High-fat diet (HFD) is a major problem causing neuronal damage. However, the mechanisms and potential therapeutic targets have not been evaluated yet. Thymoquinone (TQ) could regulate oxidative stress and inflammatory process. Hence, the present study elucidated the significant role of TQ on oxidative stress, inflammation as well as morphological changes in the cerebellum of rats with HFD. The present study establishes the role of TQ on oxidative stress, anti-inflammatory process and morphological changes in the cerebellum following high-fat diet supplementation in rats. Rats were divided into three groups as (1) Control, (2) HFD (6 mg/day) for eight weeks and (3) HFD supplementation (4 weeks) followed TQ 300 mg/kg/day treated (4 weeks). After treatment, body weight and serum cholesterol profile were measured. Also, blood samples were collected to measure oxidative stress markers glutathione (GSH), superoxide dismutase (SOD) and inflammatory cytokines. Furthermore, neuronal morphological changes were also observed in the cerebellum of the rats. HFD rats show higher in body weight as compared with the control group. TQ treatment significantly (p<0.05) lowers the body weight with significant (p<0.05) reduction in cholesterol, triglycerides, high-density lipoprotein (HDL) and low-density lipoprotein (LDL). The anti-oxidative enzymes significantly reduced in HFD rats as compared with the control group. Surprisingly, treatment with TQ could improve the level of GSH and SOD. TQ treatment significantly (p<0.05) reduced the inflammatory markers as compared with HFD alone. The histological study revealed that neuronal damage was prevented in the cerebellum following treatment of TQ. TQ treatment minimizes neuronal damage as well as reduces inflammation and improves anti-oxidant enzymes.

The use of herbal products for the treatment, prevention and cure of diseases is one of the oldest human medicinal practices. In fact, the majority of the population in developing countries depend on ancestral plant knowledge for healthcare. However, there is still a gap between progress observed in clinical pharmacy and in the field of herbal and traditional medicine, remaining many natural products with biological activity to be identified.

Plectranthus species (Lamiaceae family) have a widespread ethnobotanical use and are often cited by its medicinal properties and application, particularly in folk medicine. They contain many antioxidant compounds and exhibit several effects (anti-inflammatory, antimicrobial and antifungal) which suggest that Plectranthus may be a promising genus for the discovery of medicinal compounds.

Thus, the isolation of secondary metabolite compounds from the Plectranthus spp. is important to validate scientifically the popular uses of these plants and also to find new sources of potentially economically important products or compounds which can be transformed into active pharmaceutical ingredients. Besides, the cytotoxicity evaluation of the plant extracts and their active ingredients are required for their effective and safe therapeutic use.

This work enumerates the compounds isolated to date from Plectranthus ecklonii Benth., extracts and their biological activities. The HPLC analysis presented is part of an ongoing project at CBIOS of identification, quantification and evaluation of the bioactive components (in particular, diterpenes and hydrocinnamic acids) in different species of Plectranthus.

Essential oils (EOs), which are complex biomolecules composed of volatile compounds, have emerged as a new strategy to deal with bacterial infections and as a valid alternative to synthetic drugs. Here, we report the modification of biodegradable wet-spun microfibers composed of cellulose acetate (CA) and polycaprolactone (PCL) with EOs, aiming at their localized, controlled release. Cinnamon leaf oil (CLO), cajeput oil (CJO), and clove oil (CO) were selected from a group of 20 EOs according to their minimal inhibitory concentration (MIC) against Staphylococcus aureus (<22.4 mg/mL) and Escherichia coli (<11.2 mg/mL). CA/PCL prepared at 10% and 14%wt in a 3/1 ratio in acetic acid and acetone were processed in the form of microfibers by wet-spinning at an extrusion rate of 0.5 mL/h directly into an ethanol coagulation bath. Microfibers were modified by immersion in ethanol solutions containing EOs at 2xMIC and ampicillin (control antibiotic). Incorporation was confirmed by UV-VIS, FTIR and TGA. After 72h, fibers contained ampicillin at MIC but only 14%, 66% and 76% of MIC for CLO, CO and CJO, respectively. Unloaded and loaded microfibers were characterized as uniform and homogeneous. Data showed that even at small amounts the EO-modified microfibers were effective against the tested bacteria. Considering the amount immobilized, CLO-containing fibers were deemed the most effective from the group, suggesting a superior affinity of the EOs active groups towards the CA/PCL matrix. These results indicate that CA/PCL microfibers loaded with EOs can be easily produced and applied in scaffolds for biomedical applications.

Cyclic ADP-ribose (cADPR) is a natural occurring metabolite of NAD⁺ capable of mobilizing Ca²⁺ ions from intracellular stores. It was firstly isolated from sea urchin eggs extract, but it was later established that it is also produced in many other mammalian cells, including pancreatic β-cells, T-lymphocytes, smooth and cardiac muscle cells and cerebellar neurons, acting as a Ca²⁺-mobilizing agent. For this activity, cADPR has been classified as a second messenger that, activating the ryanodine receptors of the sarcoplasmatic reticulum, is able to mobilize the calcium ions from intracellular stores. cADPR is involved in many physiological processes related to the variation of the Ca²⁺ concentration, such as the synaptic homeostasis in neurons, as well as fertilization and cellular proliferation. This cyclic nucleotide, characterized by a very labile glycosidic bond at the N1, is rapidly hydrolysed also in neutral aqueous solutions to the inactive ADP-ribose. Matsuda and co-workers were the first who synthesized new analogues of the cADPR in which the adenine base was replaced by a hypoxanthine ring. This kind of modification produced the cyclic inosine diphosphate ribose (cIDPR) which proved to be stable in hydrolytic physiological conditions and showed significant Ca²⁺ mobilizing activity. A lot of modifications regarding the northern and southern ribose, as well as the purine base of cADPR, have been proposed so far. In our laboratories we have synthesized several analogues of cIDPR. In particular, the analogue with the northern ribose replaced by a pentyl chain (cpIDP) showed interesting Ca²⁺ mobilizing activity on the neuronal PC12 cell line. Starting from these results, we report here the synthesis of a novel analogue , in which the “northern” ribose of cIDPR was replaced by a 2”,3”-dihydroxy pentyl chain.

Since the last few decades, proteins have emerged as the major class of pharmaceuticals with more than 200 protein-based products currently available in the market, of which 90% are used as therapeutics. The protein engineering market is bolstered by the need for drugs with improved efficiency, specificity, technological capabilities, rise of antibody based drugs and steady growth in the therapeutic market. Monoclonal antibodies (mAbs) is the fastest growing segment in the therapeutic market, though the other segments comprising non-mAb recombinant proteins like Insulin, Erythropoetin (EPO), Interferons (INF),Interleukins (ILs) and Somatotropin (hGH) are also in great demand for therapy.

In this paper we propose the generation of synthetic small and more sophisticated molecule structures that optimize the binding affinity to a target (ASYNT-GAN). To achieve this we leverage on three important achievements in A.I.: Attention, Deep Learning on Graphs and Generative Adversarial Networks. Similar to text generation based on parts of text we are able to generate a molecule architecture based on an existing target.

By adopting this approach, we propose a novel way of searching for existing compounds that are suitable candidates. Similar to question and answer Natural Language solutions we are able to find drugs with highest relevance to a target. We are able to identify substructures of the molecular structure that are the most suitable for binding.

In addition, we are proposing a novel way of generating the molecule in 3D space in such a way that the binding is optimized. We show that we are able to generate compound structures and protein structures that are optimised for binding to a target.

Chronic wounds (CW) have numerous entry ways for pathogen invasion and prosperity, damaging host tissue and hindering tissue remodelling. Essential oils (EOs) exert quick and efficient antimicrobial (AM) action, unlikely to induce bacterial resistance. Cajeput oil (CJO) has strong AM properties, namely against Staphylococcus aureus and Pseudomonas aeruginosa, as previously established by the team (DOI: 10.3390/antibiotics9060314). Chitosan (CS) is a natural and biodegradable cationic polysaccharide, widely known for its AM features. CS (100-300 kDa; DA of 9.6±1.4%) and PVA (72 kDa, 88% hydrolyzed) films (ratio 30/70; 9%wt) were prepared by solvent casting and phase inversion method (similarly as in DOI: 10.1002/app.48626). CJO was added to CS solution before blending it with PVA, with loading amount of 1 and 10 wt% in relation to total polymeric mass. Loaded films with 0.89 ± 0.05 and 1.14 ± 0.10 mm in thickness were obtained, respectively, 23 and 57% thicker than the unloaded films. Degree of swelling (%) and porosity also increased. Films chemical composition and thermal stability reinforce the achievement of loaded blended films. AM activity was evaluated through the agar diffusion assay and time kill kinetics, with the biomaterials incubated with S. aureus and P. aeruginosa. Thin inhibitory zones were observed with films placed in direct contact each of the bacterium. CS films alone showed an outstanding AM activity against both bacteria, with 1h being sufficient to eradicate all P. aeruginosa colony traces. Still, CS/PVA blended films carrying 1/10% CJO, having improved mechanical properties than CS films alone, resulted in 2/3 (S. aureus) or 3/4 (P. aeruginosa) log reduction in 24h of contact. This study is a first proof of concept that CJO can be dispersed into CS/PVA films and show bactericidal effects, notably when combined with CS, and particularly against P. aeruginosa, this way opening new avenues for CW therapeutics.

Olive tree farming practices and the olive oil industry generate huge amounts of wastes every year. One of the main components of these agroindustrial residues is the extractive fraction and interestingly it contains non-structural components, including bioactive compounds. Particularly, exhausted olive pomace (EOP) is the final residue resulting from the industrial extraction of olive-pomace oil with hexane and the extractives account for more than 40% of its chemical composition.

In this work, different extraction methods, including green technologies, have been compared to evaluate the extraction of antioxidants from EOP. Hydrothermal extraction using medium (85 ºC) and high thermal conditions (200 ºC) were carried out. Aqueous accelerated extraction was also assessed at different temperatures (55 ºC and 190 ºC). Likewise, the effect of organosolv extraction (50% ethanol at 200 ºC), alkaline hydrolysis at 55 ºC and different NaCl concentrations (3-9% w/v) were also tested. Moreover, the influence of the time extraction (up to 120 min) and the solid loading (up to 25%) were evaluated in the aforementioned experiments.

The extracts obtained were characterized regarding the content of total phenols by the Folin-Ciocalteu method. In general, the solid loading showed a negative influence on the phenolic compounds extraction with all techniques used. Using water as extractive agent, the values ranged between 26.8 and 50.8 mg gallic acid equivalents (GAE)/g EOP, depending on the conditions. In particular, when the accelerated solvent extraction was used, the temperature showed a clear positive effect on the phenolic compound extraction and the extration time was considerably reduced. It enables the obtention of 40.6 mg GAE/g EOP at 190 ºC in 10 min. Nevertheless, the best results were found using organosolv extraction at 200 ºC (82.1 mg/g GAE/g EOP). After characterization, hydroxytyrosol was found to be one of the potential active compounds in EOP.

Docosahexaenoic acid (DHA) is the predominant omega-3 fatty acid in the brain and has been shown to have neuroprotective effects in Parkinson’s disease, but the underlying mechanism has not been fully elucidated. DHA is metabolized to DHA epoxides (EDPs) and hydroxides by cytochrome P450s (P450s), and EDPs are further hydroxylated to the corresponding diols (DHDPs) by soluble epoxide hydrolase (sEH). In the present study, we investigated the roles of these DHA metabolites in the beneficial effects of DHA supplementation on a rotenone-induced rat model of Parkinson’s disease. Metabolite analysis by LC-MS revealed that CYP2A1, 2C11, 2C13, 2C23, and 2E1 contributed to the formation of EDPs, and these P450s and sEH were expressed in the rat brain. We found that DHA supplementation in rats improved the motor dysfunction induced by rotenone. In addition, DHA reversed the decrease in tyrosine hydroxylase and the increase in lipid peroxidation generated by rotenone in the striatum. DHA supplementation also induced mRNA expression of antioxidant genes and Nrf2 protein expression in the striatum. However, these effects of DHA supplementation were eliminated by cosupplementation with the sEH inhibitor TPPU. DHA metabolites, including EDPs and DHDPs, in the rat brain were analyzed by LC-MS/MS. Supplementation with DHA increased the amount of DHA-diols (DHDPs) in the rat brain, while the amount of EDPs was not significantly increased. In addition, TPPU suppressed the increase in DHDPs and increased EDPs in the brain. These results indicate that DHDPs are important in the beneficial effects of DHA supplementation. The addition of DHDPs to PC12 cells increased the mRNA levels of antioxidant genes along with Nrf2 induction. This study suggests that DHA metabolites—DHDPs generated by P450s and sEH—have an important role in improving rotenone-induced Parkinson’s disease.

Glycolysis inhibitors are currently considered as potential anticancer agents due to the fact that significant changes occur in tumor cells, known as metabolic reprogramming. So, in particular, changes in carbohydrate metabolism are called the Warburg effect. However, the use of carbohydrate metabolism inhibitors is severely limited due to the fact that they have pronounced systemic toxicity. But there is a possibility of using metabolic inhibitors, subject to targeted delivery. One of the key such approaches is the use of liposomal forms of drugs.

We have carried out studies comparing the systemic toxicity of the glycolysis inhibitor iodoacetate and its liposomal form.

Using HPLC, the optimal conditions for obtaining iodoacetate encapsulated in liposomes and its final concentration in them were established. Thanks to biochemical blood analysis, it was found that iodoacetate disrupts carbohydrate metabolism and has a pronounced cardiotoxic effect, but does not have hepatotoxic manifestations. Based on the indicators of biochemical markers of the toxic effect of iodoacetate encapsulated in liposomes, it was found that when administered in a course for two weeks, iodoacetate in this form has neither cardiotoxic nor any other negative effects on target organs, that is, it is safe.

Thus, iodoacetate in the form of liposomes has great potential as an anticancer agent, especially in the case of polypharmacy.

Olive leaves are a source of valuable compounds, including phenolic compounds and triterpenic acids, with applications in the phytopharmacy sector, among others. Nevertheless, there are untapped olive resources that can give new clues for the discovery of natural bioactive compounds. Therefore, in this study the antioxidant composition of olive mill leaves (OML), extracted olive pomace (EOP) and a new byproduct, the residual fraction from olive pit cleaning (RFOPC), was characterized and compared to olive leaves, which have been extensively studied as a source of bioactive compounds. The chemical characterization showed that all these byproducts contain a high amount of extractives; in all cases it was higher than 26%. This is interesting since the extractive fraction contains non-structural compounds, such as phenolic compounds and triterpenic acids. Then, ultrasound-assisted extraction was applied to recover phenolic compounds from this fraction using an aqueous-ethanolic solution. All the extracts showed antioxidant properties and the total phenolic content ranged from 69 (RFOPC) to 140 (EOP) g of gallic acid equivalents/kg dry weight. The profile obtained by liquid chromatography-mass spectrometry was different, suggesting some chemical differences in the phenolic composition. In fact, while olive leafy biomass and EOP contained hydroxytyrosol derivatives, RFOPC was a source of novel trilignols. Other bioactive compounds were found in the extracts, including triterpenic acids, which could give an extra value due to their cardioprotective properties.