Ferulic Acid Esterase Producing Lactobacillus johnsonii from Goat Feces as Corn Silage Inoculants.

Ferulic acid esterase (FAE+)-producing lactobacilli are being studied as silage inoculants due to their potential of increasing forage fiber digestibility. In this work, three FAE+ Lactobacillus (L.) johnsonii strains were isolated from caprine feces and characterized according to their potential probiotic characteristics and as silage inoculants. Limosilactobacillus fermentum CRL1446, a human probiotic isolated from goat cheese, was also included in the experiments as a potential silage inoculant. FAE activity quantification, probiotic characterization, and growth in maize aqueous extract indicated that L. johnsonii ETC187 might have a better inoculant and probiotic aptitude. Nevertheless, results in whole-corn mini silos indicated that, although acid detergent fiber (ADF) was significantly reduced by this strain (3% compared with the uninoculated (UN) group), L. johnsonii ETC150 and CRL1446 not only induced similar ADF reduction but also reduced dry matter (DM) loss (by 7.3% and 6.5%, respectively) compared with the UN group. Additionally, CRL1446 increased in vitro DM degradability by 10%. All treatments reduced gas losses when compared with the UN group. The potential probiotic features of these strains, as well as their beneficial impact on corn fermentation shown in this study, encourage further studies as enhancers in animal production.

Aroma compounds and consumer acceptability of soybean paste fermented by lactobacilli.

Consumption of soybean-based foods is affected by the flavor of the legume; due to the presence of undesirable compounds called "beany flavors". To solve this problem, the influence of solid state fermentation by lactobacilli on the production of volatile compounds in soybean paste was determined. The volatile's production was measured by gas chromatography. Forty compounds were identified in the different soybean pastes studied. The results showed that fermentation stimulates the production of desirable volatile compounds in foods such as ketones (22-75%) and decreased unpleasant compounds (10-84%).The consumers acceptance study showed that a group of participants (30% approximately) preferred the fermented samples associated with sweet and acid aroma like yogurt. In conclusion, the fermentation positively influences the generation of desirable volatile compounds and completely reduces hexanal in one sample. Therefore, fermentation with the studied strains is a valid strategy to modify the aroma profile of a soybean-based food matrix. Supplementary Information: The online version contains supplementary material available at 10.1007/s13197-021-05210-5.

Spray dried lactobacilli maintain viability and feruloyl esterase activity during prolonged storage and under gastrointestinal tract conditions.

The use of lactobacilli with feruloyl esterase (FE) activity in the development of functional foods has gained considerable interest in recent years. Microencapsulation of FE-producing bacteria to facilitate their incorporation into food is a challenge. The aim of this study was to evaluate survival and maintenance of FE activity during storage at 4 °C and under simulated gastrointestinal tract (GIT) conditions of microcapsules of FE-producing Lactobacillus (Lb.) strains obtained by spray drying. Lb. fermentum CRL1446 and Lb. johnsonii CRL1231 powders maintained viability at concentrations ≥ 106 CFU/g (minimum probiotic dose) when stored at 4 °C for 12 months. Lb. acidophilus CRL1014 powders were only able to maintain ≥ 106 CFU/g during 4 months of storage. FE activity was conserved in three microencapsulated strains evaluated, an increase of specific activity being observed until month 12 of storage. Powders of the three strains incubated under GIT conditions maintained their viability (≥ 106 CFU/g), but specific FE activity was only detected in Lb. fermentum and Lb. johnsonii powders (0.80-0.83 and 0.21-0.56 U/mg, respectively). CRL1446 and CRL1231 microcapsules were able to resist prolonged storage and GIT conditions, retaining FE activity and preserving their probiotic potential and could be incorporated into functional foods.

The secreted esterase of Propionibacterium freudenreichii has a major role in cheese lipolysis.

Free fatty acids are important flavor compounds in cheese. Propionibacterium freudenreichii is the main agent of their release through lipolysis in Swiss cheese. Our aim was to identify the esterase(s) involved in lipolysis by P. freudenreichii. We targeted two previously identified esterases: one secreted esterase, PF#279, and one putative cell wall-anchored esterase, PF#774. To evaluate their role in lipolysis, we constructed overexpression and knockout mutants of P. freudenreichii CIRM-BIA1(T) for each corresponding gene. The sequences of both genes were also compared in 21 wild-type strains. All strains were assessed for their lipolytic activity on milk fat. The lipolytic activity observed matched data previously reported in cheese, thus validating the relevance of the method used. The mutants overexpressing PF#279 or PF#774 released four times more fatty acids than the wild-type strain, demonstrating that both enzymes are lipolytic esterases. However, inactivation of the pf279 gene induced a 75% reduction in the lipolytic activity compared to that of the wild-type strain, whereas inactivation of the pf774 gene did not modify the phenotype. Two of the 21 wild-type strains tested did not display any detectable lipolytic activity. Interestingly, these two strains exhibited the same single-nucleotide deletion at the beginning of the pf279 gene sequence, leading to a premature stop codon, whereas they harbored a pf774 gene highly similar to that of the other strains. Taken together, these results clearly demonstrate that PF#279 is the main lipolytic esterase in P. freudenreichii and a key agent of Swiss cheese lipolysis.

Differentiation of lactic acid bacteria strains by postelectrophoretic detection of esterases.

Lactic acid bacteria (LAB) comprise a diverse group of Gram-positive, non-spore-forming microorganisms. These bacteria are widely used in food technology. The species identification of LAB depends mainly on physiological and biochemical criteria. The esterolytic systems of LAB remain poorly characterized. Esterases (EC 3.1.1.3) represent a diverse group of hydrolases catalyzing the cleavage and formation of esters bonds Screening of esterases is usually performed either by employing chromophoric substances (e.g., alpha- or beta-naphthyl esters of short-chain fatty acids). The post-electrophoretic detection of esterases is a sensitive technique applied in bacterial systems, that mainly provides information on the similarity of strains within the same species or subspecies according to their esterase patterns. This technique is principally used to determine the number and substrate specificity of esterases and lipases, revealing the complexity of lipase and esterase systems. The present chapter describes the technique of polyacrylamide gel electrophoresis (PAGE; in the absence of sodium dodecyl sulfate [SDS]), in non-denaturing conditions, to find intracellular fractions for strain typing of LAB.

Determination of esterolytic and lipolytic activities of lactic acid bacteria.

Lactic acid bacteria (LAB) are considered weakly lipolytic compared with many other groups of bacteria (e.g., Pseudomonas, Bacillus, and Achromobacter). The esterolytic and lipolytic systems of dairy LAB remain poorly characterized. Esterases from lactic acid bacteria, yeasts, and Pseudomonas organisms may be involved in the development of fruity flavors in foods, and pregastric lipase and esterases are essential for the development of typical flavor in Italian cheese. Microbial lipases and esterases may improve quality or accelerate the maturation of cheeses, cured bacon, and fermented sausages. Lipases are defined as glycerol ester hydrolases (EC 3.1.1.3) that hydrolyze tri-, di-, and monoglycerides present at an oil-water interface. Esterases (EC 3.1.1.6) hydrolyze esters in solution and may also hydrolyze tri- and especially di- and monoglycerides containing short-chain fatty acids. Some probiotic strains of LAB can hydrolyze the triglycerides, releasing most short and medium chain, and essential fatty acids, which are valuable to today's health-conscious consumer. Medium chain fatty acids (C6-C14), in particular, have become accepted treatment for patients with malabsorption symptoms, a variety of metabolic disorders, cholesterol problems, and infant malnutrition. These probiotic bacteria could alleviate lipase deficiency in the digestive tract during digestion (steatorrhea). In this chapter, we describe different methods routinely used in our laboratory to determine the esterolytic and lipolytic activity of LAB. These techniques include the use of alpha- and beta-naphthyl derivatives of fatty acids (chromogenic method), the p-nitrophenyl (pNP) derivative of fatty acids (chromogenic method), and triglycerides (agar-well assay technique and titrimetric test) as substrates.