Phytochemical properties and functional characteristics of wild turmeric (Curcuma aromatica) fermented with Rhizopus oligosporus.

This study investigated the effect of solid-state fermentation of wild turmeric (Curcuma aromatica) with Rhizopus oligosporus, a common fungus found in fermented soy tempeh, on phytochemical and biological properties. Ultra-performance liquid chromatography-tandem mass spectrometry showed that fermented wild turmeric has higher concentrations of curcumin, demethoxycurcumin, bisdemethoxycurcumin, phenolic compounds and total flavonoid-curcuminoid after fermentation for 1-, 3-, and 5-day relative to non-fermented turmeric. The l-carnitine content reached 242 µg g-1 extract after fermentation for 7-day. Wild turmeric had 1.47- and 2.25-fold increases in ORAC and FRAP, respectively, after 3-day fermentation. The inhibitory effects of fermented wild turmeric on lipid accumulation from 3T3-L1 cells, nitric oxide production from lipopolysaccharide-stimulated RAW264.7 murine macrophages, and melanin formation by B16F10 mouse melanoma cells with α-MSH increased 1.08-, 1.44-, and 1.52-fold, respectively, after 3-day fermentation. Based on these results, fermented wild turmeric product can be used as a functional ingredient in the cosmeceutical and nutraceutical industries.

The bifidogenic effects and dental plaque deformation of non-digestible isomaltooligosaccharides synthesized by dextransucrase and alternansucrase.

Non-digestible isomaltooligosaccharides (NDIMOS) are functional food and beverage ingredients that contributed to human health benefits through metabolism of gastrointestinal microorganism. In this study, NDIMOS were synthesized by combine dextransucrase from Leuconostoc mesenteroides B512F/KM and alternansucrase from L. mesenteroides NRRL 1355CF10/KM using sucrose as substrate and maltose as acceptor. Their digestibility was confirmed by using digestive enzymes including α-amylase and amyloglucosidase. NDIMOS inhibited insoluble glucan formation through mutansucrase from Streptococcus mutans. The bifidogenic effect of NDIMOS was investigated by growth of four strains of Bifidobacterium in MRS broth containing NDIMOS, compared with MRS broth contain glucose and negative control. Additionally, Bifidobacterium bifidum or Bifidobacterium adolescentis inhibited the growth of Salmonella enterica serovar typhimurium when they were co-cultivation in MRS broth containing NDIMOS. These results suggested that NDIMOS is potential functional ingredient for food, beverage, and pharmaceutical application.

Brewing of glucuronic acid-enriched apple cider with enhanced antioxidant activities through the co-fermentation of yeast (Saccharomyces cerevisiae and Pichia kudriavzevii) and bacteria (Lactobacillus plantarum).

Co-fermentation using yeast (Saccharomyces cerevisiae and Pichia kudriavzevii) and the bacteria (Lactobacillus plantarum) as starters isolated from spontaneous sourdough was conducted for the brewing of glucuronic acid (GlcA)-enriched apple cider. The concentration of GlcA in the apple cider co-fermented for 14 d with commercial S. cerevisiae and L. plantarum was 37.7 ± 1.7 mg/mL while a concentration of 62.8 ± 3.1 mg/mL was recorded for fermentation with P. kudriavzevii and L. plantarum, which was higher than the corresponding single yeast fermentation. The co-fermented apple cider revealed higher 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity of 171.67 ± 0.79 µg trolox equivalents (TE)/mL using P. kudriavzevii and L. plantarum, compared to the control (143.89 ± 7.07 µg TE/mL) just using S. cerevisiae. Thus, the co-fermentation of S. cerevisiae and L. plantarum and P. kudriavzevii and L. plantarum provided a new strategy for the development of GlcA-enriched apple cider with enhanced antioxidant capacity.

The Inhibitory Effects of Plant Derivate Polyphenols on the Main Protease of SARS Coronavirus 2 and Their Structure-Activity Relationship.

The main protease (Mpro) is a major protease having an important role in viral replication of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the novel coronavirus that caused the pandemic of 2020. Here, active Mpro was obtained as a 34.5 kDa protein by overexpression in E. coli BL21 (DE3). The optimal pH and temperature of Mpro were 7.5 and 37 °C, respectively. Mpro displayed a Km value of 16 µM with Dabcyl-KTSAVLQ↓SGFRKME-Edans. Black garlic extract and 49 polyphenols were studied for their inhibitory effects on purified Mpro. The IC50 values were 137 µg/mL for black garlic extract and 9-197 µM for 15 polyphenols. The mixtures of tannic acid with puerarin, daidzein, and/or myricetin enhanced the inhibitory effects on Mpro. The structure-activity relationship of these polyphenols revealed that the hydroxyl group in C3', C4', C5' in the B-ring, C3 in the C-ring, C7 in A-ring, the double bond between C2 and C3 in the C-ring, and glycosylation at C8 in the A-ring contributed to inhibitory effects of flavonoids on Mpro.

Characteristics of sourdough bread fermented with Pediococcus pentosaceus and Saccharomyces cerevisiae and its bio-preservative effect against Aspergillus flavus.

Six lactic acid bacteria (LAB) and four yeast strains were isolated from Pyeongchang spontaneous sourdough. In combination with the segregated Saccharomycopsis fibuligera and Saccharomyces cerevisiae, Pediococcus pentosaceus was employed for sourdough bread starters because of its antifungal action against Aspergillus flavus. The sourdough bread fermented with P. pentosaceus and S. cerevisiae displayed 56.4% ± 5.5% antifungal movement counter to A. flavus expansion at 96 h. The concentration of lactic and acetic acids in the sourdough bread was 4.5- and 1.6-folds above the control bread, respectively, contributing to the balanced sensory properties with a fermentation quotient (FQ) of 2.08-2.86. SPME- GC/MS newly distinguished twenty-two volatile compounds including six aldehydes, five alcohols, one phenol, three ketones, one acid, and six esters. The results suggest the P. pentosaceus and S. cerevisiae combination as promising sourdough starters for making enhanced quality bread free of preservatives.

Enhancement of neuroprotection, antioxidant capacity, and water-solubility of crocins by transglucosylation using dextransucrase under high hydrostatic pressure.

Crocin, one of the major carotenoid pigments of Crocus sativus (saffron), is responsible for antioxidant activity, neuroprotection, and the inhibition of tumor cell proliferation. In order to improve the functionality of crocin, α-glucosyl-(1→6)-trans-crocins (C-Gs) were synthesized using sucrose and dextransucrase from Leuconostoc mesenteroides. High hydrostatic pressure (HHP) technique was applied to the synthesis process of C-Gs in order to improve its transglucosylation yield. A 100 MPa HHP condition enhanced the production yield of C-Gs by 1.95 times compared to that of 0.1 MPa atmospheric pressure. Novel C-Gs were purified by HPLC, and their chemical structures were determined using NMR analysis. Novel C-Gs increased water solubility 4.6-5.7 times and antioxidant activity 1.5-2.6 times, respectively, compared to crocin, and their neuroprotections (cell viability 92.5-100.4 %) on HT22 mouse hippocampal neuronal cells were significantly higher than that of crocin (cell viability 84.6 %). This advanced neuroprotection of novel C-Gs could be highly associated with their enhanced antioxidant activity. Thus, the enhanced water solubility and functionality of novel C-Gs can induce better clinical efficacy of neuroprotection than trans-crocin.

Composition and biochemical properties of ale beer enriched with lignans from Schisandra chinensis Baillon (omija) fruits.

To develop a beverage with high antioxidant capacity and desirable sensory characteristics, Schisandra chinensis (omija) fruits were added to ale type beer at different time points of the brewing process. The phenolic compounds contents in beer were found to be dependent at the moment of the addition of omija fruit. Addition of omija fruits at the initiation of boiling imparted highest oxidative stability to beer and resulted in highest total phenolic and flavonoid contents in ale beer (606.82 mg GAE/L and 406.75 mg QE/L, respectively). The amounts of schisandrin, gomisin A and gomisin B in beer were 12.10 mg/mL, 3.12 mg/mL and 0.86 mg/mL, respectively. Taken together, it is hypothesized that the addition of omija fruits to traditional brewing process can improve the development of value-added beer products.

Fermented Wild Ginseng by Rhizopus oligosporus Improved l-Carnitine and Ginsenoside Contents.

We conducted this study to investigate the beneficial effects of Rhizopus oligosporus fermentation of wild ginseng on ginsenosides, l-carnitine contents and its biological activity. The Rhizopus oligosporus fermentation of wild ginseng was carried out at 30 °C for between 1 and 14 days. Fourteen ginsenosides and l-carnitine were analyzed in the fermented wild ginseng by the ultra high pressure liquid chromatography-mass spectrometry (UPLC-MS) system. Our results showed that the total amount of ginsenosides in ginseng increased from 3,274 to 5,573 mg/kg after 14 days of fermentation. Among the 14 ginsenosides tested, the amounts of 13 ginsenosides (Rg1, Rb2, Rb3, Rc, Rd, Re, Rf, Rg2, Rg3, Rh1, compound K, F1 and F2) increased, whereas ginsenoside Rb1 decreased, during the fermentation. Furthermore, l-carnitine (630 mg/kg) was newly synthesized in fermented ginseng extract after 14 days. In addition, both total phenol contents and DPPH radical scavenging activities showed an increase in the fermented ginseng with respect to non-fermented ginseng. These results show that the fermentation process reduced the cytotoxicity of wild ginseng against RAW264.7 cells. Both wild and fermented wild ginseng showed anti-inflammatory activity via inhibition of nitric oxide synthesis in RAW264.7 murine macrophage cells.

Enzymatic synthesis and biological characterization of a novel mangiferin glucoside.

Mangiferin, a major constituent of Mangifera indica L., has attracted substantial attention due to its anti-oxidant, anti-diabetic, anti-inflammatory, and anti-microbial activities. However, its poor solubility in water limits its use in food and pharmaceutical industries. In this study, novel mangiferin-(1→6)-α-d-glucopyranoside (Mg-G1) was enzymatically synthesized from mangiferin and sucrose using glucansucrase from Leuconostoc mesenteroides B-512F/KM, and optimized using response surface methodology. The water solubility of Mg-G1 was found to be 824.7 mM, which is more than 2300-fold higher than that of mangiferin. Mg-G1 also showed DPPH radical scavenging activity and superoxide dismutase (SOD)-like scavenging activity, which were 4.77- and 3.71-fold higher than that of mangiferin, respectively. Mg-G1 displayed inhibitory activity against human intestinal maltase and COX-2. Thus, the novel glucosylated mangiferin may be used as an ingredient in functional food and pharmaceutical application.

Structural and functional characteristics of clustered amylopectin produced by glycogen branching enzymes having different branching properties.

Three bacterial glycogen branching enzymes (GBEs) having different branching characteristics were used to produce clustered amylopectin (CAP), and structure and functional properties of CAPs were intensively analyzed. Branch distributions of three CAPs varied from very short (DPn = 6.65) to medium (DPn = 14.1). Branch distribution showed profound correlation with hydrodynamic diameter, water solubility, digestibility, and effects on mice gut-microbiota. All the CAPs showed nearly no viscosity and retrogradation. The very short-branch CAP exhibited more than 100-fold water-solubility, 3.5-fold lower α-amylase catalytic efficiency, and 27% lower digestibility in small intestine-mimicking condition than amylopectin. Intriguingly, medium branch CAP had 1.8-fold larger hydrodynamic diameter than the very short one. Mice gut-microbiota composition statistically varied after 12-day feeding of the CAPs, but only the medium chain CAP brought clear positive changes on the gut-microbiota. Consequently, slowly digestible starch was successfully synthesized by the single GBE, but the CAP structure affects in vivo functions in complicated manner.

Synthesis and characterization of stevioside having low degree polymerized glucosides using dextransucrase and dextranase.

Transglycosylation is one of enzymatic methods to improve the physical and biochemical properties of various functional compounds. In this study, stevioside glucosides were synthesized using sucrose as a substrate, stevioside (Ste) as an acceptor, and dextransucrase from Leuconostoc mesenteroides B-512 F/KM. The highest Ste conversion yield of 98% was obtained with 50 mg/mL Ste, 800 mM sucrose, and dextransucrase 4 U/mL at 28 °C for 6 h. The concentration of Ste was unchanged while of Ste-G1 was increased from 7.7 mM to 9.1 mM as the Ste acceptor reaction digest was treated with dextranase from Lipomyces starkeyi. Ste-G1 (13-O-ß-sophorosyl-19-O-ß-isomaltosyl-steviol), Ste-G2 (13-O-(ß-(1→6) glucosyl)-ß-glucosylsophorosyl-19-O-ß-isomaltosyl-steviol), and Ste-G2' (13-O-ß-sophorosyl-19-O-ß-isomaltotriosyl-steviol) were determined by NMR. These glucosylated Ste showed increased stabilities at pH 2, 60 °C for 48 h as compared to Ste. Ste-G1, Ste-G2, and Ste-G2' inhibited the insoluble glucan synthesis from sucrose by mutansucrase from Streptococcus muntans by the transfer of the glucosyl group of sucrose to Ste-G1, Ste-G2, and Ste-G2'. The relative water solubility of curcumin, pterostilbene or idebenone was increased by Ste or Ste glucosides treatment. Ste and Ste-G1 restored cell viability in RAW264.7 cells at concentrations up to 8 mg/mL and inhibited nitric oxide production in LPS-induced RAW264.7 cells with IC50 of 3.29 and 1.87 mg/mL.

Antimelanogenesis Effects of Fungal Exopolysaccharides Prepared from Submerged Culture of Fomitopsis castanea Mycelia.

Fungal exopolysaccharides are important natural products having diverse biological functions. In this study, exopolysaccharides from Fomitopsis castanea mycelia (FEPS) were prepared, and the highest mushroom tyrosinase inhibitory activity was found. FEPS were prepared from cultivation broth by ethanol precipitation method. The extraction yield and protein concentration of FEPS were 213.1 mg/l and 0.03%, respectively. FEPS inhibited mushroom tyrosinase with the half maximal inhibitory concentration (IC50) of 16.5 mg/ml and dose-dependently inhibited cellular tyrosinase activity (63.9% at 50 µg/ml, and 83.3% at 100 µg/ml) in the cell-free extract of SK-MEL-5 human melanoma cell and α-melanocytestimulating hormone (α-MSH)-stimulated melanin formation in intact SK-MEL-5 human melanoma cell. The IC50 of FEPS against NO production from RAW264.7 macrophage cells was 42.8 ± 0.64 µg/ml. By in vivo study using a zebrafish model, exposure of FEPS at 400 µg/ml to dechorionated zebrafish embryos for 18 h decreased the pigment density, compared to that without FEPS-treated control.

Brewing Rutin-Enriched Lager Beer with Buckwheat Malt as Adjuncts.

Brewing with buckwheat as an ingredient has been proven to be successful in several previous studies. However, few studies have focused on the effects of buckwheat on the rutin content and antioxidant activity of beer. In order to develop a lager beer with high rutin content and desirable sensory characteristics, tartary buckwheat malt was used as a brewing adjunct. The results showed that the rutin-degrading enzyme was the key factor affecting the rutin content in the wort and beer. Compared to beer made using the common mashing method, the rutin content in the buckwheat beers produced using an improved mashing method was approximately 60 times higher. The total flavonoid contents in buckwheat beers also depended strongly on the mashing methods, ranging from 530.75 to 1,704.68 mg QE/l. The rutin-rich beers also showed better oxidative stability during forced-aging. Meanwhile, the buckwheat beers were found to be acceptable in terms of the main quality attributes, flavor, and taste.

Anti-cariogenic Characteristics of Rubusoside.

Streptococcus mutans plays an important role in the development of dental caries in humans by synthesizing adhesive insoluble glucans from sucrose by mutansucrase activity. To explore the anti-cariogenic characteristics of rubusoside (Ru), a natural sweetener component in Rubus suavissimus S. Lee (Rosaceae), we investigated the inhibitory effect of Ru against the activity of mutansucrase and the growth of Streptococcus mutans. Ru (50 mM) showed 97% inhibitory activity against 0.1 U/mL mutansucrase of S. mutans with 500 mM sucrose. It showed competitive inhibition with a K i value of 1.1 ± 0.2 mM and IC50 of 2.3 mM. Its inhibition activity was due to hydrophobic and hydrogen bonding interactions based on molecular docking analysis. Ru inhibited the growth of S. mutans as a bacteriostatic agent, with MIC and MBC values of 6 mM and 8 mM, respectively. In addition, Ru showed synergistic anti-bacterial activity when it was combined with curcumin. Therefore, Ru is a natural anti-cariogenic agent with anti-mutansucrase activity and antimicrobial activity against S. mutans. ELECTRONIC SUPPLEMENTARY MATERIAL ESM: The online version of this article (doi: 10.1007/s12257-018-0408-0) contains supplementary material, which is available to authorized users.

Composition and biochemical properties of l-carnitine fortified Makgeolli brewed by using fermented buckwheat.

Makgeolli is a traditional Korean alcoholic rice beverage. It is brewed of ingredients containing starch, Nuruk, and water. In order to improve the quality and functionality of Makgeolli, the Rhizopus oligosporus fermented buckwheat containing 18.7 mg/kg of l-carnitine were utilized to brew l-carnitine fortified Makgeolli with rice. Makgeolli was prepared in two-stage fermentation method and total rutin and quercetin in each fermented buckwheat Makgeolli were increased 1.8-fold greater than buckwheat Makgeolli. DPPH antioxidant activity was enhanced in fermented buckwheat Makgeolli than buckwheat Makgeolli (21.9%-65.7%). The amounts of l-carnitine in rice Makgeolli, buckwheat Makgeolli, and fermented buckwheat Makgeolli were 0.9, 0.8-1.0, and 1.0-1.9 mg/L, respectively. The fermented buckwheat Makgeolli not only promoted health benefit by increasing l-carnitine and flavonols, but also made effective alcohol production (2.8%-8.4%) compared to common buckwheat Makgeolli, indicating the potential industrial application with health benefits.

Characterization of quinoa (Chenopodium quinoa) fermented by Rhizopus oligosporus and its bioactive properties.

Quinoa is a pseudocereal that contains high quality protein, minerals, vitamins, polyphenols, and phytosterols. In this study, quinoa was fermented by Rhizopus oligosporus (R. oligosporus) up to 5 days and the functional compounds (L-carnitine, GABA, vanillic acid and gallic acid) were analyzed by LC/MS. The amounts of L-carnitine and GABA were 0.13 mg/kg and 540 mg/kg for nonfermented quinoa (NF), 3.15 mg/kg and 1040 mg/kg for fermented quinoa at 3 days (3F), and 1.54 mg/kg and 810 mg/kg for fermented quinoa at 5 days (5F). The vanillic acid and gallic acid were 1.3 and 0.1 mg/kg for NF, 1.55 and 2.37 mg/kg for 3F, and 1.83 and 0.84 mg/kg for 5F, respectively. Total phenolic contents and total flavonoids contents were 41 mg gallic acid (GAE)/kg and 13 mg quercetin equivalent (QE)/kg for NF, 74 mg GAE/kg and 16 mg QE/kg for 3F, and 80 mg GAE/kg and 19 mg QE/kg for 5F, respectively. Antioxidant activity (SC50) was 3.6 mg/mL for NF, 3.4 mg/mL for 3F, and 2.3 mg/mL for 5F. Nitric oxide production on RAW264.7 macrophages of fermented quinoa revealed 29% and 56% inhibition of nitric oxide production for NF and 5F, respectively. Therefore, fermented quinoa can be used as a healthy and valuable food product.

The use of fermented buckwheat to produce L-carnitine enriched oyster mushroom.

L-Carnitine is an essential compound that shuttles long chain fatty acids into mitochondria. The objective of this study was to produce L-carnitine enriched oyster mushroom (Pleurotus ostreatus) using common buckwheat fermented by Rhizopus oligosporus. Mushroom grown on common buckwheat medium contained 9.9-23.9% higher L-carnitine (186.3 mg/kg) than those grown on basal medium without any buckwheat addition. Those grown on fermented common buckwheat medium contained the highest L-carnitine content (201.2 mg/kg). Size index and lightness of mushroom pileus (L*) were also the highest (100.7 and 50.6, respectively) for those grown in medium added with fermented common buckwheat (20%, w/w). Antioxidant activities of both mushroom extracts (1.5 mg/mL) showed the same level as 38.7% for mushroom grown in media added with common buckwheat or fermented common buckwheat. At the treatment concentration of 300 µg/mL, viabilities of murine macrophage cell line Raw 264.7 cells treated with ethanol extract of oyster mushroom grown on buckwheat medium ranged from 58.9 to 67.8%. The oyster mushroom grown on buckwheat and fermented buckwheat medium can be used as one of the substitutes for meat based diets.

Decrease of insoluble glucan formation in Streptococcus mutans by co-cultivation with Enterococcus faecium T7 and glucanase addition.

OBJECTIVES: To develop preventive canine oral health bio-materials consisting of probiotics and glucanase to reduce insoluble glucan and volatile sulfur compound formation. RESULTS: Co-cultivation of Enterococcus faecium T7 with Streptococcus mutans at inoculation ratio of 3:1 (v/v) resulted in 25% reduction in the growth of Streptococcus mutans. Amounts of soluble and insoluble glucans produced by S. mutans were decreased to 70 and 55%, respectively. Insoluble glucan was decreased from 0.6 µg/ml in S. mutans culture to 0.03 µg/ml in S. mutans co-cultivated with E. faecium T7 in the presence of Lipomyces starkeyi glucanase. Volatile sulfur compound, a main component of halitosis produced by Fusobacteria nucleatum, was decreased by co-cultivating F. nucleatum with E. faecium. CONCLUSION: E. faecium and glucanase can be combined as potentially active ingredients of oral care products for pets by reducing plaque-forming bacteria growth and their by-products that cause cavity and periodontal disease.

Production of steviol from steviol glucosides using ß-glucosidase from a commercial pectinase, Sumizyme PX.

OBJECTIVE: To purify and characterize a specific enzyme from a commercial pectinase for the production of steviol from stevioside (Ste) without adding organic solvent and to improve steviol production. RESULTS: Commercial Sumizyme PX converted Ste to steviol with a yield of 98%. ß-Glucosidase from Sumizyme PX (ßglyPX) was purified in three steps with 12.5-fold purification and 51% yield. The specific activity of the purified ßglyPX was 141 U/mg. The molecular weight of ßglyPX was ~ 116 kDa on SDS-PAGE. Its optimum activity was at pH 3.5 and 65 °C. It was stable for 12 h up to 55 °C and for 24 h at pH 2-9.5. K m values of ßglyPX for pNPGal, oNPGlc, lactose, and Ste were 2.4, 0.7, 18, and 7.8 mM, respectively. The optimum conditions for steviol production were 55 °C, 900 U/ml, 80 mg Ste/ml, 12 h. CONCLUSION: ßglyPX contains great potential for industrial steviol production from Ste.

Enzymatic synthesis of chlorogenic acid glucoside using dextransucrase and its physical and functional properties.

Chlorogenic acid, a major polyphenol in edible plants, possesses strong antioxidant activity, anti-lipid peroxidation and anticancer effects. It used for industrial applications; however, this is limited by its instability to heat or light. In this study, we for the first time synthesized chlorogenic acid glucoside (CHG) via transglycosylation using dextransucrase from Leuconostoc mesenteroides and sucrose. CHG was purified and its structure determined by nuclear magnetic resonance and matrix-associated laser desorption ionization-time-of-flight mass spectroscopy. The production yield of CHG was 44.0% or 141mM, as determined by response surface methodology. CHG possessed a 65% increased water solubility and 2-fold browning resistance while it displayed stronger inhibition of lipid peroxidation and of colon cancer cell growth by MTT assay, compared to chlorogenic acid. Therefore, this study may expand the industrial applications of chlorogenic acid as water-soluble or browning resistant compound (CHG) through enzymatic glycosylation.