Potential Health Benefits Associated with Lunasin Concentration in Dietary Supplements and Lunasin-Enriched Soy Extract

Abstract

Lunasin has demonstrated antioxidative, anti-inflammatory, and chemopreventive properties. The objectives were to evaluate the concentration of lunasin in different lunasin-based commercial dietary supplements, to produce a lunasin-enriched soy extract (LESE) using a two-step pilot-plant-based ultrafiltration process, and to evaluate their biological potential in vitro. LESE was produced using 30 and 1 kDa membranes in a custom-made ultrafiltration skid. Lunasin was quantified in eight products and LESE. Lunasin concentrations of the lunasin-based products ranged from 9.2 ± 0.6 to 25.7 ± 1.1 mg lunasin/g protein. The LESE extract contained 58.2 mg lunasin/g protein, up to 6.3-fold higher lunasin enrichment than lunasin-based dietary supplements. Antioxidant capacity ranged from 121.5 mmol Trolox equivalents (TE)/g in Now^® Kids to 354.4 mmol TE/g in LESE. Histone acetyltransferase (HAT) inhibition ranged from 5.3% on Soy Sentials^® to 38.3% on synthetic lunasin. ORAC and lunasin concentrations were positively correlated, and HAT and lunasin concentrations were negatively correlated (p < 0.05). Melanoma B16-F10 and A375 cells treated with lunasin showed dose-dependent inhibitory potential (IC₅₀ equivalent to 330 and 370 μM lunasin, respectively). Lunasin showed protein kinase B expression (57 ± 14%) compared to the control (100%) in B16-F10. Lunasin concentration found in commercial products and lunasin-enriched soy extract could exert benefits to consumers.

Keywords: anti-melanoma activity; antioxidant potential; lunasin-enriched soy extract; lunasin-rich supplements; ultrafiltration.

Figure 1

Analysis of commercial lunasin-based dietary supplements. (a) SDS-PAGE protein electrophoretic profile and (b) Western blot analysis of different lunasin-based products. Lanes SL and MW are synthetic lunasin and molecular weight standard, respectively. Samples were used at a concentration of 100 µg protein/mL and coded as 1: Soy Sentials^®, 2: Slimplicity^®, 3: Now^® dietary supplement, 4: Provantage^®, 5: LunaRich X™, 6: GlucAffect^®, 7: Now^® kids-vanilla and 8: Now^® kids-chocolate.

Figure 2

Soluble protein and lunasin analysis of different fractions obtained from two-step ultrafiltration pilot plant production of lunasin-enriched soy extract. (a) Soluble protein concentration as measured by protein DC assay, (b) Lunasin concentration as measured by ELISA. Different letters indicate significant differences among fractions (p < 0.05).

Figure 3

Protein profile and lunasin identification in pilot plant enrich fractions. (a) Protein electrophoretic profile and (b) Western blot analysis of different fractions from pilot plant production of lunasin-enriched soy extract. Lanes MW: molecular weight standard, 1: defatted soy flour:water mixture (1:10), 2: supernatant after centrifugation, 3: 30 kDa retentate, 4: 30 kDa permeate, 5: 1 kDa retentate, and 6,7: synthetic lunasin (10 µM).

Figure 4

Biological potential of lunasin-based dietary supplements, lunasin-enriched soy extract, and synthetic lunasin. (a) Antioxidant capacity (mmol TE/g of product), (b) Histone acetyltransferase HAT (%) inhibitory potential. Samples were used at a concentration of 100 µg protein/mL and coded as 1: Soy Sentials^®, 2: Slimplicity^®, 3: Now^® dietary supplement, 4: Provantage^®, 5: LunaRich X™, 6: GlucAffect^®, 7: Now^® kids-vanilla and 8: Now^® kids-chocolate, 9: Freeze-dried lunasin enriched soy extract, and 10: Synthetic lunasin at 10 µM (n = 3). Different letters indicate significant differences among products (p < 0.05). Gallic acid (0.1 mg/mL) had an ORAC value of 220.3 ± 2.7 mmol TE/g, used for comparison purposes.

Figure 5

Lunasin content correlations with antioxidant capacity and HAT inhibitory potential. (a) Lunasin (mg/g product) positively correlated with ORAC values and (b) Lunasin (mg/g protein) negatively correlated with HAT activity. Samples were used at a concentration of 100 µg protein/mL; materials included: Soy Sentials^®, Slimplicity^®, Now^® dietary supplement, Provantage^®, Lunarich X™, GlucAffect^®, Now^® kids-vanilla and Now^® kids-chocolate (n = 3). Gallic acid (0.1 mg/mL) had an ORAC value of 220.3 ± 2.7 mmol TE/g, used for comparison purposes.

Figure 6

Melanoma cells treated with lunasin extract: (a) Cell viability: B16-F10 cell lines treated with different concentrations (0.158 ng/mL to 3.16 mg/mL). (b) Phosphorylated AKT pathway expression on cells B16-F10 after treatment with lunasin extract IC₅₀ equivalent to 330 μM. (c) Phosphorylated AKT pathway expression on cells A-375 after treatment with lunasin extract IC50 equivalent to 370 μM. p values reflect the comparison of the control and the treated for each marker * p < 0.05, ** p < 0.005.