Neuroprotective effect of Bouvardia ternifolia (Cav.) Schltdl via inhibition of TLR4/NF-κB, caspase-3/Bax/Bcl-2 pathways in ischemia/reperfusion injury in rats.

Introduction: Bouvardia ternifolia is a plant known for its traditional medicinal uses, particularly in treating inflammation and oxidative stress. Recent studies have explored its potential in neuroprotection, especially in the context of cerebral ischemia/reperfusion injury, a condition where blood supply returns to the brain after a period of ischemia, leading to oxidative stress and inflammation. This damage is a major contributor to neuronal death and neurodegenerative diseases. Methods: A BCCAO/reperfusion model was induced, followed by treatment with B. ternifolia extract. Various molecular biology methods were employed, including Western blot analysis, gene expression assessment via RT-qPCR, and the measurement of oxidative stress mediators. Results: In the BCCAO/reperfusion model, the compounds in the dichloromethane extract work by targeting various signaling pathways. They prevent the activation of iNOS and nNOS, reducing harmful reactive oxygen and nitrogen species, and boosting antioxidant enzymes like catalase and superoxide dismutase. This lowers oxidative stress and decreases the expression of proteins and genes linked to cell death, such as Bax, Bcl-2, and caspase-3. The extract also blocks the TLR4 receptor, preventing NF-κB from triggering inflammation. Additionally, it reduces the activation of microglia and astrocytes, as shown by lower levels of glial activation genes like GFAP and AiF1. Conclusion: The dichloromethane extract of B. ternifolia demonstrated significant neuroprotective effects in the BCCAO/reperfusion model by modulating multiple signaling pathways. It effectively reduced oxidative stress, inhibited inflammation, and attenuated apoptosis, primarily through the downregulation of key proteins and genes associated with these processes. These findings suggest that the extract holds therapeutic potential for mitigating ischemia/reperfusion-induced neuronal damage.

An Organic Fraction of Oenothera rosea L'Her Ex. Aiton Prevents Neuroinflammation in a Rat Ischemic Model.

BACKGROUND: Oenothera rosea L'Her Ex. Aiton, presenting antioxidant and anti-inflammatory activities, is traditionally used to treat bruises and headaches and as a healing agent. This study aimed to investigate whether its organic fraction (EAOr) has neuroprotective properties against neuroinflammation in the context of ischemia/reperfusion. METHODS: The chemical composition of EAOr was determined using HPLC techniques, and its neuroprotective activities were evaluated in a common carotid-artery ligation model for the induction of ischemia/reperfusion (I/R). The animals were supplemented with EAOR for 15 days. On the last day, the animals were rested for one hour, following which the common carotid-artery ligation procedure was performed to induce I/R. The neurological deficit was evaluated at 24 h after I/R using Bederson's scale, and the relative expression of inflammatory genes and structure of hippocampal neurons were analyzed at 48 h. RESULTS: The chemical analysis revealed five major compounds in EAOr: gallic acid, rutin, ellagic acid, and glucoside and rhamnoside quercetin. EAOr prevented neurological deficit 24 h after I/R; led to the early activation of the AIF and GFAP genes; reduced Nfkb1, IL-1beta, Il-6 and Casp3 gene expression; and protected hippocampal neurons. CONCLUSIONS: Our findings demonstrate that EAOr contains polyphenol-type compounds, which could exert a therapeutic effect through the inhibition of neuroinflammation and neuronal death genes, thus maintaining hippocampal neurons.

Etiology of Foliar Blight of Indian Paintbrush (Castilleja tenuiflora) in Mexico.

Castilleja tenuiflora is a native perennial plant used in traditional Mexican medicine. In June 2022, leaf blight symptoms were observed in a wild population of C. tenuiflora plants. Disease incidence was 80% and disease intensity reached up to 5% of the leaf area. Currently, there are no reports of pathogens causing leaf blight in this plant; therefore, this work aimed to identify the fungi responsible for the disease. The fungi recovered from the diseased tissue were characterized by means of pathogenicity tests and cultural, morphological, and molecular characterization. The information obtained revealed that Alternaria alternata and Alternaria gossypina are the pathogens responsible for the disease. This is the first report implicating species of Alternaria in causing leaf blight of C. tenuiflora in Mexico, as well as the first report of Alternaria gossypina also in Mexico. These pathogens may threaten the in situ conservation of native C. tenuiflora populations and limit their in vitro propagation. Future research lines should focus on determining the effect of these pathogens on metabolite production.

Induction of Monoterpenoid Oxindole Alkaloids Production and Related Biosynthetic Gene Expression in Response to Signaling Molecules in Hamelia patens Plant Cultures.

Hamelia patens (Rubiaceae), known as firebush, is a source of bioactive monoterpenoid oxindole alkaloids (MOAs) derived from monoterpenoid indole alkaloids (MIAs). With the aim of understanding the regulation of the biosynthesis of these specialized metabolites, micropropagated plants were elicited with jasmonic acid (JA) and salicylic acid (SA). The MOA production and MIA biosynthetic-related gene expression were evaluated over time. The production of MOAs was increased compared to the control up to 2-fold (41.3 mg g DW-1) at 72 h in JA-elicited plants and 2.5-fold (42.4 mg g DW-1) at 120 h in plants elicited with SA. The increment concurs with the increase in the expression levels of the genes HpaLAMT, HpaTDC, HpaSTR, HpaNPF2.9, HpaTHAS1, and HpaTHAS2. Interestingly, it was found that HpaSGD was downregulated in both treatments after 24 h but in the SA treatment at 120 h only was upregulated to 8-fold compared to the control. In this work, we present the results of MOA production in H. patens and discuss how JA and SA might be regulating the central biosynthetic steps that involve HpaSGD and HpaTHAS genes.

Cyclic Hexapeptide from Bouvardia ternifolia (Cav.) Schltdl. and Neuroprotective Effects of Root Extracts.

Bouvardia ternifolia (Cav.) Schltdl. is a shrub that belongs to the Rubiaceae family and is distributed throughout México; it has been used for its antioxidant, neuroprotective, and anti-inflammatory properties. This work aimed to evaluate the protective effects of B. ternifolia root extracts on the blood-brain barrier and the positive regulation of cytokines IL-1ß, IL-6, and TNF-α, and the characterization of compounds present in the dichloromethane (BtD) and hexane (BtH) extracts. Male ICR mice were orally administered with B. ternifolia extracts for 5 days before a single injection of LPS. Administration of BtH and BtD significantly decreased Evans blue leakage into brain tissue by 70% and 68%, respectively. Meloxicam (MX) decreased the concentration of IL-1ß by 39.6%; BtM by 53.9%; BtAq by 48.4%; BtD by 31.9%, and BtH by 37.7%. BtH was the only treatment that significantly decreased the concentration of IL-6 by 32.2%. The concentration of TNF-α declined with each of the treatments. The chemical composition of BtD and BtH was characterized by GC-MS, and the cyclic hexapeptide was identified by 13C, 1H NMR, and two-dimension techniques. In the BtD extract, seven compounds were found and in BtH 13 compounds were found. The methanolic (BtM) and aqueous (BtAq) extracts were not subjected to chemical analysis, because they did not show a significant difference in the BBB protection activity. Therefore, the results suggested that the extracts BtD and BtH protect the blood-brain barrier, maintaining stable its selective permeability, thereby preventing LPS from entering the brain tissue. Simultaneously, they modulate the production of IL-1ß, IL-6, and TNF-α. It is important to note that this research only evaluated the complete extracts.

Mixed elicitation with salicylic acid and hydrogen peroxide modulates the phenolic and iridoid pathways in Castilleja tenuiflora plants.

MAIN CONCLUSION: SA and H2O2, in single and mixed elicitation stimulate specialized metabolism and activate oxidative stress in C. tenuiflora plants. Single elicitation with salicylic acid (SA at 75 µM) and, hydrogen peroxide (at 150 µM), and mixed elicitation (75 µM SA + 150 µM H2O2) were evaluated on specialized metabolism in Castilleja tenuiflora Benth. plants. Total phenolic content (TPC), phenylalanine ammonia-lyase (PAL) activity, antioxidant enzymes and specialized metabolite profiles, as well as the expression levels of eight genes involved in phenolic (Cte-TyrDC, Cte-GOT2, Cte-ADD, Cte-AO3, Cte-PAL1, Cte-CHS1) and terpene pathways (Cte-DXS1 and Cte-G10H) and their correlation with major metabolite (verbascoside and aucubin) concentrations were investigated. TPC content (three-fold) and PAL activity (11.5-fold) increased with mixed elicitation, as well as catalase and peroxidase activity (11.3-fold and 10.8-fold, respectively), compared to single elicitation. Phenylethanoid accumulation was greatest under mixed elicitation, followed by SA and H2O2. Lignan accumulation was differential, depending on the plant part and the elicitor. Flavonoids only appeared after mixed elicitation. The high concentration of verbascoside under mixed elicitation was related to a high gene expression. Single elicitation induced iridoid accumulation in specific parts (H2O2 in aerial parts and SA in roots), whereas under mixed elicitation, it accumulated in both parts. A high concentration of aucubin in the aerial part was related to a high expression level of genes of the terpene pathway Cte-DXS1 and Cte-G10H, and in the root with Cte-G10H, while Cte-DXS1 was downregulated in this tissue in all treatments. Mixed elicitation with SA and H2O2 represents an interesting tool to increase the production of specialized metabolites in plants.

A Mixture of Kaempferol-3-O-sambubioside and Kaempferol-3-O-sophoroside from Malvaviscus arboreus Prevents Ethanol-Induced Gastric Inflammation, Oxidative Stress, and Histologic Changes.

Malvaviscus arboreus is used in traditional Mexican medicine to treat gastrointestinal diseases. Therefore, a mixture of Kaempferol-O-sambubioside and Kaempferol-O-sophoroside (MaSS) isolated from flowers of this species was tested as a preventive treatment on gastric lesions induced with ethanol in rats. MaSS was obtained by chromatographic methods and administered by oral pathway to male Sprague Dawley rats with ethanol-induced gastric lesions. Pretreatment with MaSS at doses of 30, 90, 120, and 180 mg/kg significantly prevents gastric lesions, inhibits the increment in relative stomach weight (%) in gastric IL-6, and also provokes an increment of IL-10 concentration and catalase activity. Finally, MaSS prevented edema in the mucosa and submucosa and diminished microscopic gastric lesions provoked by ethanol.

Anti-arthritic and anti-inflammatory effects of Baccharis conferta Kunth in a kaolin/carrageenan-induced monoarthritis model.

ETHNOPHARMACOLOGICAL RELEVANCE: Popularly known as "escoba" (broom) or "escobilla china" (Chinese brush), Baccharis conferta Kunth (Asteraceae), is a plant widely used in Mexican folk medicine for alleviating muscular and rheumatic pain. A recent study described that dichloromethane extract as well as fractions and isolated compounds, possess anti-inflammatory activity in TPA-induced acute edema. AIM OF THE STUDY: Based on the popular medicinal uses of B. conferta as well as previous studies on its anti-inflammatory activity, the aim of this research was to evaluate the anti-arthritic and anti-inflammatory effects of dichloromethane extract, fractions, and compounds from B. conferta in a monoarthritis model induced with kaolin/carrageenan (K/C). MATERIALS AND METHODS: Aerial parts of B. conferta were collected, dried, and macerated with dichloromethane. The dichloromethane extract (BcD) was separated by open column chromatography to obtain the BcD2 fraction where the diterpene kingidiol (KIN) was isolated and from the BcD3 fraction the flavonoid cirsimaritin (CIR), which are the most active compounds in the TPA model. In addition, the flavonoids acacetin, pectolinaringenin and 6-methoxykaempferide were identified and isolated from the BcD2 fraction. The content of the main compounds was estimated in BcD, BcD2 and BcD3. The anti-arthritic and anti-inflammatory effects of B. conferta were investigated by evaluating ankle joint inflammation, hyperalgesia using the hot plate test, and pro- and anti-inflammatory cytokine levels in the synovial capsule as well as histological changes in ankle joint tissue in a monoarthritis model induced with K/C in Balb/c mice. RESULTS: Oral administration of BcD2 fraction (25 mg/kg) and KIN (10 mg/kg) reduced the ankle thickness induced by K/C and decreased the levels of TNF-α, IL-1ß, IL-6 and IL-17, while BcD2 increased IL-10. In addition, BcD2 and KIN showed significant edema attenuation of the synovial membrane and decreased inflammatory infiltration and cartilage erosion compared to the VEH group. Finally, BcD (50 mg/kg), KIN (10 mg/kg) and CIR (5 mg/kg) decreased hyperalgesia. CONCLUSIONS: B. conferta constitutes a therapeutic or preventive candidate for osteoarthritis, because of decreased articular inflammation and pain accompanied with the modulation of cytokine concentrations, which confirms the anti-arthritic and anti-inflammatory activities of B. conferta and support its popular use.

New Chromones from Bouvardia ternifolia (Cav.) Schltdl with Anti-Inflammatory and Immunomodulatory Activity.

The extract, fractions, and compounds of the Bouvardia ternifolia root were evaluated as an antiarthritic using a complete Freund's adjuvant (CFA) model in mice and NF-κB inhibition in RAW 264.7 macrophages. Four active compounds, including two new compounds, ternifoliol and ternifolial, were isolated by open column chromatography and identified by spectroscopic and spectrometric techniques, resulting in benzochromone-like structures with aromatic rings and hydroxyl groups, which could be responsible for the anti-inflammatory activity and inhibitory NF-κB. Changes in the joint cytokine profile monitored the antiarthritic effect. A decrement was observed in the local concentration of the following cytokines with different treatments: IL-17 by 64% and 70.3% with the aqueous extract (BtAq), ethyl acetate extract (BtAcOEt), and M3 fraction; interleukin-1 beta (IL-1ß) by 10.2% and 15.7% with BtAq and the M4 fraction, respectively; IL-6 with M1, M2, M3, and M4 between 42% and 64%; necrosis factor-alpha (TNF-α) by 60.9% with M4. Conversely, the anti-inflammatory cytokine interleukin-10 (IL-10) increased between 94% and 99% with M1, M2, M3, and M4. Kidney IL-6 decreased with BtAq, M1, M2, M3, and M4 between 68.9% and 85.8%. TNF-α decreased with BtAcOEt, BtAq, M1, M2, and M4 between 34% and 80.2%. The NF-κB pathway was inhibited with BtAcOEt (90.1%), M1 (85%), M2 (93.5%), M3 (84.5%), M4 (90.3%), ternifoliol (75.6%), bouvardin (20.4%), and scopoletin (89%). We conclude that B. ternifolia modulated the inflammatory response at the joint and kidney levels and the NF-κB pathway.

In Vitro Bile Salt Hydrolase (BSH) Activity Screening of Different Probiotic Microorganisms.

Bile salt hydrolase (BSH) activity in probiotic strains is usually correlated with the ability to lower serum cholesterol levels in hypercholesterolemic patients. The objective of this study was the evaluation of BSH in five probiotic strains of lactic acid bacteria (LAB) and a probiotic yeast. The activity was assessed using a qualitative direct plate test and a quantitative high-performance thin- layer chromatography assay. The six strains differed in their BSH substrate preference and activity. Lactobacillus plantarum DGIA1, a potentially probiotic strain isolated from a double cream cheese from Chiapas, Mexico, showed excellent deconjugation activities in the four tested bile acids (69, 100, 81, and 92% for sodium glycocholate, glycodeoxycholate, taurocholate, and taurodeoxycholate, respectively). In the case of the commercial probiotic yeast Saccharomyces boulardii, the deconjugation activities were good against sodium glycodeoxycholate, taurocholate, and taurodeoxycholate (100, 57, and 63%, respectively). These last two results are part of the novelty of the work. A weak deconjugative activity (5%) was observed in the case of sodium glycocholate. This is the first time that the BSH activity has been detected in this yeast.

Synergism and Subadditivity of Verbascoside-Lignans and -Iridoids Binary Mixtures Isolated from Castilleja tenuiflora Benth. on NF-κB/AP-1 Inhibition Activity.

Pharmacodynamic interactions between plant isolated compounds are important to understand the mode of action of an herbal extract to formulate or create better standardized extracts, phytomedicines, or phytopharmaceuticals. In this work, we propose binary mixtures using a leader compound to found pharmacodynamic interactions in inhibition of the NF-κB/AP-1 pathway using RAW-Blue™ cells. Eight compounds were isolated from Castilleja tenuiflora, four were new furofuran-type lignans for the species magnolin, eudesmin, sesamin, and kobusin. Magnolin (60.97%) was the most effective lignan inhibiting the NF-κB/AP-1 pathway, followed by eudesmin (56.82%), tenuifloroside (52.91%), sesamin (52.63%), and kobusin (45.45%). Verbascoside, a major compound contained in wild C. tenuiflora showed an inhibitory effect on NF-κB/AP-1. This polyphenol was chosen as a leader compound for binary mixtures. Verbacoside-aucubin and verbascoside-kobusin produced synergism, while verbascoside-tenuifloroside had subadditivity in all concentrations. Verbascoside-kobusin is a promising mixture to use on NF-κB/AP-1 related diseases and anti-inflammatory C. tenuiflora-based phytomedicines.

Gastroprotective activity of kaempferol glycosides from Malvaviscus arboreus Cav.

ETHNOPHARMACOLOGICAL RELEVANCE: Malvaviscus arboreus is traditionally used in Mexico and Central America for culinary and medicinal purposes. Leaves and flowers of this species are commonly used for preparation of salads, herbal teas and herbal dyes. Panamanian, Guatemalan and Mexican healers use this medicinal plant for the management of fever, respiratory complications, dysentery, liver and gallbladder problems, stomachache and gastritis between other health troubles. AIM OF THE STUDY: Considering the traditional use of M. arboreous as well as its content in flavonoids and other polyphenols, the objective of this work was to evaluate the gastroprotective effect of an aqueous extract and identify the potential bio-active principles from flowers of this species. MATERIAL AND METHODS: Fresh flowers of Malvaviscus arboreus were collected, dried, and macerated with water. The aqueous extract (ExAq) was partitioned using an immiscible mixture of water and ethyl acetate, giving an aqueous (MaAq) and organic (MaEA) fractions. The gastroprotective effect was carried out using an ethanol-induced gastric ulcer experimental test in male rats. While tween 20 was used as a negative control, famotidine (10 mg/kg) and L-arginine (300 mg/kg) were used as positive controls. Compounds 1 and 2 were isolated by several chromatographic techniques and the chemical characterization was carried out by means of the analysis of the NMR spectra in one and two dimensions. RESULTS: The integrate extract (ExAq) to 250, 500 and 750 mg/kg showed gastroprotective effect with high levels of 97.8%, 79.5% and 91.1% respectively. The organic fraction (MaEA) displayed a protection of 91.2%, 96.0% and 99.4% when it was evaluated at 125, 250 and 500 mg/kg respectively. Comparison of these results with famotidine at 10 mg/kg (83% of gastroprotection) indicated that ethyl acetate fraction showed a better gastroprotection. The bio-guided separation of this organic mixture, allowed obtaining the most active fraction (C1F4, 60 mg/kg) which was finally purified to obtain two glycosylated flavonols: kaempferol 3-O-D-sophoroside (1) and kaempferol 3-O-D-sambubioside (2). This mixture of flavonoids (40 y 60 mg/kg) showed 93.7 and 92% of gastroprotective activity respectively. CONCLUSION: This study allowed demonstrating that an aqueous extract and its organic fraction (MaEA) from M. arboreous contain glycosylated flavonoids (1 and 2) which are responsible of the gastroprotective properties of M. arboreous. These results will be used in the future development of a standardized treatment useful in the therapeutic management of gastric ulcers.

Glucoindole alkaloid accumulation induced by yeast extract in Uncaria tomentosa root cultures is involved in defense response.

OBJECTIVE: To evaluate the induction of monoterpenoid indole alkaloids (MIA) and phenolic compound production by yeast extract (YE) and its relationship with defense responses in Uncaria tomentosa (Rubiaceae) root cultures. RESULTS: Root cultures were elicited by YE at three concentrations. The 0.5 mg YE ml-1 treatment did not affect cell viability but increased the hydrogen peroxide concentration by 5.7 times; guaiacol peroxidase activity by twofold; and the glucoindole alkaloid 3α-dihydrocadambine (DHC) content by 2.6 times (to 825.3 ± 27.3 µg g-1). This treatment did not affect the contents of monoterpenoid oxindole alkaloids or chlorogenic acids. In response to 0.5 mg YE ml-1 treatment, the transcript levels of MIA biosynthetic genes, TDC and LAMT, increased 5.4 and 1.9-fold, respectively, that of SGD decreased by 32%, and that of STR did not change. The transcript levels of genes related to phenolic compounds, PAL, CHS and HQT, increased by 1.7, 7.7, and 1.2-fold, respectively. Notably, the transcript levels of Prx1 and Prx encoding class III peroxidases increased by 1.4 and 2.5-fold. CONCLUSION: The YE elicitor induced an antioxidant defense response, increased the transcript levels of genes encoding enzymes related to strictosidine biosynthesis precursors and class III peroxidases, and decreased the transcript level of SGD. Thus, YE could stimulate antifungal DHC production in root cultures of U. tomentosa.

In Vivo Gastroprotective and Antidepressant Effects of Iridoids, Verbascoside and Tenuifloroside from Castilleja tenuiflora Benth.

Stress is an important factor in the etiology of some illnesses such as gastric ulcers and depression. Castilleja tenuiflora Benth. (Orobanchaceae) is used in Mexican traditional medicine for the treatment of gastrointestinal diseases and nervous disorders. Previous studies indicated that organic extracts from C. tenuiflora had gastroprotective effects and antidepressant activity. In this study, we aimed to evaluate the gastroprotective and antidepressant activity of fractions and isolated compounds from the methanolic extract (MECt) of C. tenuiflora in stressed mice. Chromatographic fractionation of MECt produced four fractions (FCt-1, FCt-2, CFt-3, and FCt-4) as well as four bioactive compounds which were identified using TLC, HPLC and NMR analyses. The cold restraint stress (CRS)-induced gastric ulcer model followed by the tail suspension test and the forced swim test were used to evaluate the gastroprotective effect and antidepressant activity of the extract fractions. FCt-2 and FCt-3 at 100 mg/kg had significant gastroprotective and antidepressant effects. All isolated compounds (verbascoside, teniufloroside and mixture geniposide/ musseanoside) displayed gastroprotective effects and antidepressant activity at 1 or 2 mg/kg. The above results allow us to conclude that these polyphenols and iridoids from C. tenuiflora are responsible for the gastroprotective and antidepressant effects.

A new furofuran lignan diglycoside and other secondary metabolites from the antidepressant extract of Castilleja tenuiflora Benth.

Castilleja tenuiflora has been used for the treatment of several Central Nervous System (CNS) diseases. Herein we report the antidepressant activity of the methanol extract from the leaves of this medicinal plant. The oral administration of MeOH extract (500 mg/kg) induced a significant (p < 0.05) decrement of the immobility parameter on Forced Swimming Test (FST) and an increment in the latency and duration of the hypnosis, induced by administration of sodium pentobarbital (Pbi, 40 mg/kg, i.p.). Chemical analysis of this antidepressant extract allowed the isolation of (+)-piperitol-4-O-xylopyranosyl-(1→6)-O-glucopyranoside. This new furofuran lignan diglycoside was named tenuifloroside (1) and its complete chemical structure elucidation on the basis of 1D and 2D NMR spectra analysis of the natural compound 1 and its peracetylated derivative 1a is described. This compound was found together with two flavones-apigenin and luteolin 5-methyl ether-a phenylethanoid-verbascoside-and three iridoids-geniposide, caryoptoside and aucubin. All these compounds were purified by successive normal and reverse phase column chromatography. Tenuifloroside, caryoptoside and luteolin 5-methyl ether were isolated from Castilleja genus for the first time. These findings demonstrate that C. tenuiflora methanol extract has beneficial effect on depressive behaviors, and the knowledge of its chemical constitution allows us to propose a new standardized treatment for future investigations of this species in depressive illness.

In vivo anti-inflammatory and anti-ulcerogenic activities of extracts from wild growing and in vitro plants of Castilleja tenuiflora Benth. (Orobanchaceae).

ETHNOPHARMACOLOGICAL RELEVANCE: Castilleja tenuiflora Benth. (Orobanchaceae) is a perennial shrub used since the 16(th) century in Mexican traditional medicine for the treatment of a number of health disorders including inflammation, stomach pain and tumors. The aim of the present study was to evaluate the cytotoxic, anti-inflammatory and anti-ulcerogenic activities of ethyl acetate (EaE), methanol (ME) and aqueous extracts (AE) of Castilleja tenuiflora wild grown (CtW) and in vitro plants (CtIv). MATERIAL AND METHOD: Phytochemical analysis of the phenylethanoid glycoside (PhG) and iridoid glycoside (IG) components was carried out by chromatographic methods. In vitro cytotoxic activity of the extracts was evaluated in the following four carcinoma cell lines: colon (HF-6), breast (MCF-7), prostate (PC-3), and nasopharyngeal (KB). The topical anti-inflammatory activity was evaluated in mouse ear edema induced by 12-O-tetradecanoylphorbol-13-acetate (TPA). Anti-ulcerogenic activity was evaluated in rats using an absolute ethanol-induced acute gastric ulcer model. RESULTS: The main compounds in the extracts were isoverbascoside, verbascoside and aucubin and their concentration depended both on the solvent used and on the plant material origin. None of the extracts showed cytotoxicity against the tested cell lines. In contrast, CtWEaE, CtWAE and CtIvEaE (1.6 mg/ear) showed moderate anti-inflammatory activity similar to dexamethasone (1 mg/ear) with a 38.2, 39.3 and 49.1% decrease of inflammation, respectively. CtWEaE and CtIvEaE (100 mg/kg) showed high anti-ulcerogenic activity with 88.3 and 83.1% inhibition, respectively, compared to famotidine (20 mg/kg, 32.8% inhibition). CONCLUSION: Castilleja tenuiflora extracts provided significant gastric protection in an acute ulcer induction model and topical anti-inflammatory activity in a mouse ear edema model. These activities are related to verbascoside and may explain the traditional use of Castilleja tenuiflora in the treatment of anti-inflammatory and gastrointestinal disorders. Cultured Castilleja tenuiflora plants (in vitro) exhibited pharmacological activities and also have the potential to produce bioactive compounds.

Strictosidine-related enzymes involved in the alkaloid biosynthesis of Uncaria tomentosa root cultures grown under oxidative stress.

The activity and gene expression of strictosidine-related enzymes in Uncaria tomentosa root cultures exposed to oxidative stress were studied. Elicitation with 0.2 mM hydrogen peroxide (H2 O2 ) or a combination of 0.8 mM buthionine sulfoximine and 0.2 mM jasmonic acid (BSO-JA) increased peroxidase activities by twofold at Day 8 and glutathione reductase by 1.4-fold at Day 5 in H2 O2 elicited cultures respect to the control. Production of monoterpenoid oxindole alkaloids (MOA), 3α-dihydrocadambine, and dolichantoside was stimulated after H2 O2 elicitation, reaching levels of 886.4 ± 23.6, 847.7 ± 25.4, and 87.5 ± 7.2 µg/g DW, at Day 8 which were 1.7-, 2.1-, and 2.3-fold higher relative to control. BSO-JA elicited cultures produced about twice alkaloids than H2 O2 -treated cultures, following a biphasic pattern with maxima at 0.5 and 8 days. Alkaloid production was preceded by increase in strictosidine synthase (STR) and strictosidine glucosidase (SGD) activities. After elicitation with H2 O2 or BSO-JA, the STR activity (pKat/mg protein) increased by 1.9-fold (93.8 ± 17.8 at 24 h) or 2.5-fold (102.4 ± 2.2 at 6 h) and the SGD activity (pKat/mg protein) by 2.8-fold (245.2 ± 14.4 at 6 h) or 4.2-fold (421.2 ± 1.8 at 18 h) relative to control. STR and SGD transcripts were upregulated after elicitation. H2 O2 -treated roots showed higher levels of STR at 48-192 h and SGD at 24-48 h, while BSO-JA treatments showed STR increased at 12 h and SGD at 24 h. Also, LC/ESI-MS confirmed the biosynthesis of dolichantoside from N-ω-methyltryptamine and secologanin by U. tomentosa protein extracts.

A propagation procedure for Cuphea aequipetala Cav. (Lythraceae) and antioxidant properties of wild and greenhouse-grown plants / Procedimiento para la propagación de Cuphea aequipetala Cav. (Lythraceae) y propiedades antioxidantes de plantas silvestres y crecidas en invernadero

Cuphea aequipetala Cav. (Lythraceae) is native to Mexico and is used in folk medicine to treat tumors. An efficient protocol for in vitro shoot proliferation and plant acclimatization of C. aequipetala was developed. Total phenolic compounds and flavonoids contents were determined in methanolic extracts of roots, stems, and leaves from wild and greenhouse-grown plants. Their antioxidant properties were compared using in vitro assays (scavenging of 2,2-Diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-Azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) radicals, and reducing power in the phosphomolybdenum assay). This is the first report of a successful propagation procedure for C. aequipetala. These methods offer a viable approach for long-term in vitro conservation and proliferation of this species. C. aequipetala shoots maintained their proliferation capacity during long-term subculture (3 years). The propagated shoots can successfully acclimatize and grow to maturity, and they retain the ability to accumulate antioxidants.

Cuphea aequipetala Cav. (Lythraceae) es una planta nativa de México que se utiliza en la medicina tradicional para tratar tumores. En este trabajo se desarrolló un procedimiento para la proliferación de brotes y la aclimatización de plantas de C. aequipetala. Se determinó la concentración de compuestos fenólicos totales y de flavonoides en extractos metanólicos de raíces, tallos y hojas de plantas silvestres y crecidas en invernadero. Sus propiedades antioxidantes fueron comparadas utilizando ensayos in vitro (captura de radicales DPPH y ABTS y poder reductor por el ensayo de fosfomolibdeno). Este es el primer reporte exitoso sobre un procedimiento para la propagación de C. aequipetala. Este método ofrece una alternativa viable para la conservación a largo plazo y la proliferación de esta especie. Los brotes de C. aequipetala han mantenido su capacidad de multiplicación a largo plazo (tres años). Los brotes se convirtieron en plantas adultas aclimatadas, manteniendo su habilidad para acumular compuestos antioxidantes.

Establecimiento de un protocolo de permeabilización con detergentes que incrementa la eficiencia en la producción de betacianinas en cultivos de células de Beta vulgaris L / stablishment of a premeabilization protocol with detergents to increase betacianine production efficiency in a Beta vulgaris L. Cell cultures / Estabelecimento de um protocolo de permeabilização com detergentes que incrementa a eficiência na produção de betacianinas em cultivos de células de Beta vulgaris L

Se evaluó el efecto de cuatro detergentes, Tween 20, 40 y 80, y Tritón X-100® como agentes químicos permeabilizantes para la liberación de betacianinas (BC) de células en suspensión de Beta vulgaris. Se seleccionó el agente químico permeabilizante con base en la concentración de betacianinas liberadas y el tiempo de contacto. El contenido de BC se estimó usando medición de color por análisis de imágenes. Los resultados mostraron que la adición de Tritón X-100® 0,7mM durante 10min era suficiente para liberar el 36% de BC, con una viabilidad de 60-70%, y permitiendo además un nuevo ciclo de cultivo de las células tratadas y la acumulación paulatina de betacianinas durante el segundo ciclo.

Red beet (Beta vulgaris L.) cell suspensions were permeabilized by means of four chemical detergent agents, Tween 20, 40 and 80, and Triton X-100®, to evaluate the recovery of betacianins (BC). The permeabilizating agent was selected as a function of the quantity of BC released and the contact time. Betacianin concentration was measured using digital color image analysis. The results showed that 36% of betacianins was released using Triton X-100® (0.7mM) during 10min; using these extraction conditions, the viability remained at 60-70%. This treatment allowed a second growing-cycle, as well as, an additional accumulation of betacianins.

Avaliou-se o efeito de quatro detergentes, Tween 20, 40 e 80, e Tritón X-100® como agentes químicos permeabilizantes para a liberação de betacianinas (BC) de células em suspensão de Beta vulgaris. Selecionou-se o agente químico permeabilizante com base na concentração de betacianinas liberadas e o tempo de contacto. O conteúdo de BC se estimou usando medição de cor por análise de imágens. Os resultados mostraram que a adição de Tritón X-100® 0,7mM durante 10min era suficiente para liberar 36% de BC, com uma viabilidade de 60-70%, e permitindo além disso um novo ciclo de cultivo das células tratadas e a acumulação paulatina de betacianinas durante o segundo ciclo.

La agregación celular en la producción de metabolitos secundarios en cultivos vegetales in vitro / Celular aggregation in secondary metabolite production in in vitro plant cell cultures

El cultivo de células vegetales es una alternativa biotecnológica para la producción de metabolitos secundarios. Sin embargo, la productividad de los sistemas in vitro es menor a la obtenida de plantas. En esta revisión se ilustra la diferenciación y compartamentalización celular como eventos necesarios para la síntesis de metabolitos secundarios en las plantas. Se discute la inducción de la agregación celular en los cultivos in vitro como una de las estrategias para favorecer la acumulación de estos compuestos químicos. Este efecto positivo podría ser explicado como consecuencia de la formación de estructuras morfogénicas y/o por una condición de estrés por limitaciones de oxígeno al interior de los agregados. Finalmente, se muestra que la combinación de la agregación con otras estrategias tales como la selección de líneas celulares, la elicitación y la adición de precursores constituye una alternativa para desarrollar bioprocesos a partir de células vegetales in vitro para la producción de compuestos químicos de alto valor agregado.