3.3. Urease Inhibition Assay

Urease inhibition was performed as described elsewhere [41]. Briefly, 10 μL of 10⁸ CFU/mL H. pylori suspension was incubated with equal amount of serially diluted compound in 96 well microplates for 30 min at 37 °C. Subsequently, 200 μL of detection reagent composed of 50 mM phosphate buffer, pH 6.8 containing 500 mM urea, and 0.02% phenol red was added to each well. The color development was measured at 555 nm in 5 min intervals. Controls included bacteria with the reagent and reagent with each compound. The percentage of inhibition was calculated by the equation % inhibition = [(activity without compound − activity with compound)/(activity without compound)] × 100. The activity was compared to a reference urease inhibitor acetohdyroxamic acid (Sigma Aldrich, St. Louis, MO, USA).