Rossana Coda

Long-Term Fungal Inhibitory Activity of Water-Soluble Extracts of Phaseolus vulgaris cv. Pinto and Sourdough Lactic Acid Bacteria during Bread Storage

ABSTRACT The antifungal activity of proteinaceous compounds from different food matrices was investigated. In initial experiments, water-soluble extracts of wheat sourdoughs, cheeses, and vegetables were screened by agar diffusion assays with Penicillium roqueforti DPPMAF1 as the indicator fungus. Water-soluble extracts of sourdough fermented with Lactobacillus brevis AM7 and Phaseolus vulgaris cv. Pinto were selected for further study. The crude water-soluble extracts of L. brevis AM7 sourdough and P. vulgaris cv. Pinto had a MIC of 40 mg of peptide/ml and 30.9 mg of protein/ml, respectively. MICs were markedly lower when chemically synthesized peptides or partially purified protein fractions were used. The water-soluble extract of P. vulgaris cv. Pinto showed inhibition toward a large number of fungal species isolated from bakeries. Phaseolin alpha-type precursor, phaseolin, and erythroagglutinating phytohemagglutinin precursor were identified in the water-soluble extract of P. vulgaris cv. Pinto by nano liquid chromatography-electrospray ionization-tandem mass spectrometry. When the antifungal activity was assayed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, all three proteins were inhibitory. A mixture of eight peptides was identified from the water-soluble extract of sourdough L. brevis AM7, and five of these exhibited inhibitory activity. Bread was made at the pilot plant scale by sourdough fermentation with L. brevis AM7 and addition of the water-soluble extract (27%, vol/wt; 5 mg of protein/ml) of P. vulgaris cv. Pinto. Slices of bread packed in polyethylene bags did not show contamination by fungi until at least 21 days of storage at room temperature, a level of protection comparable to that afforded by 0.3% (wt/wt) calcium propionate.

Molecular adaptation of sourdough Lactobacillus plantarum DC400 under co-cultivation with other lactobacilli.

This work was aimed at investigating the molecular mechanisms of Quorum Sensing (QS) in Lactobacillus plantarum DC400 when co-cultured with other sourdough lactobacilli. The growth and survival of L. plantarum DC400 was not affected when co-cultivated with Lactobacillus sanfranciscensis DPPMA174 or Lactobacillus rossiae A7. Nevertheless, 2-DE analysis showed that the level of protein expression of L. plantarum DC400 increased under co-culture conditions. Although several proteins were commonly induced in both co-cultures, the highest induction was found in co-culture with L. rossiae A7. Overexpressed proteins, related to QS and stress response mechanisms, were identified: DnaK, GroEL, 30S ribosomal protein S1 and S6, ATP synthase subunit beta, adenosylmethionine synthetase (MetK), phosphopyruvate hydratase, phosphoglycerate kinase, elongation factor Tu, putative manganese-dependent inorganic pyrophosphatase, d-lactate dehydrogenase, triosephosphate isomerase, fructose-bisphosphate aldolase and nucleoside-diphosphate kinase. As shown by real-time PCR, expression of the luxS gene of L. plantarum DC400 was also affected during co-cultivation. According to overexpression of MetK and luxS during co-cultivation, synthesis of AI-2-like substances was also influenced by the type of microbial co-cultures. This study showed that expression of some genes/proteins, also QS-related, in L. plantarum was influenced by co-cultivation of other sourdough lactobacilli.

Exploitation of the nutritional and functional characteristics of traditional Italian legumes: The potential of sourdough fermentation

This study aimed at evaluating the composition of nineteen traditional Italian legumes and at investigating the potential of the sourdough fermentation with selected lactic acid bacteria to improve the nutritional and functional features. Traditional Italian legumes, all with product certifications and belonging to Phaseolus vulgaris, Cicer arietinum, Lathyrus sativus, Lens culinaris and Pisum sativum species, were used in this study. Seeds were milled, and flours were analyzed for proximate composition and subjected to sourdough fermentation at 30°C for 24h. Lactobacillus plantarum C48 and Lactobacillus brevis AM7 were used as selected starters. Compared to control doughs, without bacterial inoculum, the concentrations of free amino acids (FAA), soluble fibres, and total phenols increased for all legume sourdoughs. Raffinose decreased of up to ca. 64%. During sourdough fermentation, the level of GABA markedly increased and reached values up to 624mg/kg. Condensed tannins decreased. At the same time, almost all legume sourdoughs showed increases of the antioxidant and phytase activities. As shown by PCA analysis based on data of total FAA, GABA, raffinose, soluble/insoluble dietary fibre, condensed tannins and antioxidant and phytase activities, all legume sourdoughs were clearly differentiated from control doughs. The traditional Italian legumes are bio-diverse, and all showed high levels of nutritional elements and suitability for optimal sourdough fermentation. Legume flours subjected to sourdough fermentation would be suitable to be used alone or better in mixture with cereals, and as gluten-free ingredients for making novel and healthy foods.

Proteomic Analysis by Two-Dimensional Gel Electrophoresis and Starch Characterization of Triticum turgidum L. var. durum Cultivars for Pasta Making

Criteria for durum wheat quality are continuously evolving in response to market pressure and consumers preference. Specific attributes of durum wheat for different end products require more rapid and objective means to grade and classify wheat parcels based on processing potential. A total of 10 durum wheat cultivars were compared for compositional, protein, and starch characteristics. Mean values for the gross composition differed for total protein, gluten, and starch. Two-dimensional electrophoresis (2DE) analysis showed the proteome diversity among the cultivars. As shown by the principal component analysis (PCA) applied to 2DE data of gliadin and glutenin fractions, cultivars differed mainly from the number of proteins and levels of protein expression. As determined by the rapid viscoanalyzer (RVA), swelling power, starch damage, amylose content, and starch pasting properties of 10 cultivars differed significantly. 2DE fingerprinting and amylose content seemed to distinguish specific cultivars being useful tools for selecting suitable durum wheat cultivars for pasta making.

Exopolysaccharides Production during the Fermentation of Soybean and Fava Bean Flours by Leuconostoc mesenteroides DSM 20343

Consumption of legumes is highly recommended due to their beneficial properties. Thus, there is a great interest in developing new legume-based products with good texture. In situ produced microbial exopolysaccharides (EPS) are regarded as efficient texture modifiers in the food industry. In this study, soybean and fava bean flours with different levels of added sucrose were fermented by Leuconostoc mesenteroides DSM 20343. After fermentation, a significant increase in viscosity was observed. Sugars, glucans, fructans, mannitol, lactic acid, and acetic acid were quantified to follow the EPS and metabolite production. By treating the fermented doughs selectively with dextranase or levanase, the major role of glucans in viscosity improvement was confirmed. The roles of microbial fructansucrase and endogenous α-galactosidase in degradation of raffinose family oligosaccharides (RFO) were also investigated. This study shows the potential of Ln. mesenteroides DSM 20343 in tailoring viscosity and RFO profiles in soybean and fava bean flours.

Improvement of the protein quality of wheat bread through faba bean sourdough addition

The effects of the substitution of wheat flour with faba bean flour and faba bean sourdough on the properties of composite bread were investigated. Bread was prepared by replacing wheat flour with 30% of faba bean flour, native or after sourdough fermentation. The addition of faba bean flour influenced the structure of the breads, causing a slight decrease of volume and higher hardness compared to wheat bread. However, when fermented faba bean flour was added, the crumb porosity of the bread was not affected. The addition of 30% of faba bean flour increased wheat bread protein content from 11.6 up to 16.5% of dry matter. The addition of native faba bean flour did not affect the in vitro protein digestibility, resulting similar to wheat bread (64%). On the contrary, faba bean sourdough bread showed higher protein digestibility (73%). Generally, the addition of native faba bean flour caused an improvement of the nutritional indexes of the composite bread, further enhanced when fermentation was carried out. The free amino acid profile, protein chemical score, and biological value index were the highest in faba bean sourdough bread. In addition, the predicted glycemic index was the lowest in faba bean sourdough bread.

Sourdough-type propagation of faba bean flour: Dynamics of microbial consortia and biochemical implications

The microbial ecology of faba bean sourdoughs obtained from an Italian (Ita) and a Finnish (Fi) cultivar, belonging respectively to Vicia faba major and V. faba minor groups, was described by 16S rRNA gene pyrosequencing and culture-dependent analysis. The sourdoughs were propagated with traditional backslopping procedure throughout 14days. Higher microbial diversity was found in the sourdough deriving from V. faba minor (Fi), still containing residual hulls after the milling procedure. After 2days of propagation, the microbial profile of Ita sourdough was characterized by the dominance of the genera Pediococcus, Leuconostoc and Weissella, while the genera Lactococcus, Lactobacillus and Escherichia, as well as Enterobacteriaceae were present in Fi sourdoughs. Yeasts were in very low cell density until the second backslopping and were not anymore found after this time by plate count or pyrosequencing analysis. Among the lactic acid bacteria isolates, Pediococcus pentosaceus, Leuconostoc mesenteroides and Weissella koreensis had the highest frequency of occurrence in both the sourdoughs. Lactobacillus sakei was the only lactobacillus isolated from the first to the last propagation day in Fi sourdough. According to microbiological and acidification properties, the maturity of the sourdoughs was reached after 5days. The presence of hulls and the different microbial composition reflected on biochemical characteristics of Fi sourdoughs, including acidification and phenolic compounds. Moreover, proteolysis in Fi sourdough was more intense compared to Ita. The microbial dynamic of the faba bean sourdoughs showed some differences with the most studied cereal sourdoughs.

Degradation of vicine, convicine and their aglycones during fermentation of faba bean flour

In spite of its positive repercussions on nutrition and environment, faba bean still remains an underutilized crop due to the presence of some undesired compounds. The pyrimidine glycosides vicine and convicine are precursors of the aglycones divicine and isouramil, the main factors of favism, a genetic condition which may lead to severe hemolysis after faba bean ingestion. The reduction of vicine and convicine has been targeted in several studies but little is known about their degradation. In this study, the hydrolysis kinetics of vicine and convicine and their derivatives during fermentation with L. plantarum DPPMAB24W was investigated. In particular, a specific HPLC method coupled to ESI-MS and MS/MS analysis, including the evaluation procedure of the results, was set up as the analytical approach to monitor the compounds. The degradation of the pyrimidine glycosides in the fermented flour was complete after 48 h of incubation and the aglycone derivatives could not be detected in any of the samples. The toxicity of the fermented faba bean was established through ex-vivo assays on human blood, confirming the experimental findings. Results indicate that mild and cost effective bioprocessing techniques can be applied to detoxify faba bean also for industrial applications.

Impact of Enzymatic and Microbial Bioprocessing on Protein Modification and Nutritional Properties of Wheat Bran.

Besides providing dietary fiber, wheat bran is a recognized source of protein and is considered a very valuable substitute for other protein-rich sources in the food and feed industry. Nonetheless, several factors affect protein bioavailability, including brans layered structure. This study showed the influence on the release and protein modification of wheat bran of different bioprocessing methods involving the activation of endogenous enzymes of bran, the addition of an enzyme mixture having carbohydrase activity, and microbial fermentation. Bioprocessing in acidic conditions significantly enhanced the solubilization of protein from wheat bran, reaching the highest value in the treatment where the sole endogenous protease activity was activated. Bioprocessing through controlled fermentation allowed a more intense proteolysis and strongly impacted the in vitro digestibility of proteins. The combined use of starter cultures and cell-wall-degrading enzymes was characterized by the highest increase of phytase activity and total phenols.

Exploring the Microbiota of Faba Bean: Functional Characterization of Lactic Acid Bacteria

This study investigated the metabolic traits of 27 lactic acid bacteria (LAB) strains belonging to different species, previously isolated from faba bean. The activities assayed, related to technological and nutritional improvement of fermented faba bean, included peptidases, β-glucosidase, phytase, as well as exopolysaccharides synthesis and antimicrobial properties. In addition, the bacteria performance as starter cultures during faba bean fermentation on proteolysis, antioxidant potential, and degradation of condensed tannins were assessed. Fermentative profiling showed that only 7 out of 27 strains were able to metabolize D-raffinose, particularly Leuc. mesenteroides I01 and I57. All strains of Pediococcus pentosaceus exerted high PepN activity and exhibited β-glucosidase activity higher than the median value of 0.015 U, while phytase activity was largely distributed among the different strains. All the weissellas, and in lower amount leuconostocs, showed ability to produce EPS from sucrose. None of the strains did not survive the simulated gastrointestinal tract with the exception of P. pentosaceus I56, I76, 147, I214, having a viability of 8–9 log CFU/ml at the end of the treatment. None of the strains showed antimicrobial activity toward Staphylococcus aureus, while eight strains of P. pentosaceus exhibited a strong inhibitory activity toward Escherichia coli and Listeria monocytogenes. Generally, the doughs fermented with pediococci exhibited high amount of total free amino acids, antioxidant activity, and condensed tannins degradation. These results allowed the identification of LAB biotypes as potential starter cultures for faba bean bioprocessing, aiming at the enhancement of faba bean use in novel food applications.