A home-based approach to demonstrate column and thin layer chromatography during the COVID-19 pandemic.

The COVID-19 pandemic caused several educational challenges. Conducting laboratory experiments was an uphill task during the pandemic. Here, we developed a low-cost and reliable home-based experimental setup to teach column and thin layer chromatography (TLC) using silica gel granules available at home. Powdered silica gel, prepared by grinding silica gel granules, was used as the stationary phase. Iso-propyl alcohol, purchased from a pharmacy, was diluted with water and used as the mobile phase. A food coloring was chromatographically separated using the designed column. Moreover, TLC plates were prepared using powdered silica gel and a drop of food coloring was separated on TLC plates using the same mobile phase. In the article, we show our experiences by providing methods used to implement this experimental setup. We assume that this experimental setup will be helpful for other universities, research institutes and schools to develop online laboratory curricula to demonstrate basic chromatography techniques required for subjects such as chemistry, biochemistry and biology.

Cytotoxic effects of ergone, a compound isolated from Fulviformes fastuosus.

BACKGROUND: Mushrooms inspired the cuisines of many cultures and conventional medicaments for cancer. However, a substantial number of mushroom species are yet unexplored, possessing an unknown chemical, biological and pharmacological profiles. Fulviformes fastuosus is a terrestrial mushroom, which is commonly found in Sri Lankan woodlands. The current study was aimed at isolation and characterization of a potent cytotoxic compound from F. fastuosus and investigating the apoptotic effect induced by the active principle against cancer and normal cell lines. METHODS: Bioactivity guided isolation of active principles from the methanol extract of F. fastuosus was performed by a rapid extraction and isolation method using different chromatographic techniques. Potential cytotoxic compound was identified using one and two dimensional nuclear magnetic resonance spectroscopy and mass spectrometry. Isolated compound was screened for in vitro cytotoxicity against Hepatocellular carcinoma (HepG-2), Muscle rhabdomyosarcoma (RD) and Rat Wistar liver normal (CC-1) cell lines using 3 4, 5-(dimethylthiazol-2-yl) 2-5-diphenyl tetrazolium bromide (MTT) cell viability assay. Apoptotic features of cells were observed via microscopic examination and ethidium bromide/acridine orange fluorescent staining. RESULTS: The interpretation of spectral data resulted in the identification of the chemical structure as ergosta-4,6,8 (14),22-tetraen-3-one (ergone). Ergone exhibited promising cytotoxic properties against RD cells with less cytotoxicity effect on CC-1 cells. In addition, ergone also possesses a strong cytotoxic effect against HepG-2 cells showing low toxic level for CC-1 cells. Apoptotic features of treated cells were detected via morphological characterization and ethidium bromide/acridine orange staining. CONCLUSION: The present study elaborates the isolation of a potent cytotoxic compound; ergone, from F. fastuosus via a rapid and efficient isolation method. Importantly, ergone has exhibited greater cytotoxic activity against RD cells with high selectivity index compared to cytotoxicity against HepG-2 cells. Ergone can be used in the development of therapeutic strategies for curbing rhabdomyosarcoma.

Antioxidant potential, in vitro cytotoxicity and apoptotic effect induced by crude organic extract of Anthracophyllum lateritium against RD sarcoma cells.

BACKGROUND: Macrofungi have an established history of use in traditional oriental medicine. Anthracophyllum lateritium is a terrestrial macrofungus found in the dry zone forest reserves in Sri Lanka. Yet there are no scientific reports on bioactive properties of this species. Hence, the current study was aimed at determining the antioxidant potential, in vitro antiproliferative activity and apoptotic effect induced by crude methanolic extract of A. lateritium against RD sarcoma cell line. METHOD: The crude extract of A. lateritium was dissolved in methanol (MEFCA) and antioxidant activity was evaluated using in vitro assays: inhibition of DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging, ferric ion reducing power and 2-deoxy-D-ribose degradation assay. Total phenol and flavonoid contents of MEFCA were assayed using folin Ciocalteu method and aluminium chloride colorimetric method. In vitro cytotoxicity was determined using MTT assay against RD cells after 24 h exposure to MEFCA. Ethidium bromide/ acridine orange staining, DNA fragmentation and protein synthesis experiments were used to study the apoptotic features and antiproliferative activities of the treated cells. Glutathione assay and griess nitrite assay were used to analyze the reduced glutathione content and liberation of nitric oxide from apoptotic cells. RESULTS: MEFCA showed promising antioxidant activity with EC50 values of 8.00 ± 0.35 µg/mL for DPPH scavenging and 83.33 ± 0.45 µg/mL for 2-deoxy-D-ribose degradation assay. The phenolic content was 265.15 ± 0.46 of (w/w) % of Gallic acid equivalents and flavonoid content was 173.01 ± 0.35 of (w/w) % of Epigallocatechingallate. A. lateritium showed strong in vitro cytotoxic activity with an EC50 of 18.80 ± 4.83 µg/mL for MTT assay against RD cells. Ethidium bromide/acridine orange staining and DNA fragmentation indicated the apoptotic features of treated cells. Protein levels showed a dose dependent decrease supporting the fact that A. lateritium induces apoptosis of treated cells. Glutathione content and nitric oxide content of cells exhibited a dose dependent increase suggesting the apoptosis of RD cells was mediated by both nitrie ions and nitric oxide. CONCLUSIONS: The crude extract of the A. lateritium exhibited potent antioxidant, antiproliferative activity and apoptotic effect against RD cells providing supportive evidence for the ethnopharmacological use of this fungus in control of oxidative damage and remedy of cancer.