Effect Of Ethyl Acetate Extract Research Articles

The protective effects of Zingiber zerumbet rhizome against fevers in rats.

The Zingiber zerumbet rhizomes are traditionally used to treat fever, and the in vitro inhibitory effect of ethyl acetate extract from Zingiber zerumbet rhizomes (EAEZZR) against DENV2 NS2B/NS3 (two non-structural proteins, NS2 and NS3 of dengue virus type 2) has been reported earlier. This study was carried out to establish an acute toxicity profile and evaluate the anti-fever (anti-pyretic) activities of EAEZZR in yeast-induced fever in rats. The major compound of EAEZZR, zerumbone, was isolated using chromatographic methods including column chromatography (CC) and preparative thin-layer chromatography (PTLC). Additionally, the structure of zerumbone was elucidated using nuclear magnetic resonance (NMR), liquid chromatography mass spectrometer-ion trap-time of flight (LCMS-IT-TOF), infrared (IR), and ultraviolet (UV) spectroscopy. The toxicity of EAEZZR was evaluated using Organization for Economic Cooperation and Development Test Guideline 425 (OECD tg-425) with minor modifications at concentrations EAEZZR of 2000 mg/kg, 3000 mg/kg, and 5000 mg/kg. Anti-fever effect was determined by yeast-induced fever (pyrexia) in rats. The acute toxicity study showed that EAEZZR is safe at the highest 5000 mg/kg body weight dose in Sprague Dawley rats. Rats treated with EAEZZR at doses of 125, 250, and 500 mg/kg exhibited a significant reduction in rectal temperature (TR) in the first 1 h. EAEZZR at the lower dose of 125 mg/kg showed substantial potency against yeast-induced fever for up to 2 h compared to 0 h in controls. A significant reduction of TR was observed in rats treated with standard drug aspirin in the third through fourth hours. Based on the present findings, ethyl acetate extract of Zingiber zerumbet rhizomes could be considered safe up to the dose of 5000 mg/kg, and the identification of active ingredients of Zingiber zerumbet rhizomes may allow their use in the treatment of fever with dengue virus infection.

Effect of Mucuna cochinchinensis seed extract on Alloxan-Induced Diabetic Experimental Rats

Objective: Mucuna cochinchinensis (Fabaceae)is a widely grown twiner that has been used in the traditional medicine for treating many ailments. The objective of the present study was to evaluate the hypoglycemic effects of ethyl acetate extract of M. cochinchinensis seeds on alloxan-induced diabetic rat models. Materials and Methods: M.cochinchinensis seeds were collected and macerated with ethyl acetate to obtain ethyl acetate extract of M. cochinchinensis (EMC). The obtained extract was subjected to preliminary phytochemical analysis using standard protocol and HPLC-MS study to identify the phytoconstituents present. Effect of EMC on parameters like blood glucose and plasma insulin levels were assessed using commercial kits in alloxan-induced experimental rats. Results: Preliminary phytochemical analysis of EMC showed the presence of alkaloids, flavonoids, terpenoids, saponins, tannins and glycosides. HPLC-MS study revealed the presence of 6-C-Pentosyl-8-C-hexosyl apigenin and kaempferol-3-O-rutinoside in EMC. EMC effect on alloxan-induced diabetic ratsblood glucose level on 1st, 7th and 14th day and plasma insulin levels were determined and was found to revert the blood glucose levels significantly (p<0.01). Conclusion: To conclude, EMC has potent efficacy in normalizing blood glucose and serum insulin concentration in alloxan induced diabetic rats.

Isolation of Endophytic Fungi from Adhatoda vasica and Assessing its Antibacterial Efficacy and Cytotoxic Impact on the A549 Cancer Cells

The current study investigated the anti-bacterial and  anti-cancer properties of endophytic fungi obtained from medicinal plant Adhatoda vasica. We isolated and identified the endophytic fungi Humicola fuscoatra. by examining their morphological characteristics and microscopic structures. We then extracted their secondary metabolites using ethyl acetate. The ethyl acetate extracts of the fungus H. fuscoatra have shown promising effectiveness against human pathogens, including Klebsiella pneumoniae, Escherichia coli, Bacillus subtilis, and Staphylococcus aureus. A strain of H. fuscoatra exhibited significant antibacterial activity. In addition, we have assessed the effects of ethyl acetate extracts from the fungus H. fuscoatra on the growth of the human lung cancer cell line A549 in vitro assays. Using ethyl acetate extracts from the fungus H. fuscoatra effectively suppressed the growth, survival, migration, and invasion of cancer cells. In this paper we studied in detail the identification of isolated endophytic fungi H. fuscoatra from Adhatoda vasica and characterized its active metabolite compounds. The ethyl acetate-extracted secondary metabolites from the fungal culture supernatant were analyzed using gas chromatography and mass spectrometry, which detected the presence of Hexadecanoic Acid, Hexadecane, Heptadecane, Octadecane, Octadecanoic acid, phytochemicals. The results suggest that the endophytic fungus H. fuscoatra found in medicinal plants has the potential to produce bioactive compounds. They could also be used as a biocontrol agent against bacterial pathogens and for therapeutic purposes in the A549 lung cancer cell line.

Exploring the underlying mechanism of ethyl acetate extract of Annona muricata leaves via the NRF2 and NF-ΚB pathway in type 1 diabetic Wistar rats

Type 1 diabetes mellitus is an autoimmune disorder characterized by the destruction of pancreatic beta cells, leading to impaired insulin production and regulation. Recent research has shown that medicinal plants with antioxidant and anti-inflammatory properties hold promise as potential therapeutic agents for managing complications of the disease. The aim of this study is to investigate the anti-diabetic, antioxidant, and anti-inflammatory effect of ethyl acetate extract of Annona muricata leaves in streptozotocin (STZ)-induced diabetes in Wistar rats. Thirty five (35) male Wistar rats weighing 150-180g were used for this experiment. Group 1 (Normal control), Group 2 (STZ 40 mg/kg only Diabetic control), Group 3 (STZ+ metformin 500mg/kg bw), Group 4 (STZ + ethyl acetate extract 250 mg/kg bw), Group 5 (STZ + ethyl acetate extract (500 mg/kg bw). Fasting blood samples were collected by cardiac puncture for glucose estimation and liver tissues excised for both biochemical and molecular studies. The study demonstrated the extract's ability to modulate the NF-κB pathway, leading to the suppression of inflammatory responses in diabetic rats. These findings suggest that the ethyl acetate extract of A. muricata leaves may ameliorate type 1 diabetes by promoting antioxidant defense and reducing inflammatory processes through the NRF2 and NF-κB pathways. This research sheds light on the potential of natural compounds as adjunct therapies for type 1 diabetes and provides a foundation for further investigations into the development of novel treatments targeting oxidative stress and inflammation in autoimmune diabetes.

Effects of ethyl acetate extracts of Marsilea crenata Presl. leaves on risks of atherosclerosis in dyslipidemic rats

Background: Dyslipidemia is widely known as a significant contributor to the incidence of cardiovascular diseases (CVD). Over the years, traditional therapeutic strategies, including the use of herbs, have been recognized. Marsilea crenata Presl. (MC), or a plant known for its phytoestrogen content, has shown biological activities similar to oestrogen. These activities comprise the reduction of serum cholesterol levels, thereby having potential roles in managing dyslipidaemia and its associated cardiovascular risks.Objective: This study aims to examine the effects of ethyl acetate extracts of MC leaves on the serum lipid profile of experimental rats with dyslipidaemia induced by high-fat diet (HFD). Methods: This study applied a pretest-post-test study design with control group, involving 30 rats which are divided into 5 treatment groups, namely N (normal), HFD, STA (Simvastatin), MC1, MC2, and MC3. Group N was only treated with standard feed, and the others received HFD on days 1-14. On the 15th – 28th day, the HFD group was treated with standard feed, and the STA group received simvastatin 0.36 mg/200 g BW/day. The groups of MC1, MC2, and MC3 were treated with oral MC extracts at 100, 200, and 400 mg/kgBW, respectively. Their body weight was measured at the beginning of the study, as well as after 2 and 4 weeks. Subsequently, the experimental rats were fasted overnight on the 14th and 28th day. Their blood was collected on the 15th day (pre-test) and 29th day (post-test) to measure serum concentration of total cholesterol (TC), triglycerides (TG), and high-density lipoprotein cholesterol (HDL-c). Low-density lipoprotein cholesterol (LDL-c) concentration was also calculated by using the Friedewald formula. Results: MC3 extracts significantly reduced the TC (-40.95 ± 1.77), TG (-46.12 ± 3.79), and LDL-c levels (-0.017 ± 0.010), and it also increased levels of HDL-c (11.28 ± 2.34) mg/dL in the HFD-induced rats.Conclusion: Ethyl acetate extracts of MC leaves could reduce risk factors for atherosclerosis by returning lipid profile of the dyslipidemic rats to normal conditions with an increased dose.

Apoptosis-driven synergistic anti-cancer efficacy of ethyl acetate extract of Memecylon sisparense Gamble leaves and doxorubicin in in-vitro and in-vivo models of triple-negative breast cancer

In the spectrum of breast neoplasms, approximately 15 to 20% of all diagnosed cases are triple-negative breast carcinoma. TNBC grows and spreads faster than other invasive breast cancers and has a worse prognosis. The existing therapies and chemotherapeutic drugs have several limitations, so the development of safe and affordable treatment options is currently in demand. Hence, this research focuses on scientifically evaluating the therapeutic anticancer effect of ethyl acetate extract of MSG and its combined efficacy with doxorubicin against TNBC. MSG has shown an IC50 value of 48.40±1.68µg/ml on the MDA-MB-231 cell line, and the combination of MSG with Dox demonstrated the synergistic effect. Apoptotic changes such as membrane blebbing chromatin condensation were observed in MSG alone and in combination with doxorubicin treatments. Apoptosis was confirmed with Annexin V-FITC/PI staining and increased apoptotic markers such as Cleaved caspase-3 Bax and decreased anti-apoptotic markers Bcl-2 by western blotting. The tumor burden significantly decreased in MSG and combination treatment groups while restoring their body weights. Meanwhile, the Dox-treated group indicated a decreased tumor burden combined with weight loss. The present investigation revealed that MSG and doxorubicin have a synergistic anticancer effect in TNBC.

The antioxidant, anti-inflammatory and analgesic activity effect of ethyl acetate extract from the flowers of Syringa pubescens Turcz

Ethnopharmacological relevanceSyringa Pubescens Turcz. (SP), a member of the Oleaceae family, is a species of plant known as Syringa. Flowers, as the medicinal part, are commonly used in the treatment of hepatitis and tonsillitis.Aim of the study: The research was the first to assess the antioxidant and anti-inflammatory potential of different parts of SP flowers (SPF) in vitro. The most promising fraction was ethyl acetate fraction of SP flower (SPFEA). The antioxidant, anti-inflammatory and analgesic activities of SPFEA were further studied, and the chemical components were identified. MethodsHPLC was used to identify the major components in various fraction of SPF. DPPH and ABTS + radical scavenging assays as well as FRAP test and β-carotene bleaching test were employed to assess the antioxidant potential of SPF fraction in vitro. The inhibitory effect on NO production in LPS-treated RAW264.7 cells and heat-induced protein denaturation test were used to evaluate the anti-inflammatory potential of SPF fraction. Further analysis of the biological activity of SPFEA was performed. Acute toxicity test was conducted to assess the toxicity of SPFEA. The anti-inflammatory effect was assessed by utilizing xylene induced ear edema model, carrageenan-induced foot edema model and peritonitis model in vivo. The analgesic effect of SPFEA was evaluated using hot plate test, tail immersion test, formaldehyde test as well as acetic acid-induced abdominal writhing pain experiment in vivo. In carrageenan induced foot edema model, ELISA kits were employed to measure levels of inflammation factors (NO, TNF-α, IL-6, COX-2, IL-1β) in foot tissue as well as MDA, CAT, SOD, GSH-PX levels in liver tissue. ResultsHPLC results showed that there were significant differences in bioactive substances among different fractions of SPF, and SPFEA was rich in bioacitve components. Compared with other fractions of SPF, SPFEA exhibited better antioxidant and anti-inflammatory abilities. The 3000 mg/kg SPFEA group in mice had no obvious side effects. The xylene-induced ear edema model, carrageenan-induced foot edema and peritonitis models demonstrated that the SPFEA had significant anti-inflammatory effect. Moreover, inflammation factors including NO, TNF-α, IL-6, COX-2, IL-1β were significantly reduced in SPFEA groups in foot tissue induced by carrageenan. Additionally, SPFEA effectively decreased liver tissue oxidative stress levels (MDA, SOD, GSH-PX and CAT). The bioactivities of SPFEA demonstrated a clear dose-dependent relationship. The results of the hot plate test, tail immersion test, formaldehyde test and acetic acid-induced abdominal writhing pain experiments indicated the SPFEA possessed an excellent analgesic effect, and this effect was in dose-dependent manner. ConclusionThe study provides a scientific foundation for understanding the pharmacological action of SPFEA. It has been indicated that SPFEA has excellent antioxidant, analgesic and anti-inflammatory effects.

Effect of Usnea longissima ethyl acetate extract on acute oxidative and inflammatory lung damage from Staphylococcus aureus infection in rats.

The role of oxidants and proinflammatory cytokines in the pathogenesis of pneumonia caused by Staphylococcus aureus (S. aureus) has been demonstrated. The present study aims to investigate the protective effect of ethyl acetate extract (EtOAc) obtained from Usnea longissima (UL) against acute oxidative and inflammatory lung damage due to S. aureus infection in rats. Albino Wistar-type male rats were divided into three groups: Healthy (HG), S. aureus inoculated (SaG), and S. aureus inoculated + ULEtOAc administered (SUL). SaG (n = 6) and SUL (n = 6) group rats' left nostrils (excluding HG) were inoculated with 0.1 ml bacterial mixture. After 24 hours, ULEtOAc (50 mg/kg) was administered orally to the SUL group, and the same volume of normal saline was administered orally to the HG (n = 6) and SaG groups. This procedure was performed once a day for seven days. Levels of oxidant and antioxidant parameters such as malondialdehyde (MDA) and total glutathione (tGSH), as well as pro-inflammatory cytokine levels such as nuclear factor-kappa B (NF-κB), tumor necrosis factor-alpha (TNF-α), interleukin-one beta (IL-1β), were measured in removed lung tissues. Tissues were also examined histopathologically. Biochemical results showed that ULEtOAc significantly suppressed the increase of MDA, NF-κB, TNF-α, and IL-1β levels and the decrease of tGSH caused by S. aureus in lung tissue. S. aureus inoculation caused severe mononuclear cell infiltration in interstitial areas, severe lymphoid hyperplasia in bronchial-associated lymphoid tissue and severe alveolar edema, histopathologically. Treatment with ULEtOAc had an attenuating effect on these histopathological findings. Experimental results from this study suggest that ULEtOAc may be beneficial in treating S. aureus-induced oxidative and inflammatory lung damage.

Evaluation of Anti-Escherichia coli K88 from Diarrhea Piglets and Phytochemicals of Galangal Rhizome Extracts

This study investigated the antimicrobial effect of galangal rhizome extracts against Escherichia coli K88 from diarrhea pigs. Galangal rhizome was extracted with 4 solvents: Water, ethanol, methanol and ethyl acetate. The extracts determination against E. coli K88 by disc diffusion method, minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC). Results showed that inhibition diameters of galangal extract with ethyl acetate against E. coli K88 was significantly higher than that of galangal extracted with water, ethanol and methanol. However, the inhibition was significantly lower than that of streptomycin drug. The MIC and MBC values ​​of ethyl acetate extracts against E. coli K88 were 0.39 and 0.78 mg/mL, respectively. Standard bacteria, pathogenic S. aureus ATCC6538, also used for testing antibacterial effect of galangal extract revealed the ethanol and methanol and ethyl-acetate extracts antibacterial effect against S. aureus. The phytochemicals tests performed on the extracts of galangal rhizome confirms the presence phenolic compounds exhibited antimicrobial activity. This study demonstrated antibacterial efficacy of galangal extracted with ethyl acetate to further develop as a substitute for antibiotics. HIGHLIGHTS The galangal rhizome extracted with ethyl acetate exhibited a strong antimicrobial effect against E. coli K88, which known to cause diarrhea in piglets The observed antibacterial effect of ethyl acetate extract of galangal was attributed to the high presence of specific compounds, namely phenol, 4-(2-propenyl), acetate, as well as 3 sesquiterpenoids compounds that were exclusively found in ethyl acetate extract These sesquiterpenoids include 3-(1,5-dimethyl-4-hexenyl)-6-methylene-1-cyclohexene; and 1,5-dimethyl-8-(methylethylidene)-1,5-cyclodecadiene and (-)-5-oxatricyclo[8.2.0.0(4,6)] dodecane, 12-trimethyl-9-methylene; and 3-phenyl-1,4(e)-dodecadiene GRAPHICAL ABSTRACT

Schinus terebinthifolius Raddi (Brazilian pepper) leaves extract: in vitro and in vivo evidence of anti-inflammatory and antioxidant properties.

The aim of this work was to evaluate the anti-inflammatory and antioxidant effects of ethyl acetate extract obtained from the leaves of Brazilian peppertree Schinus terebinthifolius Raddi (EAELSt). Total phenols and flavonoids, chemical constituents, in vitro antioxidant activity (DPPH and lipoperoxidation assays), and cytotoxicity in L929 fibroblasts were determined. In vivo anti-inflammatory and antioxidant properties were evaluated using TPA-induced ear inflammation model in mice. Phenol and flavonoid contents were 19.2 ± 0.4 and 93.8 ± 5.2 of gallic acid or quercetin equivalents/g, respectively. LC-MS analysis identified 43 compounds, of which myricetin-O-pentoside and quercetin-O-rhamnoside were major peaks of chromatogram. Incubation with EAELSt decreased the amount of DPPH radical (EC50 of 54.5 ± 2.4µg/mL) and lipoperoxidation at 200-500µg/mL. The incubation with EAELSt did not change fibroblast viability up to 100µg/mL. Topical treatment with EAELSt significantly reduced edema and myeloperoxidase activity at 0.3, 1, and 3mg/ear when compared to the vehicle-treated group. In addition, EAELSt decreased IL-6 and TNF-α levels and increased IL-10 levels. Besides, it modulated markers of oxidative stress (reduced total hydroperoxides and increased sulfhydryl contents and ferrium reduction potential) and increased the activity of catalase and superoxide dismutase, without altering GPx activity.

Terminalia bellirica Fruit Extract Alleviates DSS-Induced Ulcerative Colitis by Regulating Gut Microbiota, Inflammatory Mediators, and Cytokines.

Ulcerative colitis (UC) is a chronic inflammatory disease significantly impacting patients' lives. This study aimed to elucidate the alleviating effect of ethyl acetate extract (TBEA) from Terminalia bellirica fruit on UC and to explore its mechanism. TBEA was the fraction with the best anti-inflammatory activity screened using in vitro anti-inflammatory assays, and HPLC initially characterized its composition. The mice model of ulcerative colitis was established after free drinking of 2.5% dextran sulfate sodium for six days, and the experimental group was treated with 50 mg/kg and 100 mg/kg TBEA for seven days. We found that TBEA significantly alleviated symptoms in UC mice, including a physiologically significant reduction in disease activity index and pathological damage to colonic tissue. TBEA dramatically slowed down oxidative stress and inflammatory process in UC mice, as evidenced by decreasing myeloperoxidase and malondialdehyde activities and increasing glutathione and catalase levels by reducing the concentrations of IL-6, IL-1β, TNF-α, and NO in UC mice, as well as by regulating key proteins in the IL-6/JAK2/STAT3 pathway. Meanwhile, TBEA maintained intestinal homeostasis by regulating intestinal flora structure. Our study provides new ideas for developing TBEA into a new drug to treat UC.

Antimicrobial and Cytotoxic Activities of the Secondary Metabolites of Endophytic Fungi Isolated from the Medicinal Plant Hyssopus officinalis.

According to the World Health Organization, it is estimated that by 2050, drug-resistant infections could cause up to 10 million deaths annually. Therefore, finding a new generation of antibiotics is crucial. Natural compounds from endophytic fungi are considered a potential source of new-generation antibiotics. The antimicrobial and cytotoxic effects of ethyl acetate extracts of nine endophytic fungal isolates obtained from Hyssopus officinalis were investigated for bioassay-guided isolation of the natural compounds. An extract of isolate VII showed the highest antimicrobial activities against Gram-positive bacteria Bacillus subtilis and Staphylococcus aureus (30.12 ± 0.20 mm and 35.21 ± 0.20 mm) and Gram-negative bacteria Escherichia coli and Pseudomonas aeruginosa (30.41 ± 0.23 mm and 25.12 ± 0.25 mm) among the tested extracts of isolates. Molecular identification of isolate VII confirmed it as Chaetomium elatum based on sequencing of its ITS genes, and it was discovered that this was the first time C. elatum had been isolated from H. officinalis. This isolate was cultured at a large scale for the isolation and identification of the active compound. Penicillic acid was isolated for the first time from C. elatum and its chemical structure was established by NMR spectroscopy. The penicillic acid showed strong antibacterial activities against Bacillus subtilis and Staphylococcus aureus with 20.68 mm and 25.51 mm inhibition zones, respectively. In addition, MIC and MBC values and antibiofilm activities of penicillic acid were determined. It was found that penicillic acid reduced the level of biofilms in proportion to antibacterial activity.

Ethyl acetate extract of Gastrodiaelata protects Caenorhabditis elegans from oxidative stress and amyloid β peptide toxicity.

Gastrodia elata Blume is a traditional Chinese medicine with a long history, which has numerous pharmacological activities, such as anti-inflammation, anti-oxidation and protection of nerves. The present study investigated the regulatory effect of ethyl acetate extract of Gastrodia elata (EEGE) on the β-amyloid (Aβ) toxicity of Caenorhabditis elegans (C. elegans). First, the main components of EEGE were analyzed using high-performance liquid chromatography, and the total phenols, total flavonoids and total antioxidant capacity of EEGE were determined. Next, the regulation effect of EEGE on Aβ-induced toxicity of C. elegans was evaluated through experiments on nematode paralysis, lifespan, oxidative and heat stress, locomotor ability, reproductive ability, reactive oxygen species (ROS) level, Aβ aggregation test, malondialdehyde (MDA) level, catalase (CAT) activity and superoxide dismutase (SOD) activity. Finally, the mechanism of EEGE was elucidated using RNA sequencing (RNA-Seq) and the expression levels of related genes were verified using quantitative PCR. The present study revealed that the main components of EEGE included phosphorylated (p)-hydroxybenzyl alcohol, p-hydroxybenzaldehyde and 4,4'-dihydroxydiphenylmethane, possessing strong in vitro free radical scavenging and reducing abilities. In addition, after the intervention of EEGE, the paralysis of nematodes could be delayed, the survival time of the nematodes was prolonged, the survival rate of the nematodes under stress (high temperature and oxidation) conditions was improved, the activity capacity and reproductive capacity of the nematodes were improved, the activities of SOD and CAT were improved and the levels of ROS and MDA were reduced. Notably, EEGE directly inhibited Aβ plaque aggregation in nematodes. RNA-Seq analysis showed that EEGE regulated metabolism and longevity-related genes, and these genes were regulated by the insulin/IGF-1 signaling (IIS) pathway. Therefore, the present study hypothesized that the regulatory mechanism of EEGE was significantly related to the IIS pathway. The present research results demonstrated that the protective effect of EEGE on transgenic C. elegans was to reduce Aβ protein aggregation, improve the in vivo antioxidant level, effectively remove free radicals and to regulate the expression of genes related to IIS pathway, thereby reducing Aβ-induced toxicity and delaying nematode paralysis.

In vitro Evaluation of the Potent Antileishmanial Activity of Ferula tabasensis Alone or in Combination with Shark Cartilage Extract Against the Standard Iranian Strain of Leishmania major (MRHO/IR/75/ER).

The available drugs for the treatment of leishmaniasis are highly toxic and extremely expensive, with low efficiency; therefore, the development of effective therapeutic compounds is essential. The present study aimed to explore the antileishmanial effects of ethyl acetate extract, methanol extract, and fractions 1-4 (F1-F4) of Ferula tabasensis, alone or in combination with shark cartilage extract (ShCE), on L. major in vitro. In this study, ethyl acetate, methanol, and n-hexane extracts were extracted from the aerial roots of F. tabasensis by the maceration method. The silica gel column chromatography was used to separate n-hexane extracts at varying polarities (F1-F4 fractions). Subsequently, the effects of extracts and fractions against promastigotes were assessed by the parasite counting method microscopic inhibition test and MTT assay. Besides, their effects on the infected macrophage cells and the number of amastigotes were investigated. Cytotoxicity was evaluated in non-infected J774A.1 macrophage cells. Finally, apoptosis induction of promastigotes, including infected and non-infected macrophages, was evaluated. The results indicated the highly potent activity of F. tabasensis extracts and F1-F4 fractions, alone or in combination with ShCE, against L. major promastigotes and amastigotes in a dose-dependent manner (P < 0.05). The F1 fraction and methanol extract showed markedly higher toxicity compared to the other extracts and fractions, with 50% inhibitory concentrations (IC50/72h) of 2.4 ± 0.29 and 2.9 ± 0.55 µg/mL against promastigotes and 1.79 ± 0.27 µg/mL and 1.39 ± 0.27 µg/mL against amastigotes (P < 0.001). Moreover, they had a high selectivity index (SI) due to the low toxicity of macrophages (P < 0.0001). The results of flow cytometry indicated that the percentages of apoptotic promastigote cells in contact with IC50 concentrations of F1 and methanol extract alone after 72 h were 43.83 and 43.93%, as well as 78.4%, and 65.45% for their combination with ShCE, respectively.Also, apoptosis of infected macrophages induced by F1 and methanol extracts was estimated at 68.5% and 83.7%, respectively. In this study, the F1 fraction and methanol extract of F. tabasensis showed potent efficacy against L. major, associated with low toxicity and apoptosis induction. Therefore, they can be promising therapeutic candidates in future animal and even human studies.

Virucidal antiviral activity of Maytenus quadrangulata extract against Mayaro virus: Evidence for the presence of catechins.

Mayaro virus (MAYV) is an arbovirus endemic to the Amazon region, which comprises the states of the North and Midwest region of Brazil and encompasses the largest tropical forest in the world, the Amazon Forest. The confirmation of its potential transmission by Aedes aegypti and recent cases in Brazil, mainly in large centers in the northern region, led to the classification of Mayaro fever as an emerging disease. Traditional medicine is commonly used to treat various diseases, mainly by local riverside populations. Some species of the genus Maytenus, which have similar morphologies, are popularly used to treat infections and inflammations. In this context, our research group has studied and confirmed the antiviral activity of several plant-derived compounds. However, several species of this same genus have not been studied and therefore deserve attention. This study aimed to demonstrate the effects of ethyl acetate extracts of leaves (LAE) and branches (TAE) of Maytenus quadrangulata against MAYV. Mammalian cells (Vero cells) were used to evaluate the cytotoxicity of the extracts. After cell infection by MAYV and the treatment with the extracts, we evaluated the selectivity index (SI), the virucidal effect, viral adsorption and internalization, and the effect on viral gene expression. The antiviral action was confirmed by quantifying the viral genome using RT-qPCR and by analyzing the effect on virus yield in infected cells. The treatment was performed based on the effective concentration protective for 50% of the infected cells (EC50). The leaves (LAE; EC50 12.0μg/mL) and branches (TAE; EC50 101.0μg/mL) extracts showed significative selectivity against the virus, with SI values of 79.21 and 9.91, respectively, which were considered safe. Phytochemical analysis revealed that the antiviral action was associated with the presence of catechins, mainly in LAE. This extract was chosen for the subsequent studies since it reduced the viral cytopathic effect and virus production, even at high viral loads [MOI (multiplicity of infection) 1 and 5]. The effects of LAE resulted in a marked reduction in viral gene expression. The viral title was drastically reduced when LAE was added to the virus before infection or during replication stages, reducing virus production up to 5-log units compared to infected and untreated cells. Through kinetic replication, MAYV was not detected in Vero cells treated with LAE throughout the viral cycle. The virucidal effect of LAE inactivates the viral particle and can intercept the virus at the end of the cycle when it gains the extracellular environment. Therefore, LAE is a promising source of antiviral agents.

Ethyl Acetate Extract of Halymenia durvillei Induced Apoptosis, Autophagy, and Cell Cycle Arrest in Colorectal Cancer Cells.

Colorectal cancer is one of the most death-dealing cancers. However, conventional cancer treatments still have side effects. Therefore, novel chemotherapeutic agents with less side effects are still in search. A marine red seaweed, Halymenia durvillei, is recently interested in its anticancer effects. This study investigated the anticancer effect of ethyl acetate extract of H. durvillei (HDEA) on HT-29 colorectal cancer cells in association with the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling pathway. HDEA-treated HT-29 and OUMS-36 cells were used for cell viability tests by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide assay. The effects of HDEA on apoptosis and cell cycle were evaluated. The nuclear morphology and mitochondrial membrane potential (ΔΨm) were observed by Hoechst 33342 and JC-1 staining, respectively. The gene expression of PI3K, AKT, and mTOR genes was evaluated using a real-time semiquantitative reverse transcription-polymerase chain reaction. The corresponding protein expressions were assessed by western blot analysis. The result revealed that the cell viability of treated HT-29 cells diminished while that of OUMS-36 cells was non-significant. By the down-regulation of cyclin-dependent ki-nase 4 and cyclin D1, HDEA-treated HT-29 cells were arrested in the G0/G1 phase. By the up-regulation of cleaved poly(adenosine diphosphate-ribose) polymerase, caspase-9, caspase-8, caspase-3, and Bax, HDEA-treated HT-29 cells underwent apoptosis, but suppressed Bcl-2, disrupted nuclear morphology and ΔΨm. Furthermore, treated HT-29 cells underwent autophagy by up-regulation of light chain 3-II and beclin-1. Lastly, HDEA suppressed the expression of PI3K, AKT, and mTOR. Therefore, HDEA exerts anticancer effects against HT-29 cells, confirmed by apoptosis, autophagy, and cell cycle arrest induction via regulation of the PI3K/AKT/mTOR signaling pathway.

Polyphenolic Profiles and Cytotoxic Effect of Iraqi Morus alba leaves Ethyl Acetate Extract.

Morus alba, member of the Moraceae family, is a perennial tree utilized in folk medicine, preparing the modern drug, and considered the main food for silkworms. However, data on chemical content in the leaves is still limited; the main objective of this study is to detect the presence and determine the concentration of different polyphenolic constituents in the leaves of the Morus alba plant by reverse phase-high performance liquid chromatography (RP-HPLC) and evaluate the cytotoxic effect of ethyl acetate extract of this plant on human breast cancer (AMJ-13) cell line. Phytochemical analysis of the Morus alba leaves ethyl acetate extract led to identifying and quantification of six polyphenolic constituents designated as phenolic acids (caffeic, chlorogenic, and p-coumaric acid), catechins (epicatechin) and flavonoids (luteolin and apigenin) in which p-coumaric acid exhibited the highest concentration follow by luteolin (identified for the first time), chlorogenic acid, caffeic acid, apigenin and epicatechin as the least. According to estimates, the ethyl acetate extract with a high concentration of polyphenolic constituents gave the best findings as cytotoxic against breast cancer AMJ-13 cell line with an IC50 value of 129.5 µg/ml.

Immunomodulatory Effects of Phaleria macrocarpa Leaf Extract in Normal And Cyclophosphamides Induced in Wistar Rats

Abstract. Cyclophosphamide is an antineoplastic drug belonging to the alkylating agents commonly used in treating cancer. However, the use of cyclophosphamide causes a decrease in the body's immune system by reducing lymphocyte proliferation. The curent study was conducted to evaluate the immunomodulatory effect of ethyl acetate extracts of the leaf of the P. macrocarpa (EEADMD) and the ethanol extracts of the leaf of the P. macrocarpa (EEDMD) in normal and cyclophosphamide induced rats. This study used rats classified into two major groups: normal rats and rats induced by cyclophosphamide which were then given P. macrocarpa leaf extract until day 14. On day 4, the test animals were infected with 1% Staphylococcus aureus suspension in. Induced with cyclophosphamide 70 mg/kg BW was carried out on the 8th and 13th days, and then the immunomodulatory activity was tested using total leukocyte analysis, leukocyte differential, and delayed type hypersensitivity response. The results showed that EEDMD 400 mg/kg BW  in normal rats and EEDMD 100 mg/kg BW in cyclophosphamide induced rats could increase total leukocytes and leukocyte differential with a significant difference to the negative group (p &lt;0.05). The results of the delayed hypersensitivity response test of EEADMD and EEDMD at a dose of 100 mg/kg BW with cyclophosphamide induction and normal rats could give an increase in rat paw volume with a significant difference to the negative group (p &lt;0.05). This study shows that EEADMD and EEDMD have an immunomodulatory effect on increasing total and differential leukocytes and leukocyte differential and delayed type hypersensitivity response.

Wedelolactone, a Component from Eclipta prostrata (L.) L., Inhibits the Proliferation and Migration of Head and Neck Squamous Cancer Cells through the AhR Pathway.

Ecliptae prostrata (L.) L. has been widely used in East Asia with reported biological activities, including anti-cancer properties. We aimed to investigate the effect of ethyl acetate extract of Ecliptae prostrata (L.) L. (EAE) and its component wedelolactone on the proliferation and migration of head and neck squamous cancer cells. The proliferation of human SCC-4 and mouse CU110-1 tongue squamous carcinoma cells was assessed using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) method. Scratch wound assays were performed to assess cell migration rates. The levels of Ecadherin and vimentin were used as markers of the epithelial-to-mesenchymal transition (EMT). AhR, CYP1A1, and CYP1B1 levels were examined to uncover the mechanism of inhibition of cell migration by wedelolactone. We found that EAE and wedelolactone decreased the proliferation of human SCC-4 cells and mouse CU110-1 cells at doses of EAE at > 25 μg/ml and wedelolactone at > 6.25 μg/ml. Similarly, both EAE and wedelolactone produced inhibitory effects against migration, but the effective doses that significantly inhibited migration were lower than those affecting proliferation. Wedelolactone below 12.5 μg/ml inhibited the epithelial-to-mesenchymal transition (EMT) with increased expression of E-cadherin and decreased expression of vimentin in SCC-4 and CU110-1 cells. Further analysis showed wedelolactone inhibited the expression of AhR and its downstream target molecules CYP1A1 and CYP1B1 in both squamous carcinoma cells at the same doses inhibiting cell migration. The addition of benzo (a)pyrene [B(a)P], an agonist of AhR, stimulated migration, especially in the CU110-1 cells with reported cancer stem cell-like characteristics. Instructively, B(a)P reversed the inhibitory effects of wedelolactone on AhR expression and cell migration, suggesting that wedelolactone antagonizes cell migration through the AhR pathway. Moreover, the higher activity of EAE and wedelolactone against the migration of cancer stem-like CU110-1 cells relative to SCC-4 cells suggests selective activity against cancer stem cells. Our study identifies wedelolactone as a major active component of Ecliptae prostrata (L.) L. with promising anti-cancer properties against head and neck squamous cancer cells.

Inhibition Effects of Ethyl Acetate Extracts from Inonotus obliquus on Prion Replication

Inhibition Effects of Ethyl Acetate Extracts from Inonotus obliquus on Prion Replication