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Several chemical compounds of seven faba bean cultivars—Sampo, Louhi, Talia, Tiffany, Julia, Gloria, and Vroma—grown in Finland were determined. The effect of seed maturity on the levels of vicine/convicine (vc), trypsin inhibitors, condensed tannins, phytic acid, lectins, and oligosaccharides was analyzed. The overall vc ratio remained similar in all the samples, and the largest amounts of vc were found in the early stage of the bean and decreased during maturation. Talia and Gloria had lower trypsin inhibition than the other cultivars. The smallest amounts of condensed tannins were found in the Talia and Gloria cultivars, and the largest in the Tiffany and Julia cultivars. Overall, it seems that maturation decreased the amount of condensed tannins. The fully mature faba bean contained an average of 10.3 to 13.4 g/kg (DW) phytic acid, and there was no difference between the seven cultivars. The highest lectin activity was found in the Sampo, Julia, and Tiffany cultivars, and the lowest activity was in Vroma and the mature seeds of Talia and Louhi. The simple sugars (rhamnose, galactose, glucose, sucrose, and fructose) were dominant in the early developmental stage, whereas the amount of long chain oligosaccharides (stachyose and verbascose) increased during maturation of beans, and the level of raffinose was similar in all developmental stages. In conclusion, the content of chemical compounds depended on the maturation stage and the faba bean cultivar. The low content of stachyose, verbascose, and raffinose in young Tiffany and Julia beans is beneficial when consumed as green bean.

To enable the utilization of seasonal biomasses in e.g., farm-scale biogas plants, the process should be flexible and ensure stable gas production. However, information about microbial community dynamics in long-term co-digestion with versatile co-feedstocks is lacking. This study investigated the effects of co-feedstock changes on the performance and evolution of microbial consortia during 428-day anaerobic digestion of cow slurry. Co-feedstocks consisted of hydrocarbon-, protein- and lipid-rich materials. A high throughput 16S ribosomal RNA gene sequencing was used to analyze the taxonomic profile of microbial communities. Due to the low loading rate, the changes were subtle in bacteria, but a shift on archaeal genera in response to different and changing feedstock compositions was observed. Despite drastic changes in co-feedstock composition, stable and flexible anaerobic digestion with relatively constant core microbiome can be achieved with cautious operation of the process.

Consumers worldwide are increasingly interested in the authenticity and naturalness of products. At the same time, the food, agricultural and forest industries generate large quantities of sidestreams that are not effectively utilized. However, these raw materials are rich and inexpensive sources of bioactive compounds such as polyphenols. The exploitation of these raw materials increases income for producers and processors, while reducing transportation and waste management costs. Many Northern sidestreams and other underutilized raw materials are good sources of polyphenols. These include berry, apple, vegetable, softwood, and rapeseed sidestreams, as well as underutilized algae species. Berry sidestreams are especially good sources of various phenolic compounds. This chapter presents the properties of these raw materials, providing an overview of the techniques for refining these materials into functional polyphenol-rich ingredients. The focus is on economically and environmentally sound technologies suitable for the pre-treatment of the raw materials, the modification and recovery of the polyphenols, as well as the formulation and stabilization of the ingredients. For example, sprouting, fermentation, and enzyme technologies, as well as various traditional and novel extraction methods are discussed. Regarding the extraction technologies, this chapter focuses on safe and green technologies that do not use organic solvents. In addition, formulation and stabilization that aim to protect isolated polyphenols during storage and extend shelflife are reviewed. The formulated polyphenol-rich ingredients produced from underutilized renewable resources could be used as sustainable, active ingredients--for example, in food and nutraceutical industries.

Consumers worldwide are increasingly interested in the authenticity and naturalness of products. At the same time, the food, agricultural and forest industries generate large quantities of sidestreams that are not effectively utilized. However, these raw materials are rich and inexpensive sources of bioactive compounds such as polyphenols. The exploitation of these raw materials increases income for producers and processors, while reducing transportation and waste management costs. Many Northern sidestreams and other underutilized raw materials are good sources of polyphenols. These include berry, apple, vegetable, softwood, and rapeseed sidestreams, as well as underutilized algae species. Berry sidestreams are especially good sources of various phenolic compounds. This chapter presents the properties of these raw materials, providing an overview of the techniques for refining these materials into functional polyphenol-rich ingredients. The focus is on economically and environmentally sound technologies suitable for the pre-treatment of the raw materials, the modification and recovery of the polyphenols, as well as the formulation and stabilization of the ingredients. For example, sprouting, fermentation, and enzyme technologies, as well as various traditional and novel extraction methods are discussed. Regarding the extraction technologies, this chapter focuses on safe and green technologies that do not use organic solvents. In addition, formulation and stabilization that aim to protect isolated polyphenols during storage and extend shelflife are reviewed. The formulated polyphenol-rich ingredients produced from underutilized renewable resources could be used as sustainable, active ingredients--for example, in food and nutraceutical industries.

Lupine (Lupinus sp.) is a valuable source of plant proteins. There is little knowledge on the impact of food processing on composition and sensory properties of lupine products. In this research, we investigated the impact of fermentation with five starters of lactic acid bacteria on the sensory quality and flavor-active compounds in dairy analogues prepared from sweet lupine (Lupinus angustifolius L.). The sensory qualities of unfermented and fermented products were studied with generic descriptive analysis and affective tests. Acids and sugars were analyzed with GC-FID and volatiles with HS-SPME-GC-MS and GC-O. Fermentation increased sourness and 'vinegar' odor and reduced the 'beany' odor and flavor as well as the unpleasantness of flavor. Formation of volatiles during the fermentation was dependent on the starters. However, all fermentations increased the contents of lactic, acetic, and hexanoic acids, while reducing the contents of hexanal, described as 'grassy' in the unfermented lupine sample.

Faba bean (Vicia faba L.) is a grain legume that provides possibilities to increase local protein self-sufficiency both as feed and food in areas where possibilities for cultivation of other protein crops are limited. However, full ripening may not be achieved and storage of the moist beans by crimping and ensiling provides an economic alternative to drying, and is well suited for animal feeding. The objectives of the current experiment were to evaluate the effects of moisture content and various silage additives on the fermentation quality, aerobic stability and fate of bioactive compounds of ensiled crimped faba beans in a pilot scale ensiling experiment. Faba beans were combine harvested at moisture level of 443 g/kg and crimped prior to ensiling. The following additive treatments were applied using three replicates: 1) control (CON), 2) two commercial lactic acid bacteria inoculants as mixture (LABmix), 3) In-house LAB inoculant (LABLuke), 4) Formic acid based product (FA) and 5) Salt based product (Salt). Further, two of the additive treatments were prepared with additional moisture: 6) Control with tap water addition (CON + w) and 7) LABmix with tap water addition (LABmix + w). The beans ensiled without additive showed elevated pH, butyric acid and ethanol concentrations and proportion of ammonia N in total N. Addition of water further decreased the fermentation quality. The LAB inoculants improved the fermentation quality of the beans compared to CON. The fermentation was effectively restricted when FA was used. Both LAB and chemical additives improved aerobic stability compared to CON and Salt was especially effective in improving aerobic stability. Vicine disappeared totally during the ensiling period and also convicine was effectively degraded. It seems that promoting microbial activity in the silo either by LAB inoculation or increased moisture promoted the degradation of convicine, while chemical additives decreased the degradation of it. Total and particularly soluble proanthocyanidins were also degraded during the ensiling period. Crimping and ensiling is an alternative preservation method for faba beans which eliminates the energy consumption and investments required for drying. In addition, benefits regarding the concentrations of bioactive compounds may be expected.

Propionic acid bacteria (PAB) are important for the manufacture of Swiss-type cheeses since they are involved in the formation of eyes and are responsible for the characteristic flavour and aroma of these cheeses. Propionibacterium freudenreichii is most commonly encountered in Swiss-type cheeses and is the sole Propionibacterium species used as a cheese starter. Aspartase activity determined in PAB is strain-dependent and it is important to find a usable way to identify the specific strains involved in cheese making. In this work, 16S-amplified ribosomal DNA restriction analysis and pulsed-field gel electrophoresis (PFGE) were used to differentiate 107 PAB strains. Consequently, the presence and persistence of inoculated Propionibacterium starter strains can be monitored during cheese ripening. The studied metabolic features of lactose utilisation, nitrate reductase activity and aspartase activity could not be directly associated with particular genotypes, but principal component analysis (PCA) suggested a linkage between related PFGE patterns and aspartase activity.

This study focused on defining the differences in L. plantarum gene expression levels in different media and in different growth phases using an easy and cost-efficient monitoring of gene expression. A macroarray based on a group of selected L. plantarum genes, 178 genes belonging to 18 main groups, printed onto a nitrocellulose filter was designed in this work. Using the macrofilters designed, the expression of a selected set of L. plantarum genes was assayed in synthetic MRS medium and in extracted carrot juice. To compare the potential differences of starter gene expression in hygienic and contaminated cultivation media, the L. plantarum strain was cultivated in both sterile and contaminated (yeast and Escherichia coli) MRS and carrot juice. The number of genes found to be regulated as a function of growth was clearly higher in MRS-based growth medium than in carrot juice, In carrot juice, expression of the gene encoding malolactic enzyme (MLE), which makes L. plantarum an advantageous microbe in e.g. wine making, was found to be upregulated in logarithmic phase of growth. The current study demonstrated that macroarrays printed on nitrocellulose filters with simple robotic systems can be analyzed by standard laboratory equipment and methods usually available in molecular laboratories. Using this technology, rapid and cost-efficient analysis of genome function of L. plantarum can be carried out e.g. in developing regions, where lactic acid fermentation of food and feed matrices is a common practice.

The ruminal microbiome, comprising large numbers of bacteria, ciliate protozoa, archaea and fungi, responds to diet and dietary additives in a complex way. The aim of this study was to investigate the benefits of increasing the depth of the community analysis in describing and explaining responses to dietary changes. Quantitative PCR, ssu rRNA amplicon based taxa composition, diversity and co-occurrence network analyses were applied to ruminal digesta samples obtained from four multiparous Nordic Red dairy cows fitted with rumen cannulae. The cows received diets with forage:concentrate ratio either 35:65 (diet H) or 65:35 (L), supplemented or not with sunflower oil (SO) (0 or 50 g/kg diet dry matter), supplied in a 4 × 4 Latin square design with a 2 × 2 factorial arrangement of treatments and four 35-day periods. Digesta samples were collected on days 22 and 24 and combined. QPCR provided a broad picture in which a large fall in the abundance of fungi was seen with SO in the H but not the L diet. Amplicon sequencing showed higher community diversity indices in L as compared to H diets and revealed diet specific taxa abundance changes, highlighting large differences in protozoal and fungal composition. Methanobrevibacter ruminantium and Mbb. gottschalkii dominated archaeal communities, and their abundance correlated negatively with each other. Co-occurrence network analysis provided evidence that no microbial domain played a more central role in network formation, that some minor-abundance taxa were at nodes of highest centrality, and that microbial interactions were diet specific. Networks added new dimensions to our understanding of the diet effect on rumen microbial community interactions.

The preparation of sour milk products has been known in all parts of the world as a means of preserving milk against spoilage. Traditional sour milk foods were obtained by spontaneous acidi…cation with lactic acid bacteria (LAB) naturally present in milk, which during their growth reduce the pH and denature milk proteins, causing appearance and texture changes in the product. In certain cases, yeasts and molds, which occur along with LAB, give a special feature to the sour milk product. Due to the speci…c characteristics of milk and LAB and the conditions of fermentation, various types of regionally speci…c fermented milk have been developed throughout the world.

This study characterized the bacteria causing decay of carrots during storage and marketing. Spoilage strains were identified by 16S-amplified rDNA restriction analysis and intergenic transcribed spacer-PCR-restriction fragment length polymorphism (ITS-PCR-RFLP). Genotypic fingerprinting by RFLP-pulsed-field gel electrophoresis was used to assess the genetic diversity of the isolates. A total of 252 Pseudomonas isolates from carrots were identified and classified into eight separate groups. Most strains belonged to group A (Pseudomonas fluorescens, Pseudomonas marginalis, and Pseudomonas veronii) and group B (Pseudomonas putida). The strains identified as Pectobacterium carotovorum subsp. carotovorum, Pectobacterium atrosepticum, Dickeya chrysanthemi, and Erwinia rhapontici were distinguished by ITS-PCR-RFLP. All isolates belonging to the genera Pectobacterium and Erwinia were responsible for carrot spoilage. This work has led to the development of new strategies for the identification and genotyping of vegetable-spoiling strains of Pseudomonas, Pectobacterium, and Erwinia. This is also the first report describing the occurrence of carrot-spoiling E. rhapontici. Early recognition of spoilage bacteria in vegetables is important for the implementation of effective handling strategies. Pectolytic bacteria may cause considerable financial losses because they account for a large proportion of bacterial rot of fruits and vegetables during storage, transit, and marketing.

Propionic acid bacteria (PAB) are important as starter cultures for the dairy industry in the manufacture of Swiss-type cheeses, in which they are involved in the formation of eyes and are responsible for the typical flavour and aroma. These characteristics are mainly due to the classical propionic acid fermentation, but also the conversion of aspartate to fumarate and ammonia by the enzyme aspartase and the subsequent reduction of fumarate to succinate, which occur in dairy Propionibacterium freudenreichii ssp. shermanii and ssp. freudenreichii starter strains. Additionally, the metabolism of free amino acids may be partly responsible for secondary fermentation and the subsequent split defects in cheese matrix. Here a method for aspartase activity was established and a number of dairy propionibacteria belonging to P. freudenreichii ssp. shermanii and freudenreichii were screened for this enzyme activity. A wide range of aspartase activity could be found in PAB isolates originating from cheese. The majority, i.e. 70% of the 100 isolates tested, showed very low levels of aspartate activity.

Aims of the study were to characterize two Lactobacillus plantarum-related strains, Lact. plantarum and Lactobacillus paraplantarum isolated from fermented vegetables and, for their potential use as starter strains, compare their growth in various food matrices. Species-level identification of the strains belonging to the Lact. plantarum group was performed by multiplex-PCR with species-specific primers and generation of distinct genotypic profiles was carried out by PFGE-based DNA-fingerprinting. Growth profiles were determined in various food and feed matrices. Compared to Lact. plantarum, Lact. paraplantarum reached higher cell densities in all plant-based matrices and MRS broth. On the contrary to the good growth in plant-based matrices and MRS, poor growth was observed in unprocessed milk. Supplemented lactose did not improve the growth of either tested strain, while predigestion of milk proteins with Lactobacillus helveticus or addition of casitone proved to be an effective means to enhance growth. To find out the applicability of molecular methods, the strains were transformed with replicative plasmids by electroporation. To our knowledge, this is a first report of the electrotransformation of Lact. paraplantarum with a recombinant plasmid.

Development of molecular methods has enabled detailed studies of the proteolytic system of Lactobacillus helveticus, which has a central role in the release of bioactive peptides during the fermentation of milk. The impact of general aminopeptidase (PepN) and X-prolyl dipeptidyl aminopeptidase (PepX) activity of L. helveticus on the level of angiotensin I-converting enzyme (ACE) inhibitory activity in fermented milk was elucidated by taking advantage of peptidase-negative derivatives of L. helveticus CNRZ32. According to the results, increased level of ACE-inhibitory activity was attained in milk fermented by the peptidase-deficient mutants. The ACE-inhibitory activity determined with the PepN-deficient strain decreased towards the end of fermentation, suggesting that PepN gives rise to a provisional accumulation of ACE-inhibitory peptides before their hydrolysis to shorter peptides or free amino acids. The ACE-inhibitory activity determined with the PepX-deficient strain increased throughout the fermentation, indicating specific blocking of the further hydrolysis of ACE-inhibitory peptides.

Aims: Phenotypic and molecular methods were used to identify and compare the strain composition of three industrial dairy starters used for the manufacture of viili. Methods and Results: Preliminary differentiation was made by phenotypic methods. Genotypic differentiation was carried out using polymerase chain reaction (PCR) and further characterization at strain level by pulsed-field gel electrophoresis (PFGE). The isolates could be assigned as acid-producing Lactococcus lactis strains of both lactis and cremoris subspecies, and aroma producers, identified as L. lactis subsp. lactis biovar diacetylactis and Leuconostoc mesenteroides. PCR analysis discriminated between the lactococcal subspecies, and cluster analysis of the digestion patterns of PFGE analysis revealed different genotypes in each subspecies. Each Leuconostoc-genotype seemed to be specific to only a single starter mix. Conclusions: The work proved that in addition to L. lactis subsp. lactis biovar diacetylactis and Leuc. mesenteroides subsp. cremoris, commercial viili starters of traditional origin may contain (i) only L. lactis subsp. cremoris, (ii) both L. lactis subsp. cremoris and L. lactis subsp. lactis as a minority, and – as a new discovery – (iii) only L. lactis subsp. lactis. Significance and Impact of the Study: The results obtained give an overview of the microbial population of viili starters and can be exploited in the development of optimized starter cultures for industrial-scale manufacture of viili.

Lactobacillus brevis possesses a surface layer protein (SlpA) with tightly regulated synthesis. The slpA gene is expressed by two adjacent promoters, P1 and P2. The level of P2-derived transcripts was approximately 10 times higher than that of P1-derived transcripts throughout the entire growth of L. brevis. The half-lives of slpA transcripts were shown to be exceptionally long (14 min).

A cassette based on the expression signals of the Lactobacillus brevis surface (S)-layer protein gene (slpA) was constructed. The low-copy-number vector pKTH2095, derived from pGK12, was used as the cloning vector. The efficiency of slpA promoters in intracellular protein production was studied using three reporter genes, beta-glucuronidase (gusA), luciferase (luc) and aminopeptidase N (pepN) in three different lactic acid bacteria hosts: Lactococcus lactis, Lactobacillus plantarum and Lactobacillus gasseri. The S-layer promoters were recognized in each strain and especially L. lactis and Lb. plantarum exhibited high levels of transcripts. The production kinetics of reporter proteins was studied as a function of growth. The GusA, Luc and PepN activities varied considerably among the lactic acid bacterial strains studied. The highest levels of beta-glucuronidase and luciferase activity were obtained in L. lactis. The level of GusA obtained in L. lactis corresponded to over 15% of the total cellular proteins. The highest level of aminopeptidase N activity was achieved in Lb. plantarum where PepN corresponded up to 28% of the total cellular proteins at the late exponential phase of growth. This level of PepN activity is 30-fold higher than that in Lb. helveticus, which is the species from which the pepN gene originates.

The aim of this study was to examine the enzymic hydrolysis of αs2-casein by isolating and identifying the released peptides. The enzymes applied in the study were chymosin, plasmin and trypsin as well as cell free extracts from three strains of Lactobacillus helveticus and nine strains of L. casei. The findings showed that chymosin had weak proteolytic activity on αs2-casein. Plasmin, on the other hand, released numerous peptides under the used conditions. The majority of the identified fragments were released from the C terminal end of the substrate. Plasmin hydrolysed mainly Lys-X bonds. The third enzyme, trypsin, hydrolysed several bonds of αs2-casein. Peptides were released from almost all regions of the protein. Trypsin acted on the carboxyl sides of arginyl and lysyl residues. Cell free extract of lactobacilli had little activity on αs2-casein.

This study was conducted to investigate the rate of proteolysis and peptide profiles of different Finnish fermented milk products. The highest rate of proteolysis was observed in Biokefir, while the greatest change in the rate of proteolysis was observed in Gefilus®. Differences in starters and manufacturing processes reflected on the peptide profiles of the products. Most of the identified peptides originated from either the N- or C-terminal region of β-casein or from the N-terminal region of αs1-casein., Suomalaisten hapanmaitotuotteiden valkuaisaineiden pilkkoutumista tutkittiin varastoinnin aikana. Tuotteista määritettiin proteolyysiaste sekä eristettiin ja identifioitiin peptidejä peptidikartalta. Suurin proteolyysiaste todettiin biokefiirissä, kun taas suurin proteolyysiasteen muutos todettiin gefiluksessa. Erot käytetyissä hapatteissa ja valmistusmenetelmissä kuvastuivat tuotteiden peptidikartoissa. Useimmat tunnistetuista peptideistä olivat peräisin β-kaseiinin N- tai C-terminaalisesta päästä tai αs1-kaseiinin N-terminaalisesta päästä.

Vegetable by-products could potentially be suitable to be used as feeds in animal diets, which in general would strengthen the sustainability of both food and feed systems. Two experiments were performed with the objective to evaluate the aerobic spoilage via visual inspection and short term ensiling process of carrot by-products with silage additives (lactic acid bacteria [LAB] inoculants or formic acid [FA]). In the first experiment (Exp. 1), carrot by-product was treated with five additives including a control without additive, two commercial LAB inoculants (heterofermentative and homofermentative), an in-house isolated LAB mixture and a commercial FA based product. Second experiment (Exp. 2) used a 2 × 3 factorial design (type of raw material [carrot by-product with or without pieces] and additive treatment [control, heterofermentative LAB inoculant or FA based product]). Use of FA restricted lactic acid fermentation indicated by the high residual water soluble carbohydrate concentration while simultaneously stimulating ethanol production. LAB inoculants were not able to efficiently affect quality of ensiled carrot by-products. Use of FA also delayed the spoilage of the fresh carrot by-product particularly in Exp. 1.