Asymmetric Syntheses of Analogs of Kainic Acid

Kainic acid has been used for nearly 50 years as a tool in neuroscience due to its pronounced neuroexcitatory properties. However, the significant price increase of kainic acid resulting from the disruption in the supply from its natural source, the alga Digenea Simplex, as well as inefficient synthesis of kainic acid, call for the exploration of functional mimics of kainic acid that can be synthesized in a simpler way. Aza kainoids analog could be one of them. The unsubstituted aza analog of kainoids has demonstrates its ability as an ionotropic glutamate receptor agonist and showed affinity in the chloride dependent glutamate (GluCl) binding site. This opened a question of the importance of the presence of one nitrogen or both nitrogens in the aza kainoid analogs for binding to glutamate receptors. Therefore, two different pyrrolidine analogs of kainic acid, trans-4-(carboxymethyl)pyrrolidine-3-carboxylic acid and trans-2-carboxy-3-pyrrolidineacetic acid, were synthesized through multi-step sequences. The lack of the affinity of both pyrrolidine analogs in GluCl binding site indicated that both nitrogens in aza kainoid analogs are involved in hydrogen bonding with receptors, significantly enhancing their affinity in GluCl binding site. Another potential functional mimic of kainic acid is isoxazolidine analogs of kainoids whose skeleton can be constituted directly via a 1, 3 dipolar cycloaddition as the key step. The difficulty in synthesizing N-unsubstituted isoxazolidines when applying such common protecting groups as alkyl, phenyl and benzyl groups, and the requirement of a desired enantioselectivity due to the three chiral ceneters in kainic acid, pose great challenges. Hence, several different protected nitrones were studied to establish that diphenylmethine nitrone may be a good candidate as the dipole in that the generated isoxazolidines can be deprotected in mild conditions with high yields. Our investigations also indicated that the exo/endo selectivity of the 1, 3 dipolar cycloaddition can be controlled by Lewis acids, and that the application of a directing group in dipolarophiles can accomplish a satisfied enantioselectivity. Those results demonstrated the synthesis of isoxazoldines analogs of kainic acid is very promising.

Immobilized Microalgae for Nutrient Recovery from Source Separated Urine

Shortages in supply of nutrients and freshwater for a growing human population are critical global issues. Traditional centralized sewage treatment can prevent eutrophication and provide sanitation, but is neither efficient nor sustainable in terms of water and resources. Source separation of household wastes, combined with decentralized resource recovery, presents a novel approach to solve these issues. Urine contains within 1 % of household waste water up to 80 % of the nitrogen (N) and 50 % of the phosphorus (P). Since microalgae are efficient at nutrient uptake, growing these organisms in urine might be a promising technology to concomitantly clean urine and produce valuable biomass containing the major plant nutrients. While state-of-the-art suspension systems for algal cultivation have mayor shortcomings in their application, immobilized cultivation on Porous Substrate Photobioreactors (PSBRs) might be a feasible alternative. The aim of this study was to develop a robust process for nutrient recovery from minimally diluted human urine using microalgae on PSBRs. The green alga Desmodesmus abundans strain CCAC 3496 was chosen for its good growth, after screening 96 algal strains derived from urine-specific isolations and culture collections. Treatment of urine, 1:1 diluted with tap water and without addition of nutrients, was performed at a light intensity of 600 μmol photons m-2 s-1 with 2.5 % CO2 and at pH 6.5. A growth rate of 7.2 g dry weight m-² day-1 and removal efficiencies for N and P of 13.1 % and 94.1 %, respectively, were determined. Pre-treatment of urine with activated carbon was found to eliminate possible detrimental effects of pharmaceuticals. These results provide a basis for further development of the technology at pilot-scale. If found to be safe in terms human and environmental health, the biomass produced from three persons could provide the P for annual production of 31 kg wheat grain and 16 kg soybean, covering the caloric demand in food for almost one month of the year for such a household. In combination with other technologies, PSBRs could thus be applied in a decentralized resource recovery system, contributing to locally close the link between sanitation and food production.

Extracción, identificación y prueba microbiológica del agar extraído de Gracilaria fortissima dawson (rhodophyta, gigartinales, gracilariaceae) (ING)

The red seaweed Gracilaria fortissima colected in the caribean coast of Costa Rica, was studied for the extraction, identification and microbiological performance of the agar contend of this plant, as well as the mineral contend. The research was done focused on the agar quality included pH, % extraction, geling point, fusion point and gel strength, as well as infrared analysis and the performance as a microbiological culture of bacteria and fungus and compared with commercial agar from BDH chemicals.

Composizione biochimica di diverse specie microalgali e prove di feeding su Ruditapes philippinarum

L'utilizzo di diete a base di microalghe per l'allevamento di novellame di molluschi bivalvi sta ottenendo sempre maggiore interesse data la crescente domanda di mercato nel settore dell'acquacoltura. Le dimensioni cellulari, la composizione biochimica e la digeribilità della parete cellulare delle microalghe sono fattori fondamentali per la loro applicabilità nell'alimentazione dei bivalvi; anche il contenuto di acidi grassi essenziali quali EPA e DHA ha un ruolo rilevante poiché influenza il tasso di crescita e di sviluppo degli animali. Nel presente studio sono stati utilizzati due ceppi autoctoni della Sacca di Goro del genere Thalassiosira, Pheaodactylum tricornutum, l'alga verde Tetraselmis chui e l'aptofita T-Iso, quest’ultima ampiamente utilizzata nell'alimentazione di diverse specie di molluschi bivalvi. Gli esperimenti condotti hanno permesso di (1) caratterizzare la crescita e la composizione biochimica delle 5 specie microalgali e (2) di analizzare l'applicabilità delle stesse tramite prove di alimentazione su larve e novellame di Ruditapes philippinarum, al fine di ottimizzarne la produzione. I risultati migliori, per numero di sopravvissuti e taglia raggiunta, sono stati ottenuti con larve alimentate con dieta plurispecifica a base di T-Iso e Thalassiosira ceppo A in rapporto 2:1; Thalassiosira ceppo A e P. tricornutum, in diete monospecifiche, sono state identificate come le più appetibili e assimilabili dal novellame, il quale mostra buoni valori del tasso massimo di clearance (2,17±0,25 e 2,27±0,15 L h-1, rispettivamente) e di efficienza di assorbimento del cibo (67 e 46%). Si è trattato di uno studio preliminare volto a verificare eventuali miglioramenti legati all’utilizzo di diete diversificate rispetto a quelle standardizzate di solito impiego; la ricerca di nuove specie algali e ulteriori analisi sulle attuali specie disponibili e sulla loro applicabilità potranno essere utili alla crescita sostenibile del settore dell’acquacoltura.

S-nitrosylation homeostasis in plants: key enzymes and regulatory pathways

The nitrosylated form of glutathione (GSNO) has been acknowledged to be the most important nitrosylating agent of the plant cell, and the tuning of its intracellular concentration is of pivotal importance for photosynthetic life. During my time as a PhD student, I focused my attention on the enzymatic systems involved in the degradation of GSNO. Hence, we decided to study the structural and catalytic features of alcohol dehydrogenases (GSNOR and ADH1) from the model land plant Arabidopsis thaliana (At). These enzymes displayed a very similar 3D structure except for their active site which might explain the extreme catalytic specialization of the two enzymes. They share NAD(H) as a cofactor, but only AtGSNOR was able to catalyze the reduction of GSNO whilst being ineffective in oxidizing ethanol. Moreover, our study on the enzyme from the unicellular green alga Chlamydomonas reinhardtii (Cr) revealed how this S-nitrosoglutathione reductase (GSNOR) specifically use NADH to catalyze GSNO reduction and how its activity responds to thiol-based post-translational modifications. Contextually, the presence of NADPH-dependent GSNO-degrading systems in algal protein extract was highlighted and resulted to be relatively efficient in this model organism. This activity could be ascribed to several proteins whose contribution has not been defined yet. Intriguingly, protein extract from GSNOR null mutants of Arabidopsis displayed an increased NADPH-dependent ability to degrade GSNO and our quantitative proteome profiling on the gsnor mutant revealed the overexpression of two class 4 aldo-keto reductases (AKR), specifically AtAKR4C8 and AtAKR4C9. Later, all four class 4 AKRs showed to possess a NADPH-dependent GSNO-degrading activity. Finally, we initiated a preliminary analysis to determine the kinetic parameters of several plant proteins, including GSNOR, AKR4Cs, and thioredoxins. These data suggested GSNOR to be the most effective enzyme in catalyzing GSNO reduction because of its extremely high catalytic proficiency compared to NADPH-dependent systems.

Portofino Seaweed Garden: a local citizen science case study on Ericaria amentacea (C.Agardh) Molinari & Guiry, 2020 restoration

Ericaria amentacea is an endemic alga of the Mediterranean Sea that lives in the littoral rocky fringe. The species is sensitive to environmental changes, so it’s used to assess the water ecological quality. Nevertheless, E. amentacea is deeply impacted by coastal development which cause rapid regression despite data on its distribution and healthy status are still limited. Moreover, it’s little known by people outside the scientific community. In this context, Portofino Seaweed Garden was born, a conservation and citizen science project aim to involve marine outdoor enthusiasts in protecting and restoring E. amentacea, creating a submerged garden. Restoration measures have been encouraged by EU regulations. Here, using citizen science, 1) I evaluated the spatial variability of E. amentacea abundance along the central-eastern Ligurian coast, to evaluate its status and choose donor and restoration sites. 2) I carried out an E. amentacea restoration (with outplanting lab-cultured embryos on 50 clay tiles). 3) I assessed the community involvement and education of volunteers. Simple protocol was created to train them on the monitoring. Unprecedented E. amentacea reproductive mismatch affected the restoration performance, probably caused by marine heat wave that hit the Mediterranean in summer 2022. After fertile apices collection in Pontetto (GE) and during laboratory phase, gametes spawned on the discs didn’t settle as expected. Only 16 tiles showed juveniles and they were outplanted at Punta Castello (C zone of Portofino MPA). Unfortunately, they didn’t survive in the field due to an interplay of physical and biological factors. From citizen science point of view, the project demonstrated positive outcomes of collaborations between people and scientists by involving more than 100 participants. Citizen scientists became specialize in the protocol providing quality data for E. amentacea conservation. Current results suggest that outplanting should be further tested.