Stemodin (1) is a diterpene produced by Stemodia maritima L. (Scrophulariaceae) and shows antiviral and tumor cell proliferation inhibitory activities. Chemical modifications of 1 were performed to produce the known stemodinone (2), two new stemodinone oximes, Z (3) and E (4), and seven new oxime esters derivatives (5, 6, 7, 8, 9, 10, and 11). All derivatives were evaluated for their cytotoxic activity in HL60 (promyelocytic leukemia), SNB-19 (astrocytoma), HCT-116 (colon carcinoma), and PC3 (prostate) human cancer cell lines, and L929 (healthy murine cells). Oximes 3 and 4 showed lower cytotoxic values than 1 against all cancer cell lines tested. However, esters 9 and 10 exhibited cell growth inhibition percentages higher than 1 against PC3, SNB-19, and HCT-116 cancer cell lines, ranging from 62.96 to 94.27%. Many of these values exceed that presented by doxorubicin (66.05-86.87%). All compounds showed no cytotoxic effect on healthy cells at the tested concentration of 25 µg mL-1. (1 L Scrophulariaceae (Scrophulariaceae activities 2, 2 , (2) oximes (3 4, (4) 5, 5 (5 6 7 8 11. 11 . 11) HL HL6 promyelocytic leukemia, leukemia leukemia) SNB19 SNB 19 SNB-1 astrocytoma, astrocytoma (astrocytoma) HCT116 HCT 116 HCT-11 colon carcinoma, carcinoma carcinoma) PC prostate (prostate L92 cells. cells) However SNB19, 19, 6296 62 96 62.9 9427 94 27 94.27% 66.0586.87%. 66058687 66.05 86.87% 66 05 86 87 (66.05-86.87%) mL1. mL1 mL 1. mL-1 ( (2 (4 SNB1 SNB- (astrocytoma HCT11 HCT-1 L9 629 62. 942 94.27 0586 66.0586.87% 6605868 6605 66.0 8687 86.87 0 (66.05-86.87% mL- HCT1 HCT- 94.2 058 66.0586.87 660586 660 66. 868 86.8 (66.05-86.87 94. 66.0586.8 66058 86. (66.05-86.8 66.0586. (66.05-86. 66.0586 (66.05-86 66.058 (66.05-8 (66.05- (66.05 (66.0 (66. (66 (6

Abstract Introduction/objectives Systemic lupus erythematosus (SLE) is a classic prototype of the multisystem autoimmune disease and follows a relapsing and remitting course. Triptolide is a diterpene triepoxide extracted from Chinese medicine Tripterygium wilfordii Hook F, with potent immunosuppressive and anti-inflammatory properties. Our previous work observed that triptolide alleviated lupus in MRL/lpr lupus mice with the upregulation of regulatory T cells (Treg) proportion in previous study. In this study, we explored the proportion of follicular T regulatory (Tfr), follicular T helper (Tfh) and germinal center (GC) B cells in lupus mice and evaluated the efficacy of triptolide for lupus treatment in vivo. Methods 20 female MRL/lpr mice were randomly divided into 2 treatment groups and treated orally with vehicle or triptolide. C3H mice were all housed as controlled group and treated orally with vehicle. The percentage of Tfr cells, Tfh cells and GC B cells in spleen of mice were detected by Flow cytometric analysis and immunohistochemistry after 13 weeks of treatment. Results We found that the percentage of Tfr cells decreased in MRL/lpr mice compared with controlled mice. The percentage of Tfh cells in MRL/lpr mice was significantly higher compared with that in controlled mice. The ratio of Tfr/Tfh is also decreased in lupus mice. After treated with triptolide in MRL/Lpr mice in vivo, the percentage of Tfr cells and ratio of Tfr/Tfh increased. The proportion of GC B cells also decreased in mice treated with triptolide by FACS and immunohistochemistry. Conclusions Our results demonstrate that the effect of triptolide in alleviating lupus is partly by reversing immune imbalance with increased percentage of Tfr cells and ratio of Tfr/Tfh. Triptolide might also has effect on immune response through inhibiting proliferating GC B cells. Introductionobjectives Introduction objectives SLE (SLE course F antiinflammatory anti inflammatory properties MRLlpr MRL lpr Treg (Treg study Tfr, , (Tfr) (Tfh (GC vivo CH C H 1 TfrTfh MRLLpr Lpr (Tfr

Phytochemical study of the roots of Euphorbia phosphorea Mart. (Euphorbiaceae) was carried out through chromatographic techniques, resulting in the isolation of a new ent-abietane diterpene named 11β,12β-dihydroxy-ent-abieta-8(14),13(15)-dien-16,12α-olide (1), and of nine known ent-abietane diterpenes jolkinolide A (2), jolkinolide E (3), euphorin H (4), euphopilolide (5) jolkinolide F (6), ent-12-hydroxy-12[R]-abieta-8(14),13(15)-dien-16,12-olide (7), ent-11α-hydroxyabieta-8(14),13(15)-dien-16,12α-olide (8), 17-hydroxyjolkinolide B (9) and caudicifolin (10). The structures of all compounds were established using spectroscopic techniques such as 1D and 2D NMR, and the structure of the compound 1 was established also with MS, IR and ECD. All compounds were submitted to an in silico study through of a predictive model and then submitted to in vitro tests against Mycobacterium tuberculosis and M. smegmatis for evaluation of their antimycobacterial activity. Compounds 5 and 9 showed mycobacterial growth inhibition with MIC values of 62.5 μM against M. tuberculosis and M. smegmatis, respectively.

ABSTRACT The effects of different nitrogen doses on dry weight yield, stevioside and rebaudioside A contents, and yields were evaluated in stevia (Stevia rebaudiana Bertoni). This study was carried out at the Instituto Federal Catarinense, Rio do Sul (SC), Brazil, under decreasing photoperiods. A randomized complete block design with four blocks and five treatments (0, 65, 135, 200 and 270 kg ha-1 N) was used. Variables related to dry weight yield (total dry weight, leaf dry weight, stem dry weight, number of secondary branches and tertiary branches, and main stem length), growth rates (total leaf area, leaf area index, specific leaf area, leaf area ratio, leaf weight ratio), and stevioside and rebaudioside A contents and yields were evaluated. The doses of nitrogen fertilization did not affect the variables related to dry weight yield, stevioside and rebaudioside A contents and yields, or rebaudioside A:stevioside ratio in stevia genotype 8 (G8), grown under decreasing photoperiods.

RESUMEN El efecto de diferentes tasas de nitrógeno en la producción de biomasa, el contenido y rendimiento de esteviósido y rebaudiósido A en el cultivo de estevia se evaluaron en un experimento de campo en el Instituto Federal Catarinense, Rio do Sul (SC), Brasil, en condiciones de fotoperíodo decreciente. Se utilizó un diseño de bloques completos al azar con cuatro bloques y cinco tratamientos (0, 65, 135, 200 y 270 kg ha-1 N). Se evaluaron las variables relacionadas con la producción de biomasa (DM total, tallo MS, hojas MS, número de ramas secundarias, número de ramas terciarias y longitud de rama más grande), variables relacionadas con los índices de crecimiento (AFT, IAF, AFE, RAF y RPF) y también variables relacionadas con el contenido y el rendimiento de esteviósido y rebaudiósido A. Las diferentes dosis de fertilización nitrogenada no afectaron las variables relacionadas con la acumulación de materia seca en el genotipo G8 cultivado en condiciones de fotoperíodo decreciente en las condiciones de campo. Los niveles de fertilización nitrogenada no afectaron los niveles de esteviósido y rebaudiósido A, la productividad de esteviósido y rebaudiósido A y la proporción de rebaudiósido A / esteviósido en el G8 cultivado en condiciones de fotoperíodo de disminución de campo.

Kahweol is a compound derived from coffee with reported antinociceptive effects. Based on the few reports that exist in the literature regarding the mechanisms involved in kahweol-induced peripheral antinociceptive action, this study proposed to investigate the contribution of the endocannabinoid system to the peripheral antinociception induced in rats by kahweol. Hyperalgesia was induced by intraplantar injection of prostaglandin E2(PGE2) and was measured with the paw pressure test. Kahweol and the drugs to test the cannabinoid system were administered locally into the right hind paw. The endocannabinoids were purified by open-bed chromatography on silica and measured by LC-MS. Kahweol (80 µg/paw) induced peripheral antinociception against PGE2-induced hyperalgesia. This effect was reversed by the intraplantar injection of the CB1 cannabinoid receptor antagonist AM251 (20, 40, and 80 μg/paw), but not by the CB2 cannabinoid receptor antagonist AM630 (100 μg/paw). Treatment with the endocannabinoid reuptake inhibitor VDM11 (2.5 μg/paw) intensified the peripheral antinociceptive effect induced by low-dose kahweol (40 μg/paw). The monoacylglycerol lipase (MAGL) inhibitor, JZL184 (4 μg/paw), and the dual MAGL/fatty acid amide hydrolase (FAAH) inhibitor, MAFP (0.5 μg/paw), potentiated the peripheral antinociceptive effect of low-dose kahweol. Furthermore, kahweol increased the levels of the endocannabinoid anandamide, but not of the other endocannabinoid 2-arachidonoylglycerol nor of anandamide-related N-acylethanolamines, in the plantar surface of the rat paw. Our results suggested that kahweol induced peripheral antinociception via anandamide release and activation of CB1 cannabinoid receptors and this compound could be used to develop new drugs for pain relief.

Resumo Fundamento: Diversos estudos têm mostrado que as classes de diterpenos exercem efeito significativo no sistema cardiovascular. Os diterpenos, em particular, estão entre os principais compostos associados às propriedades cardiovasculares, como a propriedade vasorrelaxante, inotrópica, diurética e a atividade hipotensora. Embora o mecanismo de vasorrelaxamento do manool seja visível, seu efeito sobre a pressão arterial (PA) ainda é desconhecido. Objetivo: Avaliar o efeito hipotensor in vivo do manool e verificar o efeito de vasorrelaxamento ex vivo em anéis aórticos de ratos. Métodos: Os animais foram divididos aleatoriamente em dois grupos: normotensos e hipertensos. O grupo normotenso foi submetido à cirurgia sham e adotou-se o modelo 2R1C para o grupo hipertenso. Realizou-se monitoramento invasivo da PA para testes com manool em diferentes doses (10, 20 e 40 mg/kg). Foram obtidas curvas de concentração-resposta para o manool nos anéis aórticos, com endotélio pré-contraído com fenilefrina (Phe) após incubação com Nω-nitro-L-arginina metil éster (L-NAME) ou oxadiazolo[4,3-a]quinoxalina-1-ona (ODQ). Os níveis plasmáticos de óxido nítrico (NOx) foram medidos por ensaio de quimioluminescência. Resultados: Após a administração de manool, a PA se reduziu nos grupos normotenso e hipertenso, e esse efeito foi inibido pelo L-NAME em animais hipertensos apenas na dose de 10 mg/kg. O manool ex vivo promoveu vasorrelaxamento, inibido pela incubação de L-NAME e ODQ ou remoção do endotélio. Os níveis plasmáticos de NOx aumentaram no grupo hipertenso após a administração de manool. O manool induz o relaxamento vascular dependente do endotélio na aorta de ratos, mediado pela via de sinalização NO/cGMP e redução da PA, e também pelo aumento plasmático de NOx. Esses efeitos combinados podem estar envolvidos na modulação da resistência periférica, contribuindo para o efeito anti-hipertensivo do diterpeno. Conclusão: Esses efeitos em conjunto podem estar envolvidos na modulação da resistência periférica, contribuindo para o efeito anti-hipertensivo do diterpeno.

Abstract Background: Many studies have shown that the diterpenoid classes exert a significant effect on the cardiovascular system. Diterpenes, in particular, are among the main compound links to cardiovascular properties such as vasorelaxant, inotropic, diuretic and hypotensive activity. While the manool vasorelaxation mechanism is visible, its effect on blood pressure (BP) is still unknown. Objective: To evaluate the in vivo hypotensive effect of manool and check the ex vivo vasorelaxation effect in rat aortic rings. Methods: The animals were divided randomly into two groups: normotensive and hypertensive. The normotensive group was sham-operated, and the 2K1C model was adopted for the hypertensive group. Invasive BP monitoring was performed for manool tests at different doses (10, 20 and 40 mg/kg). Concentration-response curves for manool were obtained in the aorta rings, with endothelium, pre-contracted with phenylephrine (Phe) after incubation with Nω-nitro-L-arginine methyl ester(L-NAME) or oxadiazole [4,3-a]quinoxalin-1-one (ODQ). Nitric oxide (NOx) plasma levels were measured by chemiluminescence assay. Results: After manool administration, BP was reduced in normotensive and hypertensive groups, and this effect was inhibited by L-NAME in hypertensive animals only in 10 mg/kg dose. Ex vivo manool promoted vasorelaxation, which was inhibited by L-NAME and ODQ incubation or endothelium removal. NOx plasma levels increased in the hypertensive group after manool administration. Manool elicits endothelium-dependent vascular relaxation in rat aorta mediated by the NO/cGMP signaling pathway and BP reduction, also by NOx plasma increase. These combined effects could be involved in modulating peripheral resistance, contributing to the antihypertensive effect of diterpene. Conclusion: These effects together could be involved in modulating peripheral resistance, contributing to the antihypertensive effect of diterpene.

Copaifera species (Fabaceae) comprises approximately 70 species of large trees, from which 16 can be found in Brazil. The oleoresins obtained from their trunk are widely used in Brazilian folk medicine, which display important antitumoral potential. Chemically, these oleoresins are mainly composed of a mixture of sesquiterpenes and diterpenes. In this paper we are describing the isolation and identification of 12 already known terpenes from oleoresins obtained from three different Copaifera species (C. multijuga, C. pubiflora and C. trapezifolia) and 2 novel diterpenes (ent-16-hidroxy-3,13 clerodadien-15,18-dioic acid and ent-labda-5,13-dien-15-oic acid) from C. trapezifolia. Both new compounds were identified by nuclear magnetic resonance (NMR) spectroscopic (1H and 13C NMR, correlation 1H-1H (COSY), heteronuclear multiple quantum coherence (HMQC) and heteronuclear multiple bond correlation (HMBC)) and by high-resolution electrospray ionization mass spectrometry (HR-ESIMS) analyses. The cytotoxic potential of these oleoresins, their main non-volatile compounds and their volatile compound fractions were evaluated against a panel of tumoral (MCF-7, ACP01, A549, HeLa) and normal cell lines (MCF-10A, GM07492-A) through XTT (tetrazolium salt) and SRB (sulforhodamine B) assays. The novel diterpene ent-labda-5,13-dien-15-oic acid displayed relevant cytotoxic effect against most of the cancer cell lines with mean inhibitory concentration (IC50) values ranging from 3.57 ± 1.12 to 22.56 ± 1.03 µg mL-1, and a high selectivity level in both assays.

The phytochemical investigation of the hexane extract which was obtained from the leaves of Xylopia laevigata (Annonaceae) produced six terpenes. Among these, five diterpenes named abieta-7,13-dien-3-one, ent-7β-acetoxy-16β-hydroxy-kaurane, 4-epi-cupressic acid, powerol, and labdorffianic acid B, and one oxygenated sesquiterpene, spathulenol. This is the first time these isolated diterpenes from X. laevigata have been described. The different structures belong to the abietane, kaurane, and labdane diterpenoids, and are indicative of the chemical diversity found in X. laevigata. Moreover, the diterpene ent-7β-acetoxy-16β-hydroxy-kaurane is reported herein for the first time as a natural product, and 4-epi-cupressic acid, powerol, and labdorffianic acid B are reported for the first time in the Annonaceae family. The structures of the isolated compounds were established by extensive analyses using 1D and 2D NMR spectroscopy in combination with MS. The cytotoxic activities of compounds abieta-7,13-dien-3-one, spathulenol, 4-epi-cupressic acid, and powerol were evaluated against tumor and non-tumor cell lines, in which spathulenol was found to be the most active, mainly against K562 with an IC50 value of 17.20 μmol L-1. (Annonaceae terpenes abieta7,13dien3one, abieta713dien3one abietadienone abieta 7,13 dien 3 one, 7 13 abieta-7,13-dien-3-one ent7βacetoxy16βhydroxykaurane, ent7βacetoxy16βhydroxykaurane entβacetoxyβhydroxykaurane ent 7β acetoxy 16β hydroxy kaurane β 4epicupressic epicupressic 4 epi cupressic sesquiterpene X described abietane diterpenoids Moreover product family D MS nontumor non lines active K K56 IC IC5 1720 17 20 17.2 L1. L1 L 1. 1 L-1 abieta7 13dien3one abieta7,13dien3one 713 7,1 βacetoxy βhydroxykaurane K5 172 2 17. L- dienone 71 7,

7a-Hydroxyroyleanone is a bioactive diterpene isolated from leaves of Tetradenia riparia, active against tuberculosis and T. cruzi. In order to use this compound as starting material for the synthesis of compounds with enhanced bioactivity we aimed the dehydration of the 7-alpha-hydroxyl group of royleanone. The reaction performed in acetic acid and hydrochloric acid under reflux gave an unexpected chlorinated naphtoquinone, with aromatization and opening of one of the rings of the starting material, as major product, besides 6,7-dehydroroyleanone and a complex mixture of other products. The reaction products were purified by high-speed countercurrent chromatography with hexane-acetonitrile 1:1. Compounds were tested for their anti-leishmanial activity (L. amazonensis) pointing out a potential leishmanicidal agent which may favor the development of more effective therapies.

Copaifera multijuga Hayne is the main species of the genus in the Amazon region, being exploited for copaiba oil production mainly because of its therapeutic properties. The plant parts (leaves, branches, and barks) are widely used in folk medicine. However, they are little explored in terms of chemical composition. In this study, the polar extracts of leaves, branches, and barks of the species were analysed by direct infusion in ESI-MS with the aid of reference standards and MS2 fragmentation experiments. Twenty-six substances (carboxylic acids, flavonoids, condensed tannins, a diterpene and fifteen isomers of galoylquinic acids) were detected. Metabolic profiles were similar between extracts of leaves and branches, especially with the detection of galloylquinic acids and carboxylic acids. In bark extract, condensed tannins epicatechin-(4β-8)-epiafzelechin (dimer) and epicatechin-(4β-8)-epiafzelechin-(4β-8)-epiafzelechin (trimer) were detected. The PCA showed differences and similarities in the chemical composition of the copaíba mari-mari extracts. Thus, galloylquinic acids and flavonoids kaempferol, 7-O-methyl-kaempferol, quercetin-3-O-L-pentoside, and quercetin-3-O-L-rhamnoside are described for the first time in C. multijuga. The phytotherapeutic properties of teas of the plant parts of the species may be associated with the suggested polyphenols.

RESUMO O perfil dos compostos bioativos em bebidas café depende das espécies e variedades de café, métodos de plantio e processamento pós-colheita, processo de torra e também do método de preparação da bebida. O trabalho teve como objetivo estudar os teores de caveol e cafestol - diterpenos do café com efeitos conhecidos a saúde - comparando bebidas de café preparadas usando métodos usuais de preparo (filtrada, espresso e café solúvel). As bebidas filtradas (pano ou papel), espresso e café solúvel foram obtidas de grãos com torra média da espécie Coffea arabica (tipo 2). Cinco repetições genuínas de cada bebida foram preparadas e os extratos foram liofilizados. Os diterpenos foram quantificados por CLUE. Os resultados foram relatados em mg de diterpeno por g de sólidos ou por dose padrão de 50 mL. O teor de sólidos das bebidas de café variou de 2,06 a 2,46 g 100 mL-1. Todas as bebidas apresentaram baixo teor de diterpenos: 0,05 a 0,16 mg de kahweol e 0,11 a 0,14 mg de cafestol 50 mL-1. A bebida instantânea mostrou o menor teor de kahweol e ausência de cafestol, redução relacionada ao processo de produção de café solúvel. O teor de diterpenos foi semelhante em cafés espresso e filtrados em papel. O café filtrado em pano apresentou menor teor de sólidos, mas níveis mais elevados de diterpenos (em mg g-1 de sólidos). Teores similares de cafestol e kahweol (mg 50 mL-1) foram observados nas bebidas filtradas e no espresso.

ABSTRACT The profile of bioactive compounds in coffee brews depends on the coffee bean species and varieties, harvesting and post-harvesting practices, roasting processes, and also on the brewing method. The present research aimed to study the contents of cafestol and kahweol - coffee diterpenes with a known impact on human health - comparing coffee beverages prepared using common brewing methods (filtered, espresso, and instant coffee brews). Filtered (cloth-filtered and paper-filtered), espresso and instant brews were obtained from a medium-roasted Coffea arabica coffee (NY 2). Five genuine replicates of each coffee brew were prepared, and the extracts were lyophilized. A validated UPLC-based method provided the content of diterpenes. The results were reported in mg of diterpene per g of solids or per a standard dose of 50 mL of coffee brew. Solids content of coffee brews ranged from 2.06 to 2.46 g 100 mL-1. All coffee brews presented low diterpene contents: 0.05 to 0.16 mg of kahweol and 0.11 to 0.14 mg of cafestol 50 mL-1. Instant coffee brew showed the lowest content of kahweol and absence of cafestol; this reduction was related to the production process of soluble coffee. Diterpenes content was similar in espresso and paper-filtered brews. The cloth-filtered coffee had lower solids content, but higher levels of diterpenes (in mg g-1 of solids). Similar cafestol and kahweol contents (mg 50 mL-1) were observed in filtered and espresso brews.

Abstract The emergence of antibiotic resistance in microorganisms is a serious challenge globally. Natural hydrophobic diterpene carboxylic acids present in rosin have unsatisfactory inhibitory properties against pathogens due to their poor water solubility. Therefore, the objective of research work was to modify the natural rosin into rosin maleic anhydride adduct with improved bioinhibitory properties for methicillin-resistant Staphylococcus aureus (MRSA). Prescreened MRSA isolates were found 78.05% and 29.27% resistant to oxacillin and vancomycin antibiotics respectively. The dosage effect of 0, 25, 50, and 100 mg L-1 rosin maleic anhydride adduct revealed the best inhibition response at 25 mg L-1. Moreover, bacteriostatic as well as the inhibitory effect of rosin maleic anhydride adduct was noticed against MRSA isolates. Gompertz model predicted better uptake of maleic anhydride adduct as compared to rosin. The higher specific growth rate of MRSA at reduced lag time correlated with increased toxicity of maleic anhydride adduct. This research concludes rosin maleic anhydride adduct has superior inhibitory properties against MRSA strains.

Abstract Commercial roasted and ground coffees are usually blends of Coffea arabica and Coffea canephora. Considering the differences in price and sensory characteristics between these two species, the identification of the presence of each species in commercial blends is of great interest. The aim of this study was to describe typical profiles of caffeine and diterpenes (kahweol and cafestol) contents and the ratios among these compounds to support the characterization of Coffea species in roasted coffees. 32 good cup quality Brazilian C. arabica coffees (from coffee quality contests) produced using different postharvest treatments were studied. All analysis were performed by HPLC. Higher ranges were observed in diterpene contents - kahweol varied from 1.75 to 10.68 g/kg (coefficient of variation of 510%) and cafestol from 1.76 to 9.66 g/kg (449%) - than caffeine, that varied from 5.1 to 16.2 g/kg (coefficient of variation of 218%). Wide ranges of the kahweol/cafestol ratio (0.63 to 2.77) and the caffeine/kahweol ratio (0.84 to 5.15) were also observed. Hence it was proposed the additional use of a new parameter, the ratio of caffeine/sum of diterpenes (kahweol + cafestol) that presents values from 0.54 to 2.39. The results indicated that the combined use of these parameters could be a potential tool for discriminating Coffea species in blends of roasted and ground coffee. It was proposed as potentially indicative of C. arabica: values of kahweol/cafestol ratio above 0.50, associated with caffeine/kahweol ratio lower than 5.50 and caffeine/sum of diterpenes ratio lower than 2.50.

Abstract The chemical study of roots from Azadirachta indica A. Juss., Meliaceae, led to the isolation of two new terpenoids, limonoid morenolide and diterpene 17-hydroxy-sandaracopimar-8,15-dien-11-one, in addition to the four well-known limonoids nimbinene, nimbinal, nimbandiol and salannin, and three diterpenoids nimbidiol, ferruginol, and 6,7-dehydroferruginol. Their structural elucidations were based on one and bidimensional Nuclear Magnetic Resonance and Electrospray ionization mass spectrometry spectra data which was compared to the data found in literature. The anti-inflammatory, cytotoxic and antimycobacterial activities of the identified terpenoids were evaluated.