Chemical composition, in vitro antitumor and pro-oxidant activities of Glandora rosmarinifolia (Boraginaceae) essential oil.

The biological properties of essential oils have been demonstrated in the treatment of several diseases and to enhance the bioavailability of other drugs. In natural habitats the essential oils compounds may play important roles in the protection of the plants as antibacterials, antivirals, antifungals, insecticides and also against herbivores by reducing their appetite for such plants or by repelling undesirable others. We analyzed by gas-chromatography mass spectrometry the chemical composition of the essential oil of aerial parts of Glandora rosmarinifolia (Ten.) D.C. Thomas obtained by hydrodistillation and verified some biological activities on a panel of hepatocellular carcinoma cell lines (HA22T/VGH, HepG2, Hep3B) and triple negative breast cancer cell lines (SUM 149, MDA-MB-231). In the essential oil we detected 35 compounds. The results of the biological assays indicate that essential oil of G. rosmarinifolia induces cell growth inhibition at concentration-dependent way in all cell line models. This oil does not seem to possess antioxidant activity, while the cytotoxicity of G. rosmarinifolia essential oil appeared to involve, at least in part, a pro-oxidant mechanism. Our results show for the first time the antitumoral and pro-oxidant activities of G. rosmarinifolia essential oil and suggest that it may represent a resource of pharmacologically active compounds.

Cytotoxic, proapoptotic and antioxidative potential of flavonoids isolated from propolis against colon (HCT-116) and breast (MDA-MB-231) cancer cell lines.

Isolated and structurally confirmed, eleven flavonoids from propolis were examined for their cytotoxicity toward human colon cancer and human breast cancer cells. Their effect on induction of apoptosis and their antioxidative activities were also evaluated. Six flavonoids induced cytotoxic effects in both cell lines. Luteolin had a marked effect on both cell lines, especially on HCT-116 cells (IC50 72h, 66.86µM). Also, luteolin was observed to have the highest apoptotic potential after 72h treatment of examined cell lines (27.13% and 37.09%, respectively). Myricetin exhibited selective inhibition of cell growth (IC50 114.75µM) and induced apoptosis in MDA-MB-231 cells only. Luteolin and galangin exhibited prooxidative properties 24h after the treatment in HCT-116 cells, while myricetin induced prooxidative effects in MDA-MB-231 cells. On the other hand, selected flavonoids exhibited antioxidative properties 72h after the treatment, decreasing superoxide anion radical and nitrite levels in both cell lines. Cytotoxic and proapoptotic effects on colon and breast cancer cell lines and the influence on their redox status make tested flavonoids good candidates for developing new anticancer drugs.

The relationship between the helicobacter pylori seropositivity with systemic and local oxidative status and hyperemesis gravidarum: a pilot study.

OBJECTIVES: The aim of study was to determine the helicobacter pylori (HP) seropositivity and oxidative parameters in serum and saliva of pregnant women with poor oral hygiene and hyperemesis gravidarum (HG). METHODS: A case-control study was conducted involving 50 pregnant women in their first trimester of pregnancy. Twenty-five subjects had a diagnosis of HG, and remaining 25 were healthy pregnant women who served as control subjects were included. The groups were adjusted for age, parity and gestational week. All patients were subjected to the measurement of total oxidant status (TOS) and total antioxidant status in serum and saliva. Also HP seropositivity was investigated. RESULTS: Serum TAS and TOS values were similar, although oxidative burden in saliva of women with HG were significantly higher than controls. HP seropositivity was found to be 24% in women with HG and 4% of controls. CONCLUSIONS: Our results suggest that significantly increased oxidative burden and slightly decreased antioxidative capacity of saliva may be involved in the pathogenesis of HG and this condition may be the result of HP infection which was found to be significantly more common in women with poor oral hygiene and HG.

Gold functionalised attapulgite for discrimination of hydrogen peroxide and oxidising ions.

Aiming to monitor hydrogen peroxide and oxidizing ions in aqueous circumstance, functionalized attapulgite [i.e. gold-modified attapulgite nanocomposites (Au/ATP NCs)] as peroxidase mimics were prepared by loading ß-cysteamine-capped gold nanoparticles onto attapulgite with the use of electrostatic interactions. As-prepared Au/ATP NCs were used for the detection of H2O2. The linear range and detection limit for H2O2 were determined by quantitative experiments emerging excellent stability and recyclability. The peroxidase-like activity of Au/ATP NCs could be expanded for the detection of some oxidative ions (Fe3+ and Ag+) was also been discovered. Based on the absorption spectrum and steady-state kinetics, the mechanism for peroxidase mimic reaction is investigated. This work represents the first example of multitarget detection for molecule and cations (H2O2, Fe3+ and Ag+) using Au/ATP NCs as peroxidase enzyme mimics and holds a promise for future biomedical and analytical applications.

Oxidant effects and toxicity of Croton campestris in Drosophila melanogaster.

CONTEXT: Croton campestris A.St.-Hil. (Euphorbiaceae) is a species native to Northeast Brazil used by traditional communities for the treatment of a variety of health problems. However, potential toxicological effects of this plant are unknown. OBJECTIVE: The potential toxicity of the hydroalcoholic extract of C. campestris leaves on Drosophila melanogaster insect model, additionally with phytochemical constitution and cellular mechanisms mediating the action of extract were analysed in this study. MATERIALS AND METHODS: Constituents of the extract were evaluated by HPLC. In vitro antioxidant potential of extract was analysed by DPPH, ABTS and FRAP. Flies injected culture medium mixed with extract (0.1-50 mg/mL) for 72 h. After, ROS production was evaluated by DCF-DA oxidation. Phosphorylation of MAPK signalling pathway was investigated by Western blotting method. Activity of antioxidant enzymes was analysed in homogenates. RESULTS: Major components of the extract include quercetin (38.11 ± 0.06 mg/g), caffeic acid (20.06 ± 0.17 mg/g) and kaempferol (15.45 ± 0.05 mg/g). Consumption of the extract impaired locomotor performance and induced fly death of flies (LC50 of 26.51 mg/mL). Augmented ROS formation and SOD, CAT and GST activity were observed from 0.1 mg/mL. JNK and p38 kinases phosphorylation was modulated and Paraquat-induced toxicity was augmented by extract. DISCUSSION AND CONCLUSION: Our data show important toxicological effects of C. campestris leading to increased mortality and impaired locomotor performance accompanied by induction of cell stress markers in flies. The study draws attention to the indiscriminate use of plant extracts.

Piper nigrum ethanolic extract rich in piperamides causes ROS overproduction, oxidative damage in DNA leading to cell cycle arrest and apoptosis in cancer cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Ayurvedic and Chinese traditional medicine and tribal people use herbal preparations containing Piper nigrum fruits for the treatment of many health disorders like inflammation, fever, asthma and cancer. In Brazil, traditional maroon culture associates the spice Piper nigrum to health recovery and inflammation attenuation. AIMS OF THE STUDY: The aim of the current work was to evaluate the relationship between reactive oxygen species (ROS) overproduction, DNA fragmentation, cell cycle arrest and apoptosis induced by Piper nigrum ethanolic extract and its antitumor activity. METHODS: The plant was macerated in ethanol. Extract constitution was assessed by TLC, UV-vis and ESI-IT-MS/MS spectrometry. The cytotoxicity, proliferation and intracellular ROS generation was evaluated in MCF-7 cells. DNA damage effects were evaluated through intercalation into CT-DNA, plasmid DNA cleavage and oxidative damage in CT-DNA. Tumor growth inhibition, survival time increase, apoptosis, cell cycle arrest and oxidative stress were assessed in Ehrlich ascites carcinoma-bearing mice. RESULTS: Extraction yielded 64mg/g (36% piperine and 4.2% piperyline). Treatments caused DNA damage and reduced cell viability (EC50=27.1±2.0 and 80.5±6.6µg/ml in MCF-7 and HT-29 cells, respectively), inhibiting cell proliferation by 57% and increased ROS generation in MCF-7 cells (65%). Ehrlich carcinoma was inhibited by the extract, which caused reduction of tumor growth (60%), elevated survival time (76%), cell cycle arrest and induced apoptosis. The treatment with extract increased Bax and p53 and inhibited Bcl-xL and cyclin A expression. It also induced an oxidative stress in vivo verified as enhanced lipid peroxidation and carbonyl proteins content and increased activities of glutathione reductase, superoxide dismutase and catalase. GSH concentration was decreased in tumor tissue from mice. CONCLUSION: The ethanolic extract has cytotoxic and antiproliferative effect on MCF-7 cells and antitumor effect in vivo probably due to ROS overproduction that induced oxidative stress affecting key proteins involved in cell cycle arrest at G1/S and triggering apoptosis. Finally, the overall data from this study are well in line with the traditional claims for the antitumor effect of Piper nigrum fruits.

Pomegranate Juice Polyphenols Induce Macrophage Death via Apoptosis as Opposed to Necrosis Induced by Free Radical Generation: A Central Role for Oxidative Stress.

At high concentrations, polyphenols induce cell death, and the polyphenols-rich pomegranate juice (PJ), known for its antioxidative/antiatherogenic properties, can possibly affect cell death, including macrophage death involved in atherogenesis. In the present study, apoptotic/necrotic macrophage death was analyzed in J774A.1 macrophages and in peritoneal macrophages isolated from atherosclerotic apoE-/- mice treated with PJ. The effects of PJ were compared with those of the free radical generator 2, 2'-azobis (2-amidinopropane) dihydrochloride (AAPH). Both PJ and AAPH significantly increased J774A.1 macrophage death; however, flow cytometric and microscopic analyses using annexin V/propidium iodide revealed that PJ increased the early apoptosis of the macrophage dose dependently (up to 2.5-fold, P < 0.01), whereas AAPH caused dose-dependent increases in late apoptosis/necrosis (up to 12-fold, P < 0.001). Unlike PJ, AAPH-induced macrophage death was associated with increased intracellular oxidative stress (up to 7-fold, P < 0.001) and with lipid stress demonstrated by triglyceride accumulation (up to 3-fold, P < 0.01) and greater chromatic vesicle response to culture medium (up to 5-fold, P < 0.001). Accordingly, recombinant paraoxonase 1, which hydrolyzes oxidized lipids, attenuated macrophage death induced by AAPH, but not by PJ. Similar apoptotic and oxidative effects were found in macrophages from apoE-/- mice treated with PJ or AAPH. As macrophage apoptotic/necrotic death has considerable impact on atherosclerosis progression, these findings may provide novel mechanisms for the antiatherogenicity of PJ.

Structural elucidation and biological studies of a novel exopolysaccaride from Klebsiella pneumoniae PB12.

An exopolysaccharide (KNPS) of an average molecular weight ∼1.8×10(5) Da was isolated from the culture medium of Klebsiella pneumoniae PB12. Structural characterization of KNPS was carried out using sugar and methylation analysis, Smith degradation and 1D/2D NMR experiments. Sugar analysis showed that the KNPS composed of arabinose, galactose, 3-O-methyl-galctose and glucose in a molar ratio of nearly 4:3:1:1. The proposed repeating unit of the KNPS has a backbone chain consisting of two (1→6)-galactopyranosyl residues, two (1→5)-arabinofuranosyl residues, one (1→6)-glucopyranosyl residue and one (1→3)-arabinopyranosyl residue, out of which one (1→6)-galactopyranosyl residue was branched at O-2 position with a (1→2)-linked-galactopyranosyl residue terminated with non reducing arabinofuranosyl residue and one (1→5)-arabinofuranosyl residue branched at O-3 position with non reducing end 3-O-Me-galactopyranosyl residue. KNPS was found non-toxic toward human lymphocyte up to the dosage of 100 µg/ml. KNPS enhanced malondialdehyde (MDA), reactive oxygen species (ROS), and have the potential to alter the ratio of oxidized glutathione (GSSG) and reduced glutathione (GSH) levels in the cellular system.

Pro-oxidant activity of indicaxanthin from Opuntia ficus indica modulates arachidonate metabolism and prostaglandin synthesis through lipid peroxide production in LPS-stimulated RAW 264.7 macrophages.

Macrophages come across active prostaglandin (PG) metabolism during inflammation, shunting early production of pro-inflammatory towards anti-inflammatory mediators terminating the process. This work for the first time provides evidence that a phytochemical may modulate the arachidonate (AA) metabolism in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages, promoting the ultimate formation of anti-inflammatory cyclopentenone 15deoxy-PGJ2. Added 1 h before LPS, indicaxanthin from Opuntia Ficus Indica prevented activation of nuclear factor-κB (NF-κB) and over-expression of PGE2 synthase-1 (mPGES-1), but up-regulated cyclo-oxygenase-2 (COX-2) and PGD2 synthase (H-PGDS), with final production of the anti-inflammatory cyclopentenone. The effects were positively related with concentration between 50 and 100 µM. Indicaxanthin did not have any effect in the absence of LPS. A kinetic study investigating the redox status of LPS-stimulated macrophages between 0.5 and 12 h, either in the absence or in the presence of 50-100 µM indicaxanthin, revealed a differential control of ROS production, with early (0.5-3 h) modest inhibition, followed by a progressive (3-12 h) concentration-dependent enhancement over the level induced by LPS alone. In addition, indicaxanthin caused early (0.5-3 h) concentration-dependent elevation of conjugated diene lipid hydroperoxides, and production of hydroxynonenal-protein adducts, over the amount induced by LPS. In LPS-stimulated macrophages indicaxanthin did not affect PG metabolism when co-incubated with either an inhibitor of NADPH oxidase or vitamin E. It is concluded that LPS-induced pro-oxidant activity of indicaxanthin at the membrane level allows formation of signaling intermediates whose accumulation modulates PG biosynthetic pathway in inflamed macrophages.

Antimicrobial properties of an oxidizer produced by Burkholderia cenocepacia P525.

A compound with both oxidizing properties and antibiotic properties was extracted and purified from broth cultures of Burkholderia cenocepacia strain P525. A four step purification procedure was used to increase its specific activity ~400-fold and to yield a HPLC-UV chromatogram containing a single major peak. Size exclusion chromatography suggests a molecular mass of ~1,150 and UV spectroscopy suggests the presence of a polyene structure consisting of as many as six conjugated double bonds. Biological studies indicate that the compound is bacteriostatic. Enterobacter soli and E. aerogenes cells incubated with the compound exhibit a longer lag phase of growth. The bacteriostatic activity is greater at pH 3 than at pH 5. Bacteria such as B. cenocepacia strain P525 may have value in the agricultural industry as biocontrol agents.

Antimicrobial activity of Amazon Astrocaryum aculeatum extracts and its association to oxidative metabolism.

Several compounds present in fruits as polyphenols are able to kill or inhibit the growth of microorganisms. These proprieties are relevant mainly in tropical areas, as Amazonian region where infectious are highly prevalent. Therefore, this study investigated the antimicrobial activity of tucumã Amazonian fruit against 37 microorganisms. The potential role of oxidative metabolism imbalance was also studied as causal mechanism of antimicrobial activity. The results showed antibacterial effect of pulp and peel tucumã hydro-alcoholic extracts on three Gram-positive bacteria (Enterococcus faecalis, Bacillus cereus, Listeria monocytogenes) and antifungal effect against Candida albicans. The antimicrobial contribution of main chemical compounds (quercetin, rutin, ß-carotene and gallic, caffeic and chlorogenic acids) found in tucumã extracts was also investigated showing an inhibitory effect depending of the organism mainly by quercetin in bacteria and rutin in C. albicans. Analysis of kinetic of DNA releasing in extracellular medium by fluorescence using DNA Pico Green assay(®) and reactive oxygen species production (ROS) showed potential oxidative imbalance contribution on tucumã inhibitory effect. In B. cereus and C. albicans this effect was clear since after 24h the ROS levels were higher when compared to negative control group. In conclusion, tucumã extracts present antimicrobial activity to four microorganisms that have large problems of drug resistance, and the possible mechanism of action of this Amazon fruit is related to REDOX imbalance.

Induction of apoptosis in murine leukemia by diarylheptanoids from Curcuma comosa Roxb.

Diarylheptanoids, isolated from the rhizome of Curcuma comosa Roxb., have several biological activities including anti-oxidant and anti-inflammation. The present study investigated the effect of five diarylheptanoids isolated from C. comosa rhizome on the proliferation of murine P388 leukemic cells. Compound-092, (3S)-1-(3,4-dihydroxyphenyl)-7-phenyl-(6E)-6-hepten-3-ol, bearing a catechol moiety, was the most potent diarylheptanoid (IC(50) of 4 µM) in inhibiting P388 leukemic cell viability by causing DNA breakage and inducing apoptosis. Apoptotic cell death was characterized by the presence of chromatin condensation, formation of apoptotic bodies, DNA fragmentation, and externalization of plasma membrane phosphatidylserine. This compound increased caspase-3 activity about fivefold above the untreated control, decreased the intracellular reduced glutathione level, and impaired mitochondrial transmembrane potential. In the presence of Cu(II) ion, the compound exhibited a pro-oxidant activity causing DNA strand breakage and enhancing the anti-proliferative activity. The results provide evidence for the pro-oxidant activity of the diarylheptanoid bearing a catechol moiety in the induction of apoptosis in murine P388 leukemia.

Phloroglucinols with prooxidant activity from Garcinia subelliptica.

A new phloroglucinol, garcinielliptone HF ( 1), possessing an unprecedented skeleton, and the tautomeric pair of garcinielliptone FC ( 2/ 2a) were isolated from the heartwood and pericarp of Garcinia subelliptica, respectively. Their structures, including relative configurations, were elucidated by means of spectroscopic methods. The ability of compounds 1 and 2/ 2a to induce DNA-cleavage activity was examined using supercoiled plasmid pBR322 DNA. In the presence of Cu(II), compounds 1 and 2/ 2a caused significant breakage of pBR322 DNA. The involvement of H2O2 and O2 (*-), and H2O2, O2 (*-), and OH (*) in 1- and 2/ 2a-mediated scission, respectively, was established by inhibition or no protection of DNA breakage by various oxygen radical scavengers. Thus, in the presence Cu(II), 1 and 2/ 2a may show a prooxidant effect on DNA and induce cell death.

Estatus pro-oxidante en niños del Valle de Mezquital, Hidalgo (México) / No disponible

El objetivo ha sido demostrar que el retraso en el crecimiento de la niñez está asociado a un estatus pro-oxidante. Se estudiaron niños indígenas con edades entre 9 y 11 años, residentes en el Valle del Mezquita!, Hidalgo, México. Las mediciones clínicas y antropométricas fueron realizadas utilizando técnicas estándares. En muestras de sangre se determinaron contenidos de vitaminas E y A, contenidos de metales (Cu, Mn, Fe, Zn y Se) y se determinaron los contenidos de sustancias reactivas al ácido tio-barbitúrico (SRATE1). Los niños del grupo experimental mostraron concentración disminuida de vitamina E (643.8 ± 11.3 y 529.3 .÷, 89 pgidL) y de vitamina A (28.9 ± 3.5 y 26.4 ± 2.5 wgidIL), con respecto a los niños del grupo control. La relación vitamina EiSF?A7-8 resultó más alta en el grupo control (1458.8 I- 736) comparado con el grupo experimental (1003 ± 535.3); las niñas presentaron valores más altos de este cociente que los niños. No se ha podido asociar la prosenda de un estatus pro-oxidante, ya que no se encontraron variaciones significativas. De los resultados SRATB y concentraciones de hierro resultaron más elevados en el grupo experimental que en el grupo control, pero no estadísticamente diferentes. Se propone considerar una suplementación de vitaminas antioxidantes para prevenir dicho estado y la capacidad potencial para inducir daño en células y tejidos finos. Se recomienda realizar otros estudios que permitan demostrar la capacidad antioxidante medida en suero. con determinaciones de capacidades antioxidantes de enzimas en estos niños. Se incrementa la posibilidad de que el estatus oxidante esté presente. dada la capacidad antioxidante disminuida por déficit de vitamina E y A y disminución de la relación SRATB/vitamina relacionado con el pobre desarrollo y la baja estatura (AU)

To test the hypothesis that growth retardation in early childhood might be associated to an oxidant stress status, 9-11 aged, living in Mezquita/ Valley (Hidalgo, México) were studied. Clinical and anth-ropometric measures were made using standard techniques. Blood samples were obtained by venous puncture. Vitamins E and C, copper, iron manganese, zinc: selenium and TBARS (thiobabituic acid reactive substances) were measured. Stunted children showed a decreased vitamin E (643.8 ± 11.3 to 529.3 -± 89 ug/dL) and A (28.9 ± a5 to 26.4 ± 2.5 pg/d/L) serum concentration than control group. The ratio Vitamin E/ TBARS were loo higher in control group (1458.8 ± 736) when was compared with stunted group (1003 ± 535.3). The girls showed higher values of this ratio than boys, in both control and stunted groups. From the data presented herein is not possible to associate the presence of oxidant stress status due the absence of statistical significances. In both, TBARS and iron the concentrations measure we-re higher in stunted group, but not statistically different compared with control group. Due the potential pre-oxidative stress status of student children, we propose to take in consideration the sup-plementation of antioxidant vitamins in those children in order to prevent the oxidative stress status, and its potential capacity for induce damage of cells and tissues and dysfunctional consequences. On the other hand, it is necessary to carry out other studies conducing to measured antioxidant serum capacity including antioxidant enzymes determinations in student children. Finally our data and observations raises the possibility that oxidative status due to decreased antioxidant capacity (vitamin E, A, TBRS/vitamin E) presented in stunted children, might be related with the low development of height in this children (AU)

Targeted natural product isolation guided by HPLC-SPE-NMR: constituents of Hubertia species.

The hyphenated technique, high-performance liquid chromatography-solid-phase extraction-nuclear magnetic resonance spectroscopy (HPLC-SPE-NMR), has been applied for rapid identification of novel natural products in crude extracts of Hubertia ambavilla and Hubertia tomentosa. The technique allowed full or partial identification of all major extract constituents and demonstrated the presence of unusual quinic acid derivatives containing the (1-hydroxy-4-oxocyclohexa-2,5-dienyl)acetyl residue that exhibit strongly coupled ABXY patterns, the parameters of which were obtained by spin simulations. Using homo- and heteronuclear 2D NMR data acquired in the HPLC-SPE-NMR mode, complete structure determination of three new natural products, i.e., 3,5-di-O-caffeoyl-4-O-[(1-hydroxy-4-oxocyclohexa-2,5-dienyl)acetyl]quinic acid (1), its 2-hydroxy derivative (2), and 3,5-di-O-caffeoyl-4-O-[(4-hydroxyphenyl)acetyl]quinic acid (3), was performed. Finally, targeted isolation of 1 was achieved by SPE fractionation and preparative HPLC, followed by evaluation of its antioxidant and antimicrobial activity. In contrast to chlorogenic acid and 3,5-di-O-caffeoylquinic acid, which act as antioxidants, compound 1 proved at the same conditions to possess prooxidant activity in an assay evaluating the oxidation of human low-density lipoprotein induced by Cu(2+).

Successful removal of p-quinone with chitosan in an aqueous phase in relation to degree of deacetylation.

Phenol oxidant is successfully removed by using chitosan particles in the aqueous phase. Removal of p-quinone by chitosan from crab shells was investigated kinetically from molecular weight (MW) of chitosan, deacetylation degree (DD) and reaction temperature. The rate constant assuming first-ordered reaction on removal of p-quinone in aqueous phase primarily depended on the MW of chitosan, not on the DD. Quantities of chitosan exceeding 5 x 10(5) MW are able to obtain a sufficiently high rate constant (10(-3) s(-1)). At higher temperatures, higher rate constants were obtained in the entire experimental MW and DD. The activation energy obtained was 43.8 kJ x mol(-1).

Anti- and pro-oxidant activity of water soluble compounds in Cichorium intybus var. silvestre (Treviso red chicory).

The anti- and pro-oxidant activity of water soluble components in Cichorium intybus var. silvestre was investigated. This vegetable is domestically known as Treviso red chicory recalling its cultivation in the area of Treviso (Italy). The vegetable juices, obtained by centrifugation of the vegetable and treated at 2 and 102 degrees C, were assessed for their antioxidant activity (AA) using the micellar model system linoleic acid-beta-carotene. The obtained juice at 2 degrees C possessed either anti- or pro-oxidant activity. The boiled juice showed only strong AA, proving that the vegetable pro-oxidant components were thermally instable. Juice components were fractionated by sequential dialysis (1000-300000 Da membrane cut-off), SPE, GFC, and RP-HPLC techniques showing the presence of several highly antioxidant components with different molecular weight (MW), and polar features in Treviso red chicory. The pro-oxidant fraction capable of masking the presence of the antioxidant components in the vegetable juice is retained by a MW>300000 Da dialysis membrane.

Isolation, identification, and characterization of compounds from acer rubrum capable of oxidizing equine erythrocytes.

OBJECTIVE: To identify compounds in Acer rubrum that cause hemolysis or oxidation of equine erythrocytes and determine whether these toxins are found in other Acer spp. SAMPLE POPULATION: Equine erythrocytes. PROCEDURE: Washed erythrocytes were incubated with extracts and fractions of Acer spp that were separated by thin layer chromatography. Methemoglobin and hemolysis were measured spectrophotometrically. Compounds within Acer spp fractions associated with cell oxidation or hemolysis were identified by gas chromatography-mass spectrometry. RESULTS: Erythrocytes incubated separately with either A. rubrum, A. saccharum, or A. saccharinum extracts had increased methemoglobin formation, compared with extract-free control samples. Two Acer spp fractions had toxic effects on erythrocytes in vitro. A major component of the Acer fraction that caused a significant amount of methemoglobin formation was identified as gallic acid. An amount of gallic acid equivalent to that found in A. rubrum extract significantly increased methemoglobin, compared with extract-free control erythrocytes, but caused less methemoglobin formation than A. rubrum extracts did. A potential co-oxidant, 2,3-dihydro-3,5-dihydroxy-6-methoxy-4H-pyran-4-one, was found in the A. rubrum extract and may have been responsible for increasing methemoglobin formation. A second A. rubrum fraction caused methemoglobin formation and significant hemolysis. A. saccharum and A. saccharinum extracts caused hemolysis but less than the A. rubrum extracts did. CONCLUSION AND CLINICAL RELEVANCE: Oxidants in A. rubrum are also found in A. saccharum and A. saccharinum, and the ingestion of A. saccharum and A. saccharinum poses a potential threat to horses.