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Three novel hydrophobic deep eutectic solvents (DESs) based on oleic acid and terpenes (thymol, DL-menthol, and geraniol) were prepared, characterized, and used to extract astaxanthin from the microalga Haematococcus pluvialis without any pre-treatment of the cells. The three DES were composed of Generally Recognized As Safe (GRAS) and edible ingredients. All the tested DESs gave astaxanthin recovery values of about 60 and 30% in 6 h if applied on freeze-dried biomass or directly on algae culture, respectively. The carotenoid profile was qualitatively identical to what was obtained by using traditional organic solvents, regardless of the DES used; the monoesters of astaxanthin with C18-fatty acids were the main compounds found in all the carotenoid extracts. The thymol:oleic acid DES (TAO) could preserve astaxanthin content after prolonged oxidative stress (40% of the astaxanthin initially extracted was still present after 13.5 h of light exposure), thanks to the superior antioxidant properties of thymol. The capacity of improving astaxanthin stability combined with the intrinsic safety and edibility of the DES components makes the formulation astaxanthin-TAO appealing for the food ingredients/additives industry.

The recycling of polyhydroxybutyrate (PHB) via depolymerization-polymerization is here proposed and assessed for the first time. Effective depolymerization of PHB to crotonic acid through a thermolytic distillation at mild conditions (170 °C and 150 mbar), and without the need of any catalyst, gives a crotonic acid-enriched condensate (94% yield and 98% selectivity towards trans-crotonic acid) that was used without any further purification as a substrate for producing renewed PHB with a culture of Cupriavidus necator. The yield of PHB (0.6 g PHB per g of crotonic acid), final PHB content (63%), and PHB-molecular weight (1.5 MDa) confirmed the technical feasibility of this tandem approach, with a 55% overall yield. A preliminary evaluation of the energy consumption of various depolymerization approaches indicated that the thermolytic distillation consumes 20–25% less energy than previously reported methods; the energy requirement for producing crotonic acid from PHBwaste with 50% moisture has a still acceptable energy demand (4 MJ/kgcrotonic acid), comparable to the energy required for obtaining fermentable sugars (4.4 MJ/kgsugar) to be used for feeding bacteria that accumulate PHB.

One of the most widely used approaches for improving drug permeation across the stratum corneum barrier of the skin is the use of chemical penetration enhancers, such as surfactants. In this study, two anionic surfactants, named C12-OPK and C18-OPK, were synthesized via condensation of itaconic acid and fatty amines, with C12 and C18 alkyl chains, respectively. Assessment of impacts on HaCaT keratinocyte cell viability was used as indicator of their potential to cause skin irritation 24 h post exposure (Alamar Blue assay). The LC50 values of C12-OPK and C18-OPK (144 and 85 mg/L, respectively) were lower than LC50 values of the most used commercial surfactants (e.g. SDS). The effect of different surfactant concentrations (up to ten times the critical micellar concentration, CMC) on hydrocortisone (HC) solubility and permeation through porcine skin was also evaluated. Results showed that drug solubility increased linearly with increasing concentrations of both surfactants, as a consequence of the association between drug and micelles. In vitro permeation results showed that the permeability coefficient increased at surfactant concentrations lower than the CMC. In particular, a higher enhancement effect on drug permeation was obtained with C18-OPK, due to its hydrophobic properties that ensured a more effective HC permeation in comparison to C12-OPK.

Pyrolysis was explored as a technology for depolymerizing and charring starch-based plastic bags in a novel valorization perspective based on a chemical recycling strategy, alternative to anaerobic digestion or composting. The char obtained from the thermal treatment (420°C for 15 h) was sulfonated to give a highly active and recyclable heterogeneous acid catalyst (10 wt% yield based on starch-based plastic bags subjected to pyrolysis), comparable in terms of catalytic activity and reusability to the analog catalyst prepared from potato starch and tested on a model esterification reaction. From pyrolysis liquid of starch-based plastic bags, highly pure terephthalic acid (4 wt% yield) was isolated through self-precipitation, whereas the remaining pyrolysis liquid produced (21 wt%) was solvent-fractionated to give a polar fraction (7 wt% yield) enriched in levoglucosan, and a less polar fraction (14 wt% yield) enriched in monobutenyl adipate and terephthalate.

In view of the potential use of pyrolysis-based technologies, it is crucial to understand the environmental hazards of pyrolysis-derived products, in particular bio-oils. Here, three bio-oils were produced from fast pyrolysis of pine wood and intermediate pyrolysis of corn stalk and poultry litter. They were fully characterized by chemical analysis and tested for their biodegradability and their ecotoxicity on the crustacean Daphnia magna and the green alga Raphidocelis subcapitata. These tests were chosen as required by the European REACH regulation. These three bio-oils were biodegradable, with 40-60% of biodegradation after 28 days, and had EC50 values above 100mgL(-1) for the crustacean and above 10mgL(-1) for the alga, showing low toxicity to the aquatic life. The toxic unit approach was applied to verify whether the observed toxicity could be predicted from the data available for the substances detected in the bio-oils. The predicted values largely underestimated the experimental values.

CO2 is a renewable, abundant and cheap C1-feedstock and its conversion to cyclic carbonates starting from epoxides has been widely explored in the last years. Nevertheless, conducting this reaction under mild and sustainable conditions is still a challenging task. Herein we present the use of choline-salt based eutectic mixtures as catalysts for the reaction of CO2 with epoxides to give cyclic carbonates. Choline chloride and choline iodide have been coupled with various hydrogen bond donors (HBDs), mainly cheap and bio-based carboxylic acids and polyols, to form two classes of eutectic mixtures. Very good yields were achieved under mild conditions (80 °C in 7–22 h) for various terminal epoxides, with both classes of catalysts. While a pressure of 0.4 MPa of CO2 is required to obtain appreciable conversions using choline chloride-based mixture, atmospheric pressure of CO2 (balloon) has been successfully used with choline iodide-based mixtures. Furthermore, the catalysts could be recycled without appreciable loss of the catalytic activity. The improved catalytic performance of both choline-based eutectic mixtures is attributed to the synergistic activity of the halide, responsible for the opening of the epoxy-ring, and the HBD that has a role in the stabilization of the alkoxide intermediate.

Ionic liquids (ILs) are promising materials exploited as solvents and media in many innovative applications, some already used at the industrial scale. The chemical structure and physicochemical properties of ILs can differ significantly according to the specific applications for which they have been synthesized. As a consequence, their interaction with biological entities and toxicity can vary substantially. To select highly effective and minimally harmful ILs, these properties need to be investigated. Here we use the so called chromatophores - protein-phospholipid membrane vesicles obtained from the photosynthetic bacterium Rhodobacter sphaeroides- to assess the effects of imidazolinium and pyrrolidinium ILs, with chloride or dicyanamide as counter anions, on the ionic permeability of a native biological membrane. The extent and modalities by which these ILs affect the ionic conductivity can be studied in chromatophores by analyzing the electrochromic response of endogenous carotenoids, acting as an intramembrane voltmeter at the molecular level. We show that chromatophores represent an in vitro experimental model suitable to probe permeability changes induced in cell membranes by ILs differing in chemical nature, degree of oxygenation of the cationic moiety and counter anion. (C) 2015 Elsevier B.V. All rights reserved.

Obtainment and testing of pure enantiomers are of great importance for bioactive compounds, because of the assessed implications of enantioselectivity in receptor-mediated responses. Herein we evaluated the use of biocatalysis to obtain enantiomerically pure β-lactam intermediates further exploited in the synthesis of novel integrin ligands as single enantiomers. From a preliminary screening on a set of commercially available hydrolases, Burkholderia Cepacia Lipase (BCL)emerged as a suitable and highly performing enzyme for the kinetic resolution of a racemic azetidinone, key intermediate for the synthesis of novel agonists of integrins. Upon optimization of the biocatalytic protocol in terms of enzymes, acylating agents and procedures, the two β-lactam enantiomers were obtained in excellent enantiomeric excesses (94% and 98% ee). Synthetic elaborations on the separated enantiomers allowed the synthesis of four chiral β-lactams which were evaluated in cell adhesion assays on Jurkat cell line expressing α 4 β 1 integrin, and K562 cell line expressing α 5 β 1 integrin. Biological tests revealed that only (S)-enantiomers maintained the agonist activity of racemates with a nanomolar potency, and a specific enantio-recognition by integrin receptors was demonstrated.

Since the 1990s red-tide blooms of Fibrocapsa japonica have frequently been observed in European waters; despite their recurrence also in the Adriatic coastal areas, they have never been conclusively linked to ichthyotoxic events. Since the toxicity of F. japonica is still under debate and its effects differ among the strains, in this study all the compounds previously postulated as involved in the toxic mechanism, such as brevetoxins, fatty acids and ROS, were screened for Adriatic strains. Whole algal extracts were analyzed for brevetoxin PbTx-2, which was not found, and for a qualitative and quantitative analysis of fatty acids. The fatty acid profile evidenced the presence of PUFAs, with considerable amounts of 18:4n-3, 20:4n-6, and 20:5n-3, as already found in different strains, but with a lower amount of arachidonic acid (20:4n-6). None of the PUFAs was released in the extracellular medium. Different toxicological assays (Vibrio fischeri, Artemia sp., haemolysis of fish erythrocytes) were performed using algal cellular, extracellular or subcellular samples as well as fractions of the algal extract, in order to identify the toxic compounds. The tested cultures were found to inhibit V. fischeri bioluminescence, to affect Artemia nauplii viability and to have haemolytic effects comparable to those previously observed in different strains. The solid phase extraction (SPE) of the Adriatic F. japonica extracts resulted in a high concentration of PUFAs in two main fractions, which caused mortality in Artemia nauplii and inhibition of V. fischeri bioluminescence. Fish (Dicentrarchus labrax) assays were also performed, reporting mortality after long exposure times and the cause of fish death was investigated. A significant increase of H2O2 in the tanks where sea basses were exposed to F. japonica was obtained, indicating that the presence of fish stimulates H2O2 production by algal cells and leading to the hypothesis of H2O2 involvement in F. japonica toxicity. The presence of oxidative stress in the exposed fish was confirmed by the increased malondialdehyde concentration in their gills. Overall these results indicate that since F. japonica cells can easily cling to fish gills, due to their high polysaccharide production, it is presumable that high PUFA amounts are released in loco, as a result of cell breakage, and that exert their toxicity together with ROS. This study shows that the Adriatic strains of F. japonica can be harmful to higher level organisms. Nevertheless a high cell density and a long lasting bloom are necessary to cause severe damage to fish gills or death for crustaceans.

Lipid extraction is a critical step in the development of biofuels from microalgae. Here a new procedure was proposed to extract hydrocarbons from dried and water-suspended samples of the microalga Botryococcus braunii by using switchable-polarity solvents (SPS) based on 1,8-diazabicyclo-[5.4.0]-undec-7-ene (DBU) and an alcohol. The high affinity of the non-ionic form of DBU/alcohol SPS towards non-polar compounds was exploited to extract hydrocarbons from algae, while the ionic character of the DBU-alkyl carbonate form, obtained by the addition of CO(2), was used to recover hydrocarbons from the SPS. DBU/octanol and DBU/ethanol SPS were tested for the extraction efficiency of lipids from freeze-dried B. braunii samples and compared with n-hexane and chloroform/methanol. The DBU/octanol system was further evaluated for the extraction of hydrocarbons directly from algal culture samples. DBU/octanol exhibited the highest yields of extracted hydrocarbons from both freeze-dried and liquid algal samples (16% and 8.2% respectively against 7.8% and 5.6% with n-hexane).

We report the first temperature study on the stereoselectivity of the ‘in-water’ aldol reaction between nicotinaldehyde (3-pyridinecarbaldehyde) and cyclohexanone catalyzed by morpholine and trans -4- tert -butyldimethylsilyloxy- l -proline. Eyring plots of diastereomeric ratio anti / syn gave a constant diastereoselectivity with respect to reaction temperature using morpholine as catalyst. With O(TBS)- l -proline we observed a non-linear behaviour of the Eyring plot with the presence of an inversion temperature ( T inv ), which disclosed dynamic solvation effects in water.

In addition to their antibiotic potency, beta-lactams have recently been investigated as inhibitors of serine proteinase such as leukocyte elastase (LE), released by inflammatory cells. We describe the synthesis of a series of 4-alkylidene-beta-lactams, and investigate how substitutions on C-3, C-4, and N-1 of the beta-lactam ring affect the activity of human LE and gelatinases MMP-2 and MMP-9. LE activity was measured using a chromogenic substrate, while gelatin-zymography assay was used to evaluate gelatinase activity. We demonstrate that C-4 unsaturation on the beta-lactam ring determines the degree of biological activity, with a selectivity over LE by 3-[1-(tert-butyldimethylsilyloxy)-ethyl] derivatives (lowest IC(50) was 4 microM), and over gelatinase MMP-2 by C-3-unsubstituted 4-[1-ethoxycarbonyl]-ethylidene-beta-lactams (lowest IC(50) was 60 microM). (3S)-3-[(1R)-1-hydroxyethyl]-4-(1-ethoxycarbonyl)-ethylidene-azetidin-2-one inhibits gelatinase MMP-9. The compounds tested showed no cytotoxicity against NIH-3T3 murine fibroblasts. This is the first example of beta-lactams inhibiting metallo-proteinases instrumental in cancer invasion and angiogenesis. These molecules are good candidates for prototype drugs showing selective antibiotic, anti-inflammatory, and anti-invasion properties.

Synthesis of 3-vinyl- and 3-alkyl-Δ3-cephems was performed in a regioselective manner by palladium acetate catalyzed reaction of 3-sulfonato- or 3-bromo-Δ3-cephems with dialkylzinc or vinyltributyltin. A new access to 3-alkyl-Δ3-cephems is opened, starting from the allenyl compound 10.

N -( p -methoxyphenyl)-hexahydro-1,3,5-triazine in presence of a Lewis acid and ( R )-3-( t -butyldimethylsilyloxy)butyric acid chloride with Et 3 N directly furnish (3 S ,1′ R )- N - p -methoxyphenyl-3-(1- t -butyldimethylsilyloxy)ethylazetidin-2-one with good diastereo-selectivity. This product is transformed into the 4-acetoxy-azetidinone 1 , a key intermediate in the synthesis of β-lactam antibiotics.

A detailed study of the influence of temperature, pH and enzyme amount on the enantiomeric excess in the enzymatic resolution of N -1 protected 3-amino-azetidin-2-ones with supported PGA is presented. Both enantiomers could be obtained in good enantiomeric excess.

The cyanosilylation of a series of N-substituted lactaldehyde imines is reported. The trimethylsilylcyanide addition affords optically active α-amino nitriles. All the imines show a syn diastereofacial selectivity that is maintained irrespective to the nature of the Lewis acid employed and even remains in the noncatalyzed reaction.

Treatment of (3R,4R,1′R)-3-[1-(t-butyldimethylsilyloxy)ethyl]-4-acetoxy-azetidin-2-one with ethyl diazoacetate in the presence of a Lewis acid such as TiCl4, TiF4, AlCl3 or SnCl4 affords the novel (3S,1′R)-3-[1-(t-butyldimethylsilyloxy)ethyl]-4(1-ethoxycarbonyl-methylidene)-azetidin-2-one as an E and/or Z isomer.