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. 2024 Jun 28;33(9):2243–2254. doi: 10.1007/s10068-024-01640-x

Enhanced protective effect of whey protein fermented with Lacticaseibacillus rhamnosus IM36 on dexamethasone-induced myotube atrophy

Ji Hun Jang 1, Hyeon Ho Jung 1, Nam Su Oh 1,✉
PMCID: PMC11315874  PMID: 39130659

Abstract

This study investigated the preventive potential of whey protein fermented with Lacticaseibacillus rhamnosus IM36 (FWP) against muscle atrophy induced by dexamethasone (DEX). FWP exhibited enhanced antioxidant activities compared with those of unfermented whey protein, effectively suppressing DEX-induced reactive oxygen species production. FWP was treated before the administration of 100 μM DEX on C2C12 myotubes and compared to unfermented whey (WP). DEX significantly inhibited myotube viability and muscle protein synthesis and enhanced degradation. FWP exhibited a dose-dependent attenuation of cell viability loss compared with that of WP. Additionally, FWP stimulated the formation of myotubes and muscle protein synthesis by upregulating myogenesis and insulin-like growth factor-1 expression. Furthermore, FWP significantly attenuated forkhead box protein O3a-mediated ubiquitin ligases and autophagy of lysosomes activated by DEX, inhibiting pathways that lead to muscle protein breakdown. These findings suggest that FWP enhances antioxidant activity and prevented DEX-induced muscle atrophy by regulating muscle protein homeostasis.

Supplementary Information

The online version contains supplementary material available at 10.1007/s10068-024-01640-x.

Keywords: Fermented whey, Lacticaseibacillus rhamnosus, Muscle atrophy, Antioxidant activity, Dexamethasone

Introduction

Skeletal muscles, accounting for approximately 40% of body weight, are vital for movement and energy metabolism (Cruz-Jentoft et al., 2019). Muscle atrophy, resulting from an imbalance in protein synthesis and degradation, leads to wasting and reduced muscle strength (Yin et al., 2021). The mechanisms underlying muscle atrophy involve an intricate interplay between the protein synthesis and degradation pathways (Sartori et al., 2021). Upstream signals, such as oxidative stress and inflammation, trigger this process, whereas downstream systems, such as the ubiquitin–proteasome and autophagy-lysosome systems, degrade proteins (Yin et al., 2021). Insulin-like growth factor-1 (IGF-1) plays a pivotal role in skeletal muscle regulation by promoting protein synthesis and inhibiting the protein degradation pathway mediated by forkhead box protein O3a (FOXO3a) through the ubiquitin proteasome system (UPS) and autophagy lysosomal system (ALS) (Yoshida and Delafontaine, 2020). Dexamethasone (DEX) is widely used to induce muscle atrophy, primarily by stimulating autophagy, mitochondrial fragmentation, and inflammation in skeletal muscle cells, contributing to the breakdown of muscle proteins and the disruption of muscle homeostasis, ultimately leading to muscle wasting (Sartori et al., 2021).

Whey protein, once considered dairy waste, is now valued for its high nutrient content, particularly its rich source of essential amino acids and branched-chain amino acids, such as leucine, valine, and isoleucine (Minj and Anand, 2020). Owing to its high leucine content and digestibility, whey protein stimulates protein synthesis more effectively than other protein sources, such as soy and casein (Dijk et al., 2018). Whey proteins and their derived peptides have been reported to exhibit diverse bioactivities, including antioxidant, anti-inflammatory, and immunomodulatory effects, suggesting their potential for managing various diseases when incorporated into functional foods (Ali et al., 2022). Notably, whey protein fermentation by lactic acid bacteria (LAB) produces bioactive peptides with enhanced antioxidant activities that exhibit protective effects against muscle atrophy (Corrochano et al., 2018; Jang et al., 2023).

The aim of this study was to evaluate the preventive effects of the metabolic byproducts of whey fermentation by LAB on DEX-induced muscle atrophy. Using specific LAB strains, whey proteins were fermented, and their antioxidant activities determined. Based on its functionality, whey protein fermented with the Lacticaseibacillus rhamnosus IM36 strain (FWP) was selected, and the preventive effect of FWP on DEX-induced C2C12 myotubes was demonstrated.

Materials and methods

Preparation of microorganisms and media

Previously, LAB strains were isolated from infant feces, and their probiotic properties, such as resistance to acid and bile salts, adhesion to intestinal cells and antioxidant activities were evaluated, and 4 LAB candidates with high probiotic properties were used in this study (data not shown). These strains were identified using 16S rRNA sequencing (Supplementary Table 1). All strains were cultured in de Man, Rogosa, and Sharpe broth (Difco Laboratory, MI, USA) at 37 °C for 21 h for inoculum preparation. Then the strains were washed and resuspended thrice in sterile saline (0.85% NaCl solution). Whey protein isolate (Carbery Food Ingredients Limited, Ballineen, Ireland) was mixed with glucose at a 10:1 (w/w) ratio, dissolved in deionized water to a final concentration of 5% (w/v), and sterilized at 85 °C for 5 min.

Fermentation and sample preparation

The washed bacteria were inoculated into the sterilized whey mixtures to a 0.5% (v/v) final concentration and incubated at 85 °C for 48 h. In preparation for the control group, a solution containing the same ratio of whey protein isolate to glucose without fermentation was prepared under the same conditions (WP). Following the fermentation process, the solutions were centrifuged (26,000×g, 10 min, 4 °C), and the supernatants were lyophilized and stored at − 80 °C.

Assessment of antioxidant activity

The antioxidative activities of whey fermented with the candidate strains were evaluated using hydroxyl (OH) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assays, and ferric-reducing antioxidant power (FRAP) assays. The OH radical scavenging activity assay was performed as described by Li et al. (2014), and DPPH and FRAP assays were carried out as described by Oh et al. (2014).

Degree of hydrolysis

The degree of hydrolysis during whey fermentation was evaluated using the o-phthaldialdehyde (OPA) method described by Nielsen et al. (2001) with some modifications. A leucine standard curve (0–10 mM) was used to calculate the total amount of peptide released. Briefly, the OPA reagent was prepared by mixing 1 mL of 40 mg/mL OPA solution (in methanol), 25 mL of 0.1 M sodium tetraborate, 2.5 mL of 20% (w/w) sodium dodecyl sulfate, and 100 µL of β-mercaptoethanol. Then, 10 µL of samples and standards were mixed with 180 µL of the OPA reagent and shaken at 24 °C for 5 min. After the reaction, the absorbance was measured at 340 nm using a BioTek Epoch 2 microplate reader (BioTek Instruments Inc., Winooski, VT, USA).

pH measurement and viable cell count

During whey fermentation by the selected bacteria, pH changes were evaluated at 3-h intervals using a calibrated pH meter with standardized buffer solutions (pH 4.0, 7.0, and 10.0; Thermo Fisher Scientific, Waltham, MA, USA). In addition, the total number of bacterial cells in the fermented whey was measured using the plate counting method. The fermented solution was serially diluted with sterile saline and inoculated onto de Man, Rogosa, and Sharpe agar plates. The plates were incubated at 37 °C for 48 h. The viable cell count was expressed as log CFU/mL.

Cell culture and treatment

C2C12 myoblasts were purchased from the American Type Culture Collection (Manassas, VA, USA). Cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 10% fetal bovine serum (Gibco) and 1% penicillin–streptomycin (Gibco) at 37 °C in a humidified atmosphere containing 5% CO2. C2C12 cells were seeded in a six-well plate at a density of 5 × 104 cells/well. When the myoblasts reached 90–100% confluence, the medium was replaced with differentiation medium (DM), DMEM supplemented with 2% horse serum (Gibco), and 1% penicillin–streptomycin to differentiate myoblasts into myotubes. The media was replaced every two days. After four days of pre-differentiation, WP and fermented whey were dissolved in DM and used to treat C2C12 myotubes for 48 h. After sample treatment, DEX (Sigma-Aldrich Chemical Co.) was dissolved in serum-free DMEM containing 1% penicillin–streptomycin to a 100 μM concentration and treated for 24 h to induce myotube atrophy. Normal DM and serum-free DMEM were used as controls.

Myotube viability

The effects of the sample and DEX treatment on myotube viability were measured using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT; Sigma-Aldrich Chemical Co.) assay. After DEX treatment, the medium was replaced with 0.5 mg/mL MTT reagent and incubated in the dark at 37 °C for 2 h. Subsequently, the MTT reagent was removed, and dimethyl sulfoxide was added. The absorbance of the lysates was measured at 540 nm using a microplate reader and expressed as a percentage of that of the control group.

Intracellular reactive oxygen species (ROS) measurements

To quantitatively assess ROS generation, cells were incubated with 2.5 μM 2′,7′-dichlorofluorescein diacetate (Sigma-Aldrich Chemical Co.) at 37 °C for 30 min. Subsequently, the fluorescence intensity of 2′,7’-dichlorofluorescein was detected using a Tecan microplate reader (Tecan Life Sciences, Männedorf, Switzerland) at excitation and emission wavelengths of 480 and 530 nm, respectively.

Quantitative reverse transcription polymerase chain reaction assay (qRT-PCR)

Total RNA isolation, cDNA synthesis, and qRT-PCR were performed as previously described (Jang et al., 2023). The relative fold changes of the examined genes were analyzed using the 2−ΔΔCT method (Livak and Schmittgen, 2001) and normalized with the housekeeping gene GAPDH. The primers used in this study are listed in Supplementary Table 2.

Statistical analysis

All experimental data were expressed as means ± standard error of the mean (SEM). Statistical analysis was conducted using SPSS software (version 25.0; IBM, Chicago, IL, USA). Differences between groups were compared using one-way analysis of variance and Duncan’s multiple comparison test. Statistical significance was set at p < 0.05.

Results and discussion

Antioxidant activities of fermented whey protein by LAB strains

Recent studies have demonstrated that antioxidants have therapeutic potential for promoting muscle hypertrophy and preventing muscle atrophy through mechanisms that suppress ROS-mediated oxidative stress and gene expression related to muscle atrophy (Chen et al., 2022; Jang et al., 2023). Previous studies have demonstrated that LAB fermentation of whey produces bioactive metabolites with antioxidant activities (Mann et al., 2019). In this study, we fermented whey proteins using four probiotic strains and evaluated their antioxidant activities by using the OH radical, DPPH, and FRAP assays (Fig. 1A–C). Whey fermentation with the four LAB strains significantly increased antioxidant activity (p < 0.05). Whey fermented with L. rhamnosus IM34 and IM36 exhibited significantly higher radical-scavenging activity and reducing power than those of whey fermented with the other strains and WP (p < 0.05). Moreover, the ROS reductions in L. rhamnosus IM34 and IM36 in DEX-treated muscle cells were markedly higher than those of other candidates, with values of 75.5% and 75.1%, respectively (p < 0.05; Fig. 1D). Both L. rhamnosus strains showed significantly increased antioxidant activities during whey fermentation.

Fig. 1.

Fig. 1

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Antioxidant activities of whey fermented by selected probiotic strains. (A) Hydroxyl radical scavenging activity, (B) 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity, (C) Ferric reducing antioxidant power (FRAP) value, (D) Relative reactive oxygen species (ROS) level in dexamethasone-induced myotube. The data are presented as the mean ± standard errors of mean (SEM) with error bars indicating the standard error (n = 3). Statistically significant differences (p < 0.05) are denoted by different small letters

Antiatrophic activities of fermented whey protein by LAB strains

To confirm the preventive effect on muscle atrophy, myotubes were treated with the fermentation candidates before DEX administration by evaluating the mRNA expression of genes related to myotube atrophy was measured after DEX treatment (Fig. 2). DEX inhibits muscle protein synthesis via the IGF-1 pathway, disrupts myogenesis, and stimulates muscle proteolysis via the FOXO3a-mediated pathway, resulting in muscle atrophy (McGrath et al., 2021; Yoshida and Delafontaine, 2020). WP exhibited preventive effects against DEX-induced muscle protein degradation by decreasing the mRNA expression of AMP-activated protein kinase (AMPK), FOXO3a, Atrogin-1, and Muscle RING-finger protein-1 (MuRF1) (Fig. 2A–D). Whey fermented with L. reuteri IR02 and L. gasseri IR13 showed no significant difference in mRNA expression related to myotube atrophy compared with that of WP. However, whey fermented with L. rhamnosus IM34 and IM36 decreased the mRNA expression of AMPK, FOXO3a, Atrogin-1, and MuRF1. Notably, whey fermented by L. rhamnosus IM36 exhibited a significant decrease in AMPK, FOXO3a, and MuRF1 levels compared with those of WP (p < 0.05), restoring AMPK and FOXO3a mRNA levels to those of the normal group. Moreover, the mRNA expression levels of IGF-1 and myoblast determination protein 1 (MyoD) were significantly increased by treatment with whey fermented with L. rhamnosus IM36 (p < 0.05; Fig. 2E, F). Based on its functionality, whey fermented with L. rhamnosus IM36 (FWP) was determined to have the potential to prevent muscle atrophy and was used for further studies.

Fig. 2.

Fig. 2

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Preventive effect of whey fermented by select probiotics strains on muscle atrophy in dexamethasone-induced muscle cell. C2C12 myotubes were treated with unfermented whey (WP) or whey fermented with probiotic strains for 48 h and then 100 μM DEX for 24 h. The mRNA expression levels were quantified by quantitative reverse transcription polymerase chain reaction assay (qRT-PCR) and GAPDH was used as a housekeeping gene (A) AMP-activated protein kinase (AMPK), (B) forkhead box protein O3a (FOXO3a), (C) Atrogin-1, (D) muscle RING-finger protein-1 (MuRF1), (E) insulin-like growth factor-1 (IGF-1), (F) myoblast determination protein 1 (MyoD) expression level. The data are presented as the mean ± standard errors of mean (SEM) with error bars indicating the standard error (n = 3). Statistically significant differences (p < 0.05) are denoted by different small letters. NOR, normal group; DEX, dexamethasone only treated group; WP, unfermented whey; IR02, whey fermented by Limosilactobacillus reuteri IRO2; IR13, whey fermented by Lactobacillus gasseri IR13; IM34, whey fermented by Lacticaseibacillus rhamnosus IM34; IM36, whey fermented by Lacticaseibacillus rhamnosus IM36

Growth kinetics and changes in pH

To evaluate the changes in microbiological characteristics during whey fermentation with IM36, the degree of hydrolysis, viable cell count, and pH were measured every three hours (Fig. 3). Because the amount of free amino acids in whey is restricted, the continuous growth of L. rhamnosus IM36 depends on proteolytic enzymes (proteinases and peptidases) and specific amino acid and peptide transport systems (Pescuma et al., 2008). In this study, the degree of hydrolysis increased from 0.603 to 4.127 mM leucine equivalents, and the viable cell count of L. rhamnosus IM36 in whey increased from 6.78 to 8.96 log CFU/mL (Fig. 3A). There was an exponential increase in viable cell count between 0 and 12 h. The pH of the fermented samples decreased from 6.48 to 4.50 (Fig. 3B). To evaluate the functionality of postbiotics derived from whey fermentation by L. rhamnosus IM36, FWP supernatants were lyophilized, administered to DEX-treated C2C12 myotubes, and compared to unfermented WP.

Fig. 3.

Fig. 3

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Changes in (A) degree of hydrolysis and viable cell counts and (B) pH during whey fermentation with Lacticaseibacillus rhamnosus IM36. The data are presented as the mean ± standard errors of mean (SEM) with error bars indicating the standard error (n = 3)

Effect of FWP on myotube viability

To assess the cytotoxic and protective effects of WP and FWP on C2C12 myotubes, myotubes were treated with different concentrations of WP and FWP, and an MTT assay was performed (Fig. 4). Recent studies have shown that DEX exposure reduces myotube viability and inhibits myotube formation (Jang et al., 2023; Lee et al., 2022). DEX treatment decreased myotube viability to 58.2%; however, WP and FWP groups markedly increased myotube viability (p < 0.001). The FWP group significantly increased myotube viability compared with that of the WP group at all concentrations (100 μg/mL, p < 0.005; 500 μg/mL, p < 0.001; 1000 μg/mL, p < 0.001). Moreover, the FWP group showed a dose-dependent increase in myotube viability (p < 0.001), restoring the viability to that of the normal group.

Fig. 4.

Fig. 4

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Effect on myotube viability of whey fermented with Lacticaseibacillus rhamnosus IM36. C2C12 myotubes were treated with various concentrations (100, 500, and 1000 μg/mL) of unfermented whey (WP) or whey fermented with Lacticaseibacillus rhamnosus IM36 (FWP) for 48 h and then 100 μM dexamethasone for 24 h. The data are presented as the mean ± standard errors of mean (SEM) with error bars indicating the standard error (n = 3). ***p < 0.001, **p < 0.005 represent significant differences between marked groups. NOR, normal group; DEX, dexamethasone only treated group

Effect of FWP on muscle protein synthesis and myogenesis

Previous studies have demonstrated that IGF-1 is pivotal in regulating protein synthesis pathways in skeletal muscles, and its dysregulation under DEX-induced conditions contributes to muscle atrophy (Mishra et al., 2022; Yoshida and Delafontaine, 2020). Moreover, DEX directly inhibited muscle protein synthesis by targeting protein regulated in development and DNA damage-response 1 (REDD1) (Shimizu et al., 2011). REDD1, a direct target of the glucocorticoid receptor, inhibits muscle protein synthesis by inhibiting mammalian target of rapamycin activity, further promoting muscle degradation. DEX induces oxidative stress and activates the muscle atrophy-associated ubiquitin ligase casitas B-lineage lymphoma proto-oncogene-b (Cbl-b), initiating muscle protein ubiquitination and implicating a complex interplay between ROS, protein synthesis, and muscle atrophy pathways (Uchida et al., 2018). In this study, the mRNA expression of IGF-1, REDD1, and Cbl-b was assessed by qRT-PCR to evaluate the preventive effects of FWP on muscle synthesis (Fig. 5). DEX administration substantially decreased the IGF-1 levels. However, the FWP group showed a dose-dependent increase in IGF-1 levels (p < 0.001; Fig. 5A). Notably, FWP demonstrated a significant increase in the expression of IGF-1 compared with that of WP. The mRNA expression levels of REDD1 and Cbl-b were markedly increased by DEX treatment (p < 0.001; Fig. 5B, C). FWP suppressed the DEX-induced increase in REDD1 in a dose-dependent manner (p < 0.05). The WP group showed no significant changes in the mRNA expression of Cbl-b, while the FWP group exhibited a significant decrease at the concentration of 1000 μg/mL (p < 0.001). These results suggest that the potent anabolic effect of FWP on muscle tissue promotes protein synthesis and potentially muscle growth. Myogenesis, essential for skeletal muscle tissue development, involves myoblast fusion and differentiation into multinucleated myotubes, crucial for skeletal muscle formation and regeneration (Sampath et al., 2018). However, MyoD and myogenin expression may be dysregulated under conditions of DEX-induced muscle atrophy, potentially contributing to impaired muscle regeneration and maintenance. In the present study, the mRNA levels of MyoD and Myogenin were measured (Fig. 5D, E). DEX treatment reduced the MyoD and Myogenin mRNA levels. However, FWP showed a preventive effect on myogenic regulatory transcription from DEX treatment compared with that of WP treatment at the concentration of 1000 μg/mL (p < 0.05). These results suggest that FWP treatment prevented the suppression of myogenesis in DEX-induced myotubes.

Fig. 5.

Fig. 5

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Effect on muscle protein synthesis and myogenesis of whey fermented with Lacticaseibacillus rhamnosus IM36. C2C12 myotubes were treated with various concentrations (100, 500, and 1000 μg/mL) of unfermented whey (WP) or whey fermented with Lacticaseibacillus rhamnosus IM36 (FWP) for 48 h and then 100 μM dexamethasone for 24 h. The mRNA expression levels were quantified by quantitative reverse transcription polymerase chain reaction assay (qRT-PCR), and GAPDH was used as a housekeeping gene (A) insulin-like growth factor-1 (IGF-1), (B) regulated in development and DNA damage-response 1 (REDD1), (C) casitas B-lineage lymphoma proto-oncogene-b (Cbl-b), (D) myoblast determination protein 1 (MyoD), (E) Myogenin expression level. The data are presented as the mean ± standard errors of mean (SEM) with error bars indicating the standard error (n = 3). ***p < 0.001, **p < 0.005, *p < 0.05 represent significant differences between marked groups. NOR, normal group; DEX, dexamethasone only treated group

Effect of FWP on muscle protein degradation

DEX-induced muscle atrophy primarily involves the UPS, where key E3 ligases, such as Atrogin-1 and MuRF1, regulated by FOXO3a, significantly contribute to muscle degradation (Cohen et al., 2015). These E3 ligases control the breakdown of regulatory proteins, such as MyoD and myogenin, as well as structural proteins, such as myosin and troponin, which crucially impact muscle atrophy (Bodine and Baehr, 2014). FOXO3a also regulates ALS, which degrades cellular components essential for muscle fiber homeostasis by modulating genes, such as microtubule-associated protein light chain 3 (LC3), and responding to oxidative stress via p38 signaling (McGrath et al., 2021). Additionally, DEX treatment leads to intracellular ATP depletion and AMPK activation, thereby initiating the FOXO3a/Atrogenes pathway (Liu et al., 2016). To elucidate the preventive effect of FWP on muscle protein degradation in DEX-induced myotubes, various muscle-specific proteolytic factors, such as AMPK, FOXO3a, Atrogin-1, MuRF1, LC3, and Kruppel-Like Factor 15 (KLF15), were measured at the mRNA level (Fig. 6). DEX administration increased the mRNA expression of AMPK and FOXO3a; however, the FWP group showed dose-dependent increases in these levels (p < 0.05), restoring AMPK and FOXO3a mRNA levels to those in the normal group (Fig. 6A, B). As DEX administration increased FOXO3a levels, the mRNA expression levels of Atrogin-1, MuRF1, LC3, and KLF15 also increased (Fig. 6C–E). Increased KLF15 levels enhance FOXO expression and stimulate Atrogin-1 and MuRF1 gene expression, implying a cooperative regulatory role in the detrimental effect on muscle (Cid‐Díaz et al., 2021; Shimizu et al., 2011). The WP group showed no significant change in Atrogin-1 mRNA expression, whereas the FWP group showed a marked difference (p < 0.05; Fig. 6C). Moreover, the FWP group exhibited a significant dose-dependent reduction in the expression of MuRF1, LC3, and KLF15 (p < 0.05). Notably at the concentration of 1000 μg/mL, the FWP group showed significantly higher preventive effects on all muscle-specific proteolytic factors compared with those of the WP group (p < 0.05). The FWP group showed restoration of LC3 mRNA expression levels to those of the normal group. These results suggest that FWP treatment prevented muscle protein degradation in DEX-treated myotubes. These findings suggest that L. rhamnosus IM36 enhances the preventive effect of WP on the FOXO3a/Atrogenes pathway during fermentation. However, the mechanism of enhanced protective effect against muscle atrophy by metabolite produced by whey fermentation is unclear. Recent studies showed that fermented whey promoted more protein degradation compared to unfermented whey and result in enhanced bioactivity after digestion by gastro-intestinal digestion (Helal et al., 2023). Therefore, further studies are needed to investigate interaction between fermented whey peptide and muscle atrophy.

Fig. 6.

Fig. 6

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Effect on muscle protein degradation of whey fermented with Lacticaseibacillus rhamnosus IM36. C2C12 myotubes were treated with various concentrations (100, 500, and 1000 μg/mL) of unfermented whey (WP) or whey fermented with Lacticaseibacillus rhamnosus IM36 (FWP) for 48 h and then 100 μM dexamethasone for 24 h. The mRNA expression levels were quantified by quantitative reverse transcription polymerase chain reaction assay (qRT-PCR) and GAPDH was used as a housekeeping gene (A) AMP-activated protein kinase (AMPK), (B) forkhead box protein O3a (FOXO3a), (C) Atrogin-1, (D) muscle RING-finger protein-1 (MuRF1), (E) microtubule-associated protein light chain 3 (LC3), (F) Kruppel-Like Factor 15 (KLF15) expression level. The data are presented as the mean ± standard errors of mean (SEM) with error bars indicating the standard error (n = 3). ***p < 0.001, **p < 0.005, *p < 0.05 represent significant differences between marked groups. NOR, normal group; DEX, dexamethasone only treated group

In conclusion, whey enhanced radical-scavenging activity, reducing power, and ROS-lowering activity after fermentation with L. rhamnosus IM36. FWP with high antioxidant activity significantly promoted muscle protein synthesis and myogenesis and inhibited muscle protein degradation compared with that of normal WP. These findings highlight the potential of FWP as a preventive intervention for the management of sarcopenia and indicate a promising pathway for exploring its therapeutic implications in muscle-related disorders.

Supplementary Information

Below is the link to the electronic supplementary material.

Supplementary file1 (DOCX 12 KB) (13.3KB, docx)

Acknowledgements

This research was supported by Korea Institute of Planning and Evaluation for Technology in Food, Agriculture and Forestry (IPET) through High Value365 added Food Technology Development Program, funded by Ministry of Agriculture, Food and Rural Affairs (MAFRA) (321036-5).

Declarations

Conflict of interest

The authors do not declare any conflicts of interest.

Footnotes

Publisher's Note

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