Search Results (220)

Background/Objectives: The stability and biological safety of newly developed formulations must be established to support their therapeutic efficacy and clinical translation in pulmonary drug delivery. Therefore, this follow-up study comprehensively evaluated the short- and long-term stability and the in vitro cytotoxicity of optimized, co-spray-dried theophylline (THN) dry powders for inhalation against A549 lung epithelial cells. Methods: Two established formulations were selected: THN-RAF (raffinose–leucine–glycine based) and THN-TRE (trehalose–leucine based). Stability was assessed under accelerated conditions (40 °C/75% RH, 3 months) and long-term desiccator storage (25 °C, 1 year) using laser diffraction, SEM, XRPD, FTIR, DSC, TGA, and Andersen Cascade Impaction. As THN-TRE had been previously confirmed to be cytocompatible, only THN-RAF and its components were evaluated against A549 human alveolar epithelial cells using the MTT assay. Results: Under accelerated conditions, both formulations exhibited pronounced recrystallization (Xc up to 89.9%), agglomeration (D [0.9] up to 217.08 µm for THN-TRE), and deterioration in aerodynamic performance (FPF as low as 11.55%, MMAD up to 6.68 µm). By contrast, long-term desiccator storage induced substantial recrystallization (Xc up to 80.7%) while preserving thermal, chemical, and aerodynamic performance (FPF ≈ 40%; MMAD 4.99–5.21 µm). THN-RAF was more resistant to stress-induced agglomeration than THN-TRE. Cytotoxicity assessment confirmed cytocompatibility of THN-RAF, with cell viability exceeding 70.99% at all tested concentrations (up to 500 µg/mL). Conclusions: These findings reveal a marked discrepancy between the outcomes of ICH accelerated testing and long-term desiccator storage. They underscore the importance of considering moisture-protective packaging configurations when designing stability protocols for amorphous inhalable formulations. Full article

The plant-based diets market has grown in popularity over recent decades and is projected to reach USD 27 billion by 2030. Innovative systems such as intercropping, defined as growing two or more crop species in proximity, are promising contributors to the advancement of sustainable agriculture by improving resource efficiency, enhancing crop resilience, and cutting down the need for chemical inputs. However, legume seed lots grown under intercropping versus monocropping systems differ in composition, which, in turn, can influence seed flour quality. Furthermore, antinutritional properties of legumes, such as phytic acid, condensed tannins (CTs), and raffinose family oligosaccharides (RFOs), limit their utilisation in product formulation. Therefore, the purpose of the study was to reduce these antinutritional compounds for utilisation of the intercropped legume flour in bakery products. A harvest of intercropped peas and faba bean mix (IM) obtained from Irish farmers was soaked (S) for 8 and 16 h and germinated (G) for 24, 48, and 72 h. Non-germinated intercropped mix (IM) was used as a control. All flours were subsequently used to substitute for wheat flour and fortify wheat-based cakes. The 72 h germinated intercropped mix (pea bean; 95.5:4.5) flour showed significantly (p < 0.05) lower levels of antinutritional compounds than the non-germinated intercropped flour. Raffinose, stachyose, and verbascose contents were reduced by 43%, 38%, and 46%, respectively, while phytic acid and condensed tannins decreased by 32% and 57%, respectively. The germination process also reduced the green hue (a*) from −8.37 to −6.49 and enhanced levels of soluble dietary fibre by 2.67% of the flour, while improving their suitability for bakery applications. Wheat-based cakes fortified with germinated and non-germinated intercropped legume flours showed a significant enhancement in protein and soluble dietary fibre contents. Compared with the control cake, protein content increased from 9.52 to 12.80%, while soluble dietary fibre content increased from 0.47 to 1.99% in the fortified cakes. Cakes containing up to 40% germinated flour (G72) showed comparable specific volume values 1.84–1.86 mL/g) and slice brightness (112.50–94.23) to the control cake. Additionally, cakes formulated with 40% intercropped flour showed significantly lower condensed tannin and phytic acid contents (p < 0.05), with reductions of 38% and 31.38%, respectively. Samples containing intercropped legume flours proved suitable for bakery applications, supporting up to 40% substitution in wheat-based cakes, and the germination process was effective in reducing antinutritional properties in intercropped legume flour fortified cakes. Full article

Background/Objectives: Raffinose is a prebiotic trisaccharide fermented by human gut microbiota, yet the strain-level determinants of its degradation and associated metabolic outputs remain poorly characterized. Methods: In the present study, anaerobic fermentation, 16S rRNA gene amplicon high-throughput sequencing, culturomics, and metabolomics were conducted to address this question. Results: In vitro fermentation of raffinose was performed using fecal samples from 16 healthy donors, and substantial interindividual variation was observed in substrate consumption, microbiota composition, and short-chain fatty acid production. Through culturomics, 204 bacterial strains spanning 18 species were isolated, among which Limosilactobacillus reuteri subsp. reuteri exhibited the highest raffinose-degrading efficiency. Untargeted metabolomics comparing L. reuteri subsp. reuteri cultured on raffinose versus glucose revealed a distinct metabolic shift, with 290 metabolites significantly upregulated, including the putatively identified fructooligosaccharide 1-kestose—a compound with documented prebiotic and anti-inflammatory properties. KEGG enrichment further highlighted coordinated remodeling of nucleotide and amino acid metabolism. Conclusions: Collectively, this study provides a strain-level framework for understanding how raffinose shapes the gut microbiota and identifies L. reuteri subsp. reuteri as a key metabolic hub linking prebiotic consumption to the production of bioactive metabolites. Full article

The application of the gas chromatography coupled to mass spectrometry (GC-MS) has been applied for metabolite profiling in sprouts of Brassicaceae (broccoli, kale, radish), sunflower, and Fabaceae (alfalfa, clover, lentil, and mung bean). Soluble carbohydrates were quantitatively the major fraction of total polar metabolites in Brassicaceae and sunflower sprouts. The main sugars in sunflower, mung bean, and Brassicaceae sprouts were fructose and glucose, while in clover, alfalfa, and lentil sprouts, the dominant sugar was sucrose. Myo-inositol was found in the sprouts of all analyzed species, while d-chiro-inositol, epi-inositol, or scyllo-inositol, as well as d-pinitol, were present in the sprouts of some legumes. Among the amino acids in clover and alfalfa sprouts, asparagine, serine, and threonine were quantitatively dominant. During cold storage of sprouts or their drying, a decrease in the content of monosaccharides and an increase in the levels of sucrose were found. Furthermore, such conditions of sprouts storage caused an accumulation of raffinose, proline, and γ-aminobutyric acid. The reduction in the content of monosaccharides and the increase in the content of free amino acids in dried sprouts is new information. Moreover, information about the small effects of low-temperature storage and drying of sprouts on the content of cyclitols seems to be valuable. Full article

This study investigated lactic fermentation of green buckwheat beverages formulated at 8% (A, AA) and 10% (B, BB) solids using single- and multiple-strain cultures of Lactiplantibacillus plantarum, Lactobacillus acidophilus, Lacticaseibacillus paracasei and Lacticaseibacillus rhamnosus. Fermentation at 37 °C rapidly reduced pH from slightly alkaline values (7.44–7.57) to approximately 4.2–4.5 within 3–8 h, while viable counts increased from near-zero to 8–9 log₁₀CFU mL⁻¹, confirming efficient lactic acid bacteria (LAB) proliferation in all substrates. A general trend was observed in the sugar consumption strategy of studied LAB: sucrose (after hydrolysis) and glucose were almost completely depleted within 6–8 h, fructose was consumed more slowly, and raffinose remained largely unchanged, with the 10% substrate mainly accelerating early sugar turnover without altering final cell densities or the qualitative utilisation pattern. Dry matter changed little during fermentation, whereas total phenolic content (TPC) and total tannin content (TTC) were strongly affected in a matrix- and strain-dependent manner. At 8% solids, fermentation promoted substantial TTC reduction and, for several cultures, a net decrease in extractable phenolics. In contrast, 10% formulations, particularly those inoculated with L. acidophilus and L. paracasei (alone or in combination), partially preserved or increased TPC while achieving more moderate tannin losses. Overall, green buckwheat proved to be a promising substrate for developing fermented beverages in which solids level and starter composition can be tuned to combine rapid acidification and high LAB viability with tailored sugar depletion and favourable modulation of phenolic and tannin fractions. Full article

Rice storage under suboptimal conditions leads to rapid quality deterioration. In this study, effects of low-temperature (LT, 20 °C) combined with nitrogen-controlled atmospheric conditions (NCA, 95%) on physicochemical properties, nutrient, mycotoxins and metabolites of rice during 180 d storage were comprehensively evaluated, to explore a potential preservation technique for rice. The LT + NCA treatment significantly inhibited the accumulation in free fatty acid value, malondialdehyde and electrolyte leakage, while maintaining higher catalase activity, lower amylose content and better pasting properties (higher breakdown, lower final viscosity and setback). Crucially, mycotoxin accumulation remained within safe limits across all groups, with LT + NCA showing the lowest levels. Metabolomics analysis identified 653 metabolites, with LT + NCA significantly modulating pathways related to carbohydrate metabolism (e.g., promoting raffinose accumulation while inhibiting the pentose phosphate pathway) and amino acid metabolism (enhancing glutathione metabolism and suppressing arginine biosynthesis). These metabolic rearrangements reduced oxidative damage, stabilized membrane integrity, and preserved cooking quality. Therefore, LT + NCA could be a superior strategy for delaying oxidative stress, maintaining nutritional and cooking quality, and ensuring the safety of stored rice. Full article

Spermatogonial stem cells (SSCs) are pivotal in surrogate broodstock technology. However, species-specific protocols for the efficient enrichment and long-term preservation of SSCs in olive flounder (Paralichthys olivaceus) are not yet fully established. In this study, we evaluated and optimized methods for the isolation and cryopreservation of P. olivaceus SSCs. First, we compared two enrichment methods, including Percoll density gradient centrifugation (PDGC) and differential plating (DP). Although SSCs enriched by both methods showed increased expression of SSC-specific marker genes, PDGC resulted in significantly greater enrichment than DP. A combination of PDGC and DP did not further improve enrichment efficiency, suggesting that PDGC alone is sufficient in P. olivaceus. Second, we optimized cryopreservation conditions according to various cryoprotectants. Among the conditions, SSCs cryopreservation using 1.3 M propylene glycol (PG) as a permeating agent and 0.2 M raffinose (Raf) as a non-permeating cryoprotectant provided the highest cell viability (56.1%), demonstrating a synergistic protective effect. Finally, preliminary in vivo migration and localization ability of the cryopreserved SSCs was confirmed through xenotransplantation into zebrafish (Danio rerio) larvae. PKH26-labeled donor cells exhibited successful initial localization and short-term persistence within the presumptive gonadal ridge of the recipients at 5 days post-transplantation. These findings provide an optimized protocol for the handling and preservation of P. olivaceus germline resources, contributing to the technical advancement of surrogate reproduction strategies in this species. Full article

Germination of fenugreek (Trigonella foenum-graecum L.) seeds causes degradation of some antinutritional compounds. At the same time, the content of dietary important compounds, including some carbohydrates, in the sprouts increases. The aim of this study was to investigate changes in the soluble carbohydrate profile during germination and growth of fenugreek sprouts in the roots, hypocotyl and cotyledons. Furthermore, we assessed the effect of cold stress and desiccation on the carbohydrate profile in the main parts of the sprouts. Gas chromatography analyses of soluble carbohydrates showed that fenugreek seeds and sprouts contained sixteen soluble carbohydrates. In dry seeds, the main saccharides were raffinose family oligosaccharides (RFOs), sucrose and d-pinitol. During fenugreek germination, the drastic decomposition of RFOs and galactosides of cyclitols occurred faster in the embryonic axis than in the cotyledons. This was accompanied by an increase in the concentrations of monosaccharides and sucrose, as well as d-pinitol and myo-inositol in the developing hypocotyl and roots. Both examined stresses increased sucrose and raffinose concentration in cotyledons and roots, but in the hypocotyl similar changes were observed only under desiccation. The process of desiccation did not affect the d-pinitol content in the cotyledons of fenugreek sprouts, slightly reduced the content in the hypocotyl, but increased its level in the roots. Applied cold stress did not affect the content of d-pinitol and myo-inositol in the cotyledons and hypocotyl of fenugreek sprouts and only slightly reduced their level in the roots. The obtained results indicate different responses of fenugreek sprout organs to vegetation conditions caused by cold and/or desiccation stress. The practically insignificant effect of cold storage and desiccation on the level of d-pinitol and myo-inositol in fenugreek sprouts is new information that will probably be important for consumers. Full article

This study demonstrates a molecular size-dependent strategy to regulate the network structure of poly(vinyl alcohol) (PVA) hydrogels using a series of saccharides with increasing molecular size—glucose, maltose, raffinose, soluble starch, and amylose. FTIR, XPS, XRD, and TG analyses reveal that increasing saccharide size shifts the network from plasticization to reinforcement, which is further confirmed by mechanical testing and rheological analysis. Small-molecule saccharides disrupt hydrogen bonds and enhance chain mobility, while macromolecular starches promote network regularity through strong hydrogen bonding and crystallization induction. This structural tunability ndows the resulting hydrogels with integrated functionalities: tensile strain increases from 640% to 1500%, self-healing efficiency reaches up to 90.6%, and high-fidelity electrocardiogram (ECG) signal acquisition is achieved with a signal-to-noise ratio of 39.84 dB, comparing favorably with commercial electrodes. This work establishes a structure–property relationship linking saccharide molecular size to network architecture and provides a versatile material platform for next-generation flexible wearable sensors and bioelectrodes. Full article

Chickpea flour represents a valuable plant-based ingredient due to its high protein and fiber content; however, its application is limited by antinutritional factors and off-flavor compounds. Fermentation with LAB and yeasts, applied individually or in consortia, resulted in significant microbiological, nutritional, and aromatic changes. The fastest acidification (pH 3.9) and the most effective control of Enterobacteriaceae (<4 log CFU/g after 48 h) were observed in samples containing Lactiplantibacillus plantarum LP23, both as a monoculture and in combination with Debaryomyces hansenii Y15A. Peptide content significantly increased in all fermented samples compared to the control, with a synergistic effect in the co-culture Yarrowia lipolytica Y3 + Lacticaseibacillus paracasei L (around 230%). A pronounced reduction in raffinose-family oligosaccharides was observed, especially in the consortia Y. lipolytica Y3 + Lcb. paracasei L and D. hansenii Y15A + Lacp. plantarum LP23 (0.11–0.16 mmol/100 g). Samples with lower total volatile levels showed higher olfactory acceptability due to a marked reduction in aldehydes (up to 70–95% vs. control), and a balanced accumulation of alcohols, esters, ketones, and organic acids. Overall, LAB–yeast consortia effectively enhanced the nutritional quality, safety, and sensory properties of chickpea flour, supporting its use as a functional ingredient in plant-based foods. Full article

This study investigated the effects of a low soybean meal (SBM) diet and its supplementation with graded levels of raffinose on the growth performance, expression of genes related to nutrient transport and intestinal function, and cecal microbiota of white-feathered broilers. A total of 480 one-day-old Cobb broilers were randomly allotted to six isoenergetic and isonitrogenous dietary treatments, each with eight replicates of 10 birds. The diets consisted of a positive diet, a low SBM diet (10% reduction in SBM), and the low SBM diet supplemented with 0.10%, 0.15%, 0.20%, or 0.25% raffinose. Results indicated that, compared with the positive diet, the low SBM diet significantly increased (p < 0.05) the overall mortality and average daily feed intake (ADFI) during days 22–42, while significantly decreasing (p < 0.05) dietary ether extract (EE) availability. Raffinose supplementation to the low SBM diet linearly reduced (p < 0.05) dietary gross energy and dry matter utilization and downregulated duodenal SLC5A1 gene expression at 42 days, while linearly increasing (p < 0.05) the cecal isobutyric acid content. A decreasing tendency in mortality during days 22–42 was also observed with raffinose inclusion (p = 0.088). Notably, the low SBM diet elevated the relative abundance of Campylobacterota and Helicobacter, which was effectively reversed by raffinose supplementation. In conclusion, a 10% reduction in dietary SBM negatively affected the survival, nutrient utilization, and cecal microbial structure in broilers, whereas raffinose supplementation partially modulated these alterations. Full article

Freezing temperatures severely restrict plant growth and can cause serious damage or death to sugar beet seedlings in early spring. In this study, seedlings of two sugar beet genotypes, KWS1197 (K, freezing-tolerant) and SX1512 (S, freezing-sensitive), were subjected to RNA sequencing (RNA-seq) after exposure to −4 °C for 0, 1, 3, and 5 h. GO and KEGG enrichment and expression pattern analyses were performed to identify differentially expressed genes (DEGs) associated with freezing stress. The results showed that plant hormone signal transduction and the MAPK signaling pathway were the primary pathways involved in the freezing response in both genotypes. Notably, the galactose metabolism pathway may play an important role in freezing tolerance in KWS1197. Genes involved in galactose metabolism, including inositol 3-α-galactosyltransferase (GolS), raffinose synthase (RafS), invertase (INV), and α-galactosidase (GLA), were significantly up-regulated under freezing conditions, as confirmed by both RNA-seq and RT-qPCR analyses. These results suggest that rapid regulation of soluble sugar synthesis may represent an important mechanism contributing to freezing tolerance in sugar beet. Overall, this study provides a molecular basis for understanding freezing tolerance in sugar beet and identifies candidate genes for improving freezing resistance. Full article

Honeybees are vital pollinators that contribute substantially to global ecosystem stability and agricultural productivity. Camellia reticulata, a cross-pollinated crop species, depends on honeybees for successful reproduction. Apis cerana shows reluctance to pollinate C. reticulata, yet the molecular mechanisms underlying this phenomenon remain unexplored. In this study, we performed controlled feeding experiments in which adult worker A. cerana were supplied with stachyose, raffinose, and their combination. We assessed physiological traits including survival rate, sucrose solution consumption, and body weight gain, alongside histological changes in intestinal cell structures. We conducted RNA-seq of gut tissues as well as 16S rRNA sequencing and metabolomic profiling. Our findings revealed that the mixed oligosaccharide treatment significantly reduced the survival rate of workers, and three oligosaccharide treatments significantly reduced sucrose consumption in A. cerana. Both mixed and single-oligosaccharide treatments caused pronounced intestinal cell damage and disrupted the gut microbial community structure. Among the gut microbes, Gilliamella exhibited the most substantial decline in the stachyose group. Metabolomic analysis further demonstrated that oligosaccharide feeding significantly altered amino acid and galactose metabolism pathways, which may play critical roles in oligosaccharide utilization and directly influence honeybee survival. In summary, this study provides new insights into the molecular mechanisms underlying A. cerana mortality associated with C. reticulata pollination. These findings not only enhance our understanding of host-diet-microbiota interactions in honey bees but also offer a theoretical basis for the integrated management of A. cerana for C. reticulata pollination and the development of oligosaccharide-adapted bee diets. Full article

This study aimed to assess the variation, genetic architecture, and genome-enabled prediction of traits with nutritional and health relevance in 156 pea (Pisum sativum L.) accessions of diverse geographic origins. The traits included the total phenolic compounds (TPCs), two saponins (Ssβg, Ss1), sucrose, three raffinose-family oligosaccharides (RFOs), and the in vitro antioxidant activity (AA). An analysis of variance revealed significant effects of regional germplasm pools for all traits. Accessions from West Asia showed the highest TPC and AA levels, while those from the East Balkans and the UK displayed the lowest values. High saponin and RFO concentrations characterized accessions from Germany and the UK. Correlation and PCA analyses highlighted strong associations within compound classes and an overall negative relationship between TPCs/AA and saponins/RFOs. The accessions were clustered into seven metabolically distinct groups, partially reflecting their geographic origin. The linkage disequilibrium decayed rapidly (average of 4.7 kb). A GWAS based on 10,249 SNP markers identified 37 significant SNPs, 35 within annotated genes, associated with the metabolites, indicating a polygenic genetic architecture. Genomic prediction models showed a moderately high predictive ability (>0.40) for all traits except the raffinose content. Our findings can support line selection and the identification of genetic resources with a desired level of secondary metabolites. Full article

Pulses (edible dry seeds from legumes) are among the most important crops worldwide. These legumes contain a diverse range of carbohydrates, some of which, such as RFOs (raffinose family oligosaccharides), are considered antinutritional factors due to their negative impact on digestion. An analytical method based on high-power ultrasound-assisted extraction and HPLC analysis was developed and validated for the quantitative determination of soluble carbohydrates (verbascose, stachyose, raffinose, sucrose, galactinol, glucose, galactose, fructose, and myo-inositol) in common beans (Phaseolus vulgaris) and peas (Pisum sativum). The proposed method is fast (extraction time: 1 min), reproducible (RDS: 6.9%), accurate (97.5%), and environmentally sustainable. The method was applied to local collections of P. vulgaris (n = 12) and P. sativum (n = 34), revealing similar qualitative profiles but notable quantitative differences. In P. vulgaris, sucrose and stachyose were predominant, while in P. sativum, verbascose stood out. The total sugar content was higher in peas, especially in commercial varieties, which also showed elevated sucrose levels. Some local varieties combined high sugar content with favorable relative levels between RFOs and other sugars, making them valuable candidates for breeding programs. Linear discriminant analysis enabled classification and prediction of species and varieties, confirming the usefulness of soluble carbohydrates as tools for characterizing these plant materials. Full article