Preparation of Multifunctional Hydrogels with In Situ Dual Network Structure and Promotion of Wound Healing.

As an emerging biomedical material, wound dressings play an important therapeutic function in the process of wound healing. It can provide an ideal healing environment while protecting the wound from a complex external environment. A hydrogel wound dressing composed of tilapia skin gelatin (Tsg) and fucoidan (Fuc) was designed in this article to enhance the microenvironment of wound treatment and stimulate wound healing. By mixing horseradish peroxidase (HRP), hydrogen peroxide (H2O2), tilapia skin gelatin-tyramine (Tsg-Tyr), and carboxylated fucoidan-tyramine in agarose (Aga), using the catalytic cross-linking of HRP/H2O2 and the sol-gel transformation of Aga, a novel gelatin-fucoidan (TF) double network hydrogel wound dressing was constructed. The TF hydrogels have a fast and adjustable gelation time, and the addition of Aga further enhances the stability of the hydrogels. Moreover, Tsg and Fuc are coordinated with each other in terms of biological efficacy, and the TF hydrogel demonstrated excellent antioxidant properties and biocompatibility in vitro. Also, in vivo wound healing experiments showed that the TF hydrogel could effectively accelerate wound healing, reduce wound microbial colonization, alleviate inflammation, and promote collagen deposition and angiogenesis. In conclusion, TF hydrogel wound dressings have the potential to replace traditional dressings in wound healing.

Application of definitive screening design to optimization of the protein extraction and functional properties of proteins in Auricularia auricula.

BACKGROUND: Auricularia auricula (A. auricula) is one of the most abundant sources of plant protein in edible fungi. Problems of low protein yield exist in traditional methods of protein extraction such as alkali extraction and ultrasonic-assisted alkali after pretreatment with enzymes. Thus, the protein extraction process was investigated and optimized using a definitive screening design from A. auricula to improve the protein yield under practical operating conditions of temperature, the concentration of NaCl, meal/water ratio, extraction time and pH. RESULTS: The yield of protein isolates of the isoelectric-ammonium sulfate precipitation (9.34% w/w) was obtained almost three times and the protein content (55.23% w/w) was approximately 1.6 times that of the traditional extraction method of isoelectric precipitation. Next, the optimized method was successfully applied to the analysis of the functional properties of the protein. A. auricula protein isolate (AAPI) had better solubility, emulsification and foaming capacity than soy protein isolate (SPI) and pea protein isolate (PPI), and the oil holding capacity of AAPI exhibited extremely well, which was approximately five times that of SPI and six times that of PPI. The texture properties of AAPI gel were similar to those of PPI gels. CONCLUSION: AAPI extracted by the optimized method had a satisfactory yield and had the potential to substitute plant-originated proteins in food processing. © 2022 Society of Chemical Industry.

Structural Characterization and Immunomodulatory Activity of a Polysaccharide from Eurycoma longifolia.

A polysaccharide, Ali-1, was isolated from the roots of Eurycoma longifolia, a popular traditional medicinal herb in Malaysia. The structure of Ali-1 was characterized by monosaccharide, methylation, and NMR data analyses. The average molecular weight of Ali-1 is 14.3 ku, and it is composed of arabinose (14.31%), xylose (57.69%), galacturonic acid (13.03%), and glucuronic acid (14.86%). The main chain comprises (1→4)-linked xylose residues. It has branch points in the main chain; (1→2,4)-linked xylose residues, 1,2-linked glucuronic acid residues, and 1,2-linked arabinose residues form the branches, and the branches are terminated with T-linked galacturonic acid residues and T-linked arabinose residues. Ali-1 significantly improves the pinocytic and phagocytic abilities of RAW264.7 cells and facilitates cytokine secretion according to an immunostimulation assay. These results demonstrate that Ali-1 has potential as a functional supplement for people with compromised immune systems.

Effect of modifications on structure, physicochemical properties and lead ions adsorption behavior of dietary fiber of Flammulina velutipes.

The health effects of dietary fiber have been widely concerned, which are closely related to physicochemical properties. This study focused on soluble dietary fiber of Flammulina velutipes (FDF), evaluated the effects of modifications on structural characterization, the physicochemical properties and the heavy metal adsorption characteristics, and further clarified underlying mechanisms on Pb2+ adsorption behavior of FDFs. The results showed the modifications of extrusion and cellulase improved the yield of FDFs, increased the release of active groups and enhanced the adsorption ability in vitro. Besides, Pb2+ adsorption altered porous structure and led to the presence of carboxylate. It was a spontaneous endothermic reaction and can be fitted by the pseudo-second-order kinetic equation. The Freundlich equation was suitable to describe the adsorption isotherm. These results highlighted potential applications of the dietary fiber modification and laid the theoretical foundation for the modification processing of F. velutipes and protection from food-derived heavy metal toxicity.

Study on the physicochemical properties and antioxidant activities of Flammulina velutipes polysaccharide under controllable ultrasonic degradation based on artificial neural network.

The polysaccharide fraction (FVP2) with molecular weight of 1525.09 kDa and intrinsic viscosity of 3.43 dL/g was isolated and purified from Flammulina velutipes (F. velutipes), and the ultrasonic degradation model of FVP2 was established to predict the molecular weight and intrinsic viscosity at the same time based on artificial neural network. FVP2U1 (1149.11 kDa, 1.78 dL/g), FVP2U2 (618.91 kDa, 1.19 dL/g) and FVP2U3 (597.35 kDa, 0.48 dL/g) with different molecular weights or viscosity were produced by this model to explore the effect of ultrasound on the physicochemical properties and antioxidant activity of FVP2. The results showed that ultrasonic treatment did not change the types of characteristic functional groups, monosaccharide composition and glycosidic bond of FVP2, but changed the chemical composition ratio and the degree of polymerization. Under ultrasonic treatment, the intrinsic viscosity of FVP2 still decreased significantly when the molecular weight did not decrease. Compared to other components subjected to ultrasonic degradation, FVP2U1 demonstrated higher molecular weight and viscoelasticity, while exhibiting lower antioxidant activity. In the case of no significant difference in molecular weight and monosaccharide composition, FVP2U3 with lower intrinsic viscosity has stronger hydration ability, higher crystallization index, lower viscoelasticity and stronger antioxidant capacity than FVP2U2.

Oat protein isolate-Pleurotus ostreatus ß-glucan conjugate nanoparticles bound to ß-carotene effectively alleviate immunosuppression by regulating gut microbiota.

Individuals with immune disorders cannot establish an adequate defense to pathogens, leading to gut microbiota dysbiosis. ß-Carotene can regulate immune response, but its bioavailability in vivo is very low. Herein, we developed a glycosylated oat protein-based nanoparticle to improve the application of ß-carotene for mitigating cyclophosphamide-induced immunosuppression and gut microbiota imbalance in mice. The results showed that the nanoparticles facilitated a conversion of ß-carotene to retinol or retinyl palmitate into the systemic circulation, leading to an increased bioavailability of ß-carotene. The encapsulated ß-carotene bolstered humoral immunity by elevating immunoglobulin levels, augmenting splenic T lymphocyte subpopulations, and increasing splenic cytokine concentrations in immunosuppressed mice. This effect was accompanied by the alleviation of pathological features observed in the spleen. In addition, the encapsulated ß-carotene restored the abnormal gut microbiota associated with immunosuppression, including Erysipelotrichaceae, Akkermansia, Bifidobacterium and Roseburia. This study suggested that nanoparticles loaded with ß-carotene have great potential for therapeutic intervention in human immune disorders by specifically targeting the gut microbiota.

Structure and immunostimulatory activity studies on two novel Flammulina velutipes polysaccharides: revealing potential impacts of →6)-α-D-Glcp(1→ on the TLR-4/MyD88/NF-κB pathway.

Two novel Flammulina velutipes (F. velutipes) polysaccharides, FVPH1 and FVPH2, were isolated and purified after hot water extraction. The structural characterization revealed that the backbone of FVPH1 consisted mainly of →6)-α-D-Glcp(1→, →3,4)-α-D-Galp(1→, →4)-α-L-Fucp(1→, and →4)-ß-D-Manp(1→, while the backbone of FVPH2 consisted of →3)-α-D-Galp(1→, →3,4)-α-D-Manp(1→,→6)-α-D-Glcp(1→. The branches of FVPH1 contained →6)-α-D-Glcp(1→ and α-D-Glcp(1→ and the branches of FVPH2 consisted of →3)-α-D-Galp(1→, →6)-α-D-Glcp(1→, and ß-L-Fucp(1→. FVPH2 exhibited significantly better immunostimulatory activity than FVPH1 (P < 0.05), as evidenced by the increased expression of NO, IL-1ß, IL-6, and TNF-α and pinocytic activity of RAW264.7 cells. As the most abundant structure in the polysaccharides of F. velutipes, the content of →6)-α-D-Glcp(1→ might play a crucial role in influencing the immunostimulatory activity of F. velutipes polysaccharides. The F. velutipes polysaccharide with a lower content of →6)-α-D-Glcp(1→ and a higher branching degree could significantly enhance the immunostimulatory activity of F. velutipes polysaccharides via activating the TLR-4/MyD88/NF-κB pathway more effectively.

Improved bioavailability and antioxidation of ß-carotene-loaded biopolymeric nanoparticles stabilized by glycosylated oat protein isolate.

The limited bioavailability of ß-carotene hinders its potential application in functional foods, despite its excellent antioxidant properties. Protein-based nanoparticles have been widely used for the delivery of ß-carotene to overcome this limitation. However, these nanoparticles are susceptible to environmental stress. In this study, we utilized glycosylated oat protein isolate to prepare nanoparticles loaded with ß-carotene through the emulsification-evaporation method, aiming to address this challenge. The results showed that ß-carotene was embedded into the spherical nanoparticles, exhibiting relatively high encapsulation efficiency (86.21 %) and loading capacity (5.43 %). The stability of the nanoparticles loaded with ß-carotene was enhanced in acidic environments and under high ionic strength. The nanoparticles offered protection to ß-carotene against gastric digestion and facilitated its controlled release (95.76 % within 6 h) in the small intestine, thereby leading to an improved in vitro bioavailability (65.06 %) of ß-carotene. This improvement conferred the benefits on ß-carotene nanoparticles to alleviate tert-butyl hydroperoxide-induced oxidative stress through the upregulation of heme oxygenase-1 and NAD(P)H quinone dehydrogenase 1 expression, as well as the promotion of nuclear translocation of nuclear factor-erythroid 2-related factor 2. Our study suggests the potential for the industry application of nanoparticles based on glycosylated proteins to effectively deliver hydrophobic nutrients and enhance their application.

Dual structure on-demand release chitosan-based coating film for peach preservation.

After harvest, fruits and vegetables go through the stages of storage, transportation and sales, and how to regulate the role of preservative in different stages is an urgent problem to be solved. Herein, chitosan-stearic acid is synthesized, then interacting with trans-2-hexenal (E2H) to prepare nanoclusters. Subsequently, the temperature-sensitive liposomes were designed to encase E2H by using the special temperature-responsive open-chain phospholipids, and then combined with chitosan to obtain chitosan-based coating film for peach preservation. The thermosensitive liposomes were uniformly distributed, highly dispersed and spherical, which has excellent temperature response characteristics, and the release of active substances could be regulated in the temperature range of 10 °C-40 °C to achieve the effect of on-demand release. In addition, thermosensitive liposomes also have excellent antioxidant and antibacterial activities. Therefore, the obtained chitosan-based coating could maintain the freshness of peaches, improve the quality and extend the storage period of peaches. The edible chitosan coating and active preservative prepared in this study have potential application value in responsive intelligent active packaging.

Effects of different selenium biofortification methods on Pleurotus eryngii polysaccharides: Structural characteristics, antioxidant activity and binding capacity in vitro.

The effects of selenium biofortification methods involving sodium selenite and selenium yeast on the structural characteristics, antioxidant activity and binding capacity of Pleurotus eryngii polysaccharides were investigated. Sodium selenite Se-enriched Pleurotus eryngii polysaccharides (Se-SPEP), selenium yeast Se-enriched Pleurotus eryngii polysaccharides (Se-YPEP), and Pleurotus eryngii polysaccharides (PEP) had Se contents of 20.548 ± 1.561, 19.822 ± 0.613, and 0.052 ± 0.016 µg/g, respectively. Compared with PEP, Se-SPEP and Se-YPEP had lower molecular weight and contained the same monosaccharides in varying molar ratios. The results of FT-IR, PS, ZP, and SEM indicated significant alterations in structural characteristics following selenium biofortification. Se-PEPs exhibited superior activity against ABTS, DPPH, and ·OH radicals, as well as the higher binding capacity for Cd2+ and Cu2+ compared to natural polysaccharides. The binding capacity of the polysaccharides for Cd2+ and Cu2+ was higher at pH 6.8 compared to pH 2.0, while the opposite was observed for Pb2+. Furthermore, Se-PEPs exhibited a significantly higher binding capacity for Cd2+ and Cu2+ at both pH levels compared to natural polysaccharides (P < 0.05). Se-YPEP displayed higher antioxidant activity than Se-SPEP, with their binding capacities reversed. These data indicated that selenium biofortification methods have different positive impacts on the structure and activity of polysaccharides compared to natural polysaccharides, making Se-PEPs promising dietary supplements for safeguarding the body against the risks posed by food-derived heavy metals.

Exploring gelation properties and structural features on 3D printability of compound proteins emulsion gels: Emphasizing pH-regulated non-covalent interactions with xanthan gum.

Rational regulation of pH and xanthan gum (XG) concentration has the potential to modulate interactions among macromolecules and enhance 3D printability. This study investigated non-covalent interactions between XG and other components within compound proteins emulsion gel systems across varying pH values (4.0-8.0) and XG concentrations (0-1 wt%) and systematically explored impacts of gelation properties and structural features on 3D printability. The results of rheological and structural features indicated that pH-regulated non-covalent interactions were crucial for maintaining structural stability of emulsion gels with the addition of XG. The 3D printability of emulsion gels would be significantly improved through moderate depletion flocculation produced when XG concentration was 0.75 wt% at the pH 6.0. Mechanical properties like viscosity exhibited a strongly negative correlation with 3D printability, whereas structural stability showed a significantly positive correlation. Overall, this study provided theoretical insights for the development of emulsion gels for 3D printing by regulating non-covalent interactions.

Effects of selenylation modification on the antioxidative and immunoregulatory activities of polysaccharides from the pulp of Rose laevigata Michx fruit.

Selenylation modification has been widely utilized to improve the activity of polysaccharides and to develop novel sources of selenium (Se) supplements. A purified pulp polysaccharide of Rose laevigata Michx fruit (PPRLMF-2) was selenized into Se-PPRLMF-2 in this study. PPRLMF-2 + Se was formulated by Na2SeO3 according to the Se content of Se-PPRLMF-2. To investigate the effects of selenylation modification on the structure and functions of PPRLMF-2, the characteristics, antioxidative and immunoregulatory activities of PPRLMF-2 before and after selenylation were compared. The results showed that compared with PPRLMF-2, Se-PPRLMF-2 became an irregular fibrous network, and its Mw decreased and C-6 substitution predominated in 13C NMR spectra. Se-PPRLMF-2 significantly increased chemical antioxidant activity and reduced the oxidative damage of erythrocytes, which was not due to Se alone. Se-PPRLMF-2 significantly increased immunomodulatory activity on macrophages, which was related to Se alone. Se-PPRLMF-2 could be a good potential source of antioxidants, immune enhancers and dietary Se supplements.

Effects of ultrasound on the degradation kinetics, physicochemical properties and prebiotic activity of Flammulina velutipes polysaccharide.

The controllable ultrasonic modification was hindered due to the uncertainty of the relationship between ultrasonic parameters and polysaccharide quality. In this study, the ultrasonic degradation process was established with kinetics. The physicochemical properties and prebiotic activity of ultrasonic degraded Flammulina velutipes polysaccharides (U-FVPs) were investigated. The results showed that the ultrasonic degradation kinetic models were fitted to 1/Mt-1/M0 = kt. When the ultrasonic intensity increased from 531 to 3185 W/cm2, the degradation proceeded faster. The decrease of polysaccharide concentration contributed to the degradation of FVP, and the fastest degradation rate was at 60 °C. Ultrasound changed the solution conformation of FVP, and partially destroyed the stability of the triple helix structure of FVP. Additionally, the viscosity and gel strength of FVP decreased, but its thermal stability was improved by ultrasound. Higher ultrasonic intensity led to larger variations in physicochemical properties. Compared with FVP, U-FVPs could be more easily utilized by gut microbiota. U-FVPs displayed better prebiotic activity by promoting the growth of Bifidobacterium and Brautella and inhibiting the growth of harmful bacteria. Ultrasound could be effectively applied to the degradation of FVP to improve its physicochemical properties and bioactivities.

Purification and comparative study of bioactivities of a natural selenized polysaccharide from Ganoderma Lucidum mycelia.

The development of selenized polysaccharides is a promising strategy for the dietary selenium supplementation. The purpose of this research is to determine the influence of selenium on the structure and bioactivity of a polysaccharide fraction (MPN) isolated from Ganoderma lucidum mycelia. After biological selenium enrichment, the selenium content in the selenized polysaccharide (SeMPN) was 18.91 ± 1.8 µg/g. SeMPN had a slightly lower molecular weight than MPN, but the carbohydrate content and monosaccharide composition remained identical. Additionally, the band at 606 cm-1 in MPN changed to 615 cm-1 in SeMPN as revealed by FT-IR spectra. No significant changes were observed in the types and ratios of glycosidic linkages, as determined by NMR spectroscopy. Extracellular and intracellular antioxidant assays demonstrated that SeMPN was more effective than MPN in scavenging free radicals, inhibiting AAPH-induced erythrocyte hemolysis, and protecting catalase (CAT) and glutathione peroxidase (GSH-Px) activity in H2O2-injured PC12 cells. Additionally, SeMPN had a higher increase effect on RAW 264.7 cells's pinocytic and phagocytic capacity, as well as their production of NO, TNF-α, and IL-6. SeMPN could be as potential functional selenium supplementation.

Purification and characterization of immunomodulatory peptides from enzymatic hydrolysates of duck egg ovalbumin.

Duck egg white (DEW) is considered as an abandoned protein resource. For a higher-value utilization, preparation of immunomodulatory peptides from the extracted ovalbumin in DEW was explored. Among the hydrolysates catalyzed by five proteases, papain hydrolysate (PH) shows the highest degree of hydrolysis and the strongest immunomodulatory activity. PH could significantly enhance the phagocytic capacity and promote the NO, TNF-α, and IL-6 secretion of RAW 264.7 cells, involving toll-like receptor 2 and 4. After purification, nine identified peptides were synthesized to confirm the immunomodulatory effect, and five of them exhibit a strong activity. The peptide, TQIDKVVHFDKLPGF, presents the highest immunomodulatory activity. Moreover, the results of molecular docking indicate that nine peptides interacted with toll-like receptor 2 and 4 and all show good affinity. Furthermore, three peptides with high affinity and strong immune activity were selected for interaction site map analysis. Three peptides could form hydrogen bonds with the receptor and bind stably, which contributes to the immunomodulatory activity of the peptide. Results suggest that DEW can be a promising source of immunomodulatory peptides.

Suppression of lipopolysaccharide-induced activation of RAW 264.7 macrophages by Se-methylseleno-l-cysteine.

Se-methylseleno-l-cysteine (l-SeMC) is a natural source of organic selenium for humans. Although it has a structure similar to that of l-Cysteine (l-Cys), its anti-inflammatory properties and possible underlying mechanisms have not been explored. Here, we compared the anti-inflammatory activities of inorganic selenium (selenite), l-Cys, and l-SeMC in lipopolysaccharide (LPS)-activated RAW 264.7 murine macrophages and focused on the related molecular and biochemical events. The results showed that, anti-inflammatory activity of l-SeMC was much stronger compared to both individual l-Cys treatment and l-Cys/selenite combinations. The organic form of selenium may play a crucial role in the effects of l-SeMC. Further study confirmed that l-SeMC suppressed the RNA expression of iNOS, TNF-α, IL-1ß, IL-6, COX-2, and MMP-9, as well as the release of NO, TNF-α, IL-6, IL-12p70, COX-2, and PGE2 from LPS-activated RAW264.7 macrophages in a concentration-dependent manner. Moreover, l-SeMC prevented LPS-induced changes in cell morphology. l-SeMC concentrations between 50 and 200 µM exhibited an anti-inflammatory effect closed to that exhibited by 20 µM dexamethasone. Our results demonstrated that l-SeMC effectively inhibited the activation of RAW 264.7 macrophages induced by LPS, and suggested that l-SeMC could be a potential functional food component for the prevention or treatment of inflammatory diseases.

Structural characterization and immunomodulatory activity of a novel acid polysaccharide isolated from the pulp of Rosa laevigata Michx fruit.

A novel acid polysaccharide (PPRLMF-2) with the Mw of 137,123 Da and a triple-helix conformation was first isolated from the pulp of Rosa laevigata Michx fruit. Structural characterization showed that PPRLMF-2 consisted of rhamnose (7.6%), arabinose (26.5%), xylose (3.5%), mannose (0.9%), glucose (5.7%), galactose (31.9%) and galacturonic acid (23.9%). The methylation and NMR (1D and 2D) analysis revealed that PPRLMF-2 contained 16 types of glycosidic linkages. The immunomodulatory activity assays indicated that PPRLMF-2 could significantly enhance phagocytosis, the secretion and mRNA expression of cytokines in RAW 264.7 cells. Additionally, SR, GR, TLR-2, and TLR-4 were the main pattern recognition receptors (PRRs) of PPRLMF-2 to upregulate the p-ERK, p-JNK, p-p38, and p-p65. These results suggested that PPRLMF-2 could recognize the PRRs of the macrophages to enhance the immunomodulatory activity via activation of the MAPKs and NF-κB signaling pathways. This study provides important implications of PPRLMF-2 as an attractive immunomodulatory functional food.

Structural characterization and immunomodulatory activity of a novel polysaccharide from Pueraria lobata (Willd.) Ohwi root.

A novel polysaccharide with a molecular weight of 12.3 kDa, was isolated from the root of Pueraria lobata (Willd.) Ohwi using cold water extraction method. Methylation analysis, Periodate oxidation, Smith degradation, and nuclear magnetic resonance analysis showed that 1,4-α-D-Glcp and 1,3,6-α-D-Glcp were the main linkage types of this polysaccharide. It exhibited immunomodulatory activity by enhancing the pinocytic and phagocytic capacities and promoting the secretion of nitric oxide, interleukin (IL)-6, and tumor necrosis factor alpha by RAW 264.7 cells. The beta glucan receptor, scavenger receptor I, and toll-like receptor 4 were identified as the main receptors of the polysaccharide on the membrane of RAW 264.7 cells. These results suggested that this polysaccharide is a candidate functional food supplement for hypoimmune populations.

Structural characterization of polysaccharides with potential antioxidant and immunomodulatory activities from Chinese water chestnut peels.

Chinese water chestnut peels are a kind of vegetable processing waste containing many active components such as polysaccharides, the structure of which remains unknown. To elucidate the structure of polysaccharides from Chinese water chestnut peels, two polysaccharides named WVP-1 and WVP-2 were isolated. WVP-1 (3.16 kDa) consisted of mannose (1.75 %), glucose (84.69 %), galactose (6.32 %), and arabinose (7.24 %), while WVP-2 (56.97 kDa) was composed of mannose (3.18 %), rhamnose (1.52 %), glucuronic acid (1.42 %), galacturonic acid (4.83 %), glucose (11.51 %), galactose (36.02 %), and arabinose (41.53 %). Linkage and NMR data indicated that WVP-1 was composed mainly of →4)-α-d-Glcp(1→ and a certain proportion of →3)-ß-d-Glcp-(1→, including linear and branched polysaccharides simultaneously. WVP-2 was a pectin-like polysaccharide with →4)-α-d-GalpA6Me-(1→ units and the branch points of →3,4)-α-l-Arap-(1→, →3,6)-ß-d-Galp-(1→. WVP-2 exhibited stronger potential antioxidant and immunomodulatory activities than WVP-1 in vitro. These results provide a foundation for the further study of polysaccharides from Chinese water chestnut peels.

Comparative study of DHA-enriched phosphatidylcholine and EPA-enriched phosphatidylcholine on ameliorating high bone turnover via regulation of the osteogenesis-related Wnt/ß-catenin pathway in ovariectomized mice.

Here, we compared the effects of marine DHA-enriched phosphatidylcholine (DHA-PC) and EPA-enriched phosphatidylcholine (EPA-PC) on high bone turnover in a model of osteoporosis induced by bilateral ovariectomy in vivo, and further investigated the possible protective mechanisms. Meanwhile, DHA-PC and EPA-PC clearly ameliorated the microstructure of the trabecular bone and accelerated bone mineral apposition rate, additionally increasing bone mineral density and biomechanical properties of the bone. Furthermore, gene and protein expression levels suggest that DHA-PC and EPA-PC inhibited overactive osteogenesis via down-regulation of the expression of the osteogenesis-related Wnt/ß-catenin signaling pathway. In conclusion, DHA-PC and EPA-PC reduced excessive osteogenesis via normalization of Wnt/ß-catenin expression. These results may contribute to the elucidation of the anti-osteoporotic properties of DHA-PC and EPA-PC and further develop their potential application value as a functional food.