Orange Peel Waste as Feedstock for the Production of Glycerol-Free Biodiesel by the Microalgae Nannochloropsis oculata.

The bioconversion of agri-food waste into high-value products is gaining growing interest worldwide. Orange peel waste (OPW) is the main by-product of orange juice production and contains high levels of moisture and carbohydrates. In this study, the orange waste extract (OWE) obtained through acid hydrolysis of OPW was used as a substrate in the cultivation of the marine microalgae Nannochloropsis oculata. Photoheterotrophic (PH) and Photoautotrophic (PA) cultivations were performed in OWE medium and f/2 medium (obtained by supplementing OWE with macro- and micronutrients of f/2 medium), respectively, for 14 days. The biomass yields in PA and PH cultures were 390 mg L-1 and 450 mg L-1, while oil yields were 15% and 28%, respectively. The fatty acid (FA) profiles of PA cultures were mostly represented by saturated (43%) and monounsaturated (46%) FAs, whereas polyunsaturated FAs accounted for about 10% of the FAs. In PH cultures, FA profiles changed remarkably, with a strong increase in monounsaturated FAs (77.49%) and reduced levels of saturated (19.79%) and polyunsaturated (2.72%) FAs. Lipids obtained from PH cultures were simultaneously extracted and converted into glycerol-free biodiesel using an innovative microwave-assisted one-pot tandem protocol. FA methyl esters were then analyzed, and the absence of glycerol was confirmed. The FA profile was highly suitable for biodiesel production and the microwave-assisted one-pot tandem protocol was more effective than traditional extraction techniques. In conclusion, N. oculata used OWE photoheterotrophically, resulting in increased biomass and oil yield. Additionally, a more efficient procedure for simultaneous oil extraction and conversion into glycerol-free biodiesel is proposed.

Anti-Bacterial Adhesion on Abiotic and Biotic Surfaces of the Exopolysaccharide from the Marine Bacillus licheniformis B3-15.

The eradication of bacterial biofilm represents a crucial strategy to prevent a clinical problem associated with microbial persistent infection. In this study we evaluated the ability of the exopolysaccharide (EPS) B3-15, produced by the marine Bacillus licheniformis B3-15, to prevent the adhesion and biofilm formation of Pseudomonas aeruginosa ATCC 27853 and Staphylococcus aureus ATCC 29213 on polystyrene and polyvinyl chloride surfaces. The EPS was added at different times (0, 2, 4 and 8 h), corresponding to the initial, reversible and irreversible attachment, and after the biofilm development (24 or 48 h). The EPS (300 µg/mL) impaired the initial phase, preventing bacterial adhesion even when added after 2 h of incubation, but had no effects on mature biofilms. Without exerting any antibiotic activity, the antibiofilm mechanisms of the EPS were related to the modification of the (i) abiotic surface properties, (ii) cell-surface charges and hydrophobicity, and iii) cell-to-cell aggregation. The addition of EPS downregulated the expression of genes (lecA and pslA of P. aeruginosa and clfA of S. aureus) involved in the bacterial adhesion. Moreover, the EPS reduced the adhesion of P. aeruginosa (five logs-scale) and S. aureus (one log) on human nasal epithelial cells. The EPS could represent a promising tool for the prevention of biofilm-related infections.

Improvement of Biomass and Phycoerythrin Production by a Strain of Rhodomonas sp. Isolated from the Tunisian Coast of Sidi Mansour.

Microalgae are photoautotrophic microorganisms known as producers of a large variety of metabolites. The taxonomic diversity of these microorganisms has been poorly explored. In this study, a newly isolated strain was identified based on the 18S rRNA encoding gene. The phylogenetic analysis showed that the isolated strain was affiliated with the Rhodomonas genus. This genus has greatly attracted scientific attention according to its capacity to produce a large variety of metabolites, including phycoerythrin. Growth and phycoerythrin production conditions were optimized using a Plackett-Burman design and response surface methodology. An expression profile analysis of the cpeB gene, encoding the beta subunit of phycoerythrin, was performed by qRT-PCR under standard and optimized culture conditions. The optimization process showed that maximum cell abundance was achieved under the following conditions: CaCl2 = 2.1328 g/L, metal solution = 1 mL/L, pH = 7 and light intensity = 145 µmol photons/m2/s, whereas maximum phycoerythrin production level occurred when CaCl2 = 1.8467 g/L, metal solution = 1 mL/L, pH = 7 and light intensity = 157 µmol/m2/s. In agreement, positive transcriptional regulation of the cpeB gene was demonstrated using qRT-PCR. This study showed the successful optimization of abiotic conditions for highest growth and phycoerythrin production, making Rhodomonas sp. suitable for several biotechnological applications.

Evaluation of Betacoronavirus OC43 and SARS-CoV-2 Elimination by Zefero Air Sanitizer Device in a Novel Laboratory Recirculation System.

Indoor air sanitizers contrast airborne diseases and particularly severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)/Coronavirus disease 2019 (COVID-19). The commercial air sanitizer Zefero (Cf7 S.r.l., San Giovanni La Punta, Italy) works alternatively using a set of integrated disinfecting technologies (namely Photocatalysis/UV mode) or by generating ozone (Ozone mode). Here we evaluated the virucidal efficacy of Zefero setup modes against human Betacoronavirus OC43 and SARS-CoV-2. For this purpose, we designed a laboratory test system in which each virus, as aerosol, was treated with Photocatalysis/UV or Ozone mode and returned into a recirculation plexiglass chamber. Aerosol samples were collected after different times of exposure, corresponding to different volumes of air treated. The viral RNA concentration was determined by qRT-PCR. In Photocatalysis/UV mode, viral RNA of OC43 or SARS-CoV-2 was not detected after 120 or 90 min treatment, respectively, whereas in Ozone mode, viruses were eliminated after 30 or 45 min, respectively. Our results indicated that the integrated technologies used in the air sanitizer Zefero are effective in eliminating both viruses. As a reliable experimental system, the recirculation chamber developed in this study represents a suitable apparatus for effectively comparing the disinfection capacity of different air sanitizers.

Nutritional conditions of the novel freshwater Coccomyxa AP01 for versatile fatty acids composition.

AIMS: This study was to analyse the biomass production and fatty acids (FAs) profiles in a newly isolated chlorophyte, namely Coccomyxa AP01, under nutritionally balanced (NB) conditions (comparing nitrate and urea as nitrogen sources) and nitrogen or phosphate deprivation. METHODS AND RESULTS: Lipid yields was about 30%-40% of dried biomasses in all examined nutritional conditions. Under NB conditions, lipids were principally constituted by monounsaturated FAs, mainly represented by oleic acid, and saturated and polyunsaturated FAs at similar concentrations. Nutrients deprivation induced remarkable changes in FAs profiles, with the highest amounts of saturated (42%-46%), followed by similar amounts of monounsaturated and polyunsaturated, and the emergence of rare long-chain FAs. Under phosphate deprivation, biomass yield was similar to NB conditions, with the highest yield of saturated (mainly palmitic acid) and of polyunsaturated FAs (33%) (mainly linoleic and linolenic acids). CONCLUSIONS: Balanced or deprived nutritional conditions in Coccomyxa AP01 induced a selective production and composition of FAs. The phosphate-deprivation condition concomitantly provided high biomass yield and the production of high value saturated and polyunsaturated FAs with industrial interest. SIGNIFICANCE AND IMPACT OF THE STUDY: Coccomyxa AP01 could be considered a promising source of different FAs, including also docosapentaenoic acid, for several commercial purposes spanning from biodiesel production, pharmaceutical and cosmetic applications to innovative aquaculture fish feeds.

Sequestering ability to Cu2+ of a new bodipy-based dye and its behavior as in vitro fluorescent sensor.

A Bodipy (4,4-difluoro-4-bora-3a,4a-diaza-s-indacene) derivative has been conceived and synthesized starting from l-aspartic acid, as a selective turn-off sensor of Cu2+ ions. Its acid-base properties were determined to study the formation of metal/sensor complex species by titration of solutions each containing a different metal ion, such as Cu2+, Ca2+, Zn2+, Pb2+ and Hg2+ and different metal/sensor ratios. The speciation models allowed us to simulate the distribution of the metal/sensor complex species at the normal concentrations of the corresponding metals present in biological fluids. The distribution diagrams, obtained by varying the concentration of sensor 1, clearly indicate that sensor 1 responds selectively to Cu2+ at micromolar concentrations, even in the presence of other more abundant metal cations Ca2+. Finally, we analyzed the cellular uptake of sensor 1 on human erythrocytes and its ability to chelate Cu2+ in the cellular environment. Results indicate that it crosses the plasmatic membrane and colors the cells of a bright fluorescent red. Exposing the fluorescent cells to Cu2+ results in a complete cellular photobleaching of the red fluorescence, indicating that sensor 1 is able to detect metal changes in the cytosolic environment.

A micro-Raman spectroscopic investigation of leukemic U-937 cells in aged cultures.

Recently it has been shown that micro-Raman spectroscopy combined with multivariate analysis is able to discriminate among different types of tissues and tumoral cells by the detection of significant alterations and/or reorganizations of complex biological molecules, such as nucleic acids, lipids and proteins. Moreover, its use, being in principle a non-invasive technique, appears an interesting clinical tool for the evaluation of the therapeutical effects and of the disease progression. In this work we analyzed molecular changes in aged cultures of leukemia model U937 cells with respect to fresh cultures of the same cell line. In fact, structural variations of individual neoplastic cells on aging may lead to a heterogeneous data set, therefore falsifying confidence intervals, increasing error levels of analysis and consequently limiting the use of Raman spectroscopy analysis. We found that the observed morphological changes of U937 cells corresponded to well defined modifications of the Raman contributions in selected spectral regions, where markers of specific functional groups, useful to characterize the cell state, are present. A detailed subcellular analysis showed a change in cellular organization as a function of time, and correlated to a significant increase of apoptosis levels. Besides the aforementioned study, Raman spectra were used as input for principal component analysis (PCA) in order to detect and classify spectral changes among U937 cells.

Viable but nonculturable state of foodborne pathogens in grapefruit juice: a study of laboratory.

Several foodborne human pathogens, when exposed to harsh conditions, enter viable but nonculturable (VBNC) state; however, still open is the question whether VBNC pathogens could be a risk for public health, because, potentially, they can resuscitate. Moreover, cultural methods for food safety control were not able to detect VBNC forms of foodborne bacteria. Particularly, it has not been established whether food chemophysical characteristics can induce VBNC state in contaminating pathogen bacterial populations, especially in food, such as salads and fresh fruit juices, not subjected to any decontamination treatment. In this preliminary study, we intentionally contaminated grapefruit juice to determine whether pathogen bacteria could enter VNBC state. In fact, grapefruit juice contains natural antimicrobial compounds, has an average pH of about 3 and low content in carbohydrates. Such characteristics make grapefruit juice a harsh environment for microbial survival. For this purpose, Escherichia coli O157:H7, Salmonella enterica serovar Typhimurium ATCC 14028, Listeria monocytogenes ATCC 7644, and Shigella flexneri ATCC 12022, at two different inoculum sizes, have been used. Viability by the LIVE/DEAD BacLight Bacterial Viability kit and culturability by plate counts assay were monitored, whereas "resuscitation" of nonculturable populations was attempted by inoculation in nutrient-rich media. The data showed that L. monocytogenes lost both culturability and viability and did not resuscitate within 24 h independently on inoculum size, whereas E. coli O157:H7 was able to resuscitate after 24 h but did not after 48 h. Salmonella Typhimurium and S. flexneri, depending on inoculum size, lost culturability but maintained viability and were able to resuscitate; moreover, S. flexneri was still able to form colonies after 48 h at high inoculum size. In conclusion, entry into VBNC state differs on the species, depending, in turn, on inoculum size and time of incubation.

Synthesis and characterization of poly(3-hydroxyalkanoates) from Brassica carinata oil with high content of erucic acid and from very long chain fatty acids.

Pseudomonas aeruginosa produced medium chain length poly(3-hydroxyalkanoates) (mcl-PHAs) when grown on substrates containing very long chain fatty acids (VLCFA, C>20). Looking for low cost carbon sources, we tested Brassica carinata oil (erucic acid content 35-48%) as an intact triglyceride containing VLCFA. Oleic (C18:1), erucic (C22:1), and nervonic (C24:1) acids were also employed for mcl-PHA production as model substrates. The polymers obtained were analyzed by GC of methanolyzed samples, GPC, 1H and 13C NMR, ESI MS of partially pyrolyzed samples, and DSC. The repeating units of such polymers were saturated and unsaturated, with a higher content of the latter in the case of the PHA obtained from B. carinata oil. Statistical analysis of the ion intensity in the ESI mass spectra showed that the PHAs from pure fatty acids are random copolymers, while the PHA from B. carinata oil is either a pure polymer or a mixture of polymers. Weight-average molecular weight varied from ca. 56,000 g/mol for the PHA from B. carinata oil and oleic acid, to about 120,000 g/mol for those from erucic and nervonic acids. The PHAs from erucic and nervonic acids were partially crystalline, with rubbery characteristics and a melting point (Tm) of 50°C, while the PHAs from oleic acid and from B. carinata oil afforded totally amorphous materials, with glass transition temperatures (Tg) of -52°C and -47°C, respectively.