The elongation factor 1-alpha as storage reserve and environmental sensor in Nicotiana tabacum L. seeds.

Given their critical role in plant reproduction and survival, seeds demand meticulous regulatory mechanisms to effectively store and mobilize reserves. Within seeds, the condition of storage reserves heavily depends on environmental stimuli and hormonal activation. Unlike non-protein reserves that commonly employ dedicated regulatory proteins for signaling, proteinaceous reserves may show a unique form of 'self-regulation', amplifying efficiency and precision in this process. Proteins rely on stability to carry out their functions. However, in specific physiological contexts, particularly in seed germination, protein instability becomes essential, fulfilling roles from signaling to regulation. In this study, the elongation factor 1-alpha has been identified as a main proteinaceous reserve in Nicotiana tabacum L. seeds and showed peculiar changes in stability based on tested chemical and physical conditions. A detailed biochemical analysis followed these steps to enhance our understanding of these protein attributes. The protein varied its behavior under different conditions of pH, temperature, and salt concentration, exhibiting shifts within physiological ranges. Notably, distinct solubility transitions were observed, with the elongation factor 1-alpha becoming insoluble upon reaching specific thresholds determined by the tested chemical and physical conditions. The findings are discussed within the context of seed signaling in response to environmental conditions during the key transitions of dormancy and germination.

Phytochemical Analysis and In Vitro Antileukemic Activity of Alkaloid-Enriched Extracts from Vinca sardoa (Stearn) Pignatti.

Vinca sardoa (Stearn) Pignatti, known as Sardinian periwinkle, is widely diffused in Sardinia (Italy). This species contains indole alkaloids, which are known to have a great variety of biological activities. This study investigated the antileukemic activity against a B lymphoblast cell line (SUP-B15) of V. sardoa alkaloid-rich extracts obtained from plants grown in Italy, in Iglesias (Sardinia) and Rome (Latium). All the extracts showed a good capacity to induce reductions in cell proliferation of up to 50% at the tested concentrations (1-15 µg/mL). Moreover, none of the extracts showed cytotoxicity on normal cells at all the studied concentrations.

Advances in the Phytochemical Characterisation and Bioactivities of Salvia aurea L. Essential Oil.

The Salvia L. genus (Lamiaceae) is largely used in the pharmaceutical and food industry. Several species of biological relevance are extensively employed in traditional medicine, including Salvia aurea L. (syn. S. africana-lutea L.), which is used as a traditional skin disinfectant and in wounds as a healing remedy; nevertheless, these properties have not been validated yet. The aim of the present study is to characterise S. aurea essential oil (EO), unveiling its chemical composition and validating its biological properties. The EO was obtained by hydrodistillation and subsequently analysed by GC-FID and GC-MS. Different biological activities were assessed: the antifungal effect on dermatophytes and yeasts and the anti-inflammatory potential by evaluating nitric oxide (NO) production and COX-2 and iNOS protein levels. Wound-healing properties were assessed using the scratch-healing test, and the anti-aging capacity was estimated through the senescence-associated beta-galactosidase activity. S. aurea EO is mainly characterised by 1,8-cineole (16.7%), ß-pinene (11.9%), cis-thujone (10.5%), camphor (9.5%), and (E)-caryophyllene (9.3%). The results showed an effective inhibition of the growth of dermatophytes. Furthermore, it significantly reduced protein levels of iNOS/COX-2 and simultaneously NO release. Additionally, the EO exhibited anti-senescence potential and enhanced wound healing. Overall, this study highlights the remarkable pharmacological properties of Salvia aurea EO, which should be further explored in order to develop innovative, sustainable, and environmentally friendly skin products.

Unveiling the Chemical Composition and Biological Properties of Salvia cacaliifolia Benth. Essential Oil.

Salvia is widely recognized for its therapeutic potential. However, the biological relevance of some species remains unknown, namely Salvia cacaliifolia Benth. Therefore, the aim of this study is to unveil the chemical composition and relevant properties to its essential oil (EO). The EO was characterized by GC and GC-MS and its antifungal effect was evaluated according to the CLSI guidelines on dermatophytes and yeasts. The anti-inflammatory potential was assessed on lipopolysaccharide-stimulated macrophages, by assessing the production of nitric oxide (NO) and the effect on the protein levels of two key pro-inflammatory enzymes, iNOS and COX-2 by western blot analysis. Wound healing capacity was determined using the scratch wound healing assay, and the anti-aging potential was assessed by evaluating the senescence marker ß-galactosidase. The EO was mainly characterized by γ-curcumene, ß-bisabolene, bicyclogermacrene and curzerenone. It is effective in inhibiting the growth of dermatophytes and C. neoformans. The EO significantly decreased iNOS and COX-2 protein levels and concomitantly reduced NO release. Additionally, it demonstrated anti-senescence potential and promoted wound healing. Overall, this study highlights relevant pharmacological properties of the EO of Salvia cacaliifolia, which should be further explored envisaging the development of sustainable, innovative, and environmentally friendly skin products.

Ethnopharmacobotany and Diversity of Mediterranean Endemic Plants in Marmilla Subregion, Sardinia, Italy.

Human populations in various regions across the world exploit the medicinal properties of plants to treat a wide variety of diseases. Areas with both high rates of endemic taxa and persisting traditional uses of the local botanical resources are key sites for the investigation of Traditional Botanical Knowledge (TBK). Commonly, in these areas, information regarding the medicinal properties of native plants has been transmitted orally from generation to generation, however, a rapid decline in this knowledge has been observed, which can be attributed to socio-economic changes in recent years. The Mediterranean basin is one such site, where human history is intimately entwined with nature. The unique geographical situation and unrivaled environmental heterogeneity of the area, have allowed both the development of diverse civilizations as well as providing the basis for the evolution of extraordinary biodiversity. The Mediterranean basin can therefore be considered a global hotspot of endemic vascular plants, and of traditional knowledge of medicinal and aromatic species. This study researches the historical subregion of Marmilla (central-southern Sardinia, Italy), which was chosen because of its specific cultural and demographic characteristics: i.e., prolonged isolation and extreme longevity of the inhabitants of the area. Semi-structured interviews were conducted with 145 people from the region, and 137 medicinal plants belonging to 62 families were identified, of which around 57,3% were taxa exclusive to the Mediterranean Basin. Findings showed that the most used parts of the plant were the leaves (49%), while as far as preparations are concerned, decoction (50%) was the most used to prepare medicinal formulations, making this the highest number of medico-botanical taxa reported in a study carried out in Sardinia using a similar methodology. In addition, this study contributes towards preventing the loss of TBK by documenting the medicinal traditions, passed down orally for centuries, in the words of the participants, shedding new light on the traditional knowledge of the inhabitants of the island. The findings lay the foundations for future applied studies in the fields of phytotherapy and phytochemical investigation.

Isolation and characterization of a main porin from the outer membrane of Salinibacter ruber.

Salinibacter ruber is an extremophilic bacterium able to grow in high-salts environments, such as saltern crystallizer ponds. This halophilic bacterium is red-pigmented due to the production of several carotenoids and their derivatives. Two of these pigment molecules, salinixanthin and retinal, are reported to be essential cofactors of the xanthorhodopsin, a light-driven proton pump unique to this bacterium. Here, we isolate and characterize an outer membrane porin-like protein that retains salinixanthin. The characterization by mass spectrometry identified an unknown protein whose structure, predicted by AlphaFold, consists of a 8 strands beta-barrel transmembrane organization typical of porins. The protein is found to be part of a functional network clearly involved in the outer membrane trafficking. Cryo-EM micrographs showed the shape and dimensions of a particle comparable with the ones of the predicted structure. Functional implications, with respect to the high representativity of this protein in the outer membrane fraction, are discussed considering its possible role in primary functions such as the nutrients uptake and the homeostatic balance. Finally, also a possible involvement in balancing the charge perturbation associated with the xanthorhodopsin and ATP synthase activities is considered.

Fatty acid and triacylglycerol composition of seed and pericarp oils of the medicinal crop Withania somnifera (L.) Dunal cultivated in Sardinia (Italy).

We studied the chemical composition and the nutritional properties of fixed oils extracted from seeds and pericarps of the medicinal plant Whitania somnifera. W. somnifera wild germplasm was collected in Sardinia (Italy) and cultivated in Campidano subregion (Southern Sardinia). The main fatty acids of seed oil were 18:2 n-6 (53%, 73.6 mg/g of dry weight), 18:1 n-9 (21%, 29.3 mg/g of dry weight), 16:0 (19%), and 18:0 (4%). Trilinolein and dilinolein derivatives represented the main seed oil triacylglycerols. Linoleic (40%, 67.7 mg/g of dry weight), oleic (29%, 48.7 mg/g of dry weight), palmitic (22%), stearic (4%) and α-linolenic (3%), were the main fatty acids in pericarp oil, with higher total level of monounsaturated fatty acids than seed oil, while monolinolein and dilinolein derivatives represented the main triacylglycerols. The results of this study qualify Sardinian W. somnifera berries as an interesting source of oils with nutritional properties.

Chemical characterization and bioactivity of the essential oil from Santolina insularis, a Sardinian endemism.

Santolina insularis is a Sardinian endemism that is widely used in traditional medicine. The essential oil was obtained with a yield of 2.7% and is mainly characterized by ß-phellandrene (22.6%), myrcene (11.4%) and artemisia ketone (7.6%). The oil significantly reduced NO production without affecting macrophages viability. In addition, it substantially inhibited the expression of two key pro-inflammatory enzymes, iNOS and COX-2 (71% and 25% at 0.54 mg/mL). Furthermore, the oil had a promising antifungal activity being Cryptococcus neoformans (MIC = 0.13 mg/mL) and the majority of dermatophytes (MIC = 0.13 mg/mL) the most susceptible fungi. Moreover, it significantly decreased the yeast-to-hypha transition (80% inhibition at 0.13 mg/mL) on C. albicans long before showing inhibitory effects. Overall, these results show that S. insularis could be applied in the treatment of fungal infections and associated inflammatory response.

Chemical composition and biological activity of essential oil of Teucrium scordium L. subsp. scordioides (Schreb.) Arcang. (Lamiaceae) from Sardinia Island (Italy).

The aim of this study is to demonstrate the antifungal, anti-inflammatory and anti-migratory potential of the essential oil of Teucrium scordium subsp. scordioides (Schreb.) Arcang, a plant widely used in traditional medicine in Sardinia. The oil was rich in germacrene D (25.1%), δ-cadinene (12.9%) and alloaromadendrene (11.3%). The yeast Cryptococcus neoformans and the dermatophytes Trichophyton rubrum, T. mentagrophytes var. interdigitale and Epidermophyton floccosum were the most susceptible fungi to the action of the oil. In lipopolysaccharide (LPS)-stimulated macrophages, the oil was able to decrease nitric oxide production by ca. 30% at 1.25 µL/mL, without affecting cell viability. In the scratch wound assay, it allowed for ca. 36% of wound closure after 18 h, thus showing anti-migratory properties. Overall, this study highlights the potential of this species to mitigate fungal infections associated with an inflammatory response. Furthermore, we also reported for the first time its anti-migratory capacity, thus suggesting anticancer properties.

Characterization of Essential Oils from Different Taxa Belonging to the Genus Teucrium in Sardinia Island, Italy.

The genus Teucrium L. (Lamiaceae) is a genus growing in mild climate zones, particularly in the Mediterranean Basin and Central Asia. It is represented by 11 taxa in Sardinia (Italy), living commonly in sunny habitats. In this study, the following eight Sardinian Teucrium taxa were selected, and the essential oils (EOs), obtained by stem distillation, were analyzed by GC-FID and GC-MS: T. capitatum subsp. capitatum, T. chamaedrys subsp. chamaedrys, T. flavum subsp. glaucum, T. marum, T. massiliense, T. scordium subsp. scordioides, T. scorodonia, and T. subspinosum. The comprehensive analyses led to the identification of 87 constituents representing the majority of the volatile compounds. Significant differences, both qualitative and quantitative, were observed between the taxa. Overall, monoterpenes and sesquiterpenes characterized all Teucrium EOs: T. capitatum subsp. capitatum and T. flavum subsp. glaucum revealed the highest content of monoterpene hydrocarbons, while in the other Teucrium taxa sesquiterpene hydrocarbons prevailed. Worthy of note, diterpenes were found only in T. marum and T. subspinosum, whereas T. massiliense was rich in non-terpenic oxygenated compounds. To the best of our knowledge, this is the first comprehensive report on the chemical composition of EOs obtained from Sardinian Teucrium species.

Antifungal activity and chemical composition of the essential oil from the aerial parts of two new Teucrium capitatum L. chemotypes from Sardinia Island, Italy.

The chemical composition of two populations of Teucrium capitatum L. one from the coastline and the other one from the mountainous area of Sardinia (Italy) was assessed. Two chemotypes were identified: limonene/α-pinene/(E)-nerolidol chemotype predominant in the coastline, and limonene/α-pinene/α-trans-bergamotene/humulene epoxide II chemotype common in plants growing in the mountainous area. In addition, our results showed that the sample growing in the coastline had a more promising antifungal activity. Furthermore, this sample was highly effective in inhibiting C. albicans germ tube formation, at doses well below its MIC. Overall, this study shows that the edaphoclimatic characteristics play an important role on the essential oil composition and biological activity of Teucrium capitatum L.

So Uncommon and so Singular, but Underexplored: An Updated Overview on Ethnobotanical Uses, Biological Properties and Phytoconstituents of Sardinian Endemic Plants.

The last decades have recorded an increase of plant-based drug discovery processes. Indeed, natural products possess a superior chemical diversity as compared to synthetic ones, leading to a renewal in searching for new therapeutic agents from the plant kingdom. In particular, since the structural variety of natural compounds reflects the biodiversity of their source organisms, regions of the world with high biodiversity and endemism deserve particular interest. In this context, Sardinia Island (Italy), with 290 endemic taxa (12% of the total flora), is expected to provide unique and structurally diverse phytochemicals for drug development. Several research groups built up a large program dedicated to the analysis of Sardinian endemic species, highlighting their peculiar features, both in respect of phytochemical and biological profiles. On this basis, the aim of this review is to provide an up-to-date and comprehensive overview on ethnobotanical uses, biological properties and phytoconstituents of Sardinian endemic plants in order to support their beneficial potential and to provide input for future investigations. We documented 152 articles published from 1965 to June 2020 in which a broad range of biological activities and the identification of previously undescribed compounds have been reported, supporting their great value as sources of therapeutic agents.

Intra-specific variation in the little-known Mediterranean plant Ptilostemon casabonae (L.) Greuter analysed through phytochemical and biomolecular markers.

Ptilostemon casabonae (L.) Greuter is a Mediterranean endemism traditionally used for its health-giving properties. Little is known about this species, therefore this study provides additional information about the phytochemical and biomolecular patterns of this plant, to have a combined fingerprint as a taxonomic tool. Several P. casabonae specimens were therefore collected from three different sites, two from Sardinia (Italy) and one from Corsica and the hydroalcoholic extracts of their aerial parts were investigated through HPLC-PDA-MS/MS analysis to study the phenolic composition. Quercetin, luteolin, kaempferol, apigenin and diosmetin O-glycosides, and caffeoylquinic acid derivatives were found as main components. Samples from the three sites showed similar phenolic profiles, although statistical analyses highlighted some quantitative differences for several compounds. The biomolecular analysis included amplification and sequencing of ITS, 5S-rRNA-NTS and psbA regions. No difference was found in the nucleotides among the P. casabonae samples from different geographical origins; however, a comparison with other Ptilostemon species sequences from Genbank, revealed an interspecific variability of ITS and psbA regions. The combination of the results of the phytochemical and biomolecular studies provide information on P. casabonae useful to depict this little-known plant, which can also be applied for future investigations and to obtain a fingerprint of it. Moreover, the stability of the phenolic profile within the species affords to identify a set of specialised metabolites useful for its chemotaxonomic characterization. At the same time, the stability of the biomolecular profile of P. casabonae, and the identification of sequences specific for this species, enables to identify useful biomolecular markers to distinguish it unequivocally.

Chemical and biomolecular analyses to discriminate three taxa of Pistacia genus from Sardinia Island (Italy) and their antifungal activity.

This work reports the results and the comparison concerning the chemical and biomolecular analyses and the antifungal activity of three wild Pistacia species (Anacardiaceae) from Sardinia. Volatile oils from leaves and twigs of Pistacia x saportae, Pistacia lentiscus and Pistacia terebinthus were characterised using GC-FID and GC-MS techniques and tested against some fungal strains. Two DNA nuclear regions (ITS and 5S-rRNA-NTS) were amplified through PCR technique and sequenced. The three **Pistacia have similar chemical profile, although there are some important quantitative differences. The analysis of ITS and 5S-rRNA-NTS regions, reveals a species-specific nucleotide variation among the three **taxa. This method could emerge as a powerful tool for the species identification, especially because the discrimination of these three **taxa appears difficult for non-expert botanists. Concerning the antifungal activity, P. lentiscus and P. x saportae show the highest activity against Cryptococcus neoformans, with a MIC value of 0.32 µL/mL.

The hydro-alcoholic extracts of Sardinian wild thistles (Onopordum spp.) inhibit TNFα-induced IL-8 secretion and NF-κB pathway in human gastric epithelial AGS cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Thistles species (Family: Compositae) are traditionally used in the Mediterranean area, particularly in Sardinia. They are usually gathered from the wild and used for both food and therapeutic purposes, including gastrointestinal disorders. AIM OF THE STUDY: This work aims to evaluate the anti-inflammatory activity of eight wild thistles from Sardinia, in an in vitro model of gastric inflammation, and to identify the major active compounds in the extracts. MATERIALS AND METHODS: The hydro-alcoholic extract of the aerial part of each species was prepared. After the induction of inflammation by the addition of tumor necrosis factor-α (TNFα) (10ng/mL), AGS cells were treated with extracts/pure compounds under study. The inhibition of interleukin-8 (IL-8) release, IL-8 and NF-κB promoter activities and NF-κB nuclear translocation were evaluated. Extracts main components were identified by HPLC-PDA-MS/MS. RESULTS: Only Onopordum horridum Viv. and Onopordum illyricum L. hydro-alcoholic extracts reduced, in a concentration-dependent fashion, the IL-8 release and promoter activity in human gastric epithelial cells AGS. The effect was partially due to the NF-κB pathway impairment. Onopordum hydro-alcoholic extracts were also chemically profiled, and caffeoylquinic acid derivatives were the main compounds identified in the extract. Further investigations showed that 3,5 dicaffeoylquinic acid highly inhibited IL-8 secretion in AGS cells (IC50 0.65µM), thus suggesting that this compound contributed, at least in part, to the anti-inflammatory activity elicited by O. illyricum extracts. CONCLUSIONS: Our results suggest that Onopordum species may exert beneficial effects against gastric inflammatory diseases. Thus, these wild plants deserve further investigations as preventive or co-adjuvant agents in gastric diseases.

Characterization of four wild edible Carduus species from the Mediterranean region via phytochemical and biomolecular analyses.

Carduus species (Compositae) are widely distributed in the Mediterranean area, and traditionally used for both food and medicinal purposes. The hydroalcoholic extracts of four wild edible Carduus species collected in Sardinia (Carduus argyroa Biv., Carduus nutans subsp. macrocephalus (Desf.) Nyman, Carduus pycnocephalus L., Carduus cephalanthus Viv.) were analyzed and characterized by HPLC-PDA-MS/MS and PCR-RFLP of the nrDNA internal transcribed spacer (ITS). Flavonoids and caffeoylquinic acid derivatives were the predominant classes of secondary metabolites characterizing the extracts. The ITS region was sequenced in parallel, and a PCR-RFLP method was applied with three selective restriction enzymes. Statistical analyses, on both chemical and biomolecular results, revealed that individuals clustered according to their taxonomic classification. The combination of the two techniques discriminates the four species within the genus, giving further information on these little-investigated plants, traditionally used in the Mediterranean area and in Sardinia.

Chemical composition of Lycium europaeum fruit oil obtained by supercritical CO2 extraction and evaluation of its antioxidant activity, cytotoxicity and cell absorption.

We studied the total phenols and flavonoids, liposoluble antioxidants, fatty acid and triacylglycerol profiles, and oxidative status of oil obtained from Lycium europaeum fruits following supercritical CO2 extraction (at 30MPa and 40°C). Linoleic (52%), palmitic (18%), oleic (13%), and α-linolenic (6%) were the main oil fatty acids, while trilinolein and palmitodilinolein/oleodilinolein represented the main triacylglycerols. The oil was characterized by high levels of all-trans-zeaxanthin and all-trans-ß-carotene (755 and 332µg/g of oil, respectively), α-tocopherol (308µg/g of oil), total phenols (13.6mg gallic acid equivalents/g of oil), and total flavonoids (6.8mg quercetin equivalents/g of oil). The oil showed radical scavenging activities (ABTS and DPPH assays) and inhibited Caco-2 cell growth. Moreover, the incubation of differentiated Caco-2 cells with a non-toxic oil concentration (100µg/mL) induced a significant intracellular accumulation of essential fatty acids. The results qualify L. europaeum oil as a potential source for food/pharmaceutical applications.

Evaluation of antioxidant and tyrosinase inhibitory activities of the extracts of Sarcopoterium spinosum (L.) Spach fruits.

Sarcopoterium spinosum fruits have been used to get extracts of different nature; two fixed oils were obtained by means of supercritical fluid extraction (SFE) with CO2 at 250 bar and 40°C and using n-hexane in a Soxhlet extraction (SE) apparatus. Aqueous solutions: an aromatic water (AW) and a residual water (RW) were obtained by hydrodistillation (HD). In the RW, following have been identified: quercetin glucuronide, luteolin 7-O-glucuronide, isorhamnetin 3-O-glucuronide, quercetin sulfate and quercetin. Among all tested plant extracts, the RW had the highest content of polyphenol (378 mg GAE/g of weight) and of flavonoids (26 mg QE/g of weight), and the highest antioxidant activity, comparable to that of Trolox. It was also the most active extract of this series (IC50 = 0.292 mg/mL) in the tyrosinase activity assays performed with L-3,4-dihydroxyphenylalanine (L-DOPA) as substrate.

Chemical characterisation and biological activity of leaf essential oils obtained from Pistacia terebinthus growing wild in Tunisia and Sardinia Island.

In the present work the chemical compositions, measured by GC and GC-MS, of the essential oils obtained by hydrodistillation from leaves of Pistacia terebinthus collected in Bizerte (Tunisia) and Baunei (Italy) are reported. Both essential oils possessed high content of monoterpene hydrocarbons (86.3% and 90.9%, respectively), being α-pinene (62.4 vs. 35.0)%, camphene (3.0 vs. 2.4)%, ß-pinene (12.1 vs. 4.5)%, terpinolene (1.7 vs. 35.2)% and ß-phellandrene (3.8 vs. 4.5)% the main components. The Tunisian essential oil exhibited higher antifungal activity than the Italian one. Cryptococcus neoformans and the majority of dermatophyte strains showed more sensitivity to the Tunisian oil, when compared to Candida strains, in particular Trichophyton rubrum, Microsporum canis and Epidermophyton floccosum, with MIC and MLC values in the range (0.16-0.32) µL/mL. The results obtained support the use of the oil from Tunisia for the treatment of dermatophytosis.

Untargeted Metabolomics of Tomato Plants after Root-Knot Nematode Infestation.

After 2 months from the infestation of tomato plants with the root-knot nematode (RKN) Meloidogyne incognita, we performed a gas chromatography-mass spectrometry untargeted fingerprint analysis for the identification of characteristic metabolites and biomarkers. Principal component analysis, and orthogonal projections to latent structures discriminant analysis suggested dramatic local changes of the plant metabolome. In the case of tomato leaves, ß-alanine, phenylalanine, and melibiose were induced in response to RKN stimuli, while ribose, glycerol, myristic acid, and palmitic acid were reduced. For tomato stems, upregulated metabolites were ribose, sucrose, fructose, and glucose, while fumaric acid and glycine were downregulated. The variation in molecular strategies to the infestation of RKNs may play an important role in how Solanum lycopersicum and other plants adapt to nematode parasitic stress.