Insights into the soybean protein isolate hydrolysates: Performance characterization, emulsion construction and in vitro digestive behavior.

In this experiment, the co-constructed O/W emulsions of different soy protein hydrolysates (SPHs) and gum arabic (GA) were investigated. SPHs were prepared by hydrolyzing soy protein isolate (SPI) using different enzymes, and investigated the effects of enzyme types and hydrolysis time on the physicochemical properties of SPHs. Moreover, SPI/GA and SPHs/GA were prepared and used as hydrophilic emulsifiers to construct O/W emulsions. The results showed that the optimal hydrolysis times for bromelain, pepsin and trypsin were 2 h (BSPH2), 3 h (PSPH3) and 3 h (TSPH3), respectively. Compared with SPI/GA emulsions, SPHs/GA emulsions had smaller particle size, more negative charge, higher interfacial adsorbed protein, and more stable emulsion systems. During the digestion process, SPHs/GA emulsions were effective in realizing the release of bioactives. In conclusion, enzymatic hydrolysis can be an effective modification technique, and SPHs/GA can be used as an effective emulsifier for the emulsion system.

Effects of Fermentation with Tetragenococcus halophilus and Zygosaccharomyces rouxii on the Volatile Profiles of Soybean Protein Hydrolysates.

The effects of fermentation with lactic acid bacteria (LAB) and yeast on the aroma of samples were analyzed in this work. The volatile features of different soybean hydrolysates were investigated using both GC-MS and GC-IMS. Only 47 volatile flavor compounds (VFCs) were detected when using GC-IMS, while a combination of GC-MS and GC-IMS resulted in the identification of 150 compounds. LAB-yeast fermentation could significantly increase the diversity and concentrations of VFCs (p < 0.05), including alcohols, acids, esters, and sulfurs, while reduce the contents of aldehydes and ketones. Hierarchical clustering and orthogonal partial least squares analyses confirmed the impact of fermentation on the VFCs of the hydrolysates. Seven compounds were identified as significant compounds distinguishing the aromas of different groups. The partial least squares regression analysis of the 25 key VFCs (ROAV > 1) and sensory results revealed that the treatment groups positively correlated with aromatic, caramel, sour, overall aroma, and most of the key VFCs. In summary, fermentation effectively reduced the fatty and bean-like flavors of soybean hydrolysates, enhancing the overall flavor quality, with sequential inoculation proving to be more effective than simultaneous inoculation. These findings provided a theoretical basis for improving and assessing the flavor of soybean protein hydrolysates.

Covalent modification of soy protein hydrolysates by EGCG: Improves the emulsifying and antioxidant properties.

In this study, the effect of EGCG conjugation on the emulsifying and antioxidant properties of SPHs was investigated to improve the functional characteristic of soy protein hydrolysates (SPHs) and develop a novel hydrolysates/peptides-EGCG conjugates. Enzymatic hydrolyzed SPHs (DH 5%, 8%, 10%) covalent with 1% EGCG to prepare conjugates at pH 9.0. The free amino group and tryptophan content of SPHs-EGCG conjugates significantly decreased, indicating the successful preparation of SPHs-EGCG conjugates. Additionally, 5% SPHs-EGCG conjugates showed the highest EGCG binding capacity. EGCG conjugation increased the particle sizes and charge of SPHs. Compared with non-covalent SPHs, the covalent modification of EGCG increased the emulsifying and antioxidant capacity, especially for 5% SPHs-EGCG, it exhibited much higher surface hydrophobicity, ESI (emulsifying stability index), EAI (emulsifying activity index), and antioxidant activity than others. This result revealed that SPHs and EGCG played a synergistic effect in improving the emulsifying and antioxidant capacity. Fluorescence spectroscopy analysis showed that the combination of EGCG conjugation significantly decreased the fluorescence intensity and caused maximum emission red-shift. The formation of a covalent bond between SPHs and EGCG was verified through Fourier transform infrared spectroscopy (FTIR), and the results also showed a significant increase in the α-helix and random coil contents of the conjugation, and a significant decrease in the ß-sheet and ß-turn contents. These results indicate that EGCG conjugation with SPHs induced the unfolding and stretching of protein flexibility. Overall, SPHs-EGCG conjugates can be applied as a promising emulsifier to fabricate emulsion systems and would be helpful in designing functional beverages containing polyphenols and peptides with enhanced functional nutritional properties.

Debittering effect of partially purified proteases from soybean seedlings on soybean protein isolate hydrolysate produced by alcalase.

To explore the potential application of proteases from soybean seedlings in the debittering of soybean protein hydrolysates, soybean seeds were germinated from 1 to 10 days. It was found that the sixth day seedlings exhibited highest proteases activity (130 U/g). After partial purification, the activity of proteases (PSP) from the sixth day seedlings further increased to 2675 U/g. In addition, PSP exhibited maximum activity at 50 â„ƒ and pH 5.5, and mainly comprised of two proteins with the molecular weight of 64.57 and 25.12 kDa respectively. PSP could decrease the bitterness score of the soybean protein isolate hydrolysate (SPIH) produced by Alcalase 2.4L from 3.45 to 0 in 3 h. Meanwhile, the degree of hydrolysis of SPIH slightly increased from 11.87% to 15.61% without reducing the antioxidant activity. This study may provide a solution to the contradiction between removing the bitterness of soybean protein hydrolysates and maintaining the bioactivity.

In vitro antioxidant activities of the novel pentapeptides Ser-His-Glu-Cys-Asn and Leu-Pro-Phe-Ala-Met and the relationship between activity and peptide secondary structure.

BACKGROUND: Using high-performance liquid chromatography/tandem mass spectrometry, two novel antioxidant pentapeptides [Ser-His-Glu-Cys-Asn (SHECN) and Leu-Pro-Phe-Ala-Met (LPFAM)] were identified from 1-3-kDa soybean protein hydrolysates (SPH). The MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay was used to evaluate cytotoxicity in HepG2 cells. Antioxidant activity was measured using in vitro assays, including the cellular antioxidant activity assay (CAA), 2,2-diphenyl-1-picrylhydrazyl or 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) inhibition, and oxygen radical absorbance capacity (ORAC) assays. Finally, the secondary structure was determined using circular dichroism (CD). RESULTS: The results revealed that two novel peptides were nontoxic and possessed antioxidant activity. SHECN had significantly higher antioxidant activity than LPFAM (P < 0.05). The CAA value of SHECN was 776.22 µmol QE 100 g-1 . SHECN also showed significant DPPH inhibition (70.18 ± 4.06%) and ABTS inhibition (88.16 ± 0.76%). It had normalized ORAC values of 0.3000 ± 0.0070 µmol GE mg-1 and 0.0900 ± 0.0020 µmol TE mg-1 , respectively. The results of the CD analysis demonstrated that, compared to LPFAM, which had much lower antioxidant activity, SHECN had a high ß-sheet content and reduced α-helix content. CONCLUSION: The results indicated that SHECN possessed high antioxidant activity. A higher ß-sheet content and lower content levels of α-helix appear to be correlated with antioxidant activity. © 2016 Society of Chemical Industry.

Rapid determination of oligopeptides and amino acids in soybean protein hydrolysates using high-resolution mass spectrometry.

INTRODUCTION: Soybean protein hydrolysates (SPHs), especially oligopeptides, have shown a variety of functional properties, including immunomodulatory and anti-oxidant effects. Soybean protein hydrolysate products have been used as functional ingredients in food, sports nutrition or clinical nutrition. However, the mixture is mostly undefined due to its complex nature, containing peptides and minor amino acids as well as small proteins. OBJECTIVES: To develop a specific and efficient method for the identification and structural characterisation of oligopeptides in SPHs, and to determine free amino acids in SPHs in the same analytical run, for evaluation of the chemical profile of SPH products. METHODS: Accurate mass spectrometry (MS) datasets of SPH samples were recorded on a high-performance liquid chromatography (HPLC) tandem high-resolution (HR) MS system. Potential oligopeptides were tentatively characterised based on their elemental compositions and ring double bond equivalent (RDBE) values, as well as HRMS/MS data. The analytical method to determine amino acids was evaluated in terms of linearity, precision, apparent recovery and limits of detection and quantitation. RESULTS: In total, 186 oligopeptides spanning the mass range of m/z 200-1500 and three major free amino acids could be determined in SPH samples in a single sample injection. Ninety-nine oligopeptides were tentatively characterised. The sensitive and specific instrumental performances also permitted the determination of 19 amino acids with a limit of quantitation of ≤ 0.1 µg/mL. CONCLUSION: The HPLC-HRMS technique has proven to be an advantageous tool for the rapid characterisation of oligopeptides and determination of amino acids in soybean protein hydrolysates.

Purification and characterization of calcium-binding soybean protein hydrolysates by Ca2+/Fe3+ immobilized metal affinity chromatography (IMAC).

Soybean protein hydrolysates (SPHs) can bind calcium in order to form soluble peptide-calcium complexes. However, amino acid composition and structural characteristics of the calcium chelating SPHs are still unclear. This study separated SPHs with calcium and iron immobilized metal affinity chromatography (IMAC), and examined the effects of SPHs with different amino acid composition on calcium binding capacity. Three fractions (FFe-1, FFe-2 and FFe-3) isolated with IMAC-Fe(3+) were shown possessing increased Glu, Gln, Lys and Pro content from FFe-1 to FFe-3, and improved amount of bound calcium. Furthermore, the fractions adsorbed on IMAC-Ca(2+) (Fe(3+)) were separated and identified with reverse-phase (RP)-HPLC and MALDI-TOF MS/MS. The results showed that the sequence of peptides from FCa-2 and FFe-3 fractions was DEGEQPRPFPFP.