Chloroplast genomes of Simarouba Aubl., molecular evolution and comparative analyses within Sapindales.

Simarouba, a neotropical genus in the family Simaroubaceae, currently lacks comprehensive genomic data in existing databases. This study aims to fill this gap by providing genomic resources for three Simarouba species, S. amara, S. versicolor, and S. glauca. It also aims to perform comparative molecular evolutionary analyses in relation to other species within the order Sapindales. The analysis of these three Simarouba species revealed the presence of the typical quadripartite structure expected in plastomes. However, some pseudogenization events were identified in the psbC, infA, rpl22, and ycf1 genes. In particular, the CDS of the psbC gene in S. amara was reduced from 1422 bp to 584 bp due to a premature stop codon. Nucleotide diversity data pointed to gene and intergenic regions as promising candidates for species and family discrimination within the group, specifically matK, ycf1, ndhF, rpl32, petA-psbJ, and trnS-trnG. Selection signal analyses showed strong evidence for positive selection on the rpl23 gene. Phylogenetic analyses indicated that S. versicolor and S. glauca have a closer phylogenetic relationship than S. amara. We provide chloroplast genomes of three Simaruba species and use them to elucidate plastome evolution, highlight the presence of pseudogenization, and identify potential DNA barcode regions.

The complete chloroplast genome of Marupa (Simarouba amara Aubl., Simaroubaceae).

Marupa (Simarouba amara Aublet 1775) is a tropical tree of the family Simaroubaceae. It is commonly used for its wood in the Amazonian forest, and it is an important species for restoring degraded environments. Yet, very little genetic resources are available to study this plant. In this paper, we sequenced for the first time the complete chloroplast genome of Marupa, using Oxford Nanopore long-read technology. The genome is 159,838 bp, includes 131 genes in total and presents a classic quadripartite structure. Its length and structure are similar to those of sister species of the Simaroubaceae family. A maximum likelihood phylogeny of the order Sapindale reveals that Simarouba amara is well positioned in its family. This complete plastome is a first step towards a better analysis of Marupa future evolution.

Glaucarubinone from Simarouba glauca DC. as a potential biocontrol agent for mosquito vector management.

The study explored Simarouba glauca DC. for mosquito larvicidal potential by performing bioactivity-guided chemical investigation of its root extract resulting in isolation of the known bioactive metabolite glaucarubinone (1). Mosquito larvicidal activity of glaucarubinone (1) against the three vector species viz. Anopheles stephensi, Aedes aegypti, and Culex quinquefasciatus was determined using a modified WHO 2005 protocol. It was observed that Culex quinquefasciatus larvae were the most susceptible species with LC50 13.88 ppm and LC90 70.01 ppm followed by Aedes aegypti and Anopheles stephensi at 24 h of exposure. The mode of action as observed microscopically is the lysis of midgut and thorax cells of the third instar larvae. The crystal structure of the glaucarubinone (1) is reported for the first time using X-ray crystallography. This phytochemical product has the potential to act as a green alternative to existing chemical-based insecticides for integrated vector management.

GC-MS analysis and anti-oxidant activity of bioactive compounds of Simarouba glauca leaf extracts.

The aim of this study is to analyse the phytochemical composition, anti-oxidant activity, FT-IR and GC-MS analysis of Simarouba glauca leaf extracts. The chloroform extract exhibited highest T.P.C. (139.82 ± 0.06 mg/g GAE) and highest T.F.C. (41.95 ± 0.50 mg/g QE). The GC-MS analysis confirmed the presence of ten compounds in acetone extract and twenty-two compounds in methanol extract. The GC-MS analysis of acetone fraction showed the major peaks such as Glycerine (13.20%), 1,1,1-trichloro-2-methyl-2-propanol (8.92%), Cyclohexanol,2-methyl-5-(1-methylethenyl)- (6.09%), Tetradecane, 2,6,10-trimethyl (7.68%) and Phytol (7.53%). Some of these compounds exhibit anti-oxidant activities. GC-MS analysis of methanol fraction exhibited the presence of following compounds: Undecane (8.52%), 2,3-Trimethylene-4-pyrone (1.79%), cis-Sinapyl alcohol (2.33%) and Umckalin (1.19%). The acetone and methanol fractions exhibited higher percentage of inhibition in DPPH assay. The overall results of this study indicate that the acetone and methanol fractions have been demonstrated to be efficacious against various diseases.

Experimental investigation on use of hydrotreated Simarouba oil (green diesel) and hydrogen gas in a dual-fuel CI engine.

Hydrogen additives to Simarouba glauca vegetable oil (SO) are a common method for addressing the difficulties in combustion caused by SO's poor physical qualities. This research intends to examine parameters such as performance, emission, and combustion characteristics when hydrogen is used as a direct/indirect addition in a SO-fuelled compression ignition (CI) engine. Hydrogen was directly introduced along with intake air until the knocking limit. Experiment was conducted at different load conduction with SO. Through the hydrotreatment process, hydrogen was used in a roundabout way to convert SO to green diesel. Hydrotreated Simarouba vegetable oil (HTSO) enhanced brake thermal efficiency by 23% at full load, whereas direct hydrogen induction improved it by 9% at knock limit. Hydrogen induction resulted in higher NOx emissions than HTSO, at the expense of a slight rise in smoke emissions. Addition of hydrogen lowered both HC and CO emissions directly and indirectly.

Comparison of Simarouba glauca seed shell carbons for enhanced direct red 12B dye adsorption: Adsorption isotherm and kinetic studies.

This study aims at the transformation of the waste lignocellulosic biomass, Simarouba glauca seed shell obtained from biofuel industries, into a value-added adsorbent for the removal of dyes from aqueous media. The basic dye direct red (DR) 12B was adsorbed using chemically (ZnCl2) and thermally activated Simarouba seed shell carbon (ZASRC and SRC, respectively). Dye removal in batch mode was studied by evaluating adsorbent dosage, contact time, pH, adsorption isotherm and kinetics. Enhanced adsorption of DR12B was attained within 80 min at pH 5 with the maximal adsorption capacity (Q0) of 17.48 and 64.94 mg g-1, for SRC and ZASRC, respectively. Further, the dye removal followed Freundlich isotherm model and pseudo second-order kinetics. The mean-free energy of adsorption demonstrated that dye adsorption onto ZASRC occurs through ion-exchange. Thus, ZASRC can be safely and easily applied for the removal of direct red 12B from aqueous solutions.

Caspase-Dependent Apoptosis Induced by Simarouba Glauca on Human Non-Small-Cell Lung Cancer, A549 Cells.

BACKGROUND: The leaves of Simarouba glauca (S. glauca) have been used as a potential source of anticancer agents in traditional medicine. Attempts have been made to isolate anticancer agents from the leaves of S. glauca. The objective of the present study was to demonstrate the anticancer and apoptotic effect of the leaf extract of petroleum ether (LPE) on human non-small-cell lung cancer A549 cells. METHODS: MTT assay was used to investigate the effect of LPE on the viability of A-549 cells. The apoptotic effect of human lung cancer cells was evaluated using fluorescence staining, acridine orange/ ethidium bromide staining, Hoechst staining, flow cytometry analysis, annexin V staining, and caspase assay. RESULTS: The results showed a direct correlation between the dose and the rate of cytotoxicity. Fluorescence staining revealed apoptotic features, such as blebbing and chromatin condensation. Flow cytometry analysis and annexin V staining revealed phosphatidyl serine externalization. Caspase assay confirmed that the extract inhibited cell death. Caspase 3 expressions indicated that the cell death occurred either through the mitochondrial pathway or the death receptor. CONCLUSIONS: The study revealed that the LPE induced the apoptosis of human non-small-cell lung cancer, A549 cells, either through mitochondrial or death receptor pathway.

Broad-Spectrum Antimicrobial, Antioxidant, and Anticancer Studies of Leaf Extract of Simarouba glauca DC In Vitro.

The current study aimed to screen the preliminary phytochemicals in the leaf extract of the medicinal plant Simarouba glauca and to analyze its potential antimicrobial, antioxidant and anticancer properties. The phytochemical profile of the methanol extract was analyzed, and bioactive compounds were identified using chromatography, FTIR and GCMS. Antimicrobial activity and Minimum Inhibitory Concentration (MIC) were determined against 14 bacterial and 6 fungal strains. Moreover, the synergistic effect of a plant extract with commercially available antibiotics was also evaluated using the checkerboard method. The ethanolic and methanolic extracts showed exclusive activity against S. aureus and profound activity against E. coli and S. marcescens. Upon comparing breakpoints, methanolic extract demonstrated higher antimicrobial activity with a MIC value of 3.2 mg/mL against the test pathogens. Furthermore, the extracts demonstrated potential antioxidant activity; methanol extract had higher antioxidant potential compared to the ethanol extract. The major proactive bioactive compound with maximum antioxidant capacity was observed to be terpenoids. The methanol extract of S. glauca showed significant cytotoxicity against the MCF-7 breast cancer cell line with an IC50 value of 16.12 µg/mL. The overall results of our work provide significant evidence for the usage of methanolic extract of S. glauca as an efficient ethnomedicinal agent and a potential candidate for relieving many human ailments.

The Apoptotic Properties of Leaf Extracts of Simarouba glauca against Human Leukemic Cancer Cells.

BACKGROUND AND OBJECTIVE: Simarouba glauca is a plant belonging to the family of Simaroubaceae. It is a potent source of secondary metabolites. The aim of this study was to evaluate the apoptotic properties of leaf extracts of Simarouba glauca against human leukemic cancer cells. MATERIALS AND METHODS: Cytotoxicity of Simarouba glauca was assessed in the leaf extract of petroleum ether against leukemic cells by MTT assay. To detect the apoptotic features, fluorescence microscopy analysis was done with dual acridine orange/ethidium bromide fluorescent staining and Hoechst staining. To determine the externalization of phosphatidylserine, annexin v staining was done. Mitochondrial or death receptor activation was confirmed by caspase 3 analysis by flow cytometry. RESULTS: This study revealed that Simarouba glauca was able to treat leukemia. Among the four extracts, petroleum ether extract showed a higher order of in vitro anticancer activity. The petroleum ether extract strongly inhibited the proliferation of K562 cell lines with IC50 values of 186 µg/ml. Dual acridine orange/ethidium bromide fluorescent staining and Hoechst staining revealed the characteristic features of apoptosis. Annexin V confirmed early and late stage apoptosis. Caspase-3 analysis revealed that cell death was due to mitochondrial or death receptor activation in mitochondrial pathway. CONCLUSION: These findings suggested that Simarouba glauca leaf extracts inhibited leukemic cells in a time- and dose-dependent manner either through mitochondrial or death receptor activation. The leaf extracts of Simarouba glauca was found to be nontoxic to lymphocytes. It can be concluded that Simarouba glauca is an important source of phytochemicals posing efficacy against leukemic cancer cells.
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Oral acute and sub-chronic toxicity assessment of aqueous leaf extract of Simarouba glauca DC (Paradise tree).

Simarouba glauca has been widely reported to be effective against a number of diseases and possesses medicinal benefits. Thus, the study was conducted to evaluate the toxic effect of aqueous leaf extract of Simarouba glauca (AESG) on relevant organs of male Wistar rats. The oral acute toxicity of AESG was evaluated according to the method described by Lorke. Sub-chronic toxicity of AESG was carried out in line with the guidelines of the Organization for Economic Co-operation and Development (OECD), using a total of twenty-four (24) male Wistar rats divided into four groups of six rats each. Test rats were orally administered AESG at doses of 500, 1000 and 2000 mg /kg body weight, respectively, daily for thirty (30) days. At the end of the study, rats were fasted overnight and sacrificed; the relevant biochemical and histopathology evaluation was carried out. Statistical analysis was conducted using the GraphPad Prism®, version 7. The data obtained indicated that the LD50 exceeded 5000 mg/kg. There were significant increases (P < 0.05) in percentage (%) body weight of test rats. There were no significant differences (P < 0.05) in mean liver, kidney, and heart weight/body weight (IOW/BWT) ratios. The AST activity was significantly lowered (P < 0.05) in rats administered AESG 2000 mg/kg. The ALP activities were significantly elevated (P < 0.05), while the GGT activities were significantly lowered (P < 0.05) in all groups of rats administered AESG. Plasma conjugated and unconjugated bilirubin were significantly lowered and elevated (P < 0.05), respectively in rats administered AESG 1000 and 2000 mg/kg. Plasma urea was significantly elevated (P < 0.05) in rats given AESG 1000 mg/kg. Test rats given AESG 2000 mg/kg recorded significant reduction (P < 0.05) in plasma sodium ions concentration. Rats given AESG 500 mg/kg recorded significant reduction (P < 0.05) in plasma bicarbonate ion levels. The findings suggest that AESG was not significantly toxic to the liver, kidney, and heart.

Structure of intact chitinase with hevein domain from the plant Simarouba glauca, known for its traditional anti-inflammatory efficacy.

Chitinase from the leaves of Simarouba glauca, a plant used in traditional anti-inflammatory therapy is purified and characterized. Peptide mass finger print analysis revealed the protein as an endo-chitinase which was further confirmed using chitin-agar assay. The enzyme exhibited significant anti-fungal efficacy against phyto-pathogens such as Macrophomina phaseolina, Fusarium oxysporum and Sclerotium rolfsii. Chitinolysis was also examined against insoluble chitin using SEM. Using X-ray diffraction data up to 1.66 Å, the structure was determined by Molecular Replacement using crystal structure of GH19 Chitinase-like protein from Hevea brasiliensis. During structure refinement, an extra domain could be traced and identified as hevein domain. To our knowledge, this is the first report of any chitinase with intact hevein domain. The GH19 chitinase and hevein domains though connected by a lengthy loop, are restricted to be close by disulfide bridges. These bridges connecting each domain with the loop may be important for proper chitin feeding into the active site. By considering reports on hevein and chitinase domains as well as the traditional use of the plant, this report of an intact hevein-chitinase protein and their relative orientation may add further insights for the usefulness of this protein.

Anti-proliferative potential of phytochemical fractions isolated from Simarouba glauca DC leaf.

Emerging advancements in anticancer drug discovery research are leaning towards the plant-based bioactive fractions, which is a cocktail of naturally abundant two or more substances with unique proportions, exhibiting greater potential to combat cancers than the individual molecules. Thus, isolation and characterization of anti-cancer activity enriched fractions from plants is gaining scientific attention. Consistent with this view, one of the evidence-based traditional medicinal plants, well known for its anti-cancer potential, Simarouba glauca (SG) leaf has been scientifically examined to identify and isolate the potent anti-cancer fraction. The dried SG leaves were extracted successively with the solvents of increasing polarity. The phytochemical characterization of obtained extracts and fractions were carried out to determine the phenolic acid composition. All fractions were individually examined for anti-cancer property in cancer cells representing lungs, cervix, breast, colon and rectum in vitro. Among all fractions tested, the chloroform (SGC) and ethyl acetate (SGEA) extracts showed potent anti-proliferative effects by triggering apoptosis. In summary, our findings demonstrate that the extracts SGC and SGEA have potent anti-cancer activities compared to other fractions of SG leaf and thus warrant further pre-clinical studies to establish scientific basis for the anticancer potentials of SG.

Simalikalactone D, a Potential Anticancer Compound from Simarouba tulae, an Endemic Plant of Puerto Rico.

Species of the genus Simarouba have been studied because of their antimalarial and antileukemic activities. A group of oxygenated terpenes called quassinoids have been isolated from species of the Simarouba genus, and are responsible for its therapeutic properties. We hypothesized that Simarouba tulae, an endemic plant from Puerto Rico, is a natural source rich in quassinoid compounds with anticancer activity. The leaves were processed and extracted with solvents of different polarities. The extracts were screened for their antiproliferative activity, and it was shown that the chloroform extract was the most active extract. This extract was purified using different chromatographic techniques to afford the quassinoid simalikalactone D (SKD). This compound was further characterized using NMR and X-ray diffraction analysis. A reassessment of original structural assignments for SKD is proposed. SKD showed high cytotoxicity activity, with an IC50 of 55, 58, and 65 nM in A2780CP20 (ovarian), MDA-MB-435 (breast), and MDA-MB-231 (breast) cell lines, respectively. Exposure to SKD led to 15% inhibition of the migration of MDA-MB-231 cells.

Tricaproin Isolated From Simarouba glauca Inhibits the Growth of Human Colorectal Carcinoma Cell Lines by Targeting Class-1 Histone Deacetylases.

While anticancer properties of Simarouba glauca (SG, commonly known as Paradise tree) are well documented in ancient literature, the underlying mechanisms leading to cancer cell death begin to emerge very recently. The leaves of SG have been used as potential source of anticancer agents in traditional medicine. Recently attempts have been made to isolate anticancer agents from the leaves of SG using solvent extraction, which identified quassinoids as the molecules with tumoricidal activity. However, it is not known whether the anti-cancer potential of SG leaves is just because of quassinoids alone or any other phytochemicals also contribute for the potency of SG leaf extracts. Therefore, SG leaves were first extracted with hexane, chloroform, ethyl acetate, 70% ethanol, water and anti-cancer potential (for inhibiting colorectal cancer (CRC) cells HCT-116 and HCT-15 proliferation) determined using Sulforhodamine-B (SRB) assay. The chloroform fraction with maximal anticancer activity was further fractionated by activity-guided isolation procedure and structure of the most potent compound determined using spectral analysis. Analysis of the structural characterization data showed the presence of tricaproin (TCN). TCN inhibited CRC cells growth in a time- and dose dependent manner but not the normal cell line BEAS-2B. Mechanistically, TCN reduced oncogenic Class-I Histone deacetylases (HDACs) activity, followed by inducing apoptosis in cells. In conclusion, the anti-cancer potential of SG is in part due to the presence of TCN in the leaves.

Testing the monophyly of Simaba (Simaroubaceae): Evidence from five molecular regions and morphology.

Generic circumscriptions in the mostly pantropical family Simaroubaceae are somewhat controversial. Simaba is the largest genus, currently defined as exclusively neotropical, with around 25 species of trees and shrubs, but both its limits and infrageneric classification have been a matter of discussion and divergence. Traditionally, species of the genus have been treated in three sections: Simaba sect. Tenuiflorae, S. sect. Floribundae and S. sect. Grandiflorae, but a phylogenetic analysis suggested that the latter two may not be monophyletic. To test the monophyly of Simaba and its infrageneric classification, we used a molecular approach based on DNA sequence data from two nuclear ribosomal spacer regions (ITS and ETS) and three plastid regions (rps16 intron, and intergenic spacers psbA-trnH and trnL-trnF), including a comprehensive sampling of species from Simaba and closely related genera. We also performed ancestral character reconstructions to identify morphological characters that could serve as synapomorphies for major clades and to explore patterns of homoplasy in the morphological dataset. Our results show Simaba as traditionally circumscribed is not monophyletic, with taxa segregated into two strongly supported but distinct clades, one of which is more closely related to Simarouba. The three main clades that emerged in the phylogeny include a mostly Amazonian Simaba clade (which includes the type species of Simaba and the remaining species of S. sect. Tenuiflorae, here proposed to be recognized as Simaba sensu stricto), a mostly extra-Amazonian Simaba clade (a distinct lineage that will be recognized as Homalolepis, a genus currently treated in synonymy and equivalent to Simaba sections Grandiflorae and Floribundae), and the Simarouba clade (including all of its current species). These three clades are characterized by a combination of morphological characters, described in detail herein, some of which are novel features for Simaba not previously reported in the literature. Mapping character-states on the phylogenetic tree provides tests for evolutionary hypotheses. For example, our reconstruction of habit and geographic distribution suggests that the diversification of several shrubby species within the extra-Amazonian lineage in the South American cerrados probably occurred from ancestors inhabiting tropical forests, involving transitions in morphological and ecological traits.

Antibacterial Activity of Leaf Extract of Annona muricata and Simarouba glauca on Enterococcus faecalis.

AIM: To determine the antimicrobial effect of water extracts of leaves of Annona muricata and Simarouba glauca on Enterococcus faecalis using agar diffusion method. MATERIALS AND METHODS: Dried leaves of A. muricata and S. glauca were powdered and extracted in a soxhlet apparatus. Enterococcus faecalis was grown overnight in Trypticase soy agar plates. About 10 µl of each extract was placed on agar plates and incubated overnight. The zone of inhibition was measured after 24 hours. About 1% sodium hypochlorite and distilled water were used as positive and negative controls. RESULTS: The leaf extract of A. muricata showed similar effectiveness as that of sodium hypochlorite, whereas the leaf extract of S. glauca showed only a slight reduction in growth of E. faecalis. CONCLUSION: Leaf extract of A. muricata can be developed as an alternative to sodium hypochlorite for root canal irrigants. CLINICAL SIGNIFICANCE: Success of endodontic treatment depends on complete disinfection of the root canals. Root canal irrigants have a major role in complete disinfection of the root canals. Chemical root canal irrigants are more or less toxic to the oral environment. In this study, naturally derived leaf extracts of A. muricata and S. glauca are compared with sodium hypochlorite for its effectiveness against E. faecalis - the most common pathogen found in the root canals.

Evidence of the late lignification of the G-layer in Simarouba tension wood, to assist understanding how non-G-layer species produce tensile stress.

To recover verticality after disturbance, angiosperm trees produce 'tension wood' allowing them to bend actively. The driving force of the tension has been shown to take place in the G-layer, a specific unlignified layer of the cell wall observed in most temperate species. However, in tropical rain forests, the G-layer is often absent and the mechanism generating the forces to reorient trees remains unclear. A study was carried out on tilted seedlings, saplings and adult Simarouba amara Aubl. trees-a species known to not produce a G-layer. Microscopic observations were done on sections of normal and tension wood after staining or observed under UV light to assess the presence/absence of lignin. We showed that S. amara produces a cell-wall layer with all of the characteristics typical of G-layers, but that this G-layer can be observed only as a temporary stage of the cell-wall development because it is masked by a late lignification. Being thin and lignified, tension wood fibres cannot be distinguished from normal wood fibres in the mature wood of adult trees. These observations indicate that the mechanism generating the high tensile stress in tension wood is likely to be the same as that in species with a typical G-layer and also in species where the G-layer cannot be observed in mature cells.

Simaroubaceae family: botany, chemical composition and biological activities

The Simaroubaceae family includes 32 genera and more than 170 species of trees and brushes of pantropical distribution. The main distribution hot spots are located at tropical areas of America, extending to Africa, Madagascar and regions of Australia bathed by the Pacific. This family is characterized by the presence of quassinoids, secondary metabolites responsible of a wide spectrum of biological activities such as antitumor, antimalarial, antiviral, insecticide, feeding deterrent, amebicide, antiparasitic and herbicidal. Although the chemical and pharmacological potential of Simaroubaceae family as well as its participation in official compendia; such as British, German, French and Brazilian pharmacopoeias, and patent registration, many of its species have not been studied yet. In order to direct further investigation to approach detailed botanical, chemical and pharmacological aspects of the Simaroubaceae, the present work reviews the information regarding the main genera of the family up to 2013.

Safety evaluation of Simarouba glauca seed fat.

Simarouba glauca DC is a tree of the family Simaroubaceae, which grows well up to 1,000 m above sea level in all types of well-drained soils (pH 5.5 to 8.0) and in places with 250 to 2,500 mm annual rainfall. The seed oil has been extracted both by mechanical expelling and solvent extraction. The fatty acid composition and iodine value of the oil indicate that it possesses saturated (40.8-42.6%), monounsaturated (52.9-55.0%), and polyunsaturated (2.5-3.4%) fatty acid in ratios close to that of palm oil. These characteristics are suitable for its use as edible oil. Acute oral toxicity and safety evaluation in a 13-week feeding trial on albino rats showed that the oil is comparable to groundnut oil in all the parameters.

Compounds from Simarouba berteroana which inhibit proliferation of NF1-defective cancer cells.

A neurofibromatosis type 1 (NF1) based bioassay-guided phytochemical investigation on Simarouba berteroana led to the isolation of one new canthin-6-one-9-methoxy-5-O-ß-D-glucopyranoside (1), seven known canthine alkaloids (2-8), two known quassinoids (9-10) and a known neo-lignan (11). The structures of all compounds were established by HRMS, 1D- and 2D-NMR analysis and comparison with previously reported data. Most of the compounds inhibited the proliferation of an Nf1- and p53-deficient mouse glioma cell line at non-cytotoxic concentrations.