Thereafter, the plate was shaken carefully and absorbance was measured using a Labsystem Multiscan MCC/340 plate reader,

at 340 nm every 15 s over 10 min to monitor the oxidation of NADH. Aspartase activity was calculated according to: To validate the repeatability of the method, six randomly selected strains were grown independently in triplicate for protein extraction and aspartase activity, which were repeated three times for each cultivation (data not shown; maximum SD=±14.08%). Based on the results showing good repeatability of the technique, aspartase activity from the remaining strains was determined as the mean of three parallel assays. To compare aspartase activity determined in PD-0332991 mw single isolates it was important to fix the growth conditions, as the highest enzyme activity was encountered at the late log phase of growth (data not shown).

Given the nature of the assay, the lowest quantification limit of aspartase activity was set at 100 units. Therefore, strains displaying aspartase activity lower than 100 units are considered as one group without a precisely determined activity. Figure 2 shows the aspartase activity of the PAB strains analysed in this study as well as the percentage of ABT199 strains belonging to the chart segments representing aspartase activity levels of 0–25%, 25–50%, 50–75% and 75–100% with respect to the highest activity detected in this study. More than 70% of the strains tested belonged to the segment representing the lowest aspartase activity (0–25%). Of this group, the aspartase activity of 42 strains was assayed as being lower than Cytidine deaminase 100 units. Of the remaining strains, the percentage categorized to the segments representing higher aspartase activity was decreased in parallel to the increase in activity (19% in activity level group 25–50%, 5% in activity level

group 50–75% and 3% in activity level group 75–100%). Thus, low aspartase activity was a common characteristic of propionibacteria of Swiss-type cheese origin studied here. Some strains with high aspartase activity were found, but at a low proportion compared with the isolates with low activity. Although a wide range of aspartase activity was detected between different strains, the commonly used dairy P. freudenreichii ssp. freudenreichii and shermanii could not be differentiated on the basis of enzyme activity. The role of aspartase activity in Swiss-type cheese manufacture has been recognized to have considerable impact on the formation of eyes and flavour (Langsrud & Reinbold, 1973; Thierry et al., 2005). Yet, as shown here, aspartase activity is strain dependent and so each strain must be tested separately in order to be able to choose the most suitable starter culture for cheese production.