Intermittent fasting along with hydroalcoholic extract of Centella-asiatica ameliorates sub-acute hypoxia-induced ischemic stroke in adult zebrafish.

Reduced blood flow (hypoxia) to the brain is thought to be the main cause of strokes because it deprives the brain of oxygen and nutrients. An increasing amount of evidence indicates that the Centella-Asiatica (HA-CA) hydroalcoholic extract has a variety of pharmacological benefits, such as antioxidant activity, neuroprotection, anti-inflammatory qualities, and angiogenesis promotion. Intermittent fasting (IF) has neurological benefits such as anti-inflammatory properties, neuroprotective effects, and the ability to enhance neuroplasticity. The current study evaluates the combined effect of IF (for 1, 6, and 12 days) along with HA-CA (daily up to 12 days) in adult zebrafish subjected to hypoxia every 5 min for 12 days followed by behavioral (novel tank and open-field tank test), biochemical (SOD, GSH-Px, and LPO), inflammatory (IL-10, IL-1ß, and TNF-α), mitochondrial enzyme activities (Complex-I, II, and IV), signaling molecules (AMPK, MAPK, GSK-3ß, Nrf2), and imaging/staining (H&E, TTC, and TEM) analysis. Results show that sub-acute hypoxia promotes the behavioral alterations, and production of radical species and alters the oxidative stress status in brain tissues of zebrafish, along with mitochondrial dysfunction, neuroinflammation, and alteration of signaling molecules. Nevertheless, HA-CA along with IF significantly ameliorates these defects in adult zebrafish as compared to their effects alone. Further, imaging analysis significantly provided evidence of infarct damage along with neuronal and mitochondrial damage which was significantly ameliorated by IF and HA-CA. The use of IF and HA-CA has been proven to enhance the physiological effects of hypoxia in all dimensions.

Integrating High-Resolution Mass Spectral Data, Bioassays and Computational Models to Annotate Bioactives in Botanical Extracts: Case Study Analysis of C. asiatica Extract Associates Dicaffeoylquinic Acids with Protection against Amyloid-ß Toxicity.

Rapid screening of botanical extracts for the discovery of bioactive natural products was performed using a fractionation approach in conjunction with flow-injection high-resolution mass spectrometry for obtaining chemical fingerprints of each fraction, enabling the correlation of the relative abundance of molecular features (representing individual phytochemicals) with the read-outs of bioassays. We applied this strategy for discovering and identifying constituents of Centella asiatica (C. asiatica) that protect against Aß cytotoxicity in vitro. C. asiatica has been associated with improving mental health and cognitive function, with potential use in Alzheimer's disease. Human neuroblastoma MC65 cells were exposed to subfractions of an aqueous extract of C. asiatica to evaluate the protective benefit derived from these subfractions against amyloid ß-cytotoxicity. The % viability score of the cells exposed to each subfraction was used in conjunction with the intensity of the molecular features in two computational models, namely Elastic Net and selectivity ratio, to determine the relationship of the peak intensity of molecular features with % viability. Finally, the correlation of mass spectral features with MC65 protection and their abundance in different sub-fractions were visualized using GNPS molecular networking. Both computational methods unequivocally identified dicaffeoylquinic acids as providing strong protection against Aß-toxicity in MC65 cells, in agreement with the protective effects observed for these compounds in previous preclinical model studies.

Centella asiatica improves memory and executive function in middle-aged rats by controlling oxidative stress and cholinergic transmission.

ETHNOPHARMACOLOGICAL RELEVANCE: Centella asiatica (L.) Urban, is a medicinal herb with rich history of traditional use in Indian subcontinent. This herb has been valued for its diverse range of medicinal properties including memory booster, and also as a folk treatment for skin diseases, wound healing and mild diuretic. AIM OF STUDY: Aging is a gradual and continuous process of natural decay in the biological systems, including the brain. This work aims to evaluate the effectiveness of ethanolic extract of Centella asiatica (CAE) on age-associated cognitive impairments in rats, as well as the underlying mechanism. MATERIAL AND METHODS: Rats were allocated into five distinct groups of 5 animals each: Young rats (3 months old rats), middle-aged (m-aged) rats (13-14 months old), and the remaining three groups were comprised of m-aged rats treated with different concentrations of CAE, viz., 150, 300, and 450 mg/kg b. w., orally for 42 days. Y-maze, open field, novel object recognition, and elevated plus maze tests were used to assess animal behavior. The malondialdehyde (MDA), superoxide dismutase (SOD), and acetylcholinesterase (AChE) assays; and H&E staining were done in the rat brain to assess the biochemical and structural changes. CAE was also subjected to HPLC analysis, in vitro antioxidant and anti-cholinergic activity. The active compounds of CAE were docked with AChE and BuChE in molecular docking study. RESULTS: The results showed that CAE treatment improves behavioral performance; attenuates the age-associated increase in MDA content, SOD, and AChE activity; and reduces neuronal loss. In vitro study showed that CAE has concentration-dependent antioxidant and anti-AChE activity. Furthermore, the presence of Asiatic acid and Madecassic acid in CAE and their good binding with cholinergic enzymes (in silico) also suggest the anticholinergic effect of CAE. CONCLUSION: The findings of the current study show that the anticholinergic and antioxidant effects of CAE are attributable to the presence of Asiatic acid and Madecassic acid, which not only provide neuroprotection against age-associated cognitive decline but also reverse it.

Wound healing efficacy of a polymeric spray film solution containing Centella asiatica leaf extract on acute wounds.

OBJECTIVE: To study the clinical efficacy of a polymeric spray film containing Centella asiatica extract to heal acute wounds. METHOD: A polymeric spray film solution for wound healing was formulated using Centella asiatica extract, which contained triterpenes, including asiatic acid, madecassic acid, asiaticoside and madecassoside. The stability and physicochemical properties of the formulation were evaluated, and a multicentre, randomised, controlled trial was conducted to assess its clinical wound-healing efficacy. The Pressure Ulcer Scale for Healing (PUSH Tool) score was used to evaluate wound healing on days 0, 3, 5 and 7. RESULTS: The cohort consisted of 60 volunteers with clean-contaminated wounds (class 1), randomly assigned to the Control (n=30) and Testing (n=30) groups. The spray product contained asiatic acid, madecassic acid, asiaticoside and madecassoside at 0.20±0.02mg/ml, 0.16±0.01mg/ml, 0.32±0.03mg/ml and 0.10±0.00mg/ml, respectively. The pH value was 5.5±0.01, and the viscosity was 33±4cP. The product was stable for six months when stored at 30±2°C and at 40±2°C, in 75±5% relative humidity. The tested product significantly reduced the total PUSH and exudate scores, indicating that the polymeric spray film solution containing Centella asiatica improved wound healing. The average healing recovery times for the Testing and Control groups were 4.6±1.1 days and 4.87±1.0 days, respectively. CONCLUSION: In this study, Centella asiatica extract-containing polymeric spray film solution was beneficial as an acute wound medication, which could shorten healing time with no adverse effects.

The standardized extract of Centella asiatica L. Urb attenuates the convulsant effect induced by lithium/pilocarpine without affecting biochemical and haematological parameters in rats.

BACKGROUND: Status epilepticus (SE) is a type of epileptic activity characterized by a failure of the inhibitory mechanisms that limit seizures, which are mainly regulated by the GABAergic system. This imbalance increases glutamatergic neurotransmission and consequently produces epileptic activity. It is also associated with oxidative stress due to an imbalance between reactive oxygen species (ROS) and antioxidant defences. Unfortunately, long-term treatment with anti-epileptic drugs (AEDs) may produce hepatotoxicity, nephrotoxicity, and haematological alterations. In this way, some secondary metabolites of plants have been used to ameliorate the deterioration of nervous system disorders through their antioxidant properties, in addition to their anticonvulsant effects. An example is Centella asiatica, a plant noted to have a reputed neuroprotective effect related to its antioxidant activity. However, similar to conventional drugs, natural molecules may produce side effects when consumed in high doses, which could occur with Centella asiatica. Therefore, we aimed to evaluate the effect of a standardized extract of Centella asiatica L. Urb with tested anticonvulsant activity on biochemical and haematological parameters in rats subjected to lithium/pilocarpine-induced seizures. METHODS: Twenty-eight adult male Wistar rats were randomly divided into four groups (n = 7 each): vehicle (purified water), Centella asiatica (200 and 400 mg/kg), and carbamazepine (CBZ) (300 mg/kg) as a pharmacological control of anticonvulsant activity. Treatments were administered orally every 24 h for 35 consecutive days. On Day 36, SE was induced using the lithium/pilocarpine model (3 mEq/kg, i.p. and 30 mg/kg s.c., respectively), and the behavioural and biochemical effects were evaluated. RESULTS: Centella asiatica 400 mg/kg increased the latency to the first generalized seizure and SE onset and significantly reduced the time to the first generalized seizure compared to values in the vehicle group. Biochemical parameters, i.e., haematic cytometry, blood chemistry, and liver function tests, showed no significant differences among the different treatments. CONCLUSION: The dose of Centella asiatica that produces anticonvulsant activity in the lithium/pilocarpine model devoid of hepatotoxicity, nephrotoxicity, and alterations in haematological parameters suggests that the standardized extract of this plant could be of utility in the development of new safe therapies for the treatment of convulsions associated with epilepsy.

Investigating high throughput phenotyping based morpho-physiological and biochemical adaptations of indian pennywort (Centella asiatica L. urban) in response to different irrigation regimes.

Indian pennywort (Centella asiatica L. Urban; Apiaceae) is a herbaceous plant used as traditional medicine in several regions worldwide. An adequate supply of fresh water in accordance with crop requirements is an important tool for maintaining the productivity and quality of medicinal plants. The objective of this study was to find a suitable irrigation schedule for improving the morphological and physiological characteristics, and crop productivity of Indian pennywort using high-throughput phenotyping. Four treatments were considered based on irrigation schedules (100, 75, 50, and 25% of field capacity denoted by I100 [control], I75, I50, and I25, respectively). The number of leaves, plant perimeter, plant volume, and shoot dry weight were sustained in I75 irrigated plants, whereas adverse effects on plant growth parameters were observed when plants were subjected to I25 irrigation for 21 days. Leaf temperature (Tleaf) was also retained in I75 irrigated plants, when compared with control. An increase of 2.0 °C temperature was detected in the Tleaf of plants under I25 irrigation treatment when compared with control. The increase in Tleaf was attributed to a decreased transpiration rate (R2 = 0.93), leading to an elevated crop water stress index. Green reflectance and leaf greenness remained unchanged in plants under I75 irrigation, while significantly decreased under I50 and I25 irrigation. These decreases were attributed to declined leaf osmotic potential, increased non-photochemical quenching, and inhibition of net photosynthetic rate (Pn). The asiatic acid and total centellosides in the leaf tissues, and centellosides yield of plants under I75 irrigation were retained when compared with control, while these parameters were regulated to maximal when exposed to I50 irrigation. Based on the results, I75 irrigation treatment was identified as the optimum irrigation schedule for Indian pennywort in terms of sustained biomass and a stable total centellosides. However, further validation in the field trials at multiple locations and involving different crop rotations is recommended to confirm these findings.

Prediction of Translational Regulation by Network Interaction in Synaptic Plasticity Induced with Centella asiatica.

Background: Recently, human life expectancy, aging, and age-related health disorders, especially neurodegenerative diseases such as Alzheimer's disease (AD), have increased. The increasing number of AD patients causes a heavy social and economic burden on society. Since there is no treatment for AD, utilization of natural products is currently accepted as an alternative or integrative treatment agent against AD. Methods: Selection of protein databases related to synaptic plasticity was obtained from a gene bank. The protein-protein interaction (PPI) analysis was performed using Cytoscape 3.9.1. Prediction of Centella asiatica target constituents and their relationship with target synaptic plasticity was performed using STITCH, followed by GO and KEGG pathway enrichment analysis and molecular binding of ligands to presynaptic and postsynaptic receptors afterwards. Results: From the protein database, 446 protein coding genes related to synaptic plasticity were found. PPI and KEGG pathway analysis showed potentiality to inhibit AKT and mTORC1 pathways. The targeted proteins were TSC1, Rheb, and FMRP. Conclusion: This study showed potentiality of Centella asiatica in AD through its binding to several proteins such as TSC1, Rheb, and FMRP. This compound in Centella asiatica was able to bind to the AKT1 and mTOR signaling pathways. Centella asiatica may behold greater potency in AD therapy.

Centella asiatica alleviates psoriasis through JAK/STAT3-mediated inflammation: An in vitro and in vivo study.

ETHNOPHARMACOLOGICAL RELEVANCE: Centella asiatica (L.) Urban (CA) is a dry herb of the Umbelliferae family, first mentioned in Shennong's Herbal Classic. It is known for its ability to clear heat and dampness, detoxify, and reduce swelling, making it a popular treatment for dermatitis, wound healing, and lupus erythematosus. Psoriasis is a chronic inflammatory skin disease that is characterized by clearly delineated erythema and squamous skin lesions. However, the effect of CA on regulating inflammation and its mechanism in the pathogenesis of psoriasis is still not fully understood. AIM OF THE STUDY: This study evaluated the effects of CA on inflammatory dermatosis by in vitro and in vivo studies. And clarified the important role of the JAK/STAT3 signaling pathway in the treatment of psoriasis with CA. METHODS AND MATERIALS: Different components of CA were extracted and analyzed for their total flavonoid and polyphenol contents. The antioxidant capacity of the CA extracts was determined using DPPH, ABTS, and FRAP methods. In vitro, HaCaT cells were induced by lipopolysaccharide (LPS, 20 µg·mL-1) to establish an inflammatory injury model, and the effects of CA extracts on oxidative stress, inflammation and skin barrier function were evaluated systematically. Annexin V-FITC/PI staining was utilized for detecting cell apoptosis, while the expression of NF-κB and JAK/STAT3 pathways were detected by RT-PCR and western blot. Combined with an in vivo mice model of Imiquimod (IMQ) induced psoriasis-like skin inflammation, the most effective CA extract for alleviating psoriasis was identified and its potential mechanism was investigated. RESULTS: CA extracts showed high antioxidant capacity and were able to increase the content of GSH and SOD while reducing intracellular ROS generation. Notably, CA ethyl acetate extract (CAE) was found to be the most effective. Furthermore, CA extracts effectively downregulate inflammatory factors (IFN-γ, CCL20, IL-6 and TNF-α) mRNA levels and improved the gene expressions of barrier protective factors AQP3 and FLG, among them CAE and n-hexane extract of CA (CAH) had better effects. Western blot analysis indicated that CAE and CAH had anti-inflammatory effects by inhibiting the activation of NF-κB and JAK/STAT3 pathways, and CAE exhibited the best regulatory effect at the dose of 25 µg·mL-1. In vivo experiment, the psoriasis-like skin inflammation mice model was established by 5% IMQ and treated CAE solution (10, 20, 40 mg·mL-1) for 7 days, the results showed that CAE intervention reduced the skin scale and blood scab, and significantly inhibited the secretion of inflammatory factors in both serum and skin lesions at the dose of 40 mg·mL-1. CONCLUSION: Centella asiatica extracts were effective in improving skin inflammation and skin barrier dysfunction, and also alleviated psoriasis through JAK/STAT3 pathway. The results provided experimental support for the potential use of Centella asiatica in functional food and skin care products.

Centella asiatica (L.) Urban. Attenuates Cell Damage in Hydrogen Peroxide-Induced Oxidative Stress in Transgenic Murine Embryonic Stem Cell Line-Derived Neural-Like Cells: A Preliminary Study for Potential Treatment of Alzheimer's Disease.

BACKGROUND: Centella asiatica (L.) (C. asiatica) is commonly known in South East and South East Asia communities for its nutritional and medicinal benefits. Besides being traditionally used to enhance memory and accelerate wound healing, its phytochemicals have been extensively documented for their neuroprotective, neuroregenerative, and antioxidant properties. OBJECTIVE: The present study aims to investigate the effects of a standardized raw extract of C. asiatica (RECA) on hydrogen peroxide (H2O2)-induced oxidative stress and apoptotic death in neural-like cells derived from mouse embryonic stem (ES) cell line. METHODS: A transgenic mouse ES cell (46C) was differentiated into neural-like cells using 4-/4+ protocol with addition of all-trans retinoic acid. These cells were then exposed to H2O2 for 24 h. The effects of RECA on H2O2-induced neural-like cells were assessed through cell viability, apoptosis, and reactive oxygen species (ROS) assays, as well as neurite length measurement. The gene expression levels of neuronal-specific and antioxidant markers were assessed by RT-qPCR analysis. RESULTS: Pre-treatment with H2O2 for 24 hours, in a dose-dependent manner, damaged neural-like cells as marked by a decrease in cell viability, substantial increase in intracellular ROS accumulation, and increase in apoptotic rate compared to untreated cells. These cells were used to treat with RECA. Treatment with RECA for 48 h remarkably restored cell survival and promoted neurite outgrowth in the H2O2- damaged neurons by increasing cell viability and decreasing ROS activity. RT-qPCR analysis revealed that RECA upregulated the level of antioxidant genes such as thioredoxin-1 (Trx-1) and heme oxygenase-1 (HO-1) of treated cells, as well as the expression level of neuronal-specific markers such as Tuj1 and MAP2 genes, suggesting their contribution in neuritogenic effect. CONCLUSION: Our findings indicate that RECA promotes neuroregenerative effects and exhibits antioxidant properties, suggesting a valuable synergistic activity of its phytochemical constituents, thus, making the extract a promising candidate in preventing or treating oxidative stress-associated Alzheimer's disease.

Polyacetylenes from the adventitious roots of Centella asiatica with glucose uptake stimulatory activity.

Centella asiatica (L.) Urban is an herbaceous perennial plant of the Apiaceae family that has diverse medicinal uses. Its active components are saponin, phenolics, and polyacetylenes. Plant cell cultures have been exploited for the efficient production of metabolites with pharmacological activity. In this work, we prepared adventitious root cultures of C. asiatica and analyzed their content and biological activity. Adventitious root extracts were found to increase glucose uptake by differentiated L6 skeletal muscle cells and to be more efficient than the extract of whole plants. Chromatographic fractionation of the extracts from adventitious roots of C. asiatica led to the isolation of two known polyacetylenes, araliadiol (1) and 8-acetoxy-1,9-pentadecadiene-4,6-diyn-3-ol (2), in addition to a new polyacetylene, which we have named centellidiol (3). All the three polyacetylenes stimulated glucose uptake in a dose-dependent manner. The methanol extract of adventitious roots contained 0.53% and 0.82% of compounds 1 and 2, respectively, which are values that were 15 and 21 times higher that are found in mother plants. We therefore suggest that the high content of these polyacetylenes contributes to the high efficacy of C. asiatica adventitious root cultures. Overall, adventitious root cultures of C. asiatica can be part of a secure supply of effective ingredients including polyacetylenes.

Efecto de unas gotas a base de ácido hialurónico, Aloe vera y Centella asiática en la calidad de vida de pacientes con ojo seco / Effect of eye drops based on hyaluronic acid, Aloe vera and Centella asiatica on quality of life of patients with dry eye

Introducción: la enfermedad de ojo seco (EOS) es una patología común y un motivo de consulta frecuente en farmacia comunitaria. Esta tiene un gran impacto en la calidad de vida (QoL) de los pacientes; por ello, un tratamiento adecuado debería mejorarla. Las lágrimas artificiales constituyen la base para el manejo de la EOS y el principal tratamiento de indicación farmacéutica.Objetivo: estudiar el efecto de unas gotas oculares a base de ácido hialurónico, Centella asiática y Aloe vera en la QoL de pacientes con EOS. Paralelamente, la adherencia al tratamiento, la tolerancia y seguridad del producto, fueron evaluados.Metodología: estudio clínico posautorización, abierto, prospectivo y multicéntrico. Los pacientes recibieron el tratamiento durante 60 días. Para estudiar el efecto sobre la QoL, los pacientes cumplimentaron el cuestionario OSDI© antes de iniciar el tratamiento y a los 30 y 60 días de tratamiento. La adherencia se evaluó mediante la reconciliación de los envases y la seguridad mediante un seguimiento de los eventos adversos. Resultados: los pacientes tratados con las gotas oculares, población por intención de tratar (ITT, n=44), obtuvieron mejoras clínicamente importantes en la QoL, sintomatología y función visual, pues la puntuación OSDI© disminuyó de 32,80 (DE=23,00) (basal, EOS severo) a 12.64 (DE=15,32) (estado normal, p<0,001), y a 9.22 (DE=10,37) (estado normal, p<0,001), a los 30 y 60 días, respectivamente. Más del 70 % de los pacientes se adhirieron al tratamiento. El perfil de seguridad fue favorable. Se observaron 4 efectos adversos oculares de intensidad leve. No se plantearon problemas de seguridad. Conclusiones: Las gotas oculares objeto de estudio podrían ser una opción de tratamiento efectiva y segura para mejorar la QoL de pacientes con EOS.

Centella asiatica (L.) Urb. Extract ameliorates branched-chain amino acid (BCAA) metabolism in acute reserpine-induced stress zebrafish model via 1H Nuclear Magnetic Resonance (NMR)-based metabolomics approach.

Depression is a common mental disorder that can adversely affect psychosocial function and quality of life. However, the exact aetiology and pathogenesis of depression are still unclear. Stress plays a major role in the pathogenesis of depression. The use of currently prescribed antidepressants has many side effects. Centella asiatica (C. asiatica) has shown promising antidepressant activity in rodent models. Here, we developed a reserpine-induced zebrafish stress-like model and performed behavioural analysis, cortisol measurement and 1H-Nuclear Magnetic Resonance (1H NMR) spectroscopy-based metabolomics analysis to test the anti-stress activity of ethanolic extract of C. asiatica (RECA). A significant increase in total distance travelled (F(8,8) = 8.905, p = 0.0054) and a reduction in freezing duration (F(9, 9) = 10.38, p = 0.0018) were found in the open field test (OFT). Asiaticoside, one of tested C.asiatica's triterpenoid gives a significant increase in contact duration (F(5,5) = 142.3, (p = 0.0330) at 2.5 mg/kg). Eight biomarkers were found, i.e. ß-hydroxyisovaleric acid, leucine, threonine, scylloinositol, lactate, betaine, valine, choline and l-fucose, to be responsible for the class separation between stress and RECA-treated groups. Metabolic pathway alteration in zebrafish brain upon treatment with RECA was identified as valine, leucine and isoleucine biosynthesis, while alanine, aspartate, glutamate and glycerophospholipid metabolism was involved after fluoxetine treatment.

Influence of two natural cross-linkers on microtensile bond strength durability ­ An in vitro study / Influência de dois reticuladores naturais na durabilidade da resistência de união à microtração ­ Um estudo in vitro

Objective: to investigate the effect of two natural cross-linkers on microtensile bond strength (µTBS) and evaluate their influence on the durability of the resin dentin bonds. Material and Methods: the Moringa oleifera and Centella asiatica plant extracts were qualitatively tested with high-performance thin layer chromatography (HPTLC) for the presence of phenols. The phenolic content ranged from 27 to 30 gallic acid equivalents (GAE), µg/mg of dry weight. After etching, two concentrations (5% and 1%) of these two extracts were prepared and used as pretreatment liners on dentin. They were applied for a min. After restoration with resin composite, dentin resin beams were prepared. The study groups were 5% Moringa, 1% Moringa 5% Centella 1% Centella, and control (without cross-linker application). For each group, half of the samples underwent µTBS testing after 24 hours, while the remaining half were immersed in artificial saliva to assess the bond's longevity after 6 months of ageing. Statistical analysis was performed using one-way ANOVA followed by Tukey's post hoc test. Results: both 5% and 1% Moringa showed a significant difference (p<0.05) compared to the other groups at both intervals. However, after ageing, the specimens in the control and 1% Centella groups resulted in a significant decrease in µTBS. Conclusion: overall, both concentrations of Moringa (5% and 1%) were effective in stabilising the bond during both intervals.(AU)

Objetivo: investigar o efeito de dois reticuladores naturais na resistência de união (µTBS) à microtração e avaliar sua influência na durabilidade da adesão da resina à dentina. Material e Métodos: extratos das plantas Moringa oleifera e Centella asiatica foram qualitativamente testados através de cromatografia em camada fina de alta performance (HPTLC) para a presença de fenóis. O conteúdo fenólico alcançou entre 27 a 30 equivalentes de ácido gálico (GAE), µg/mg de peso seco. Após o condicionamento, duas concentrações (5% e 1%) dos extratos foram preparadas e utilizadas como forros de pré-tratamento em dentina. Eles foram aplicados por um minuto. Após a restauração com resina composta, palitos de dentina e resina foram preparados. Os grupos foram 5% Moringa, 1% Moringa, 5% Centella, 1% Centella e controle (sem aplicação de reticulador). Para cada grupo, metade das amostras foram submetidas ao teste µTBS após 24 horas, enquanto a outra metade foi imersa em saliva artificial para avaliar a longevidade adesiva após 6 meses de envelhecimento. Foi realizada análise estatística através de ANOVA 1-fator, seguido do teste post hoc de Tukey. Resultados: ambas as concentrações de 5% e 1% de Moringa demonstraram diferença significativa (p<0.05) comparadas aos outros grupos em ambos os intervalos. No entanto, após o envelhecimento, os espécimes dos geupos controle e 1% de Centella resultaram em uma redução significativa de µTBS. Conclusão: no geral, ambas as concentrações de Moringa (5% e 1%) foram efetivas em estabelecer a adesão em ambos os intervalos (AU)

Mass spectrometry and synchrotron-FTIR microspectroscopy reveal the anti-inflammatory activity of Bua Bok extracts.

INTRODUCTION: Bua Bok or Centella asiatica (CA) is an Asian vegetable with anti-inflammatory benefits. Asiaticoside, asiatic acid, madecassoside and madecassic have been characterised as major active ingredients with a wide range of pharmacological advantages. In manufacturing processes, high-performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LCMS) are used to routinely determine the active compounds in raw materials. OBJECTIVES: This research aims to explore anti-inflammatory properties, characterise metabolites and observe the biochemical changes of the inflammatory induced macrophages after pretreatment with the potential extracted fractions. METHODS: Bua Bok leaf extracts were prepared. Macrophages were pretreated with non-toxic fractions to determine the anti-inflammatory action. Tentative metabolites of effective fractions were identified by LC-MS. Synchrotron fourier-transform infrared (S-FTIR) microspectroscopy was utilised to observe the biochemical change of the lipopolysaccharide (LPS)-induced cells after pretreatment with potential fractions. RESULTS: Fractions of ethyl acetate, 30% and 100% ethanol highly increased the nitrile scavenging and suppressed the function of phospholipase A2 . Fractions of 70% and 100% ethanol strongly decreased nitric oxide production. The comparison of 39 chemical compounds was presented. The change of proteins was improved after pretreatment of macrophages with fraction 70% ethanol. Fraction of 100% ethanol revealed the lipid accumulation was lower than 70% ethanol and diclofenac. CONCLUSION: While the anti-inflammatory actions of 70% and 100% ethanol were similar. S-FTIR expressed they inhibited inflammatory response with the distinct features of biomolecules. The S-FTIR, LC-MS and biological assay confidently provided the efficient strategies to inform the advantage of herbal extract on cellular organisation instead of a single compound.

Centella asiatica Modulates Nrf-2 Antioxidant Mechanisms and Enhances Reactive Oxygen Species-Mediated Apoptotic Cell Death in Leukemic (THP-1) Cells.

Centella asiatica is commonly used in traditional medicine owing to its many therapeutic properties including but not limited to antioxidant and antitumor potential. This study examined the antioxidant and antiproliferative effects of its crude (C) and fractionated (C3) ethanolic leaf extracts in THP-1 cells. In THP-1 cells, C and C3 cytotoxicity was evaluated (WST-1 viability assay; 24 h; [0.2-3 mg/mL]) and half maximal inhibitory concentration was obtained. Malondialdehyde (MDA; spectrophotometry), mitochondrial depolarization (Δψm), intracellular reactive oxygen species (IROS; flow cytometry), glutathione (GSH), oxidized GSH (GSSG) concentrations, adenosine triphosphate (ATP) levels, caspase activities (luminometry) and DNA fragmentation (single cell gel electrophoresis assay) were evaluated. Protein expression and gene expression was quantified by Western blotting and quantitative polymerase chain reaction, respectively. THP-1 cell viability was dose-dependently reduced by C and C3. MDA, IROS, GSH, and Δψm were increased and ATP was decreased by C and C3 (P < .01). Antioxidant gene expression, Nrf-2 protein expression, and GSSG levels (P < .01) were increased by C, but were decreased by C3. C and C3 elevated caspase activity and DNA damage (P < .0001), whereas they decreased glutathione peroxidase and Bcl-2 protein expressions (P < .003). c-PARP protein expression and c-myc gene expression was decreased by C, whereas they were increased by C3 (P < .002). C3 reduced OGG-1 gene expression (P < .0003). Antioxidant responses were increased by C, whereas they were decreased by C3. Both C and C3 exerted antiproliferative effects in THP-1 cells by enhancing apoptosis. Of note, C3 more effectively induced apoptosis.

Asiatic acid from centella asiatica exert anti-invasive ability in human renal cancer cells by modulation of ERK/p38MAPK-mediated MMP15 expression.

BACKGROUND: Asiatic acid (AA) is a naturally pentacyclic triterpenoids extracted from traditional medicine Centella asiatica l. that has demonstrated possesses potential health benefits and antitumor ability. However, the precise anticancer effects and mechanisms by which AA impact RCC cells remains unclear. METHODS: Cell proliferation and cell cycle distribution were detected by MTT, colony formation assay and PI stain by flow cytometry, respectively. Cell mobility and invasiveness were determined by in vitro migration and invasion assay. The secretory MMP15 was detected by ELISA assay. Quantitative RT-PCR, siRNA, and immunoblot were used to determine gene expression/regulation and protein expression, respectively. Antimetastatic effect of AA were performed to lung nodule numbers in vivo metastasis mice model. MMP15, pERK1/2 and p-p38MAPK expressions were determined by immunohistochemistry. RESULTS: Our findings indicated cell proliferation and cell cycle distribution of RCC cells were not significantly influenced by AA treatment. AA suppressed cell migration, invasion and significantly down-regulated mRNA and protein expression of MMP-15 (Matrix Metallopeptidase-15). Activation of ERK1/2 and p38MAPK were inhibited with AA, whereas combined AA with siRNA-ERK or siRNA-p38MAPK markedly reduced the metastatic effect and decreased MMP-15 expression in 786-O and A498 cells. Finally, AA significantly reduced the lung metastasis formation and metastasis-related proteins of human 786-O cells in vivo metastasis mice model. CONCLUSION: AA inhibits the metastatic properties of RCC cells via inhibition of the p-ERK/p-p38MAPK axis and the subsequent down-regulation of MMP-15 in vitro and in vivo. Further study of AA as a potential anti-metastatic agent for RCC is warranted.

Phytochemical profile of Centevita®, a Centella asiatica leaves extract, and isolation of a new oleanane-type saponin.

Centella asiatica is a popular medicinal plant and several phytotherapic products in the market include its extracts as active constituents. A LC-MS guided phytochemical investigation on the commercial C. asiatica leaves extract named Centevita® allowed characterization and quantification of 24 secondary metabolites including 10 polyphenols and 14 ursane- or oleanane-type triterpenoids in the sapogenin or saponin form. This metabolomic analysis, besides confirming that the triterpenoid fraction roughly accounts for 45% of the extract weight, also resulted in the discovery of isoterminoloside, a new triglycoside saponin of the unprecedented 2α,3ß,6ß,23-tetrahydroxyolean-13(18)-en-28-oic acid (isoterminolic acid). The structure of isoterminoloside was characterized by a detailed ESI-MS and NMR investigation.

Increase water solubility of Centella asiatica extract by indigenous bioenhancers could improve oral bioavailability and disposition kinetics of triterpenoid glycosides in beagle dogs.

A newly standardised extract of Centella asiatica (Centell-S) with better water solubility than the previous standardised extract of C. asiatica (ECa 233) was developed, and pharmacokinetic profiles of bioactive triterpenoids were investigated in beagle dogs. The test substances were administered via intravenous or oral administration with single and multiple doses for 7 days. The concentrations of major bioactive triterpenoids, including madecassoside, asiaticoside, madecassic acid, and asiatic acid, in biological samples were measured by liquid chromatography-tandem mass spectrometry. The dogs in this study showed good tolerability to all test substances, based on the physical appearance and blood chemistry 24 h after dosing. The major bioactive triterpenoids found in systemic blood circulation were madecassoside, asiaticoside, and asiatic acid; the concentration of these components ranged from 1 to 10,000 µg/L after intravenous administration of 1.0 mg/kg Centell-S. Oral administration of 10 and 20 mg/kg Centell-S generated approximately twofold higher plasma levels of both madecassoside and asiaticoside compared with equivalent doses of ECa 233. In addition, there was an accumulation of triterpenoid glycosides after multiple oral administrations of Centell-S for 7 days, while triterpenic acids showed little tendency for accumulation. Beagles had good tolerability to both standardised extracts of C. asiatica, and showed a similar pattern of bioactive triterpenoids to humans. Centell-S increased oral bioavailability of major triterpenoid glycosides and can be further developed into a phytopharmaceutical product.

Vascular relaxing effect of Hydrocotyle umbellata L. is mediated by blocking of l-type Ca2+ channels.

ETHNOPHARMACOLOGICAL RELEVANCE: Hydrocotyle umbellata L. is a medicinal herb for the treatment of some health problems including hypertension, according to traditional medicine. Even so, its vascular effects and the pharmacological action mechanisms have not been analyzed. AIM OF THE STUDY: This experiment aimed to analyze the effects of hydroalcoholic extract of Hydrocotyle umbellata L. (HEHU) on isolated vessels and verify the interaction of hibalactone (chemical marker) against Cav1.2 channels using molecular docking. MATERIALS AND METHODS: Vascular reactivity experiments were performed using rat aortas with (E+) or without endothelium (E-) in an isolated organ bath. Computational molecular docking approaches were used to show the direct effect on L-type Ca2+ Channels. RESULTS: HEHU (0-560 µg/mL) induced relaxation of the pre-contracted arteries in a concentration-dependent manner. The maximum effect was higher in E+ (76.8 ± 4.1%) as compared to E- (47.3 ± 5.5%). Pre-treatment of E+ arteries with L-NAME or ODQ reduced the relaxation to similar level of E- arteries. The treatment of arteries with MDL-12,330 A, diclofenac, propranolol and atropine did not change the relaxation induced by HEHU. The contraction caused by internal Ca2+ release induced by caffeine was reduced after HEHU treatment. Moreover, the HEHU also impaired the contraction induced by Ca2+ influx stimulated with phenylephrine or high KCl. The docking study demonstrated the effectiveness of hibalactone in blocking the Cav1.2 channel. CONCLUSIONS: These findings show that HEHU induces vascular relaxation which is potentiated (but not dependent) by endothelial cells. Blocking of Ca2+ influx seems to be the main mechanism for the vascular effects of HEHU.

Effect of Acidic Electrolysed Water and Pulsed Light Technology on the Sensory, Morphology and Bioactive Compounds of Pennywort (Centella asiatica L.) Leaves.

Pennywort (Centella asiatica) is a herbaceous vegetable that is usually served in the form of fresh-cut vegetables and consumed raw. Fresh-cut vegetables are in high demand as they offer convenience, have fresh-like quality and are potentially great for therapeutic applications. However, it could be the cause of foodborne outbreaks. Pulsed light is known as a decontamination method for minimally processed products. The aim of this study was to determine the influence of pulsed light in combination with acidic electrolysed water on the sensory, morphological changes and bioactive components in the leaves of pennywort during storage. A combination of soaking with acidic electrolysed water (AEW) at pH 2.5 and pulsed light (PL) treatment (1.5 J/cm2) was tested on the leaves of pennywort. After treatment, these leaves were refrigerated (4 ± 1 °C) for two weeks and evaluated on the basis of sensory acceptance, the visual appearance of the epidermal cell and bioactive compounds. In terms of sensorial properties, samples treated with the combined treatment were preferred over untreated samples. The combination of AEW and PL 1.5 J/cm2 was the most preferred in terms of purchasing and consumption criteria. Observations of the epidermal cells illustrated that PL treatment kept the cell structure intact. The bioactive phytocompounds found in the leaves of pennywort are mainly from the triterpene glycosides (asiaticoside, madecassoside, asiatic acid and madecassic acid) and are efficiently preserved by the combined treatment applied. In conclusion, the combination of acidic electrolysed water and pulsed light treatment is beneficial in retaining the sensory quality and bioactive compounds in the leaves of Pennywort during storage at 4 ± 1 °C.