De Novo Transcriptome Assembly of Cedar (Cedrela odorata L.) and Differential Gene Expression Involved in Herbivore Resistance.

Timber trees are targets of herbivorous attacks. The identification of genes associated with pest resistance can be accomplished through differential expression analysis using transcriptomes. We reported the de novo assembly of cedar (Cedrela odorata L.) transcriptome and the differential expression of genes involved in herbivore resistance. The assembly and annotation of the transcriptome were obtained using RNAseq from healthy cedar plants and those infested with Chrysobothris yucatanensis. A total of 325.6 million reads were obtained, and 127,031 (97.47%) sequences were successfully assembled. A total of 220 herbivory-related genes were detected, of which 170 genes were annotated using GO terms, and 161 genes with 245 functions were identified-165, 75, and 5 were molecular functions, biological processes, and cellular components, respectively. To protect against herbivorous infestation, trees produce toxins and volatile compounds which are modulated by signaling pathways and gene expression related to molecular functions and biological processes. The limited number of genes identified as cellular components suggests that there are minimal alterations in cellular structure in response to borer attack. The chitin recognition protein, jasmonate ZIM-domain (JAZ) motifs, and response regulator receiver domain were found to be overexpressed, whereas the terpene synthase, cytochrome P450, and protein kinase domain gene families were underexpressed. This is the first report of a cedar transcriptome focusing on genes that are overexpressed in healthy plants and underexpressed in infested plants. This method may be a viable option for identifying genes associated with herbivore resistance.

The putative cannabinoid-secreting trichome of Trema micrantha (L.) Blume (Cannabaceae).

Trema, a genus of the popularly known Cannabaceae, has recently been the subject of cannabinoid bioprospection. T. micrantha is a tree with pharmacological potential widely used in folk medicine. It has two types of glandular trichomes, bulbous and filiform, spread throughout the plant body. Considering the proximity of this species to Cannabis sativa and Trema orientalis, species containing cannabinoids, the glandular trichomes of T. micrantha are also expected to be related to the secretion of these compounds. Thus, this study aims to detail the morphology of secretory trichomes during the synthesis, storing and release of metabolites in T. micrantha. We tested the proposition that they could be a putative type of cannabinoid-secreting gland. Pistillate and staminate flowers and leaves were collected and processed for ontogenic, histochemical, and ultrastructural analyses. Both types of glandular trichomes originate from a protodermal cell. They are putative cannabinoid-secreting sites because: (1) terpene-phenols and, more specifically, cannabinoids were detected in situ; (2) their secretory subcellular apparatus is consistent with that found in C. sativa: modified plastids, polyribosomes, an extensive rough endoplasmic reticulum, and a moniliform smooth endoplasmic reticulum. Plastids and smooth endoplasmic reticulum are involved in the synthesis of terpenes, while the rough endoplasmic reticulum acts in the phenolic synthesis. These substances cross the plasma membrane by exocytosis and are released outside the trichome through cuticle pores. The study of the cell biology of the putative cannabinoid glands can promote the advancement of prospecting for natural products in plants.

Neither lysigenous nor just oil: Demystifying myrtaceous secretory cavities.

PREMISE: Leaf subepidermal secretory cavities are a notable trait in Myrtaceae, but their formation is still controversial because of the lack of consensus on their ontogeny among authors. Knowledge about the compounds present in these cavities has grown over the last few years, demonstrating that terpenoid-rich oils are not their unique content. These two points are the focus of this study on the ontogeny, structure, and contents of secretory cavities in neotropical Myrtaceae. METHODS: We used histochemical tests and Raman analysis to verify the basic chemical composition of the cavity contents of nine species. We studied the ontogeny of glands in one species, comparing aldehyde-fixed tissues and fresh sections mounted in an inert medium. RESULTS: We observed schizogenous development and appearance of the secretory cavities and found that sample processing may induce cell breakdown, which can be misinterpreted as lysigeny. The content of these cavities contains putative terpenes, resins, carbonyl groups, and flavonoids. CONCLUSIONS: Our findings support the hypothesis that the lysigenous appearance of the oil glands is a technical artifact. These tissue distortions must be considered when interpreting the development of this type of secretory structure. Moreover, the basic analyses of chemical constituents show for the first time that the glands of neotropical Myrtaceae are potential reservoirs of some compounds such as flavonoids previously reported as novelties for a few other myrtaceous species. Because some of them are non-lipid compounds, the idea that the glands are just oil repositories is no longer applicable.

Inhibition of Acinetobacter baumannii Biofilm Formation by Terpenes from Oregano (Lippia graveolens) Essential Oil.

Acinetobacter baumannii is a nosocomial pathogen known for its ability to form biofilms, leading to persistent infections and antibiotic resistance. The limited effective antibiotics have encouraged the development of innovative strategies such as using essential oils and their constituents. This study evaluated the efficacy of oregano (Lippia graveolens) essential oil (OEO) and its terpene compounds, carvacrol and thymol, in inhibiting A. baumannii biofilms. These treatments showed a minimum inhibitory concentration of 0.6, 0.3, and 2.5 mg/mL and a minimum bactericidal concentration of 1.2, 0.6, and 5 mg/mL, respectively. Sub-inhibitory doses of each treatment and the OEO significantly reduced biofilm biomass and the covered area of A. baumannii biofilms as measured by fluorescence microscopy. Carvacrol at 0.15 mg/mL exhibited the most potent efficacy, achieving a remarkable 95% reduction. Sub-inhibitory concentrations of carvacrol significantly reduced the biofilm formation of A. baumannii in stainless steel surfaces by up to 1.15 log CFU/cm2 compared to untreated bacteria. The OEO and thymol exhibited reductions of 0.6 log CFU/cm2 and 0.4 log CFU/cm2, respectively, without affecting cell viability. Moreover, the terpenes inhibited twitching motility, a crucial step in biofilm establishment, with carvacrol exhibiting the highest inhibition, followed by OEO and thymol. The study provides valuable insights into the potential of terpenes as effective agents against A. baumannii biofilms, offering promising avenues for developing novel strategies to prevent persistent infections and overcome antibiotic resistance.

Fine resolution analysis of bacterial communities associated with Neochloris oleoabundans culture and insights into terpenes as contamination control agents.

Biological contamination is one of the main bottlenecks in microalgae production, reducing quality and productivity and sometimes leading to the complete loss of the cultures. Selecting terpenes can be a pathway toward eco-friendly contamination control in microalgae cultures. This work evaluated the presence of bacterial contaminants in N. oleoabundans cultures through HTS and 16 S analysis and their susceptibility to six natural terpenes (α-pinene, ß-pinene, limonene, trans-cinnamaldehyde, linalool, and eugenol). The principal phyla identified were Proteobacteria, Bacteroidetes, and Actinobacteria, and based on these data, 89 bacterial isolates of seven genera were obtained (36 Aureimonas sp., 27 Microbacterium sp., 5 Pseudomonas sp., 9 Bacillus sp., 14 Shinella sp., 1 Brevundimonas sp., and 1 Exiguobacterium sp.) at 25ºC in the presence of light. It was possible to observe that Beta-pinene 50 mg L- 1 only inhibited Bacillus sp. In contrast, Alpha-pinene, Linalool, and Trans-cinnamaldehyde, at a concentration of 6.25 mg L- 1 efficiently inhibited most isolates. The inhibition percentages found were 79-99%.

Centella asiatica and Its Metabolite Asiatic Acid: Wound Healing Effects and Therapeutic Potential.

An intense effort has been focused on new therapeutic approaches and the development of technologies for more efficient and rapid wound healing. The research for plants used for long time in traditional medicine in the treatment of wound has become a promising strategy to obtain drugs therapeutically useful in the acute and chronic wound management. In this context, Centella asiatica (Apiaceae) has been used to treat a variety of skin diseases, such as leprosy, lupus, varicose ulcers, eczema and psoriasis, in Asiatic traditional medicine for thousands of years. Studies have shown that Centella asiatica extracts (CAE) display activity in tissue regeneration, cell migration and wound repair process by promoting fibroblast proliferation and collagen synthesis. Preliminary findings have shown that the asiatic acid is one of the main active constituents of C. asiatica, directly associated with its healing activity. Thus, this study discusses aspects of the effects of Centella asiatica and its active component, asiatic acid, in different stages of the healing process of cutaneous wounds, including phytochemical and antimicrobial aspects that contribute to its therapeutic potential.

Functional characterization of a bark-specific monoterpene synthase potentially involved in wounding- and methyl jasmonate-induced linalool emission in rubber (Hevea brasiliensis).

Rubber (Hevea brasiliensis) is a latex-producing plant that often encounters mechanical wounding, as well as pathogen and pest attacks through wound sites during and after tapping. Terpenoids play an important role in the ecological interactions of many plant species, and their diversity is mainly generated by enzymes known as terpene synthases (TPS). In this study, one cDNA sequence encoding a putative terpene synthase, HbTPS20, was obtained from the bark tissues of H. brasiliensis. The encoded protein contains 610 amino acids with a putative N-terminal plastid transit peptide of approximately 70 residues. It belongs to the TPS-b subfamily. Further phylogenetic analysis showed that HbTPS20 formed a separate branch that diverged from the progenitor of all other potentially functional terpene synthases of the rubber TPS-b subfamily. The truncated HbTPS20 without the signal peptide coding sequence was successfully expressed in E. coli and in vitro enzymatic assays with geranyl diphosphate (GPP) or neryl diphosphate (NPP) as a substrate defined HbTPS20 as an active linalool synthase (HbLIS) with the ability to produce linalool as the principal product. RT-qPCR analysis showed that the highest transcript levels of HbTPS20 were found in barks, and this gene was expressed at 2.26- and 250-fold greater levels in the bark tissues of wounded and MeJA-treated plants, respectively, than in those of the control plants. This indicates that this gene may be involved in the induced stress responses of rubber.

Determination of variability of terpenes and terpenoids in Cannabis sativa by gas chromatography-flame ionization detection and gas chromatography-mass spectrometry.

Absolute content of terpenes in inflorescences of two strains of Cannabis sativa L., CAT 1 and CAT 3, has been determined. Twenty terpenes commonly present in these samples were quantified by solid phase microextraction combined with gas chromatography and flame ionization detection (SPME/GC-FID). High amounts of ß-myrcene, α-pinene, ß-pinene, limonene, (E)-ß-ocimene, ß-caryophyllene, α-humulene, (E)-nerolidol, and linalool, were found in both strains. Lower concentrations (< 20 µg·g-1) of other terpenes were also determined. Only (E)-ß-ocimene was detected at 50 µg·g-1 in CAT 3 whereas it was below the LOD in CAT 1. Concentrations of other compounds for which standards were not available, were estimated based on a response factor obtained from the calibration curves of compounds with similar chemical structures. Fingerprints of both CAT strains were obtained and the identities of most volatile compounds were assigned using gas chromatography coupled to mass spectrometer detector (GC-MS). Additionally, an assessment of variability of terpenes was achieved by analyzing ten plants of each strain grown under controlled conditions and harvested at the same time. This variability was about 20%, considering terpenes at concentration above 20 µg·g-1.

In Silico Genome-Wide Mining and Analysis of Terpene Synthase Gene Family in Hevea Brasiliensis.

Terpene synthases (TPSs) catalyze terpenoid synthesis and affect the intracellular isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) concentration. In this study, we mined the in silico genome-wide TPS genes of Hevea brasiliensis and identified 47 full-length TPS genes. They had DDXXD, DXDD, NSE/DTE, RR(X)8 W, EA(X)W, and other conserved motifs. The phylogenetic tree analysis revealed that the TPSs of H.brasiliensis (HbTPSs) were divided into five subfamilies, TPS-a, TPS-b, TPS-c, TPS-e/f, and TPS-g. HbTPSs were predicted to have functions in the cellular components, molecular functions, and biological processes. HbTPSs were involved in seven pathways, which were K14173, K14175, K15803, K04120, K04121, K17982, and K12742 in the secondary metabolite pathway prediction. Three-dimensional structures of HbTPSs of 7 pathways were predicted, and DDXXD, NSE/DTE, and EA(X)W conserved motifs near the binding sites were found. Cis-acting elements analysis showed that they had more cis-acting elements related to phytohormone responsiveness, which indicated that terpenoid biosynthesis might be related to phytohormone regulation. RNA-Seq analysis showed that different HbTPSs were expressed differentially in different tissues. This study's results help reveal the role of HbTPSs and their molecular mechanism and help resolve the regulatory mechanism of terpenoid biosynthesis in H.brasiliensis.

Bioactive compounds of the Opuntia monacantha fruit.

Opuntia monacantha is a Cactaceae found in regions with few nutrients and water. This study aimed to identify bioactive compounds in the fruit of O. monacantha in the southern region of Brazil. In the fruit of O. monacantha, five phenolic compounds were identified: protocatechuic acid, ferulic acid, p-coumaric acid, catechin and quercetin. The ferulic acid and catechin were found in higher concentration in the fruit. The terpenes α-amyrin, ß-amyrin, oleanolic acid and cycloartenol were also identified in the hexane fractions of the fruit. Unsaturated fatty acids were found in the fruit seed, with the highest concentration of linoleic acid (69.0%).

Essential Oils of Aromatic Plant Species from the Atlantic Rainforest Exhibit Extensive Chemical Diversity and Antimicrobial Activity.

Microbial resistance, caused by the overuse or inadequate application of antibiotics, is a worldwide crisis, increasing the risk of treatment failure and healthcare costs. Plant essential oils (EOs) consist of hydrophobic metabolites with antimicrobial activity. The antimicrobial potential of the chemical diversity of plants from the Atlantic Rainforest remains scarcely characterized. In the current work, we determined the metabolite profile of the EOs from aromatic plants from nine locations and accessed their antimicrobial and biocidal activity by agar diffusion assays, minimum inhibitory concentration, time-kill and cell-component leakage assays. The pharmacokinetic properties of the EO compounds were investigated by in silico tools. More than a hundred metabolites were identified, mainly consisting of sesqui and monoterpenes. Individual plants and botanical families exhibited extensive chemical variations in their EO composition. Probabilistic models demonstrated that qualitative and quantitative differences contribute to chemical diversity, depending on the botanical family. The EOs exhibited antimicrobial biocidal activity against pathogenic bacteria, fungi and multiple predicted pharmacological targets. Our results demonstrate the antimicrobial potential of EOs from rainforest plants, indicate novel macromolecular targets, and contribute to highlighting the chemical diversity of native species.

Avaliação da atividade antibacteriana do monoterpeno (r)-(+)-citronelal contra cepas de Pseudomonas aeruginosa

Introdução: Os monoterpenos, que são os principais constituintes da maioria dos óleos essenciais, compreendem uma série ampla de substâncias naturais e/ou sintéticas que já apresentaram diversas atividades biológicas. Objetivo: O presente estudo teve como objetivo avaliar a atividade antibacteriana do monoterpeno (R)-(+)-citro nelal contra cepas de Pseudomonas aeruginosa. Metodologia: Utilizou-se a técnica de diluição seriada para a determinação da concentração inibitória mínima (CIM) e também realizou-se a determinação da concentração bactericida mínima (CBM) do (R)-(+)-citronelal. Resultados: Os resultados obtidos para o monoterpeno testado foi de CIM igual à 512 µg/mL, apresentando assim um efeito antibacteriano forte e CBM variação de 512 a 1024 µg/mL, sendo assim considerado bactericida. Conclusão: Conclui-se que o monoterpeno (R)-(+)-citronelal devido ao seu potencial antimicrobiano pode ser considerado muito importante para o tratamento de várias enfermidades causadas pela Pseudomonas aeruginosa.

SUMMARY Introduction: Monoterpenes, which are the main constituents of most essential oils, comprise a wide range of natural and/or synthetic substances that have already exhibited diverse biological activities. Objective: The present study aimed to evaluate the antibacterial activity of monoterpene (R)-(+)-citronellal against Pseudomonas aeruginosa strains. Methodology: The serial dilution technique was used to determine the Minimum Inhibitory Concentration (MIC) and the Minimal Bactericidal Concentration (CBM) of (R)-(+)-citronellal was also determined. Results: The results obtained for the tested monoterpene were MIC50 equal to 512 µg/mL, thus presenting a strong antibacterial effect and CBM ranging from 512 to 1024 µg/mL, thus being considered bactericidal. Conclusion : Thus, it is concluded that the monoterpene (R)-(+)-citronellal, due to its antimicrobial potential, can be considered very important for the treatment of several diseases caused by Pseudomonas aeruginosa.

Introducción: Los monoterpenos, que son los componentes principales de la mayoría de los aceites esenciales, comprenden una amplia gama de sustancias naturales o sintéticas que ya han exhibido diversas actividades biológicas. Objetivo: El presente estudio tuvo como objetivo evaluar la actividad antibacteriana del monoterpeno (R)-(+)-citronelal contra cepas de Pseudomonas aeruginosa. Metodología: Se utilizó la técnica de dilución en serie para determinar la Concentración Inhibitoria Mínima (MIC) y también se determinó la concentración bactericida mínima (CBM) de (R)-(+)-citronelal. Resultados: Los resultados obtenidos para el monoterpeno ensayado fueron MIC50 igual a 512 µg/mL, presentando así un fuerte efecto antibacteriano y CBM que van desde 512 a 1024 µg/mL, por lo que se considera bactericida. Conclusión: así, se concluye que el monoterpeno (R)-(+)-citronelal, por su potencial antimicrobiano, puede considerarse muy importante para el tratamiento de diversas enfermedades causadas por Pseudomonas aeruginosa.

Myrtenol Reduces Orofacial Nociception and Inflammation in Mice Through p38-MAPK and Cytokine Inhibition.

Orofacial pain is one of the commonest and most complex complaints in dentistry, greatly impairing life quality. Preclinical studies using monoterpenes have shown pharmacological potential to treat painful conditions, but the reports of the effects of myrtenol on orofacial pain and inflammation are scarce. The aim of this study was to evaluate the effect of myrtenol in experimental models of orofacial pain and inflammation. Orofacial nociceptive behavior and the immunoreactivity of the phosphorylated p38 (P-p38)-MAPK in trigeminal ganglia (TG) and spinal trigeminal subnucleus caudalis (STSC) were determined after the injection of formalin in the upper lip of male Swiss mice pretreated with myrtenol (12.5 and 25 mg/kg, i.p.) or vehicle. Orofacial inflammation was induced by the injection of carrageenan (CGN) in the masseter muscle of mice pretreated with myrtenol (25 and 50 mg/kg, i.p.) or its vehicle (0.02% Tween 80 in saline). Myeloperoxidase (MPO) activity and histopathological changes in the masseter muscle and interleukin (IL)-1ß levels in the TG and STSC were measured. The increase in face-rubbing behavior time induced by formalin and P-p38-MAPK immunostaining in trigeminal ganglia were significantly reduced by myrtenol treatment (12.5 and 25 mg/kg). Likewise, increased MPO activity and inflammatory histological scores in masseter muscle, as well as augmented levels of IL-1ß in the TG AND STSC, observed after CGN injection, were significantly decreased by myrtenol (25 and 50 mg/kg). Myrtenol has potential to treat orofacial inflammation and pain, which is partially related to IL-1ß levels in the trigeminal pathway and p38-MAPK modulation in trigeminal ganglia.

A Bioinformatics Tool for Efficient Retrieval of High-Confidence Terpene Synthases (TPS) and Application to the Identification of TPS in Coffea and Quillaja.

Terpenoids are a class of compounds that are found in all living organisms. In plants, some terpenoids are part of primary metabolism, but most terpenes found in plants are classified as specialized metabolites, encoded by terpene synthases (TPS). It is not obvious how to assign the putative product of a given TPS using bioinformatics tools. Phylogenetic analyses easily assign TPS into families; however members of the same TPS family can synthetize more than one terpenoid-and, in many biotechnological applications, researchers are more interested in the product of a given TPS rather than its phylogenetic profile. Automated protein annotation can be used to classify TPS based on their products, despite the family they belong to. Here, we implement an automated bioinformatics method, search_TPS, to identify TPS proteins that synthesize mono, sesqui and diterpenes in Angiosperms. We verified the applicability of the method by classifying wet lab validated TPS and applying it to find TPS proteins in Coffea arabica, C. canephora, C. eugenioides, and Quillaja saponaria. Search_TPS is a computational tool based on PERL scripts that carries out a series of HMMER searches against a curated database of TPS profile hidden Markov models. The tool is freely available at https://github.com/liliane-sntn/TPS .

Tropane alkaloids and terpenes synthase genes of Datura stramonium (Solanaceae).

BACKGROUND: Plants have evolved physical-chemical defense to prevent/diminish damage by their enemies. Chemical defense involves the synthesis' pathways of specialized toxic, repellent, or anti-nutritive metabolites to herbivores. Molecular evolutionary studies have revealed the origin of new genes, acquisition and functional diversification along time in different plant lineages. METHODS: Using bioinformatic tools we analyze gene divergence of tropane alkaloids (TAs) and terpene synthases (TPSs) in Datura stramonium and other species of Solanaceae; compared gene and amino acids sequence of TAs and TPSs on genomes, cDNA and proteins sequences of Viridiplantae. We analyzed two recently assembled genomes of D. stramonium (Ticumán and Teotihuacán), transcriptomes of Datura metel and genomes of other Solanaceae. Hence, we analyzed variation of TAs and TPSs to infer genes involved in plant defense and plant responses before stress. We analyzed protein modeling and molecular docking to predict interactions between H6H and ligand; we translated the sequences (Teo19488, Tic8550 and Tic8549) obtained from the two genomes of D. stramonium by using Swiss-Model and Ramachandran plot and MolProbity structure validation of Teo19488 protein model. RESULTS: For TAs, we detected an expansion event in the tropinone reductase II (TRII) and the ratio synonymous/non-synonymous substitutions indicate positive selection. In contrast, a contraction event and negative selection was detected in tropinone reductase I (TRI). In Hy-oscyamine 6 b-hydroxylase (H6H), enzyme involved in the production of tropane alkaloids atropine and scopolamine, the synonymous/non-synonymous substitution ratio in its dominion indicates positive selection. For terpenes (TPS), we found 18 DsTPS in D. stramomiun and seven in D. metel; evolutionary analyses detected positive selection in TPS10.1 and TPS10.2 of D. stramonium and D. metel. Comparison of copies of TPSs in D. stramonium detected variation among them in the binding site. Duplication events and differentiation of TAs and TPSs of D. stramonium, as compared to other Solanaceae, suggest their possible involvement on adaptive evolution of defense to herbivores. Protein modeling and docking show that the three protein structures obtained of DsH6H from Teo19488, Tic-8550 and Tic8549 maintain the same interactions and the union site of 2OG-FeII_Oxy with the Hy-o ligand as in 6TTM of D. metel. CONCLUSION: Our results indicate differences in the number of gene copies involved in the synthesis of tropane alkaloids, between the genomes of D. stramonium from two Mexican populations. More copies of genes related to the synthesis of tropane alkaloids (TRI, TRII, H6H, PMT) are found in D. stramonium as compared to Viridiplantae. Likewise, for terpene synthases (TPS), TPS-10 is duplicated in D. stramonium and D. metel. Further studies should be directed to experimentally assess gain (overexpression) or loss (silencing) of function of duplicated genes.

Brazilian essential oils as source for the discovery of new anti-COVID-19 drug: a review guided by in silico study.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in China and its spread worldwide has become one of the biggest health problem due to the lack of knowledge about an effective chemotherapy. Based on the current reality of the SARS-CoV-2 pandemic, this study aimed to make a review literature about potential anti-coronavirus natural compounds guided by an in silico study. In the first step, essential oils from native species found in the Brazilian herbal medicine market and Brazilian species that have already shown antiviral potential were used as source for the literature search and compounds selection. Among these compounds, 184 showed high antiviral potential against rhinovirus or picornavirus by quantitative structure-activity relationship analysis. (E)-α-atlantone; 14-hydroxy-α-muurolene; allo-aromadendrene epoxide; amorpha-4,9-dien-2-ol; aristochene; azulenol; germacrene A; guaia-6,9-diene; hedycaryol; humulene epoxide II; α-amorphene; α-cadinene; α-calacorene and α-muurolene showed by a molecular docking study the best result for four target proteins that are essential for SARS-CoV-2 lifecycle. In addition, other parameters obtained for the selected compounds indicated low toxicity and showed good probability to achieve cell permeability and be used as a drug. These results guided the second literature search which included other species in addition to native Brazilian plants. The majority presence of any of these compounds was reported for essential oils from 45 species. In view of the few studies relating essential oils and antiviral activity, this review is important for future assays against the new coronavirus. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s11101-021-09754-4.

Rerouting and Improving Dauc-8-en-11-ol Synthase from Streptomyces venezuelae to a High Yielding Biocatalyst.

The dauc-8-en-11-ol synthase from Streptomyces venezuelae was investigated for its catalytic activity towards alternative terpene precursors, specifically designed to enable new cyclisation pathways. Exchange of aromatic amino acid residues at the enzyme surface by site-directed mutagenesis led to a 4-fold increase of the yield in preparative scale incubations, which likely results from an increased enzyme stability instead of improved enzyme kinetics.

Photoprotective Activity of Buddleja cordata Cell Culture Methanolic Extract on UVB-irradiated 3T3-Swiss Albino Fibroblasts.

The research on compounds exhibiting photoprotection against ultraviolet radiation (UVR) is a matter of increasing interest. The methanolic extract of a cell culture of Buddleja cordata has potential photoprotective effects as these cells produce phenolic secondary metabolites (SMs). These metabolites are attributed with biological activities capable of counteracting the harmful effects caused by UVR on skin. In the present work, the methanolic extract (310-2500 µg/mL) of B. cordata cell culture showed a photoprotective effect on UVB-irradiated 3T3-Swiss albino fibroblasts with a significant increase in cell viability. The greatest photoprotective effect (75%) of the extract was observed at 2500 µg/mL, which was statistically comparable with that of 250 µg/mL verbascoside, used as positive control. In addition, concentrations of the extract higher than 2500 µg/mL resulted in decreased cell viability (≤83%) after 24 h of exposure. Phytochemical analysis of the extract allowed us to determine that it was characterized by high concentrations of total phenol and total phenolic acid contents (138 ± 4.7 mg gallic acid equivalents and 44.01 ± 1.33 mg verbascoside equivalents per gram of extract, respectively) as well as absorption of UV light (first and second bands peaking at 294 and 330 nm, respectively). Some phenylethanoid glycosides were identified from the extract.

Nerolidol-beta-cyclodextrin inclusion complex enhances anti-inflammatory activity in arthritis model and improves gastric protection.

Rheumatoid arthritis is an autoimmune inflammatory disease with progressive degradation of cartilage and joints. Additionally, gastric ulcer affects many patients who make prolonged use of non-steroidal anti-inflammatory drugs widely used in the symptomatic treatment of rheumatoid arthritis. Nerolidol, a natural sesquiterpene, has several biological activities including anti-inflammatory and antiulcerogenic action. This study aims to develop and characterize a nerolidol ß-cyclodextrin inclusion complex and to evaluate its activity in an experimental arthritis model. Inclusion complex was prepared by the lyophilization method and characterized by NMR, term analysis, XRD and SEM. Neutrophil migration assays and histopathological analysis were performed on zymosan-induced arthritis model using Swiss mice. And the gastroprotective effect was evaluated in two models of gastric ulcers: induced by ethanol and indomethacin. Inclusion complex showed no cytotoxicity and free nerolidol at a dose of 100 mg/kg (p.o.) in the arthritis model reduced neutrophil migration in 56% in relation to vehicle, and this inhibition was more expressive in the inclusion complex (67%) at the same dose. Histopathological analysis of the joint tissue confirmed the reduction of inflammatory signs. In the ethanol-induced gastric ulcer model, free nerolidol reduced the relative ulcer area more expressively (4.64%) than the inclusion complex (21.3%). However, in the indomethacin induction model, the inclusion complex showed better results in gastric protection compared to free nerolidol. The action of nerolidol complexed in beta-cyclodextrin in reducing arthritis inflammation combined with its gastroprotective action make it a potential new drug.

Bioactive metabolites in of Ginkgo biloba leaves: variations by seasonal, meteorological and soil / Metabólitos bioativos em folhas de Ginkgo biloba: variações sazonais, meteorológicas e do solo

Ginkgo biloba is a traditional Chinese herbal medicine containing multiple components that contribute to its notable bioactivities. Variations of seasonal, meteorological and planting soil on the phytochemicals contents in G. biloba leaves due to the effects of growth meteorological and soil parameters were investigated in this study. The leaves of G. biloba were collected from different months and place in Zhejiang province, the contents of flavones (quercetin, kaempferol and isorhamnetin) and terpene lactones (bilobalide, ginkgolides A, B and C) were quantified by high performance liquid chromatography (HPLC) and the evaporative light scattering detector (ELSD) method. The established methods were validated with good linearity, precision, repeatability, stability, and recovery. Comprehensive analysis suggested the proper harvest time for G. biloba was in October of Zhejiang province. The result of correlation analysis with meteorological factors shows that the temperature and precipitation have non-significant effect on the main components of G. biloba. In addition, the type and content (Mn and Zn) of the soil showed significantly effect on the content of flavonoids and terpene lactones. This study enriched the knowledge on the development and utilization value of the G. biloba leaves and was useful for determining the optimal harvest time and growing condition.(AU)

Ginkgo biloba é um fitoterápico tradicional da medicina chinesa que contém vários componentes que contribuem para suas notáveis bioatividades. Variações sazonais, meteorológicos e de plantio do solo sobre os teores fitoquímicos em folhas de G. biloba, devido aos efeitos do crescimento meteorológico e parâmetros do solo, foram investigadas neste estudo. As folhas de G. biloba foram coletadas em diferentes meses e na província de Zhejiang, os teores de flavonas (quercetina, kaempferol e isorhamnetina) e lactonas terpênicas (bilobalida, ginkgolídeos A, B e C) foram quantificados por Cromatografia Líquida de Alta Eficiência (CLAE) e pelo método do detector de dispersão da luz por evaporação. Os métodos estabelecidos foram validados com boa linearidade, precisão, repetibilidade, estabilidade e recuperação. Uma análise abrangente sugeriu que o tempo de colheita adequado para G. biloba foi em outubro na província de Zhejiang. O resultado da análise de correlação com fatores meteorológicos mostra que a temperatura e a precipitação não têm efeito significativo sobre os principais componentes de G. biloba. Além disso, o tipo e composição (Mn e Zn) do solo apresentaram efeito significativo sobre o teor de flavonoides e lactonas terpênicas. Este estudo enriqueceu o conhecimento sobre o valor de desenvolvimento e utilização das folhas de G. biloba e foi útil para determinar o melhor tempo de colheita e condição de crescimento.(AU)