Liyan Zhao

Effect of Nano‐Packing on Preservation Quality of Fresh Strawberry (Fragaria ananassa Duch. cv Fengxiang) during Storage at 4 °C

UNLABELLED A novel nano-packing material with lower relative humidity, oxygen transmission rate and high longitudinal strength was synthesized by blending polyethylene with nano-powder (nano-Ag, kaolin, anatase TiO(2), rutile TiO(2)), and its effect on preservation quality of strawberry fruits (Fragaria ananassa Duch. cv Fengxiang) was investigated during storage at 4 degrees C. Results showed that nano-packaging was able to maintain the sensory, physicochemical, and physiological quality of strawberry fruits at a higher level compared with the normal packing (polyethylene bags). After a 12-d storage, decreases in the contents of total soluble solids, titratable acidity, and ascorbic acid of nano-packing were significantly inhibited. Meanwhile, decay rate, anthocyanin, and malondialdehyde contents were decreased to 16.7%, 26.3 mg/100g, 66.3 micromol/g for nano-packing and 26.8%, 31.9 mg/100g, 75.4 micromol/g for normal packing; polyphenoloxidase (PPO) and pyrogallol peroxidase (POD) activities were significantly lower in nano-packing than the control. These data indicated that the nano-packaging might provide an attractive alternative to improve preservation quality of the strawberry fruits during extended storage. PRACTICAL APPLICATION Nano-packing exhibited identified quality benefits applicable to the preservation of fresh strawberry. Furthermore, nano-packing has the advantages of simple processing and feasibility to be industrialized in contrast with other storages. Thus, the utilization of nano-packing will likely assist commercial producers and retailers in extending the shelf life of products over a broader range in the future.

A Tricholoma matsutake Peptide with Angiotensin Converting Enzyme Inhibitory and Antioxidative Activities and Antihypertensive Effects in Spontaneously Hypertensive Rats

Hypertension is a major risk factor for cardiovascular disease. A crude water extract of the fruiting bodies of a highly prized mushroom Tricholoma matsutakei exerted an antihypertensive action on spontaneously hypertensive rats (SHRs) at a dosage of 400 mg/kg. An angiotensin converting enzyme (ACE) inhibitory peptide with an IC50 of 0.40 μM was purified from the extract and designated as TMP. Its amino acid sequence was elucidated to be WALKGYK through LC-MS/MS analysis. The Lineweaver-Burk plot suggested that TMP was a non-competitive inhibitor of ACE. A short-term assay of antihypertensive activity demonstrated that TMP at the dosage of 25 mg/kg could significantly lower the systolic blood pressure (SBP) of SHRs. TMP exhibited remarkable stability over a wide range of temperatures and pH values. It also demonstrated 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity. The aforementioned activities of TMP were corroborated by utilizing the synthetic peptide. Hence T. matsutake can be used as a functional food to help prevent hypertension- associated diseases.

Effects of Selenium and Methionine Supplementation of Breeder Hen Diets on Selenium Concentration and Oxidative Stability of Lipids in the Thigh Muscles of Progeny

The effects of dietary supplementation to female chickens with selenium (Se) and methionine (Met) on the next generation were studied. Lang-shan breeding hens (450) were obtained at 52 wk of age and randomly allotted to 9 treatments; 5 replicates of each treatment were carried out. The breeders were fed a basal corn-soybean meal diet (0.13 mg Se/kg) supplemented with 0, 0.30, or 0.60 mg/kg Se from Sel-Plex and 0.32%, 0.40%, or 0.54% Met for the 30-d adapting period and 70-d experiment period. Se and glutathione (GSH) concentrations, glutathione peroxidase (GSH-Px) activity, and the oxidative stability of muscular lipids of 90-d progeny were determined by testing the TBARS values. When breeders received the highest levels of Met or Se, GSH-Px activity was decreased, the Se concentration and the oxidative stability of muscular lipids were increased with the supplementation of Se or Met. When breeder hens were given a Met-deficient diet, supplementing with Se decreased the Se deposition in progeny thigh. With regard to lipid oxidation, 0.3 mg/kg maternal dietary Se supplementation decreased the oxidative stability of muscle lipid and 0.6 mg/kg Se supplementation showed no difference from the control. When breeders were fed a Se-deficient diet, the GSH-Px activity was increased significantly and the oxidative stability of progeny muscles was decreased with the supplementation of Met. It was concluded that supplementation of the maternal diet with higher Se and Met can increase Se deposition in progeny muscle and lead to more effective protection against lipid oxidation in progeny thighs.

A protease-resistant α-galactosidase from Pleurotus djamor with broad pH stability and good hydrolytic activity toward raffinose family oligosaccharides.

An acidic α-galactosidase designated as PDGI (Pleurotus djamor α-galactosidase) was purified to homogeneity with 290-fold purification and a specific activity of 52.18 units/mg by means of ion exchange chromatography and gel filtration chromatography. PDGI is a monomeric protein exhibiting a molecular mass of 60kDa in SDS-PAGE and gel filtration. The optimum pH and temperature of the enzyme with pNPGal as substrate were 5.0 and 53.5°C, respectively. It displayed great pH stability within the pH range 3.0-10.0. Besides, the enzyme harbored remarkable resistance to acid protease and varying degrees of tolerance to other proteases: trypsin>collagenase Type-I>α-chymotrypsin neutral protease>proteinaseK. It was strongly inhibited by K+, Cd2+, Cu2+, Hg2+, Al3+, Fe3+ and Ag + ions. The chemical modification reagents diethypyrocarbonate (DEPC), 2,3-butanedione (DIC) and trinitrophenol (TNBS) increased the activity of PDGI 1.5-fold whereas N-bromosuccinimide (NBS) and parachloro-mercuri-benzoate (PCMB) drastically suppressed its activity. PDGI displayed activity toward stachyose and raffinose. The Km values for hydrolysis of pNPGal, stachyose and raffinose were 0.76, 7.63 and 6.29mM, respectively. Furthermore, PDGI degraded raffinose and sthachyose. These results suggest that PDGI has great potential for elimination of the non-digestible and flatulence-causing oligosaccharides stachyose and raffinose from legumes.

Isolation and Characterization of a Novel Lectin from the Edible Mushroom Stropharia rugosoannulata

To date, only a few steroids have been isolated from the mushroom Stropharia rugosoannulata which can be cultivated. In this paper, a novel lectin (SRL) with a molecular weight of 38 kDa, and a unique IKSGVYRIVSWQGALGPEAR N-terminal sequence was isolated from S. rugosoannulata, which represents the first protein isolated from the mushroom. The purification methods included (NH4)2SO4 precipitation, ion exchange chromatography on CM-cellulose, Q-Sepharose, and SP-Sepharose, and gel- filtration on Superdex-75. The lectin was adsorbed on all three types of ion exchangers and was purified more than 450-fold. The lectin was stable below 70 °C (with half of the activity preserved at 80 °C), and in the presence of NaOH and HCl solutions up to a concentration of 12.5 mM and 25 mM, respectively. The hemagglutinating activity of SRL was inhibited by inulin. Cd2+ and Hg2+ ions strongly reduced the hemagglutinating activity at concentrations from 1.25 mM to 10 mM. SRL exhibited anti-proliferative activity toward both hepatoma Hep G2 cells and leukemia L1210 cells, with an IC50 of 7 μM and 19 μM, respectively. The activity of HIV-1 reverse transcriptase could also be inhibited by SRL, with an IC50 of 10 μM.

Biochemical Characteristics of Three Laccase Isoforms from the Basidiomycete Pleurotus nebrodensis

The characterization of three laccase isoforms from Pleurotus nebrodensis is described. Isoenzymes Lac1, Lac2 and Lac3 were purified to homogeneity using ion exchange chromatography on DEAE-cellulose, CM-cellulose and Q-Sepharose and a gel filtration step on Superdex 75. The molecular weights of the purified laccases were estimated to be 68, 64 and 51 kDa, respectively. The isoenzymes demonstrated the same optimum pH at 3.0 but slightly different temperature optima: 50–60 °C for Lac1 and Lac3 and 60 °C for Lac2. Lac2 was always more stable than the other two isoforms and exposure to 50 °C for 120 min caused 30% loss in activity. Lac2 was relatively less stable than the other two isoforms when exposed to the pH range of 3.0–8.0 for 24 h, but inactivation only occurred initially, with around 70% residual activity being maintained during the whole process. Oxidative ability towards aromatic compounds varied substantially among the isoforms and each of them displayed preference toward some substrates. Kinetic constants (Km, Kcat) were determined by using a 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) assay, with Lac3 showing the best affinity and Lac2 displaying the highest catalytic efficiency. Amino acid sequences from peptides derived from digestion of isoenzymes showed great consistency with laccases in the databases.

A Fungal α-Galactosidase from Tricholoma matsutake with Broad Substrate Specificity and Good Hydrolytic Activity on Raffinose Family Oligosaccharides

An acidic α-galactosidase designated as TMG was purified from the fruiting bodies The purification protocol entailed ion exchange chromatography on Q-Sepharose and of Tricholoma matsutake with 136-fold purification and a specific activity of 909 units/mg. Mono-Q and fast protein liquid chromatography on Superdex 75. TMG is a monomeric protein exhibiting a molecular mass of 47 kDa in SDS-PAGE and gel filtration. The purified enzyme was identified by LC-MS/MS and three inner amino acid sequences were obtained. The optimum pH and temperature for TMG with pNPGal as substrate were pH 4.5 and 55 °C, respectively. The α-galactosidase activity was strongly inhibited by K+, Ca2+, Cd2+, Hg2+, Ag+ and Zn2+ ions. The enzyme activity was inhibited by the chemical modification agent N-bromosuccinimide (NBS), indicating the importance of tryptophan residue(s) at or near the active site. Besides hydrolyzing pNPGal, TMG also efficaciously catalyzed the degradation of natural substrates such as stachyose, raffinose, and melibiose. Thus TMG can be exploited commercially for improving the nutritional value of soy milk by degradation of indigestible oligosaccharides.

Isolation of an Angiotensin I-Converting Enzyme Inhibitory Protein with Antihypertensive Effect in Spontaneously Hypertensive Rats from the Edible Wild Mushroom Leucopaxillus tricolor

An 86-kDa homodimeric angiotensin I-converting enzyme (ACE) inhibitory protein designated as LTP was isolated from fruit bodies of the mushroom Leucopaxillus tricolor. The isolation procedure involved ultrafiltration through a membrane with a molecular weight cutoff of 10-kDa, ion exchange chromatography on Q-Sepharose, and finally fast protein liquid chromatography-gel filtration on Superdex 75. LTP exhibited an IC50 value of 1.64 mg∙mL−1 for its ACE inhibitory activity. The unique N-terminal amino acid sequence of LTP was disclosed by Edman degradation to be DGPTMHRQAVADFKQ. In addition, seven internal sequences of LTP were elucidated by liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. Results of the Lineweaver-Burk plot suggested that LTP competitively inhibited ACE. Both LTP and the water extract of L. tricolor exhibited a clear antihypertensive effect on spontaneously hypertensive rats.

Isolation of a protease-resistant and pH-stable α-galactosidase displaying hydrolytic efficacy toward raffinose family oligosaccharides from the button mushroom Agaricus bisporus

A 45-kDa monomeric acidic α-galactosidase with a specific activity of 193.12 units/mg was isolated from the fruiting bodies of Agaricus bisporus. Blast search of internal peptide sequences suggested that it is a member of GH family 27 and it is most similar to hypothetical protein AGABI2DRAFT_70106. The enzyme displayed maximal activity at pH 4.0 and 60°C, respectively. The enzyme remained stable within the pH range 2.0-9.0 but its activity was markedly suppressed in the presence of Cu2+, Hg2+, Fe3+ and Ag+ ions. It displayed resistance to α-chymotrypsin and neutral protease. Moreover, it manifested degradative activity toward both oligosaccharides and polysaccharides. The enzyme manifested Km values of 0.30mM, 10.65mM and 19.21mM, toward pNPGal, stachyose and raffinose respectively. These results suggest that Agaricus bisporus α-galactosidase is a promising candidate for elimination of raffinose oligosaccharides (RFOs) in biotechnological applications.

A novel ribonuclease with HIV-1 reverse transcriptase inhibitory activity purified from the fungus Ramaria formosa

A purification protocol that encompassed anion exchange chromatography (EC) on DEAE‐cellulose, cation EC on CM‐cellulose, anion EC on Q‐Sepharose, and fast protein liquid chromatography‐gel filtration of Superdex 75 was used to isolate a ribonuclease from dried fruiting bodies of Ramaria formosa. The ribonuclease was unadsorbed on CM‐cellulose but adsorbed on both DEAE‐cellulose and Q‐Sepharose. It displayed a molecular mass of 29‐kDa in both gel filtration and sodium dodecyl sulfate–polyacrylamide gel electrophoresis. The ranking of its ribonucleolytic activity toward polyhomoribonucleotides was poly(U) > poly(C) > poly(G) > poly(A). It exhibited a pH optimum of pH 5 and a temperature optimum at 60 °C. The ribonuclease inhibited HIV‐1 reverse transcriptase with an IC50 of 3 µM.