Molecular networking-based dereplication of strictosidine-derived monoterpene indole alkaloids from the curare ingredient Strychnos peckii.

RATIONALE: Monoterpene indole alkaloids (MIAs) are a large group of biologically active compounds produced by hundreds of plant species in numerous plant families, such as Apocynaceae, Loganiaceae and Rubiaceae. Although this diversity is biosynthetically intermediated by strictosidine, there are no works focused on the fragmentation patterns under collision-induced dissociation of strictosidine-derived alkaloids. METHODS: Initially, the alkaloid fingerprint of Strychnos peckii was established using leaf spray with tandem mass spectrometry (LS-MS/MS). Then, high-performance liquid chromatography coupled to tandem mass spectrometry (HPLC/MS/MS) analyses were carried out to focus on the patterns of neutral losses in product ion scan experiments with the leaf aqueous extract. Finally, the product ion spectra from a set of presumable strictosidine-type derivatives were analyzed and organized via molecular networking (MN), and dereplicated by manual interpretation of MS/MS spectra. RESULTS: LS-MS/MS allowed the tentative identification of strictosidine-derived alkaloids in the leaves of S. peckii, showing useful neutral losses for the dereplication of strictosidine analogues by HPLC/MS/MS experiments. The use of MN combined with manual interpretation of the fragmentation patterns highlighted characteristic fragmentation pathways, and allowed the tentative identification of strictosidine, desoxycordifoline, strictosidinic acid, 10-hydroxystrictosidine, 5-carboxystrictosidine, lyaloside, 3,4-dehydrostrictosidine and strictosidine lactam. CONCLUSIONS: The use of MN combined with the analysis of the fragmentation patterns proved to be a useful strategy for the dereplication of strictosidine-derived MIAs from S. peckii, highlighting known and unprecedented structures, as well as useful diagnostic product ions. Therefore, this workflow is an effective approach for the characterization of strictosidine-type alkaloids in future dereplication works.

Assessment of the phenolic content, mutagenicity and genotoxicity of ethanolic extracts of stem bark and leaves from Strychnos pseudoquina A. St.-hil.

Strychnos pseudoquina is a plant species whose stem bark is used as bitter tonic beverage. The phytochemical analysis, as well as quantification of phenolic constituents and antioxidant activity of ethanolic extracts from S. pseudoquina stem bark, and leaves were conducted. The extracts were tested for mutagenicity (Ames test) and DNA-damaging activity (Plasmid Cleavage test). Leaves recorded the largest amount of flavonoids. The performed high-performance liquid chromatography (HPLC) showed flavonoids such as isorhamnetin and strychnobiflavone (phytochemical markers of the investigated species) in stem barks, but not in leaves. The proanthocyanidin content and antioxidant activity were significantly higher in stem barks than in leaves. Stem bark and leaf extracts presented mutagenic activity against TA98 and TA100 strains with, and without, metabolic activation (S9). The Plasmid Cleavage test did not indicate DNA-damaging activity. Our results suggest that extracts deriving from S. pseudoquina should be used with extreme caution, mainly the stem bark extract, which is widely used in folk medicine.

Strychnos pseudoquina modulates the morphological reorganization of the scar tissue of second intention cutaneous wounds in rats.

Natural substances are used in folk medicines to treat injuries. Strychnos pseudoquina has scarring, antipyretic, and antimalarial actions. The present study aimed to analyze the effect of S. pseudoquina on cutaneous wound healing in rats. The S. pseudoquina extract was submitted to phytochemical prospection. The levels of flavonoids and total phenolic compounds in the extract were 50.7 mg/g and 2.59 mg/g, respectively. Thirty Wistar rats were individualized in cages with food and water ad libitum (registration no. 730/2014). After anesthesia, three circular wounds (12mm diameter) were made in the animals, which were randomly separated into five treatments: Sal, saline; VO, ointment vehicles (lanolin and Vaseline); SS, positive control (silver sulfadiazine 1%); LE 5, freeze-dried extract 5%; and LE 10, lyophilized extract 10%. The animals were treated with the ointment daily for 21 days. Every seven days, the area and the rate of wound contraction were evaluated. Tissue samples were removed for histopathological analysis of the number of mast cells, elastic and collagen fibers, and biochemical analyses, quantification of malondialdehyde (MDA), carbonylated proteins (PCN), superoxide dismutase (SOD), catalase (CAT), transforming growth factor ß (TGF-ß), Interleukin 10 (IL-10) and tumor necrosis factor (TNF). The number of mast cells, collagen and elastic fibers in the rat wounds were higher in the treatments with the plant. The extract also stimulated the activity of antioxidant enzymes, particularly SOD, presenting high levels, and maintained low levels of PCN. The TGF-ß and IL-10 concentration was higher in the LE5 and LE10 treatment of the extract than in the Sal, OV and SS treatments on day 7. The ointment based on S. pseudoquina closed the wound faster and accelerated wound healing in animals.

Hydroethanolic Extract of Strychnos pseudoquina Accelerates Skin Wound Healing by Modulating the Oxidative Status and Microstructural Reorganization of Scar Tissue in Experimental Type I Diabetes.

The effect of topical application of ointment based on Strychnos pseudoquina hydroethanolic extract in the cutaneous wounds healing in diabetic rats was evaluated. Samples of S. pseudoquina were submitted to phytochemical prospection and in vitro antioxidant assay. Thirty Wistar rats were divided into 5 groups: Sal-wounds treated with 0.9% saline solution; VH-wounds treated with 0.6 g of lanolin cream (vehicle); SS-wounds treated with silver sulfadiazine cream (10 mg/g); ES5- and ES10-wounds treated with an ointment of S. pseudoquina extract, 5% and 10%, respectively. Fragments of wounds were removed for histological and biochemical analysis every 7 days during 21 days. ES showed equivalent levels per gram of extract of total phenols and flavonoids equal to 122.04 mg for TAE and 0.60 mg for RE. The chlorogenic acid was one of the major constituents. S. pseudoquina extract presented high antioxidant potential in vitro. ES5 and ES10 showed higher wound healing rate and higher amount of cells, blood vessels, and type III and I collagen. The oxidative stress markers were lower in the ES5 and ES10 groups, while the antioxidants enzymes levels were higher. Ointment based on S. pseudoquina extract promotes a fast and efficient cutaneous repair in diabetic rats.

Strychnos pseudoquina A. St. Hil.: a Brazilian medicinal plant with promising in vitro antiherpes activity.

AIMS: To investigate the anti-HSV and anti-inflammatory effects of a standardized ethyl acetate extract (SEAE) prepared with the stem bark of Strychnos pseudoquina, along with two isolated compounds: quercetin 3-O-methyl ether (3MQ) and strychnobiflavone (SBF). METHODS AND RESULTS: The mechanisms of action were evaluated by different methodological strategies. SEAE and SBF affected the early stages of viral infection and reduced HSV-1 protein expression. Both flavonoids elicited a concentration-dependent inhibition of monocyte chemoattractant protein-1 (MCP-1), whereas 3MQ reduced the chemokine release more significantly than SBF. Conversely, both compounds stimulated the production of the cytokines TNF-α and IL-1-ß in LPS-stimulated cells, especially at the intermediate and the highest tested concentrations. CONCLUSIONS: SEAE and SBF interfered with various steps of HSV replication cycle, mainly adsorption, postadsorption and penetration, as well as with ß and γ viral proteins expression; moreover, a direct inactivation of viral particles was observed. Besides, both flavonoids inhibited MCP-1 selectively, a feature that may be beneficial for the development of new anti-HSV agents. SIGNIFICANCE AND IMPACT OF THE STUDY: The results indicated that the samples present anti-HSV and anti-inflammatory activities, at different levels, which is an interesting feature since cold and genital sores are accompanied by an inflammation process.

A new indole-alkaloid and a new phenolic-glycoside with cytotoxic activity from Strychnos fendleri.

A new indole alkaloid strychnosinol (1) and a new phenolic-glycoside (2) were isolated from the bark and leaves of Strychnos fendleri Sprague & Sandwith, together with six known compounds reported for the first time in this species. The structures of these compounds were determined on the basis of spectroscopic data; mainly those obtained by using (1)H and (13)C NMR (1D and 2D) and mass spectrometry. Strychnosinol (1) and the phenolic glycoside (2) together with compounds 3-8 were evaluated for cytotoxicity against a panel of five tumour cell lines; IC50 values between 0.090 and 0.227 µM for the human tumour cell lines were observed for compound 2.

Antileishmanial activity and evaluation of the mechanism of action of strychnobiflavone flavonoid isolated from Strychnos pseudoquina against Leishmania infantum.

The present study aimed to investigate the in vitro antileishmanial activity of strychnobiflavone flavonoid against Leishmania infantum, as well as its mechanism of action, and evaluate the ex vivo biodistribution profile of the flavonoid in naive BALB/c mice. The antileishmanial activity (IC50 value) of strychnobiflavone against stationary promastigote and amastigote-like stages of the parasites was of 5.4 and 18.9 µM, respectively; with a 50% cytotoxic concentration (CC50) value of 125.0 µM on murine macrophages, resulting in selectivity index (SI) of 23.2 and 6.6, respectively. Amphotericin B, used as a positive control, presented SI values of 7.6 and 3.3 for promastigote and amastigote-like stages of L. infantum, respectively. The strychnobiflavone was also effective in reducing in significant levels the percentage of infected macrophages, as well as the number of amastigotes per macrophage, after the treatment of infected macrophages using the flavonoid. By using different fluorescent probes, we investigated the bioenergetics metabolism of L. infantum promastigotes and demonstrated that the flavonoid caused the depolarization of the mitochondrial membrane potential, without affecting the production of reactive oxygen species. In addition, using SYTOX(®) green as a fluorescent probe, the strychnobiflavone demonstrated no interference in plasma membrane permeability. For the ex vivo biodistribution assays, the flavonoid was labeled with technetium-(99m) and studied in a mouse model by intraperitoneal route. After a single dose administration, the scintigraphic images demonstrated a highest uptake by the liver and spleen of the animals within 60 min, resulting in low concentrations after 24 h. The present study therefore demonstrated, for the first time, the antileishmanial activity of the strychnobiflavone against L. infantum, and suggests that the mitochondria of the parasites may be the possible target organelle. The preferential distribution of this compound into the liver and spleen of the animals could warrant its employ in the treatment of visceral leishmaniasis.

Can a Strychnos species be used as antiulcer agent? Ulcer healing action from alkaloid fraction of Strychnos pseudoquina St. Hil. (Loganiaceae).

ETHNOPHARMACOLOGICAL RELEVANCE: Strychnos pseudoquina St. Hil. (Loganiaceae) is one Brazilian native medicinal species described in the first edition of the Brazilian Official Pharmacopoeia in 1929. This medicinal plant, popularly known as "quina-quina", "quina-branca" or "casca aromatica was very commonly used in folk medicine in tea form obtained from the bark and/or leaves as tonic, antipyretic, antimalarial and mainly against diseases of the liver, spleen and stomach. AIM OF THE STUDY: Previous study already characterized the gastroprotective action of this species The aim of the present study is to elucidate the mechanism of the healing process mediated by the methanolic extract (ME) and their enriched alkaloid fraction (EAF) from Strychnos pseudoquina in chronic gastric ulceration induced by 5% acetic acid in rats, an experimental model that accurately reflects human gastrointestinal disease. MATERIAL AND METHODS: The ME and EAF was administered orally in a single dose (based on previously study of dose-response curve) for 14 days after chronic ulceration was induced in rats. The healing effect of ME and EAF was evaluated by macroscopic and morphometric analyses, immunohistochemical assay (PCNA and SOD) and anti-Helicobacter pylori effect was evaluated by in vitro assay. RESULTS: Our results demonstrated that EAF significantly reduced border internal (42%) and external (38%) lesion area (mm(2)) by macroscopic analyses (P<0.05). Animals treated with EAF stimulated some proliferative factors by increasing the height of epithelial regenerative area and the expression of PCNA-positive nuclei. The number of vessels in gastric mucosa of rats treated with EAF reveals an expressive increase (4 times more than vehicle treatment) of vessels that stimulate cells proliferation in the healing region. These results suggest that the recovery of vascularization of the ulcerated area is involved in the healing action of alkaloid fraction of Strychnos pseudoquina. The MIC (minimum inhibitory concentration) of 75 µg/ml from EAF showed an effective in vitro anti-Helicobacter pylori action of this fraction. EAF also was quite effective in the process of SOD release that is an important protective factor against bacterial agents. The efficacy of EAF was accomplished safely without presenting any alteration of toxicological parameters during 14 day of treatment. CONCLUSIONS: The expressive gastric healing effect by increasing cellular proliferation together with expression of SOD activity and antibacterial action against Helicobacter pylori confirm the efficacy of this species in heal gastric mucosa and these results are a important contribution to the knowledge of a crude drug presents at the Brazilian Official Pharmacopoeia since 1929.

Evaluation of hypoglicemic activity and healing of extract from amongst bark of "Quina do Cerrado" (Strychnos pseudoquina ST. HILL) / Avaliação da atividade hipoglicemiante e cicatrizante de extrato da entrecasca da Quina do Cerrado (Strychnos pseudoquina ST. HILL)

PURPOSE: Evaluation the hypoglycemic and healing effects of the Strychnos pseudoquina. METHODS: 33 Wistar rats were divided in the following groups, group 1 not sensitized with alloxan and untreated by aqueous extracts; group 2 sensitized with aloxana and untreated by aqueous extracts of Quina; group 3 sensitized by aloxana ad treated with the aqueous extract of quina. Diabetes was induced by alloxan diabetogenic drugs at a dose of 42 mg/kg of weight. The glycemias was evaluated by glycemic measuring Accu-check®. To verify the healing, was made a longitudinal cut of 1 cm in the back of the mouse. The group 3 was treated with a microemulsion containing Strychnos pseudoquina. Wounds were macroscopically evaluated during pre-determined days after the cut (1st, 3rd, 7th, 9th, 14th). RESULTS: The glycemia levels in the group treated with Strychnos pseudoquina were lower than the group sensitized by alloxan and not treated with Strychnos pseudoquina. There was no difference between the healing of the wounds treated with quina and another groups. CONCLUSIONS: Strychnos pseudoquina presents hypoglycemic effect. Meanwhile the topical use of the microemulsion of Strychnos pseudoquina presents no macroscopically significant effect on the healing of wounds in diabetic rats.

OBJETIVO: Avaliar o efeito hipoglicemiante e cicatrizante de Strychnos pseudoquina. MÉTODOS: Utilizou-se 33 ratos Wistar, divididos nos seguintes grupos: grupo 1 não sensibilizado pela aloxana e não tratado pelo extrato aquoso; grupo 2 sensibilizado com aloxana e não tratado pelo extrato aquoso de Quina; grupo 3 sensibilizado pela aloxana, tratado com o extrato aquoso de Quina. O diabetes foi induzido pela aloxana, droga diabetôgenica, na dose de 42mg/Kg de peso de rato. As glicemias foram avaliadas pelo medidor de glicemia Accu-check®. Para verificar a cicatrização, foi realizado um corte de 1 cm longitudinal na região dorsal do rato, sob anestesia inalatória. O grupo 3 foi tratado com uma microemulsão contendo "Quina do Cerrado". As feridas foram avaliadas macroscopicamente nos períodos pré-determinados (1º, 3°, 7°, 9º e 14° dias após o corte). RESULTADOS: Os níveis glicêmicos do grupo com "Quina do Cerrado" foram menores que os do grupo sensibilizado pela aloxana e não tratados. Não houve diferença entre a cicatrização das feridas tratadas com Quina comparada à dos outros grupos. CONCLUSÕES: Strychnos pseudoquina apresenta efeito hipoglicemiante. Entretanto o seu uso tópico em microemulsão não apresenta efeito significativo na cicatrização de feridas em ratos diabéticos.

Evaluation of hypoglicemic activity and healing of extract from amongst bark of "Quina do Cerrado" (Strychnos pseudoquina ST. HILL).

PURPOSE: Evaluation the hypoglycemic and healing effects of the Strychnos pseudoquina. METHODS: 33 Wistar rats were divided in the following groups, group 1 not sensitized with alloxan and untreated by aqueous extracts; group 2 sensitized with aloxana and untreated by aqueous extracts of Quina; group 3 sensitized by aloxana ad treated with the aqueous extract of quina. Diabetes was induced by alloxan diabetogenic drugs at a dose of 42 mg/kg of weight. The glycemias was evaluated by glycemic measuring Accu-check. To verify the healing, was made a longitudinal cut of 1 cm in the back of the mouse. The group 3 was treated with a microemulsion containing Strychnos pseudoquina. Wounds were macroscopically evaluated during pre-determined days after the cut (1st, 3rd, 7th, 9th, 14th). RESULTS: The glycemia levels in the group treated with Strychnos pseudoquina were lower than the group sensitized by alloxan and not treated with Strychnos pseudoquina. There was no difference between the healing of the wounds treated with quina and another groups. CONCLUSIONS: Strychnos pseudoquina presents hypoglycemic effect. Meanwhile the topical use of the microemulsion of Strychnos pseudoquina presents no macroscopically significant effect on the healing of wounds in diabetic rats.

Isolation of cytotoxic metabolites from targeted peruvian amazonian medicinal plants.

The antiproliferative bioassay-guided fractionation of five Peruvian plants, Doliocarpus dentatus, Picramnia sellowii, Strychnos mitscherlichii, Iryanthera juruensis, and Croton alnifolius, led to the isolation and identification of their different major cytotoxic constituents, betulinic acid (1), nataloe-emodin (2), bisnordihydrotoxyferine (4), 2',4'-dihydroxy-6'-methoxy-3,4-methylenedioxydihydrochalcone (5), and 2',4'-dihydroxy-4,6'-dimethoxydihydrochalcone (6) and 12-O-tetradecanoylphorbol-13-acetate (7), respectively. Eight human tumor cell lines and two nontumorigenic cell lines were used in this investigation. Their in vitro activity against Mycobacterium tuberculosis is also reported.

Assessment of DNA damage by extracts and fractions of Strychnos pseudoquina, a Brazilian medicinal plant with antiulcerogenic activity.

Strychnos pseudoquina St. Hil. is a native plant of the Brazilian Savannah, used in popular medicine to treat a number of conditions. Since it contains large quantities of alkaloids with proven antiulcer activity, we tested the genotoxic potential of crude extracts and fractions containing alkaloids and flavonoids from the leaves of this plant, on Salmonella typhimurium and performed the micronucleus test on peripheral blood cells of mice treated in vivo. The results showed that the methanol extract of the leaves of S. pseudoquina is mutagenic to the TA98 (-S9) and TA100 (+S9, -S9) strains of Salmonella. The dichloromethane extract was not mutagenic to any of the tested strains. Fractions enriched with alkaloids or flavonoids were not mutagenic. In vivo tests were done on the crude methanol extract in albino Swiss mice, which were treated, by gavage, with three different doses of the extract. The highest dose tested (1800 mg/kgb.w.) induced micronuclei after acute treatment, confirming the mutagenic potential of the methanol extract of the leaves of S. pseudoquina. In high doses, constituents of S. pseudoquina compounds act on DNA, causing breaks and giving rise to micronuclei in the blood cells of treated animals.

Evaluation of Strychnos pseudoquina ST. HIL. leaves extract on gastrointestinal activity in mice.

Strychnos pseudoquina ST. HIL. (Loganiaceae) was investigated for its ability to protect the gastric mucosa against injuries caused by non-steroidal anti-inflammatory drugs (piroxicam) and a necrotizing agent (HCl/EtOH) in mice. The MeOH extract and enriched alkaloidic fraction (EAF) provided significant protection in experimental models wheer used at doses of 250 and 1000 mg/kg. In vivo tests were carried out to evaluate for possible toxic effects and no mortality was observed up to the 5 g/kg dose level. Phytochemical investigation led to the isolation of a new indole alkaloid, which elucidated the observed pharmacological effects.

A new strychnobrasiline base of Strychnos mattogrossensis.

A new strychnobrasiline derivative, 12-hydroxy-10,11-dimethoxystrychnobrasiline 1, has been isolated from the stem barks of Strychnos mattogrossensis. Its structure was established by spectroscopy analysis (1D and 2D NMR, MS, IR, UV) and literature data comparison. Two other known indoline alkaloids were also obtained from the heartwood, 12-hydroxy-11-methoxystrychnobrasiline 2 and strychnobrasiline 3.

On the alkaloids of Strychnos. XXXV. The occurrence of akagerine in South American Strychnos.

Akagerine (3) was isolated from three different species of South American Strychnos: from S. gardneri A. D.C. (Rio de Janeiro, Brazil), together with 11-methoxydiaboline (2); from S. jobertiana Baillon (Brazil), together with diaboline (1): and from S. parvifolia D.C. (Bahia, Brazil).