Fermentation doi: 10.3390/fermentation12100477

Authors: Nasib Qureshi Thaddeus C. Ezeji Haibo Huang Ronald E. Hector

Sustainable production of biofuels and chemicals by green fermentation routes is becoming attractive. In this paper, the production of butyric acid by fermentation of wheat straw is studied. This conversion process requires biomass milling, pretreatment, hydrolysis, fermentation, and recovery. Of these five processing steps, pretreatment, hydrolysis, fermentation, and recovery are particularly cost-intensive. In the current study we combined simultaneous saccharification and fermentation in one unit operation for the substrate of wheat straw. In the integrated process, 90 g L−1 of wheat straw was simultaneously saccharified and fermented to 20.8 g L−1 of total acetic and butyric acids, of which butyric acid is the main product. As a result of the integration of saccharification with fermentation into a single streamlined workflow, the economics of the process significantly improved. In the fermentation process in which water was recycled, 39.9 g L−1 total acid (acetic acid + butyric acids) was produced. This is comparable to 40.8 g L−1 total acid produced in the glucose-fed control fermentation. Using cell-recycling technology, acid productivity was increased by 8–10 fold (800–1000%) from wheat straw hydrolysate as a feed. In this article, product yield, productivity, and selectivity were deemed to have high potential.

Fermentation doi: 10.3390/fermentation12100476

Authors: HyunHo Yun Hwee Won Ji Yeonju Lim Ju-Sik Min Sang Hyun Kim Geon-Hee Lee Seonggon Kim Sung-Oh Sohn

Gastric mucosal injury is associated with acute epithelial damage, inflammatory cytokine release, and disruption of the protective mucus barrier. NPKE6-FM is a fermented metabolite preparation produced from puffed soybean and rice inoculated with Bacillus amyloliquefaciens NPKE6, a strain isolated from Korean water kimchi. Although NPKE6-FM has shown antioxidant and anti-inflammatory potential in previous in vitro and in vivo studies, its gastroprotective efficacy against acute gastric injury has not been fully evaluated. This study investigated the protective effects of NPKE6-FM in an HCl/ethanol-induced acute gastritis model in ICR mice. Male ICR mice were randomly assigned to six groups after weight measurement (n = 5 per group): normal control (distilled water only), model group (HCl/ethanol plus distilled water), positive control (HCl/ethanol plus rebamipide, 50 mg/kg), and NPKE6-FM treatment groups receiving 10, 50, or 100 mg/kg. Test substances or vehicle were orally administered for 14 consecutive days. Following an approximately 18 h fast on Day 15, acute gastric mucosal injury was induced by oral administration of 200 mM HCl in 80% ethanol. Gastric tissues were collected approximately 1 h after injury induction for macroscopic lesion assessment, histopathological examination, cytokine analysis, and quantitative real-time PCR analysis of inflammation- and mucin-related genes. HCl/ethanol administration induced marked hemorrhagic and erosive gastric lesions, histopathological mucosal damage, and increased levels of TNF-α and IL-6. NPKE6-FM treatment reduced gross gastric lesion scores in a dose-dependent manner, with the 100 mg/kg group showing the greatest protective effect. Histological evaluation further indicated improved preservation of gastric mucosal architecture in NPKE6-FM-treated mice. In addition, NPKE6-FM significantly suppressed TNF-α and IL-6 levels and downregulated the mRNA expression of inflammatory mediators, including COX-2, iNOS, TNF-α, and IL-6. Conversely, the HCl/ethanol-induced reduction in Muc5ac expression was substantially restored by NPKE6-FM treatment. These findings demonstrate that NPKE6-FM protects against HCl/ethanol-induced acute gastric mucosal injury through anti-inflammatory activity and reinforcement of the gastric mucus barrier. NPKE6-FM may therefore represent a promising functional food ingredient or nutraceutical candidate for the prevention or attenuation of gastric mucosal injury.

Fermentation doi: 10.3390/fermentation12100475

Authors: Gilson C. A. Chagas-Junior César R. Balcázar-Zumaeta Nelson Rosa Ferreira Roberto Mori Efrain M. Castro-Alayo Guilherme Chagas Neto Emanuela da Conceição Cardoso Alessandra Santos Lopes Gustavo Araujo Pereira Rosinelson da Silva Pena

Defined starter cultures may improve the control and consistency of traditional fermentations. Therefore, this study evaluated the effects of Pediococcus acidilactici inoculation (Pa) on the physicochemical and chemical characteristics of traditional Amazonian cassava flour fermentation compared with spontaneous fermentation (CT). Changes in pH, total titratable acidity (TTA), sugars, and hydrogen cyanide (HCN) were monitored throughout 120 h of fermentation, and the resulting flour was characterized by physicochemical properties, color, HCN, and biogenic amines. Pa promoted pronounced acidification during the final fermentation stages, reaching pH 4.77 and TTA 6.49 mEq NaOH/100 g at 120 h, and produced distinct temporal profiles of sugars and cyanogenic compounds. However, enhanced acidification did not correspond to greater cyanide removal during fermentation. Subsequent flour processing markedly reduced residual total HCN to 0.44 and 0.42 mg/kg in CT and Pa, respectively, while free HCN responses were below the calibration range in both flour samples. Among the ten biogenic amines investigated, four were quantitatively determined. Compared with CT, Pa showed significantly higher concentrations of putrescine and spermidine and a significantly lower concentration of cadaverine, whereas phenylethylamine did not differ significantly between treatments. Overall, P. acidilactici modulated fermentation and final-product characteristics, supporting further investigation of its technological potential as a starter culture for traditional fermented cassava flour production.

Fermentation doi: 10.3390/fermentation12100474

Authors: İnci Cerit Ayşe Avcı Recep Gunes Elif Sezer Eliza Altynova Omca Demirkol

This study evaluated sunflower meal, a protein- and carbohydrate-rich by-product of sunflower oil processing, as a sustainable feedstock for the supplement-free cultivation of Saccharomyces cerevisiae var. boulardii. Sequential acid and enzymatic hydrolysis were applied to release fermentable sugars. During this process, the formation of fermentation inhibitors, such as hydroxymethyl furfural (HMF) and furfural, was also monitored. Hydrolysates were analyzed by high-performance liquid chromatography over 72 h to determine sugar profiles and inhibitor concentrations. The selected hydrolysis conditions yielded up to 25.76 g/L glucose, whereas HMF and furfural concentrations remained relatively low, ranging from 0.09 to 0.52 g/L and 0.02 to 0.11 g/L, respectively. The resulting hydrolysate was evaluated as a cultivation medium for S. cerevisiae var. boulardii under submerged fermentation conditions (30 °C, 120 rpm) without additional nutrient supplementation. Viable cell counts reached 8.3–8.4 log CFU/mL, comparable to those obtained in a yeast extract-peptone-malt extract (YPM) control medium. The sunflower meal hydrolysate exhibited substantially higher antioxidant activity than YPM throughout the cultivation period, with high initial values indicating a major contribution from sunflower-derived compounds. These findings demonstrate that sunflower meal can be converted into a hydrolysate with relatively low HMF and furfural concentrations that supports substantial S. cerevisiae var. boulardii growth. The resulting fermentation broth also exhibited high antioxidant activity, highlighting the potential of sunflower meal hydrolysate as a sustainable alternative medium for probiotic yeast production.

Fermentation doi: 10.3390/fermentation12100473

Authors: Ji-Eung Kim Sung-Chul Jung Mi-Hwa Lee Su-Kyoung Lee

Nuruk, a traditional Korean fermentation starter, largely determines the quality of makgeolli; however, data on nuruk produced from novel wheat cultivars remain limited. In this descriptive case study, nuruk made from the novel wheat cultivar ‘Wooju’ (A) was characterized alongside commercial Jinju-gokja (B) and a Korea Food Research Institute reference nuruk (C), with three independently manufactured batches per type. Physicochemical, enzymatic, antioxidant, and microbiological properties were analyzed, together with qualitative Q-ToF LC-MS/MS metabolite profiles. Nuruk A showed the highest total acidity (1.19%), total polyphenol (1090 ± 15 µg GAE/g) and flavonoid (1927 ± 10 µg QE/g) contents, total fungal count, and levels of phenylalanine, tryptophan, and GABA, but the lowest α-amylase and protease activities. Of the 22 compounds detected (20 tentatively identified), six differed in detection: two lysophospholipids and a nucleotide sugar were detected only in C, two oxylipins only in A and B, and LPI(18:2) was not detected in A. Wooju-based nuruk may be suited to antioxidant-oriented applications, although its lower saccharification and proteolytic activities may require process optimization. Because the samples differed simultaneously in cultivar, starter type, and manufacturing conditions, these differences should be regarded as preliminary and cannot be attributed to the Wooju cultivar alone.

Fermentation doi: 10.3390/fermentation12100472

Authors: Tomás García-Cayuela

Food systems are currently under increasing pressure to reduce waste and by-product generation, improve resource-use efficiency, and transition towards more sustainable production models [...]

Fermentation doi: 10.3390/fermentation12100471

Authors: Ángel González-Amador Luis-Fernando Patlan-Velázquez Gabriela Rodríguez-Serrano Lorena Gómez-Ruiz Francisco Guzmán-Rodríguez Sergio Alatorre-Santamaría Aurora Pintor-Jardines Alma Elizabeth Cruz-Guerrero

Sweet whey can be valorized through fermentation and subsequent production of Requeson, a fresh whey cheese obtained by heat-induced protein precipitation. This study evaluated the physicochemical (moisture, water activity, protein, ash, chloride, and EPS content), textural (hardness, cohesiveness, adhesiveness, springiness, chewiness, and fracturability), sensory (overall liking and JAR saltiness), and in vitro bioactive characteristics of Requeson produced from sweet whey fermented with exopolysaccharide (EPS)-producing Lactobacillus delbrueckii subsp. bulgaricus NCFB 2772 and Streptococcus thermophilus SY-102, individually or in co-culture. Unfermented, citric-acid-acidified whey served as the control. The co-culture (FC) showed the highest EPS content (3.5 mg/g), higher moisture, lower hardness and fracturability, and greater springiness than the control. Consumer evaluation (n = 130) showed favorable acceptance of FC. Among FC formulations containing 0.5–1.5% NaCl, 0.5% NaCl yielded the highest overall liking, although it did not differ significantly from 1.0%. FC showed the highest DPPH radical-scavenging (22.76%) and ACE-I inhibitory (31.00%) activities. Overall, co-culture fermentation represents a potential strategy for producing value-added Requeson with modified technological and sensory properties and measurable in vitro bioactivities.

Fermentation doi: 10.3390/fermentation12100469

Authors: Mehmet Mükerrem Rençber Zijing Zhou Jingrui Deng Rutao Liu Qigui Niu

Single-stage dark fermentation offers a promising route to convert lignocellulosic green waste into biohythane, a mixture of biohydrogen (H2) and biomethane (CH4), but conditions maximizing hydrogen selectivity may increase inputs, with unclear economic consequences. This study examines the effect of food-to-microorganism (F/M) ratio, pH, agitation, and NaOH pretreatment on biohythane yield and evaluates the resulting process economics at campus, district, and city scales. The most H2-selective condition (F/M 0.5, pH 6.0, no agitation) produced 50.3 mL H2/g volatile solids (VS) and 245.9 mL CH4/g VS; alkaline operation with agitation and NaOH raised biodegradability to 81.1% of theoretical maximum but suppressed H2 to near zero. Techno-economic analysis showed the acidic, non-agitated condition achieved the highest net present value at district scale under two independent capital-cost methods (NPV = US$19.6 million and US$10.7 million; internal rate of return (IRR) = 55%): its CH4 revenue gain (+US$290,000/yr) was outweighed 3-fold by higher costs (+US$966,000/yr). This ranking held across scales and under H2-purification and tax-incentive scenarios. A low-input strategy therefore achieves superior returns by preserving CH4 revenue while avoiding agitation and pretreatment costs, with H2 better viewed as a co-benefit than the primary target, challenging the assumption that maximizing total biogas output is economically preferable within a scale-sensitive techno-economic framework.

Fermentation doi: 10.3390/fermentation12100470

Authors: Saisai Su Yuxuan Wang Longjun Shi Zuoliang Chen Xiaoyan Yu Yanqing Duan Zhihong Liu

Recovering volatile fatty acids (VFAs) from anaerobic fermentation broth as preferred carbon sources is often hindered by the poor recoverability of powdered adsorbents and the slow mass transfer within conventional hydrogel beads. Herein, we report a MgAl-layered double hydroxide (LDH) and sodium alginate (SA) composite ink tailored for extrusion-based 3D printing, followed by Ca2+ crosslinking and freeze-drying to fabricate monolithic adsorbents. Rheological tests show that the ink exhibits shear-thinning behavior and elastic-dominated viscoelasticity (G′ > G″), ensuring smooth extrusion and high shape fidelity during printing. Under optimized conditions (25 °C, 3.0 g/L, 120 min, pH 7.0), the 3D-printed monolith removed 55.74% of total VFAs. Kinetic analysis reveals that the adsorption follows the pseudo-second-order model (R2 = 0.915), consistent with a chemically controlled adsorption contribution and yielding a calculated equilibrium capacity of 286.5 mg/g. Notably, the adsorbent exhibits a pronounced selectivity toward acetic acid, which accounts for 62% of the total adsorbed VFAs. These findings demonstrate that 3D printing offers a viable strategy to overcome the intrinsic limitations of LDH-based adsorbents, namely, difficult separation and limited internal diffusion, thereby enabling efficient and selective VFA recovery from complex fermentation streams.

Fermentation doi: 10.3390/fermentation12100468

Authors: Lucía M. Mendoza Emilse Bentencourt Guillermo A. Vega-López Silvina Fadda

Oenococcus oeni is the main lactic acid bacterium responsible for malolactic fermentation (MLF) during winemaking, a critical step for obtaining high-quality wines. However, the harsh wine environment, characterized by low pH, ethanol, sulfur dioxide, and inhibitory yeast metabolites, can compromise bacterial growth and MLF performance. To investigate the mechanisms underlying wine stress tolerance, O. oeni CRL 1947 was grown in unfermented and fermented grape juice media and analyzed using a proteomic (two-dimensional electrophoresis) and RT-qPCR. A total of 38 proteins displayed significant differential expression under all conditions, with 23 up-regulated and 15 down-regulated. One protein spot (fructokinase) was exclusively detected in fermented medium, whereas aspartate carbamoyltransferase was completely inhibited. Proteins related to carbohydrate, lipid, nucleotide, and amino acid metabolism, as well as stress-response chaperones and peptidases, were over-expressed in fermented grape juice medium. In contrast, enzymes involved in lipid biosynthesis, glycolysis, amino acid metabolism, hydrolase, and dehydrogenase activities were down-regulated. Differential expression of genes associated with malic acid and citrate metabolism as well as in stress response was also observed. These findings provide further insights into the molecular adaptations of O. oeni to wine-stressful conditions and support the selection of the best adapted starter cultures for efficient MLF during winemaking.

Fermentation doi: 10.3390/fermentation12100467

Authors: Anthony Jose Balcazar-Sinailin Melanie Ashley Ochoa-Ocampo Leslie Galeny Cordero-Castillo Juan A. Castillo-Garit Patricio J. Espinoza-Montero Carlos Méndez-Durazno Noroska G. S. Mogollón Karel Diéguez-Santana

Trametes polyzona is a white-rot fungus whose laccase, manganese peroxidase, and lignin peroxidase system supports fermentation-based bioprocesses: pollutant bioremediation, effluent treatment, and enzyme production under solid-state and submerged fermentation. Its extracts also show antioxidant, antimicrobial, and cytotoxic activity. How this field is structured, and how far it has moved toward application, has not been examined. We analyzed 64 publications indexed in Scopus and Web of Science (1991–2025) with Bibliometrix R-Tool (v4.3.2) and VOSviewer (v1.6.20). Output has grown steadily since 2005, led by Thailand, Belgium, and South Africa. Keyword co-occurrence (119 terms, 3253 links) resolved five clusters: laccase catalysis and immobilization (39 terms); ligninolytic enzyme systems and strain characterization (27); comparative white-rot mycology and effluent treatment (23); dye decolorization with toxicity assessment (21); and enzyme purification and characterization (9). The strategic diagram placed laccase-mediated phenol oxidation and dye biodegradation among the motor themes, whereas bioactivity research remained a niche theme. The field has moved from enzymatic description toward immobilization and reactor-based treatment, but within the 64 records analyzed, species-specific reactor evidence rests on a single laboratory airlift study and no volumetric productivity, oxygen-transfer, or economic data are reported. Genomic and metabolomic resources remain scarce, defining clear priorities for fermentation-oriented research.

Fermentation doi: 10.3390/fermentation12100466

Authors: Emilse Bentencourt Florencia Mohamed Miguel Fernández de Ullivarri Lucía M. Mendoza

Bioinformatics has become an important tool for characterizing the properties of potential starter cultures such as Oenococcus oeni, providing insights into metabolism, wine adaptation, and genetic diversity. This study aimed to characterize the genomes of three native O. oeni strains (X2L, ST, and Sb10) isolated from the wine-producing region of Northwestern Argentina and to compare them with strains from other wine-related environments. The analyzed genomes showed features consistent with the species, including genome sizes of approximately 1.7 Mb and GC contents close to 38%. Most coding sequences were related to carbohydrate and amino acid metabolism. No genes associated with biogenic amine production or antibiotic resistance were detected, supporting their potential suitability as starter cultures for winemaking. ANI values above 97% supported their classification as O. oeni. Despite this high genomic similarity, differences were observed among strains. X2L and Sb10 exhibited an ANI of 99.99% and no SNP differences, suggesting that these isolates may represent clonal variants. In contrast, X2L and ST showed lower genomic similarity and belonged to different phylogroups, despite sharing the same origin. Strain ST exhibited greater genomic similarity to several reference strains, highlighting the genetic diversity and evolutionary complexity of O. oeni populations.

Fermentation doi: 10.3390/fermentation12100465

Authors: Gulmira T. Kassenova Zhanara K. Tulemissova Sagyman Zhadyra Zubaira Kozhakhmetova Aigerim Tuganbay Talgat Ikombayev Akkenzhe Omarova Ayaulym E. Muratbekova Botagoz Kulzhanova Per E. J. Saris

Traditional Suzbe is a fermented dairy product widely prepared at the household level in Kazakhstan, but the composition and functional potential of its microbiota remain poorly characterized. In this study, shotgun metagenomic sequencing was used to investigate the microbial communities of 20 traditional Suzbe samples collected from five geographically distinct regions of Kazakhstan. Lactic acid bacteria dominated the microbial communities, with annotations assigned to Lactococcus lactis, Lactococcus cremoris, and Streptococcus thermophilus present in all samples. Alpha diversity did not differ significantly among the five regions, and PERMANOVA did not detect statistically significant regional differences in overall microbial community composition (R2 = 0.253, p = 0.216). Nevertheless, LEfSe analysis identified several region-associated taxa, mainly among less abundant members of the microbiota. Functional annotation using the KEGG database showed that carbohydrate metabolism, amino acid metabolism, energy metabolism, cofactor and vitamin metabolism, and membrane transport were among the major functional categories. Functional PERMANOVA did not detect statistically significant regional differences in overall functional composition (R2 = 0.249, p = 0.276), although several metabolic pathways differed in relative abundance. Overall, the sampled Suzbe microbiomes were dominated by lactic acid bacteria, with several species detected in all samples and major functional categories detected across regions. Region-associated differences were identified among specific taxa and metabolic pathways; the nonsignificant PERMANOVA results do not establish taxonomic or functional conservation. This study provides new metagenomic data on traditional Suzbe and contributes to a better understanding of fermented dairy microbiota in Central Asia.

Fermentation doi: 10.3390/fermentation12100464

Authors: Yujie Wang Chuyu Zheng Gaoyu Ren Han Zhang Shuna Zhao Guanghui Shen

Production of functional exopolysaccharides (EPS) by lactic acid bacteria (LAB) is a promising strategy for the value-added utilization of beet molasses. This study aimed to isolate high-sucrose-tolerant LAB strains from beet molasses and evaluate their potential for crude EPS production. A total of 13 LAB strains were initially isolated from beet molasses, among which seven isolates were selected as high-sucrose-tolerant candidates based on their growth performance under high-sucrose conditions. Further crude EPS-production screening showed that two isolates, Leuconostoc mesenteroides C4 and Pediococcus pentosaceus T1, produced relatively high crude EPS yields. These two isolates were then identified by 16S rRNA gene sequence analysis. Further, L. mesenteroides C4 was selected as a candidate, and preliminary OFAT screening followed by Box–Behnken response surface methodology (RSM) was employed to optimize the conditions for crude EPS production. The antioxidant activity and antibiofilm activity of the crude EPS-C4 fraction were also evaluated. The RSM-predicted optimal conditions for crude EPS production by L. mesenteroides C4 were 350 g/L of beet molasses, an initial pH of 6.0, a fermentation time of 30 h, a fermentation temperature of 37 °C, an inoculum size of 2.3%, a loading volume of 85%, and a shaking speed of 108 r/min. Under these optimal conditions, a maximum crude EPS yield of 16.99 ± 0.22 g/L was obtained, which was approximately 5.0-fold higher than the yield obtained under the initial fermentation condition (3.40 g/L). Furthermore, crude EPS-C4 exhibited in vitro free-radical-scavenging activity against DPPH and ABTS+ radicals, with scavenging rates of 97.00% and 67.35% at 6.0 mg/mL, respectively. The crude EPS-C4 fraction also inhibited biofilm formation by Escherichia coli and Staphylococcus aureus, with inhibition rates of 60.68% and 68.69% at 8.0 mg/mL, respectively. These findings provide preliminary evidence that beet molasses can be used as a substrate for LAB-EPS production and suggest that crude EPS-C4 is a promising candidate for further purification, structural characterization, safety assessment, and application-oriented evaluation.

Fermentation doi: 10.3390/fermentation12100463

Authors: Jinyu Du Zhenchao Wu Tong Yang Xuan Tang Yu Bai Xinbo Cao Zhiyu Yang Hongzhi Ling Baiyan Cai Jingping Ge

Blue honeysuckle (Lonicera caerulea L.) is a cold-adapted berry rich in bioactive polyphenols. The association of phenolic compounds with the plant cell-wall matrix may limit their extractability. To promote polyphenol release and enhance antioxidant properties, a blue honeysuckle matrix was co-fermented with Lactobacillus rhamnosus 6224 and Saccharomyces cerevisiae W5. Process parameters were optimized using response surface methodology, with total phenolic content (TPC) as the response variable. Under the optimized conditions, TPC reached 1374.79 mg/100 g. Compared with the unfermented control, DPPH radical, ABTS radical, H2O2, and hydroxyl radical scavenging activities increased by 4.27, 15.10, 12.89, and 12.63 percentage points, respectively. LC-MS profiling showed that co-fermentation markedly altered the phenolic profile of the blue honeysuckle matrix and increased the relative abundances of several putatively annotated phenolic compounds, including quercetin, protocatechuic acid, catechin, and caffeic acid. These findings indicate that the optimized co-fermentation process increased TPC, altered phenolic composition, and enhanced in vitro antioxidant activity, offering a potential approach to the value-added processing of blue honeysuckle.

Fermentation doi: 10.3390/fermentation12100462

Authors: Yajing Yin Zhemin Yuan Meng Zhao Weiting Li Jingqi Wang Yueru Zhao

Polyunsaturated fatty acids (PUFAs) play critical physiological roles in maintaining cardiovascular health, neurodevelopment, and immune regulation, with continuously growing market demand. Traditional supply chains based on fish oil and plant oils face severe challenges including resource depletion, environmental pollution, and sustainability concerns. Single cell oil production via fermentation using filamentous fungi has emerged as a sustainable and economically viable alternative. This review provides a comprehensive overview of research progress on PUFA synthesis by filamentous fungi, covering strain resources, biosynthetic pathways, regulatory mechanisms, major product development, and industrialization prospects. Filamentous fungi have formed a complete spectrum from native high-yielding strains (e.g., Mortierella alpina, Mucor circinelloides, Umbelopsis isabelline, Thamnidium elegans, Cunninghamella echinulata) to genetically tractable chassis cells (e.g., Aspergillus oryzae). In addition, the oleaginous yeast Yarrowia lipolytica also plays a non-negligible role in the heterologous synthesis of polyunsaturated fatty acids. Starting from acetyl-CoA, filamentous fungi synthesize saturated fatty acids via the fatty acid synthase system and subsequently convert them to important PUFAs such as arachidonic acid (ARA), γ-linolenic acid (GLA), and eicosapentaenoic acid (EPA) through the sequential catalysis of Δ9-, Δ12-, Δ6-, and Δ5-desaturases and elongases. The SNF1 energy sensor, the nitrogen metabolism regulator AreA, and environmental factors constitute a complex multi-layer regulatory network that coordinately modulates fatty acid unsaturation and yield. The 2A peptide-based multigene co-expression platform, cofactor engineering, competitive pathway blockade, and fermentation process optimization have synergistically enabled the targeted accumulation of specific PUFAs. Nevertheless, industrialization of filamentous fungal PUFA production still faces major bottlenecks. We also provide a perspective on the industrial potential of filamentous fungi as “lipid cell factories” for the production of polyunsaturated fatty acids (PUFAs). This review aims to provide a comprehensive knowledge framework for researchers in related fields and to inform future fundamental research and industrial translation.

Fermentation doi: 10.3390/fermentation12100461

Authors: Kristin Rang Wolfgang Büscher Yurui Sun Jessica Paßmann Gerd-Christian Maack

As temperatures rise worldwide, the effects of ensiling and storage temperature on silage need to be elucidated. In this paper, we aim to illuminate the effect the storage temperature has on the aerobic stability of whole-plant maize silage. The study was designed to identify the effect that temperatures of 23 °C and 33 °C have on untreated or maize silage inoculated with a biological additive. Furthermore, the study elucidates how temperature affects the ensiling phase of untreated maize silage. The silage was incorporated into the novel nine-sample live measuring system, which enables us to constantly measure the temperature pH and oxygen content 10 cm behind the silo face. The second experiment proved that silage that was fermented and stored at 23 °C remained aerobically stable longer than silage fermented at 33 °C. Switching the variant from 33 °C during fermentation to 23 °C during storage resulted in aerobic stability matching that of silage continuously stored at 33 °C, with both differing significantly from the variant continuously stored at 23 °C. In the first experiment, it was demonstrated that the biological additive used in this study improved several indicators of aerobic stability independent of fermentation temperature.

Fermentation doi: 10.3390/fermentation12100460

Authors: Jia Li Panpan Wang Ziwen Qu Ying Xu Wenrui Li Yuhan Shan Yuping Quan

This study examined how targeted mutations in two pyruvate-associated genes, d-ldh and budA, affected adaptive phenotypes and fermented milk quality in Companilactobacillus crustorum MN047. Premature stop codons were introduced into d-ldh and budA by CRISPR-assisted base editing and verified by Sanger sequencing. The resulting nonsense mutants, designated ΔbudA and Δd-ldh, retained growth comparable to the wild-type strain (wt) but showed contrasting phenotypes. At 24 h, auto-aggregation was 4.44-fold the wt value in Δd-ldh and 0.71-fold the wt value in ΔbudA, whereas ΔbudA showed lower aggregation and stress survival. Viable counts remained 7.2–7.7 log10 CFU/mL, although titratable acidity and textural properties differed among fermented milk groups. ΔbudA produced higher firmness and consistency, whereas Δd-ldh showed lower values for both parameters. Electronic nose PCA clearly separated the three groups, while GC-MS identified strain-dependent differences in selected relative volatile compounds. These findings show that the two pyruvate-associated mutations affected bacterial robustness and fermented milk quality in distinct ways.

Fermentation doi: 10.3390/fermentation12100459

Authors: Julia Kruyeniski Carolina Mónica Mendieta Fernando Esteban Felissia María Cristina Area

Error in Figure [...]

Fermentation doi: 10.3390/fermentation12100458

Authors: Leqi Tang Yujue Wang Jianghua Chen Qiang Wang Yan He Miaomiao Zeng Xian Zhang Hailing Zhang

The growing global demand for flavor stability and terroir expression in the specialty coffee industry has positioned postharvest fermentation as a core driver of coffee quality upgrading, second in importance only to cultivar genetic improvement. Coffee fermentation is shifting from an empirical spontaneous pectin-removal process to a precision biotechnological approach for targeted flavor regulation, with microbial communities acting as core mediators of biochemical transformation. This review compiles key research progress in coffee fermentation from 2018 to 2026, spanning processing paradigms, microbial ecology, flavor metabolic mechanisms, sensory safety, and industrial iteration. It compares the microenvironmental characteristics and microbial succession of four mainstream processing modes; clarifies functional divergence and terroir effects of the core microbiota; dissects the multi-stage cascade flavor metabolic network; and explains how fermentation precursors transfer to roasted coffee flavor. Importantly, most microbiota–flavor associations in current studies remain correlative rather than experimentally proven causal relationships. Evidence indicates that inoculated fermentation can improve cupping quality, reduce batch-to-batch variation, and mitigate biosafety risks. Several proposed precision fermentation approaches remain at the laboratory and pilot-scale stage and are not yet widely deployed industrially. Current field bottlenecks include poor regional adaptability of commercial starters, limited real-time monitoring tools, and ambiguous microbiota–flavor causal mechanisms. Future research will focus on terroir-adapted synthetic consortia, AI-driven precision fermentation, and multi-omics integrated studies, to advance coffee processing from empirical craft to precision biomanufacturing.

Fermentation doi: 10.3390/fermentation12100457

Authors: Sasiprapa Kullachonphuri Nuttapon Khongdee Suwimon Wicharuck Yupa Chromkaew Yun-Hsin Lin Kuan-Chen Cheng

Drought stress is one of the most damaging abiotic stressors impacting global agricultural productivity and soil ecosystem integrity, especially in the semi-arid and arid areas. Drought results in a cascade of physiological and biochemical disturbances in plants, such as inhibition of photosynthesis, loss of turgor pressure, and impairment of osmoregulation, while also causing a degradation of soil health through losses in organic carbon, nutrient availability, and microbial community structure. This calls for integrated soil management approaches that can restore soil health and support plant resilience simultaneously under water-limited conditions. Biochar, generated through the thermal decomposition of agro-waste materials under oxygen-limited conditions, represents a promising circular bioeconomy strategy for valorizing agricultural residues into high-value soil amendments. Biochar is widely known to enhance the physicochemical properties of soils, including water retention, cation exchange capacity, and microbial habitat provision. Plant growth-promoting rhizobacteria (PGPR) have been established as an environmentally friendly means of enhancing plant growth through phytohormone production, ACC deaminase activity, and antioxidant enzyme activation under stressed conditions. The combined application of biochar and PGPR has garnered considerable scientific attention due to their complementary modes of action and, in some cases, beneficial combined effects. Several studies reported significant improvement in plant water status, photosynthetic efficiency, osmoregulation, soil enzyme activity, and nutrient retention. This article summarizes the present research on the soil and physiological effects of drought stress before exploring the basic mechanisms of biochar–PGPR interaction and their combined efficacy under drought conditions. Furthermore, the review calls attention to the future research priorities, such as field-scale validation, region-specific formulation development, and omics-based mechanistic investigations to maximize the full potential of PGPR and biochar combined as a sustainable approach for alleviating drought stress.

Fermentation doi: 10.3390/fermentation12100456

Authors: Juan Fernando Campaña-Perez Sascha Bernard Lokman Alpsoy Marta Mollerach Thomas Brandstetter Betiana Lerner Maximiliano Sebastian Perez Jürgen Rühe

This study addresses some limitations of standard cell encapsulation methods by producing hydrogel beads via UV-induced C,H-insertion crosslinking (CHic) in a microfluidic setup and the simultaneous encapsulation of yeast cells. It describes a simple one-step production of monodisperse beads with consistent dimensions of ~1000 × 500 µm and the encapsulation of Saccharomyces cerevisiae within these beads. Glucose consumption profiles show that the encapsulated yeast consumes glucose at rates comparable to free yeast, indicating that encapsulation does not impair metabolic activity. The encapsulated cells maintain high retained metabolic activity for up to 15 days at 4 °C and even at 30 °C, whereas free yeast stored at this temperature suffers a significant loss of activity. This highlights the potential of the platform for storing cells without refrigeration. Another advantage is that the fermentation solutions remain clear with minimal turbidity, and the beads can be easily separated from the solution by simple sieving, thus avoiding the need to filter the solution after fermentation. The results obtained suggest that the platform is robust and versatile for use in fermentation bioreactors and cell storage.

Fermentation doi: 10.3390/fermentation12100455

Authors: Emma P. Fukuda Emily C. Fowler Merritt L. Drewery

Previous research has established distinctions between microbial communities that associate with either the solid or liquid rumen microenvironments utilizing 16S rRNA sequencing. Whole genome metagenomic sequencing (WGS) facilitates comprehensive analysis of a microorganism’s entire genome, producing strain level data and providing insights into microbial functionality. The objective of our study was to utilize WGS to characterize microbial species and enzyme profiles associated with the liquid versus solid rumen microenvironments in beef steers consuming forage. Four steers consuming forage were utilized in a 4 × 4 Latin Square experiment; on d 14 of each period, rumen contents were separated into solid and liquid samples and DNA was extracted then sequenced using WGS. Alpha and beta diversity measures revealed differences between solid and liquid microenvironments. There were also differences in enzyme profiles: carbohydrate-active enzyme classes were different across microenvironments, except glycosyl transferase. Further, the solid microenvironment was enriched in enzymes related to energy production/conversion and nutrient transport and metabolism, while the liquid microenvironment was enriched in coenzyme transport and metabolism enzymes. Species associated with cellulolytic activities were generally more abundant in the solid microenvironment while those associated with amylolytic activity were more abundant in the liquid. Genes related to methanogenesis were also different across microenvironments. Our findings offer valuable insights into the stratification of microorganisms across rumen microenvironments, expanding on existing literature with findings from next-generation sequencing and application to forage-based beef systems.

Fermentation doi: 10.3390/fermentation12100454

Authors: Tuba Buyuksirit-Bedir

The occurrence of pesticide residues in vegetables remains an important food safety concern, while their behavior during traditional fermentation is less well characterized. This study evaluated changes in pesticide residues during 35 days of spontaneous pepper fermentation. Ten commercial pepper samples were fermented using a traditional household-scale procedure, and pesticide residues and microbial populations were monitored throughout fermentation. A total of 120 pesticides were analyzed by liquid chromatography-tandem mass spectrometry, and processing factors (PFs) were calculated. Four pesticides, acetamiprid, boscalid, cypermethrin, and metalaxyl, were detected in seven of the ten fresh pepper samples. The concentrations of all detected pesticides decreased during spontaneous fermentation, although the magnitude of reduction varied considerably among compounds. Cypermethrin showed the greatest reduction, reaching 92.4% by day 21 (PF = 0.076), whereas acetamiprid and metalaxyl decreased by 56.6% (PF = 0.434) and 49.7% (PF = 0.503), respectively, by day 35. Boscalid showed comparatively greater persistence, with a 20% reduction (PF = 0.800) on day 35. Exploratory correlation analysis showed predominantly negative associations between pesticide concentrations and microbial counts. However, these findings should not be interpreted as evidence of a causal relationship with pesticide dissipation. The magnitude of dissipation varied among the detected pesticides and may also have been influenced by sample- and matrix-specific factors.

Fermentation doi: 10.3390/fermentation12100453

Authors: Aris Melloni Michal Dabros Helena Mylise Copeland Yoann Fink Keith D. Rochfort Brian Freeland

This study presents the development of an advanced control system for regulating the specific growth rate of Lactobacillus rhamnosus in fed-batch cultures using an advanced soft sensor approach. A predictive Artificial Neural Network (ANN) model was developed to estimate growth rate in real time, utilizing input variables including total cell density, oxygen uptake rate, carbon evolution rate, dissolved oxygen, and bioreactor volume. The ANN model demonstrated high accuracy, with a validation-subset RMSE of 0.073 h−1 and an R2 of 0.99. To improve control stability, the predicted growth rate was integrated into a closed-loop Proportional-Integral (PI) controller that dynamically adjusted the substrate feed rate. The controller parameters were optimized using the Ziegler-Nichols method, achieving a mean control error of 18.2% ± 5.26% across different growth rate setpoints. The implementation reduced process variability and enhanced control precision compared to traditional control methods. These findings highlight the potential of ANN-based predictive models for improving bioprocess automation, indicating the potential of ANN-based predictive models for bioprocess automation, subject to validation under industrial conditions.

Fermentation doi: 10.3390/fermentation12100452

Authors: Romana Heralecka Karolina Kostelnikova Mojmir Baron Jiri Sochor

Climate change accelerates grape ripening, intensifies malic acid degradation, and promotes potassium accumulation, which results in lower acidity and increased pH in musts and wines. This review summarizes the mechanisms underlying these changes and critically evaluates the available approaches to compensate for them. Traditional chemical and physical acidification methods are associated with several technological, sensory, and economic limitations. Biological acidification is a promising alternative, particularly through the use of Lachancea thermotolerans for lactic acid production and selected Saccharomyces cerevisiae strains capable of producing malic acid. The potential of targeted selection and genetic modification is also discussed. Biotechnological approaches therefore represent a promising tool to maintain acid–base balance and wine quality in a warming climate.

Fermentation doi: 10.3390/fermentation12100451

Authors: Anastasia Thanou Anna Ofrydopoulou Vasileios D. Prokopiou Katie Shiels Alexandros Tsoupras

Commercially available caprine dairy matrices are rich sources of lipid-associated bioactive constituents; however, comparative evaluations of their polarity-separated fractions remain limited. This study aimed to characterize and compare the chemical composition, antioxidant capacity, and inhibitory effects on thrombo-inflammatory agonist-induced platelet aggregation of amphiphilic (total amphiphilic content, TAC) and lipophilic (total lipophilic content, TLC) fractions isolated from three independently produced commercial goat milk and three independently produced commercial kefir-based goat yogurt-type fermented products, with different LOT numbers. Total lipid (lipophilic and amphiphilic) extracts were obtained by Bligh and Dyer extraction and then fractionated to TAC and TLC by counter-current separation using a green hexane/ethanol–water solvent system. Analytical determinations included spectrophotometric quantification of total carotenoid and phenolic contents, attenuated total reflectance Fourier-transform infrared (ATR–FTIR) structural profiling, and liquid chromatography–mass spectrometry (LC–MS) relative fatty-acid profiling. Antioxidant capacity was assessed using DPPH and ABTS radical-scavenging assays, while ex vivo light-transmission aggregometry in human platelet-rich plasma, obtained from six independent healthy donors (n = 6) for each fraction, was employed to evaluate potential inhibitory effects against platelet-activating factor (PAF) and adenosine diphosphate (ADP). Across both matrices, TAC fractions exhibited markedly higher radical-scavenging responses than the corresponding TLC fractions, particularly in the ABTS assay. In platelet aggregometry, TAC fractions showed significantly stronger inhibition of both PAF- and ADP-induced aggregation than their corresponding TLC fractions. Specifically, the TAC fraction from the kefir-based yogurt product exhibited the lowest IC50 value against PAF-induced aggregation, whereas the two TAC fractions showed comparable inhibitory potency against ADP-induced aggregation. ATR–FTIR spectroscopy revealed spectral features consistent with lipid-associated, hydroxyl-containing, and phosphate-containing amphiphilic structures, while LC–MS profiling identified palmitic, stearic, oleic, and α-linolenic acids among the predominant fatty acids in the free and esterified fatty-acid pools. Overall, the TAC fractions exhibited substantial in vitro radical-scavenging activity and ex vivo inhibition of agonist-induced platelet aggregation, suggesting potential anti-inflammatory and antithrombotic relevance for these activities. Further in vivo studies are required to determine their physiological relevance.

Fermentation doi: 10.3390/fermentation12100450

Authors: Ronnie G. Willaert

Microorganisms are, above all, chemists [...]

Fermentation doi: 10.3390/fermentation12100449

Authors: Yaodi Han Yudan Li Yunfei Zhai Xinling Wang Jiaxuan Song Wudong Liu Zhaoyu Han

This study aimed to determine the effects of dietary supplementation with a compound additive containing guanidinoacetic acid (GAA) and cysteamine on rumen metabolism, fermentation, and the microbiota of fattening Holstein bulls. Sixty Holstein dairy bulls (294.33 ± 3.42 kg) were divided into three groups (n = 20). Each treatment comprised three independently fed pens (n = 3 pens per treatment), and one bull was sampled from each pen for the rumen measurements. Group A served as the control group, whereas Groups B and C received a concentrated supplement containing 1 kg/t and 2 kg/t of a compound additive (GAA combined with cysteamine), respectively, over 90 days. Kyoto Encyclopedia of Genes and Genomes identified five key pathways. Specifically, indole-3-acetaldehyde, 2-amino-3-methoxybenzoic acid, 2-succinyl-5-enolpyruvyl-6-hydroxy-3-cyclohexene-1-carboxylate, and cholesterol glucuronide were upregulated, whereas 4-fumarylacetoacetic acid and pseudouridine 5′-phosphate were downregulated. Compared with those in the control group, acetic acid, butyric acid, total volatile fatty acids (VFAs), propionic acid, and microbial crude protein (MCP) increased, whereas pH, acetic acid/propionic acid (A/P) ratio, and NH3-N did not differ. At the genus level, Weissella abundance increased, Methanobrevibacter abundance decreased, and Xylanibacter and Ruminococcus abundances increased in the low-supplementation group (1 kg/t). Methanobrevibacter correlated negatively with VFAs, MCP, and NH3-N but correlated positively with A/P. Weissella and Ruminococcus showed the opposite trend. Increasing the inclusion level of the compound additive improved rumen metabolism, fermentation, and the microbial environment. This ultimately reduced stress and enhanced rumen health in fattening Holstein bulls.

Fermentation doi: 10.3390/fermentation12100448

Authors: Redife Aslihan Uçar Ömer Şimşek

The cosmetic and dermocosmetic industries are increasingly incorporating fermentation-derived ingredients for their benefits on skin barrier function, hydration, pigmentation, oxidative stress, and microbiome homeostasis. Advances in fermentation biotechnology have enabled the production of bioactive compounds, including organic acids, postbiotics, microbial lysates, bioactive peptides, bacteriocins, and fermentation-enhanced phytochemicals. These bioactives may support skin health through barrier reinforcement, modulation of melanogenesis, antioxidant activity, and microbiome interactions. Beyond topical applications, fermented functional foods and microbiome-derived metabolites may influence skin physiology through the gut–skin axis, linking microbial metabolism with systemic immune and barrier regulation. Fermentation also offers opportunities for sustainable production of dermocosmetic ingredients from renewable resources and agro-industrial by-products. However, evidence on fermentation-derived cosmetic ingredients, fermented foods, microbiome science, nutritional dermatology, and cosmetic biotechnology remains fragmented across disciplines. This review therefore introduces Fermentocosmetology as an integrative conceptual terminology, rather than a new discipline, to connect these research areas within a unified framework. Attention is given to bacteriocins as fermentation-derived antimicrobial peptides with emerging dermatological relevance, including activity against Cutibacterium acnes and Staphylococcus aureus. Overall, the proposed framework positions fermentation as a versatile platform connecting topical and systemic approaches to skin health while highlighting opportunities for microbiome-oriented, sustainable, and precision dermocosmetic innovation.

Fermentation doi: 10.3390/fermentation12100447

Authors: Tevfik Koçak Tuba Eda Arpa Zemzemoğlu Özlem Baran Elif Melek Avci Aziz Alper Biten Bence Raposa Duygu Ağagündüz

Fermentation is among the oldest food technologies, yet its relevance now extends well beyond preservation. Fermented foods are increasingly viewed as matrix-dependent biological systems whose microbial and chemical composition evolves during fermentation and post-fermentation processing, and whose functional components may undergo further transformation during gastrointestinal digestion and host–microbiome interaction, rather than as passive carriers of single nutrients or microorganisms. This review synthesizes mechanistic, compositional, and clinical evidence on fermented foods as functional foods through three core scientific pillars: (i) the biological and matrix-related determinants of functionality, including microbial genotype and community ecology, enzymatic transformation, matrix restructuring, fermentation and post-fermentation processing, gastrointestinal digestion, and host phenotype; (ii) the principal fermentation-derived bioactive systems, including bioactive peptides, exopolysaccharides, organic acids, microbially synthesized vitamins, bioaccessible minerals, biotransformed phenolics, γ-aminobutyric acid and related amino acid derivatives, bacteriocins, bioactive lipids, and postbiotic components; and (iii) the mechanistic pathways linking these systems to immune, metabolic, endocrine, gastrointestinal, neuroimmune, and aging-related outcomes. Building on these foundations, the review further examines two translational dimensions: precision nutrition, with emphasis on interindividual variability, host–microbiome interactions, responder phenotypes, and product-specific human evidence; and next-generation functional fermented foods, integrating systems-level host–microbiome frameworks, multiomics, emerging predictive technologies, sustainability, safety, and regulatory translation. Microbial–epithelial–immune signaling, short-chain fatty acid receptor activation, regulatory T-cell modulation, intestinal barrier reinforcement, bile acid signaling, and selected fermentation-derived bioactive systems are supported by mechanistic evidence. Strong translational evidence is concentrated in well-characterized product–endpoint combinations, while broad metabolic, endocrine, neurocognitive, neurodegenerative, and healthy-aging claims are heterogeneous, limited, or mostly supported by preclinical and observational evidence. Poor strain-level and process characterization, varied product formulations, limited intervention periods, weak comparators, and extrapolation from isolated microbes or metabolites to whole fermented meals continue to hinder causal interpretation in the literature. Thus, future research should replace generic fermented food categories with reproducible product fingerprints integrating matrix composition, strain-resolved microbial identity, fermentation parameters, viable and non-viable fractions, metabolomic profiles, dose, and host characteristics. To determine which fermented products have clinically meaningful effects, in whom, through which mechanisms, and under what conditions, standardized controlled human interventions, multiomics with causal mediation approaches, precision nutrition frameworks, and integrated safety and regulatory evaluation are needed.

Fermentation doi: 10.3390/fermentation12100446

Authors: Gary Meza-Bone Verónica Andrade-Yucailla Iván González-Puetate Jessica Meza-Bone Marcos Barros-Rodríguez Edison Barros-Rodríguez

Forage scarcity, especially during dry periods, limits feed availability for livestock systems, affecting the productivity and sustainability of animal production. The aim of this investigation was to determine the effect of three harvest ages (8, 10 and 12 days) and five nutrient solution concentrations (0, 25, 50, 75 and 100%) on the morpho-agronomic performance, chemical composition, digestibility, rumen degradation kinetics and mitigation of gas, CH4 and CO2 emissions of hydroponic maize forage. Digestibility, degradation, and the reduction in total gas, CH4, and CO2 emissions were evaluated using in vitro techniques. A completely randomized design in a factorial arrangement (3 × 5) was used (8, 10 and 12 days of harvest) (0, 25, 50, 75 and 100% nutrient solution). Four Brahman bulls weighing 550 ± 30.5 kg were used, each fitted with a ruminal cannula. The animals were kept in separate pens, where they were fed Megathyrsus maximus and had access to water ad libitum. Biomass production demonstrated significant differences (p < 0.0001) for the effect of harvest age (12 days), concentration (100%), and their interaction. Dry matter, crude protein, neutral detergent fiber, and acid detergent fiber indicated significant differences (p < 0.0001) for the effect of harvest age, concentration, and their interaction. Dry matter digestibility yielded an effect of harvest age (8 days), concentration (75% and 100%), and their interaction (p < 0.0001). Rumen dry matter degradation of the soluble fraction, potential degradation, and effective degradation (0.02, 0.05, and 0.08, respectively) indicated an effect of harvest age, concentration, and their interaction (p < 0.0001). Gas, CH4 and CO2 production had an effect on harvest age (8 days) and fertilization concentration (75% and 100%; p < 0.0001). It can be concluded that at early harvest ages (8–10 days) combined with intermediate to high concentrations of nutrient solution (75–100%), the biomass production and chemical composition, and thus the increase in ruminal digestibility and degradability and therefore the mitigation of CH4 and CO2, of hydroponic maize forage are optimized, making it a feeding strategy for sustainable livestock production.

Fermentation doi: 10.3390/fermentation12100445

Authors: Xiaohui Li Jinxiu Yu Ke Tong Jianfeng Mei

This study compared two inoculum preparation protocols for the mixed fermentation of unsterilized soybean meal (SBM) with Aspergillus niger and Candida tropicalis. The mono-culture-derived (MO-C) inoculation protocol involved separately cultivating A. niger and C. tropicalis and subsequently inoculating both cultures into SBM, whereas the co-culture-derived (CO-C) inoculation protocol involved cultivating both microorganisms together in the same medium before inoculating into SBM. Compared with the MO-C treatment, the CO-C treatment resulted in a slightly higher apparent crude protein content and significantly increased the acid-soluble protein and free amino acid contents in fermented soybean meal (FSBM), with increases of 9.22% and 141%, respectively. The CO-C treatment was also associated with greater reductions in β-conglycinin and raffinose-family oligosaccharides (RFOs), as well as higher DPPH and ABTS radical-scavenging activities and ferric-reducing power of FSBM. These findings suggest that the co-culture inoculum protocol provides a simplified approach for preparing mixed inocula and may improve selected nutritional and functional properties of FSBM under the tested conditions.

Fermentation doi: 10.3390/fermentation12090444

Authors: Karine Rabelo Fonseca Roberta Passini Sérgio Botelho de Oliveira Danns Pereira Barbosa Luana Alves Akamine Silvia Robles Reis Duarte

This study evaluated the potential for biogas production from anaerobic digestion of cattle manure using bioaugmentation in batch-type bioreactors. A completely randomized design was adopted, with four treatments and three replications: CM, cattle manure only, as control; CMI, cattle manure plus inoculum; CMI-3, cattle manure plus inoculum and 3 mL of bioremediation additive; and CMI-9, cattle manure plus inoculum and 9 mL of bioremediation additive. The inoculum originated from a UASB (Upflow Anaerobic Sludge Blanket) reactor used in the effluent treatment plant of a food industry. The bioremediation additive was a gel containing environmental bacterial spores composed of different concentrations of Bacillus megaterium, Bacillus mycoides, Bacillus amyloliquefaciens, and Bacillus thermoglucosidasius. Substrates were characterized by pH, dry mass, organic and inorganic dry mass, total organic carbonate/total inorganic carbonate ratio, and nutrient concentration. Biogas volume, gas characteristics, and gas concentrations were assessed. Data were analyzed by ANOVA and F tests, followed by Tukey’s test at 5% significance. Inoculum, alone or combined with bioremediation, improved biogas production, with 9 mL addition of bioremediator resulting in 55.40% methane.

Fermentation doi: 10.3390/fermentation12090443

Authors: Na Jiang Yukun Zhang Manabu Ishikawa Xiaoxiao Zhang

Marine macroalgae and terrestrial halophytes represent a structurally distinct class of feed resources that can be produced without the arable land, fresh water, and synthetic fertilizer inputs required by conventional protein crops. Their direct incorporation into animal diets is, however, constrained by a convergent set of physical and toxicological barriers—viscous polysaccharides, lignocellulosic recalcitrance, anti-nutritional secondary metabolites, and accumulated heavy metals and halogens—that collectively impede practical use. Microbial fermentation can transform some of these barriers into functional resources through enzymatic deconstruction, catabolite conversion, and biosorption—a duality described here as the Barrier–Resource Duality. This critical narrative review synthesizes findings from publications across fermentation microbiology, animal nutrition, food safety, life cycle assessment, and regulatory science. The review analyzes cell-wall architectures of Chlorophyta, Rhodophyta, and Ochrophyta alongside the lignocellulosic matrix of terrestrial halophytes, maps the requisite carbohydrate-active enzyme families and polysaccharide-utilization loci, and evaluates the postbiotic mechanisms—TLR/NF-κB, histone deacetylase inhibition, and metabolite-mediated immunomodulation—by which fermentation-derived compounds may exert host benefit. The evidence base is assessed through a critical narrative synthesis that identifies knowledge gaps across five areas: (i) the absence of co-fermentation studies combining seaweeds and halophytes despite predicted substrate complementarity; (ii) the lack of pilot-scale fermentation data (no study exceeds 500 L); (iii) the single-study dependency of core in vivo efficacy claims; (iv) the absence of validated analytical methods for postbiotic quantification; and (v) the lack of a dedicated regulatory category for fermented marine biomass. It concludes that the field, while mechanistically rich, remains at Technology Readiness Level 2 (technology concept formulated), and that coordinated pilot-scale consortia, multi-institutional replication initiatives, and regulatory pathway design are prerequisites for commercial translation.

Fermentation doi: 10.3390/fermentation12090442

Authors: Hülya Hanoğlu Oral

Essential oils (EOs) have been extensively investigated as feed additives; however, their effects on ruminal degradation kinetics remain inconsistent, particularly for thymol- and carvacrol-rich EOs. This study evaluated the effects of concentrate feed with added thymol- and carvacrol-rich EOs, providing different levels of thymol and carvacrol, on ruminal degradation kinetics and effective degradability of dry matter (DM), organic matter (OM), and crude protein (CP) using an in situ nylon bag technique. Treatments included the concentrate feed (control); the concentrate feed with added carvacrol-rich EO to provide 300 or 100 mg carvacrol/kg concentrate (C300 and C100); the concentrate feed with added thymol-rich EO to provide 300 or 100 mg thymol/kg concentrate (T300 and T100); and the concentrate feed with both EOs added to provide 150 mg carvacrol + 150 mg thymol/kg concentrate (CT300) or 50 mg carvacrol + 50 mg thymol/kg concentrate (CT100). The carvacrol treatments generally maintained DM and OM degradability at levels comparable to the control, whereas the thymol and combined EO treatments showed lower degradability, particularly during the early incubation periods (p < 0.05). Thymol treatments also showed lower CP degradability than the control and carvacrol treatments at several incubation times (p < 0.05). Planned contrasts showed significant differences in DM and OM degradation kinetics between the carvacrol and thymol treatments, with thymol treatments showing lower effective degradability of DM, OM, and CP at all evaluated passage rates (p < 0.05), whereas no differences in CP kinetic parameters were detected. Degradation kinetics and effective degradability were similar between the individual EO treatments providing 300 and 100 mg carvacrol or thymol/kg concentrate. Combined EO treatments showed lower effective degradability of DM and OM than the individual EO treatments (p < 0.05), whereas CP effective degradability did not differ between the combined and individual EO treatments. Overall, these findings highlight the importance of considering EO type and nutrient-specific responses when selecting EOs for targeted use in ruminant nutrition.

Fermentation doi: 10.3390/fermentation12090441

Authors: Qingchun Luo Pengju Zhao Jianghua Li Xi Li Jian Chen Xuejun Lei Yanping Lu Jian Su Dong Zhao Jia Zheng Xinrui Zhao

The quality of Baobaoqu directly determines the flavor quality and yield of baijiu. In this work, metagenomic approaches combined with machine learning, including a multilayer perceptron (MLP) neural network model and the SHapley Additive exPlanations (SHAP) method, were employed to investigate the succession patterns and differential characteristics of microbial communities and potential enzymatic systems between two grades of Baobaoqu during storage. The results showed that Premium Baobaoqu possessed a more diverse microbial community than Normal Baobaoqu: a total of 1764 genera and 6901 species were annotated in Premium Baobaoqu, versus 1656 genera and 6440 species in Normal Baobaoqu. The top five dominant microbial genera, namely Weissella, Staphylococcus, Thermoactinomyces, Limosilactobacillus, and Pediococcus, collectively occupied 46.3–87.1% of the total microbial abundance. Ten microbial taxa were identified as key differential candidate microbial markers—including Weissella confusa, Kluyveromyces marxianus, and Staphylococcus lloydii—eight of which exhibited significantly higher abundance in Premium Baobaoqu. Glycoside hydrolases (GHs) constituted the largest enzyme class, accounting for 40.99% of the total. The top ten potential enzyme families significantly differentiated between them were GH43, GH31, GT9, GH2, CBM50, GH5, GH4, GT4, and GT26, with higher abundance of GH43, GH31, GH5, and CBM50 in Premium Baobaoqu. This study systematically uncovered the distinct microbial and enzymatic profiles of Baobaoqu of different qualities throughout storage. The findings provide theoretical guidance for the optimization of industrial production workflows and storage strategies for Baobaoqu manufacturing.

Fermentation doi: 10.3390/fermentation12090440

Authors: Yaling Zhu Rouf Ahmad Dar Xiaojie Mei Ning Fang Chen Sun Weixing Cao Ronghou Liu Adam Smoliński Le Zhang

The global fish industry is an essential contributor to food security and the economy; however, it produces considerable quantities of fish waste. Depending on species and processing methods, 20–80% of fish biomass is discarded as waste, amounting to almost 64 million tons annually. Conventional disposal methods such as landfilling, incineration, and wastewater discharge cause environmental pollution and greenhouse gas emissions. Therefore, for sustainable development, it is important to valorize fish waste into renewable energy and value-added products. Hence, this review aims to encourage the technical development of fish waste into bioenergy and bioresources by thoroughly studying key technologies for converting it into biogas, biodiesel, fertilizer, animal feed, and biochar. Although several studies have explored these technologies, most remain disjointed and lack a systematic assessment. Therefore, this work attempted to critically evaluate the technological principles, advantages, limitations, and optimization strategies of current valorization approaches. Furthermore, several recommendations have been proposed, including conducting pilot-scale trials, producing higher value-added products within a biorefinery system. Overall, this review provides a holistic outlook on the energy and resource conversion of fish waste, supporting sustainable waste management.

Fermentation doi: 10.3390/fermentation12090439

Authors: Zhicun Sheng Die Zhou Lin Niu Yi Zheng

A static magnetic field (SMF)-assisted liquid-state fermentation strategy using Lacticaseibacillus paracasei (L. paracasei) is proposed to improve the comprehensive utilization value of Ginkgo biloba leaves. Taking total flavonoid extraction yield as the objective, Box-Behnken response surface methodology was adopted to optimize the fermentation process. Meanwhile, the nutritional components and hypoglycemic activity of Ginkgo biloba leaves in vitro before and after fermentation were compared and analyzed. The optimal fermentation conditions were determined as follows: 3-fold-diluted De Man, Rogosa, and Sharpe (MRS) medium, an SMF intensity of 4.2 mT, an SMF duration of 2.9 h, and an inoculum size of 6.5%. Under these conditions, the total flavonoid extraction yield reached (3.41 ± 0.11)%, which was 3.33 times that of unfermented raw material. Analysis of the nutritional components revealed that after SMF-assisted fermentation, crude protein and crude fat contents increased by 59.58% and 13.72%, while ash and crude fiber contents decreased by 24.84% and 35.66%, respectively. SMF application also significantly affected the crude fiber content (p < 0.05). Meanwhile, the contents of phytic acid, tannins, phenolic acids, and ginkgolic acids in the SMF-assisted group declined by 29.56%, 43.10%, 23.42%, and 50.37%, respectively, and the SMF further promoted the degradation of anti-nutritional factors. Hypoglycemic activity analysis indicated that fermentation markedly enhanced the inhibitory effect of Ginkgo biloba leaf-derived flavonoids against α-glucosidase and α-amylase, and that SMF stimulation may also contribute to strengthening this inhibitory activity. This study demonstrates that SMF-coupled probiotic fermentation is an effective and environmentally friendly approach for processing Ginkgo biloba leaves, providing experimental support for high-value utilization of plant-based resources through physical-field-assisted microbial fermentation.

Fermentation doi: 10.3390/fermentation12090438

Authors: Bartłomiej Zieniuk

Non-conventional yeasts (NCYs) are promising microbial platforms for converting agro-industrial residues into value-added enzymes, yet their use for lipase production under solid-state fermentation (SSF) remains fragmented across the literature. This scoping review systematically mapped studies on lipase production by NCYs using agro-industrial residues under SSF. Scopus and Web of Science Core Collection were searched according to a PRISMA-ScR-based protocol, yielding 38 eligible original studies published between 1993 and 2025. Yarrowia lipolytica dominated the evidence base, accounting for 21 studies (55.3%), followed by Candida rugosa with six studies. Oilseed- and vegetable-oil-processing residues were used in 31 studies (81.6%), while cereal brans and fruit-processing by-products were also common substrates. Considerable variation was observed in substrate formulation, moisture, supplementation, fermentation time, optimization strategy, enzyme recovery, lipase assays, and activity units. This methodological heterogeneity precluded meaningful meta-analysis and limited direct comparison of reported lipase activities. Although several studies demonstrated applications in ester synthesis, lipid modification, polymer degradation, detergents, and wastewater treatment, most processes remained at the laboratory scale. Future development should prioritize standardized activity reporting, broader NCY diversity, improved reactor-scale control, and integrated downstream strategies to advance SSF-derived yeast lipases toward industrial application.

Fermentation doi: 10.3390/fermentation12090437

Authors: Yao Yan Xinyi Mao Fen Liu Shan Li

Ursodeoxycholic acid (UDCA) is the first-line therapy for primary biliary cholangitis and an important active constituent of bear bile. Whole-cell microbial conversion of inexpensive lithocholic acid (LCA) offers a promising alternative to conventional chemical synthesis owing to its high regioselectivity and mild reaction conditions, yet the fermentation process of Fusarium equiseti HG18 (CCTCC M2023160), a natural fungal catalyst for LCA 7β-hydroxylation, has not been systematically engineered for scalable production. This study developed a whole-cell process for UDCA production by F. equiseti HG18. Systematic optimization by single-factor experiments, Plackett–Burman design and Box–Behnken response surface methodology raised the shake-flask UDCA titer from 0.19 to 0.59 mg mL−1. Scale-up in a 3 L stirred-tank bioreactor shortened the fermentation time from 144 h in shake-flask cultivation to 96 h in the bioreactor under optimized operating conditions (pH 8.5, aeration 2.5 L min−1, and agitation 200 rpm). Experiments across a range of initial LCA loadings revealed a progressive decline in UDCA molar yield at elevated substrate concentrations, consistent with substrate-related inhibitory effects, and this loading-dependent behavior was used to design a two-stage fed-batch feeding strategy. Under the optimized condition (2.0 mg mL−1 LCA fed at 0 and 48 h), UDCA titer reached 1.71 mg mL−1, corresponding to a volumetric productivity of 17.8 mg L−1 h−1 and a UDCA molar yield of 41%. This study establishes a laboratory-scale whole-cell bioprocess for UDCA production from LCA using a wild-type fungal catalyst, integrating statistical medium optimization, bioreactor process development, and substrate-feeding strategies. The findings provide a practical framework for improving fungal whole-cell steroid biotransformation and highlight the potential of wild-type fungal platforms for scalable biocatalytic production.

Fermentation doi: 10.3390/fermentation12090436

Authors: Guangyao Hu Meng Wang Qingsheng Qi Junhong Yu Hua Yin Shumin Hu

Oxidative stress is a major physiological constraint on industrial lager yeast, compromising fermentation efficiency and flavor quality. Unsaturated fatty acids are known to influence membrane fluidity, but whether exogenous fatty acid supplementation can actively reprogram the yeast defense system, beyond serving as a passive membrane component, remains unclear. Here, we compared the effects of four fatty acids (palmitic, palmitoleic, oleic, and linoleic acid) on oxidative stress tolerance and fermentation performance in industrial lager yeast, under a defined chemical oxidative challenge (2.0 mM H2O2) in 15 °P wort, combining physiological assays with targeted gene-expression and untargeted metabolomic analyses; the dose was selected from a 0–2.0 mM gradient, with an ethanol-vehicle control included throughout. Among the fatty acids tested, palmitoleic acid (POA) most markedly enhanced oxidative stress tolerance, maintaining 91% cell viability, restoring intracellular pH to 6.1 by 24 h after transient acidification, and reducing ROS accumulation by 41.2%. Mechanistically, POA upregulated the antioxidant system, increasing catalase and glutathione peroxidase activities by 35.6% and 85.5%, respectively, and restoring glutathione levels by 35.5%. Metabolic profiling revealed a global reconfiguration, including a 2.6-fold increase in the stress-protectant proline and elevated pantothenate and coenzyme A levels, accompanied by a shift in the volatile profile toward esters, which rose from 26.1% to 60.3% of the total pool, alongside a 52.1% increase in total volatiles that did not reach significance after correction for multiple testing (q = 0.070); sensory evaluation confirmed higher fruity-estery intensity and lower soapy and staling notes in the POA beer. These findings identify POA as an active metabolic modulator, not merely a passive structural lipid, pointing to a non-transgenic nutritional strategy whose industrial value now requires validation under high-gravity, pilot-scale, and serial-repatching conditions.

Fermentation doi: 10.3390/fermentation12090435

Authors: Luca Nerva Walter Chitarra Alessandro Romano Giovanni Mian Lorenzo Lovat Raul Romor Diego Tomasi Tiziana Nardi

Grape-associated microbial communities are key contributors to wine quality and microbial terroir, yet their dynamics during withering, a critical post-harvest stage for premium sweet and fortified wine production, remain poorly characterized. Using high-throughput metabarcoding (16S rRNA and ITS), we monitored bacterial and fungal communities on Corvina grape berries throughout withering under controlled (C) and non-controlled (NC) dehydration conditions, integrating microbiome data with previously determined volatile organic compound (VOC) and stilbene profiles to uncover taxon–metabolite relationships. Microbial communities underwent progressive restructuring during dehydration. Moreover, the two withering conditions exhibited distinct dehydration kinetics, with C reaching 30% weight loss 14 days earlier than NC, with a significant effect on the abundance of several wine-relevant and spoilage-associated taxa. Metschnikowia was significantly more abundant under C conditions at 10% and 20% weight loss, while Botrytis reached 26.5% relative abundance at 30% weight loss under NC compared with 2.4% under C. A correlation network analysis identified significant associations between Lactobacillus, Acinetobacter, Clostridium, and metabolite accumulation, with Metschnikowia showing positive associations with bioactive compounds and negative associations with spoilage fungi. This first joint time-course analysis of mycobiome and bacteriome dynamics alongside metabolome changes during withering highlights significant associations between microbial succession and grape biochemical changes during withering.

Fermentation doi: 10.3390/fermentation12090434

Authors: Ahmed E. Kholif Abdelkader M. Kholif

The rumen is a complex microbial ecosystem in which anaerobic fermentation determines the availability of energy and protein to ruminant hosts and influences feed efficiency, animal productivity, and environmental emissions. This review integrates current knowledge of host–rumen microbiome interactions with particular emphasis on microbial fermentation, nutrient utilization, feed efficiency, methane (CH4) production, and sustainable ruminant production. We examine how dietary factors, including forage-to-concentrate ratio, carbohydrate fermentability, protein degradability, and lipid supplementation, alter microbial community structure, hydrogen metabolism, volatile fatty acid production, microbial protein synthesis, and nitrogen utilization. The review further evaluates microbiome-targeted strategies, including 3-nitrooxypropanol, red seaweeds (Asparagopsis spp.), nitrate, direct-fed microbials, and plant-derived bioactive compounds, with emphasis on their effects on fermentation pathways and methanogenesis; these strategies differ substantially in evidence base and mechanistic specificity, with 3-nitrooxypropanol supported by the most consistent mechanistic and in vivo evidence and several plant-derived compounds and direct-fed microbials showing more variable responses. Evidence from microbiome-wide and genome-wide association studies indicates that host genetics contributes to variation in rumen microbial composition and function, with potential consequences for feed efficiency and CH4 emissions. Host-side determinants of the rumen environment, such as feed intake, digesta passage rate, saliva production, and epithelial and immune function, further shape microbial responses. However, responses to microbiome-targeted interventions remain variable because of microbial functional redundancy, dietary context, adaptation, and host-specific effects. We therefore discuss the major constraints limiting the consistent translation of microbiome research into practical feeding strategies and propose an integrated framework combining functional microbiome indicators, precision nutrition, host–microbiome-informed selection, and real-time monitoring. Understanding and manipulating rumen microbial fermentation through coordinated nutritional and host-based approaches may provide a pathway toward improving ruminant productivity while reducing the environmental footprint of livestock production. Among the strategies reviewed, 3-nitrooxypropanol currently has the strongest and most reproducible evidence base, whereas plant-derived bioactives and several direct-fed microbials remain promising but inconsistent, and validated on-farm microbiome biomarkers remain a major gap.

Fermentation doi: 10.3390/fermentation12090433

Authors: Aleksandar Dolashki Lyudmila Velkova Tsvetan Berovski Maria Todorova Ventseslav Atanasov Maria Gerginova Ekaterina Krumova Nadya Armenova Penka Petrova Kaloyan Petrov Pavlina Dolashka

Bacillus velezensis is a promising candidate for the biocontrol of plant pathogens due to its production of broad-spectrum antimicrobial metabolites. The present study evaluates the antifungal potential and metabolic profile of the non-pathogenic rhizobacterial strain Bacillus velezensis 5RB. Modifying the mineral composition across three different growth media (A, B, and C) resulted in distinct secondary-metabolite profiles of B. velezensis 5RB. Notably, the free-cell supernatant of strain 5RB cultivated in medium A and C demonstrated the most potent antifungal activity against the gray mold pathogen Botrytis cinerea. For the first time, the antimicrobial metabolites produced by B. velezensis 5RB across these three media were comprehensively characterized using ultra-performance liquid chromatography-quantitative time-of-flight mass spectrometry (UHPLC-QTOF-MS). Mass spectrometric analyses (LC-ESI-MS) identified predominantly lipopeptides from the surfactin and fengycin families, alongside three classes of polyketides: macrolactins, difficidins, and bacillaenes. Multiple lipopeptide homologues, including a novel surfactin isoform (B*), were confirmed by tandem mass spectrometry (LC-ESI-MS/MS). Additionally, this study provides the first experimental evidence for subtilosin A production by B. velezensis 5RB. The multicomponent fermentation matrix of 5RB conferred robust biocontrol protection to tomato leaves against B. cinerea and Phytophthora infestans in an in vitro disease model, underscoring its strong potential for sustainable agricultural applications.

Fermentation doi: 10.3390/fermentation12090432

Authors: Esra Orenlili Yaylagul Adem Yavaş Mustafa Dikme Seda Gezgin Ecem Akan

Paraprobiotics offer safer alternatives to live probiotics without requiring cell viability. Here, heat-inactivated bacterial preparations (paraprobiotics) were produced from Lacticaseibacillus paracasei subsp. paracasei 431 (L. casei 431) using thermal treatments (65–121 °C). No viable colonies were detected by conventional plating in any treatment, and flow cytometry indicated that more than 96% of cells were membrane-compromised or dead. The most favorable condition (65 °C, 60 min) was identified using a TOPSIS multi-criteria ranking approach integrating loss of culturability with antimicrobial activity against Enterococcus faecalis and Escherichia coli with inactivation efficiency. This paraprobiotic and the corresponding probiotic were subjected to in vitro gastrointestinal digestion and colonic fermentation, followed by short-chain fatty acid (SCFA) and 16S rRNA-based gut microbiota analysis. The fecal inoculum was pooled from five healthy donors and used to run three replicate fermentation vessels per group. Both probiotic and paraprobiotic supplementation significantly increased acetate, butyrate, and total SCFA concentrations compared with the control, and propionate and valerate concentrations were also significantly higher in the paraprobiotic group than in the control. Both interventions increased Chao1 richness and Simpson diversity (p < 0.05), although Shannon diversity did not differ significantly, and induced significant shifts in beta diversity, alongside marked enrichment of butyrate- and SCFA-producing taxa such as Roseburia, Anaerostipes, and Akkermansia muciniphila. However, not all observed microbial changes were unequivocally beneficial; increases in Bilophila wadsworthia and Collinsella aerofaciens warrant cautious interpretation given their context-dependent associations with host health. Spearman correlation analysis identified positive associations between enriched taxa and SCFA concentrations, which should be interpreted as exploratory rather than evidence of causal metabolic cross-feeding. Under the conditions of this in vitro fermentation model, no statistically significant differences were detected between the viable and heat-inactivated preparations for most measured outcomes, indicating that thermal inactivation did not markedly diminish the capacity of L. casei 431 to influence gut microbial composition and short-chain fatty acid production. The multi-criteria selection strategy offers a practical approach for optimizing paraprobiotic production for functional food applications.

Fermentation doi: 10.3390/fermentation12090431

Authors: Raúl Ricardo Gamba Daniela Sedan Dario Andrinolo Angela León Peláez Marina A. Golowczyc

Aflatoxin B1 (AFB1), a potent carcinogenic metabolite produced by Aspergillus flavus and A. parasiticus, constitutes a significant threat to food safety. Conventional decontamination methods are costly or ineffective. Biological detoxification has emerged as a promising alternative. Kefir, a fermented milk beverage, comprises lactic acid bacteria (LAB) and yeasts with documented antifungal properties. This study evaluated whey permeate (WP) as a fermentation substrate for kefir microorganisms and assessed their AFB1-binding capacity. Kefir microorganisms cultivated in WP supported active fermentation, reaching 8.47 log CFU/mL LAB and 7.57 log CFU/mL yeasts, with concomitant lactose consumption and organic acid production. The ability of 13 individual kefir-derived microorganisms and the total kefir microbial community to bind AFB1 was evaluated under diverse conditions. All microorganisms exhibited AFB1 binding ability, although binding depended on strain, growth medium, pH, microbial concentration, and toxin concentration. The highest binding was observed for the total kefir community grown in WP. Binding remained stable over time for the total kefir community and Lactiplantibacillus plantarum CIDCA 83114 and was independent of viability and hydrophobicity. AFB1 binding by kefir microorganisms significantly reduced AFB1-induced cytotoxicity in HepG2 cells. These findings demonstrate that WP can be valorized into a value-added fermented product containing microorganisms with strong AFB1 binding and cytoprotective properties.

Fermentation doi: 10.3390/fermentation12090430

Authors: Qiting Wu Jiarui Zhao Huirong Zhu Yunle Liu Chaowan Guo Lin Ye

Sea rice harbors a diverse array of bioactive constituents, among which the efficient liberation of phytic acid is pivotal for unlocking its full functional potential. In this study, a co-fermentation system integrating yeast and lactic acid bacteria was established to generate a sea rice fermentation filtrate (SRF) with enhanced phytic acid (PA) content, aiming to broaden its application as a cosmeceutical ingredient and to facilitate the high-value utilization of sea rice. Response surface methodology was employed to optimize the inoculation ratios of the two microbial strains and the fermentation duration, and the PA levels were quantified using a commercial assay kit. The anti-melanogenic activity of SRF was assessed in murine B16 melanoma cells, and the underlying molecular mechanisms were elucidated with particular emphasis on the PI3K/AKT/GSK3β/MITF signaling cascade. Under the optimal conditions, specifically Saccharomyces cerevisiae (SC) inoculation at 4.5% and Lactobacillus plantarum (LP) at 6.0% with a fermentation time of 18 h, the PA concentration reached 576.61 μg/mL. Mechanistically, SRF treatment enhanced GSK3β phosphorylation, which was associated with reduced MITF phosphorylation and diminished the expression of downstream melanogenic enzymes, thereby effectively curtailing melanin synthesis, suggesting that SRF, as a complex fermentation product containing multiple bioactive constituents including PA, exhibits potent whitening efficacy. Collectively, these results provide novel perspectives for the valorization of sea rice and the development of natural skin-lightening agents, while also contributing to the extension of the sea rice industrial value chain.

Fermentation doi: 10.3390/fermentation12090429

Authors: Nathalia Fernandes Alessandra De Cesare Valentina Indio Ursula Gonzales-Barron Vasco Cadavez

Alheira is a traditionally smoked, fermented, non-ready-to-eat meat sausage produced in the Trás-os-Montes region of Portugal, where lactic acid bacteria (LAB) are major determinants of product safety and quality. This study used whole-genome sequencing to characterize 59 LAB isolates collected from artisanal alheira produced in six municipalities. Eight species belonging to six genera were identified, and average nucleotide identity analysis resolved 24 non-redundant strain groups. Functional annotation revealed broad repertoires of carbohydrate-active enzymes, proteases, lipases, transport systems, and genes associated with tolerance to acid, oxidative, bile, heat, and salt stress. Genotype–phenotype inference was performed with a subset of 22 strain groups represented by unflagged genome assemblies. After false-discovery-rate correction, carbon-metabolism, transport, energy-metabolism, total functional-gene, acid-stress, bile-stress, and total-stress counts were positively associated with total pH decline. A mixed-effects model showed a positive association between standardized total functional-gene count and acidification (b=0.137±0.059, p=0.049; marginal R2=0.266, conditional R2=0.582), but within–between decomposition indicated that the association was detectable between species rather than within species. Regularized regression, Random Forest, AICc model comparison, and partial least-squares regression converged on carbon metabolism, transport, energy metabolism, and acid- or bile-stress functions as the principal exploratory signals; however, sparse LASSO regression for variable selection was unstable, and prediction of unobserved species was limited. None of the five targeted pathways: acetaldehyde, citrate, diacetyl, exopolysaccharide, and lactose metabolism, remained significant after multiple-testing correction. These findings provide a genomic and phenotypic basis for selecting native starter-culture candidates while emphasizing the need for independent functional and safety validation.

Fermentation doi: 10.3390/fermentation12090428

Authors: Elisa Aiello Mattia Pia Arena Andrea Pulvirenti Maria Gullo

Non-Saccharomyces yeasts shape the aroma, freshness, and reduced-alcohol profiles of wines, yet their oenological contribution is curtailed by a poor tolerance to ethanol. Because ethanol acts on the plasma membrane, the cell’s selective barrier, this limitation is rooted in the membrane lipidome, whose remodeling has been documented but never integrated into a wine-specific, predictive framework. This review provides that synthesis, keeping the non-Saccharomyces wine yeasts as the subject and Saccharomyces cerevisiae as a reference frame. It first describes how ethanol perturbs the bilayer and how the cell compensates through unsaturated fatty acids, ergosterol, phospholipid-class balance, and membrane fluidity. It then traces the concentration- and time-dependent remodeling of the lipidome, together with the medium-chain fatty-acid, temperature, and oxygen co-stressors accompanying it, and the comparative membrane biology distinguishing these yeasts from S. cerevisiae, including the markedly reduced uptake of exogenous sterols under oxygen limitation. Finally, it weighs the nutritional, process, and non-genetically modified strain-improvement strategies that reinforce them. The same lipid descriptors recur throughout, yet are seldom measured in improved wine strains. We argue that the membrane lipidome can be read as a candidate multivariate predictive signature of ethanol tolerance and of fermentation performance, and outline the validation it still requires.

Fermentation doi: 10.3390/fermentation12090427

Authors: Tekmile Cankurtaran Kömürcü Miguel Angel Alvarez Gonzales İhsan Sarıoğlan Stephen R. Lindemann Yunus Emre Tunçil

This study investigated the effects of yeast-mediated pre-fermentation on the composition and functional characteristics of modern and ancient wheat genotypes, and their subsequent impact on gut microbial fermentation. Whole wheat flours were pre-fermented using Saccharomyces cerevisiae (6%, w/w) at 30 °C for 6 h. Raw and pre-fermented whole wheat samples were subjected to in vitro upper gastrointestinal digestion, followed by monosaccharide composition analysis, in vitro fecal fermentation using a pooled inoculum prepared from three healthy donors, microbial profiling, and short-chain fatty acid (SCFA) quantification. Fermentation increased Folin–Ciocalteu-reactive values and antioxidant capacity, while reducing residual starch and protein after digestion, indicating alterations in the composition of digestion-derived residues. After fermentation by S. cerevisiae, monosaccharide analysis revealed a relative enrichment of glucose, accompanied by decreased proportions of arabinose and xylose. Following in vitro fecal fermentation, pre-fermented wheat samples exhibited significantly (p < 0.001, AMOVA) different microbial community structures compared with their raw counterparts, based on Bray–Curtis dissimilarities, whereas differences in α-diversity were comparatively limited. In addition, taxa associated with carbohydrate fermentation and SCFA production, including Parabacteroides distasonis, Butyrivibrio crossotus, and Kineothrix, showed treatment- and genotype-dependent responses. All wheat samples supported SCFA production, although SCFA accumulation was more gradual than with inulin. Overall, yeast-mediated pre-fermentation was associated with changes in wheat composition and subsequent shifts in fecal microbial community structure and fermentation dynamics, with responses varying among wheat genotypes.

Fermentation doi: 10.3390/fermentation12090426

Authors: Gbeminiyi Olamiti Shonisani Eugenia Ramashia

Processing tubers such as potato, sweet potato, cassava, and yam, as well as legumes such as soybean, pea, chickpea, bean, and lentil, generates substantial quantities of peels, pulps, husks, and press cakes that retain considerable nutritional and functional value despite often being discarded. Microbial fermentation has emerged as a low-cost and scalable approach for valorising these residues into functional ingredients with improved digestibility and enhanced bioactive-compound content. This study synthesises the composition of major tuber and legume by-products, the solid-state and submerged fermentation systems employed for their bioconversion, and the biochemical and functional transformations achieved through fermentation, including improvements in antioxidant activity, prebiotic potential, antihyperglycaemic properties, and protein quality. The emerging applications of these fermented by-products in bakery products, dairy alternatives, snacks, nutraceuticals, and the conceptually related field of gut-microbiome-guided precision nutrition are also examined. Persistent challenges, including compositional variability, safety monitoring, and regulatory classification, are critically evaluated alongside strategies for addressing these limitations. Overall, fermentation-based valorisation of tuber and legume residues represents a scientifically credible and environmentally sustainable strategy for transforming agri-food waste streams into value-added, functional nutrition ingredients.

Fermentation doi: 10.3390/fermentation12090425

Authors: Victoria Kleinjan Melisa González Flores María Eugenia Rodriguez Christian Ariel Lopes

Torulaspora delbrueckii is a promising non-Saccharomyces yeast for beverage diversification, although its diversity and application in cider production remain poorly explored. This study characterized Patagonian T. delbrueckii isolates from wine- and cider-associated environments and evaluated their fermentative potential in apple must. Mitochondrial DNA-RFLP analysis showed that mitochondrial profiles varied according to the isolation source, while physiological characterization revealed differences in temperature, sulfite, and ethanol responses among isolates from different sources. Compared with wine isolates, cider isolates generally exhibited greater tolerance to high temperatures, lower sulfite tolerance, and shorter lag phases under ethanol stress. Eleven representative strains completed apple must microfermentations. Compared with a commercial Saccharomyces cerevisiae strain, T. delbrueckii produced higher glycerol and erythritol concentrations while achieving comparable or higher ethanol levels. Two contrasting strains were further evaluated in sequential mixed fermentations with S. cerevisiae. NPCC1340 enhanced glycerol and erythritol production, whereas NPCC1608 increased malic and succinic acids while maintaining low acetic acid levels. In mixed fermentations, ciders retained the characteristic metabolic profiles of each T. delbrueckii strain, whereas S. cerevisiae improved fermentation performance, producing intermediate fermentation kinetics. These findings reveal phenotypic diversity among Patagonian T. delbrueckii strains and suggest differences associated with their isolation sources, highlighting their potential as starter cultures for producing ciders with distinct chemical characteristics.

Fermentation doi: 10.3390/fermentation12090424

Authors: Aminur Rahman Pottathil Shinu JB Senthil Kumar Md Azizul Haque

Chlorpyrifos (CPF) is the most commonly used organophosphorus insecticide in agriculture globally. This has raised concerns due to its persistence, bioaccumulation, neurotoxicity, and environmental and human health effects. Until 2020, CPF was the most commonly used pesticide in European Union (EU) food production. CPF is also used in other parts of the world, though it has been discontinued in the EU, as national pesticide surveillance programs indicate it is found in soil, water, and food. This type of persistence is potentially harmful to farmers, consumers, and animals because CPF is toxic. Markedly, CPF has the potential to change the microbiota composition of soils, i.e., fungal, bacterial, and actinomycete communities, and inhibit the mineralization of nitrogen. The key CPF activity is associated with the inhibition of acetylcholinesterase (AChE), leading to reproductive, neurotoxic, and genotoxic effects. Microbial degradation, especially when applied by means of bacteria, has become one of the promising alternatives to the traditional physicochemical means since it is inexpensive, does not harm the environment and may possibly be fully detoxified. This review summarizes the latest developments in the study of CPF-degrading bacteria, enzyme pathways, microbial diversity, and the evaluation of the environmental impact of microbial remediation. Recent research findings, genomic research developments, and potential applications of CPF-degrading bacteria are addressed. In addition, this study reveals the current knowledge gaps, presents biotechnological challenges, and suggests future directions in the application of field-scale studies, focusing on the application of microbial solutions in sustainable agriculture.

Fermentation doi: 10.3390/fermentation12090423

Authors: Zhonghao Wang Hongyan Hou Weibing Zhang Wei Zhang Yulong Zhao Lianqing Wei Jie Cheng Yuxuan Jiang Feier Ren Jiajin Sun Qinghong Li Wenjie Zhang

Skatole is a harmful, odorous pollutant in livestock manure. Skatole-degrading strains are mostly harmful Gram-negative bacteria, whereas safe Gram-positive strains demonstrate poor degradation performance, limiting bioremediation applications. To solve this problem, this study was conducted to enhance the skatole degradation capacity of the food-grade strain Lactococcus lactis NZ9000 via heterologous expression of the skatole-degrading Ska enzyme from Acinetobacter piscicola p38. Three gene sequences, designated Ska-Y (original sequence from A. piscicola p38), Ska-D (E. coli codon-optimized), and Ska-R (L. lactis codon-optimized), were separately expressed using constitutive pMG36e and nisin-inducible pNZ8148 plasmids. The constitutive system only transcribed mRNA but produced misfolded, nonfunctional inclusion bodies. By contrast, the codon-optimized pNZ8148-Ska-R strain exhibited prominent skatole degradation ability, even under non-inductive conditions. Under optimal conditions (30 ℃, ultra-low nisin induction), the engineered strain increased the 24 h degradation rate of 50 mg/L skatole from 20% to 88% and completely degraded 25 mg/L skatole. Field tests verified that the strain effectively reduced skatole accumulation in manure compost and lagoon manure. This study achieved efficient functional heterologous expression in Gram-positive bacteria, provides new insights into ultra-low-dose induction and codon optimization, and offers a safe and efficient microbial agent for livestock manure odor remediation.

Fermentation doi: 10.3390/fermentation12090422

Authors: Vicente Barros Maribel Mamani Benjamín Castillo Sara Cuadros-Orellana Nidia Torres-Ponce Cristian Valdés

Native Lactic Acid Bacteria (LAB) isolated from fruits and vegetables in Chile’s Maule Region were identified as Lactiplantibacillus plantarum and L. pentosus. Their growth and lactic acid production were evaluated under furan inhibitor stress (furfural and 5-methylfurfural (5-MF)) relevant to agroindustrial waste valorization. In fruit hydrolysates containing natural furans (11–57 mg/L), LAB grew similarly to inhibitor-free controls and produced 17–25 g/L lactic acid. Under controlled conditions, furfural at 12 g/L reduced bacterial growth by over 80% and decreased lactic acid production to only 2 g/L. In contrast, 5-MF at the same concentration allowed substantially higher tolerance, with up to 7 g/L lactic acid. Benchmarked directly against the commercial reference strain Lactiplantibacillus plantarum PL8014 under identical conditions, the native isolate B1 matched or exceeded PL8014’s lactic acid output in most furfural and 5-MF concentrations tested, including the two most severe 5-MF conditions. Logistic growth and Luedeking–Piret models accurately described both bacterial growth dynamics and lactic acid production kinetics under all inhibitory conditions (R2 > 0.96). Inhibitor concentration proved more critical than inhibitor type, although furfural consistently showed stronger potency. These findings confirm the biotechnological potential of native Maule LAB and provide quantitative kinetic parameters for optimizing lactic acid production from lignocellulosic waste, supporting circular economy strategies.

Fermentation doi: 10.3390/fermentation12090421

Authors: Zekeriya Safa İnanç Mustafa Aslan Battal Yılmaz Deniz Şişman Erhan Çetin Oğuzhan Kahraman Mustafa Selçuk Alataş Huzur Derya Arık

Background: Plant flavonoids are candidate alternatives to ionophores for modulating rumen fermentation, yet the intact glycoside rutin (quercetin-3-O-rutinoside) has received little attention. The objective of this study was to determine whether rutin changes the in vitro ruminal fermentation of total mixed rations (TMR) differing in forage-to-concentrate ratio; monensin was included at a single dose as a reference additive rather than a direct comparator. Methods: In a 2 × 4 factorial batch-culture experiment, a roughage-based (60:40) and a concentrate-based (40:60) TMR were incubated for 24 h without additive (control) or with rutin at 2.5% or 5% of substrate dry matter or monensin at 30 ppm. Results: Rutin did not change the gas production kinetics, ruminal pH, the volatile fatty acid (VFA) profile, or the stoichiometrically estimated methane and carbon dioxide at either dose (p > 0.05), and no dose response was found. Substrate type likewise left the VFA profile and the estimated gases unchanged (p > 0.05). Ammonia nitrogen (NH3-N) responded to the additive group (p = 0.027) and to the substrate × additive interaction (p = 0.022), but Tukey comparisons separated no individual means, so this response is preliminary. Conclusions: Neither rutin nor monensin markedly altered fermentation of these silage-, alfalfa- and cereal-based rations in the 24 h batch system; the substrate-dependent NH3-N signal warrants confirmation, and direct methane measurement is required before any anti-methanogenic potential can be claimed.

Fermentation doi: 10.3390/fermentation12090420

Authors: Şükrüye Selin Koyun Gamze Nur Müjdeci

In this study, the potential of oilseed processing residues, namely plum seed pulp, pumpkin seed pulp, pomegranate seed pulp, wheat germ pulp, and coconut pulp, was evaluated as alternative substrates for pulcherrimin production by indigenous yeast strain Metschnikowia pulcherrima ELM-GS-3. Among the tested substrates, wheat germ pulp supported the highest biomass and pulcherrimin concentrations and was therefore selected for process optimization. The effects of temperature, agitation rate, fermentation time, and initial pH on pulcherrimin concentration were investigated using Response Surface Methodology (RSM) based on a Central Composite Rotatable Design (CCRD). The developed quadratic model was statistically significant (p < 0.0001) with an R2 of 0.9441. The numerically selected conditions were determined as 25 °C, 71 rpm, 12 days, and an initial pH of 2 where pulcherrimin concentration reached 5.5 ± 0.3 g/L, representing a 1.79-fold increase compared with the non-optimized process. Bioprocess performance analysis revealed a volumetric productivity (Qp) of 0.459 g/L/day, a specific productivity (qp) of 0.029 g/g/day, and a product yield coefficient (YP/X) of 0.346 g/g biomass. The experimentally obtained value was in good agreement with the model prediction. To the best of our knowledge, this is the first report evaluating wheat germ pulp as a substrate for pulcherrimin production by M. pulcherrima. The results demonstrate the potential of using wheat germ pulp, an oilseed processing residue, as a substrate for natural pigment production and provide a basis for further studies on process sustainability, scale-up, downstream processing, and economic feasibility.

Fermentation doi: 10.3390/fermentation12090419

Authors: Griselda Méndez-Marcial José Alfredo Carrillo-Salazar Alejandra Miranda-Carrazco Martha Hernández-Rodríguez

Pulque is a traditional Mexican beverage produced by the spontaneous fermentation of “aguamiel”, the sap of several Agave species. Fermentation is driven by adding a previously fermented inoculum, locally known as “semilla”, whose microbial community contributes to the sensory and physicochemical properties of the beverage. The objective of this study was to provide a preliminary characterization of the microbial taxonomic composition and functional potential of pulque inoculant prepared from Agave mapisaga and Agave salmiana using shotgun metagenomic sequencing. Six inoculum samples were sequenced on the DNBSEQ using 150 bp paired-end reads. Metagenomic DNA was extracted using a CTAB-based protocol and analyzed in the Galaxy platform. The workflow included quality filtering, host-sequence removal, taxonomic classification with Kraken2, assembly with MEGAHIT, and functional annotation with eggNOG Mapper. Bacterial communities in inoculum from A. mapisaga and A. salmiana sap were dominated by Acetobacter (68.1% and 58.3%) and Leuconostoc (19.5% and 21.9%). Komagataeibacter was more abundant in A. mapisaga inoculum (3.8%), whereas Zymomonas was more abundant in A. salmiana inoculum (11.7%). The greatest species-level difference was observed for Zymomonas mobilis, whose mean relative abundance was 6.4-fold higher in A. salmiana. Fungal communities were dominated by Saccharomyces (91.4% and 72.7%) and Kluyveromyces (6.6% and 25.5%) in A. mapisaga and A. salmiana, respectively. Across all replicates, the most abundant species were Acetobacter sp. AC2005 (23.9%), Saccharomyces paradoxus (17.0%), and Zymomonas mobilis (12.9%), together accounting for approximately 54% of the total relative abundance. Kluyveromyces marxianus was 3.9-fold more abundant in A. salmiana inoculum, whereas Saccharomyces paradoxus was 1.26-fold more abundant in A. mapisaga inoculum. Alpha-diversity analysis indicated higher bacterial diversity in A. mapisaga inoculum, with a Shannon index of 2.74 and 35 exclusive species, whereas A. salmiana inoculum showed greater fungal diversity, with a Shannon index of 0.68. These differences were not statistically significant (p > 0.05). However, beta-diversity analysis suggested substantial separation between the microbial communities associated with the two Agave species (R2 ≈ 0.92). Functional annotation identified genes potentially associated with carbohydrate metabolism, sucrose degradation, and secondary metabolite biosynthesis. These findings suggest that the agave species used as the sap source may influence the taxonomic composition and functional potential of microbial communities involved in pulque fermentation.

Fermentation doi: 10.3390/fermentation12090418

Authors: Meng Chai Rui-Tao Yu Yi-Tong Li Feng-Qing Wang Dong-Zhi Wei

2-Pyrone-4,6-dicarboxylic acid (PDC) is a promising pseudo-aromatic bio-based monomer for the production of biodegradable polyesters. Here, we combined metabolic engineering with fermentation process optimization to establish efficient de novo PDC biosynthesis in Corynebacterium glutamicum. To enhance the availability of protocatechuic acid (PCA), the direct precursor of PDC, endogenous PCA degradation was blocked, and precursor supply through the shikimate pathway was strengthened. Subsequently, the ligABC cluster from Sphingomonas paucimobilis SYK-6 was heterologously introduced to enable PDC formation from glucose. Medium screening identified a corn steep liquor-containing formulation that increased the PDC titer by 1.4-fold in shake-flask cultures. Further optimization of corn steep liquor concentration, dissolved oxygen, and glucose feeding rate in a 5-L bioreactor increased the PDC titer to 71.1 g/L at 90 h, with an average productivity of 0.8 g/L/h, the highest productivity for C. glutamicum-based PDC production reported to date.

Fermentation doi: 10.3390/fermentation12090417

Authors: Eren Kuter Derya Merve Karagöz Muhammad Shazaib Ramay Anas Tahir Syed Umer Akhter Bekir Tosun Ifrah Raza Murat Er Dilan Kuter Umair Ahsan

Triticale, a hybrid cereal crop derived from the cross of wheat and rye, is widely used for silage production owing to its high nutritional qualities and tolerance to varying climatic conditions. The genotype and maturity stage of forage crops govern the efficiency of microbial inoculants used for ensiling. The effects of homofermentative (HMF; Lactiplantibacillus plantarum and Enterococcus faecium) and heterofermentative (HTF; Lentilactobacillus buchneri) inoculants, either alone or in combination, on the fermentation quality, nutrient composition, and in vitro rumen digestibility of ensiled triticale varieties (Alperbey and Karma 2000) were assessed in this study. The study was carried out using a 2 × 2 × 2 factorial arrangement of two triticale varieties, two levels of HMF [0 or 1 × 105 CFU/g fresh forage (0.8 mg/kg)], and two levels of HTF [0 or 0.5 × 105 CFU/g fresh forage (500 mg/kg)]. Forages were harvested at the early dough stage, vacuum-packed in 110 μm-thick plastic bags (35 cm × 25 cm), and allowed to ferment for 120 days. Fermentation characteristics, chemical composition, volatile fatty acids, and digestibility traits were evaluated after ensiling. Application of HMF inoculant significantly reduced silage pH compared to the control and HTF treatments in Alperbey, whereas in Karma 2000, terminal pH remained statistically similar between control and HMF treatments. Generally, inoculated silages had lower ammonia nitrogen levels than control silages, reflecting better protein conservation. The HTF significantly increased the acetic acid and decreased the butyric acid concentrations. There were significant two-way (variety × HMF and variety × HTF) and three-way (variety × HMF × HTF) interactions for crude protein content, fiber contents, volatile fatty acids profile, and digestibility characteristics (p < 0.05). For Karma 2000, the application of HMF increased crude protein concentration; however, it also increased lignin and fiber contents, thereby reducing organic matter digestibility. In contrast, dual inoculation raised neutral detergent fiber digestibility. The results show that variety-specific inoculant selection can improve triticale silage quality, underscoring the importance of aligning microbial additives with the forage’s biochemical properties.

Fermentation doi: 10.3390/fermentation12090416

Authors: Évellin do Espirito Santo Ana Carolini Fernandes Mota Rafael Boffo Agatha Gonçalves Araújo Julia Bolognesi Andrade Stephanie França Carneiro Aline Kirie Gohara-Beirigo Maria Clara Arco e Flexa Fortuna Aline Mello Carvalho Daniel Pecoraro Demarque Livia Seno Ferreira-Camargo João Carlos Monteiro de Carvalho

This study assessed the effects of aqueous extracts from turmeric (Curcuma longa), mint (Mentha sp.), grape peel (Vitis sp.), and mango peel (Mangifera indica) on the growth and total carotenoid content of the microalga Dunaliella salina. A two-stage cultivation strategy was employed, in which aqueous extracts prepared by decoction were added at 0.5% (v/v) either at the inoculation or after the exponential growth phase. Microalgal growth was monitored by direct cell counting, and total carotenoid content was determined spectrophotometrically in the harvested biomass. The chemical profiles of the extracts were characterised by HPLC-MS/MS, enabling the annotation of key phenolic and flavonoid compounds. Specific orange turmeric and mint formulations enhanced microalgal cell proliferation by up to 31.88% and 28.95%, respectively, compared to the control. However, despite the higher biomass, the total carotenoid content did not increase correspondingly; orange turmeric extract even led to reductions of 90.84 to 92.86% in carotenoid content. These findings suggest that exogenous antioxidants may suppress the oxidative stress signals required to trigger carotenoid biosynthesis in D. salina. Additionally, grape peel extracts prepared at higher temperatures promoted better algal growth than those obtained at lower temperatures. These findings highlight the dual role of plant extracts in modulating microalgal growth and secondary metabolism, demonstrating that antioxidant-rich extracts may be used to enhance biomass accumulation before the subsequent induction of stress-mediated carotenogenesis, thereby potentially improving biotechnological yields.

Fermentation doi: 10.3390/fermentation12090415

Authors: Marina Edith Navarro Emma Elizabeth Tymczyszyn Carolina Soledad Martinez María Jimena Prieto Bárbara Mercedes Bravo-Ferrada

Wine lees (WL) are one of the main byproducts generated during winemaking and are usually disposed of in landfills or incorporated into the soil as an organic amendment; nevertheless, these practices can have negative environmental impacts. Reusing agro-industrial byproducts is a key step toward more sustainable production cycles, and WL represent a promising alternative for formulating culture media for the production of lactic acid bacteria (LAB) biomass, microorganisms widely used in the food and wine industries. In this work, we aim to evaluate the ecotoxicity of WL using a bioassay with zebrafish (Danio rerio) embryos and to analyze their potential as a source of nitrogen and growth factors for the production of UNQLp 11 Lactiplantibacillus plantarum strain biomass in whey permeate. Trials conducted with zebrafish showed that embryos exposed to WL exhibited increased mortality in a concentration-dependent response and the appearance of sublethal effects, such as alterations in embryonic development. Regarding WL biotechnological application, the results obtained confirm that the growth of Lpb. plantarum depends largely on the composition of the culture medium. Whey permeate media (WP) without supplementation showed nutritional limitations, reflected in lower cell counts. While supplementation with WL significantly improved bacterial growth, in some cases reaching values comparable to those obtained with MRS. The results demonstrated that WL can act as an efficient nutrient source, with the potential to replace commercial yeast extract in Lpb. plantarum culture media.

Fermentation doi: 10.3390/fermentation12090414

Authors: Vicente Peña-Caballero Pablo Antonio López-Pérez María José Enríquez-Arredondo Elizabeth Quintana-Rodríguez Adán Topiltzin Morales-Vargas José Luis Zárate-Castrejón

A formal framework of bioprocesses enables accurate prediction of reaction outcomes, thereby optimizing resource allocation and reducing production costs. This study aimed to establish an approximate stoichiometric equation and determine the bioenergetics growth parameters of the pink oyster mushroom (Pleurotus djamor) using a “black box” modeling approach. A commercial strain was cultivated in polypropylene bags at 28 °C and 75% relative humidity. The harvested mushroom biomass was dried and analyzed for C, H, and N content, with O determined by difference. The resulting empirical formulas were CH1.33O0.36N0.02 for the dry wheat straw substrate and CH1.81O0.41N0.09 for the fungal biomass. The bioprocess exhibited a primordia initiation period of 20.5 days, a total harvest window of 50.0 days, a maximum biological efficiency of 16.77%, a model yield (Y) of 0.90%, and a productivity of 20.5 g/100 g substrate. In conclusion, this biotechnological framework provides a robust predictive tool for industrial scaling, enabling mass and energy balance optimization in real time without reliance on costly intracellular measurements. Thus, it establishes a reliable and sustainable pathway to convert low-cost agricultural residues into high-value bioproducts, supporting the goals of a circular economy.

Fermentation doi: 10.3390/fermentation12090413

Authors: Bereket Zeleke Tunkala Kristy DiGiacomo Pablo S. Alvarez Hess Jeremy J. Cottrell Frank R. Dunshea Brian J. Leury

Excessive protein degradation reduces protein-use efficiency in ruminants. This in vitro study evaluated brown-sorghum-bran extract (SBE; 20 or 40 g/kg DM), BioProtect (15 or 30 mL/kg DM), and tannic acid (30 g/kg DM; positive control) in soybean and canola meals. Gas production and ammonia-N were assessed over 24 h; protein digestion and amino acid concentrations were also determined. Gas production and ammonia-N were generally lower in treated meals, with responses dependent on substrate, additive and incubation time (p ≤ 0.003). Substrate × additive interactions affected RDP, RUP and IVIDP (p < 0.001). The higher inclusion rates, 40 g/kg DM SBE and 30 mL/kg DM BioProtect, did not differ from tannic acid in RDP, RUP or IVIDP within either substrate. After in vitro intestinal digestion, total EAA showed a substrate × additive interaction (p = 0.033); 30 mL/kg DM BioProtect and 40 g/kg DM SBE produced among the highest total EAA concentrations in both substrates. Among the individual acid-hydrolysable EAA recovered from the liquid fraction after in vitro intestinal digestion, only methionine concentration was affected by additive treatment (p < 0.001). Overall, 30 mL/kg DM BioProtect and 40 g/kg DM SBE produced the most consistent responses for RDP, RUP and IVIDP; however, total digested CP decreased slightly (p < 0.001), and amino-acid responses were variable. In vivo studies are required to determine effects on amino-acid supply and animal responses.

Fermentation doi: 10.3390/fermentation12090412

Authors: Yaru Feng Mengjie An Jie Cao Ruirong Li Jinling Cai

Organic waste causes severe pollution, while agriculture lacks quality fertilizers. Solid-state fermentation solves both issues. Nevertheless, traditional methods are slow and lead to nitrogen loss. They also emit greenhouse gases and yield uneven products. This fails to reach modern low-carbon standards. We urgently need efficient solid-state fermentation systems. Therefore, this review summarizes recent advances in parameter optimization and cutting-edge innovations. Firstly, key solid-state fermentation operational parameters, such as carbon-to-nitrogen ratio, aeration frequency, temperature, humidity and pH, are analyzed. The optimization of these parameters facilitates nitrogen retention and mitigates pollutant emissions. Secondly, emerging enhancement strategies are discussed. Elaboration on the electron-shuttle effect of modified biochar and advances in synthetic microbial consortia is provided. For process coupling, the mechanisms of hydrothermal carbonization (HTC) combined with solid-state fermentation are explored. Bioelectrochemically assisted solid-state fermentation (MCFT/BFC) based on Direct Inter-Species Electron Transfer (DIET) is also examined. Furthermore, feasible mitigation approaches for pollutants including greenhouse gases and antibiotic resistance genes (ARGs) are summarized. Subsequently, the applications of mathematical models, the Internet of Things (IoT), and artificial intelligence in solid-state fermentation are introduced. Finally, current challenges in large-scale application and risk management are analyzed. Future prospects involving multi-omics, life cycle assessment (LCA), and functional customization are discussed. Low-carbon solutions for high-value waste utilization are provided in this review.

Fermentation doi: 10.3390/fermentation12090411

Authors: Peng Wang Mengli Han Qian Zhang Bin Zhang Jingzhe Wang Yan Zheng Wenjuan Zhao Ning Chen

This study used pepper straw as the fermentation substrate to investigate the effects of different molasses addition levels on the nutritional quality, fermentation quality, and microbial community of pepper straw in micro-storage. A control group (CON) and groups with different molasses addition levels (PA (2.0%), PB (4.0%), PC (6.0%)) were established. The results showed that the addition of molasses effectively reduced the dry matter (DM) loss rate in micro-storage of pepper straw. Compared with the control group, the 4% and 6% treatment groups significantly reduced the dry matter (DM) loss rate. In the 6% treatment group, crude protein (CP) content increased by 1.51 percentage points; at the same time, it significantly increased the relative abundance of Lactobacillus in the fermentation system, lowered the pH, and increased the concentrations of lactic acid (LA) and acetic acid (AA), with the 6% treatment group exhibiting the lowest pH value of 4.18. In summary, the optimal addition range is 4.0–6.0%. Considering production costs, we recommend an addition level of 4.0%.

Fermentation doi: 10.3390/fermentation12090410

Authors: Caner Caliskan Celia Segura Godoy Arianna Ressa Aylin W. Sahin Emanuele Zannini Laura Nyhan Elke K. Arendt

Brewers’ spent grain (BSG) is a brewing by-product with potential as a sustainable protein source. However, its limited amino acid profile and weak gelation properties limit its use in plant-based yoghurt alternatives (YAs). This study evaluated the compositional and techno-functional properties of brewers’ spent grain protein isolate (BSGI), pea protein isolate (PPI), mung bean protein isolate (MBP), and chickpea protein concentrate (CPC), and investigated their suitability in fermented YAs. BSGI was deficient in lysine, while PPI and MBP were limited in sulfur amino acids, highlighting the potential nutritional complementarity of cereal-legume protein blends. BSGI was blended with each legume ingredient at a fixed 50:50 ratio and fermented to a pH of 4.5 using a Streptococcus thermophilus culture. All YAs contained over 3.0% protein and showed improved indispensable amino acid profiles. The BSGI + MBP blend demonstrated the highest water retention, firmness, consistency, and rheological strength, followed by BSGI + PPI, whereas BSGI + CPC had the weakest gel structure. These formulation-level differences cannot be attributed solely to the intrinsic functionality of the individual protein ingredients, as protein content and non-protein components, particularly dietary fibre, differed among formulations. Overall, BSGI shows potential for incorporation into mixed plant-protein YAs and supports further optimisation of sustainable cereal-legume protein systems.

Fermentation doi: 10.3390/fermentation12090409

Authors: Xiaolong Li Weiliang Qi Xiaoyan Zhang Zhilong Yao Qian Li Yulong Bai Zongbing Zhan

Aerobic co-composting of spent mushroom substrate (SMS) and cattle manure is a vital technology for agricultural waste recycling. However, high refractory lignocellulose content and low indigenous microbial activity usually hinder compost heating and maturation. Exogenous inoculation and raw material sterilization are common microbiome regulation strategies, while their divergent functional mechanisms and the contributions of native versus exogenous microbiota remain unclear. Herein, three treatments (control with intact indigenous microbiota, sterilization pretreatment, and 0.5% w/w fungal–bacterial inoculation) were established to explore their effects on composting performance and fungal community succession over 34 days. Compost stability and maturity were evaluated via thermophilic duration and germination index (GI), with fungal communities analyzed by ITS high-throughput sequencing. The results revealed that sterilization severely suppressed composting, causing delayed heating, a low peak temperature of 55.2 °C, inhibited lignocellulose degradation, and disordered fungal community structure with increased richness but decreased evenness due to random colonization by weak degraders. In contrast, microbial inoculation prominently improved composting efficiency, with a peak temperature of 67.5 °C, eight days of sustained high temperature, and a final GI of 89.67%. The degradation rates of cellulose, hemicellulose and lignin reached 68.32%, 72.15% and 35.28%, respectively. Inoculation directionally enriched core lignocellulose-degrading fungi (Chaetomium, Aspergillus fumigatus, Thermoascus) and maintained functional community balance. Core fungal genera dominated lignocellulose decomposition and humification driven by environmental factors. This study clarifies that indigenous microbiota underpin spontaneous composting, while targeted inoculation optimizes functional microbiota, and single sterilization impairs composting. It provides mechanistic guidance for efficient agricultural waste composting.

Fermentation doi: 10.3390/fermentation12090408

Authors: Dilara Yeniterzi Mustafa Selçuk Alataş

Pulse-processing plants generate large volumes of screenings, that is, undersized, broken and foreign-matter-contaminated grain that is removed before the material enters the human food chain. Their feeding value for ruminants is still poorly documented. This study characterised the chemical composition, Cornell Net Carbohydrate and Protein System (CNCPS) nitrogen fractions, in vitro ruminal dry matter degradability (IVDMD) and rumen fermentation behaviour of screenings from five pulse species: soybean, dry bean, chickpea, green lentil and red lentil. Six independent batches per species (n = 30) were obtained from processing plants in different regions of Türkiye. Total digestible nutrients (TDNs) and energy values were estimated with NRC (2001) equations, gas production was recorded for 48 h in a semi-automatic modular system and fitted to a logistic model, IVDMD was measured in a DaisyII incubator and volatile fatty acids (VFAs), pH and NH3-N were determined after 24 h of incubation. Differences were declared at p < 0.05 throughout. Soybean screenings had the highest crude protein (35.99%), recalculated TDNs (75.26%) and 48 h gas production (65.16 mL per 460 mg of incubated sample), a ranking that largely disappears once gas is expressed per gram of incubated organic matter; chickpea screenings combined the highest starch (50.67%), IVDMD (71.27%) and fractional degradation rate. Dry bean screenings fermented most slowly, with a lag time of 10.46 h. Acetate, propionate and total VFAs did not differ among species. No parent grain was analysed alongside the screenings, so any comparison with clean pulse grain rests on published values for other samples and cultivars. Within that limit, the screenings carry enough protein, calculated energy and rumen-undegradable protein to warrant testing as partial replacements for conventional concentrates in feeding trials.

Fermentation doi: 10.3390/fermentation12090407

Authors: Ling Hua Yujie Deng Qinlin Yang Jingzhu Zhao Min Song

To explore coffee’s effect on sweet rice wine quality, this study developed a novel fermented beverage using round glutinous rice and coffee powder as raw materials. The fermentation process was optimized using an orthogonal array design combined with fuzzy comprehensive evaluation. Flavor, physicochemical properties, and antioxidant activities were systematically evaluated via electronic tongue (E-tongue), gas chromatography–mass spectrometry (GC-MS), and gas chromatography–ion mobility spectrometry (GC-IMS). The optimal fermentation parameters were 0.6% fermentation starter (Jiuqu), 10% coffee powder, a fermentation time of 56 h, and a temperature of 28 °C. Adding coffee powder significantly increased total flavonoid content. Compared to the control group, the scavenging activities against 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS+), as well as hydroxyl radicals, increased by 18.36%, 7.40%, and 10.96%, respectively, compared to the control group. Flavor analysis identified 173 volatile organic compounds, demonstrating strong complementarity between GC-MS and GC-IMS. The addition of coffee powder preserved traditional sweet rice wine components while introducing roasted heterocyclic compounds, such as 2-methylpyrazine and 2-ethylfuran, thereby imparting nutty and roasted characteristics. E-tongue analysis revealed enhanced sensor responses for sweetness, bitterness, and saltiness, along with decreased umami. In conclusion, the addition of coffee powder is an effective strategy to enhance the in vitro antioxidant activity and enrich the flavor profile of sweet rice wine. This study provides a scientific basis for the application of coffee in traditional fermented foods.

Fermentation doi: 10.3390/fermentation12090406

Authors: Scott Simonin Valentina Bianconi Federico Sizzano Christine Monnard Sylvain Schnée Gilles Bourdin Esteban Alfonso Benoit Bach

The phenomenon of global warming has been demonstrated to induce the premature ripening of grapes, resulting in a concomitant increase in sugar and a decrease in acidity. This can result in the production of wines that are characterised by an imbalance in terms of alcohol content and acidity. The utilisation of lactic acid-producing Lachancea thermotolerans yeast in fermentation represents a promising strategy for reducing the pH of must without the need for chemical acidification. However, it is important to note that the fermentation rate and lactic acid production can be influenced by several factors present in the must. The present study focused on the effect of copper, used in organic vineyards to limit mildew, on biomass and lactic acid production by two L. thermotolerans oenological strains in laboratory-scale fermentations. The methodology used is based on an innovative integrative approach combining conventional microbiological, spectroscopic, cytometric and transcriptomic analyses. The results obtained demonstrated that the addition of copper increased lactic acid production in comparison with the control condition and this effect was strain-dependent, reaching up to 16.15 g/L for strain Lt1 under conditions of high copper concentration (20 mg/L). The monitoring of microbial populations demonstrated discrepancies in the fermentation kinetics in conditions exhibiting varying concentrations of copper and showed that an increased duration of cell generation is evident under conditions exhibiting elevated copper concentrations, particularly in the context of the Lt1 strain. Furthermore, the analysis of the samples using flow cytometry revealed the oxidative status of the two strains throughout the fermentation process. Transcriptomic data indicated that metabolic pathways involved in cell communication, ribosomal and cell wall activity of L. thermotolerans were modulated by the presence of copper. The findings of this study demonstrate strain-specific effects of copper on L. thermotolerans, which may be of interest to winemakers, particularly those engaged in organic viticulture who intend to use acidifying yeasts during warm vintages.

Fermentation doi: 10.3390/fermentation12090405

Authors: Cristian Avilés-Ramírez Nelda X. Martínez-Galero Diana I. Orbe-Díaz Natividad Castro-Alarcón Yanet Romero-Ramírez Verónica I. Martínez-Santos Arturo Ramírez-Peralta Roxana Reyes-Ríos Alberto Patricio-Hernández Jeiry Toribio-Jiménez

Spontaneous fermentation is central to ancestral mezcal production, yet the bacterial and fungal communities associated with Agave cupreata must remain insufficiently characterized. This exploratory study described community composition and predicted functional profiles at three stages of a traditional fermentation defined by sugar depletion: 10 °Brix (0 h), 5 °Brix (72 h), and 2 °Brix (144 h). Ambient temperature ranged from 21.7 to 23.7 °C, must temperature from 28.5 to 29.9 °C, pH from 4.6 to 5.2, and relative humidity from 51.3% to 58.7%. At each stage, material collected from two wooden vats and three vertical positions was pooled into one composite sample. Bacterial 16S rRNA gene and fungal ITS1 amplicons were sequenced and analyzed using QIIME 2. At 10 °Brix, five dominant bacterial genera accounted for more than 93% of relative abundance. Lactic and acetic acid bacteria represented more than 91% at 5 °Brix, while Pediococcus, Lactiplantibacillus, and Loigolactobacillus were prominent at 2 °Brix. Seven fungal genera represented more than 98% of the initial profile, whereas Saccharomyces, Debaryomyces, and Maudiozyma accounted for more than 98% at 2 °Brix. PICRUSt2 predicted pathways related to carbohydrate degradation and fermentation, whereas FUNGuild assignments were predominantly saprotrophic at the later stages. Because one pooled sample was analyzed per stage, these patterns are descriptive and require validation in replicated fermentations. Despite these limitations, this study provides baseline information on the microbial communities associated with ancestral A. cupreata fermentation and identifies taxa and predicted functions that merit future isolation and experimental validation.

Fermentation doi: 10.3390/fermentation12090404

Authors: Meng Song Huangfeng Qiu Donghua Tan Jingqi Liu Yuting Liang Yu Yang

Citric acid is one of the most effective organic acids for rare earth bioleaching. Yarrowia lipolytica represents a promising chassis due to its inability to produce oxalic acid; however, the metabolic bottlenecks between citrate synthesis and mitochondrial export remain to be systematically addressed. In this study, a heterologous “push–pull” strategy was applied to simultaneously reinforce citrate synthesis and mitochondrial efflux by co-overexpressing YlAMPD (encoding AMP deaminase) and AnYHM2 (encoding a mitochondrial citrate carrier from Aspergillus niger) under the strong constitutive promoters. Three recombinant strains were constructed in Yarrowia lipolytica Po1f: YlAMPD-overexpressing Po1f-pJQ08, AnYHM2-expressing Po1f-pJQ09, and the co-expression strain Po1f-pJQ12. RT-qPCR confirmed successful transcriptional activation of the target genes. Shake-flask fermentation revealed that YlAMPD and AnYHM2 single-gene overexpression strains increased citric acid production by 34.23% and 29.62%, respectively, whereas the co-expression strain achieved a titer of 22.33 g·L−1, representing a 94.51% increase over the control. This enhancement substantially exceeded the arithmetic sum of the individual increases (63.85%), thereby exhibiting a distinct supra-additive effect. This study provides a candidate strain with application potential for rare earth bioleaching and offers a referable metabolic engineering strategy for citric acid production in Yarrowia lipolytica.

Fermentation doi: 10.3390/fermentation12090403

Authors: Camilo Molina Jhon Edinson Valencia Cristina Ramírez-Toro Liliana Londoño-Hernández German Bolívar Anna María Polanía Rivera

The shortage of protein-rich foods is a major challenge due to the rapid growth of the world’s population. For this reason, efforts are being made to achieve sustainability in the food system to produce nutritious foods with better qualities. The present study aimed to utilize a combination of widely consumed legumes with nutritional properties—such as lentils, chickpeas, and beans—considering their physicochemical and functional characteristics, to produce tempeh as a model. A simple mixture design was employed to develop a legume-based product through the production of fermented flour using Rhizopus oligosporus ATCC 22959. For this purpose, proximate composition, water absorption index (WAI), pH, phenolic content, and antioxidant capacity, via the DPPH (2,2-diphenyl-1-picrylhydrazyl) and ABTS (2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid)) radicals, were determined in the raw legumes. Optimal fermentation conditions were determined through digital image analysis, and fermentations were carried out according to the design. The crude and soluble protein content, phenolic content, and DPPH of the fermented samples were determined, and a statistical optimization was performed by maximizing each variable. Through optimization, it was found that a formulation of 80.81% lentil and 19.19% chickpea presented the best desirability (D = 0.75) according to the criteria mentioned above. These results were also compared with those obtained from the preparation of an original soy tempeh using the microorganism Rhizopus oligosporus; it was found that the protein differences between the original tempeh and the one made from the legume blend were 19 g/100 g dry matter for the original tempeh and 27.5 g/100 g dry matter for the one made with the legume blend, demonstrating that the combination of legumes exerts a favorable interaction within the mixture model on the physicochemical properties of tempeh and could represent significant potential for the production of flours applicable to the development of food products as part of alternative protein sources.

Fermentation doi: 10.3390/fermentation12090402

Authors: Shiyun Wang Yuanyuan Li Yanan Shi Benhong Xu Mingtao Huang

Collagen functional fragments retain selected activities of parent collagens while allowing greater flexibility in sequence design and precision fermentation. Although recent reviews have covered recombinant collagen production technologies, expression platforms, purification strategies, quality control, and biomedical applications, fragment selection, host–process matching, production, and characterization have received less integrated attention. This review focuses primarily on collagen-derived functional fragments, while collagen-mimetic peptides and collagen-like proteins are discussed as related design systems. The biological basis for fragmentation includes receptor-recognition motifs, matrikines and matricryptins, and basement membrane-derived fragments. The review further examines how motif context, Gly-X-Y organization, stabilizing sequence features, protease susceptibility, post-translational modification requirements, and host compatibility influence fragment stability, expression performance, production feasibility, and product integrity. Microbial production using Escherichia coli, Komagataella phaffii, and Saccharomyces cerevisiae is discussed from the perspectives of construct–host matching, secretory or intracellular production, prolyl 4-hydroxylase configuration, fermentation optimization and scale-up, and product characterization. Finally, we discuss AI-assisted, quality-guided design-build-test-learn workflows that integrate computational prediction, curated structural, extracellular-matrix, interaction, and protease resources, two-tier candidate evaluation, and format-appropriate experimental testing to support iterative sequence, host, and process optimization. The development of collagen functional fragments therefore depends on coordinated optimization of biological function, molecular design, microbial host performance, fermentation processes, and product characterization.

Fermentation doi: 10.3390/fermentation12090401

Authors: Yanjie Jia Wanting Yang Lulu Zhang Xinkang Hu Huanhuan Zhang Bo Zhang

Text Correction [...]

Fermentation doi: 10.3390/fermentation12090400

Authors: Kittaporn Rumjuankiat Nipon Sonhom Sujitra Techo Ratha-korn Vilaichone Sittiruk Roytrakul Janthima Jaresitthikunchai Thitiphorn Janyaphisan Wonnop Visessanguan Weerapong Woraprayote

Functional fermented foods are increasingly recognized as sources of microbial metabolites with potential biological activity. In this study, MRK2-3, a protease-sensitive anti-Helicobacter pylori component produced by Lactiplantibacillus pentosus MRK2-3 isolated from fermented mushroom sausage, was enriched and preliminarily characterized for in vitro anti-H. pylori activity. Activity appeared during the stationary phase. Sequential ethyl acetate extraction, Sep-Pak C18 chromatography, and reverse-phase HPLC increased specific activity, although the final activity recovery was 0.72%. MALDI-TOF analysis of the active fraction showed a dominant ion at approximately m/z 1848 together with several lower-intensity ions; therefore, chemical homogeneity and sequence identity were not established. The MRK2-3 fraction retained activity after incubation at pH 2–6 and after heating at 80 °C and 100 °C for 30 min, whereas activity decreased under alkaline conditions and after treatment at 121 °C for 15 min. Trypsin, α-chymotrypsin, pepsin and proteinase K reduced activity. The fraction inhibited all tested H. pylori strains (agar-dilution MIC ranged from 12.5 to 100 μg/mL). In a simulated gastric fluid, higher concentrations produced a rapid reduction in viable H. pylori 3949 counts. These findings support MRK2-3 as a fermentation-derived putative antimicrobial peptide candidate with anti-H. pylori activity under gastric-like conditions. However, its amino acid sequence, structural identity, and novelty require confirmation. Antimicrobial mechanism, cytotoxicity, and in vivo efficacy also remain to be established.

Fermentation doi: 10.3390/fermentation12090399

Authors: Julia Kruyeniski Carolina Mónica Mendieta Fernando Esteban Felissia María Cristina Area

The production of second-generation bioethanol from lignocellulosic biomass requires efficient enzymatic hydrolysis and fermentation processes that remain effective at industrially relevant solids loadings. In this study, bleached Pinus taeda kraft pulp was evaluated as a model substrate for bioethanol production at bench scale (4 L reactor) under high-consistency conditions (12.5–13.9% solids). Three process configurations were compared: separate hydrolysis and fermentation (SHF), simultaneous saccharification and fermentation (SSF), and pre-hydrolysis followed by simultaneous saccharification and fermentation (pSSF). Enzymatic hydrolysis in the SHF and SSF configurations stabilized between 54% and 58%, indicating that hydrolysis was the main process bottleneck under the evaluated conditions. In contrast, Saccharomyces cerevisiae efficiently fermented the available glucose, achieving nearly complete conversion of glucose. Among the evaluated strategies, pSSF showed the highest ethanol yield and volumetric productivity, achieving an ethanol yield of 61.8% and a productivity of 0.61 g L−1 h−1. While laboratory-scale SSF experiments conducted at 2% solids achieved complete conversion, the ethanol yield decreased to approximately 58% at the bench scale, highlighting the impact of high-solids operation on process performance. The lower performance observed at high solids may be associated with factors commonly reported during scale-up, including increased slurry viscosity, reduced mixing efficiency, limited enzyme accessibility, and mass-transfer constraints. Overall, the results manifest the need to enhance hydrolysis performance through improved reactor design, more effective mixing strategies, and optimized high-solids processing to facilitate the scale-up of lignocellulosic bioethanol production.

Fermentation doi: 10.3390/fermentation12090398

Authors: Sameh S. El-Hadad Amira A. Ayad Leonard L. Williams Ahmed Y. Okda Laila K. Hassan Mahmoud Abd El-Aziz

Consumers increasingly favor frozen yogurt as a healthier alternative to ice cream due to its lower fat content and probiotic benefits. Baked fermented milk (BFM), with its caramelized flavor, brown color, and enhanced nutritional properties, has emerged as a promising ingredient for improving the functional and sensory qualities of frozen yogurt. This study aimed to develop a novel probiotic baked frozen yogurt (PBFY) using BFM instead of traditional fermented milk (TFM) and to evaluate its physicochemical, microbiological, and sensory characteristics compared with traditional probiotic frozen yogurt (PTFY). Reconstituted skim milk and fresh cream were heat-treated (90 °C, 5 min) for TFM or baked (115 °C, 20 min) for BFM prior to culture inoculation. Both bases were inoculated with Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus, along with the probiotic strains Lactobacillus acidophilus and Bifidobacteriumanimalis subsp. lactis. PTFY and PBFY were formulated by blending ice cream mix with TFM or BFM at 25, 50, and 75% substitution levels. Increasing fermented milk concentration significantly decreased pH and surface tension while elevating mix viscosity. High inclusion levels (75%) impaired whipping ability, reduced overrun, increased fat destabilization, decreased the melting rate, and reduced culture survival during storage. Compared with PTFY, PBFY exhibited a higher melting rate, lower viscosity, and superior antioxidant activity (1.31- and 1.33-fold higher DPPH and ABTS radical-scavenging activity, respectively); however, the fermented milk type did not significantly affect overrun or fat destabilization. During storage, S. thermophilus was the most stable strain, with a decline rate of 0.6–0.69 log10 CFU/mL/month, whereas B. animalis subsp. lactis showed a more rapid decline (0.80–0.89 log10 CFU/mL/month). Sensory evaluation revealed that a 50% BFM inclusion level was optimal, producing PBFY with a smooth texture, light brown color, and appealing caramel-like flavor.

Fermentation doi: 10.3390/fermentation12090397

Authors: Yuan Fang Wenting Song Xuefeng Guo

Poly(β-L-malic acid) (PMLA) is a water-soluble, biodegradable aliphatic polyester whose pendant carboxyl groups support chemical functionalization for biomedical, packaging, and materials applications. Microbial fermentation can use pure sugars and biomass-derived carbon sources under mild conditions, but industrial translation remains constrained by feedstock cost and variability, strain performance, oxygen and pH control, pretreatment-derived inhibitors, and downstream recovery. This review therefore focuses on the fermentative production of PMLA from refined and renewable carbon sources, the microorganisms and metabolic routes involved, and the process variables that govern titer, yield, productivity, molecular weight, and purification. Agricultural and forestry feedstocks are compared according to their actual carbohydrate class and processing requirements. Bamboo shoot shell hydrolysate is treated as an emerging case study rather than an established production platform: one accepted shake-flask study directly demonstrated PMLA production by Aureobasidium pullulans NRRL Y-2311-1, but controlled bioreactor validation, reproducibility, techno-economic analysis, and application-specific product qualification remain to be further investigated. The review also examines autohydrolysis, low-molecular-weight PMLA for biomedical use, furan inhibition, membrane and ion-exchange purification, and the limits of current economic comparisons. This evidence-based framing identifies where bamboo-processing residues may contribute to renewable PMLA production while distinguishing laboratory feasibility from industrial readiness.

Fermentation doi: 10.3390/fermentation12090396

Authors: Da Ma Ruidong Yang Yuanchi Wang Yin Zheng

The development of plant-based functional beverages is often limited by inherent matrix defects, particularly undesirable off-flavors, astringency, and colloidal instability. Water kefir (WK), a highly resilient multispecies symbiotic consortium, offers a robust biorefining platform to address these challenges. This review systematically elucidates the underlying micro-ecological logic and biochemical mechanisms of WK-mediated plant matrix remodeling. We first detail how spatial niche differentiation and cross-feeding networks among lactic acid bacteria, yeasts, and acetic acid bacteria drive ecological homeostasis. Next, we highlight core molecular events that elevate sensory quality: protein unfolding for off-flavor elimination, enzymatic depolymerization of phenolics to mitigate astringency, and exopolysaccharide synthesis for rheological and flavor diffusion control. Finally, to overcome industrial scale-up challenges, we outline a precision fermentation framework, integrating systems multi-omics, real-time biomimetic monitoring, and sensory topological modeling. Ultimately, this synthesis provides theoretical guidance for the reverse flavor engineering and targeted nutritional design of novel plant-based beverages.

Fermentation doi: 10.3390/fermentation12090395

Authors: Chengyue Sun Xiaomeng Liu Qiulong Zou Ruwen Yang Roujia Kang Dixiang Bing Lei Zhang Ziyuan Ding Xianlong Zhou Wei Jiang

Low-glutamic-acid monosodium glutamate wastewater (L-MSGW), characterized by high (NH4)2SO4 concentrations, presents significant challenges for conventional treatment. γ-Polyglutamic acid (γ-PGA) production using industrial wastewater is an economical and environmentally friendly strategy. In the current study, we isolated and identified Bacillus subtilis GB, which exhibited exceptional tolerance to (NH4)2SO4, and capability for the high-efficiency biosynthesis of γ-PGA using untreated L-MSGW. Fermentation conditions were optimized using single-factor experiments coupled with response surface methodology, followed by scale-up validation in a 5 L fermenter. Under optimal conditions, the maximum γ-PGA yield reached 16.57 g/L with a minimal glutamate consumption of only 4.9 g/L. The study validated the feasibility of efficient γ-PGA production from L-MSGW by B. subtilis GB, providing a novel technical approach and theoretical basis for low-cost treatment and high-value resource utilization of L-MSGW. This study not only demonstrates the low-cost L-MSGW can be used for the high-value γ-PGA by B. subtilis GB but also provides a sustainable and economically viable solution for industrial wastewater treatment.

Fermentation doi: 10.3390/fermentation12080394

Authors: Silvia Stredanská Janka Kubincová Mária Kopuncová Eugen Kiss Stanislav Baxa Miroslav Stredanský

Arachidonic acid (AA) is a high-value long-chain polyunsaturated fatty acid commonly produced by submerged fermentation of oleaginous fungi. Solid-state fermentation (SSF) using low-cost agro-industrial by-products represents a promising alternative approach for sustainable microbial lipid production. In this study, spent malt, a by-product of the brewing industry, was evaluated as a solid substrate for AA production by Mortierella alpina under SSF conditions. Four M. alpina strains were screened for growth, lipid accumulation, and AA production, and M. alpina 959 was selected as the most promising strain. The effects of nitrogen supplementation and oil incorporation on biomass formation, lipid accumulation, and AA productivity were subsequently investigated. Sunflower oil incorporation into spent malt-based substrates improved AA productivity under the most favorable conditions, resulting in an AA proportion of 38.6% of total fatty acids and 102.5 mg AA g−1 of final dry mass of the SSF system (FDW-SC). Comparison of selected vegetable oils further demonstrated that the lipid source affected AA production, while having only a limited effect on fungal growth and total lipid accumulation. Overall, this study demonstrates the feasibility of spent malt-based SSF for AA production by M. alpina and highlights the valorization of brewery spent malt as a low-cost agro-industrial by-product for the production of value-added microbial lipids.

Fermentation doi: 10.3390/fermentation12080393

Authors: Fernando Sánchez-Suárez Isidoro Lucena Nieves López de Lerma Rafael A. Peinado

Pedro Ximénez sweet wines are traditionally produced from raisined grapes and are characterised by very high sugar and ethanol contents. However, there is growing interest in wines with lower sweetness and improved sensory balance. The aim of this study was to evaluate the effect of controlled partial fermentation of Pedro Ximénez raisined grape must prior to fortification on the chemical and sensory properties of the resulting wines. Fermentations were conducted until ethanol concentrations of 3.0, 5.0, 6.5, and 7.7% (v/v) were reached, after which fermentation was arrested, and all wines were standardized to a final ethanol concentration of 9% (v/v). Fermentation progression resulted in a reduction of residual sugar content from 445 to 313 g/L and significant increases in titratable acidity, volatile acidity, glycerol, higher alcohols, ethyl acetate, and 2-phenylethanol. Hierarchical cluster analysis confirmed that fermentation degree was the main factor driving wine differentiation. Sensory evaluation showed that partial fermentation reduced the intensity of characteristic descriptors such as honey, raisin, and fig while simultaneously decreasing sweetness perception and increasing freshness and overall balance. Wines fermented to 6.5 and 7.7% (v/v) ethanol received the highest preference scores from the expert sensory panel. These results indicate that controlled partial fermentation is a feasible technological strategy for modifying the composition and sensory profile of Pedro Ximénez wines and provide a basis for further optimization of alternative, less sweet Pedro Ximénez-style wines.

Fermentation doi: 10.3390/fermentation12080392

Authors: Qian Zhang Zonglu Yao Lixin Zhao Jing Feng Juan Luo Jiadong Yu Ruixia Shen

Aerobic composting converts agricultural waste into stable, humus-rich products, and the application of exogenous additives is an efficient strategy to enhance humification. This systematic review synthesized studies published between 2023 and 2026 on additives that promote humification during agricultural waste composting. Based on the distribution of the retrieved literature, additives are categorized into inorganic additives, organic additives, biological strategies, and composite systems, and the effects and mechanisms of each category are systematically discussed. Iron-based additives achieve the highest humic acid (HA) increases of 82–267% through Fenton-like redox catalysis. Clay minerals and biochar produce moderate HA enhancements of 25–163% via physical structuring and surface adsorption with broader applicability. Small-molecule precursors and exogenous humic substances achieve HA gains exceeding 100% at sub-percent doses. Biological strategies provide self-sustaining catalytic activity but are sensitive to environmental conditions. Composite additives, the largest category, generally outperform single additives through functional complementarity, though antagonistic effects have also been documented. Cross-study patterns suggest that different feedstocks respond preferentially to distinct additive types, though systematic experimental validation is lacking. Critical gaps between laboratory findings and practical application are identified, including the predominance of small-scale studies, the absence of techno-economic analysis, and the unassessed environmental fate of metal-based additives. Future research priorities include pilot-scale validation under industrial conditions, the establishment of standardized humification metrics, and long-term field monitoring.

Fermentation doi: 10.3390/fermentation12080391

Authors: Murat Emre Terzioğlu Zeynep Çağla Tekgül

In dairy production, the fermentation process is a complex biochemical system that plays a significant role in determining the quality criteria of the final product. Traditional methods for controlling fermentation rely on limited and non-standard process parameters. In recent years, omics technologies have come to the forefront, enabling the monitoring of fermentation dynamics at the molecular level with their current, efficient, and reliable approaches. Thanks to omics approaches such as metabolomics, metagenomics, proteomics, and lipidomics, starter culture behavior, metabolite formation, aroma–texture formation, and microbial interactions in the fermentation process can be characterized more comprehensively. On the other hand, evaluating or calculating high-dimensional omics data using traditional statistical methods presents a challenge. Artificial intelligence applications are overcoming this challenge, offering significant opportunities for the accurate and reliable evaluation of data. Artificial intelligence-powered models hold promise in areas such as predicting fermentation kinetics, process control, optimizing quality criteria, and revealing the therapeutic potential of products through metabolites. This compilation aims to comprehensively address current approaches to modeling the fermentation process and quality parameters of dairy products using multi-omics technologies and artificial intelligence applications. In this respect, it will provide current and important perspectives for industrial applications and future studies.

Fermentation doi: 10.3390/fermentation12080390

Authors: Cristobal A. Onetto Jane McCarthy Simon A. Schmidt

Hanseniaspora uvarum is consistently observed as the dominant non-Saccharomyces species in grape juice leading to spontaneous fermentations and is often considered a detrimental factor due to the increased volatile acidity associated with its abundance. This study investigated how fermentation temperature affects the competitive interaction between H. uvarum and Saccharomyces cerevisiae, and how these shifts influence microbial community structure during spontaneous grape juice fermentations. Defined single- and co-inoculated fermentations were conducted across a range of temperatures (17, 23, and 25 °C), and cell abundance was quantified by flow cytometry. In parallel, spontaneous fermentations were monitored using ITS metabarcoding to assess temperature-driven changes in fungal community composition. At 17 °C, H. uvarum dominated early fermentation due to its faster growth rate, resulting in slower sugar consumption and prolonged fermentations. Increasing fermentation temperature enhanced the early growth of S. cerevisiae. Consistent patterns were observed in spontaneous fermentations, where higher temperatures increased the relative abundance of S. cerevisiae and shortened fermentation time. These results demonstrate that fermentation temperature is a major driver of yeast competition and suggest that moderate increases in fermentation temperature may help limit excessive H. uvarum proliferation during spontaneous fermentations.

Fermentation doi: 10.3390/fermentation12080389

Authors: Aytac Perihan Akan Kenan Dalkilic Aysenur Ugurlu

Rapid population growth, urbanization, and industrialization are continuously increasing global energy demand while intensifying climate change associated with fossil fuel consumption. In this context, renewable energy production from organic waste has gained increasing attention as a sustainable and environmentally friendly strategy. Anaerobic digestion (AD) offers significant potential for simultaneous waste stabilization and biomethane generation. However, many previous studies investigating lignocellulosic or nutrient-rich substrates have relied on physical, chemical, or thermal pretreatment methods to enhance biodegradability, despite their additional operational costs, energy consumption, and environmental impacts. Therefore, developing low-cost and pretreatment-free co-digestion strategies remains an important research need. This study investigated the biomethane production potentials of untreated chicken manure (CM) and duckweed (Lemna minor-LM) collected from the final sedimentation tanks of wastewater treatment plants under mono-digestion and co-digestion conditions. The study hypothesized that rapidly growing and widely available LM biomass could enhance methane production without requiring pretreatment. Among all reactors, CM0.75 (75% of the total TS derived from CM and 25% from LM and inoculum) achieved the highest performance with a cumulative biogas production of 5350 mL (74.2% of CH4) and a methane yield of 327 mL CH4/g VS, while mono-digestion of CM resulted in the lowest methane yield of 104 mL CH4/g VS. The results demonstrated that LM biomass naturally proliferating in wastewater treatment plants can be directly utilized as an effective co-substrate to improve biomethane production from poultry wastes. The proposed approach provides a cost-efficient, eco-friendly, and circular-economy-oriented alternative by eliminating the need for pretreatment while simultaneously valorizing problematic biomass generated in wastewater treatment facilities.

Fermentation doi: 10.3390/fermentation12080388

Authors: Gulhan Turk Gorkem Ozulku Saeideh S. Fatemizadeh Osman Sagdic Ömer Şimşek

The aim of this study is to provide an alternative bakery practice by evaluating the effects of cold fermentation (backslopping at 7 and 15 °C) and cold storage (stored at 7 and 15 °C) after backslopping on Type I sourdough characteristics and the technological performance of the corresponding breads. Sourdough characteristics (pH, acidity, and microbial counts), bread quality (texture and color), storage behavior, and volatile compound profiles (VoC) were investigated to compare the extent to which these treatments improve product quality. No significant difference was observed in pH and total titratable acidity among the sourdoughs. However, the fermentation quotient of the cold storage group was higher than that of the cold fermented group. Backslopping at cold fermentation temperatures (7 and 15 °C) caused a significant reduction in lactic acid bacteria counts. Conversely, cold fermentation and cold storage at 15 °C led to a significant increase in yeast counts. From a technological perspective, bread produced with sourdough backslopped at 15 °C exhibited a higher specific volume than breads produced with the other cold-treated sourdoughs. At the end of the storage period, the bread made with cold-backslopped sourdough at 7 °C had the highest hardness value, while others remained similar. Principal component analysis (PCA) of volatile compounds revealed that cold-stored sourdoughs were clustered apart from room-temperature-stored sourdoughs, mainly driven by ethyl hexanoate and hexyl acetate. Heptanoic acid was among the key VOCs contributing to the positioning of breads made with cold-stored sourdoughs in the PCA. In conclusion, these findings demonstrate that backslopping and storage at 15 °C offer an industrially relevant production strategy while improving textural properties and VoC profiles.

Fermentation doi: 10.3390/fermentation12080387

Authors: Willian de S. M. Reis Gabriel L. de Arruda Silvio S. da Silva Arnaldo M. R. Prata Júlio C. dos Santos

Bioenergy biorefineries generate lignocellulosic by-products rich in fermentable sugars that can serve as renewable feedstocks for the production of high-value bioproducts, including microbial pigments with promising bioactive properties (antioxidant, antimicrobial, and anticancer). Therefore, this study aimed to identify a suitable inoculum preparation strategy, to optimize the culture medium, and to evaluate biopigment production using xylose-based media derived from sugarcane bagasse hemicellulosic hydrolysate (SBHH). Different inoculation strategies were evaluated (cell suspension, whole mycelial discs, and fractionated mycelial discs) and supplementation with Tween 80 (TW80). The medium composition was optimized using a Box–Behnken design, with xylose, yeast extract, and TW80 as variables, and fermentations were then conducted under selected inoculum conditions in semi-defined media and SBHH. The mycelial disc inoculation strategy was selected due to its high biopigment production and lower operational complexity, yielding 8.99, 8.48, and 11.78 AU of yellow, orange, and red biopigments, respectively. The optimized culture composition consisted of 55.65 g/L of xylose, 4.18 g/L of yeast extract, and 15.38 g/L of Tween 80. The cultivation of M. ruber in SBHH resulted in 12.73, 10.75, and 14.56 AU of yellow, orange, and red biopigments, respectively. Thus, the strategy of inoculum preparation associated with non-ionic surfactant proved promising for application in bioenergy biorefineries.

Fermentation doi: 10.3390/fermentation12080386

Authors: Mamica Ruci Renata Kongoli Rosaria Cozzolino Cristina Matarazzo Bruno Testa Onejda Kyçyk Julian Karaulli Massimo Di Renzo Catello Di Martino Fatbardha Lamçe Massimo Iorizzo

The growing demand for clean-label and low-sulphite wines has increased interest in alternative preservation strategies capable of reducing sulphur dioxide (SO2) usage while maintaining wine quality and stability. In this study, the effectiveness of Vitis vinifera leaf extract as a natural alternative to sulphur dioxide was evaluated in High Hydrostatic Pressure (HHP)-treated Fiano wines. Three experimental wines were produced: CW (control wine), SW (sulphited wine), and LW (leaf-extract wine). Following alcoholic fermentation, all wines were subjected to HHP treatment (600 MPa for 5 min) and stored at 4 °C for 60 days. Physicochemical parameters, volatile organic compounds (VOCs), and sensory characteristics were evaluated immediately after alcoholic fermentation and after HHP treatment followed by refrigerated storage. Compared with the control wine, LW exhibited approximately 20% higher total polyphenol concentrations, whereas SW showed the greatest preservation of fermentation-derived esters. HHP treatment induced only moderate changes in the volatile fraction, confirming the suitability of this non-thermal technology for wine stabilization. LW wines were characterized by higher abundances of terpene-related compounds, C6 alcohols, medium-chain fatty acids, and phenolic-associated volatiles, resulting in more pronounced floral, balsamic, herbaceous, and vegetal sensory attributes. Principal Component Analysis (PCA) explained 79.0% of the total VOC variability and clearly differentiated LW wines from CW and SW according to their volatile profiles, while sensory analysis confirmed the development of a distinctive aromatic identity associated with grapevine leaf extract. Overall, the results indicate that the combined application of V. vinifera leaf extract and HHP represents a promising strategy for the partial replacement of sulphur dioxide in white winemaking. This integrated approach contributes to wine stabilization while promoting the valorization of grapevine leaves as a sustainable winery by-product within a circular economy framework.

Fermentation doi: 10.3390/fermentation12080385

Authors: Pablo Mompean José Ramón Acosta-Motos Llanos Martínez-Martínez Luis Noguera-Artiaga Angel A. Carbonell-Barrachina Patricia Navarro Antonio José Pérez-López

Oxygen management during alcoholic fermentation can redirect phenolic reactions and yeast-derived aroma formation, but the use of mild hyperbaric conditions as a micro-oxygenation strategy remains poorly characterized. This study evaluated the effects of hyperbaric micro-oxygenation at 1.40 ATA in a stainless-steel chamber on the fermentation of Vitis vinifera L. cv. Monastrell must, comparing treated and non-micro-oxygenated samples at the initial, mid-fermentation, and final stages. Physicochemical parameters, CIELAB color coordinates, total phenolic content, volatile organic compounds, and descriptive sensory attributes were analyzed. Hyperbaric micro-oxygenation did not impair fermentation completion, as both treatments reached final residual sugar values of 2.2 g/L and alcohol contents of 15.2–15.4% v/v. The treatment promoted a darker final chromatic profile, with lower L*, the highest overall color difference, and a marked increase in total phenolic content, reaching 1900.9 mg gallic acid equivalents/L compared with 1593.2 mg gallic acid equivalents/L in control. Volatile changes were compound, and stage-dependent, indicating modulation rather than generalized enhancement of aroma formation. Ethyl esters, particularly ethyl octanoate and ethyl decanoate, increased markedly under micro-oxygenation, while acetate esters such as ethyl acetate and hexyl acetate decreased relative to the initial must, reflecting a shift in the balance of aroma-active compounds rather than a uniform increase across all volatile families. These findings support mild hyperbaric micro-oxygenation as a promising non-thermal strategy to modulate Monastrell fermentation quality.

Fermentation doi: 10.3390/fermentation12080384

Authors: Soumya Gupta Annabel Fernandes Laura Grasa Carlos Rubio Jesús Salafranca

The integration of coupled dark fermentation (DF) and photofermentation (PF) systems represents a promising approach for concurrent H2 production and organic load reduction in carbohydrate–rich wastewater. However, their behavior under non–optimized, continuous industrial operation remains inadequately characterized, hindering practical implementation. This case study evaluated an integrated DF–PF system treating a synthetic sugar mixture mimicking acidic fruit and dairy processing wastewater. The system presented herein (5-L reactors) serves as an initial prototype to facilitate scaling to both pilot (40-L reactors) and, ultimately, industrial (600-L) scales using real effluents within the framework of a research project. The bioreactors were operated continuously with a 10 h hydraulic retention time and an organic loading rate of 2.5 g COD L−1 d−1. The pH and temperature were monitored but intentionally left uncontrolled. The DF stage facilitated by hydrogen–producing bacteria achieved H2 concentrations of up to 57% (v/v) and a maximum production rate of 177 mL H2 L−1 d−1; however, it demonstrated notable process instability due to the absence of controls. In contrast, the PF stage exhibited negligible H2 production (1.8% v/v) attributable to the displacement of Rhodopseudomonas species by microbial competition, thereby functioning predominantly as a polishing step. Overall, the coupled system achieved an average COD removal of 34%, highlighting the functional differentiation between stages and identifying microbial competition as the primary constraint under suboptimal conditions.

Fermentation doi: 10.3390/fermentation12080383

Authors: Chaoyang Zhu Dina Zhu Bei Liao Zhanlei Fan Yongmei Hu Shumiao Zhao Yunxiang Liang Jinshan Li

Black rice wine is a traditional cereal-based fermented beverage valued for its colour, flavour and bioactive constituents. However, product innovation is needed to meet increasing consumer demand for fruit-derived aroma, nutritional quality and functional attributes. To address this demand, we developed an integrated processing strategy combining enzymatic pretreatment of pitaya pulp with stage-specific co-fermentation, an approach that has not been systematically evaluated in rice wine systems. Pitaya pulp was first pretreated with pectinase using an orthogonal design to optimise juice yield and betacyanin retention; the enzymatically pretreated pitaya pulp was then introduced either as pulp or as fermented pitaya wine at the saccharification, tank-flushing or ageing stage. The resulting wines were compared with a pitaya-free control and three commercial black rice wines on the basis of physicochemical indices, phenolic and pigment contents, amino acid composition, mineral composition and fuzzy mathematics-based sensory evaluation. Pitaya pulp added at saccharification (Sample THJ) yielded the strongest functional and nutritional profile, with the highest total phenolic content (629.74 mg/L); high flavonoid content (152.07 mg/L); enrichment in Mg, K and Mn; and a broader taste-active amino acid profile. By contrast, pitaya pulp added during tank flushing (Sample CHJ) achieved the highest sensory score (85.3) and had the highest anthocyanin content. These findings indicate that the timing and form of pitaya addition determine the balance between functional enrichment, pigment retention and sensory quality. Co-fermentation with red-fleshed pitaya, particularly the early addition of enzymatically pretreated pitaya pulp, provides a practical route for the development of fruit-flavoured black rice wine with improved nutritional composition and consumer appeal, suggesting the potential for enhanced functional value.

Fermentation doi: 10.3390/fermentation12080382

Authors: Umair Ahsan Derya Merve Karagöz Muhammad Shazaib Ramay Syed Umer Akhter Anas Tahir Bekir Tosun Ifrah Raza Murat Er Muhammad Kashif Yar Eren Kuter

A 2 × 2 × 2 factorial experiment was conducted to study the individual and combined effects of barley variety (Tarım 92 and Bravo), homofermentative inoculant (HMF; Lactobacillus plantarum and Enterococcus faecium), and heterofermentative inoculant (HTF; Lactobacillus buchneri) on the fermentation characteristics, chemical composition, nutritive value indices, and in vitro digestibility of whole-crop barley silages which were harvested at the early dough stage. Forty-eight laboratory silos (six replicates per treatment) were ensiled for 120 days. Tarım 92 silage exhibited superior fermentation stability across all treatments, with pH ranging from 4.11 to 4.30, Flieg points of 102.94–115.47, and lower NH3-N concentrations (142.01–173.72 mg/kg TN (total nitrogen)), reflecting better protein preservation than Bravo silage. In contrast, the poorest fermentation outcome (pH 5.24; Flieg 47.06; NH3-N 307.48 mg/kg TN) was exhibited by the Bravo silage inoculated with only homofermentative inoculants. Significant three-way interaction effects were detected among barley variety, homofermentative inoculant, and heterofermentative inoculant (p < 0.05). The dual-inoculated Tarım 92 silage showed the optimum in vitro digestibility and nutritional value (TDN 77.14%; NEL 1.81 Mcal kg−1 DM; RFV 176.25; ADF 18.75%; IVOMD 65.72%; predicted DMI 3.06% BW), while higher amounts of butyric acid and NH3-N suggested that better nutritional value was not always accompanied by better protein preservation. The combined inoculation also yielded the best outcomes within Bravo treatments (RFV 144.00), whereas HTF inoculation alone reduced IVDMD and IVOMD below uninoculated controls. Overall, these results show that the response to microbial inoculation is genotype-dependent and a combination of homofermentative and heterofermentative inoculation for high-dry-matter barley varieties can enhance nutritive value and digestibility. However, fermentation quality indicators, particularly butyric acid and NH3-N, require consideration in the selection of technique.

Fermentation doi: 10.3390/fermentation12080381

Authors: Yotsapon Yangngam Seangla Cheas Chanon Suntara Metha Wanapat Juan J. Loor Anusorn Cherdthong

This study evaluated molasses-based liquid feeds containing conventional urea or a commercial fat-coated urea product marketed as slow-release urea (SRU). The effects on in vitro gas-production kinetics, rumen fermentation, and substrate degradability were examined. Treatments followed a 5 × 3 factorial design. The five formulations were 0% urea, 5% urea, 10% urea, 5% SRU, and 10% SRU. Each formulation was supplied at 0.9%, 1.8%, or 2.7% of the 0.5 g basal substrate dry matter (DM). Four independent incubation runs were conducted. A formulation × inclusion-level interaction was detected for gas-production lag time (p = 0.0046). The shortest lag time occurred with 10% SRU supplied at 2.7%. At 48 h, the 0.9% inclusion level resulted in greater dry matter and organic matter degradability than the 1.8% and 2.7% levels. The 5% SRU formulation had the lowest 24 h degradability, whereas 10% SRU had the greatest 48 h dry matter degradability. Ammonia nitrogen (NH3-N) was analyzed separately at each sampling time. The 10% conventional urea formulation had the greatest NH3-N concentration at 1 and 2 h. The 10% SRU formulation had the greatest concentration at 4, 6, and 12 h. These within-time differences do not demonstrate different ammonia-release patterns over time. At 12 h, the 10% SRU formulation had the greatest total volatile fatty acid concentration and propionate molar proportion. Protozoal counts were unaffected. The results reflect differences among the complete liquid-feed formulations. They do not provide direct evidence of controlled ammonia release or improved nitrogen–carbohydrate synchronization. Further in vivo studies using compositionally balanced formulations are required.

Fermentation doi: 10.3390/fermentation12080380

Authors: Nisa Alfilasari Nattha Tampanna Nualpun Sirinupong Santad Wichienchot

Bioactive peptides (BPs) are increasingly recognized for modulating the gut microbiome, metabolome, and brain function via the gut–brain axis. Kapi-pla, a traditional Thai fermented freshwater fish paste rich in proteins and peptides, is widely consumed in Southern Thailand. This study profiled the peptides of Phatthalung Kapi-pla (PK) and Songkhla Kapi-pla (SK) and investigated their impacts on gut microbiota and metabolome using a simulated colonic fermentation model with fecal samples from patients with Alzheimer’s disease (AD). Microbial composition and metabolites were assessed by 16S rRNA sequencing and LC–MS/MS, respectively. After 24 h fermentation, PK modestly increased Shannon diversity relative to the unsupplemented control, with richness indices unchanged, reduced Proteobacteria abundance and opportunistic pathogens such as Escherichia–Shigella and Klebsiella, and selectively increased short- and branched-chain fatty acids, including acetate, propionate, butyrate, and iso-valerate. PK further enhanced neuroactive metabolites relevant to AD pathology, underscoring its potential as a functional food ingredient to ameliorate AD-associated dysbiosis and support gut–brain axis health.

Fermentation doi: 10.3390/fermentation12080379

Authors: Luis F. Castro Tessa A. J. Murphree Kate P. Perry Sean Kuster L. Federico Casassa

The aim of this study was to investigate the effects of thermo-maceration and apple pomace addition on phenolic compound extraction during cider fermentation and on the volatile profile of the final product. Jonagold apples were milled, and the juice was divided into four groups: (1) Control: no apple pomace or thermo-maceration; (2) Control TM: thermo-maceration only; (3) CiderAP: apple pomace addition only; and (4) CiderAPTM: apple pomace addition combined with thermo-maceration. Apple pomace was added at 50% of the average pomace weight created during milling, and thermo-maceration consisted of heating the juice to 60 °C for 1 h prior to fermentation. Ciders were fermented at 20 ± 1 °C for 14 days. CiderAP and CiderAPTM showed higher total phenolic content (TPC) and antioxidant activity than the control treatments. The volatile profile was also modified, with CiderAPTM showing the highest phenylethyl alcohol concentration and esters exhibiting compound-specific responses. Statistical analysis showed that only apple pomace addition significantly increased TPC and antioxidant activity, while both apple pomace addition and thermo-maceration affected the volatile profile. Under the conditions evaluated, thermo-maceration did not increase TPC or antioxidant activity beyond the effect of apple pomace addition but did modify the volatile profile of the ciders. Further research should evaluate different thermo-maceration temperatures and durations to determine its potential application in cider production. The findings of this study provide valuable insights for cider makers on the valorization of apple pomace and potential use of thermo-maceration in cider production.

Fermentation doi: 10.3390/fermentation12080378

Authors: Soha Ghzayel Ahmed E. Kholif Alexey Díaz-Reyes Bassam Abu Aziz Halimeh Zoabi Raouia Ben Rhouma Sawsan Hassan Secundino López Adel M. M. Kholif Silvia Parrini Andrea Confessore Hajer Ammar

This study examined the effects of replacing 25% (P25) or 50% (P50) of concentrate dry matter (DM) with sun-dried carob (Ceratonia siliqua L.) pods on growth performance, apparent nutrient digestibility, carcass traits, meat quality, serum biochemistry, and rumen microbiology in growing Assaf lambs. Twenty-four weaned male Assaf lambs (initial body weight [BW] 27.0 ± 0.5 kg; 2.5 months of age) were randomly assigned to three dietary treatments (n = 8 per group) in a completely randomized design and fed for 16 weeks. P50 achieved the highest ANCOVA-adjusted least squares mean final BW (53.0 kg) and average daily gain (ADG) (220.8 g/d), followed by P25 (51.1 kg; 203.3 g/d) and the control (46.2 kg; 160.2 g/d) (p < 0.001). Feed conversion ratio (FCR) improved from 8.99 in the control to 6.16 and 6.11 in P25 and P50, respectively, with no significant difference between the two carob-supplemented groups (p < 0.001). Apparent DM and organic matter (OM) digestibility increased with carob inclusion at both 3 and 6 months of age (p ≤ 0.0001). Cold carcass weight (CCW) was higher in carob-supplemented lambs (p < 0.001), whereas carcass muscle proportion did not differ among treatments (p = 0.688), and carcass fat proportion was higher in P25 than in the control (p = 0.039). Warner–Bratzler shear force declined progressively with carob inclusion (p < 0.001), indicating improved meat tenderness. Serum total protein was highest in P25, whereas blood urea nitrogen (BUN), low-density lipoprotein (LDL), and glutamate oxaloacetate transaminase (GOT) decreased with carob inclusion (p < 0.001). Rumen pH was highest in P25 (6.40), total bacterial and lactic acid bacteria (LAB) counts increased, and protozoa counts declined (p < 0.001). Because carob pods replaced concentrate, rather than being added to an isonitrogenous diet, the combined effects of pods per se, reduced crude protein supply, and altered energy density must all be considered when interpreting the results. These findings support carob pods as a practical, locally available partial substitute for concentrate feed in Assaf lamb production under Mediterranean and Near Eastern conditions.