Abstract
Uptake of the fluorescent probe 1‐N‐phenylnaphthylamine (NPN), as
adapted to an automated spectrofluorometer enabling multiwell reading
of microtitre plates, was applied to determine permeability changes in
Gram‐negative bacteria. An intact outer membrane is a permeability
barrier, and excludes hydrophobic substances such as NPN but, once
damaged, it can allow the entry of NPN to the phospholipid layer,
resulting in prominent fluorescence. With Escherichia coli O157, Pseudomonas aeruginosa, and Salmonella typhimurium
as test organisms and ethylenediaminetetraacetic acid and sodium
hexametaphosphate as the model permeabilizers, quantitative and highly
reproducible NPN uptake levels were obtained that differed
characteristically between the test bacteria. Furthermore, citric acid
was shown to be a potent permeabilizer at millimolar concentrations, its
effect being partly (Ps. aeruginosa, Salm. typhimurium) or almost totally (E. coli O157) abolished by MgCl2,
suggesting that part of the action occurs by chelation. Sodium citrate
induced weak NPN uptake, which was totally abolished by MgCl2.
In conclusion, the NPN uptake assay with the automated
spectrofluorometer serves as a convenient method in analysing and
quantifying the effects of external agents, including potential food
preservatives, on Gram‐negative bacteria.
Original language | English |
---|---|
Pages (from-to) | 213-219 |
Journal | Journal of Applied Microbiology |
Volume | 88 |
Issue number | 2 |
DOIs | |
Publication status | Published - 2000 |
MoE publication type | A1 Journal article-refereed |