Selection of β-glucosidase positive strains from naturally fermented table olives

Oleuropein is a β-glucosidase compound responsible for intense bitter flavour of drupes and its hydrolysis is necessary to make the final product edible for the consumer. The use of microbial starter cultures, able to degrade bitter compounds, is useful to accelerate, control and lead the fermentative process, maintaining the original organoleptic characteristic of the final product. Lactobacillus plantarum species is considered a potential starter culture in fermented table olives, for the high versatility, adaptation in different environments, ability to degrade oleuropein and for salt tolerance. The aim of the present study was to select β-glucosidase positive strains able to conduct the fermentation and to accelerate the debittering process of table olives. For this purpose, the oleuropein-degrading ability of Lactobacillus plantarum strains was tested under different stress conditions and the gene encoding for the β-glucosidase enzyme was detected. The beta-glucosidase activity of L. plantarum strains, previously isolated from Nocellara Etnea table olives and identified by sequencing, was determined enzymatically by using p-nitrophenyl-β-D-glucoside substrate, under the following stress condition: salt concentration (4.0, 5.0, 6.0), pH (4.5, 5.5) temperature (16 °C and 32 °C). The enzymatic test was also performed in simulated brine medium at the same stress conditions described above. In addition, the ability to degrade oleuropein was estimated using MRS broth added with pure oleuropein (1g/L), under the stress condition mentioned above. For each L. plantarum strain the presence of the gene encoding for the β-glucosidase enzyme was investigated through PCR assay. The results obtained showed good growth performances under different stress conditions for all tested strains. Furthermore, the strains demonstrated both good β-glucosidase activity and oleuropein-degrading ability. In particular, the strains C11C8, F3.6, F3.7, F3.8 and F3.9 exhibited the highest β-glucosidase activity in brine at 5% NaCl at 16 °C, while the strains F1.8M and F1.10 in brine at 4% NaCl at 32 °C. The presence of the gene encoding for β-glucosidase enzyme was revealed only in the strains F1.16, F3.2, F3.6, F3.9 and C11C8. The present study allowed to select L. plantarum strains with promising oleuropein-degrading ability under stress conditions that could be considered as potential starter cultures for table olives fermentation.