Fermentation, Volume 3, Issue 4 (December 2017) – 21 articles

Surplus hops (Humulus lupulus L.) that are not needed by the brewing industry could be used as a feed supplement for cattle and other ruminants. Previous research indicates that antimicrobial hops plant secondary metabolites (i.e., α- and β-acids) inhibit methane and ammonia production and promote the growth of ruminant animals. The goal was to determine that hop pellets produced for brewing still possessed the requisite antimicrobial activity after 5-year storage. HPLC (high performance liquid chromatography) analysis indicated that the α- and β-acid concentrations in two varieties of hops were relatively stable after 5 years of storage under N₂. Either hop variety inhibited the growth of the ruminal hyper ammonia-producing bacterium, Clostridium sticklandii SR, in broth culture and Petri plate bioassays. Either hop variety inhibited ammonia production from amino acids or peptides by mixed rumen microorganisms from Holstein steers. These results are similar to those previously obtained with fresh hops, hops extracts, other antimicrobial phytochemicals and typical feed ionophores, such as monensin. The rumen-active antimicrobial phytochemicals in hops can still be present and active after years under certain storage conditions. Further investigation is warranted to determine how surplus and older hops can be used to benefit ruminant nutrition and ruminant industries. Full article

Probiotics are live microorganisms that, when administered in adequate amounts, confer a health benefit on the host, mainly through the process of replacing or including beneficial bacteria in the gastrointestinal tract. Fermented dairy foods such as yogurt, fermented milk and cheese are the major vehicle in delivering probiotics, and probiotic delivery have been traditionally associated with these fermented dairy foods. Additionally, many other non-dairy probiotic products and non-food form such as capsules, pills and tablets are also available and some of these non-food forms are highly popular among the consumers. Certain non-dairy probiotic foods, especially beverages that are non-fermented products, can also play an important role in probiotic delivery. There is an increasing demand for non-dairy probiotic foods (both fermented and non-fermented) including fruit and vegetable juices, soy and certain cereal products due to vegetarianism, lactose intolerance and dairy allergies, as well as interest in low cholesterol foods. In this context, this review mainly focus on the different types of probiotic food products including beverages with special reference to their viability followed by a brief account on the applicability of using fermented and non-fermented beverage products in probiotic delivery. Full article

In this study, an innovative green biorefinery system was successfully developed to process the green biomass into multiple biofuels and bioproducts. In particular, fresh giant miscanthus was separated into a solid stream (press cake) and a liquid stream (press juice) using a screw press. The juice was used to cultivate microalga Chlorella vulgaris, which was further thermochemically converted via thermogravimetry analysis (TGA) and pyrolysis-gas chromatography/mass spectrometry (Py-GC/MS) analysis, resulting in an approximately 80% conversion. In addition, the solid cake of miscanthus was pretreated with dilute sulfuric acid and used as the feedstock for bioethanol production. The results showed that the miscanthus juice could be a highly nutritious source for microalgae that are a promising feedstock for biofuels. The highest cell density was observed in the 15% juice medium. Sugars released from the miscanthus cake were efficiently fermented to ethanol using Saccharomyces cerevisiae through a simultaneous saccharification and fermentation (SSF) process, with 88.4% of the theoretical yield. Full article

Biocontrol strategies for the limitation of undesired microbial developments in foods and beverages represent a keystone toward the goal of more sustainable food systems. Brettanomyces bruxellensis is a wine spoilage microorganism that produces several compounds that are detrimental for the organoleptic quality of the wine, including some classes of volatile phenols. To control the proliferation of this yeast, sulfur dioxide is commonly employed, but the efficiency of this compound depends on the B. bruxellensis strain; and it is subject to wine composition and may induce the entrance in a viable, but nonculturable state of yeasts. Moreover, it can also elicit allergic reactions in humans. In recent years, biological alternatives to sulfur dioxide such as the use of yeasts and lactic acid bacteria starter cultures as biocontrol agents are being investigated. The controlled inoculation of starter cultures allows secure, fast and complete alcoholic and malolactic fermentations, limiting the residual nutrients that B. bruxellensis utilizes to survive and grow in wine. The current study is focused on the assessment of the effect of autochthonous yeasts and bacterial strains from the Apulia Region on the development of B. bruxellensis in wine, in terms of both growth and volatile phenols’ production. The investigation evidences the positive role of indigenous mixed cultures in the control of this spoilage yeast, either co-inoculating different strains of Saccharomyces cerevisiae, S. cerevisiae/non-Saccharomyces or co-inoculating S. cerevisiae/Oenococcus oeni. Our findings expand the existing knowledge of the application of protechnological microbial diversity and of non-Saccharomyces as a biocontrol agent in oenology. We report a further demonstration of the interest in selecting indigenous strains as a strategic tool for winemakers interested in the improvement of regional wines. Full article

The use of non-Saccharomyces yeasts to improve complexity and diversify wine style is increasing; however, the interactions between non-Saccharomyces yeasts and lactic acid bacteria (LAB) have not received much attention. This study investigated the interactions of seven non-Saccharomyces yeast strains of the genera Candida, Hanseniaspora, Lachancea, Metschnikowia and Torulaspora in combination with S. cerevisiae and three malolactic fermentation (MLF) strategies in a Shiraz winemaking trial. Standard oenological parameters, volatile composition and sensory profiles of wines were investigated. Wines produced with non-Saccharomyces yeasts had lower alcohol and glycerol levels than wines produced with S. cerevisiae only. Malolactic fermentation also completed faster in these wines. Wines produced with non-Saccharomyces yeasts differed chemically and sensorially from wines produced with S. cerevisiae only. The Candida zemplinina and the one L. thermotolerans isolate slightly inhibited LAB growth in wines that underwent simultaneous MLF. Malolactic fermentation strategy had a greater impact on sensory profiles than yeast treatment. Both yeast selection and MLF strategy had a significant effect on berry aroma, but MLF strategy also had a significant effect on acid balance and astringency of wines. Winemakers should apply the optimal yeast combination and MLF strategy to ensure fast completion of MLF and improve wine complexity. Full article

Ethanol, carbon dioxide and glycerol are the major products produced by yeast during wort fermentation but they have little impact on beer and spirit flavour. It is the type and concentration of secondary metabolites that can determine overall beer flavour. These compounds are (but not only) primarily: higher alcohols, esters, carbonyls and sulphur compounds—inorganic and organic. There are a number of factors that can modify the balance of these compounds most of which are discussed in this review paper. Full article

Banana is one of the most important fruits in the Brazilian diet and is mainly consumed naturally. Losses from crop to final consumer are high and estimated in about 30%. The aim of this work was to elaborate a distilled alcoholic beverage from discarded banana and to compare with commercial trademarks. Initially, yeast strains were isolated from banana fruit and characterized by their production of volatile aroma compounds. The highest aroma-producing yeast isolate was identified by ITS-rRNA gene sequencing as Pichia kluyveri. Pasteurized banana pulp and peel was fermented by the selected P. kluyveri at approximately 10⁷ cells/mL. The sugars were converted quickly, and a high ethanol concentration (413 mg/L) was achieved after 24 h of fermentation. The fermented banana must was distilled in a Femel Alambic, and the head, heart and tail fractions were collected. The banana brandy produced had highest concentration of volatile compounds compared to trademarks, such as isoamyl acetate (13.5 mg/L), ethyl hexanoate (0.8 mg/L) and others. The results showed that whole banana must could be a good substrate for fermentation and distillation, and the sensory analysis performed revealed that the produced beverage had good acceptance by the tasters. This study demonstrates the potential of banana as a possible alternative to reduce waste and increase income to farmers. Full article

Acetic acid is undesired in Icewine. It is unclear whether its production by fermenting yeast is linked to the nicotinamide adenine dinucleotide (NAD⁺/NADH) system or the nicotinamide adenine dinucleotide phosphate (NADP⁺/NADPH) system. To answer this question, the redox status of yeast cytosolic NAD(H) and NADP(H) were analyzed along with yeast metabolites to determine how redox status differs under Icewine versus table wine fermentation. Icewine juice and dilute Icewine juice were inoculated with commercial wine yeast Saccharomyces cerevisiae K1-V1116. Acetic acid was 14.3-fold higher in Icewine fermentation than the dilute juice condition. The ratio of NAD⁺ to total NAD(H) was 24-fold higher in cells in Icewine fermentation than the ratio from the dilute juice condition. Conversely, the ratio of NADP⁺ to total NADP(H) from the dilute fermentation was 2.9-fold higher than that in the Icewine condition. These results support the hypothesis that in Icewine, increased NAD⁺ triggered the catalysis of NAD⁺-dependent aldehyde dehydrogenase(s) (Aldp(s)), which led to the elevated level of acetic acid in Icewine, whereas, in the dilute condition, NADP⁺ triggered NADP⁺-dependent Aldp(s), resulting in a lower level of acetic acid. This work, for the first time, analyzed the yeast cytosolic redox status and its correlation to acetic acid production, providing a more comprehensive understanding of the mechanism of acetic acid production in Icewine. Full article

l-arginine, an amino acid with a growing range of applications within the pharmaceutical, cosmetic, food, and agricultural industries, can be produced by microbial fermentation. Although it is the most nitrogen-rich amino acid, reports on the nitrogen supply for its fermentation are scarce. In this study, the nitrogen supply for the production of l-arginine by a genetically modified Escherichia coli strain was optimised in bioreactors. Different nitrogen sources were screened and ammonia solution, ammonium sulphate, ammonium phosphate dibasic, and ammonium chloride were the most favourable nitrogen sources for l-arginine synthesis. The key role of the C/N ratio for l-arginine production was demonstrated for the first time. The optimal C/N molar ratio to maximise l-arginine production while minimising nitrogen waste was found to be 6, yielding approximately 2.25 g/L of l-arginine from 15 g/L glucose with a productivity of around 0.11 g/L/h. Glucose and ammonium ion were simultaneously utilized, showing that this ratio provided a well-balanced equilibrium between carbon and nitrogen metabolisms. Full article

Arthrospira platensis (Spirulina) is the most cultivated microalga worldwide. Improving its cultivation in terms of biomass productivity, quality, or production cost could significantly impact the Spirulina industry. The objectives of this paper were defined as to contribute to this goal. Spirulina biomass productivity was investigated through medium choice. A modified Zarrouk’s medium was selected as it gave higher final dry weights and longer sustained growth than Hiri’s and Jourdan’s media. Then, in order to reduce Spirulina production cost, modified Zarrouk’s medium was rationalized by testing different dilutions. It was found that modified Zarrouk’s medium could be diluted up to five times without impacting the growth rates in a 28-days batch cultivation. Higher dry weights were even observed after 21 days of batch cultivation (1.21 g/L for 20%-modified Zarrouk’s medium in comparison to 0.84 g/L for modified Zarrouk’s medium). Iron uptake was then investigated as one of the major contributors to Spirulina nutritional quality. An increase in iron content was obtained by replacing iron sulfate by iron EDTA at a concentration of 10 mg_Fe/L (2.11 ± 0.13 mg_Fe/g_biomass for EDTA-FeNa, 3 H₂O at 10 mg_Fe/L compared to 0.18 ± 0.13 for FeSO₄,6H₂O at 2 mg_Fe/L). Impact of light intensity on Spirulina biomass productivity was also investigated in a 2 L Photobioreactor (PBR). Specific growth rates were calculated for Photosynthetically Photon Flux Densities (PPFD) from 85 to 430 µmol/m²/s. At 430 µmol/m²/s, photoinhibition was not observed and the specific growth rate was maximum (0.12/day). Finally, a 40-day cultivation experiment was conducted in a 1000 L PBR giving a maximum daily areal productivity of 58.4 g/m²/day. A techno-economic analysis gave production cost two to 20 times higher for PBR (from 18.71 to 74.29 €/kg) than for open ponds (from 3.86 to 9.59 €/kg) depending on Spirulina productivity. Full article

Yeasts involved in veil formation during biological ageing of Sherry wines are mainly Saccharomyces cerevisiae, and they have traditionally been divided into four races or varieties: beticus, cheresiensis, montuliensis and rouxii. Recent progress in molecular biology has led to the development of several techniques for yeast identification, based on similarity or dissimilarity of DNA, RNA or proteins. In view of the latest yeast taxonomy, there are no more races. However, molecular techniques are not enough to understand the real veil-forming yeast diversity and dynamics in Sherry wines. We propose a reliable method, using a microtiter reader, to evaluate the fermentation and assimilation of carbon and nitrogen sources, the osmotolerance and the antibiotic resistance, using 18 S. cerevisiae and 5 non-Saccharomyces yeast strains, to allow correct identification and classification of the yeast strains present in the velum of flor complex. Full article

Microbially induced mineral precipitation is recognized as a widespread phenomenon in nature. A diverse range of minerals including carbonate, sulphides, silicates, and phosphates can be produced through biomineralization. Calcium carbonate (CaCO₃) is one of the most common substances used in various industries and is mostly extracted by mining. In recent years, production of CaCO₃ by bacteria has drawn much attention because it is an environmentally- and health-friendly pathway. Although CaCO₃ can be produced by some genera of bacteria through autotrophic and heterotrophic pathways, the possibility of producing CaCO₃ in different environmental conditions has remained a challenge to determine. In this study, calcium alginate was proposed as a protective carrier to increase the bacterial tolerance to extreme environmental conditions. The model showed that the highest concentration of CaCO₃ is achieved when the bacterial cells are immobilized in the calcium alginate beads fabricated using 1.38% w/v Na-alginate and 0.13 M CaCl₂. Full article

Potato liquor, a byproduct of potato starch production, is steam-treated to produce protein isolate. The heat treated potato liquor (HTPL), containing significant amounts of organic compounds, still needs to be further treated before it is discarded. Presently, the most common strategy for HTPL management is concentrating it via evaporation before using it as a fertilizer. In this study, this scenario was compared with two biotreatments: (1) fermentation using filamentous fungus R. oryzae to produce a protein-rich biomass, and (2) anaerobic digestion of the HTPL to produce biogas. Technical, economic and environmental analyses were performed via computational simulation to determine potential benefits of the proposed scenarios to a plant discarding 19.64 ton/h of HTPL. Fungal cultivation was found to be the preferred scenario with respect to the economic aspects. This scenario needed only 46% of the investment needed for the evaporation scenario. In terms of the environmental impacts, fungal cultivation yielded the lowest impacts in the acidification, terrestrial eutrophication, freshwater eutrophication, marine eutrophication and freshwater ecotoxicity impact categories. The lowest impact in the climate change category was obtained when using the HTPL for anaerobic digestion. Full article

Lactobacillus rhamnosus GR-1 is a probiotic that has been shown to reduce the risk of urogenital problems and urinary tract infections. Rice pudding is a popular gluten-free dairy product, and could be a vehicle to deliver L. rhamnosus GR-1 to a broader population. The purpose of this study was to investigate the growth and viability of L. rhamnosus GR-1 in six fermented rice pudding samples, each one supplemented with one type of prebiotic (short-chain inulin-2% w/w, 4% w/w; long-chain inulin-2% w/w, 4% w/w) and oat-0.5% w/w, 1% w/w, along with control, over a 21-day storage period. The objective was to determine if the supplementation would have a positive effect on the microbial viability of L. rhamnosus GR-1, and to evaluate the sensory properties of the samples. All of the samples had viable levels of L. rhamnosus GR-1. Bacterial counts were at least 1 × 10⁸ CFU/mL over the 21-day storage period. The probiotic rice pudding sample supplemented with 4% w/w short-chain inulin had the highest hedonic score for flavour, sweetness, texture, and overall acceptability. This study shows that the addition of short-chain inulin, long-chain inulin, and oat had no adverse supplementation effects on the viability of L. Rhamnosus GR-1. There is the potential for the production of a novel functional food. Full article

Volatile Fatty Acids (VFA) are small organic compounds that have attracted much attention lately, due to their use as a carbon source for microorganisms involved in the production of bioactive compounds, biodegradable materials and energy. Low cost production of VFA from different types of waste streams can occur via dark fermentation, offering a promising approach for the production of biofuels and biochemicals with simultaneous reduction of waste volume. VFA can be subsequently utilized in fermentation processes and efficiently transformed into bioactive compounds that can be used in the food and nutraceutical industry for the development of functional foods with scientifically sustained claims. Microalgae are oleaginous microorganisms that are able to grow in heterotrophic cultures supported by VFA as a carbon source and accumulate high amounts of valuable products, such as omega-3 fatty acids and exopolysaccharides. This article reviews the different types of waste streams in concert with their potential to produce VFA, the possible factors that affect the VFA production process and the utilization of the resulting VFA in microalgae fermentation processes. The biology of VFA utilization, the potential products and the downstream processes are discussed in detail. Full article

The fungi Botrytis cinerea, Fusarium pallidoroseum, and Fusarium moniliforme are the causative agents of several plant diseases and can cause significant crop loss both before and after harvest. Fungicides are employed to control these phytopathogens, but fungicide use has led to an increase in resistance and may negatively affect the environment and human health. Hence, more environmentally sustainable solutions such as biological control methods are needed. The purpose of this study was to screen 22 bacterial isolates for inhibitory activity against fungal phytopathogens. To evaluate antifungal activity, the bacterial isolates were individually spot-inoculated onto Tryptic Soy Agar or de Man, Rogosa, Sharpe agar, and then a plug of fungal-colonized agar was placed onto the center of the isolate-inoculated plate. Plates were incubated at 24 °C for 10 days and fungal growth was evaluated. Nine of the 22 isolates screened inhibited all three fungi; inhibition by these isolates ranged from 51–62%, 60–68%, and 40–61% for B. cinerea, F. pallidoroseum, and F. moniliforme, respectively. Isolates were also screened for biosurfactant activity using the drop-collapse test. Bacillus megaterium, Bacillus coagulans, Bacillus thuringiensis and three Bacillus amyloliquefaciens isolates demonstrated strong biosurfactant activity and suppression of all three fungi, and therefore are recommended for further study. Full article

Microbial interactions are fundamental during milk fermentation, determining the product final characteristics. Galactomyces geotrichum, Lactobacillus plantarum and Enterococcus faecalis are among the most common microorganisms in the Colombian Kumis. The aim of the research was to evaluate the yeast–bacteria interactions in milk fermentation at 28 °C. UHT (Ultra-High Temperature) milk was inoculated with single- or multiple-strains associations and analysed periodically to determine the microbial counts, organic acids and total free amino acids (FAA). The results evidenced different growth performance of the strains in single or co-culture, with a positive effect of G. geotrichum KL20B on the lactic acid bacteria (LAB) growth performance. All the strains consumed citric acid after 6 h of incubation with E. faecalis KE06 as the major consumer; however, all the co-cultures showed an early metabolism of citrate but with a low intake rate. In addition, the interaction between G. geotrichum KL20B and E. faecalis KE06 led to a low accumulation of acetic acid. Formic acid fluctuated during fermentation. The strains interaction also led to an increase in ethanol content and a lower accumulation of FAA. In conclusion, the three strains co-culture enhances the LAB viability, with high production of lactic acid and ethanol, as a consequence of adaptation to the environment and substrate exploitation. To our knowledge, this is the first time in which it is showed that G. geotrichum KL20B could be used to compensate for the slow acid-producing ability of Lb. plantarum and E. faecalis in milk, underlining that this consortium applies some mechanisms to regulate the growth and milk composition in acids and ethanol content. Full article

Grape musts sometimes reveal excess acidity. An excessive amount of organic acids negatively affect wine yeasts and yeast fermentation, and the obtained wines are characterized by an inappropriate balance between sweetness, acidity or sourness, and flavor/aroma components. An appropriate acidity, pleasant to the palate is more difficult to achieve in wines that have high acidity due to an excess of malic acid, because the Saccharomyces species in general, cannot effectively degrade malic acid during alcoholic fermentation. One approach to solving this problem is biological deacidification by lactic acid bacteria or non-Saccharomyces yeasts, like Schizosaccharomyces pombe that show the ability to degrade L-malic acid. Excessive volatile acidity in wine is also a problem in the wine industry. The use of free or immobilized Saccharomyces cells has been studied to solve both these problems since these yeasts are wine yeasts that show a good balance between taste/flavor and aromatic compounds during alcoholic fermentation. The aim of this review is to give some insights into the use of Saccharomyces cerevisiae strains to perform biological demalication (malic acid degradation) and deacetification (reduction of volatile acidity) of wine in an attempt to better understand their biochemistry and enological features. Full article

The production of several agricultural products and foods are linked with fermentation. Traditional methods used to control and monitor the quality of the products and processes are based on the use of simple chemical analysis. However, these methods are time-consuming and do not provide sufficient relevant information to guarantee the chemical changes during the process. Commonly used methods applied in the agriculture and food industries to monitor fermentation are those based on simple or single-point sensors, where only one parameter is measured (e.g., temperature or density). These sensors are used several times per day and are often the only source of data available from which the conditions and rate of fermentation are monitored. In the modern food industry, an ideal method to control and monitor the fermentation process should enable a direct, rapid, precise, and accurate determination of several target compounds, with minimal to no sample preparation or reagent consumption. Here, state-of-the-art advancements in both the application of sensors and analytical tools to monitor beverage and food fermentation processes will be discussed. Full article

Fermented foods and beverages are a heterogeneous class of products with a relevant worldwide significance for human economy, nutrition and health for millennia. A huge diversity of microorganisms is associated with the enormous variety in terms of raw materials, fermentative behavior and obtained products. In this wide microbiodiversity it is possible that the presence of microbial pathogens and toxic by-products of microbial origin, including mycotoxins, ethyl carbamate and biogenic amines, are aspects liable to reduce the safety of the consumed product. Together with other approaches (e.g., use of preservatives, respect of specific physico-chemical parameters), starter cultures technology has been conceived to successfully dominate indigenous microflora and to drive fermentation to foresee the desired attributes of the matrix, assuring quality and safety. Recent trends indicate a general return to spontaneous food fermentation. In this review, we point out the potential risks for human health associated with uncontrolled (uninoculated) food fermentation and we discuss biotechnological approaches susceptible to conciliate fermented food safety, with instances of an enhanced contribution of microbes associated to spontaneous fermentation. Full article

Dairy waste is a complex mixture of nutrients requiring an integrated strategy for valorization into various products. The present work adds insights into the conversion of fat-rich dairy products into biomass, glycerol, and fatty acids via submerged cultivation with edible filamentous fungi. The pH influenced fat degradation, where Aspergillus oryzae lipase was more active at neutral than acidic pH (17 g/L vs. 0.5 g/L of released glycerol); the same trend was found during cultivation in crème fraiche (12 g/L vs. 1.7 g/L of released glycerol). In addition to glycerol, as a result of fat degradation, up to 3.6 and 4.5 g/L of myristic and palmitic acid, respectively, were released during A. oryzae growth in cream. The fungus was also able to grow in media containing 16 g/L of lactic acid, a common contaminant of dairy waste, being beneficial to naturally increase the initial acidic pH and trigger fat degradation. Considering that lactose consumption is suppressed in fat-rich media, a two-stage cultivation for conversion of dairy waste is also proposed in this work. Such an approach would provide biomass for possibly feed or human consumption, fatty acids, and an effluent of low organic matter tackling environmental and social problems associated with the dairy sector. Full article