The Radical Scavenging Activities and Anti-Wrinkle Effects of Soymilk Fractions Fermented with Lacticaseibacillus paracasei MK1 and Their Derived Peptides

Abstract

Soybean-derived peptides exert several beneficial effects in various experimental models. However, only a few studies have focused on the radical scavenging and anti-wrinkle effects of soymilk-derived peptides produced via different processes, such as fermentation, enzymatic treatment, and ultrafiltration. Therefore, in this study, we investigated the radical scavenging and antiwrinkle effects of soymilk fractions produced using these processes. We found that 50SFMKUF5, a 5 kDa ultrafiltration fraction fermented with Lacticaseibacillus paracasei MK1 after flavourzyme treatment, exhibited the highest radical scavenging activity using the 2,2-diphenyl-1-picrylhydrazyl radical scavenging assay as well as potent anti-wrinkle effects assessed by type 1 procollagen production and tumor necrosis factor-α production in ultraviolet B (UVB)-treated human dermal fibroblasts and HaCaT keratinocytes. To identify potential bioactive peptides, candidate peptides were synthesized, and their anti-wrinkle effects were assessed. APEFLKEAFGVN (APE), palmitoyl-APE, and QIVTVEGGLSVISPK peptides were synthesized and used to treat UVB-irradiated fibroblasts, HaCaT keratinocytes, and α-melanocyte-stimulating hormone-induced B16F1 melanoma cells. Among these peptides, Pal-APE exerted the strongest effect. Our results highlight the potential of soymilk peptides as anti-aging substances.

Keywords: anti-aging; anti-wrinkle; fermentation; lactic acid bacteria; peptide; radical scavenging activity; soybean milk.

Figure 1

2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activities of various soymilk fractions. (a) 50S, 50SMK, 50SMKUF30, and 50SMKUF5. (b) 50SFMK, 50SFMKUF30, and 50SFMKUF5.

Figure 2

Effects of various soymilk fractions on the viability of human dermal fibroblasts. (a) 100SPMK, 100SPMKUF30, and 100SPMKUF5. (b) 50SFMK, 50SFMKUF30, and 50SFMKUF5.

Figure 3

Cytotoxicities of various soymilk fractions in HaCaT keratinocytes. (a) 50SMKP, 50SMKPUF30, and 50SMKPUF5. (b) 50SFMK, 50SFMKUF30, and 50SFMKUF5.

Figure 4

Production of type 1 procollagen in ultraviolet B (UVB)-irradiated human dermal fibroblasts treated with (a) 50S and (b) 50SFMKUF5. The mean ± standard deviation (SD) values of three independent experiments were analyzed using a two-way analysis of variance (ANOVA) and Dunnett’s test (* p < 0.05 and ** p < 0.01).

Figure 5

Inhibition of tumor necrosis factor (TNF)-α production in UVB-irradiated HaCaT keratinocytes treated with (a) 50S and (b) 50SFMKUF5. The mean ± SD values of three independent experiments were analyzed using two-way ANOVA and Dunnett’s test (* p < 0.05).

Figure 6

Reducing sugar contents of various soymilk fractions. The mean ± SD values of three independent experiments were analyzed using a t-test (** p < 0.01 and *** p < 0.001).

Figure 7

Liquid chromatography–tandem mass spectrometry (LC-MS/MS) chromatograms of 50SFMKUF5. (a) APEFLKEAFGVN and (b) QIVTVEGGLSVISPK.

Figure 8

Production of type 1 procollagen based on the peptide concentration in UVB-irradiated fibroblast cells. Cells were treated with (a) 50SFMKUF5, (b) QIVTVEGGLSVISPK, (c) APEFLKEAFGVN, (d) palmitoyl-APEFLKEAFGVN. The mean ± SD values of three independent experiments were analyzed using two-way ANOVA and Dunnett’s test (* p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001).

Figure 9

TNF-α inhibition according to the peptide concentration in UVB-irradiated HaCaT keratinocytes. Cells were treated with (a) 50SFMKUF5, (b) QIVTVEGGLSVISPK, (c) APEFLKEAFGVN, and (d) palmitoyl-APEFLKEAFGVN. The mean ± SD values of three independent experiments were analyzed using two-way ANOVA and Dunnett’s test (* p < 0.05 and ** p < 0.01).

Figure 10

Melanin inhibition according to the peptide concentration in α-melanocyte-stimulating hormone (α-MSH)-treated B16F1 melanoma cells. Cells were treated with (a) 50SFMKUF5, (b) QIVTVEGGLSVISPK, (c) APEFLKEAFGVN, and (d) palmitoyl-APEFLKEAFGVN. The mean ± SD values of three independent experiments were analyzed using two-way ANOVA and Dunnett’s test (* p < 0.05 and **** p < 0.0001).