Validation of a New HPLC-DAD Method to Quantify 3-Deoxyanthocyanidins Based on Carajurin in Medicinal Plants and for Chemical Ecology Studies.

Anthocyanidins are found in nature mainly as pelargonidin, cyanidin, peonidin, delphinidin, petunidin, and malvidin derivatives. These compounds are found free or as glycoside derivatives which are responsible for the color (red, blue, and violet) of some foods and are responsible for attracting seed dispersers. They are grouped into 3-hydroxyanthocyanidins, 3-deoxyanthocyanidins (3D-anth), and O-methylated anthocyanidins. A new method was developed and validated to quantify 3D-anth in plant-rich extracts. To test the new method, Arrabidaea chica Verlot was selected as it is widely used in folk medicine, and it is rich in 3D-anth. The new method was developed by HPLC-DAD and expressed 3D-anth as carajurin content. Carajurin was chosen as the reference standard due to its role as a biological marker for the antileishmanial activity for A. chica. The selected method used a silica-based phenyl column, a mobile phase composed of potassium dihydrogen phosphate buffer, acetonitrile, and methanol, in a gradient elution mode and detection at 480 nm. The method reliability was confirmed by verifying selectivity, linearity, precision, recovery, and robustness. This method contributes to quality control and development of a possible active pharmaceutical ingredient from A. chica as well as it can be used to evaluate 3D-anth in plant extracts with chemical ecology interest.

Carajurin Induces Apoptosis in Leishmania amazonensis Promastigotes through Reactive Oxygen Species Production and Mitochondrial Dysfunction.

Carajurin is the main constituent of Arrabidaea chica species with reported anti-Leishmania activity. However, its mechanism of action has not been described. This study investigated the mechanisms of action of carajurin against promastigote forms of Leishmania amazonensis. Carajurin was effective against promastigotes with IC50 of 7.96 ± 1.23 µg.mL-1 (26.4 µM), and the cytotoxic concentration for peritoneal macrophages was 258.2 ± 1.20 µg.mL-1 (856.9 µM) after 24 h of treatment. Ultrastructural evaluation highlighted pronounced swelling of the kinetoplast with loss of electron-density in L. amazonensis promastigotes induced by carajurin treatment. It was observed that carajurin leads to a decrease in the mitochondrial membrane potential (p = 0.0286), an increase in reactive oxygen species production (p = 0.0286), and cell death by late apoptosis (p = 0.0095) in parasites. Pretreatment with the antioxidant NAC prevented ROS production and significantly reduced carajurin-induced cell death. The electrochemical and density functional theory (DFT) data contributed to support the molecular mechanism of action of carajurin associated with the ROS generation, for which it is possible to observe a correlation between the LUMO energy and the electroactivity of carajurin in the presence of molecular oxygen. All these results suggest that carajurin targets the mitochondria in L. amazonensis. In addition, when assessed for its drug-likeness, carajurin follows Lipinski''s rule of five, and the Ghose, Veber, Egan, and Muegge criteria.

Transplacental Transfer of Primaquine and Neurobehavioral Development of Prenatally Exposed Rats.

Primaquine (PQ) not only eliminates P. falciparum gametocytes but also kills liver dormant forms of P. vivax and P. ovale. Owing to these unique therapeutic properties, it is an essential drug. Although PQ has been used for over 70 years, its toxicological database has gaps such as the absence of studies on its reproductive and developmental toxicity and kinetics in pregnancy. This study investigated the transplacental transfer of PQ and the effects of intrauterine exposure on the postnatal growth, survival, and neurobehavioral development of the offspring. PQ kinetics and transplacental transfer were investigated in rats treated orally (40 mg.kg·bw-1) on gestation day (GD) 21. PQ was analyzed by high-performance liquid chromatography with diode array ultraviolet detection. To evaluate effects of intrauterine exposure on postnatal development, dams were treated orally with PQ (20 mg.kg·bw-1·d-1) or water (controls) on GD 0-21. Postnatal survival, body weight gain, somatic maturation, and reflex acquisition were evaluated. The open field test (OF) was conducted on PND 25. PQ concentration in the fetal plasma was nearly half that in maternal plasma. Except for increase in pregnancy loss, no effects of PQ were noted at term pregnancy and first days of life. Prenatal PQ did not affect postnatal weight gain nor did it impair somatic and neurologic development of the offspring. Pups born to PQ-treated dams showed reduced exploration and enhanced emotionality in the OF. PQ given in pregnancy, at doses greater than those recommended for malaria therapy, may affect pup postnatal survival and emotional behavior.

Carajurin: a anthocyanidin from Arrabidaea chica as a potential biological marker of antileishmanial activity.

Leishmaniasis is a group of neglected tropical diseases whose treatment with antimonials bears limitations and has changed little in over 80 years. Medicinal plants have been evaluated as a therapeutic alternative for leishmaniasis. Arrabidaea chica is popularly used as a wound healing and antiparasitic agent, especially as leishmanicidal agent. This study examined the leishmanicidal activity of a crude extract (ACCE), an anthocyanidin-rich fraction (ACAF), and three isolated anthocyanidins from A. chica: carajurin, 3'-hydroxy-carajurone, and carajurone. We evaluated the antileishmanial activity against promastigote and intracellular amastigote forms of Leishmania amazonensis and determined cytotoxicity in BALB/c peritoneal macrophages, as well as nitrite quantification, using the Griess method. Molecular docking was carried out to evaluate interactions of carajurin at the nitric oxide synthase enzyme. All compounds were active against promastigotes after 72 h, with IC50 values of 101.5 ± 0.06 µg/mL for ACCE and 4.976 ± 1.09 µg/mL for ACAF. Anthocyanidins carajurin, 3'-hydroxy-carajurone, and carajurone had IC50 values of 3.66 ± 1.16, 22.70 ± 1.20, and 28.28 ± 0.07 µg/mL, respectively. The cytotoxicity assay after 72 h showed results ranging from 9.640 to 66.74 µg/mL for anthocyanidins. ACAF and carajurin showed selectivity against intracellular amastigote forms (SI> 10), with low cytotoxicity within 24 h, a statistically significant reduction in all infection parameters, and induced nitrite production. Molecular docking studies were developed to understand a possible mechanism of activation of the nitric oxide synthase enzyme, which leads to an increase in the production of nitric oxide observed in the other experiments reported. These results encourage us to suggest carajurin as a biological marker of A. chica.

Cytotoxicity and selectiveness of Brazilian Piper species towards oral carcinoma cells.

BACKGROUND: Oral squamous cell carcinoma (OSCC) is one of the ten most common types of cancer worldwide. Plants of the genusPiper are used in traditional medicine to treat cancer, and they have a vast diversity of phytochemicals with cytotoxic potential. Purpose and Study Design: In this work, we analyzed the cytotoxic and selective potential of extracts and semipurified fractions of Piper mollicomum (PM), Piper truncatum (PT), Piper cernuum (PC), Piper arboreum (PA), and Piper cabralanum (PCa) using three different OSCC cell lines (SCC4, SCC9 and SCC25), and we measured their in vivo toxicities and conducted chemical analyses of their active fractions. RESULTS: The dichloromethane fractions of the crude methanolic extracts of the leaves of PM(-L-D), PC(-L-D) and PCa(-L-D) exhibited notable IC50 values of 94.2, 47.2 and 47.5 µg/mL, respectively, and all three of these extracts were more active than carboplatin (172.3 µg/mL). The most selective fraction was PC-L-D, which exhibited SI > 4.5; less than 5% hemolysis; and no significant alterations in in vivo acute toxicology. The major constituents in active fractions were lignans (PC-L-D and PCa-L-D) and chromenes (PM-L-D). CONCLUSION: PC-L-D demonstrated great potential for further development as an anticancer drug and could be the key to developing more effective and less toxic therapies against oral cancer.

Antifungal activities of the essential oil and its fractions rich in sesquiterpenes from leaves of Casearia sylvestris Sw.

Casearia genus (Salicaceae) is found in sub-tropical and tropical regions of the world and comprises about 160-200 species. It is a medicinal plant used in South America, also known as "guaçatonga", "erva-de-tiú", "cafezinho-do-mato". In Brazil, there are about 48 species and 12 are registered in the State of Rio de Janeiro, including Casearia sylvestris Sw. There are many studies related to the chemical profile and cytotoxic activities of extracts from these plants, although few studies about the antifungal potential of the essential oil have been reported. In this work, we have studied the antifungal properties of the essential oil of C. sylvestris leaves, as well as of their fractions, against four yeasts (Saccharomyces cerevisae, Candida albicans, C. glabrata and C. krusei) for the first time. The chemical analysis of the essential oil revealed a very diversified (n = 21 compounds) volatile fraction composed mainly of non-oxygenated sesquiterpenes (72.1%). These sesquiterpenes included α-humulene (17.8%) and α-copaene (8.5%) and the oxygenated sesquiterpene spathulenol (11.8%) were also identified. Monoterpenes were not identified. The fractions are mainly composed of oxygenated sesquiterpenes, and the most active fraction is rich in the sesquiterpene 14-hydroxy -9-epi-ß-caryophyllene. This fraction was the most effective in inhibiting the growth of three yeast strains.

Antifungal activities of the essential oil and its fractions rich in sesquiterpenes from leaves of Casearia sylvestris Sw

ABSTRACT Casearia genus (Salicaceae) is found in sub-tropical and tropical regions of the world and comprises about 160-200 species. It is a medicinal plant used in South America, also known as "guaçatonga", "erva-de-tiú", "cafezinho-do-mato". In Brazil, there are about 48 species and 12 are registered in the State of Rio de Janeiro, including Casearia sylvestris Sw. There are many studies related to the chemical profile and cytotoxic activities of extracts from these plants, although few studies about the antifungal potential of the essential oil have been reported. In this work, we have studied the antifungal properties of the essential oil of C. sylvestris leaves, as well as of their fractions, against four yeasts (Saccharomyces cerevisae, Candida albicans, C. glabrata and C. krusei) for the first time. The chemical analysis of the essential oil revealed a very diversified (n = 21 compounds) volatile fraction composed mainly of non-oxygenated sesquiterpenes (72.1%). These sesquiterpenes included α-humulene (17.8%) and α-copaene (8.5%) and the oxygenated sesquiterpene spathulenol (11.8%) were also identified. Monoterpenes were not identified. The fractions are mainly composed of oxygenated sesquiterpenes, and the most active fraction is rich in the sesquiterpene 14-hydroxy -9-epi-β-caryophyllene. This fraction was the most effective in inhibiting the growth of three yeast strains.

Chemical study and larvicidal activity against Aedes aegypti of essential oil of Piper aduncum L. (Piperaceae).

Piper aduncum L. is used in folk medicine to treat respiratory and inflammatory diseases. The aim of this study was to analyze the essential oil from leaves of P. aduncum collected in the Brazilian Cerrado, North of Minas Gerais, as well as to evaluate the larvicidal activity of this oil and of its major constituent. The essential oil was analyzed by gas chromatography coupled to flame ionization detector and gas chromatography coupled to mass spectrometry that allowed characterizing 23 compounds (monoterpenes: 90.4%; sesquiterpenes: 7.0%). The major component was 1,8-cineole (53.9%). This oil showed to be very different from those obtained from the same species. Larvae of A. aegypti were exposed to different concentrations of the essential oil and 1,8-cineole. The mortality rate of 100% was obtained after 24 h of treatment with the oil at concentrations of 500 and 1,000 ppm. After 48 h of treatment, the mortality rate was 80% and 50% for concentrations of 250 and 100 ppm, respectively. The LC50 obtained after 24h was estimated in 289.9 ppm and after 48 h was 134.1 ppm. The major compound 1,8-cineole showed no larvicidal activity.

Aristolactams from roots of Ottonia anisum (Piperaceae).

The Piperaceae species are known worldwide for its medicinal properties and its chemical compounds. In Brazil, many species of this family are distributed mainly in Amazon Region and in the Atlantic Forest. The genus Ottonia is known as source of amides, flavonoids, arilpropanoids and terpenes with record biological activities. Six aristolactams, including, aristolactam BII, piperolactam C, goniothalactam, stigmalactam, aristolactam AII and aristolactam BIII were isolated from roots of this species. GC-MS, 1H NMR and NOESY techniques were used to characterize these compounds. This is the first report about the occurrence of aristolactams in the Ottonia anisum Sprengel.

C-glycosyl flavones from Peperomia blanda.

The methanol extract from aerial parts of the Peperomia blanda (Piperaceae) yielded two C-glycosyl-flavones. Their structures were elucidated on the basis of extensive spectroscopic analysis, including 1D and 2D NMR, chemical transformation and comparison with the related known compounds. The structure of the new flavonoids were established as 4'-methoxy-vitexin 7-O-beta-D-xylopyranoside (1) (7-O-beta-D-xylopyranosyl-8-C-beta-D-glucopyranosyl-4'-methoxy-apigenin) and vicenin-2 (2). The antioxidant activity of both compounds was investigated using the DPPH assay. Both compounds showed only modest activity, with IC50 values of 357.2 microM for 1, and 90.5 microM for 2.

Unusual chromenes from Peperomia blanda.

From the methanol extract of the aerial parts of Peperomia blanda (Piperaceae), two chromenes were isolated and characterized mainly through application of 2D-NMR spectroscopy. The structures were 2S-(4-methyl-3-pentenyl)-6-formyl-8-hydroxy-2,7-dimethyl-2H-chromene and 2S-(4-methyl-3-pentenyl)-5-hydroxy-6-formyl-2,7-dimethyl-2H-chromene named as blandachromenes I and II, respectively.

Effect of leaf essential oil from Piper solmsianum C.DC. in mice behaviour

The essential oil from Piper solmsianum leaves and its major compound (sarisan) were tested to verify their influences upon mice behaviour. The essential oil was obtained by hydrodistillation in a modified Clevenger extractor and analysed by GC/ MS. This analysis revealed in the oil the presence of monoterpenes, sesquiterpenes and of arylpropanoids. The compound sarisan, a myristicin analogue, was isolated from the oil to perform the pharmacological tests. Emulsions of the oil and of sarisan (5.0 and 10.0 percent v/v) were used in the tests. Pentobarbital (30 mg/ kg s.c.) or diazepam (2.5 mg/ kg s.c.) were tested as standard drugs to verify depressant or anxiolytic effects, respectively. Both essential oil and sarisan showed to have exciting and depressant effects in the tested animals