Use of o-phthalaldehyde to detect O-phosphorylethanolamine in bacterial lipopolysaccharide.
We have developed a method to measure O-phosphorylethanolamine groups in bacterial lipopolysaccharide using a fluorescent reagent, o-phthalaldehyde. The optimal excitation and emission wavelengths were 335 nm and 450 nm, respectively. The reaction was pH-dependent with an optimum at pH 10.5. The maximum fluorescence intensity occurred two min after mixing lipopolysaccharide with the reagent at pH 10.5. The assay was linear over a range of 1 microgram to 100 micrograms of lipopolysaccharide. When we compared the amount of primary amine (as O-phosphorylethanolamine) in native and p-hydroxyphenylacetic acid-derivatized lipopolysaccharide, we found that 97% of amine groups in native lipopolysaccharide were derivatized by p-hydroxyphenylacetic acid in the presence of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide.
Yeh
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Department of Microbiology, School of Medicine and Biomedical Sciences, State University of New York, Buffalo.
Price
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RM
Jacobs
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D004983:Ethanolamines / Q000032:analysis*; D005453:Fluorescence; D006863:Hydrogen-Ion Concentration; D008070:Lipopolysaccharides / Q000737:chemistry* / Q000378:metabolism; D008722:Methods; D012486:Salmonella typhimurium / Q000378:metabolism; D013053:Spectrophotometry; D013696:Temperature; D009764:o-Phthalaldehyde
D004983:Ethanolamines; D008070:Lipopolysaccharides; D009764:o-Phthalaldehyde; C005448:phosphorylethanolamine