Cellular Antioxidant, Anti-Inflammatory, and Antiproliferative Activities from the Flowers, Leaves and Fruits of Gallesia integrifolia Spreng Harms.

Gallesia integrifolia, a notable species in the Atlantic Forest, has been traditionally employed in folk medicine for treating rheumatism, asthma, and worms. This study investigated the cellular antioxidant, antiproliferative, and anti-inflammatory activities of the essential oils (EOs) and crude extracts (CEs) from G. integrifolia flowers, fruits, and leaves. The chemical identification of EOs was performed by GC-MS and CEs by UHPLC-MS. Cellular antioxidant and anti-inflammatory activities were assessed through mouse macrophage cell culture. In addition, the antiproliferative potential was evaluated in gastric, colorectal, breast, and lung tumor cell lines and non-tumor VERO cells. EOs predominantly contained organosulfur compounds in flowers (96.29%), fruits (94.94%), and leaves (90.72%). We found the main compound is 2,2'-Disulfanediyldiethanethiol in the EOs of flowers (47.00%), leaves (41.82%), and fruits (44.39%). Phenolic compounds were identified in CEs. The EOs and CEs demonstrated potential against the tumor cell lines tested (GI50 between 51 and 230 µg/mL). The selectivity index values were greater than 1.0 (1.01 to 3.37), suggesting a relative safety profile. Moreover, the anti-inflammatory activity IC50 ranged from 36.00 to 268 µg/mL, and the cellular oxidation inhibition ranged from 69% to 82%. The results suggest that oils and extracts derived from G. integrifolia have potential for use in various industrial sectors.

Lithium bioaccumulation in Lentinus crinitus mycelia grown in media with different lithium sources and pH values.

Lentinus crinitus bioaccumulates lithium in mycelia, but bioaccumulation may be affected by pH of the culture medium. Lithium is used in clinical practice as a mood stabilizer and antidepressant. This study aimed to assess the effect of culture medium pH and lithium source (LiCl or Li2CO3) on lithium bioaccumulation in vegetative mycelia of L. crinitus grown in malt extract broth. Lentinus crinitus U9-1 was cultured in malt extract broth supplemented with Li2CO3 or LiCl (50 mg L-1 lithium) in the pH range of 3.0 to 6.0. The pH was adjusted using HCl solution. The results showed that medium pH affected mycelial biomass production, lithium bioaccumulation in mycelial biomass, and lithium transfer from the culture medium to mycelial biomass. The effect of lithium source on the bioaccumulation capacity of mycelial biomass varied according to pH. At pH 4.0, both lithium sources stimulated mycelial biomass production compared to the control without the addition of lithium. At pH 5.5, Li2CO3 provided the highest lithium bioaccumulation in mycelial biomass. Lithium transfer from the culture medium to mycelia was highest in Li2CO3-supplemented cultures at pH 4.5. LiCl reduced hyphal width compared with Li2CO3 and the control. However, pH and lithium sources did not affect the formation of clamp connections in hyphae. For the first time, the influence of the pH of the culture medium on lithium bioaccumulation by Lentinus crinitus is reported. Finally, we conclude that the culture medium pH affected lithium transfer and bioaccumulation in mycelial biomass differently depending on the lithium source. Additionally, we report the presence of clamp connections in the hyphae of L. crinitus as an indicator of even growth.

Long-term cryopreservation of Lentinus crinitus strains by wheat grain technique.

Lentinus crinitus (Basidiomycota: Polyporales) is a saprophytic fungus with biotechnological importance described more than 20 years ago. However, there are few studies on the long-term preservation of this basidiomycete. Cryopreservation is a long-term storage technique that reduces the metabolic activity of microorganisms, but its success depends on the adjustment of the freezing process, the cryoprotectants, and the protective substrates for each species. This study aimed to assess the mycelial viability and genetic stability of L. crinitus strains cryopreserved at -86 °C for two years by the wheat grain technique using different cryoprotectants and freezing methods. Three strains of L. crinitus (U9-1, U13-5, and U15-12) were subjected to different concentrations and types of cryoprotectants (dimethyl sulfoxide, glycerol, glucose, and sucrose), freezing methods such as immediate freezing from 25 to -86 °C and progressing freezing from 25 to -86 °C in a freezing container with isopropyl alcohol to control the rate of cell freezing at -1 °C min-1, protective substrate (wheat grain and 2% malt extract agar), and cryopreservation period (1, 6, 12, and 24 months). After thawing, samples were evaluated for mycelial viability, time to mycelial recovery, mycelial stability, and genetic stability of the fungus. All techniques achieved effective cryopreservation at -86 °C, mainly with the wheat grain technique. All cryoprotectants (3.5% glycerol, 1.5% dimethyl sulfoxide, 25% sucrose, and 5% glucose), freezing methods (immediate and gradual), and protective substrate (wheat grain and malt extract agar) were effective for cryopreservation of the three L.crinitus strains in an ultra-low temperature freezer for two years. Mycelial viability, mycelial stability, and genetic stability of the fungus were not affected after two-year cryopreservation, evidencing the robustness of the long-term cryopreservation technique and the fungus.

Endophytic fungi of Brunfelsia uniflora: isolation, cryopreservation, and determination of enzymatic and antioxidant activity.

Brunfelsia uniflora (Pohl.) D. Don (Solanaceae), commonly known as manacá-de-cheiro, is widely distributed in Brazil and used by local indigenous peoples as an antirheumatic, antisyphilitic, depurative, emetic, vermifuge, and purgative agent. Several studies have examined the biological activities and phytochemical profile of Brunfelsia; however, few have focused on the diversity of endophytic microorganisms that colonize members of the genus. This study aimed to isolate and cryopreserve endophytic fungi from B. uniflora and determine their cellulase, laccase, and antioxidant activities. Endophytic fungi were isolated from B. uniflora stems, cultured on wheat grains, immersed in a 150 g L-1 aqueous sucrose solution, and cryopreserved at - 80 °C for 1 and 6 months. Cellulase activity was determined by a qualitative test using carboxymethylcellulose medium and laccase activity by a quantitative test based on the oxidation of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonate). Prior to antioxidant activity assays, fungi were grown in malt extract broth for production of mycelial biomass. A methanolic extract was prepared for evaluation of DPPH· scavenging activity, FRAP activity, and total phenolic content. A total of 46 endophytic fungal isolates were obtained from B. uniflora stems and classified into 24 groups according to morphological similarities. B. uniflora was shown to harbor different genera of ascomycete fungi as endophytic organisms. Mycelial viability was observed after 1 and 6 months of cryopreservation at - 80 °C. Fungi exhibited cellulase and laccase activities. Isolate CE23 had the highest laccase activity after 7 days of cultivation. Twelve isolates were found to have low total phenolic contents and DPPH· and FRAP activities.

Bioaccumulation of Lithium (Li2CO3) in Mycelia of the Culinary-Medicinal Oyster Mushroom, Pleurotus ostreatus (Agaricomycetes).

Pleurotus ostreatus is a white-rot mushroom that bioaccumulates metals in basidiocarps and vegetative mycelia. This fungus has been used in soil and water bioremediation of several heavy metals; however, there are few studies of lithium mycelial bioaccumulation for pharmacological use. The aim of this study was to evaluate lithium bioaccumulation in P. ostreatus mycelia grown in a liquid malt extract cultivation medium with Li2CO3 or LiCl. Each lithium source was added to the medium to obtain a concentration of 0, 5, 10, 15, 20, 25, 30, 40, 50, 100, or 200 mg · L-1 lithium. The highest bioaccumulation of lithium in mycelia was 1575.29 µg · g-1 upon treatment with 40 mg · L-1 Li2CO3. P. ostreatus mycelia produce biomass and bioaccumulate both lithium sources, but more lithium bioaccumulates when in the form of Li2CO3. This study provides a prospective for the development of biotechnological products with high aggregate values and alternative ways to deliver lithium and eventually other salts for pharmacological use.

Mycelial antineoplastic activity of Agaricus blazei.

Basidiocarp of Agaricus blazei (=Agaricus brasiliensis; =Agaricus subrufescens) is used as teas or capsules due to its antineoplastic effect but there are few reports of using mycelium for this purpose. The objective of this study was to evaluate the antineoplastic activity on sarcoma 180 cells implanted in mice of two forms of preparation of the mycelium from two A. blazei strains grown in culture medium with different concentrations of isolated soy protein. Mycelia were grown in Pontecorvo medium with different concentrations of isolated soybean protein (ISP). Mycelial hot water extract, moistened mycelial powder, hot water extract of green tea, Ifosfamida(®) (ifosfamide drug), and saline solution were administered daily by gavage in mice with sarcoma 180 cells to evaluate antineoplastic activity. It was concluded that antineoplastic activity was the same for both strains, except when used as moistened mycelial powder, which rules out the use of mycelial powder in capsules. Mycelial hot water extract had high antineoplastic activity with lower metabolic demand on the spleen and maintenance of normal blood parameters. Mycelial growth in different ISP concentrations had the same antineoplastic activity. Also the vegetative mycelium was as effective as the basidiocarp for sarcoma 180 tumor inhibition. Green tea was as effective as mycelial hot water extract.

Effects of Agaricus brasiliensis mushroom in Walker-256 tumor-bearing rats.

Agaricus brasiliensis is a mushroom native to São Paulo State, Brazil, that is studied for its medicinal proprieties. This work aimed to investigate the antitumoral activity of A. brasiliensis extracts and pure powdered basidiocarp preparation using Walker-256 (W256) tumor-bearing rats, a model for cancer-related cachexia studies. The rats were treated for 14 days by gavage (136 mg/kg) and at the end of the experiment tumors were collected to calculate mass and volume. Blood was collected for determination of plasma glucose, albumin, alanine aminotransferase (ALT), and aspartate aminotransferase (AST). Hepatic and tumor enzymes indicating oxidative stress were also evaluated. The results showed that all 4 treatments (pure powdered basidiocarp and aqueous, acid, and alkaline extracts) significantly reduced tumor size and promoted gain in body weight. Plasmatic analysis showed a reduction in AST level and increased glycemia in the treated rats. Pure basidiocarp preparations improved the liver catalase and superoxide dismutase activity, but did not change the glutathione S-transferase activity. The data collected from the W256 tumor-bearing rats revealed the beneficial effects of A. brasiliensis in tumor treatment, mainly related to cachexia. The benefits can be partly related to antioxidant activity and to reduction of weight loss and tumor growth.