Enzymatic Hydrolysis Optimization of Yak Whey Protein Concentrates and Bioactivity Evaluation of the Ultrafiltered Peptide Fractions.

Yak whey protein concentrates (YWPCs) have good functional properties, but there is still a gap in the study of their peptides. In this study, peptides were obtained by enzymatic hydrolysis, and the bioactivity of each ultrafiltration fraction was evaluated using an optimal process. YWPCs were isolated and purified from yak milk as the raw material. Alkaline protease, trypsin, and papain were used to hydrolyze YWPCs. The protease with the highest degree of hydrolysis (DH) and peptide concentration was selected as the most suitable enzyme. The effects of pH, temperature, time, and the enzyme-to-substrate ratio (E/S) on the DH and peptide concentration were investigated, and response surface methodology was utilized to optimize the hydrolysis process. The hydrolysate was separated using ultrafiltration membranes with molecular weight cut-offs of 10 kDa, 5 kDa, 3 kDa, and 1 kDa. The bioactivity of each ultrafiltration component was analyzed, including the inhibition rates of α-amylase and xanthine oxidase (XOD) activities and the scavenging rates of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) cation radicals. The results indicated that alkaline protease was the best enzyme for hydrolyzing YWPCs. The peptide concentration in the YWPC hydrolysate was the highest (17.21 mg/mL) at a pH of 8 and a concentration of 7500 U/g, after 2.5 h at 62 °C. The enzymatic hydrolysate was ultrafiltered to yield four peptide fractions, of which the <1 kDa peptides exhibited the highest α-amylase inhibitory activity (22.06%), XOD inhibitory activity (17.15%), and ABTS cationic free radical scavenging rate (69.55%). This demonstrates the potential of YWPC hydrolyzed peptides for hypoglycemic, uric acid-lowering, and antioxidant applications, providing a theoretical basis for the high-value utilization of YWPCs.

Effect of Hydrothermal Treatment on the Structure and Functional Properties of Quinoa Protein Isolate.

The aim of this study was to investigate the effects of hydrothermal treatment at different temperatures and times on the structure and functional properties of quinoa protein isolate (QPI). The structure of QPI was investigated by analyzing changes in the intrinsic fluorescence spectrum, ultra-violet (UV) spectrum, and Fourier transform infrared spectrum. The solubility, water/oil-holding capacity, emulsifying activity, and emulsion stability of QPI were studied, as were the particle size and the thermogravimetric properties of QPI. The results showed that the average particle size of QPI gradually increased with the increase in hydrothermal treatment time and temperature, and reached a maximum value of 121 °C for 30 min. The surface morphology also became rough and its thermal stability also increased. The endogenous fluorescence and UV spectral intensity at 280 nm decreased gradually with increasing hydrothermal treatment time and temperature, and reduced to the minimum values at 121 °C for 30 min, respectively. After hydrothermal treatment, the secondary structure of QPI tended to be disordered. The functional properties of QPI after treatment were all superior to those of the control. The results of this study might provide a basis for the processing and utilization of QPI.

Ultrasonic Assisted Extraction of Quinoa (Chenopodium quinoa Willd.) Protein and Effect of Heat Treatment on Its In Vitro Digestion Characteristics.

To extract and utilise the protein in quinoa efficiently, we investigated the effect of rate of quinoa protein isolate (QPI) extraction by ultrasound-assisted alkaline extraction and traditional alkaline extraction methods using single-factor experiments and Box-Behnken design. The effect of different heat treatment temperature and time on QPI functional properties and in vitro digestion characteristics were also investigated. The results showed that the optimal conditions of ultrasound- assisted alkaline extraction process were: ultrasonic time 99 min, solid-liquid ratio 1:20 w:v, ultrasonic temperature 47 °C, and pH 10, and its extraction rate and purity were 74.67 ± 1.08% and 87.17 ± 0.58%, respectively. It was 10.18% and 5.49% higher than that of the alkali-soluble acid precipitation method, respectively. The isoelectric point (pI) of QPI obtained by this method was 4.5. The flexibility and turbidity of QPI had maximum values at 90 °C, 30 min, and 121 °C, 30 min, which were 0.42 and 0.94, respectively. In addition, heat treatment changed the 1.77-2.79 ppm protein characteristic region in QPI's nuclear magnetic resonance hydrogen spectroscopy (1H NMR). After heating at 90 °C and 121 °C for 30 min, the hydrolysis degree and total amino acid content at the end of digestion (121 °C, 30 min) were significantly lower than those of untreated QPI by 20.64% and 27.85%. Our study provides basic data for the efficient extraction and utilisation of QPI.

Effects of Lactobacillus plantarum Fermentation on the Chemical Structure and Antioxidant Activity of Polysaccharides from Bulbs of Lanzhou Lily.

Recently, Lanzhou lily has attracted more attention because of its bioactive components specifically polysaccharides. We studied in vitro the effects of Lactobacillus plantarum fermentation on the physicochemical properties, chemical structure, and antioxidant activity of the Lanzhou lily polysaccharide. The results showed that compared with the unfermented Lanzhou lily polysaccharide (LP-W), the molecular weight (M w) of the fermented Lanzhou lily polysaccharide (LPF-W) decreased from 4334 to 1684 kDa, the particle size decreased from 300.8 ± 6.38 to 141.9 ± 4.96 nm, and the solubility increased from 72.33 ± 3.58 to 104.27 ± 2.91 mg/mL. In addition, after fermentation, the monosaccharide composition of LPF-W changed, and the alternation of mannose residues and glucose residues disappeared. The results of the analysis of the antioxidant activity in vitro showed that compared with LP-W, the fermented LPF-W had higher DPPH radical ability, superoxide anion radical scavenging ability, and reducing efficiency, but the hydroxyl radical scavenging ability decreased. These findings provide a reference for the potential application of the lily polysaccharide as a plant-derived antioxidant in functional foods.

A freeze-thawing method applied to the fabrication of 3-d curdlan/polyvinyl alcohol hydrogels as scaffolds for cell culture.

In this work, an innovative composite hydrogel composed of curdlan (CD)/polyvinyl alcohol (PVA) hydrogels with a 3-d network structure was successfully prepared by freeze-thaw processing. The presence of interactions, changes in crystallinity, and thermal behaviour were investigated by Fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), and thermogravimetry (TGA and DTG), respectively. The morphology of the hydrogels was investigated by scanning electron microscopy (SEM). With the increase of PVA concentration, the composite hydrogel had a greater mechanical strength while remaining remarkably ductile as evinced by tensile test results. PVA content affects the swelling and water retention of CD/PVA hydrogels. The results of CCK-8 assay showed that CD/PVA hydrogels have no cytotoxic effect on the mouse fibroblast L929 cells. The AO/EB double-staining experiment further proved that the cells in the composite hydrogels had good cytocompatibility. The porous biohydrogels developed in the present work can provide an ideal cell growth environment as a scaffold. CD/PVA hydrogels highlight the value of this system for cell adhesion and proliferation, and further soft tissue engineering application.

Isolation of antioxidant peptides from yak casein hydrolysate.

Enzymatic hydrolysis of protein is a principal method to obtain antioxidant peptides. A yak casein hydrolysate (YCH) was prepared by alcalase and trypsin digestion. An ultrafiltration membrane system was used to divide the hydrolysate into four molecular weight fractions; YCH-4 (<3 kDa) had the highest antioxidant activity. Fraction YCH-4 was separated into six subfractions by gel filtration chromatography; reverse-phase high performance liquid chromatography (RP-HPLC) was then used to partition sixteen antioxidant peptide subfractions. Liquid chromatography/electrospray tandem mass spectrometry (LC-ESI-MS/MS) was used to determine the amino acid sequence of a purified antioxidant peptide to be Arg-Glu-Leu-Glu-Glu-Leu (787.41 Da). Finally, a synthetic Arg-Glu-Leu-Glu-Glu-Leu peptide was evaluated for its superoxide anion and hydroxyl radical scavenging activity (IC50 = 0.52 and 0.69 mg mL-1), which confirmed the activity of the native purified peptide. Our results suggested that isolation and purification of antioxidant peptides from yak casein could be an important means to obtain natural antioxidant peptides.

Purification, structure and anti-oxidation of polysaccharides from the fruit of Nitraria tangutorum Bobr.

In this paper, polysaccharides were extracted from the fruits of Nitraria tangutorum Bobr. (NTWP) using a hot water extraction method and extraction conditions were optimized by RSM. The optimal conditions were determined as follows: extraction time 7 h, extraction temperature 60 °C, ratio of water to raw material 15 : 1, and with these conditions, the yield was 14.01 ± 0.11%. After purification using DEAE-cellulose column and Sephadex G-200 column, NTWP-II was successfully obtained. The results of GC-MS and SEC-LLS analysis suggested that monosaccharide composition of NTWP-II was composed of Rha, Ara, Man, Glc and Gal with the molar ratio of 1.14 : 2.5 : 3.00 : 2.69 : 5.28 and M w, M w/M n and R z 2.29 × 105, 1.32, 15.22. The detailed structure of NTWP-II was characterized by FT-IR, NMR. Based on these analyses, the structure of the repeating unit of NTWP-II was established.

Ultrasound-assisted extraction of polysaccharides from Rhododendron aganniphum: Antioxidant activity and rheological properties.

In this study, we aimed to optimize the extraction of polysaccharides from the leaves of Rhododendron aganniphum and investigate its rheological properties and antioxidant activity. After optimizing the operating parameters using a Box-Behnken design (BBD), the results showed that the optimal ultrasound-assisted extraction conditions were as follows: extraction temperature, 55°C; liquid-solid ratio, 25:1; extraction time, 2.2h; and ultrasound treatment power, 200W. The optimized experimental yield of polysaccharides by ultrasound-assisted extraction (PUAE) was 9.428%, higher than that obtained by hot water extraction (PHWE) for 12h at the same liquid-solid ratio and extraction temperature. In the in vitro antioxidant activity tests, PUAE had higher positive radical scavenging activity for hydroxyl, superoxide and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals than PHWE. However, PUAE and PHWE solutions had similar intermolecular interactions in the steady-shear flow and dynamic viscoelasticity tests, resulting in similar macroscopic behaviour. With respect to the apparent viscosity, storage modulus (G') and loss modulus (G″) of PUAE were lower at the same shear rate or angular frequency. All PUAE solutions exhibited non-Newtonian shear-thinning pseudoplastic behaviour that was accurately described by the Carreau model but was better fit by the power-law model at high shear rates (≥1/s), which demonstrated that the variation in the apparent viscosity dependence was greater at higher concentrations and shear rates. The G' and G″ of the solutions increased as the experimental frequency increased from 0.05 to 500rad/s under all experimental concentrations, and the modulus crossover point decreased gradually with increasing PUAE concentration. The above results demonstrated that the ultrasound-assisted extraction methods gave a higher yield of polysaccharides from the leaves of R. aganniphum with a shorter extraction time than the hot water extraction method, which could affect the apparent viscosity and dynamic viscoelasticity. PUAE presented good radical scavenging activity for DPPH, superoxide and hydroxyl radicals in vitro and could be used as a natural antioxidant in the food and medical industries.

Catalytic synthesis of sulfated polysaccharides. II: comparative studies of solution conformation and antioxidant activities.

Sulfated derivatives of Artemisia sphaerocephala polysaccharide (ASP) with high degree of substitution (DS) were synthesized using 4-dimethylaminopyridine (DMAP)/dimethylcyclohexylcarbodiimide (DCC) as catalyst. Size exclusion chromatography combined with multi-angle laser light scattering (SEC-LLS) results showed a decrease in fractal dimension (df) values of sulfated ASP (SASP). Compared to ASP and SASP with low DS (0.51-1.01), SASPcata2 exhibited an internal structure between rigid rod and random coil with a DS of 1.24. DS had greater influence on its conformation in aqueous solution. Circular dichroism (CD), methylene blue (MB) and Congo red (CR) spectrophotometric method and atomic force microscopy (AFM) results confirmed that the degradation of ASP and SO3H groups improved significantly the stiffness of the chains due to the electrostatic effect. Furthermore, antioxidant experiments revealed that high DS could enhance the scavenging activities of radicals and reducing power of SASP in vitro. The extended chain conformation was beneficial to enhance the biological activity of sulfated polysaccharides.

Regioselective synthesis of sulfated guar gum: comparative studies of structure and antioxidant activities.

We reported here a new synthesis of C-2 and C-3 sulfated guar gum (SRSGG) with low degree of substitution (DSS) of 0.58, employing triphenylchloromethane (TrCl) as a protected precursor. The yield and DSTr (calculated from the weight of triphenylmethanol) of triphenylmethylated GG (GGTr) was 165.6% and 0.71, respectively. In addition, low ratio (1:4) of chlorosulfuric acid to pyridine (1:4) was chosen in sulfation reaction since the protecting group was slightly sensitive to acid. Results of FT-IR and (13)C NMR spectroscopy indicated that C-2 and C-3 substitution was predominant but not fully sulfated in SRSGG. Size-exclusion chromatograph combined with multi-angle laser photometer (SEC-LLS) showed a decrease in molecular weight in the reaction. This might be due to the degradation in sulfation and deprotection process. Finally, we investigated the effect of structure features on the antioxidant activities in vitro. Vitro antioxidant experiments revealed that the regioselective sulfation at C-2 and C-3 and low molecular weight afforded strong antioxidant activities showing a much higher scavenging abilities of 1,1-diphenyl-2-picrylhydrazyl (DPPH) and hydroxyl radical than that by the known C-6-sulfated derivative. The antioxidant activities of sulfated polysaccharides were not a function of a single factor but a combination of molecular weight, DSS and substitution positions.

Selenylation modification can enhance antioxidant activity of Potentilla anserina L. polysaccharide.

Potentilla anserina L. polysaccharide had been selenizingly modified by nitric acid-selenious acid method with microwave-assistant. The optimal reaction parameters on selenylation modification were time 116 min, temperature 63°C, microwave power 190 W and the actual value of selenium content was 2690.1±7.2 µg/g. The scavenging ability of DPPH, OH and superoxide radicals, the metal chelating ability and reducing power in vitro were compared by antioxidant assay taking the non-modified PAP as a control. The results showed that selenylation modification could significantly enhance the antioxidant activity of SePAP and SePAP could be used as a potential antioxidant.

Synthesis of selenium-containing polysaccharides and evaluation of antioxidant activity in vitro.

Selenium (Se) is an essential trace element for human beings and many other forms of life. Organic Se from natural foods has higher bioavailability and is safer than inorganic Se species. In the present study, selenium-containing derivatives of Artemisia sphaerocephala polysaccharide with the highest Se content of 1703 µg/g were synthesized using H(2)SeO(3)/HNO(3) and BaCl(2) as a catalyst. The effects of the ratio of HNO(3) concentration and reaction temperature were investigated. Results of Raman and (13)C NMR spectroscopy indicated that C-6 substitution was predominant in selenized polysaccharide. A sharp decrease in M(w) was observed in the selenized reaction due to the acid environment. The in vitro antioxidant results suggest that selenium-containing polysaccharide should be applied as a novel selenium source in dietary supplements, with potent antioxidant properties.

Structure and antioxidant activities of sulfated guar gum: homogeneous reaction using DMAP/DCC catalyst.

It was essential to understand the chemical structure of polysaccharides for further research and biochemical or medical application of this natural biopolymer. In the present study, sulfated derivatives of guar gum with high degree of sulfation (DS) were synthesized using 4-dimethylaminopyridine (DMAP)/dimethylcyclohexylcarbodiimide (DCC) as catalyst in homogeneous conditions. The effects of the ratio of chlorosulfuric acid to pyridine, the content of catalyst and reaction temperature were investigated. Results of FT-IR, (1)H and (13)C NMR indicated that C-6 substitution was predominant in sulfated polysaccharide. In the sulfation reaction, a sharp decrease in M(W) was observed. The enhanced antioxidant activities of sulfated polysaccharides were not a function of a single factor but a combination of high DS and low molecule weight.

Structure-antioxidant relationships of sulfated galactomannan from guar gum.

Sulfated polysaccharides exerted potential biological property which was relative to degree of sulfation (DS), M(w), substitution position and chain conformation. In the present study, commercial guar gum was purified and its sulfated derivates with different DS and M(w) were synthesized. FT-IR and 13C NMR analysis indicated that C-6 substitution was predominant in sulfated samples compared with other positions. In the sulfation reaction, a sharp decrease in M(w) was observed. The d(f) values from 1.92 to 2.85 indicated that the -SO3H groups led to the relatively expanded conformation of sulfated polysaccharides. Antioxidant assays showed that sulfated polysaccharides had better antioxidant activities. The data obtained in in vitro models indicated that high DS and low M(w) showed the best antioxidant capacities.

A comparison study on microwave-assisted extraction of Artemisia sphaerocephala polysaccharides with conventional method: Molecule structure and antioxidant activities evaluation.

The conventional extraction methods for polysaccharides were time-consuming, laborious and energy-consuming. Microwave-assisted extraction (MAE) technique was employed for the extraction of Artemisia sphaerocephala polysaccharides (ASP), which is a traditional Chinese food. The extracting parameters were optimized by Box-Behnken design. In microwave heating process, a decrease in molecular weight (M(w)) was detected in SEC-LLS measurement. A d(f) value of 2.85 indicated ASP using MAE exhibited as a sphere conformation of branched clusters in aqueous solution. Furthermore, it showed stronger antioxidant activities compared with hot water extraction. The data obtained showed that the molecular weights played a more important role in antioxidant activities.