragit®RLPO or Eudragit®L100 up to 25% in the core. Microcapsules were characterized using differential scanning calorimertry (DSC), X-ray diffractometry (XRD), Fourier transform infrared spectroscopy (FTIR) and In vitro release. Results: DSC, XRD and FTIR showed no change in crystalline properties and functional groups of microcapsules and excipients in the formulations. In vitro extract release showed retardation of extract release in acid medium after 2 h and between 80 to 100% at pH 6.8. Generally, the microcapsules fitted more closely to first order model than the other models. Conclusion: Inclusion of these excipients did not impart negatively in interaction and release profile on microcapsules containing GK and HU aqueous extracts and can therefore be added to facilitate microcapsule production and enhance dosage form performance.

We investigate the structure, the morphology and mechanical properties of crystalline-crystalline and crystalline-amorphous di-block copolymers synthesized by using living, stereoselective polymerization catalysts. Cystalline-crystalline block copolymers (BCP) are formed by polyethylene (PE) block linked to isotactic polypropylene (iPP) or syndiotactic polypropylene (sPP) blocks while crystalline-amorphous samples are formed by PE or sPP block linked to an amorphous rubbery ethylene-co-propylene (EP) segment. A detailed structural characterization has been performed for samples crystallized in the bulk, thin films and stretched fibers and stress-induced phase transformations have been also analyzed. Thin films of BCPs have been epitaxially crystallized onto crystals of organic substrates to achieve a control over the crystallization of both blocks and to obtain ordered nanostructures where the two blocks are organized in separated alternating lamellar domains guided by the orientation of the crystalline lamellae.

The concept of “catalyst ligand design” for olefin polymerization has been deeply modified by the use of high throughput screening techniques in the last ten years. A special case is the new class of Hf(IV)-pyridylamido-based olefin polymerization catalysts discovered by a Dow/Symyx collaboration. These systems have attracted a lot of attention because of several unique features: a) they are industrially relevant producing highly isotactic polypropylenes with a solution process technology at temperature > 100 °C; b) in combination with a second catalyst they undergo reversible trans-alkylation, performing novel olefin block copolymers via ‘chain shuttling’. The stereoselectivity mechanism of propene polymerization promoted by such systems is still unknown . In this communication by means of computational methods we investigated the enantioselectivity of propene insertion for systems I-III reported in Chart 1. Surprisingly, the generally accepted model of "chiral growing chain orientation" was ineffective for this class of catalysts and a new model for the propene enantioselectivity has been sorted out.

The transformations related to phase changes of the crystals, and at lamellar length scales by effect of tensile deformation are studied in the case of some isotactic polypropylene samples having high molecular mass, polydispersity index ≈2, and stereodefects at different concentrations and with a uniform distribution, The stress induced transformations are followed in real time during stretching through wide and small angle X-ray scattering measurements. The data analysis evidences that during the transformations of the spherulitic into the fibrillar morphology, stress-induced phase transitions occurring during plastic deformation are regulated by the same factors that govern the textural and morphological changes, that is the ability of the entangled amorphous chains to transmit the stress and the intrinsic stability of the lamellar crystals. Since the relative stability of the different polymorphic forms involved in the structural transformations and the intrinsic flexibility of the chains depend on the stereoregularity, precise correlations between the stereoregularity of the chains, and the deformation behavior are outlined, paving the way for understanding the material properties at molecular level

This study investigates the properties of microparticles prepared from Andrographis paniculata (AP) and Annona muricata (AM) aqueous extracts for controlled release. Extracts obtained by maceration of the dried powdered plant leaves were microencapsulated by counterion coacervation method. Microcapsules were characterized using Fourier-transform infrared-spectroscopy (FTIR), x-ray difractometry (XRD) and differential scanning calorimetry (DSC).In vitro release studies were carried out at pH 1.2 for 2 h and 6.8 for a further 10 h. Release was monitored at274 and 230 nm for AM and AP, respectively. Encapsulation efficacy was less than 52% for AP and 70% for AM. In vitro drug release at pH 1.2 showed less than 40% release from the microcapsules after 2h while over 90% of extract was released after 6h at pH 6.8. Conventional capsules released the content within 1 h in simulated gastric fluid. FTIR, XRD and DSC results indicate the stable character of the extract within the microcapsules. Microencapsulation with chitosan- alginate controlled the release of Andrographis paniculata (AP) and Annona muricata (AM) aqueous extracts.

Palladium (Pd) and palladium oxide (PdO) nanoparticles (NPs) synthesis has gained considerable interest in the last few decades due to their numerous potential utilizations [1-3]. Systems based on Pd show high activity and selectivity in numerous catalytic processes [2, 3], like the catalytic formation of C-C bonds, one of the most useful transformations in organic synthesis [2]. Because sustainable development involves the utilization of reusable catalysts, the search for new catalytic systems to replace existing homogeneous ones is one important issue [4]. In this context, the immobilization of NPs on a solid support is an interesting alternative since the catalyst can be recycled by simple filtration and the atom/ion leaching from the particles can be minimized. Also, many electronic devices, such as resistors, use compositions containing palladium. All these applications may be more suitable with NPs in a patterned form and, since the majority of unique NPs properties are highly microstructure-dependent, to form Pd and PdO arrays on solid supports with a precise control of nanoparticles dimensions and spacing is a an important issue. Here, we report a simple method to fabricate arrays of palladium (Pd) nanoclusters and palladium oxide (PdO) nanoparticles with tunable dimensions and lateral spacing by using polystyrene-block-poly(ethylene oxide) copolymers (PS-b-PEO) as both stabilizers for Pd NPs and templates controlling the distribution of them. The cylindrical self-assembled nanostructure formed from PS-b-PEO has been used as host for selectively sequestering a nanoparticles precursor (palladium(II) acetate) in cylindrical domains of PEO. Then, Pd NPs have been obtained by reduction of the precursor in presence of the BCPs. In this way, nanocomposite thin films of well-defined morphology based on PS-b-PEO matrices and Pd NPs have been obtained. Furthermore, PdO NPs of improved stability against aggregation have been obtained on a solid and conductive support (silicon wafer) by treating the Pd containing films in air at elevated temperatures. We were able to modulate the dimensions of Pd nanoclusters and PdO NPs and their inter-distance by using as template two different PS-b-PEO copolymers having different molecular weights of both PS and PEO blocks.

We report the morphology, structure and mechanical properties of semicrystalline, polyolefin-based block copolymers (BCPs) with the aim of obtaining ordered nanostructures and innovative thermoplastic elastomers. By using living, stereoselective insertion polymerization catalysts, syndiotactic polypropylene-block-poly(ethylene-co-propylene) (sPP-b-EP) and syndiotactic polypropylene-block-polyethylene (sPP-b-PE) were synthetized.1 Thin films of BCPs were prepared from solution by dip-casting. The formation of large-sized, well-oriented lamellar nanostructures of BCPs composed of a crystallizable sPP block linked to an amorphous rubbery block EP (sPP-b-EP) was successfully achieved by using epitaxial crystallization2 with p-terphenyl as substrates. TEM bright field images and electron diffraction patterns of gold decorated nanostructures reveals the presence of well oriented crystalline lamellae ordered over large area alternated to amorphous layers. Crystalline-crystalline block copolymers (sPP-b-PE) epitaxially crystallized onto crystals of p-terphenyl show a more complex lamellar morphology. For BCPs with high volume fraction of sPP block only one lamellar orientation can be observed. In this case sPP crystallize first. For sPP-PE BCPs with similar block lengths, TEM images show regions characterized by double orientation of PE lamellae due to the epitaxial relationship of PE with p-terphenyl. In these regions PE crystallize first. They alternate with regions presenting a single lamellar orientation of sPP according to the epitaxial relationship with p-terphenyl. In these regions sPP crystallize first. In other words the block that crystallize first determines the local morphology. An analysis of the mechanical properties and a detailed structural study of BCPs have been performed in order to clarify the effects of the presence of crystalline (PE) or amorphous (EP) block on the crystallization behavior of sPP, and in particular on the stress-induced transformations.

In the last ten years, the more relevant catalysts for stereoselective olefin polymerization have been discovered by high throughput screening techniques (HTS) [1,2]. The catalyst discovery workflow using the HTS infrastructure reached such a level [3] that the “familiar” concept of “catalyst ligand design” by basic principles [4] seems to play a marginal role at the present stage. By using this approach a greater number of pyridyl-amine ligands with enhanced structural diversity were synthesized (see Figure 1). In designing the expanded ligand array, three convenient positions for structural amplification were identified, labeled in the Figure as R1, R2 and the presence of unusual reactivity of metal-aryl bond (see Hf-CAryl and Hf-CAlkyl bonds in system 3 of Figure 1). However, the stereoselectivity mechanism of propene polymerization promoted by such systems is still unknown [1,2]. In this communication, by means of computational methods, we investigated the enantioselectivity of propene insertion for systems 1-3 reported in Figure 1. Surprisingly, the generally accepted model of "chiral growing chain orientation" [5] was ineffective for this class of catalysts and a new model for the propene enantioselectivity has been sorted out. Despite the computational results tuned by DFT methods combined with dispersion and solvent corrections [6], there are two aspects that we would like to stress: a) our model is able to explain the subtle interplay between the steric and electronic effects as well as the reactivity of metal-aryl bond; b) it appears a general model, able after 35 years to reconnect the Ziegler-Natta catalysis to the more general asymmetric catalysis [7]. Let us make a final comment: we are well aware that discoveries of such systems were possible thanks to experimental HTS technologies because of the huge number of variables involved; nevertheless, we are still convinced that a rational catalysts design is not over the game as soon as new models are developed.

A study of the crystallization of the mesomorphic form of isotactic polypropylene (iPP) in samples of different stereoregularity prepared with metallocene catalysts is presented. Contrary to what claimed in the literature, we have found that the mesomorphic form can be obtained by quenching of the melt at 0 °C also in the case of low stereoregular samples, provided that the samples are kept at 0 °C for long time. The key is the formation of mesomorphic bundles with chains in ordered 3/1 helical conformation of size large enough to be stable and inhibit the crystallization of the alpha form at room temperature. For stereoirregular samples the concentration of long ordered helical stretches is low and this requires long residence time at 0 °C for the formation of mesomorphic aggregates of size larger than the critical size. This result provides evidence that the mesomorphic aggregates are not formed as a result of a cooperative process driven by entropy, as in the case of liquid crystals, but rather they form via a nucleation and growth mechanism, as in the normal crystallization processes, according to the idea that the mesophase of iPP is a highly defective crystalline form.

The effects of the presence of hexene and octene on the crystallization behavior of syndiotactic polypropylene (sPP), and in particular on the stress-induced transformations occurring during stretching have been analyzed. For hexene or octene concentrations higher than 4-5 mol% the helical form I transforms by stretching at high deformation into a different mesomorphic form, characterized by chains in the ordered 2/1 helical conformation, and disorder in the lateral packing of the chains.

Il presente lavoro di tesi è incentrato sullo studio di nuovi materiali a base di copolimeri a blocchi amorfi e semicristallini. Tali studi hanno evidenziato come sia possibile controllare il tipo di nanostruttura che si ottiene per separazione di fase, attraverso il controllo della cristallizzazione, utilizzando tecniche innovative basate sulla solidificazione direzionale da solventi cristallizzabili e sulla cristallizzazione epitassiale su opportuni supporti. Sono stati studiati copolimeri a blocchi semiscristallini a base di polietilene (PE), polipropilene sindiotattico (sPP), poli(L-lattide) (PLLA) e poliossietilene (PEO). I copolimeri semicristallini a base di PE e sPP sono stati sintetizzati mediante nuovi catalizzatori metallorganici, ed è stato effettuato uno studio strutturale e morfologico su tutti i campioni. In particolare gli studi morfologici condotti su questi campioni hanno evidenziato come sia possibile controllare il tipo di nanostruttura che si ottiene per separazione di fase, attraverso il controllo della cristallizzazione in modo da indurre la formazione di nanostrutture orientate a morfologia lamellare con ordine direzionale e periodicità dei nanodomini dei due blocchi a lungo raggio. Oltre ai copolimeri a blocchi semicristallini sono stati studiati copolimeri formati da due blocchi amorfi, in particolare è stato studiato un copolimero costituito da un blocco di polistirene (PS) legato ad un blocco di polimetilmetacrilato (PMMA). E’ stato messo appunto un semplice metodo che permette di realizzare devices elettronici con funzioni di memoria utilizzando come layer attivo la matrice polimerica nanostrutturata generata dal copolimero a blocchi PS-b-PMMA.

Homogeneous catalysts for propylene polymerization are now available for production of isotactic polypropylene (PP) with stereoselectivities approaching that of heterogeneous Ziegler-Natta (ZN) catalysts. Ewen1 and Kaminsky2 demonstrated that the microstructure of PP was predictable when produced with catalysts operating by site control mechanisms and established the relationships, known as “Ewen’s symmetry rules”, which correlate the chirality and symmetry of the catalyst to the microstructures of PP. In this communication we report a theoretical analysis by DFT calculations on the propene polymerization promoted by systems formed by N-heteroaryl-pyridylamido tridentate ligands and the corresponding (N_,N,N_)MIV(Me2)2] pentacoordinate complexes (M=Zr, Hf) (systems 1 and 2 in Chart 1).3 Our results suggest that for these systems: a) the mechanisms of stereoselectivity contradict the “Ewen’s symmetry rules”; b) the tacticity control is much more subtle and difficult to predict than the one performed by tetrahedral ansametallocene4 as well as post-metallocene systems.5 Nevertheless, breaking the generally accepted symmetry-rules open new perspectives to achieve stereoselective catalysis.

For their easy and high-yield recombinant production, their high stability in a wide range of physico-chemical conditions and their characteristic hollow structure, ferritins (Fts) are considered useful scaffolds to encapsulate bioactive molecules. Notably, for the absence of immunogenicity and the selective interaction with tumor cells, the nanocages constituted by the heavy chain of the human variant of ferritin (hHFt) are optimal candidates for the delivery of anti-cancer drugs. hHFt nanocages can be disassembled and reassembled in vitro to allow the loading of cargo molecules, however the currently available protocols present some relevant drawbacks. Indeed, protein disassembly is achieved by exposure to extreme pH (either acidic or alkaline), followed by incubation at neutral pH to allow reassembly, but the final protein recovery and homogeneity are not satisfactory. Moreover, the exposure to extreme pH may affect the structure of the molecule to be loaded. In this paper, we report an alternative, efficient and reproducible procedure to reversibly disassemble hHFt under mild pH conditions. We demonstrate that a small amount of sodium dodecyl sulfate (SDS) is sufficient to disassemble the nanocage, which quantitatively reassembles upon SDS removal. Electron microscopy and X-ray crystallography show that the reassembled protein is identical to the untreated one. The newly developed procedure was used to encapsulate two small molecules. When compared to the existing disassembly/reassembly procedures, our approach can be applied in a wide range of pH values and temperatures, is compatible with a larger number of cargos and allows a higher protein recovery.

Highly accurate high-throughput experimentation (HTE) data for a set of 21 silicon-bridged C2-symmetric ansa-zirconocenes in propene homopolymerization were collected and were used to develop quant...

A study of the structure and morphology of diblock copolymers composed of crystallizable blocks of polyethylene (PE) and syndiotactic polypropylene (sPP) having different lengths is reported. In both analyzed samples, the PE block crystallizes first by cooling from the melt (at 130 °C) and the sPP block crystallizes after at a lower temperature. Small angle X-ray scattering (SAXS) recorded during cooling showed three correlation peaks at values of the scattering vector, q1 = 0.12 nm−1, q2 = 0.24 nm−1 and q3 = 0.4 nm−1, indicating development of a lamellar morphology, where lamellar domains of PE and sPP alternate, each domain containing stacks of crystalline lamellae of PE or sPP sandwiched by their own amorphous phase of PE or sPP. At temperatures higher than 120 °C, when only PE crystals are formed, the morphology is defined by the formation of stacks of PE lamellae (17 nm thick) alternating with amorphous layers and with a long period of nearly 52 nm. At lower temperatures, when crystals of sPP are also well-formed, the morphology is more complex. A model of the morphology at room temperature is proposed based on the correlation distances determined from the self-correlation functions extracted from the SAXS data. Lamellar domains of PE (41.5 nm thick) and sPP (8.2 nm thick) alternate, each domain containing stacks of crystalline lamellae sandwiched by their own amorphous phase, forming a global morphology having a total lamellar periodicity of 49.7 nm, characterized by alternating amorphous and crystalline layers, where the crystalline layers are alternatively made of stacks of PE lamellae (22 nm thick) and thinner sPP lamellae (only 3.5 nm thick).

Plant-derived essential oils (EOs) represent a green alternative to conventional antimicrobial agents in food preservation. Due to their volatility and instability, their application is dependent on the development of efficient encapsulation strategies allowing their protection and release control. Encapsulation in Polyhydroxyalkanoate (PHA)-based nanoparticles (NPs) addresses this challenge, providing a biodegradable and biobased material whose delivery properties can be tuned by varying polymer composition. In this work, EO from Mexican oregano was efficiently encapsulated in Polyhydroxybutyrate (PHB) and Poly-3-hydroxybutyrate-co-hydroxyhexanoate (PHB-HHx)-based NPs by solvent evaporation technique achieving high encapsulation efficiency, (>60%) and loading capacity, (about 50%). The obtained NPs displayed a regular distribution with a size range of 150–210 nm. In vitro release studies in food simulant media were fitted with the Korsmeyer–Peppas model, indicating diffusion as the main factor controlling the release. The cumulative release was affected by the polymer composition, possibly related to the more amorphous nature of the copolymer, as confirmed by WAXS and DSC analyses. Both the EO-loaded nanosystems displayed antimicrobial activity against Micrococcus luteus, with PHB-HHx-based NPs being even more effective than the pure EO. The results open the way to the effective exploitation of the developed nanosystems in active packaging.

Protein concentrates were obtained from hemp seed oilcakes (OCs) and investigated as potential waste-derived source of biodegradable films at different protein and glycerol concentrations and at different pH values. These studies indicated that hemp protein (HP) film forming solutions gave rise to higher performance films when cast at pH 12 in the presence of 50% glycerol (w/w protein) used as plasticizer. Since HPs were demonstrated to act as both acyl donor and acceptor substrates of microbial transglutaminase (mTGase), they have been used as raw material to obtain films also after enzyme treatment. Film morphological characterization demonstrated that mTGase treatment was effective to produce more homogeneous and smoother films, influencing in turn positively their properties. In fact, mTGase-crosslinked films were shown to be more resistant, still flexible and exhibited a higher heat-sealing strength. In addition, the enzymatic treatment of HPs originated bio-plastics with a higher gas permeability and a greater hydrophobicity. These findings suggest the possibility to exploit the mTGase-crosslinked proteins derived from hemp OC as a promising source to produce bio-based materials useful as packaging systems for protecting food products from physical contamination and, thus, for extending their shelf-life.

Isotactic polypropylene-block-polyethylene block copolymers (iPP-b-PE) composed of crystalline blocks of isotactic polypropylene (iPP) and linear polyethylene (PE) of different block lengths have been synthesized with a living and stereoselective pyridylamidohafnium catalyst. The structure and morphology of iPP-b-PE that develop upon crystallization from the melt are presented. Temperature and time resolved X-ray diffraction experiments in wide (WAXS) and small angles (SAXS) recorded during cooling from the melt indicate that PE blocks crystallize first upon cooling and the iPP block crystallizes at lower temperatures and nucleates over the crystals of PE formed before. The SAXS data indicate that PE lamellar stacks formed first at high temperature have higher periodicity of 30 nm, whereas iPP lamellar stacks formed after at lower temperature have lower periodicity of 11–12 nm. The crystallization of the iPP and PE blocks drives microphase separation and determines the final morphology leading to a lamellar morphology characterized by alternation of amorphous and crystalline layers. At nanoscale the morphology is characterized by alternating iPP and PE microdomains, each domain including stacks of lamellar crystals of iPP or PE of different periodicities, separated by the corresponding amorphous regions of iPP or PE. Polarized optical micrographs of the melt-crystallized samples show the presence of bundle-like crystals in samples with longer block of iPP and of small banded spherulites in samples with longer block of PE. The observation of banded spherulites typical of PE suggests that the crystallization of PE defines the global morphology and is not altered by the structure of the melt, confirming that crystallization of PE occurs first upon cooling from the melt.