Alternative methods of globotrioside production using Vero cells: a microcarrier system procedure

<p>Abstract</p> <p>Background</p> <p>Glycolipids are one component of cell membranes, and are found most prevalently at the surface of the plasma membrane. Animal cells take in amphipathic glycosides, which are later glycosylated after assimilation in biosynthetic pathways. Gycosylated glycosides are released outside of cells to the surrounding culture medium. This represents an accessible method of obtaining complex glycosides.</p> <p>Results</p> <p>Vero cells are sensitive to Shiga toxins and are known to express the glycosides globotriaosyl ceramide (Gb3) and globotetraosyl ceramide (Gb4) on the surface of the plasma membrane. By administering amphipathic lactosides to Vero cells, the above mentioned glycolipids could be produced by the action of cellular enzymes. In our study, the optimum conditions (seeded cell number, incubated time period, 12-azidododecyl lactoside concentration and medium volume) for the production of Gb3 analogue were investigated. The 87.9 μg/100 mm dish (11.7 % yield) Gb3 analogue was produced under appropriate conditions. The large-scale culture of Vero cells using a microcarrier culture method with repetitions produced about 30 mg of the Gb3 analogue.</p> <p>Conclusion</p> <p>The mass production of glycosides in Vero cells was carried out on a microcarrier with repeated administration of 12-azidododecyl lactoside. The results indicated that the use of both a microcarrier culture and repetition were highly effective in the production of Gb3, Gb4 and sialyl lactoside (GM3) type-oligosaccharides.</p

Cytotoxic activity of triazole-containing alkyl β-D-glucopyranosides on a human T-cell leukemia cell line

BACKGROUND: Simple glycoside surfactants represent a class of chemicals that are produced from renewable raw materials. They are considered to be environmentally safe and, therefore, are increasingly used as pharmaceuticals, detergents, and personal care products. Although they display low to moderate toxicity in cells in culture, the underlying mechanisms of surfactant-mediated cytotoxicity are poorly investigated. RESULTS: We synthesized a series of triazole-linked (fluoro)alkyl β-glucopyranosides using the copper-catalyzed azide-alkyne reaction, one of many popular “click” reactions that enable efficient preparation of structurally diverse compounds, and investigate the toxicity of this novel class of surfactant in the Jurkat cell line. Similar to other carbohydrate surfactants, the cytotoxicity of the triazole-linked alkyl β-glucopyranosides was low, with IC(50) values decreasing from 1198 to 24 μM as the hydrophobic tail length increased from 8 to 16 carbons. The two alkyl β-glucopyranosides with the longest hydrophobic tails caused apoptosis by mechanisms involving mitochondrial depolarization and caspase-3 activation. CONCLUSIONS: Triazole-linked, glucose-based surfactants 4a-g and other carbohydrate surfactants may cause apoptosis, and not necrosis, at low micromolar concentrations via induction of the intrinsic apoptotic cascade; however, additional studies are needed to fully explore the molecular mechanisms of their toxicity. [Figure: see text] ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13065-014-0072-1) contains supplementary material, which is available to authorized users