Lysozyme-glucose stearic acid monoester conjugate formed through the Maillard reaction as an antibacterial emulsifier

Lysozyme-glucose stearic acid monoester (HEL-GE) conjugate was prepared through the Maillard reaction as an antibacterial emulsifier. The molar ratio of GE to HEL was 1:1. The isoelectric point was 6-7, which is lower than that of native HEL. Spectroscopic studies indicated that the alpha-helix content was slightly lower but the conformation around Trp had not changed and that the surface of the conjugate was covered with the GE moiety. The conjugate maintained approximately 53-57% of the enzymatic activity of native HEL at 40-60 degrees C and exhibited considerable resistance to proteolysis. The denaturation temperature of the conjugate was approximately 74 degrees C, somewhat higher than that of control HEL, whereas the enthalpy was about one-third of that of control HEL. The emulsifying activity of the conjugate and the emulsion stability were much enhanced compared to those of native HEL, and the conjugate maintained approximately 70% of the bactericidal activity.     

Novel fibrinolytic enzyme in fermented shrimp paste, a traditional asian fermented seasoning

A novel fibrinolytic enzyme was purified from fermented shrimp paste, a popular seasoning used in Asian countries. The enzyme is a monomer with an apparent molecular weight of 18 kDa, and it is composed primarily of beta-sheet and random coils. The N-terminal amino acid sequence was determined to be DPYEEPGPCENLQVA. It is a neutral protease with an optimal activity from pH 3 to 7. No inhibition was observed with PMSF, Pepstatin A, E64, and 1,10-phenanthroline, but the enzyme was slightly inhibited by EDTA and Cu(2+). It was relatively specific to fibrin or fibrinogen as a protein substrate, yet it hydrolyzed none of the plasma proteins in the studies. In vitro, the enzyme was resistant to pepsin and trypsin digestion. It also had an anticoagulant activity measured with activated partial thrombin time and prothrombin time tests. The novel fibrinolytic enzyme derived from traditional Asian foods is useful for thrombolytic therapy. In addition, this enzyme has a significant potential for food fortification and nutraceutical applications, such that its use could effectively prevent cardiovascular diseases.     

Release and activity of allelochemicals from allelopathic rice seedlings

3-Isopropyl-5-acetoxycyclohexene-2-one-1 (1), momilactone B (2), and 5,7,4'-trihydroxy-3',5'-dimethoxyflavone (3) were isolated and identified from an allelopathic rice accession PI312777. These three compounds at low concentrations could inhibit the growth of weeds Echinochloa crusgalli and Cyperus difformis associated with rice, especially mixtures of the compounds had stronger inhibitory activity than did individual compounds. Studies with hydroponic culture, continuous root exudates trapping system (CRETS), and direct resin adsorption methods showed that a total of 7.6 n moles 1, 2, and 3 were exuded from living roots of each seedling into the environment at 10 days after seedlings were transplanted. Furthermore, 1, 2, and 3 were found in the soil growing PI312777 seedlings at day 15 after seedlings emergence and reached a total of 39.5 microg/g soil at day 30. The results indicated that PI 312777 seedlings could release sufficient quantities of 1, 2, and 3 into the environment to act as allelochemicals inhibiting the growth of associated weeds. Investigations on the distribution of 1, 2, and 3 in PI 312777 plant, and its root exudates showed that the levels of 1, 2, and 3 were significantly higher in the shoots and root exudates than in the roots, and only trace 1 was observed in the roots. The results suggest that the roots of rice seedlings are not major site of synthesis or accumulation 1, 2, and 3, but a pathway for their release into the environment. The levels of 1, 2, and 3 in the root exudates were over 2-folds higher under direct resin adsorption than under hydroponic culture and CRETS, and hence, it is the preferred method to collect and identify active allelochemicals in rice exudates in future studies on rice allelopathy.     

Antimicrobial peptides released by enzymatic hydrolysis of hen egg white lysozyme

This work was aimed at the isolation, purification, and characterization of novel antimicrobial peptides from chicken egg white lysozyme hydrolysate, obtained by peptic digestion and subsequent tryptic digestion. The hydrolysate was composed of over 20 small peptides of less than 1000 Da, and had no enzymatic activity. The water-soluble peptide mixture showed bacteriostatic activity against Gram-positive bacteria (Staphylococcus aureus 23-394) and Gram-negative bacteria (Escherichia coli K-12). Two bacteriostatic peptides were purified and sequenced. One peptide, with the sequence Ile-Val-Ser-Asp-Gly-Asp-Gly-Met-Asn-Ala-Trp, inhibited Gram-negative bacteria E. coli K-12 and corresponded to amino acid residues 98-108, which are located in the middle part of the helix-loop-helix. Another novel antimicrobial peptide inhibited S. aureus 23-394 and was determined to have the sequence His-Gly-Leu-Asp-Asn-Tyr-Arg, corresponding to amino acid residues 15-21 of lysozyme. These peptides broadened the antimicrobial activity of lysozyme to include Gram-negative bacteria. The results obtained in this study indicate that lysozyme possesses nonenzymatic bacteriostatic domains in its primary sequence and they are released by proteolytic hydrolysis.     

Method for analysis of tannic acid and its metabolites in biological samples: application to tannic acid metabolism in the rat

A new analytical method for measuring tannic acid (TA) using tannase was developed and applied to the investigation of TA metabolism in the rat following oral administration at a dose of 1.0 g/kg. The proposed method for TA determination was based on the enzymatic hydrolysis of TA to gallic acid (GA) and subsequent determination by HPLC. TA metabolites were determined by HPLC. 4-O-Methylgallic acid (4-OMGA), pyrogallol (PY), and resorcinol (RE) were detected in serum. TA was excreted into urine as GA (0.01%), 4-OMGA (0.10%), PY (0.24%), and RE (2.06%) and into feces as TA (62.74%), GA (0.19%), PY (0.02%), and RE (0.76%) within 54 h after oral administration. It was suggested that >60% of TA remained unchanged but that some was hydrolyzed to GA by tannase in the intestine and further metabolized to 4-OMGA, PY, and RE.     

Bacteriological and genetic assessment of game meat from Japanese wild boars

Bacterial tests were used to assess bacterial contamination of game meat from Japanese wild boars. The bacterial contamination of wild boar meat was less than that of domestic pork, as determined by aerobic plate counts (APC) and coliform counts. None of the meat examined in this study was contaminated by Salmonella or E. coli O-157. To detect adulteration by domestic pig meat or European wild boar meat, 46 samples of game meat sold as Japanese wild boar were examined genetically. A total of 17 samples showed genetic haplotypes of European and Asian domestic pigs in the D-loop of mitochondrial DNA (mtDNA), and 16 samples showed nuclear glucosephosphate isomerase-processed pseudogene (GPIP) genotypes of European domestic pigs. The European GPIP genotypes of these samples were confirmed by PCR-RFLP analysis. These results indicate that some game meat sold as Japanese wild boar is adulterated by cross-breeding between pigs and wild boars or by contamination with meat from domestic pigs or European wild boars.     

Functional properties of oat globulin modified by a calcium-independent microbial transglutaminase

Oat globulin was modified by a calcium-independent microbial transglutaminase (TG). The TG-polymerized protein had higher solubility than the control at acidic pH and had improved water- and fat-binding properties. Incubation of 10% (w/v) oat globulin dispersions in the presence of TG at 37 degrees C led to the formation of a well-developed viscoelastic gel network with a microstructure characterized by thick strands and large clusters. The TG-induced gels had higher modulus values, lower loss tangent values, and lower frequency dependency than the heat-induced gels. The TG-induced gel system has the characteristics of classical polymer gel with permanent "chemical" cross-links, whereas the heat-denatured system has the characteristics of a temporary "physical" gel with breakable cross-links. Fourier transform infrared spectroscopy showed marked shift and intensity changes in several major bands, suggesting pronounced changes in protein conformation during TG-induced gelation. Aggregation of protein molecules was also indicated by the progressive increases in two infrared bands (1679-1682 and 1622-1625 cm(-)(1)) associated with the formation of intermolecular beta-sheets and strands. Results suggest that new food polymers with unique functionality can be produced from oat globulin treated with TG and that elastic gels can be formed near neutral pH, instead of the alkaline pH required for thermally induced oat globulin gels.     

Suppressive effect of saturated acyl L-ascorbate on the oxidation of linoleic acid encapsulated with maltodextrin or gum arabic by spray-drying

6-O-Palmitoyl L-ascorbate was added to linoleic acid at various molar ratios of the ascorbate to the acid, the mixtures were emulsified with a maltodextrin or gum arabic solution, and the emulsions were spray-dried to produce microcapsules. At higher molar ratios, the oil droplets in the emulsions were smaller, and the oxidative stabilities of the encapsulated linoleic acid were higher for both the maltodextrin- and gum arabic-based microcapsules. 6-O-Capryloyl, caproyl, and lauroyl L-ascorbates, which were synthesized through lipase-catalyzed condensation in acetone, were also used for the microencapsulation of linoleic acid. Except for capryloyl L-ascorbate, the addition of a saturated acyl ascorbate, especially caproyl ascorbate, to linoleic acid was effective for preparing oil droplets of small particle diameter and for suppressing the oxidation of the encapsulated linoleic acid.     

Oxidative degradation of bisphenol a by crude enzyme prepared from potato

When crude enzymes prepared from some vegetables and fruits were incubated with bisphenol A (2,2-bis(4-hydroxyphenyl)propane, BPA) at 37 degrees C, BPA was oxidized by crude enzymes from potato, eggplant, and lettuce. The crude enzyme prepared from potato (Solanum tuberosum) had the strongest oxidative activity for BPA. Its optimal temperature and pH were 40-45 degrees C and 8.0, respectively. More than 95% of BPA was oxidized after the incubation with potato enzyme for 60 min. BPA gave two oxidation products besides insoluble compounds during the oxidation by potato enzyme. The oxidation products were identified to be 4[1-(4-hydroxyphenyl)-1-methyl-ethyl]-benzene-1,2-diol and 4[1-(4-hydroxyphenyl)-1-methyl-ethyl]-benzene-1,3-diol. Enzymatically oxidized BPA lost the estrogen-like activity to enhance the growth of human breast cancer (MCF-7) cells.     

Aggregates and gel network structure of globin hydrolysates

A gel with excellent functional properties was prepared successfully using the hydrolysates of globin. In the present study, the structures of intermediate aggregates and gel network were observed directly with an electronic microscope. It was shown clearly that the intermediate aggregates were in a thin rod shape with a length of 130--140 nm, which was in good accordance with the results of the light scattering obtained in a previous study. The diameter of intermediate aggregates was 4--5 nm. Each unit of the intermediate aggregate was composed of beta-chain and peptide beta-1 in a ratio of 1:1. Its molecular weight was 26922 Da, and it had a diameter of 4.1 nm. The thin rod-shaped aggregates were formed with units through the hydrophobic interaction. The length of intermediate aggregate was >30--33 times the diameter. Furthermore, the cross-linked structure formed by peptide alpha-1 and the thin rod-shaped aggregates was also confirmed by the photography of the electronic microscope. These results supported the model proposed in previous papers as proper to depict exactly the formation and structure of the gel network of globin hydrolysates.