Inhibition of Uropathogenic Escherichia coli by Cranberry Juice:  A New Antiadherence Assay

Abstract

A combination of microplate technology and turbidity assessment for testing the adherence of P-fimbriated Escherichia coli to human uroepithelial cell line T24, validated with the addition of the known inhibitor 4-O-alpha-D-galactopyranosyl-alpha-D-galactopyranose (galabiose), resulted in a high-throughput, biologically relevant assessment of cranberry (Vaccinium macrocarpon). P-fimbriated ATCC E. coli strains 25922, 29194, and 49161 were inhibited by galabiose. ATCC 29194, a representative urine isolate containing the papGII allele (Class II fimbrial adhesin) and demonstrating the most significant inhibition in the presence of galabiose, was chosen for further testing. In this assay, a low-polarity fraction of cranberry juice cocktail demonstrated dose-dependent inhibition of E. coli adherence. Reported here, for the first time in V. macrocarpon, are 1-O-methylgalactose, prunin, and phlorizin, identified in an active fraction of cranberry juice concentrate. This in vitro assay will be useful for the standardization of cranberry dietary supplements and is currently being used for bioassay-guided fractionation of cranberry juice concentrate.

Coumaroyl Iridoids and a Depside from Cranberry (Vaccinium macrocarpon)

“…40 In brief, p-fimbriated E. coli were grown on CFA agar to promote fimbrial growth. Human T-24 cells were grown to confluence in 96-well microplates.…”

Antioxidant and Anti-nitric Oxide Components from <i>Quercus glauca</i>

“…Our research group recently found that the crude extract of title plant and its sub-fractions had promising antioxidant activities (IC 50 =26.44, 15.72 µg/mL, for crude extract and EtOAc layer, respectively). Herein, we report the bioassay-directed fractionation leading to the isolation of two new lignan glycosides (1, 2), along with fourteen known compounds including 4 lignanoids: dihydrodehydodiconiferyl alcohol (3), 10) (+ )-lyoniresinol-3α-O-α-L-rhamnoside (4), 11) 5-methoxy-(+ )-isolariciresinol-9′-O-α-L-rhamnoside (5), 12) (+ )-acviculin (6) 12) ; 5 triterpenoids: batrogenic acid (7), 13) trachelosperogenin A (8), 14) paradrymonoside (9), 15) 24-epipinfaensin (10), 16) rosamultin (11) 17) ; 2 flavonoids: arimadendrin (12), 18) naringenin-7-O-β-D-glucoside (13) 19) ; 2 aromatics:…”

An experimental implementation of chemical subtraction

“…In designing an assay to measure inhibition of E. coli adherence to a human uroepithelial cell line [15], it was determined that, at sufficiently low pH, the cranberry constituent benzoic acid and its urinary metabolite hippuric acid have bacteriostatic and/or bactericidal activity. The search for anti-adherent compounds was thus confounded by the presence of benzoic acid, which apparently killed the bacteria before they could be inhibited from adhering.…”

“…The full details of this assay are described elsewhere [15] but, in brief, the procedure is as follows: Immortalized human uroepithelial T24 cells (ATCC HTB4) were grown to confluence in wells of a microplate. A urinary E. coli (ATCC 29194) isolate containing the uropathogenic papGII gene was grown on CFA agar, suspended in saline to 10 7 bacteria per mL, mixed with test cranberry fractions, and incubated with the T24 cells for 1 hour.…”

“…The initial methanol fraction prior to CCC separation was moderately active, with 36% inhibition of adherence of a uropathogenic strain of E. coli to human uroepithelial cells [15] at 100 μg/mL (p = 0.04). The most polar fraction from the CCC experiment retained this activity (58%, p = 0.03) at the same concentration.…”