Fermentation, Volume 5, Issue 2 (June 2019) – 22 articles

Spent yeast from beer manufacturing is a cost-effective and nutrient-rich starting material for the production of yeast extracts. In this study, it is shown how physiologically important ingredients in a yeast extract are influenced by the composition of the spent yeast from the brewing process. In pilot fermentations, the time of cropping (primary fermentation, lagering) of the spent yeast and the original gravity (12 ˚P, 16 ˚P, 20 ˚P) of the fermentation medium was varied, and four alternative non-Saccharomyces yeast strains were compared with two commercial Saccharomyces yeast strains. In addition, spent yeast was contaminated with the beer spoiler Lactobacillus brevis. The general nutrient composition (total protein, fat, ash) was investigated as well as the proteinogenic amino acid spectrum, the various folate vitamers (5-CH₃-H₄folate, 5-CHO-H₄folate, 10-CHO-PteGlu, H₄folate, PteGlu) and the biological activity (reduction, antioxidative potential) of a mechanically (ultrasonic sonotrode) and an autolytically produced yeast extract. All the investigated ingredients from the yeast extract were influenced by the composition of the spent yeast from the brewing process. The biodiversity of the spent yeast from the brewing process therefore directly affects the content of physiologically valuable ingredients of a yeast extract and should be taken into consideration in industrial manufacturing processes. Full article

The basidiomycete brown rot fungus Neolentinus lepideus is capable of assimilating and fermenting lactose to ethanol with a conversion yield comparable to those of lactose-fermenting yeasts. The ability of the fungus to ferment lactose is not influenced by the addition of glucose or calcium. Therefore, N. lepideus may be useful in ethanol production from materials composed mainly of lactose, such as cheese whey or expired cow’s milk. Whey is a by-product of cheese manufacturing, and approximately 50% of the total worldwide production of whey is normally disposed of without being utilized. We found that N. lepideus produced ethanol directly from cheese whey with a yield of 0.35 g of ethanol per gram of lactose consumed, and it also fermented expired milk containing lactose, protein, and fat with a similar yield. Our findings revealed that the naturally occurring basidiomycete fungus possesses a unique ability to produce ethanol from cheese whey and expired milk. Thus, N. lepideus may be useful in facilitating ethanol production from dairy wastes in a cost-effective and environmentally friendly manner. Full article

The term biorefinery is related to the sustainable production of value-added bioproducts and bioenergy from biomass. Esters from fatty acids are important compounds synthesized from by-products of the oleochemical industry. In agreement with the biorefinery concept, it is important to search for catalysts that reduce the consumption of energy and water, using moderate operation conditions and low reaction times. In this work, response surface methodology (RSM) was used to optimize the enzymatic synthesis of pentyl oleate using Candida antarctica lipase B (CALB) immobilized on a polyethylene-aluminum structured support. A factorial design was employed to evaluate the effects of several parameters on the ester yield. To obtain a model with a good fit, an approach to reaction mechanism and enzyme kinetics was taken into consideration. Experimental findings were correlated and explained using equations of a ping-pong bi-bi kinetic model and considering the inhibitory effects of both substrates. The developed model was consistent with the experimental data predicting an increase in pentyl oleate production with increasing temperature and a decrease with higher oleic acid amounts and alcohol to acid molar ratios. This model could be useful in a future industrial application of CALB/LLDPE/Al to minimize the costs in oleochemical biorefineries. Full article

Most monosaccharides in nature are hexoses, which have six carbon atoms; the most well-known hexose is d-glucose. Various hexoses with distinct characteristics can be produced from inexpensive polysaccharides for applications in the food industry. Therefore, identification of the health-related functions of hexose will facilitate the consumption of hexoses in food products to improve quality of life. The hexoses available in foods include N-acetyl glucosamine, d-glucosamine, d-fructose, d-mannose, d-galactose, other d-hexoses, and l-hexoses. Here, an updated overview of food industrial production methods for natural hexoses by microbial, enzymatic, and chemical methods is provided. Full article

Non-Saccharomyces are important during wine fermentation once they influence wine composition. In the early stages of wine fermentation, and together with indigenous or commercial strains of Saccharomyces cerevisiae, non-Saccharomyces are able to transform grape-must sugars into ethanol, CO₂, and other important secondary metabolites. A better understanding of yeast biochemistry will allow the selection of yeast strains that have defined specific influences on fermentation efficiency, wine quality, and the production of human health-promoting compounds. Yeast metabolism produces compounds derived from tryptophan, melatonin, and serotonin, which are found in fermented beverages, such as wine and beer. Melatonin is a neurohormone secreted from the pineal gland and has a wide-ranging regulatory and neuroprotective role, while serotonin, as well as being a precursor of melatonin synthesis, is also a neurotransmitter. This review summarizes the importance of some conventional and nonconventional yeast strains’ alcoholic fermentations, especially in the production of metabolites that promote human health and thus, attract consumers attention towards fermented beverages. A brief reference is also made on fermented beverages containing probiotics, namely kombucha, also known as kombucha tea, and its interesting microorganism’s symbiotic relationships named SCOBY. Full article

Previously, we developed a novel production technique for giant masu salmon (GMS). This study aimed to develop a fish sauce from GMS to explore ways to efficiently utilize the salmon and to enrich the fish sauce with γ-aminobutyric acid (GABA) by microbial fermentation. The minced bodies of GMS were autolyzed by endogenous protease at 55 °C and 60 °C. During autolysis, the changes in total free amino acids and protein size was monitored by LC-MS and SDS-PAGE analysis, respectively. After 96 h, fish sauce was prepared by heating, and the amino acid composition was analyzed by LC-MS. To enrich the fish sauce with GABA, Lactobacillus plantarum strain N10 was added and incubated at 28 °C for 48 h. The total free amino acids content significantly increased for 96 h. SDS-PAGE analysis showed that major bands at 200 kDa and 48 kDa detected at 0 h gradually disappeared over time. The ratio of anserine to total amino acids in the fish sauce was approximately 36%. The concentration of GABA in the fish sauce significantly increased through the addition of strain N10. Thus, anserine-rich fish sauce could be quickly produced from GMS, and the fish sauce was enriched with GABA by microbial fermentation. Full article

Solid-state fermentation (SSF) of rice bran (RB) employing the edible fungus Pleurotus sapidus was investigated as a process strategy to improve the nutritional quality of this low-cost and abundant substrate. During fermentation, samples were withdrawn at different time intervals (4, 6, and 10 days) and further analyzed. Established methods were deployed to monitor the changes in nutritional composition (carbohydrates, proteins, ash, and lipids). Additionally, changes in fatty acid composition was studied as a function of culture progress. Results showed that the SSF of rice bran increased total carbohydrates from 36.6% to 50.2%, total proteins from 7.4% to 12.8%, and ash from 7.6% to 11.5%. However, the total lipid content was reduced from 48.5% to 27.8%. The fatty acid (FA) composition of RB included mainly oleic, linoleic, and palmitic acids. Upon fermentation with P. sapidus, small differences were found: linoleic acid and oleic acid content were increased by 0.4% and 1.1%, respectively, while palmitic acid content was reduced by 0.8%. This study demonstrated an improvement in the nutritional quality of RB after fermentation with P. sapidus, since protein, carbohydrates, minerals, and specific FA components were increased. As a whole, our results indicate that fermented rice bran could be used as a high-quality animal feed supplement. Full article

Koji, which is used for manufacturing Japanese traditional fermented foods, has long been safely used as a cosmetic product. Although its cosmetic effect has been empirically established, the underlying mechanism has not been reported. We and other groups have previously elucidated that koji contains glycosylceramides, including N-2′-hydroxyoctadecanoyl-1-O-β-d-glucosyl-9-methyl-4,8-sphingadienine and N-2′-hydroxyoctadecanoyl-1-O-β-d-galactosyl-9-methyl-4,8-sphingadienine. This led us to hypothesise that koji exerts its cosmetic effect by acting on the keratinocytes through glycosylceramides on the gene level. Therefore, in this study, we investigated the effects of glycosylceramides from various sources on gene expression in normal human epidermal keratinocytes. The results revealed that glycosylceramides purified from white koji and the white koji-producing non-pathogenic fungus Aspergillus luchuensis and A. oryzae increased the expression of occludin (OCLN, an epidermal tight junction protein) and ATP-binding cassette sub-family A member 12 (ABCA12, a cellular membrane transporter), albeit the effect was modest relative to that of ceramides. Indeed, ceramide was increased in the keratinocytes upon koji lipid extract addition. These results indicate that glycosylceramides, which are the major sphingolipids of most natural materials, have an effect of increasing ABCA12 and OCLN expression, and suggest that koji exerts its cosmetic effect by increasing ceramide and tight junctions via glycosylceramides. Full article

Microbial fermentation of plant extracts with Lactobacteria is an option to obtain microbiologically stable preparations, which may be applied in complementary medicine. We investigated the metabolic conversion of constituents from Mercurialis perennis L. extracts, which were prepared for such applications. For this purpose, aqueous extracts were inoculated with two Lactobacteria strains, namely Pediococcus sp. (PP1) and Lactobacillus sp. (LP1). Both were isolated from a fermented M. perennis extract and identified by 16S rRNA sequencing. After 1 day of fermentation, an almost complete conversion of the genuine piperidine-2,6-dione alkaloids hermidine quinone (3) and chrysohermidin (4)—both of them being oxidation products of hermidin (1) —was observed by GC-MS analysis, while novel metabolites such as methylhermidin (6) and methylhermidin quinone (7) were formed. Surprisingly, a novel compound plicatanin B (bis-(3-methoxy-1N-methylmaleimide); 8) was detected after 6 days, obviously being formed by ring contraction of 4. An intermediate of a postulated reaction mechanism, isochrysohermidinic acid (14), could be detected by LC-MS. Furthermore, an increase in contents of the metabolite mequinol (4-methoxyphenol; 9) upon fermentation points to a precursor glycoside of 9, which could be subsequently detected by GC-MS after silylation and identified as methylarbutin (15). 15 is described here for M. perennis for the first time. Full article

Scientific and commercial interest of probiotics, prebiotics and their effect on human health and disease has increased in the last decade. The aim of this review article is to evaluate the role of pro- and prebiotics on the normal function of healthy skin as well as their role in the prevention and therapy of skin disease. Lactobacilli and Bifidobacterium are the most commonly used probiotics and thought to mediate skin inflammation, treat atopic dermatitis (AD) and prevent allergic contact dermatitis (ACD). Probiotics are shown to decolonise skin pathogens (e.g., P. aeruginosa, S. aureus, A. Vulgaris, etc.) while kefir is also shown to support the immunity of the skin and treat skin pathogens through the production of antimicrobial substances and prebiotics. Finally, prebiotics (e.g., Fructo-oligosaccharides, galacto-oligosaccharides and konjac glucomannan hydrolysates) can contribute to the treatment of diseases including ACD, acne and photo aging primarily by enhancing the growth of probiotics. Full article

A large volume of food is being wasted every year, while the pulp and paper industry also generate a large amount of solid wastes on a daily basis, causing environmental challenges around the world. Dry anaerobic digestion (AD) of these solid wastes is a cost-effective method for proper management. However, dry digestion of these waste streams has been restricted due to their complex structure, the presence of possible inhibitors and inappropriate operating conditions. In light of this fact, dry digestion of food waste (FW) and paper wastes (PW) was conducted at different total solid (TS) concentrations of reactor mixtures of 14%, 16%, 18% and 20% TS, corresponding to substrate to inoculum (S/I) ratio of 0.5 and 1; investigating the optimum operating conditions for effective dry digestion of these complex wastes. The highest methane yields of 402 NmlCH₄/gVS and 229 NmlCH₄/gVS were obtained from digestion of FW and PW, respectively at 14%TS corresponding to an S/I ratio of 0.5. Increasing the S/I ratio from 0.5 to 1 and thereby having a TS content of 20% in the reactor mixtures was unfavorable to the digestion of both substrates. Full article

Biocatalysis (the use of biological molecules or materials to catalyse chemical reactions) has considerable potential. The use of biological molecules as catalysts enables new and more specific syntheses. It also meets many of the core principles of “green chemistry”. While there have been some considerable successes in biocatalysis, the full potential has yet to be realised. This results, partly, from some key challenges in understanding the fundamental biochemistry of enzymes. This review summarises four of these challenges: the need to understand protein folding, the need for a qualitative understanding of the hydrophobic effect, the need to understand and quantify the effects of organic solvents on biomolecules and the need for a deep understanding of enzymatic catalysis. If these challenges were addressed, then the number of successful biocatalysis projects is likely to increase. It would enable accurate prediction of protein structures, and the effects of changes in sequence or solution conditions on these structures. We would be better able to predict how substrates bind and are transformed into products, again leading to better enzyme engineering. Most significantly, it may enable the de novo design of enzymes to catalyse specific reactions. Full article

Lactic acid bacteria (LAB) alone or with special adjunct probiotic strains are inevitable for the preparation of specific functional foods. Moreover, because of their growth and metabolism, final products are preserved for a certain time. Thus, in this work, growth and metabolic activity of novel animal origin isolates and culture collection strains of Bifidobacterium spp. were investigated. The influence of milk media (reconstituted or ultra-high-temperature (UHT) milk), compared with synthetic modified Wilkins–Chalgren (WCH) broth under aerobic conditions was investigated. All tested bifidobacterial strains (n = 10) were grown well (1–2 log colony-forming units (CFU)/mL for 24 h at 37 °C) in all substrates and levels higher than 5 log CFU/mL remained during the cold storage period. Generally, different substrates determined almost the same maximal population densities (MPD) after 24 h that range within the average values of 8.96 ± 0.43 log CFU/mL, 8.87 ± 0.52 log CFU/mL, and 8.75 ± 0.54 log CFU/mL in reconstituted milk, UHT milk, and WCH broth, respectively. After 28 days of storage, the pH levels in milk media and broth were reduced to 4.50–5.60 and 4.60–4.90, respectively, representing a decrease of 0.8–2.13 units. Full article

Yeasts are very important microorganisms for food production. The high fermentative capacity, mainly of the species of the genus Saccharomyces, is a key factor for their biotechnological use, particularly to produce alcoholic beverages. As viability and vitality are essential to ensure their correct performance in industry, this review addresses the main aspects related to the cellular aging of these fungi as their senescence impacts their proper functioning. Laboratory strains of S. cerevisiae have proven a very successful model for elucidating the molecular mechanisms that control life span. Those mechanisms are shared by all eukaryotic cells. S. cerevisiae has two models of aging, replicative and chronological. Replicative life span is measured by the number of daughter cells a mother can produce. This kind of aging is relevant when the yeast biomass is reused, as in the case of beer fermentations. Chronological life span is measured by the time cells are viable in the stationary phase, and this is relevant for batch fermentations when cells are most of the time in a non-dividing state, such as wine fermentations. The molecular causes and pathways regulating both types of aging are explained in this review. Full article

Sweet sorghum juice (SSJ) was evaluated as fermentation substrate for the production of l-lactic acid. A thermophilic Bacillus coagulans isolate was selected for batch fermentations without the use of additional nutrients. The first batch of SSJ (Batch A) resulted on higher lactic acid concentration, yield and productivity with values of 78.75 g∙L⁻¹, 0.78 g∙g⁻¹ and 1.77 g∙L⁻¹ h⁻¹, respectively. Similar results were obtained when the process was transferred into the pilot scale (50 L), with corresponding values of 73 g∙L⁻¹, 0.70 g∙g⁻¹ and 1.47 g∙L⁻¹ h⁻¹. A complete downstream process scheme was developed in order to separate lactic acid from the fermentation components. Coarse and ultra-filtration were employed as preliminary separation steps. Mono- and bipolar electrodialysis, followed by chromatography and vacuum evaporation were subsequently carried out leading to a solution containing 905.8 g∙L⁻¹ lactic acid, with an optical purity of 98.9%. The results of this study highlight the importance of the downstream process with respect to using SSJ for lactic acid production. The proposed downstream process constitutes a more environmentally benign approach to conventional precipitation methods. Full article

The effects of various carbon sources and cultivation conditions on the growth kinetics, lipid accumulation, and medium-chain fatty acid (MCFA) production of Mucor circinelloides (MC) was investigated for 72 h in shake flask cultivation. Our previous investigation reported increments of 28 to 46% MCFAs among total cell lipids when the MC genome was genetically modified, in comparison to the wild-type. However, the growth of the engineered strain M65-TE-04 was adversely affected. Therefore, the current study was designed to enhance the growth, lipid production, and MCFA productivity of engineered M. circinelloides by optimizing the pH, agitation speed, temperature, and carbon sources. The findings for individual variables disclosed that the highest biomass (17.0 g/L) was obtained when coconut oil mixed with glucose was used as a carbon source under normal culture conditions. Additionally, the maximum lipid contents (67.5% cell dry weight (CDW)), MCFA contents (53% total fatty acid (TFA)), and overall lipid productivity (3.53 g/L·d) were attained at 26 °C, pH 6.0, and 150 rpm, respectively. The maximum biomass (19.4 g/L), TFA (14.3g/L), and MCFA (4.71 g/L) contents were achieved with integration of a temperature of 26 °C, pH 6.0, agitation speed 300 rpm, and coconut oil mixed medium as the carbon source. This work illustrates that biomass, TFA, and MCFA contents were increased 1.70–2.0-fold by optimizing the initial pH, agitation speed, temperature, and carbon sources in the M. circinelloides engineered strain (M65-TE-04) in comparison to initial cultivation conditions. Full article

The industrial production of lactic acid (LA) is mainly based on bacterial fermentation. This process can result in enantiopure or racemic mixture according to the producing organism. Between the enantiomers, L-lactic acid shows superior market value. Recently, we reported a novel anaplerotic pathway called capnophilic lactic fermentation (CLF) that produces a high concentration of LA by fermentation of sugar in the anaerobic thermophilic bacterium Thermotoga neapolitana. The aim of this work was the identification of the enantiomeric characterization of the LA produced by T. neapolitana and identification of the lactate dehydrogenase in T. neapolitana (TnLDH) and related bacteria of the order Thermotogales. Chemical derivatization and GC/MS analysis were applied to define the stereochemistry of LA from T. neapolitana. A bioinformatics study on TnLDH was carried out for the characterization of the enzyme. Chemical analysis showed a 95.2% enantiomeric excess of L-LA produced by T. neapolitana. A phylogenetic approach clearly clustered the TnLDH together with the L-LDH from lactic acid bacteria. We report for the first time that T. neapolitana is able to produce almost enantiopure L-lactic acid. The result was confirmed by bioinformatics analysis on TnLDH, which is a member of the L-LDH sub-family. Full article

The undesirable effects of some hazardous compounds involved in the different steps of the winemaking process may pose health risks to consumers; hence, the importance of compliance with recent international food safety standards, including the Hazard Analysis and Critical Control Point (HACCP) standards. In recent years, there has been a rise in the development of new technologies in response to the hazardous effects of chemical compounds detected during the winemaking process, whether naturally produced or added during different winemaking processes. The main purpose was to reduce the levels of some compounds, such as biogenic amines, ethyl carbamate, ochratoxin A, and sulfur dioxide. These technological advances are currently considered a necessity, because they produce wines free of health-hazardous compounds and, most importantly, help in the management and prevention of health risks. This review shows how to prevent and control the most common potential health risks of wine using a HACCP methodology. Full article

A direct bioconversion of gelatinized starchy waste (GSW) to lactic acid by amylolytic lactic acid bacterium Lactobacillus plantarum S21 was investigated. Corn steep liquor (CSL) was selected as the most suitable low-cost nitrogen source for replacing yeast extract, beef extract, and peptone in De Man, Rogosa and Sharpe (MRS) medium. Plackett–Burman design results indicated that GSW and CSL were the two most nutrients that significantly influence lactic acid production, among eight medium components, including GSW, CSL, K₂HPO₄, CH₃COONa, (NH₄)₂HC₆H₅O₇, MgSO₄, MnSO₄, and Tween 80. A new low-cost medium containing only GSW (134.4 g/L) and CSL (187.7 g/L) was achieved as omitting other six components from the optimized medium had no effect on lactic acid yield. Batch fermentation at 37 °C both in 1 L and 10 L jar fermenters showed non-significantly different productivity. A by-product, maltose-forming α-amylase, was successfully achieved up to 96% recovery yield using an ultrafiltration unit equipped with a 50 kDa cut-off membrane. Crude lactic acid exhibited the additional benefit of antimicrobial activity against food and feed pathogens Salmonella enterica serovar Typhimurium TISTR 292, Vibrio cholerae TH-001, and also E. coli ATCC 25922. This study presents a promising bioprocess for the simultaneous production of lactic acid, and a value-added food enzyme, using only two industrial wastes, GSW and CSL, as the medium components. Full article

Biological itaconic acid production can by catalyzed by Aspergillus terreus (a filamentous fungi) where the fermentation medium pH is of prominent importance. Therefore, in this work, we investigated what benefits the different pH regulation options might offer in enhancing the process. The batch itaconic acid fermentation data underwent a kinetic analysis and the pH control alternatives were ranked subsequently. It would appear that the pH-shift strategy (initial adjustment of pH to 3 and its maintenance at 2.5 after 48 h) resulted in the most attractive fermentation pattern and could hence be recommended to achieve itaconic acid production with an improved performance using A. terreus from carbohydrate, such as glucose. Under this condition, the itaconic acid titer potential, the maximal itaconic acid (titer) production rate, the length of lag-phase and itaconic acid yield were 87.32 g/L, 0.22 g/L/h, 56.04 h and 0.35 g/g glucose, respectively. Full article

Fungal pretreatment is a biological process that uses rotting fungi to reduce the recalcitrance and enhance the enzymatic digestibility of lignocellulosic feedstocks at low temperature, without added chemicals and wastewater generation. Thus, it has been presumed to be low cost. However, fungal pretreatment requires longer incubation times and generates lower yields than traditional pretreatments. Thus, this study assesses the techno-economic feasibility of a fungal pretreatment facility for the production of fermentable sugars for a 75,700 m³ (20 million gallons) per year cellulosic bioethanol plant. Four feedstocks were evaluated: perennial grasses, corn stover, agricultural residues other than corn stover, and hardwood. The lowest estimated sugars production cost ($1.6/kg) was obtained from corn stover, and was 4–15 times as much as previous estimates for conventional pretreatment technologies. The facility-related cost was the major contributor (46–51%) to the sugar production cost, mainly because of the requirement of large equipment in high quantities, due to process bottlenecks such as low sugar yields, low feedstock bulk density, long fungal pretreatment times, and sterilization requirements. At the current state of the technology, fungal pretreatment at biorefinery scale does not appear to be economically feasible, and considerable process improvements are still required to achieve product cost targets. Full article