Probiotic Lactic Acid Bacteria for Vaginal Application. Optimization of Biomass Production and Freeze-Drying Conditions

MARCHESI, ANTONELLA; NADER-MACÍAS, MARÍA ELENA FÁTIMA

Applied Microbiology

MDPI

Lugar: Basel; Año: 2023 vol. 3 p. 519 - 535

bacteriathat maintain their characteristics during the production and further storage, in order to exert theclaimed probiotic effect. Four probiotic strains—Lactobacillus gasseri CRL1320, Limosilactobacillus reuteriCRL1324, Ligilactobacillus salivarius CRL1328 and Lacticaseibacillus rhamnosus CRL1332—originallyisolated from vagina of healthy women exhibiting beneficial properties were evaluated. Therefore, theoptimization of growth parameters and dry biomass production with high viability while preservingprobiotic features of strains is a great challenge. Different growth conditions in MRS medium wereset up in a laboratory bioreactor; two initial pH and agitation were recorded speeds during static orcontrolled fermentations. Production conditions of 37 C with controlled pH 6.5 and 5.5-MRS with150 and 75 rpm stirring speeds were used for CRL1329 and CRL1332 strains, respectively, while staticand free pH MRS for CRL1324 and CRL1328 probiotics allowed us to obtain maximal cell viabilitycounts. However, during 12 h of fermentation, biomass yields of 19.3, 16.2 and 15.2 g/L were achievedwhen CRL1329, CRL1328 and CRL1324 probiotic strains were grown in static and free pH MRS. Thehighest biomass yield for CRL1332 strain was produced under controlled MRS-initial pH 6.5 and75 rpm fermentation conditions. To preserve probiotic high biomass viability, freeze-drying wascarried out in the presence of different cryoprotective agents. Thus, the highest viable numbers(10.9–11.8 log CFU/g) with survival rates between 91.3 and 95.6% were attained in the presence of10% trehalose (L. reuteri and L. salivarius), lactose (L. rhammosus) and lactose + trehalose + sucrosemix (L. gasseri). When stability during post-freeze-drying storage was evaluated, probiotic strainsshowed a remarkably higher viability recovery when stored at 4 C than at 25 C for 12 and 3 months,respectively. In addition, surface characteristics of vaginal probiotics were affected to different extentsduring storage depending on the strain, protective agent and storage time/temperature. Criticalfactors for growth conditions, drying process and storage stability of probiotic lactobacilli strainswere optimized in view to preserve cell high viability and surface features for the design of vaginalprobiotic formula.