Production of fructooligosaccharides by Bacillus subtilis natto CCT7712 and their antiproliferative potential.

AIMS: Fructooligosaccharides (FOSs) known for their health properties and ß-(2→6)-levan-type FOSs have shown prebiotic and immunomodulatory activities that overcome those of commercial ß-(2→1)-FOSs, but costs do not favour their use. Moreover, FOSs can reach the bloodstream through the diet, and little is known about their direct effect on cells. The aim of this work was to produce high-content FOSs by Bacillus subtilis natto CCT7712 in a bioreactor using commercial sucrose and to evaluate their antiproliferative effects in OVCAR-3 cells. METHODS AND RESULTS: FOS production reached 173·60 g l-1 , 0·2 vvm aeration and uncontrolled pH. Levan-type FOSs, composed of ß-(2 â†’ 6) links and mainly GF3 (6-nystose), were identified using RMN spectroscopy, FT-IR and ESI-MS. FOSs decreased the viability and proliferation of OVCAR-3 cells, and the effects were associated with an increased pro-inflammatory response by the induction of IL-8 and TNF-α, and the repression of ER-ß genes. The metabolic profiles showed disruption of cellular homeostasis that can be associated with a decrease in proliferation. CONCLUSIONS: The high production of levan-type FOSs from B. subtilis natto CCT7712 in a bioreactor was achieved, and they showed antiproliferative potential in OVCAR-3 cells. SIGNIFICANCE AND IMPACT OF THE STUDY: FOS could be a good target for future therapeutic studies and commercial use.

6-Dimethylaminopurine and cyclohexamide are mutagenic and alter reproductive performance and intrauterine development in vivo.

The compounds 6-dimethylaminopurine (6-DMAP) and cyclohexamide (CHX) are currently used to stimulate the development of embryos produced by nuclear transfer in the production of cloned mammals with a great deal success. This study investigated the effects of 6-DMAP and CHX in vivo using biological assays to evaluate reproductive performance in females, teratogenesis, and mutagenesis. The results of this study demonstrated that the activating agents of oocyte cytoplasm, 6-DMAP and CHX, altered the reproductive performance of the experimental animals, as well as increased the rate malformations. In addition to these adverse effects, the administration of these compounds in pregnant females resulted in genotoxic and mutagenic toxicity, as determined by comet and micronucleus assays carried out in peripheral blood samples, respectively. Based on these findings and that alterations in DNA are important, morpho-functional teratogenesis and diminished embryonic viability, suggesting that 6-DMAP and CHX, which are utilized during the cloning of mammals, are responsible for the fact that embryos produced by nuclear transfer show low viability after implantation in utero or after birth because of congenital malformations and/or alterations in their DNA.

Cytotoxicity and mutagenicity of fluoxetine hydrochloride (Prozac), with or without vitamins A and C, in plant and animal model systems.

Fluoxetine, commonly known as Prozac, is the first representative of the so-called new generation of antidepressants that promise efficacy, with few side effects, against deep depression, nervous bulimia, and anxiety. As there is a growing number of people suffering from anxiety and depression; consequently, the use of fluoxetine is also increasing. Verifying absence of drug effects such as cytotoxicity or mutagenicity is of great importance. Certain vitamins, such as vitamin A (retinol, retinoids) and vitamin C (ascorbic acid) protect and are extremely active against mutagens. We evaluated the cytotoxic and mutagenic activity of fluoxetine, with and without concomitant administration of vitamin A or C, in Allium cepa meristem cells and Wistar rat bone marrow cells. The A. cepa meristem cells showed fluoxetine cytotoxicity; concomitant treatment with vitamin A or C proved non-protective. Treatment of Wistar rats with fluoxetine intraperitoneally or via gavage did not affect cell division or cause clastogenic effects. Vitamin A and C did not affect the cytotoxicity or mutagenicity of fluoxetine in the rat cells.

Evaluation of the antimutagenic activity and mode of action of the fructooligosaccharide inulin in the meristematic cells of Allium cepa culture.

This study evaluated the mutagenicity and antimutagenicity of inulin in a chromosomal aberration assay in cultures of the meristematic cells of Allium cepa. The treatments evaluated were as follows: negative control--seed germination in distilled water; positive control--aqueous solution of methyl methanesulfonate (10 µg/mL MMS); mutagenicity--aqueous solutions of inulin (0.015, 0.15, and 1.50 µg/mL); and antimutagenicity--associations between MMS and the different inulin concentrations. The antimutagenicity protocols established were pre-treatment, simultaneous simple, simultaneous with pre-incubation, and post-treatment. The damage reduction percentage (DR%) was 43.56, 27.77, and 55.92% for the pre-treatment; -31.11, 18.51, and 7.03% for the simultaneous simple; 30.43, 19.12, and 21.11% for the simultaneous with pre-incubation; and 64.07, 42.96, and 53.70% for the post-treatment. The results indicated that the most effective treatment for inhibiting damages caused by MMS was the post-treatment, which was followed by the pre-treatment, suggesting activity by bioantimutagenesis and desmutagenesis. The Allium cepa assay was demonstrated to be a good screening test for this type of activity because it is easy to perform, has a low cost, and shows DR% that is comparable to that reported studies that evaluated the prevention of DNA damage in mammals by inulin.

Mutagenic and antimutagenic effects of aqueous extract of rosemary (Rosmarinus officinalis L.) on meristematic cells of Allium cepa.

Polyphenolic compounds present in rosemary were found to have antioxidant properties, anticarcinogenic activity, and to increase the detoxification of pro-carcinogens. The aim of the study was to determine the effect the aqueous extract of rosemary (AER) on mutagenicity induced by methylmethane sulfonate in meristematic cells of Allium cepa, as well as to describe its mode of action. Anti-mutagenicity experiments were carried out with 3 different concentrations of AER, which alone showed no mutagenic effects. In antimutagenicity experiments, AER showed chemopreventive activity in cultured meristematic cells of A. cepa against exposure to methylmethane sulfonate. Additionally, post-treatment and simultaneous treatment using pre-incubation protocols were the most effective. Evaluation of different protocols and the percent reduction in DNA indicated bioantimutagenic as well desmutagenic modes of action for AER. AER may be chemopreventive and antimutagenic.

Antigenotoxic and antimutagenic effects of glutamine supplementation on mice treated with cisplatin.

We evaluated the effects of glutamine on clastogenic and genotoxic damage prevention caused by the administration of cisplatin. Forty Swiss mice were divided into 8 experimental groups: G1 and G2, which were control groups; G3, G4, and G5, which were administered [2 doses of glutamine (orally)] separated by a 24-h period (150, 300, and 600 mg/kg, respectively), and a dose of phosphate-buffered saline by intraperitoneal injection; G6, G7, and G8, which were treated in the same manner as the previous groups, but received cisplatin rather than phosphate-buffered saline. The antimutagenicity groups showed damage reduction percentages of 79.05, 80.00, and 94.27% at the time point T1, 53.18, 67.05, and 64.74 at time point T2 for the 150, 300, and 600 mg/kg doses of glutamine, respectively. Antigenotoxic activity was evident for all 3 doses with damage reduction percentages of 115.05, 119.06, and 114.38 for the doses of glutamine of 150, 300, and 600 mg/ kg, respectively. These results suggest that further studies are needed to confirm the clastogenic activity of glutamine. However, our results may lead to rational strategies for supplementation of this antioxidant as an adjuvant in cancer treatment or for preventing genomic lesions.

Evaluation of the mutagenicity and antimutagenicity of soy phytoestrogens using micronucleus and comet assays of the peripheral blood of mice.

Studies show that soy imparts many favorable properties in the human body, including the prevention of chronic diseases such as osteoporosis, heart disease, cancer, and diabetes. Soy is rich in isoflavones, and it is a candidate for the chemoprevention of diseases owing to its low toxicity. In this study, a soy phytoestrogen (with high levels of the isoflavones genistin and daidzein) was tested in mice to investigate its mutagenicity and genotoxicity using micronucleus and comet assays of mouse peripheral blood. Phytoestrogen (0.083, 0.83 and 8.3 mg/kg body weight) was evaluated with and without the chemotherapeutic agent cyclophosphamide. For the micronucleus assay, blood was collected before treatment and after 24 and 48 h. For the comet assay, blood was collected only after 24 h. Phytoestrogen was not mutagenic and reduced cyclophosphamide-induced DNA damage. The results from the comet assay revealed a reduction of DNA damage; however, phytoestrogen did induce genotoxic damage during the 24-h treatment. This genotoxic damage could have been repaired and was therefore not identified in the micronucleus assay, which detects mutations. The results suggested that the reduction of DNA damage observed in associated treatments could also reduce the side effects of chemotherapy. Moreover, they suggested that phytoestrogen might be a candidate of interest for the chemoprevention of cancer because it protects against DNA damage.

Antimutagenic and anticarcinogenic effects of wheat bran in vivo.

Previous studies in rodents treated with the pro-carcinogen 1,2-dimethylhydrazine suggested that the consumption of wheat bran protected against DNA damage in the colon and rectum. Based on this information, we evaluated wheat bran as a functional food in the prevention and treatment of colon cancer. We used the aberrant crypt focus assay to evaluate the anticarcinogenic potential of wheat bran (Triticum aestivum variety CD-104), the comet assay to evaluate its antigenotoxicity potential, and the micronucleus assay to evaluate its antimutagenic potential. The wheat bran gave good antimutagenic and anticarcinogenic responses; the DNA damage decreased from 90.30 to 26.37% and from 63.35 to 28.73%, respectively. However, the wheat bran did not significantly reduce genotoxicity. Further tests will be necessary, including tests in human beings, before this functional food can be recommended as an adjunct in the prevention and treatment of colon cancer.

Genotoxic evaluation of the antimalarial drugs artemisinin and artesunate in human HepG2 cells and effects on CASP3 and SOD1 gene expressions.

The malaria treatment recommended by the World Health Organization involves medicines derived from artemisinin, an active compound extracted from the plant Artemisia annua, and some of its derivatives, such as artesunate. Considering the lack of data regarding the genotoxic effects of these compounds in human cells, the objective of this study was to evaluate the cytotoxicity and genotoxicity, and expressions of the CASP3 and SOD1 genes in a cultured human hepatocellular liver carcinoma cell line (HepG2 cells) treated with artemisinin and artesunate. We tested concentrations of 2.5, 5, 7.5, 10, and 20 µg/mL of both substances with a resazurin cytotoxicity assay, and the concentrations used in the genotoxicity experiments (2.5, 5, and 10 µg/mL) and gene expression analysis (5 µg/mL) were determined. The results of the comet assay in cells treated with artemisinin and artesunate showed a significant dose-dependent increase (P < 0.001) in the number of cells with DNA damage at all concentrations tested. However, the gene expression analysis revealed no significant change in expression of CASP3 or SOD1. Our data showed that although artemisinin and artesunate exhibited genotoxic effects in cultured HepG2 cells, they did not significantly alter expression of the CASP3 and SOD1 genes at the doses tested.

Evaluation of the antimutagenic and anticarcinogenic effects of inulin in vivo.

The incidence of colorectal cancer is growing worldwide. The characterization of compounds present in the human diet that can prevent the occurrence of colorectal tumors is vital. The oligosaccharide inulin is such a compound. The aim of this study was to evaluate the antigenotoxic, antimutagenic and anticarcinogenic effects of inulin in vivo. Our study is based on 3 assays that are widely used to evaluate chemoprevention (comet assay, micronucleus assay, and aberrant crypt focus assay) and tests 4 protocols of treatment with inulin (pre-treatment, simultaneous, post-treatment, and pre + continuous). Experiments were carried out in Swiss male mice of reproductive age. In order to induce DNA damage, we used the pro-carcinogenic agent 1,2-dimethylhydrazine. Inulin was administered orally at a concentration of 50 mg/kg body weight following the protocols mentioned above. Inulin was not administered to the control groups. Our data from the micronucleus assay reveal antimutagenic effects of inulin in all protocols. The percentage of inulin-induced damage reduction ranged from 47.25 to 141.75% across protocols. These data suggest that inulin could act through desmutagenic and bio-antimutagenic mechanisms. The anticarcinogenic activity (aberrant crypt focus assay) of inulin was observed in all protocols and the percentages of damage reduction ranged from 55.78 to 87.56% across protocols. Further tests, including human trials, will be necessary before this functional food can be proven to be effective in the prevention and treatment of colon cancer.

Pre-treatment with glutamine reduces genetic damage due to cancer treatment with cisplatin.

Cisplatin is an effective antineoplastic drug. However, it provokes considerable collateral effects, including genotoxic and clastogenic activity. It has been reported that a diet rich in glutamine can help inhibit such collateral effects. We evaluated this activity in 40 Swiss mice, distributed into eight experimental groups: G1 - Control group (PBS 0.1 mL/10 g body weight); G2 - cisplatin group (cisplatin 6 mg/kg intraperitoneally); G3, G4, G5 - glutamine groups (glutamine at 150, 300, and 600 mg/kg, respectively; orally); G6, G7, G8 - Pre-treatment groups (glutamine at 150, 300, and 600 mg/kg, respectively; orally and cisplatin 6 mg/kg intraperitonially). For the micronucleus assay, samples of blood were collected (before the first use of the drugs at T0, then 24 (T1) and 48 (T2) hours after the first administration). For the comet assay, blood samples were collected only at T2. The damage reduction percentages for the micronucleus assay were 90.0, 47.3, and 37.3% at T1 and 46.0, 38.6, and 34.7% at T2, for G6, G7, and G8 groups, respectively. For the comet assay, the damage reduction percentages were 113.0, 117.4, and 115.0% for G6, G7, and G8, respectively. We conclude that glutamine is able to prevent genotoxic and clastogenic damages caused by cisplatin.

In vitro genotoxicity assessment of caffeic, cinnamic and ferulic acids.

Phenols are a large and diverse class of compounds, many of which occur naturally in a variety of food plants; they exhibit a wide range of biological effects, including antibacterial, anti-inflammatory, antiallergic, hepatoprotective, antithrombotic, antiviral, anticarcinogenic, and vasodilatory actions. We examined the genotoxic and clastogenic potential of three phenolic compounds: caffeic, cinnamic and ferulic acids, using the comet and micronucleus assays in vitro. Drug-metabolizing rat hepatoma tissue cells (HTCs) were used. Three different concentrations (50, 500 and 1500 µM) of these phenolic acids were tested on the HTCs for 24 h. The caffeic, cinnamic and ferulic acids were not genotoxic by the comet assay (P > 0.05). However, the micronucleus test showed an increase in the frequency of micronucleated cells for the three compounds, indicating that these substances have clastogenic effects in HTC.

Cyclosporin A treatment and decreased fungal load/antigenemia in experimental murine paracoccidioidomycosis.

Paracoccidioidomycosis (PCM) is a systemic mycosis caused by the fungus Paracoccidioides brasiliensis (Pb). The cyclosporin A (CsA) is an immunosuppressant drug that inhibits calcineurin and has been described as a potential antifungal drug. The present study investigated the effect of CsA on the immune response, fungal load/antigenemia in experimental murine PCM. It was used four groups of BALB/c mice: (a) infected with 1 x 105 Pb18 yeast cells (Pb), (b) infected and treated with CsA every other day 10 mg/kg of CsA (s.c.) during 30 days (Pb/CsA), (c) treated with CsA (CsA) and (d) no infected/treated (PBS). The immune response was evaluated by lymphocyte proliferation, DTH assays to exoAgs, ELISA for IgG anti-gp43 (specific immune responses) and cytokine serum levels (IFN-γ, TNF-α, IL-4 and IL-10). Fungal load was determined by lung colony-forming units (CFU) counts, lung and liver histopathology analysis and antigenemia determined by inhibition-ELISA. As expected, CsA was able to inhibit the specific cellular and humoral immune response (P < 0.05), with decrease in serum IFN-γ, TNF-α and IL-4 levels (P < 0.05). Cyclosporin A treatment also resulted in significantly decreased lung Pb CFU (P < 0.05) as well as a lower number of yeasts in the lung and liver (P < 0.05) by histopathology. In concordance, the decreased antigenemia was observed in Pb/CsA group (P < 0.05). In conclusion, even with immunosuppressive action, treatment with CsA results in decreased lung fungal load/antigenemia in experimental PCM in BALB/c mice. Further study is required to determine whether this represents less severe disease or protection by CsA.

Chlorination treatment of aqueous samples reduces, but does not eliminate, the mutagenic effect of the azo dyes Disperse Red 1, Disperse Red 13 and Disperse Orange 1.

The treatment of textile effluents by the conventional method based on activated sludge followed by a chlorination step is not usually an effective method to remove azo dyes, and can generate products more mutagenic than the untreated dyes. The present work evaluated the efficiency of conventional chlorination to remove the genotoxicity/mutagenicity of the azo dyes Disperse Red 1, Disperse Orange 1, and Disperse Red 13 from aqueous solutions. The comet and micronucleus assays with HepG2 cells and the Salmonella mutagenicity assay were used. The degradation of the dye molecules after the same treatment was also evaluated, using ultraviolet and visible absorption spectrum measurements (UV-vis), high performance liquid chromatography coupled to a diode-array detector (HPLC-DAD), and total organic carbon removal (TOC) analysis. The comet assay showed that the three dyes studied induced damage in the DNA of the HepG2 cells in a dose-dependent manner. After chlorination, these dyes remained genotoxic, although with a lower damage index (DI). The micronucleus test showed that the mutagenic activity of the dyes investigated was completely removed by chlorination, under the conditions tested. The Salmonella assay showed that chlorination reduced the mutagenicity of all three dyes in strain YG1041, but increased the mutagenicity of Disperse Red 1 and Disperse Orange 1 in strain TA98. With respect to chemical analysis, all the solutions showed rapid discoloration and a reduction in the absorbance bands characteristic of the chromophore group of each dye. However, the TOC was not completely removed, showing that chlorination of these dyes is not efficient in mineralizing them. It was concluded that conventional chlorination should be used with caution for the treatment of aqueous samples contaminated with azo dyes.

Apoptotic and cell cycle response to homoharringtonine and harringtonine in wild and mutant p53 hepatocarcinoma cells.

This study aimed to evaluate the modes of action of harringtonine (HT) and homoharringtonine (HHT) alkaloids in cell with wild (HepG2/C3A) and mutant p53 (HuH-7.5). We performed assays for cytotoxicity, genotoxicity, induction of apoptosis, cell cycle phase, and membrane integrity. Obtained data were compared with the relative expression of mRNA of genes related to proliferation, apoptosis, cell cycle control, metabolism of xenobiotics, and reticulum endoplasmic stress. The relative expression of the genes showed an increase in apoptosis-inducing mRNAs, such as TNF and BBC3, as well as a reduction in BCL2 and BAK. The mRNAs of CYP2E1 and CYP2C19 xenobiotic metabolism genes increased in both lineages, while CYP3A4 increased only in the HuH-7.5 lineage. The mRNA expression of endoplasmic reticulum (ER) stress genes (ERN1 and EIF2AK3) was shown to increase in HHT and HT treatments. A similar increase was recorded in the mRNA expression of the TRAF2 gene. The changes observed in this study support the hypothesis that ER stress was more strongly associated with TNF induction, causing cell death by apoptosis in p53 mutant cells. This result with wild and mutant p53 cells may have clinical implications in the use of these compounds.

Wood versus fossil fuel as a source of excess carbon dioxide in the atmosphere: a preliminary report.

If the amounts of wood consumed in deforestation to increase agricultural land and as firewood in underindustrialized countries are added to the amount consumed by the money economies as forest products, the estimates of the net amount of wood removed from the biosphere in this century should be revised upward. The per capita ratio of the weight of carbon from net wood burned to the weight of carbon from fossil fuel burned in this century has been at least 0.1 and may have approached 1.0.

Effects of chlorophyllin on replication of poliovirus and bovine herpesvirus in vitro.

AIMS: Chlorophyllin (CHLN), a synthetic derivative of chlorophyll, was assayed in the replication of poliovirus (PV-1) and bovine herpesvirus (BoHV-1) in HEp-2 cell cultures. METHODS AND RESULTS: Virucidal activity of CHLN was evaluated and the time-of-addition assay was performed as follows: before the infection (-1 and -2 h), at the time of the infection (0 h) and after the infection (1 and 2 h). Plaque reduction assay (PRA) showed that CHLN inhibited BoHV-1 and PV-1 infection and the 50% inhibitory concentrations (IC(50)) against BoHV-1 and PV-1 infection were 8.6 and 19.8 microg ml(-1), respectively. The time-of-addition study demonstrated that the CHLN was effective inhibiting viral replication in 51% and 66.5% for PV-1 and BoHV-1, respectively, at the highest concentration of 20.0 microg ml(-1), when added during the infection. The directed effect of CHLN on viral strains demonstrated an inhibition of 62% and 66.4% for PV-1 and BoHV-1, respectively, by PRA. CONCLUSIONS: These results demonstrated that CHLN could be used as an antiviral suggesting directed activity on virus particles and on virus-receptor sites to BoHV. For poliovirus, CHLN also demonstrated virucide activity, moreover, showed to inhibit early steps of the replication cycle. SIGNIFICANCE AND IMPACT OF THE STUDY: CHLN demonstrated promising selectivity index for both virus strains; therefore, it can be used for the development of an antiviral agent.

Genotoxic, mutagenic and cytotoxic effects of the commercial dye CI Disperse Blue 291 in the human hepatic cell line HepG2.

Textile dyes are discarded into the aquatic ecosystem via industrial effluents and potentially expose humans and local biota to adverse effects. The commercial dye CI Disperse Blue 291 which contains the aminoazobenzene 2-[(2-bromo-4,6-dinitrophenyl)azo]-5(diethylamino)-4-methoxyacetanilide (CAS registry no. 56548-64-2), was tested for genotoxicity and cytotoxicity in the human hepatoma cell line HepG2, using the comet assay, micronucleus (MN) test and a cell viability test. Five different concentrations of the test compound were examined: 200 microg/ml, 400 microg/ml, 600 microg/ml, 800 microg/ml and 1000 microg/ml. An increase in comet tail length and in the frequency of MN was detected with exposure of cells to concentrations of the commercial dye from 400 microg/ml. Furthermore, the dye was found to decrease cell viability. The results of this study demonstrate for the first time the genotoxic and mutagenic effects of the dye CI Disperse Blue 291 in mammalian cells, thus stressing the need to develop non-mutagenic dyes and to invest in improving the treatment of effluents. These measures will help to prevent harmful effects that these compounds can have on humans and aquatic organisms that come in contact with them.

Anticlastogenic Activity of Aqueous Extract of Agaricus blazei in Drug-Metabolizing Cells (HTCs) During Cell Cycle.

ABSTRACT The mushroom Agaricus blazei has been extensively investigated because of evidence of its antimutagenic, antitumor, and anticarcinogenic activities. This study investigated the clastogenic and/or anticlastogenic activity of aqueous extract of Agaricus blazei (10% w/v) in drug-metabolizing rat hepatoma tissue cells (HTCs), with continuous treatment and treatment during different phases of the cell cycle. DNA damage was induced utilizing two direct-acting agents-methyl methane sulfonate and ethyl methane sulfonate-and two indirect-acting agents-2-aminoanthracene and cyclophosphamide. The aqueous extract of A. blazei with either continuous treatment or treatment during different phases of the cell cycle showed clastogenic activity. The results with continuous treatment showed that A. blazei does not protect against DNA damage-inducing agents that are direct acting. Meanwhile, when combined with indirect-acting agents, a protective effect was demonstrated. A protective effect was also found during different phases of the cell cycle when cells were treated with indirect-acting agents. The protective effects against indirect-acting agents (continuous treatment and during the different phases of the cell cycle) suggest that A. blazei may provide some health benefits to the public when used as a functional food.

Antigenotoxicity of Agaricus blazei mushroom organic and aqueous extracts in chromosomal aberration and cytokinesis block micronucleus assays in CHO-k1 and HTC cells.

Agaricus blazei (Ab) has become popularly known for its medicinal properties. Scientifically, it has been tested with regard to its capacity to protect genetic material against damage. We examined different organic extracts (methanolic extract -- ME, hexanic extract -- HE and n-butanolic extract -- BE) and an aqueous extract (AE) of Ab, for their capacity to induce DNA damage as well as for their protective effect. Genetic damage was determined by the chromosomal aberration assay (CA) in CHO-k1 cells for all extracts and the cytokinesis block micronucleus assay (CBMN) in non drug-metabolizing (CHO-k1) and drug-metabolizing (HTC) cell lines for extract BE only. The extracts did not show clastogenicity but showed anticlastogenicity. The greatest percent reduction obtained were with BE (105%) and AE (126%) treatments in CA. BE treatment did not display genotoxicity in CHO-k1, but was genotoxic in HTC. However, BE was shown to be antigenotoxic causing decreased micronucleus frequency in HTC and CHO-k1 cells. These results suggest that all the extracts contained protective substances, but in some cases they could show a genotoxic effect with regard to metabolism. Therefore, these findings warrant caution in the use of this mushroom by the population.