Chemical Characterization and Multidirectional Biological Effects of Different Solvent Extracts of Arum elongatum: in Vitro and in Silico Approaches.

Arum elongatum (Araceae) is widely used traditionally for the treatment of abdominal pain, arterial hypertension, diabetes mellitus, rheumatism and hemorrhoids. This study investigated the antioxidant properties, individual phenolic compounds, total phenolic and total flavonoid contents (HPLC/MS analysis), reducing power and metal chelating effects of four extracts obtained from A. elongatum (ethyl acetate (EA), methanol (MeOH), methanol/water (MeOH/water) and infusion). The inhibitory activity of the extracts were also determined against acetylcholinesterase, butyrylcholinesterase, tyrosinase, amylase and glucosidase enzymes. The MeOH/water extracts contained the highest amount of phenolic contents (28.85 mg GAE/g) while the highest total flavonoid content was obtained with MeOH extract (36.77 mg RE/g). MeOH/water demonstrated highest antioxidant activity against DPPH⋅ radical at 38.90 mg Trolox equivalent per gram. The infusion extract was the most active against ABTS+ ⋅ (133.08 mg TE/g). MeOH/water extract showed the highest reducing abilities with the CUPRAC value of 102.22 mg TE/g and the FRAP value of 68.50 mg TE/g. A strong metal chelating effect was observed with MeOH/water extract (35.72 mg EDTAE/g). The PBD values of the extracts ranged from 1.01 to 2.17 mmol TE/g. EA extract displayed the highest inhibitory activity against AChE (2.32 mg GALAE/g), BChE (3.80 mg GALAE/g), α-amylase (0.56 mmol ACAE/g) and α-glucosidase (9.16 mmol ACAE/g) enzymes. Infusion extract was the most active against tyrosinase enzyme with a value of 83.33 mg KAE/g. A total of 28 compounds were identified from the different extracts. The compounds present in the highest concentration were chlorogenic acids, 4-hydroxybenzoic acid, caffeic acid, p-coumaric acid, ferulic acid, isoquercitrin, delphindin 3,5-diglucoside, kaempferol-3-glucoside and hyperoside. The biological activities of A. elongatum extracts could be due to the presence of compounds such as gallic acid, chlorogenic acids, ellagic acid, epicatechin, catechin, kaempferol, 4-hydroxybenzoic acid, caffeic acid, p-coumaric acid, ferulic acid, quercetin, isoquercitrin, and hyperoside. Extracts of A. elongatum showed promising biological activities which warrants further investigations in an endeavor to develop biopharmaceuticals.

Antibacterial and cytotoxicity evaluation of Arum hygrophilum Bioss.

OBJECTIVE: Arum hygrophilum Bioss is a plant native to Asia, Europe, and Northern Africa. It is consumed as beverages, spices, or cooked leaves to cure gastrointestinal infections and cancer. This study aims to determine the antibacterial and anticancer effectivenesss of A. hygrophilum Bioss. MATERIALS AND METHODS: Using the well-diffusion method, the antimicrobial activity of the plant's aqueous extract and five other organic extracts were evaluated against bacteria often associated with food poisoning. The assessment of the antiproliferative activity by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay was done on five cancerous cell lines and on fibroblasts as a reference cell line. RESULTS: The growth of L. monocytogenes was significantly inhibited by the aqueous and ethanolic extracts. Both extracts had a minimum inhibitory concentration (MIC) of 62.5 mg/mL. The inhibition caused by the methanolic extract had a MIC of 500 mg/mL. The growth of S. aureus and MRSA were inhibited by the aqueous extract with a MIC of 500 mg/mL, while the inhibition caused by the ethanolic extract had a MIC of 250 mg/mL on MRSA and 500 mg/mL on S.aureus. Both strains of S.aureus were also inhibited by the 3-pentanon extract, while the butanol extract only exhibited a moderate growth inhibition against MRSA. The MTT assay showed that the aqueous extract had not affected the proliferation of cancer cell lines. The cytotoxicity of the ethanolic and methanolic extracts had no concentration-inhibition relationship and the IC50 values were above 800 µg/mL for all extracts. CONCLUSIONS: L. monocytogenes, S. aureus, and methicillin-resistant Staphylococcus aureus (MRSA) were inhibited by different Arum extracts. The antibacterial activity of Arum hygrophilum Bioss against foodborne pathogens makes it safe to use as a natural food preservative, and as a source for sanitizers and antimicrobials. Further investigation is recommended to determine the cytotoxicity of the plant against additional cancer cell lines.

Arum conophalloides Aqueous Extract Induced Hepatotoxicity in Rat; Histopathological, Biochemical, and mir-122 Assessments.

BACKGROUND: Arum conophalloides (A. conophalloides) is a wild edible delicate plant, widely used in traditional medicine. OBJECTIVE: This study aimed to examine the effects of A. conophalloides extracts on biochemical, molecular, and histopathological changes in the rat. METHODS: Fifty adult male Sprague-Dawley rats were divided into 5 groups (10 each) as follows: G1 or control, received distilled water; G2 and G3, treated with the aqueous extract at doses of 200 and 400 mg/kg; G4 and G5, treated with the hydroalcoholic extract at doses of 200 and 400 mg/kg. Prior to and at the end of the experiments, the serum levels of biochemistry parameters and the relative expression of miR-122 were assessed. Moreover, the liver and kidney tissues were examined microscopically. RESULTS: Liver and kidney tissues showed normal structure in all groups. There were no significant changes in biochemical indices or the expression of miR-122 in the extract-treated groups at the dose of 200 mg/kg. However, the group that received the aqueous extract at the dose of 400 mg/kg exhibited a significantly lower level of HDL, LDL, ALT, and ALP in comparison to the control. Additionally, miR-122 expression in this group exhibited a 10-fold increase (P=0.009). CONCLUSION: The serum level of hepatocyte-specific miR-122 will be more helpful in detecting hepatic changes in early stages than ALT and AST activity or histopathological evaluations of liver sections. Our findings highlight the potential hepatotoxicity of A. conophalloides aqueous extract in a rat model.

Investigation of the effects of natural compounds isolated from Arum rupicola var. rupicola on glutathione reductase enzyme purified from bovine liver.

Glutathione reductase (GR, E.C. 1.8.1.7), a flavoenzyme, is responsible for recycling of oxidized glutathione disulfide. This study was performed in two main sections. In the first GR was purified from bovine liver by affinity column chromatography and the purification rate and specific activity of the enzyme were calculated as 1832-fold and 141 EU/mg protein, respectively. The subunit molecular weight of the enzyme was determined as 55 kDa by means of SDS-PAGE. The second section isolated natural components of Arum rupicola Boiss. var. rupicola using column chromatography. The isolation protocol for this plant was performed with a series of different-sized columns with hexane-ethyl acetate. According to the thin-layer chromatography plate, seven substances (R1-R7) were isolated. Our study's aim was to find new activators or inhibitors for GR activity. With this aim, all isolated substances were tested for GR activity. R6 showed competitive inhibition, while R4 had noncompetitive inhibition of GR activity. R1 played a role as an activator of GR activity. The inhibitory activity percentage vs. concentration graph was plotted. Values of IC50 for R4 and R6 were calculated as 0.193 mg/mL and 3.98 µg/mL, respectively, from the equation of this graph.

'Last In-First Out': seasonal variations of non-structural carbohydrates, glucose-6-phosphate and ATP in tubers of two Arum species.

Knowledge on the metabolism of polysaccharide reserves in wild species is still scarce. In natural sites we collected tubers of Arum italicum Mill. and A. maculatum L. - two geophytes with different apparent phenological timing, ecology and chorology - during five stages of the annual cycle in order to understand patterns of reserve accumulation and degradation. Both the entire tuber and its proximal and distal to shoot portion were utilised. Pools of non-structural carbohydrates (glucose, sucrose and starch), glucose-6-phosphate and ATP were analysed as important markers of carbohydrate metabolism. In both species, starch and glucose content of the whole tuber significantly increased from sprouting to the maturation/senescence stages, whereas sucrose showed an opposite trend; ATP and glucose-6-phosphate were almost stable and dropped only at the end of the annual cycle. Considering the two different portions of the tuber, both ATP and glucose-6-phosphate concentrations were higher in proximity to the shoot in all seasonal stages, except the flowering stage. Our findings suggest that seasonal carbon partitioning in the underground organ is driven by phenology and occurs independently of seasonal climate conditions. Moreover, our results show that starch degradation, sustained by elevated ATP and glucose-6-phosphate pools, starts in the peripheral, proximal-to-shoot portion of the tuber, consuming starch accumulated in the previous season, as a 'Last In-First Out' mechanism of carbohydrate storage.

Physicochemical properties and combustion behavior of duckweed during wet torrefaction.

Wet torrefaction of duckweed was carried out in the temperature range of 130-250°C to evaluate the effects on physicochemical properties and combustion behavior. The physicochemical properties of duckweed samples were investigated by ultimate analysis, proximate analysis, FTIR, XRD and SEM techniques. It was found that wet torrefaction improved the fuel characteristics of duckweed samples resulting from the increase in fixed carbon content, HHVs and the decrease in nitrogen and sulfur content and atomic ratios of O/C and H/C. It can be seen from the results of FTIR, XRD and SEM analyses that the dehydration, decarboxylation, solid-solid conversion, and condensation polymerization reactions were underwent during wet torrefaction. In addition, the results of thermogravimetric analysis (TGA) in air indicated that wet torrefaction resulted in significant changes on combustion behavior and combustion kinetics parameters. Duckweed samples after wet torrefaction behaved more char-like and gave better combustion characteristics than raw sample.

In vitro and in vivo comparison of the biological activities of two traditionally and widely used Arum species from Jordan: Arum dioscoridis Sibth & Sm. and Arum palaestinum Boiss.

Arum dioscoridis and A. palaestinum (Araceae) are indigenous plant species in Jordan. HPLC-MS analysis of A. dioscoridis revealed the presence of apigenin, luteolin, quercetin, quercetin-3-O-ß-glucoside, vitexin, isoorientin, esculin, and caffeic and ferulic acids. Both Arum spp., influenced gastrointestinal carbohydrate and lipid digestion and absorption. Orlistat inhibited dose dependently and highly substantially pancreatic lipase (PL) in vitro. Similar to orlistat, Arum species aqueous extracts (AEs), apigenin, caffeic acid and esculin exhibited a concentration related PL inhibition. Comparable to acarbose, dual inhibition of α-amylase/α-glucosidase was observed for both Arum species. Like guar gum, A. dioscoridis AE minimised substantially area under 24 h glucose curve. Acute starch-induced postprandial hyperglycaemia in overnight fasting rats was highly significantly (p < 0.001) decreased by A. dioscoridis AE. A. palaestinum could not perform effectively in either starch- or glucose-fed fasting rats. No antiproliferative effects against colorectal cancer cell lines HT29, HCT116 and SW620 were detected for tested Arum spp.

Arum Palaestinum with isovanillin, linolenic acid and ß-sitosterol inhibits prostate cancer spheroids and reduces the growth rate of prostate tumors in mice.

BACKGROUND: Arum palaestinum is a plant commonly found in the Middle East that is ingested as an herbal remedy to fight cancer. However, no studies have examined the direct effect of the plant/plant extract on tumor growth in an animal model. METHODS: Verified prostate cancer cells were plated as 3D spheroids to determine the effect of extract from boiled Arum Palaestinum Boiss roots. In addition, male NU/NU mice (8 weeks old) with xenograft tumors derived from the prostate cancer cell line were treated daily with 1000 mg/kg body weight gavage of the suspension GZ17. The tumor growth was measured repeatedly with calipers and the excised tumors were weighed at the termination of the 3 week study. Control mice (10 mice in each group) received vehicle in the same manner and volume. RESULTS: The number of live prostate cancer cells declined in a dose/dependent manner with a 24 h exposure to the extract at doses of 0.015 to 6.25 mg/mL. A fortified version of the extract (referred to as GZ17) that contained higher levels of isovanillin, linolenic acid and ß-sitosterol had a stronger effect on the cell death rate, shifting the percentage of dead cells from 30 % to 55 % at the highest dose while the vehicle control had no effect on cell numbers. When GZ17 was applied to non-cancer tissue, in this case, human islets, there was no cell death at doses that were toxic to treated cancer cells. Preliminary toxicity studies were conducted on rats using an up-down design, with no signs of toxic effect at the highest dose. NU/NU mice with xenograft prostate tumors treated with GZ17 had a dramatic inhibition of tumor progression, while tumors in the control group grew steadily through the 3 weeks. The rate of tumor volume increase was 73 mm(3)/day for the vehicle group and 24 mm(3)/day for the GZ17 treated mice. While there was a trend towards lower excised tumor weight at study termination in the GZ17 treatment group, there was no statistical difference. CONCLUSIONS: Fortified Arum palaestinum Boiss caused a reduction in live cells within prostate cancer spheroids and blocked tumor growth in xenografted prostate tumors in mice without signs of toxicity.

Total phenolic content, ferric reducing and DPPH scavenging activity of Arum dioscoridis.

Arum dioscoridis, locally called 'Gavur pancari', is a wild plant the leaves of which have been used as vegetable and for preparing special soup which has a sour taste. This study was set up to determine in vitro antioxidant activities and total phenolic and flavonoid contents of different extracts of A. dioscoridis. Free radical scavenging activity was evaluated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing activity with different concentrations of ethanol, methanol, acetone and water extracts of the plant leaves. Total phenolic and flavonoid contents were widely variable depending on solvents. Ethanol and methanol extractions of the plant material showed better performances with respect to both phenolic and flavonoid contents, respectively. The highest phenolic and flavonoid contents of ethanol and methanol extracts were 100.890 mg/g GAE and 72.643 mg/g QE, respectively. The lower DPPH scavenging and ferric reducing activities were determined in comparison with previous reports and standard synthetic chemicals.

Evidence for early intracellular accumulation of volatile compounds during spadix development in Arum italicum L. and preliminary data on some tropical Aroids.

Staining and histochemistry of volatile organic compounds (VOCs) were performed at different inflorescence developmental stages on nine aroid species; one temperate, Arum italicum and eight tropical from the genera Caladium, Dieffenbachia and Philodendron. Moreover, a qualitative and quantitative analysis of VOCs constituting the scent of A. italicum, depending on the stage of development of inflorescences was also conducted. In all nine species, vesicles were observed in the conical cells of either the appendix or the stamens (thecae) and the staminodes. VOCs were localised in intracellular vesicles from the early stages of inflorescence development until their release during receptivity of gynoecium. This localisation was observed by the increase of both number and diameter of the vesicles during 1 week before receptivity. Afterwards, vesicles were fewer and smaller but rarely absent. In A. italicum, staining and gas chromatography analyses confirmed that the vesicles contained terpenes. The quantitatively most important ones were the sesquiterpenes, but monoterpenes were not negligible. Indeed, the quantities of terpenes matched the vesicles' size evolution during 1 week. Furthermore, VOCs from different biosynthetic pathways (sesquiterpenes and alkanes) were at their maximum quantity 2 days before gynoecium receptivity (sesquiterpenes and alkanes) or during receptivity (isobutylamine, monoterpenes, skatole and p-cresol). VOCs seemed to be emitted during gynoecium receptivity and/or during thermogenesis, and FADs are accumulated after thermogenesis in the spadix. These complex dynamics of the different VOCs could indicate specialisation of some VOCs and cell machinery to attract pollinators on the one hand and to repulse/protect against phytophagous organisms and pathogens after pollination on the other hand.

Engineering plant alternative oxidase function in mammalian cells: substitution of the motif-like sequence ENV for QDT diminishes catalytic activity of Arum concinnatum AOX1a expressed in HeLa cells.

Alternative oxidase (AOX) is a nonproton motive quinol-oxygen oxidoreductase which is a component of the mitochondrial respiratory chain in higher plants. In this study, we have characterized the catalytic activity and regulatory behaviors of Arum concinnatum AOX isoforms, namely AcoAOX1a and AcoAOX1b, and their artificial mutants in HeLa cells. We demonstrated that substitution of the motif-like sequence ENV on the C-terminal half of AcoAOX1a for QDT diminishes its activity and proposed that the innate inactivity of AcoAOX1b in HeLa cells is, at least in part, attributable to its QDT motif. Furthermore, we show that introduction of F130L in the hydrophilic N-terminal extension of AcoAOX1a resulted in greater activity in the presence of pyruvate. This result indicates that functional significance of the N-terminal extension is not particular to the conventional regulatory cysteine. On the basis of these findings, we discuss new insights into the structural integrity of AOX in HeLa cells and the applicability of mammalian cells for functional analysis of this enzyme.

Chemosensory ecology: deceiving Drosophila.

The Solomon's lily arum mimics the odours of yeast to attract drosophilid flies as unrewarded pollinators.

A deceptive pollination system targeting drosophilids through olfactory mimicry of yeast.

In deceptive pollination, insects are bamboozled into performing nonrewarded pollination. A prerequisite for the evolutionary stability in such systems is that the plants manage to generate a perfect sensory impression of a desirable object in the insect nervous system [1]. The study of these plants can provide important insights into sensory preference of their visiting insects. Here, we present the first description of a deceptive pollination system that specifically targets drosophilid flies. We show that the examined plant (Arum palaestinum) accomplishes its deception through olfactory mimicry of fermentation, a strategy that represents a novel pollination syndrome. The lily odor is composed of volatiles characteristic of yeast, and produces in Drosophila melanogaster an antennal detection pattern similar to that elicited by a range of fermentation products. By functional imaging, we show that the lily odors target a specific subset of odorant receptors (ORs), which include the most conserved OR genes in the drosophilid olfactome. Furthermore, seven of eight visiting drosophilid species show a congruent olfactory response pattern to the lily, in spite of comprising species pairs separated by ∼40 million years [2], showing that the lily targets a basal function of the fly nose, shared by species with similar ecological preference.

Supramolecular structure of the OXPHOS system in highly thermogenic tissue of Arum maculatum.

The protein complexes of the mitochondrial respiratory chain associate in defined ways forming supramolecular structures called respiratory supercomplexes or respirasomes. In plants, additional oxidoreductases participate in respiratory electron transport, e.g. the so-called "alternative NAD(P)H dehydrogenases" or an extra terminal oxidase called "alternative oxidase" (AOX). These additional enzymes were previously reported not to form part of respiratory supercomplexes. However, formation of respiratory supercomplexes might indirectly affect "alternative respiration" because electrons can be channeled within the supercomplexes which reduces access of the alternative enzymes towards their electron donating substrates. Here we report an investigation on the supramolecular organization of the respiratory chain in thermogenic Arum maculatum appendix mitochondria, which are known to have a highly active AOX for heat production. Investigations based on mild membrane solubilization by digitonin and protein separation by blue native PAGE revealed a very special organization of the respiratory chain in A. maculatum, which strikingly differs to the one described for the model plant Arabidopsis thaliana: (i) complex I is not present in monomeric form but exclusively forms part of a I + III(2) supercomplex, (ii) the III(2) + IV and I + III(2) + IV supercomplexes are detectable but of low abundance, (iii) complex II has fewer subunits than in A. thaliana, and (iv) complex IV is mainly present as a monomer in a larger form termed "complex IVa". Since thermogenic tissue of A. maculatum at the same time has high AOX and I + III(2) supercomplex abundance and activity, negative regulation of the alternative oxidase by supercomplex formation seems not to occur. Functional implications are discussed.

A new pyrrole alkaloid isolated from Arum palaestinum Boiss. and its biological activities.

The phytochemical analysis of the ethyl acetate fraction of Arum palaestinum Boiss. (Araceae) led to the isolation and identification of a new polyhydroxy alkaloid compound; (S)-3,4,5-trihydroxy-1 H-pyrrol-2(5H)-one (1), and other five known compounds; caffeic acid (2), isoorientin (3), luteolin (4) and vicenin 11 (5), as well as the rare compound 3,6,8-trimethoxy, 5,7,3',4'-tetrahydroxy flavone (6). The structural elucidations of all the compounds were based on spectroscopic data (1H- and 13C-NMR, DEPT, HSQC, HMBC and NOE difference techniques) and comparison with literature data. Investigation of the antioxidant activity of the ethyl acetate fraction indicated its strong scavenging capacity for 1,1 -diphenyl-2-picrylhydrazyl (DPPH) radicals (SC50 3.1+/-0.82 microg/mL). Moreover, the treatment of different human cancer cell lines with the ethyl acetate fraction led to dose-dependant suppression in the proliferation of both breast carcinoma cells (MCF-7; IC50 59.09+/-4.1 microg/mL) and lymphoblastic leukemia cells (1301; IC50 53.1+/-2.9 microg/mL); however, it was found to have no effect on the growth of hepatocellular carcinoma cells (Hep G2).

Insecticidal activity of Arum maculatum tuber lectin and its binding to the glycosylated insect gut receptors.

Mannose binding approximately 50 kDa homotetrameric lectin, purified from edible Arum maculatum tuber, was analyzed through SDS-PAGE and studied for its agglutination property using rabbit erythrocytes. Cross reactivity of the purified lectin was verified through western blot using Colocasia esculantum(Family, Araceae) tuber lectin antibody. The insecticidal activity of Arum maculatum tuber lectin (ATL) was tested against two economically important sucking pests, Lipaphis erysimi and Aphis craccivora, in an artificial diet. The LC(50) values for L. erysimi and A. craccivora were determined to be 21 microg/mL and 16 microg/mL, respectively. Addition of alpha-d-mannose in ATL-supplemented diet reduced the aphid mortality. Two major receptor proteins of ATL (approximately 40 kDa and approximately 35 kDa) were detected from the brush border membrane vesicle (BBMV) protein of L. erysimi and A. craccivora guts, respectively, using ligand-binding assay. Alpha-d-Mannose was found to be a deterrent to such binding of ATL to the BBMV receptors.

Pro-inflammatory effect of Arum maculatum lectin and role of resident cells.

Arum maculatum agglutinin (AMA) is a monocot lectin isolated from tubers of Arum maculatum L. (Araceae) which exhibits different specificity towards oligo-mannosidic-type and N-acetyllactosaminic-type glycans. We have investigated the effect of this lectin on the cells of the immune system. Models of neutrophil migration in vivo, neutrophil chemotaxis in vitro and macrophage cultures were used to study the lectin inflammatory activity. When administered into rat peritoneal cavities, AMA (80, 200 and 500 microg/mL/cavity) induced significant and dose-dependent neutrophil migration. This effect was inhibited by incubation with alpha-methyl-d-mannoside. A 83% depletion in the number of resident cells following peritoneal lavage did not reduce the AMA-induced neutrophil migration, as compared to sham animals (not washed). However, pre-treatment with 3% thioglycolate which increases the peritoneal macrophage population by 236%, enhanced the neutrophil migration induced by AMA (200 microg/mL/cavity) (119%, p < 0.05). Reduction of peritoneal mast cell population by chronic treatment of cavities with compound 48/80 did not modify AMA-induced neutrophil migration. The neutrophil chemotaxy assay in vitro shows that the lectin (300 microg/mL) induces neutrophil chemotaxy (368% p < 0.05) compared to RPMI. Finally, injection into peritoneal cavities of supernatants from macrophage cultures obtained after stimulation with AMA (300 microg/mL) enhanced neutrophil migration (110% p < 0.05). Summarizing, our data suggest that A. maculatum agglutinin presents pro-inflammatory activity, inducing neutrophil migration by two ways, one which is independent on resident cells and another one dependent on the presence of these cells.