Mineral stress response of Vitis cell wall transcriptome: identification, characterization and expression of genes from key families involved in wall biosynthesis and modification

Doutoramento em Engenharia Agronómica - Instituto Superior de Agronomia - UL

S-Alkylated/aralkylated 2-(1H-indol-3-yl-methyl)-1,3,4- oxadiazole-5-thiol derivatives. 1. Synthesis and characterization

Purpose: To synthesize and characterize S-alkylated/aralkylated 2-(1H-indol-3-ylmethyl)-1,3,4- oxadiazole-5-thiol derivatives. Methods: 2-(1H-indol-3-yl)acetic acid (1) was reacted with absolute ethanol and catalytic amount of sulfuric acid to form ethyl 2-(1H-indol-3-yl)acetate (2) which was transformed to 2-(1H-indol-3- yl)acetohydrazide (3) by refluxing with hydrazine hydrate in methanol. Ring closure reaction of 3 with carbon disulfide and ethanolic potassium hydroxide yielded 2-(1H-indol-3-ylmethyl)-1,3,4-oxadiazole-5- thiol (4) which was finally treated with alkyl/aralkyl halides (5a-u) in DMF and NaH to yield Salkylated/ aralkylated 2-(1H-indol-3-ylmethyl)-1,3,4-oxadiazole-5-thiols (6a-u). Structural elucidation was done by IR, 1H-NMR and EI-MS techniques Results: 2-(1H-indol-3-ylmethyl)-1,3,4-oxadiazole-5-thiol (4) was synthesized as the parent molecule and was characterized by IR and the spectrum showed peaks resonating at (cm-1) 2925 (Ar-H), 2250 (S-H ), 1593 (C=N ) and 1527 (Ar C=C ); 1H-NMR spectrum showed signals at δ 11.00 (s, 1H, NH-1ʹ), 7.49 ( br.d, J = 7.6 Hz, 1H, H-4\'), 7.37 (br.d, J = 8.0 Hz, 1H, H-7\'), 7.34 (br.s, 1H, H-2\'), 7.09 (t, J = 7.6 Hz, 1H, H-5\'), 7.00 (t, J = 7.6 Hz, 1H, H-6\') and 4.20 (s, 2H, CH2-10ʹ). EI-MS presented different fragments peaks at m/z 233 (C11H9N3OS)˙+ [M+2]+, 231 (C11H9N3OS)˙+ [M]+, 158 (C10H8NO)+, 156 (C10H8N2)˙+, 130 (C9H8N)+. The derivatives (6a-6u) were prepared and characterized accordingly. Conclusion: S-alkylated/aralkylated 2-(1H-indol-3-ylmethyl)-1,3,4-oxadiazole-5-thiols (6a-u) were successfully synthesized.

S-Alkylated/aralkylated 2-(1H-indol-3-yl-methyl)-1,3,4- oxadiazole-5-thiol derivatives. 2. Anti-bacterial, enzymeinhibitory and hemolytic activities

Purpose: To evaluate the antibacterial, enzyme-inhibitory and hemolytic activities of Salkylated/ aralkylated 2-(1H-indol-3-ylmethyl)-1,3,4-oxadiazole-5-thiol derivatives. Methods: Antibacterial activities of the compounds were evaluated using broth dilution method in 96 well plates. Enzyme inhibitory activities assays were investigated against α-glucosidase, butyrylcholinesterase (BchE) and lipoxygenase (LOX) using acarbose, eserine and baicalien as reference standards, respectively. A mixture of enzyme, test compound and the substrate was incubated and variation in absorbance noted before and after incubation. In tests for hemolytic activities, the compounds were incubated with red blood cells and variations in absorbance were used as indices their hemolytic activities. Results: The compounds were potent antibacterial agents. Five of them exhibited very good antibacterial potential similar to ciprofloxacin, and had minimum inhibitory concentrations (MIC) of at least 9.00 ± 4.12 μM against S. aureus, E.coli, and B. subtilis. One of the compounds had strong enzyme inhibitory potential against α-glucosidase, with IC50 of 17.11 ± 0.02 μg/mL which was better than that of standard acarbose (IC50 38.25 ± 0.12 μg/mL). Another compound had 1.5 % hemolytic activity. Conclusion: S-Alkylated/aralkylated 2-(1H-indol-3-ylmethyl)-1,3,4-oxadiazole-5-thiol deviratives with valuable antibacterial, anti-enzymatic and hemolytic activities have been successfully synthesized. These compounds may be useful in the development of pharmaceutical products.

Surface photochemistry: photodegradation study of pyrene adsorbed onto microcrystalline cellulose and silica

Ground-state diffuse reflectance, time resolved laser-induced luminescence, diffuse reflectance laser flash-photolysis transient absorption and chromatographic techniques were used to elucidate the photodegradation processes of pyrene adsorbed onto microcrystalline cellulose and silica. Ground-state diffuse reflectance showed that on both substrates low concentrations display absorption of pyrene monomers. At high concentrations spectral changes attributed to aggregate formation were observed. Laser induced fluorescence showed that pyrene onto microcrystalline cellulose mainly presents fluorescence from monomers, while for silica, excimer-like emission was observed from low surface loadings (greater than or equal to 0.5 mumol g(-1)). Transient absorption and photodegradation studies were performed at concentrations where mainly monomers exist. On silica, pyrene presents transient absorption from its radical cation. On microcrystalline cellulose both radical cation, radical anion and pyrene triplet-triplet absorption were detected. Irradiation followed by chromatographic analysis showed that pyrene decomposes on both substrates. For pyrene on microcrystalline cellulose 1-hydroxypyrene was the main identified photoproduct since in the absence of oxygen further oxidation of 1-hydroxypyrene was very slow. For pyrene on silica photodegradation was very efficient. Almost no 1-hydroxypyrene was detected since in the presence of oxygen it is quickly oxidized to other photooxidation products. On both substrates, pyrene radical cation is the intermediate leading to photoproducts and oxygen it is not involved in its formation.

A diffuse reflectance comparative study of benzil inclusion within microcrystalline cellulose and beta-cyclodextrin

Diffuse reflectance and laser-induced techniques were used to study photochemical and photophysical processes of benzil adsorbed on two solid powdered supports, microcrystalline cellulose and beta-cyclodextrin. In both substrates, a distribution of ground-state benzil conformers exists, largely dominated by skew conformations where the carbonyl groups are twisted one to the other. Room temperature phosphorescence was observed in air-equilibrated samples in both cases. The decay times vary greatly and the largest lifetime was obtained for benzil/beta-cyclodextrin, showing that this host's cavity accommodates benzil well, enhancing its room temperature phosphorescence. Triplet - triplet absorption of benzil entrapped in cellulose was detected and benzil ketyl radical formation also occurred. With benzil included into beta-cyclodextrin, and following laser excitation, benzoyl radicals were detected on the millisecond timescale. Product analysis and identification of laser-irradiated benzil samples in the two hosts clearly showed that the main degradation photoproducts were benzoic acid and benzaldehyde. The main differences were a larger benzoic acid/benzaldehyde ratio in the case of cellulose and the formation of benzyl alcohol in this support.

Photochemistry of 4-chlorophenol on cellulose and silica

The photochemistry of 4-chlorophenol (4-CP) was studied on silica and cellulose, using time-resolved diffuse reflectance techniques and product degradation analysis. The results have shown that the photochemistry of 4-CP depends on the support, on the concentration, and also on the sample preparation method. Transient absorption and photoproduct results can be understood by assuming the formation of the carbene 4-oxocyclohexa-2,5-dienylidene in both supports. On cellulose, at concentrations lower than 10 mumol g(-1), the carbene leads to the unsubstituted phenoxyl radical, and phenol is the main degradation product. At higher concentrations a new transient resulting from phenoxyl radicals coupling was also observed, and dihydroxybiphenyls are also formed. The reaction of the carbene with ground-state 4-CP was also detected through the formation of 5-chloro-2,4'-dihydroxybiphenyl. 4-Chlorophenoxyl radical and degradations products resulting from its coupling were also detected. Oxygen has little effect on the photochemistry of 4-CP on cellulose. On silica the transient benzoquinone O-oxide was formed in the presence of oxygen. Benzoquinone and hydroquinone are the main degradation products. In well-dried samples the formation of hydroquinone is reduced. At higher concentrations the same products as detected on cellulose were observed. 4-CP undergoes slow photochemical decomposition under solar radiation in both supports. The same main degradation products were observed in these conditions.

Exploring the role of the α-carboxyphosphonate moiety in the HIV-RT activity of α-carboxy nucleoside phosphonates

As α-carboxy nucleoside phosphonates (α-CNPs) have demonstrated a novel mode of action of HIV-1 reverse transcriptase inhibition, structurally related derivatives were synthesized, namely the malonate 2, the unsaturated and saturated bisphosphonates 3 and 4, respectively and the amide 5. These compounds were evaluated for inhibition of HIV-1 reverse transcriptase in cell-free assays. The importance of the α-carboxy phosphonoacetic acid moiety for achieving reverse transcriptase inhibition, without the need for prior phosphorylation, was confirmed. The malonate derivative 2 was less active by two orders of magnitude than the original α-CNPs, while displaying the same pattern of kinetic behavior; interestingly the activity resides in the “L”-enantiomer of 2, as seen with the earlier series of α-CNPs. A crystal structure with an RT/DNA complex at 2.95 Å resolution revealed the binding of the “L”-enantiomer of 2, at the polymerase active site with a weaker metal ion chelation environment compared to 1a (T-α-CNP) which may explain the lower inhibitory activity of 2.

Cellulose: A review as natural, modified and activated carbon adsorbent

Cellulose is a biodegradable, renewable, non-meltable polymer which is insoluble in most solvents due to hydrogen bonding and crystallinity. Natural cellulose shows lower adsorption capacity as compared to modified cellulose and its capacity can be enhanced by modification usually by chemicals. This review focuses on the utilization of cellulose as an adsorbent in natural/modified form or as a precursor for activated carbon (AC) for adsorbing substances from water. The literature revealed that cellulose can be a promising precursor for production of activated carbon with appreciable surface area (∼1300 m2 g−1) and total pore volume (∼0.6 cm3 g−1) and the surface area and pore volume varies with the cellulose content. Finally, the purpose of review is to report a few controversies and unresolved questions concerning the preparation/properties of ACs from cellulose and to make aware to readers that there is still considerable scope for future development, characterization and utilization of ACs from cellulose.

Cellulose: A review as natural, modified and activated carbon adsorbent

Cellulose is a biodegradable, renewable, non-meltable polymer which is insoluble in most solvents due to hydrogen bonding and crystallinity. Natural cellulose shows lower adsorption capacity as compared to modified cellulose and its capacity can be enhanced by modification usually by chemicals. This review focuses on the utilization of cellulose as an adsorbent in natural/modified form or as a precursor for activated carbon (AC) for adsorbing substances from water. The literature revealed that cellulose can be a promising precursor for production of activated carbon with appreciable surface area ( 1300 m2 g 1) and total pore volume ( 0.6 cm3 g 1) and the surface area and pore volume varies with the cellulose content. Finally, the purpose of review is to report a few controversies and unresolved questions concerning the preparation/properties of ACs from cellulose and to make aware to readers that there is still considerable scope for future development, characterization and utilization of ACs from cellulose.

Alginate / cellulose beads as supports for slowrelease of nutrient.

2016

Polyaniline-based materials and nanocomposites: from energy storage to sensing applications

The research work described in this thesis concerns materials for both energy storage and sensoristics applications. Firstly, the synthesis and characterization of magnetite (Fe3O4) functionalyzed with [3-(2-propynylcarbamate)propyl]triethoxysilane (PPTEOS) capable to reduce the gold precursor chloroauric acid (HAuCl4) without the need of additional reducing or stabilising agents is described. These nanoparticles were tested to improve performances of symmetric capacitors based on polyaniline and graphite foil. Energy storage applications were investigated also during six months stay at EPFL University of Lausanne where an investigation about different tailored catalysts for Oxygen Evolution Reaction in a particular Redox Flow Battery was carried out. For what concerns sensing applications, new materials based on cellulose modified with polyaniline and poly(2-acrylamido-2-methyl-1-propanesulfonic acid) (PAAMPSA) were synthesized, characterized and applied to monitor pressure, humidity, heart rate and lastly, bread fermentation in collaboration with the University of Fribourg and Zurich. The characterizations of all the materials investigated compriseed numerous techniques such as infrared attenuated total reflectance spectroscopy (IR-ATR), thermogravimetric analysis (TGA), scanning and transmission electron microscopy (SEM and TEM), alongside linear and cyclic voltammetry (LSV and CV), electrochemical impedance spectroscopy (EIS) and chronoamperometric analyses.

Towards the sustainable production of methyl methacrylate: A comparative analysis of industrial chemical production processes using life cycle assessment methodology

Increasing environmental awareness has been a significant driving force for innovations and process improvements in different sectors and the field of chemistry is not an outlier. Innovating around industrial chemical processes in line with current environmental responsibilities is however no mean feat. One of such hard to overhaul process is the production of methyl methacrylate (MMA) commonly produced via the acetone cyanohydrin (ACH) process developed back in the 1930s. Different alternatives to the ACH process have emerged over the years and the Alpha Lucite process has been particularly promising with a combined plant capacity of 370,000 metric tonnes in Singapore and Saudi Arabia. This study applied Life Cycle Assessment methodology to conduct a comparative analysis between the ACH and Lucite processes with the aim of ascertaining the effect of applying principles of green chemistry as a process improvement tool on overall environmental impacts. A further comparison was made between the Lucite process and a lab-scale process that is further improvement on the former, also based on green chemistry principles. Results showed that the Lucite process has higher impacts on resource scarcity and ecosystem health whereas the ACH process has higher impacts on human health. On the other hand, compared to the Lucite process the lab-scale process has higher impacts in both the ecosystem and human health categories with lower impacts only in the resource scarcity category. It was observed that the benefits of process improvements with green chemistry principles might not be apparent in some categories due to some limitations of the methodology. Process contribution analysis was also performed and it revealed that the contribution of energy is significant, therefore a sensitivity analysis with different energy scenarios was performed. An uncertainty analysis using Monte Carlo analysis was also performed to validate the consistency of the results in each of the comparisons.

Formulation of ionogel, hydrogel and films using a cellulose rich solid obtained from Pinus radiata

The purpose of my internship, carried out during my Erasmus period at the Complutense University of Madrid, was focused on the formulation of ionogels and hydrogels for the obtainment of films with high lignin content, and on their characterization measuring their antibacterial properties. For biomass formulation I used lignocellulosic biomass (Pinus Radiata) as raw material and ionic liquid as solvent. The two ionic liquids proposed were: 1-ethyl-3-methylimidazoliumdimethylphosphate [Emim][DMP] and 1-ethyl-3-methylimidazoliumdiethylphosphate [Emim][DEP]. The two-starting cellulose-rich solids were obtained from Pinus radiata wood that had been submitted to an organosolv process, to reduce its lignin content to fifteen (ORG15) and twenty per cent (ORG20). Having two ionic liquids and two solids available, the first phase of the project was devoted to the screening of both solids in both ionic liquids. Through this, it was possible to identify that only the [Emim][DMP] ionic liquid fulfils the purpose. It was also possible to discard the cellulose-rich solid ORG20 because its dissolution in the ionic liquid was not possible (after the time fixed) and, additionally, a Pinus radiata cellulose-rich solid bleached with hydrogen peroxide and containing ten per cent of lignin (ORG10B) was included in the screening. After screening, a total of five ionogels were subsequently formulated: two gels were formulated with the starting raw material ORG15 (with 1% and 1.75% cellulose, respectively) and three with ORG10B (with 1%, 1.75% and 3% cellulose, respectively). Five hydrogels were obtained from the ionogels. Rheological tests were performed on each ionogel and hydrogel. Finally, films were formulated from hydrogels and they were analysed by antibacterial testing to see if they could be applied as food packaging. In addition, antioxidant and properties such as opacity and transparency were also studied.