Sourdough production: fermentation strategies, microbial ecology, and use of non-flour ingredients.

Sourdough production is an ancient method to ferment flour from cereals for the manufacturing of baked goods. This review deals with the state-of-the-art of current fermentation strategies for sourdough production and the microbial ecology of mature sourdoughs, with a particular focus on the use of non-flour ingredients. Flour fermentation processes for sourdough production are typically carried out by heterogeneous communities of lactic acid bacteria and yeasts. Acetic acid bacteria may also occur, although their presence and role in sourdough production can be criticized. Based on the inoculum used, sourdough productions can be distinguished in fermentation processes using backslopping procedures, originating from a spontaneously fermented flour-water mixture (Type 1), starter culture-initiated fermentation processes (Type 2), and starter culture-initiated fermentation processes that are followed by backslopping (Type 3). In traditional recipes for the initiation and/or propagation of Type 1 sourdough productions, non-flour ingredients are often added to the flour-water mixture. These ingredients may be the source of an additional microbial inoculum and/or serve as (co-)substrates for fermentation. An example of the former is the addition of yoghurt; an example of the latter is the use of fruit juices. The survival of microorganisms transferred from the ingredients to the fermenting flour-water mixture depends on the competitiveness toward particular strains of the microbial species present under the harsh conditions of the sourdough ecosystem. Their survival and growth is also determined by the presence of the appropriate substrates, whether or not carried over by the ingredients added.

An in-depth multiphasic analysis of the chocolate production chain, from bean to bar, demonstrates the superiority of Saccharomyces cerevisiae over Hanseniaspora opuntiae as functional starter culture during cocoa fermentation.

Hanseniaspora opuntiae is a commonly found yeast species in naturally fermenting cocoa pulp-bean mass, which needed in-depth investigation. The present study aimed at examining effects of the cocoa isolate H. opuntiae IMDO 040108 as part of three different starter culture mixtures compared with spontaneous fermentation, regarding microbial community, substrate consumption, and metabolite production dynamics, including volatile organic compound (VOC) and phytochemical compositions, as well as compositions of the cocoa beans after fermentation, cocoa liquors, and chocolates. The inoculated H. opuntiae strain was unable to prevail over background yeasts present in the fermenting cocoa pulp-bean mass. It led to under-fermented cocoa beans after four days of fermentation, which was however reflected in higher levels of polyphenols. Cocoa fermentation processes inoculated with a Saccharomyces cerevisiae strain enhanced flavour production during the fermentation and drying steps, which was reflected in richer and more reproducible aroma profiles of the cocoa liquors and chocolates. Sensory analysis of the cocoa liquors and chocolates further demonstrated that S. cerevisiae led to more acidic notes compared to spontaneous fermentation, as a result of an advanced fermentation degree. Finally, different VOC profiles were found in the cocoa beans throughout the whole chocolate production chain, depending on the fermentation process.

Lemon juice and apple juice used as source of citrate and malate, respectively, enhance the formation of buttery aroma compounds and/or organic acids during Type 2 and Type 3 sourdough productions performed with Companilactobacillus crustorum LMG 23699.

Extra ingredients are often used in traditional sourdough production recipes by artisan bakeries. These ingredients may be the source of microorganisms or stimulate the growth and/or the metabolic activities of the microorganisms added to or naturally present in the flour-water mixture. The present study examined the influence of the addition of lemon juice or apple juice as source of citrate or malate, respectively, on the growth and activity of the citrate- and malate-positive Companilactobacillus crustorum LMG 23699 strain (formerly known as Lactobacillus crustorum LMG 23699), used to initiate firm (dough yield of 200) wheat sourdough productions, and on the flavour of the baked goods produced. Three fermentation strategies were applied, namely one-step long fermentation sourdough production processes with the addition of juice at the start (Type 2) and backslopped fermentations with the addition of juice either only at the start of the sourdough productions or at the start of the sourdough productions and at the beginning of each subsequent refreshment step during the whole backslopping process (both Type 3). It turned out that the starter culture strain used prevailed during all sourdough productions performed. Yeasts were particularly present in Type 3 sourdough productions, although lemon juice retarded their growth. Due to high yeast activity, high concentrations of ethanol and glycerol were produced toward the end of the sourdough productions. Addition of lemon juice stimulated the production of lactic acid, acetic acid, and the buttery flavour compounds acetoin and diacetyl, because of citrate conversion, during the Type 2 and Type 3 sourdough productions. In Type 3 sourdough productions, these compounds were found in higher concentrations only when lemon juice was added at each backslopping step. Alternatively, the addition of apple juice led to high concentrations of lactic acid because of malolactic fermentation in both Type 2 and Type 3 sourdough productions. Moreover, the addition of apple juice increased the initial concentrations of the carbohydrates (fructose, glucose, and sucrose) and sugar alcohols (mannitol and sorbitol), which were exhausted upon backslopping or accumulated in the sourdough matrix, respectively. Baked goods produced using sourdoughs obtained from the Type 2 and Type 3 sourdough productions with the addition of juice at each backslopping step were significantly different in flavour from doughs supplemented with the respective juices and lactic acid and/or Type 3 sourdough productions with the addition of juice only at the start.

Potential of Bacteria from Alternative Fermented Foods as Starter Cultures for the Production of Wheat Sourdoughs.

Microbial strains for starter culture-initiated sourdough productions are commonly isolated from a fermenting flour-water mixture. Yet, starter culture strains isolated from matrices other than sourdoughs could provide the dough with interesting metabolic properties and hence change the organoleptic properties of the concomitant breads. Furthermore, the selection of sourdough starter cultures does not need to be limited to lactic acid bacteria (LAB), as other food-grade microorganisms are sometimes found in sourdoughs. Therefore, different strains belonging to LAB, acetic acid bacteria (AAB), and coagulase-negative staphylococci (CNS) that originated from different fermented food matrices (fermenting cocoa pulp-bean mass, fermented sausage, and water kefir), were examined as to their prevalence in a wheat sourdough ecosystem during 72-h fermentations. Limosilactobacillus fermentum IMDO 222 (fermented cocoa pulp-bean mass isolate) and Latilactobacillus sakei CTC 494 (fermented sausage isolate) seemed to be promising candidates as sourdough starter culture strains, as were the AAB strains Acetobacter pasteurianus IMDO 386B and Gluconobacter oxydans IMDO A845 (both isolated from fermented cocoa pulp-bean mass), due to their competitiveness in the wheat flour-water mixtures. Wheat breads made with G. oxydans IMDO A845 sourdoughs were significantly darker than reference wheat breads.

Diverse Microbial Composition of Sourdoughs From Different Origins.

Hundreds of sourdoughs have been investigated in the last decades. However, many studies used a culture-dependent and/or culture-independent microbiological approach [mainly based on denaturing gradient gel electrophoresis (DGGE) of PCR amplicons], seldomly combined with a metabolite target analysis, to characterize the microbial species communities of the sourdoughs examined. Moreover, attention was mainly paid on lactic acid bacteria (LAB) and yeast species. In the present study, distinct household-scale (including an artisan lambic brewery) and artisan bakery-scale backslopped sourdoughs (17 in total), obtained from different regions (Belgium, France, United Kingdom, and USA), were examined through a multiphasic approach, encompassing a culture-dependent analysis [targeting LAB, acetic acid bacteria (AAB), and yeasts], different culture-independent techniques [rRNA-PCR-DGGE, metagenetics, and metagenomics (four bakery sourdoughs)], and metabolite target analysis. It turned out that the microbial species diversity of the sourdoughs was influenced by the house microbiota of the producer. Further, when the producer made use of different flours, the sourdoughs harbored similar microbial communities, independent of the flour used. AAB were only present in the Belgian sourdoughs, which might again be related to the processing environment. Fructilactobacillus sanfranciscensis (formerly known as Lactobacillus sanfranciscensis) was the prevalent LAB species of the eight sourdoughs produced by two of the three bakeries of different countries analyzed. These sourdoughs were characterized by the presence of either Saccharomyces cerevisiae or Kazachstania humilis. Moreover, the presence of Fl. sanfranciscensis was positively correlated with the production of mannitol and negatively correlated with the presence of other LAB or AAB species. Sourdoughs produced in an artisan lambic brewery were characterized by the presence of the yeast species Dekkera anomala and Pichia membranifaciens. One household sourdough was characterized by the presence of uncommon species, such as Pediococcus parvulus and Pichia fermentans. Metagenomic sequencing allowed the detection of many more LAB and AAB species than the other methods applied, which opened new frontiers for the understanding of the microbial communities involved during sourdough production processes.

Curing of Cocoa Beans: Fine-Scale Monitoring of the Starter Cultures Applied and Metabolomics of the Fermentation and Drying Steps.

Starter culture-initiated cocoa fermentation processes can be applied to improve the quality of cured cocoa beans. However, an accurate monitoring of the microbial strains inoculated in fresh cocoa pulp-bean mass to assess their contribution to the cocoa bean curing process is still lacking. In the present study, eight different cocoa fermentation processes were carried out with Trinitario cocoa in vessels in Costa Rica to assess the contribution of two candidate yeast starter culture strains, namely Saccharomyces cerevisiae IMDO 050523 and Pichia kudriavzevii IMDO 020508, inoculated in combination with Limosilactobacillus fermentum IMDO 0611222 and Acetobacter pasteurianus IMDO 0506386. A multiphasic approach, consisting of culture-dependent selective plating and incubation, rRNA-PCR-DGGE community profiling of agar plate washes, and culture-independent high-throughput amplicon sequencing, combined with a metabolite target analysis of non-volatile and volatile organic compounds (VOCs), was performed on samples from the fermentation and/or drying steps. The different starter culture mixtures applied effectively steered the cocoa fermentation processes performed. Moreover, the use of an amplicon sequence variant (ASV) approach, aligning these ASVs to the whole-genome sequences of the inoculated strains, allowed the monitoring of these inoculated strains and their differentiation from very closely related variants naturally present in the background or spontaneous fermentation processes. Further, traits such as malolactic fermentation during the fermentation step and acetoin and tetramethylpyrazine formation during the drying step could be unraveled. Finally, the yeast strains inoculated influenced the substrate consumption and metabolite production during all starter culture-initiated fermentation processes. This had an impact on the VOC profiles of the cured cocoa beans. Whereas the P. kudriavzevii strain produced a wide range of VOCs in the cocoa pulp, the S. cerevisiae strain mostly influenced the VOC composition of the cured cocoa beans.

The addition of citrate stimulates the production of acetoin and diacetyl by a citrate-positive Lactobacillus crustorum strain during wheat sourdough fermentation.

In traditional sourdough fermentation recipes of artisan bakeries, often extra ingredients are added to the flour-water mixture. This may accentuate the aroma and taste of the baked products produced from such sourdoughs. This is made possible, for instance, by stimulating certain microbial activities during fermentation. This study examined the effects of the addition of citrate (a food-grade organic acid present in milk and lemons) on wheat sourdough fermentation in the presence of a citrate-positive, homofermentative, lactic acid bacterial starter culture strain, namely Lactobacillus crustorum LMG 23699. Both liquid and firm wheat sourdoughs were produced. The starter culture strain was able to steer all wheat sourdough fermentations performed, as it always prevailed due to its competitiveness, as shown through culture-dependent microbiological plating and culture-independent bacterial community profiling. Moreover, it possessed all enzyme-encoding genes (as unraveled through genome mining) necessary to convert citrate into desirable compounds such as lactic acid, acetic acid, succinate, acetoin, diacetyl, and 2,3-butanediol. Indeed, citrate addition to the wheat flour-water mixture had an impact on the sourdough fermentation dynamics and thus on the aroma profile of the liquid and firm sourdoughs produced and breads made thereof. A higher final pH, higher total titratable acidity values, and low yeast counts were found in wheat sourdoughs produced with citrate. In particular, the starter culture strain added converted the supplemented citrate into more l-lactic acid as well as acetoin and diacetyl (buttery aroma compounds), which was independent of the dough yield. The buttery aroma compounds were also accentuated in the concomitant breads produced. Further, organic acid production was stimulated in the sourdoughs, whereas increased pyrazine concentrations occurred in the breads. Consequently, citrate supplementation to wheat sourdoughs could be of interest to produce baked goods with enhanced buttery aroma compounds and notes.

Impact of starter culture, ingredients, and flour type on sourdough bread volatiles as monitored by selected ion flow tube-mass spectrometry.

This study deals with the detection of volatile compounds originating from the crumb of breads made with sourdoughs obtained through starter culture-initiated fermentations, which differed in flour type (wheat and teff), ingredients (citrate and malate), fermentation time (24h or 72h), and starter culture strains (homo- and heterofermentative lactic acid bacteria species and acetic acid bacteria species) applied. Therefore, selected ion flow tube-mass spectrometry (SIFT-MS) was used. SIFT-MS is an easy-to-use and promising technique in the field of food and flavor analysis. Volatile compounds of crumb samples from the breads with sourdough were measured and compared with those of reference bread crumb samples. In general, sourdough addition had a positive effect on the concentrations of the volatile compounds measured by SIFT-MS. Furthermore, a trend toward higher concentrations of several volatiles was seen upon the addition of sourdoughs that were fermented up to 72h, compared to the addition of sourdoughs that were fermented for a shorter time. Ethanol was the major volatile compound identified tentatively, next to alcohols, aldehydes, esters, terpenes, and heterocyclic compounds. Also acetoin/ethyl acetate could be identified, but these compounds could not be distinguished. Higher alcohols showed an increase upon the use of sourdough fermented for a long time. High concentrations of acetic acid were found in breads made with Gluconobacter oxydans IMDO A845-initiated sourdough, indicating its potential for sourdough production. Breads made with teff sourdoughs were distinct from wheat-based sourdough breads as to their volatile compound profiles.

Monitoring of starter culture-initiated liquid wheat and teff sourdough fermentations by selected ion flow tube-mass spectrometry.

BACKGROUND: Selected ion flow tube-mass spectrometry (SIFT-MS) is a direct-injection mass spectrometric technique that has been introduced recently into the field of food and flavor analysis. It also shows potential for use in the monitoring of food fermentations. Therefore, this study aimed at the online monitoring of different volatile compounds produced during starter culture-initiated liquid sourdough fermentations by SIFT-MS, for which a new workflow was developed. RESULTS: The online monitoring of the volatile sample compounds acetoin and ethyl acetate, diacetyl, and ethanol was made possible during the production of sourdoughs obtained through fermentation with several interesting strains belonging to the species Lactobacillus crustorum, Lactobacillus fermentum, Lactobacillus hilgardii, Lactobacillus nagelii, Lactobacillus sakei, and Gluconobacter oxydans. Acetoin and ethyl acetate could not be distinguished based solely on SIFT-MS data. Diacetyl production was monitored in the case of Lb. crustorum LMG 23699 as a starter culture strain, thereby making the distinction between those volatiles produced in sourdough without extra ingredients added or after the addition of citrate or malate. CONCLUSION: Starter culture-initiated liquid sourdough fermentations were monitored successfully. The volatile compound production of the different starter culture strains tested reflected differences in their metabolism and/or competitiveness in a sourdough matrix. © 2018 Society of Chemical Industry.

Candida milleri species reveals intraspecific genetic and metabolic polymorphisms.

Candida milleri, together with Candida humilis, is the most representative yeast species found in type I sourdough ecosystems. In this work, comparison of the ITS region and the D1/D2 domain of 26S rDNA gene partial sequences, karyotyping, mtDNA-RFLP analysis, Intron Splice Site dispersion (ISS-PCR) and (GTG)5 microsatellite analyses, assimilation test of different carbohydrates, and metabolome assessment by FT-IR analysis, were investigated in seventeen strains isolated from four different companies as well as in type strains CBS6897(T) and CBS5658(T). Most isolates were ascribed to C. milleri, even if a strong relatedness was confirmed with C. humilis as well, particularly for three strains. Genetic characterization showed a high degree of intraspecific polymorphism since 12 different genotypes were discriminated. The number of chromosomes varied from 9 to 13 and their size ranged from less than 0.3 to over 2 Mbp. Phenotypic traits let to recognize 9 different profiles of carbon sources assimilation. FT-IR spectra from yeast cells cultivated in different media and collected at different growth phases revealed a diversity of behaviour among strains in accordance with the results of PCR-based fingerprinting. A clear evidence of the polymorphic status of C. milleri species is provided thus representing an important feature for the development of technological applications in bakery industries.