Structural Characterization and Antioxidant Activity of ß-Glucans from Highland Barley Obtained with Ultrasonic-Microwave-Assisted Extraction.

In order to efficiently extract ß-glucan from highland barley (HBG) and study its structural characterization and antioxidant activity, ultrasonic-microwave-assisted extraction (UME) was optimized by the response surface method (RSM). Under the optimal extraction conditions of 25.05 mL/g liquid-solid ratio, 20 min ultrasonic time, and 480 W microwave intensity, the DPPH radical scavenging activity of HBG reached 25.67%. Two polysaccharide fractions were purified from HBG, namely HBG-1 and HBG-2. Structural characterization indicated that HBG-1 and HBG-2 had similar functional groups, glycosidic linkages, and linear and complex chain conformation. HBG-1 was mainly composed of glucose (98.97%), while HBG-2 primarily consisted of arabinose (38.23%), galactose (22.01%), and xylose (31.60%). The molecular weight of HBG-1 was much smaller than that of HBG-2. Both HBG-1 and HBG-2 exhibited concentration-dependent antioxidant activity, and HBG-1 was more active. This study provided insights into the efficient extraction of HBG and further investigated the structure and antioxidant activities of purified components HBG-1 and HBG-2. Meanwhile, the results of this study imply that HBG has the potential to be an antioxidant in foods and cosmetics.

Effects of liposoluble components of highland barley spent grains on physiological indexes, intestinal microorganisms, and the liver transcriptome in mice fed a high-fat diet.

The purpose of this study was to investigate the effects of the active ingredients of barley lees on the physiological indexes, intestinal flora, and liver transcriptome of mice fed a high-fat diet. Twenty-four male C57BL/6J mice were randomly divided into 4 groups and fed the experimental diets for 5 weeks. The results showed that the fat-soluble components of distillers' grains significantly reduced body weight, abdominal fat, perirenal fat, blood glucose, low-density lipoprotein cholesterol, triglycerides, and total cholesterol in the high-fat diet-fed mice (p < .05), significantly decreased alanine aminotransferase and malondialdehyde levels, and significantly increased total superoxide dismutase, catalase, reduced glutathione and glutathione peroxidase levels (p < .05). At the phylum level, lipid-soluble components significantly increased the abundance of Bacteroidetes and decreased the Firmicutes/Bacteroidetes ratio. At the genus level, the relative abundances of Bacteroidetes and Clostridium were increased. Transcriptomic analysis showed that lipid-soluble components of spent grains reduced the mRNA expression of ANGPTL8, CD36, PLTP, and SOAT1 and increased the mRNA expression of CYP7A1 and ABCA1 in the cholesterol metabolism pathway, promoted the transport of cholesterol, and inhibited the absorption of cholesterol, which can decrease cholesterol levels by speeding up the conversion of cholesterol into bile acids.

Molecular Organization and Functional Analysis of a Novel Plasmid-Borne cps Gene Cluster from Lactiplantibacillus plantarum YC41.

Capsular polysaccharide (CPS) can tightly attach to bacterial surfaces and plays a critical role in protecting microorganisms from environmental stresses. However, the molecular and functional properties of some plasmid-borne cps gene clusters are poorly understood. In this study, comparative genomics of the draft genomes of 21 Lactiplantibacillus plantarum strains revealed that the specific gene cluster for CPS biosynthesis was observed only in the 8 strains with a ropy phenotype. Furthermore, the complete genomes showed that the specific gene cluster cpsYC41 was located on the novel plasmid pYC41 in L. plantarum YC41. In silico analysis confirmed that the cpsYC41 gene cluster contained the dTDP-rhamnose precursor biosynthesis operon, the repeating-unit biosynthesis operon, and the wzx gene. The insertional inactivation of the rmlA and cpsC genes abolished the ropy phenotype and reduced the CPS yields by 93.79% and 96.62%, respectively, in L. plantarum YC41 mutants. These results revealed that the cpsYC41 gene cluster was responsible for CPS biosynthesis. Moreover, the survival rates of the YC41-rmlA- and YC41-cpsC- mutants under acid, NaCl, and H2O2 stresses were decreased by 56.47 to 93.67% compared to that of the control strain. Furthermore, the specific cps gene cluster was also confirmed to play a vital role in CPS biosynthesis in L. plantarum MC2, PG1, and YD2. These findings enhance our understanding of the genetic organization and gene functions of plasmid-borne cps gene clusters in L. plantarum. IMPORTANCE Capsular polysaccharide is well known to protect bacteria against various environmental stresses. The gene cluster for CPS biosynthesis is typically organized in the chromosome in bacteria. It is worth noting that complete genome sequencing showed that a novel plasmid pYC41-borne cpsYC41 gene cluster was identified in L. plantarum YC41. The cpsYC41 gene cluster included the dTDP-rhamnose precursor biosynthesis operon, the repeating-unit biosynthesis operon, and the wzx gene, which was verified by the significantly decreased CPS yield and the absent ropy phenotype in the corresponding mutants. The cpsYC41 gene cluster plays an important role in bacterial survival under environmental stress, and the mutants had decreased fitness under stress conditions. The vital role of this specific cps gene cluster in CPS biosynthesis was also confirmed in other CPS-producing L. plantarum strains. These results advanced a better understanding of the molecular mechanisms of plasmid-borne cps gene clusters and the protective functionality of CPS.

Identification of peptides in Qingke baijiu and evaluation of its angiotensin converting enzyme (ACE) inhibitory activity and stability.

The active peptides in Qingke baijiu fermented from Qingke (highland barley) are rarely reported. This work was designed to accurately identify peptides in Qingke baijiu and evaluate their angiotensin-converting enzyme inhibitory activities in vitro. Four novel peptides, Val-Val-Thr-Gly-Val-Gly-Gly-Gln (VVTGVGGQ), Leu-Pro-Val-Gly-Pro (LPVGP), Leu-Leu-Ser-Pro-Pro (LLSPP), and Phe-Pro-Leu-Gln-Pro-His-Gln-Pro (FPLQPHQP) were identified by Nano-UPLC-MS/MS. Molecular docking showed that LPVGP and FPLQPHQP had a high affinity with ACE (binding energy -8.78, -10.02 kcal mol-1), which matched its in vitro ACE inhibitory activity (IC50 9.05, 5.03 µM). This might be related to their high hydrophobicity. Moreover, three peptides have C-terminal proline, which may contribute to their anti-digestive activity. The content of LPVGP was over 91.23% after digestion, while the content of VVTGVGGQ dropped to 55.47%. Finally, the four peptides have no obvious toxicity to Caco-2 cells. This study clarifies the ACE inhibitory activity and the structure-activity relationship of the four peptides identified in Qingke baijiu.

Raw Material Regulates Flavor Formation via Driving Microbiota in Chinese Liquor Fermentation.

Raw material is important for flavors in fermented foods. Here, the effect of hulless barley on the microbiota in Chinese liquor was studied using two main cultivars (heilaoya and dulihuang). Six genera (Lactobacillus, Saccharomyces, Komagataella, Aspergillus, Pichia, and Weissella) were identified as flavor producers. Komagataella, mainly correlated with esters, dominated in heilaoya, and Pichia, mainly correlated with carbonyls, dominated in dulihuang. The Mantel test indicated reducing sugar drove the succession of microbiota (heilaoya: P = 0.001; dulihuang: P = 0.006). Especially, glucose (P = 0.0226) and fructose (P = 0.0168) presented the most significant correlations with Pichia and Komagataella, respectively. The simulative fermentation confirmed Komagataella phaffii QK2 grew better in heilaoya with more fructose, whereas Pichia fermentans PF grew better in dulihuang with more glucose. This work highlighted the effect of raw material on microbiota, which would be beneficial for regulating the quality of fermented foods.

Hormesis of glyceollin I, an induced phytoalexin from soybean, on budding yeast chronological lifespan extension.

Glyceollin I, an induced phytoalexin isolated from soybean, has been reported to have various bioactivities, including anti-bacterial, anti-nematode, anti-fungal, anti-estrogenic and anti-cancer, anti-oxidant, anti-inflammatory, insulin sensitivity enhancing, and attenuation of vascular contractions. Here we show that glyceollin I has hormesis and extends yeast life span at low (nM) doses in a calorie restriction (CR)-dependent manner, while it reduces life span and inhibits yeast cell proliferation at higher (µM) doses. In contrast, the other two isomers (glyceollin II and III) cannot extend yeast life span and only show life span reduction and antiproliferation at higher doses. Our results in anti-aging activity indicate that glyceollin I might be a promising calorie restriction mimetic candidate, and the high content of glyceollins could improve the bioactivity of soybean as functional food ingredients.

Secondary metabolites in durian seeds: oligomeric proanthocyanidins.

Ethanolic extract of durian seeds was fractionated by reverse phase flash column chromatography and the fractions characterized by electrospray ionization mass spectroscopy. Among a few unknown compounds collected, oligomeric proanthocyanidins (OPCs) were found to be one of the main compounds. Based on this result, the OPCs were purified the first time from the durian seeds using standard procedures and gave a yield of 1.8 mg/g dry matter after fractionation by Sephadex LH-20 column. Structural analysis by ¹³C{¹H} NMR and ESI-MS spectra showed the presence of primarily B-type procyanidins with mainly epicatechin as the extension units, which was further verified by matrix assisted laser desorption/ionization-time of flight mass spectra (MALDI-TOF MS), which shows a distribution of dimers to decamers. In addition, hydroxylated peaks with molecular weight 16 units more than the poly-epicatechins represented significant peaks. We suggest this might be due to hydroxylation occurring under the MALDI-TOF MS conditions. Consistently, depolymerization with α-toluenethiol resulted in epicatechin thioether as the major product, but undetectable amount of gallocatechin or its α-toluenethiol derivatives. The oligomershave a mean degree of polymerization of 7.30.

Hypoglycemic activities of commonly-used traditional Chinese herbs.

The α-amylase and α-glucosidase inhibition activity of 92 Traditional Chinese Medicinal (TCM) herbs, which are permitted to be used as food ingredients, were evaluated using the high throughput assay developed in our laboratory. Among these herbs, twenty-seven of them possessed significant α-amylase inhibition activities ranging from 2.4 to 349.2 µmol AE/g (AE = acarbose equivalent) with inhibition concentrations at 50% inhibition (IC50) from 16.0 to 2342.2 µg/mL, respectively. In addition, they showed α-glucosidase inhibition activities ranging from 0.5 to 31.6 µmolAE/g (IC50 from 49.0 to 3385.5 µg/mL). The extracts of Rhizoma fagopyri dibotryis (Jinqiáomài), Rosa rugosa (Méiguihua), Caulis polygoni multiflori (Shǒuwuténg), Fructus amomi (Sharén), Rhizoma alpiniae officinarum (Gaoliángjiang), Folium ginkgo (Yínxìngyè) and Cortex cinnamomi (Ròuguì) showed the better inhibitory activities against both α-amylase and α-glucosidase. Our results illustrated that these food grade herbs are potent natural hypoglycemic agents and can be used as active ingredients for low glycemic index food production or TCM herbal formulations for controlling hyperglycemia.

Characterization of proanthocyanidins in stems of Polygonum multiflorum Thunb as strong starch hydrolase inhibitors.

Characterization of polyphenolic compounds in the stems of P. multiflorum was conducted using HPLC, high resolution LC-MS and LC-MSn. Proanthocyanidins in particular were isolated in 4.8% yield using solvent extraction followed by Sephadex LH-20 fractionation. HPLC analysis using a diol column revealed oligomers (from dimer to nonamer) as minor components, with (epi)catechin monomeric units predominating, and oligomers with higher degree of polymerization being dominant. Thiolysis treatment of the proanthocyanidins using mercaptoacetic acid produced thioether derivatives of (epi)catechin as the major product and a mean value of the degree of polymerization of 32.6 was estimated from the ratio of terminal and extension units of the (epi)catechin. The isolated proanthocyanidins were shown to strongly inhibit α-amylase with an acarbose equivalence (AE) value of 1,954.7 µmol AE/g and inhibit α-glucosidase with an AE value of 211.1 µmol AE/g.

Germination dramatically increases isoflavonoid content and diversity in chickpea (Cicer arietinum L.) seeds.

The effect of germination on bioactive components in legume seeds was investigated in terms of the antioxidant capacity and total phenolic contents. Germination increased the total phenolic content and antioxidant capacity of most seeds. Particularly in chickpea seeds, the isoflavone contents increased by over 100 fold, mainly due to the increase of formononetin and biochanin A level. As a result, these two compounds were conveniently isolated from the germinated seeds in preparative scale and structurally confirmed by UV-vis, ESI-MS, and (1)H NMR spectroscopies. Isoflavonoid fingerprints analyzed by HPLC-PDA and LC-ESI-MS demonstrated that germination could significantly increase isoflavonoids diversity. Twenty-five isoflavonoids were detected and identified tentatively. These include 20 isoflavones, 2 isoflavanones, and 3 pterocarpan phytoalexins. Total isoflavonoid content of germinated chickpea was approximately 5-fold of that of germinated soybean. Our findings suggest that the germinated chickpea seeds could serve as a promising functional food rich in isoflavonoids.

The effects of fungal stress on the antioxidant contents of black soybeans under germination.

Black soybeans were germinated with or without Rhizopus oligosporus for 3 days. The samples collected from each day were freeze-dried and extracted with chloroform-methanol-water mixture to simultaneously obtain organic phase (lipophilic extract) and aqueous phase (hydrophilic extract). In the lipophilic extract, α-, γ-, and δ-tocopherol were drastically decreased by 51%, 97%, and 88%, respectively, after 3 day germination under stress. On the other hand, the lipid peroxide concentration was increased in the fungi stressed, germinating beans from day 2 onward, while it was continually decreased in the nonstressed germinating counterpart. The change of antioxidant capacity (ORACoil value) was correlated to the changes of tocopherols and lipid peroxide concentrations. In nonstressed germinatng beans, ORACoil value dropped by 38% after 3 days, while in the fungi-stressed germinating beans, ORACoil was decreased by 80%. In hydrophilic extract, the antioxidant activity, measured by ORAChydro assay, is about 80 times higher than the ORACoil values, but there is no significant change before and after germination (either with or without stress). The total phenolic content in the hydrophilic extract increased only slightly by 9% in germinated beans and 15% in fungus-stress germinated samples, respectively. The advantage of stress germination of soybeans in enriching phytoalexins is traded off by loss of tocopherols.

Novel process of fermenting black soybean [Glycine max (L.) Merrill] yogurt with dramatically reduced flatulence-causing oligosaccharides but enriched soy phytoalexins.

Black soybeans [Glycine max (L.) Merrill] were germinated under fungal stress with food grade R. oligosporus for 3 days and were homogenized and fermented with lactic acid bacteria (LAB) to produce soy yogurt. Fungal stress led to the generation of oxylipins [oxooctadecadienoic acids (KODES) isomers and their respective glyceryl esters] and glyceollins--a type of phytoalexins unique to soybeans. In soy yogurt, the concentrations of total KODES and total glyceollins were 0.678 mg/g (dry matter) and 0.953 mg/g, respectively. The concentrations of other isoflavones (mainly genistein and daidzein and their derivatives) in soy yogurt remained largely unchanged after the processes compared with the control soy yogurt. Germination of black soybean under fungal stress for 3 days was sufficient to reduce stachyose and raffinose (which cause flatulence) by 92 and 80%, respectively. With a pH value of 4.42, a lactic acid content of 0.262%, and a maximum viable cell count of 2.1 x 10 (8) CFU/mL in the final soy yogurt, soy milk from germinated soybeans under fungal stress was concluded to be a suitable medium for yogurt-making. The resulting soy yogurt had significantly altered micronutrient profiles with significantly reduced oligosaccharides and enriched glyceollins.

Fungal-stressed germination of black soybeans leads to generation of oxooctadecadienoic acids in addition to glyceollins.

Microbial-stressed germination of black soybeans leads to generation of a group of oxylipins, oxooctadecadienoic acids (KODEs, including 13- Z, E-KODE, 13- E, E-KODE, 9- E, Z-KODE, and 9- E, E-KODE), and their respective glyceryl esters in addition to glyceollins, a known phytoalexins present in wild and fungi-infected soybeans. Four fungi, Aspergillus niger, Aspergillus oryzae, Rhizopus oligosporus, and white rice yeast ( Aspergillus niger wry), were applied to compare their efficiency on inducing these compounds during black soybean germination. Overall, R. oligosporus, the starter culture used in tempeh fermentation, gives the highest amounts of KODEs and glyceollins. The glyceollins and KODEs were isolated by preparative HPLC, and the structures were determined by (1)H NMR, UV-Vis, and MS spectra. On the basis of the unequal distribution of the KODEs isomers, an enzymatic reaction, instead of a nonenzymatic free radical chain reaction, is responsible for their formations. Together with other oxylipins and glyceollins, the KODEs may contribute to the soybean's defensive response to fungal infection via reaction with protein thiol groups and cell membranes. The stress-germinated black soybeans may be used as ingredients for further processing of novel functional food products with unique nutritional and flavor profiles.

Red grapefruit positively influences serum triglyceride level in patients suffering from coronary atherosclerosis: studies in vitro and in humans.

The contents of the bioactive compounds in red and blond grapefruits and their influence on humans suffering from hypertriglyceridemia were studied. It was found that red grapefruit has a higher content of bioactive compounds and a higher antioxidant potential than blond grapefruit, determined by oxygen radical scavenging capacity, 1,1-diphenyl-2-picrylhydrazyl, carotenoid bleaching, and Folin-Ciocalteu assays. Fifty-seven hyperlipidemic patients, ages 39-72 years, after coronary bypass surgery, recruited from the Institute's pool of volunteers, were randomly divided into three equal in number (19) groups: two experimental (red and blond groups) and one control group (CG). During 30 consecutive days of the investigation the diets of the patients of the red and blond dietary groups were daily supplemented with one equal in weight fresh red or blond grapefruit, respectively. Before and after this trial, serum lipid levels of all fractions and serum antioxidant activity were determined. It was found that serum lipid levels in patients of the red and blond groups versus the CG after treatment were decreased: (a) total cholesterol, 6.69 versus 7.92 mmol/L, 15.5%, and 7.32 versus 7.92 mmol/L, 7.6%, respectively; (b) low-density lipoprotein cholesterol, 5.01 versus 6.29 mmol/L, 20.3%, and 5.62 versus 6.29 mmol/L, 10.7%, respectively; (c) triglycerides, 1.69 versus 2.32 mmol/L, 17.2%, and 2.19 versus 2.32 mmol/L, 5.6%, respectively. No changes in the serum lipid levels in patients of the CG were found. In conclusion, fresh red grapefruit contains higher quantities of bioactive compounds and has significantly higher antioxidant potential than blond grapefruit. Diet supplemented with fresh red grapefruit positively influences serum lipid levels of all fractions, especially serum triglycerides and also serum antioxidant activity. The addition of fresh red grapefruit to generally accepted diets could be beneficial for hyperlipidemic, especially hypertriglyceridemic, patients suffering from coronary atherosclerosis.