PROTOCOL] Applications of different solvents and conditions for differential extraction of lipopolysaccharide in Gram-negative bacteria

Mai Phuong Nguyen; Le Viet Ha Tran; Hyun Namgoong; Yong-Hak Kim

doi:10.1007/s12275-019-9116-5

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PROTOCOL] Applications of different solvents and conditions for differential extraction of lipopolysaccharide in Gram-negative bacteria: Mai Phuong Nguyen ¹, Le Viet Ha Tran ¹, Hyun Namgoong ², Yong-Hak Kim ¹; Journal of Microbiology 2019;57(8):644-654.
DOI: https://doi.org/10.1007/s12275-019-9116-5
Published online: May 23, 2019

¹Department of Microbiology, Daegu Catholic University School of Medicine, Daegu 42472, Republic of Korea, ²Kolon R&D Division, Seoul 07793, Republic of Korea¹Department of Microbiology, Daegu Catholic University School of Medicine, Daegu 42472, Republic of Korea, ²Kolon R&D Division, Seoul 07793, Republic of Korea

Corresponding author: Yong-Hak Kim , Tel: +82-53-650-4338,

Received: 5 March 2019 • Revised: 10 April 2019 • Accepted: 11 April 2019

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Abstract

Lipopolysaccharide (LPS) is one of the major components in the outer membrane of Gram-negative bacteria. However, its heterogeneity and variability in different bacteria and differentiation conditions make it difficult to extract all of the structural variants. We designed a solution to improve quality and biological activity of LPS extracted from various bacteria with different types of LPS, as compared to conventional
methods
. We introduced a quality index as a simple measure of LPS purity in terms of a degree of polysaccharide content detected by absorbance at 204 nm. Further experiments using gel electrophoresis, endotoxin test, and macrophage activation test were performed to evaluate the performance and reliability of a proposed ‘T-sol’ method and the biological effectiveness and character of the LPS products. We presented that the T-sol method had differential effects on extraction of a RAW 264.7 cell-activating LPS, which was effective in the macrophage activation with similar effects in stimulating the production of TNF-alpha. In conclusion, the T-sol method provides a simple way to improve quality and biological activity of LPS with high yield.

Keywords: Gram-negative bacteria;lipopolysaccharide (LPS);quality index;effective does (ED50);macrophage activation

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PROTOCOL] Applications of different solvents and conditions for differential extraction of lipopolysaccharide in Gram-negative bacteria

J. Microbiol. 2019;57(8):644-654. Published online May 23, 2019

DOI: https://doi.org/10.1007/s12275-019-9116-5

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