Isolation and identification of indigestible pyroglutamyl peptides in an enzymatic hydrolysate of wheat gluten prepared on an industrial scale

An enzymatic hydrolysate of wheat gluten was further digested in vitro with porcine pepsin and pancreatin to obtain an indigestible peptide. Indigestible pyroglutamyl peptide was isolated from the digest by strong cation-exchange, size-exclusion, and reversed-phase chromatographies. The pyroglutamyl peptide was digested with pyroglutamate aminopeptidase, and the digest was reacted with phenyl isothiocyanate. The resultant phenylthiocarbamyl (PTC) peptides were purified by reversed-phase HPLC by using binary gradient elution with ammonium acetate buffer, pH 6.0, and acetonitrile. The PTC peptides were analyzed with an automatic peptide sequencer on the basis of the Edman degradation method with a modified program. Some pyroglutamyl peptides were also analyzed by fast-atom bombardment ionization mass spectrometry without the pyroglutamate amino peptidase digestion. Consequently, pyroGlu-Asn-Pro-Gln, pyroGlu-Gln-Gln-Pro-Gln, pyroGlu-Gln-Pro-Gln, pyroGlu-Gln-Pro-Gly-Gln-Gly-Gln, pyroGlu-Gln, pyroGlu-Gln-Pro, pyroGlu-Ile-Pro-Gln, pyroGlu-Ile-Pro, pyroGlu-Gln-Pro-Leu, pyroGlu-Gln-Phe-Pro-Gln, pyroGlu-Ser-Phe-Pro-Gln, pyroGlu-Phe-Pro-Gln, and pyroGlu-Gln-Pro-Pro-Phe-Ser were identified.     

Characterization of <Emphasis Type="Italic">Inago1</Emphasis> and <Emphasis Type="Italic">Inago2</Emphasis> retrotransposons in <Emphasis Type="Italic">Magnaporthe oryzae</Emphasis>

From the genome of a Japanese field isolate of the rice blast fungus, Magnaporthe oryzae, we newly identified Inago1 and Inago2 LTR retrotransposons. Both elements were found to be Ty3/gypsy-like elements whose copies were dispersed within the genome of Magnaporthe spp. isolates infecting rice and other monocot plants. Southern hybridization patterns of nine re-isolates derived from conidia of the strain Ina168 produced after a methyl viologen treatment were not changed, indicating that the insertion pattern of Inago elements is relatively stable.     

Analysis of phenolic compounds in white rice, brown rice, and germinated brown rice

Two hydroxycinnamate sucrose esters, 6'-O-(E)-feruloylsucrose and 6'-O-(E)-sinapoylsucrose, were isolated from methanol extracts of rice bran. Soluble and insoluble phenolic compounds as well as 6'-O-(E)-feruloylsucrose and 6'-O-(E)-sinapoylsucrose from white rice, brown rice, and germinated brown rice were analyzed using HPLC. The results demonstrated that the content of insoluble phenolic compounds was significantly higher than that of soluble phenolics in rice, whereas almost all compounds identified in germinated brown rice and brown rice were more abundant than those in white rice. 6'-O-(E)-Feruloylsucrose (1.09 mg/100 g of flour) and 6'-O-(E)-sinapoylsucrose (0.41 mg/100 g of flour) were found to be the major soluble phenolic compounds in brown rice. During germination, an approximately 70% decrease was observed in the content of the two hydroxycinnamate sucrose esters, whereas free phenolic acid content increased significantly; the ferulic acid content of brown rice (0.32 mg/100 g of flour) increased to 0.48 mg/100 g of flour and became the most abundant phenolic compound in germinated brown rice. The content of sinapinic acid increased to 0.21 mg/100 g of flour, which is nearly 10 times as much as that in brown rice (0.02 mg/100 g of flour). In addition, the total content of insoluble phenolic compounds increased from 18.47 mg/100 g of flour in brown rice to 24.78 mg/100 g of flour in germinated brown rice. These data suggest that appropriate germination of brown rice may be a method to improve health-related benefits.     

High mobility group box 1 (HMGB1) protein is present in the cerebrospinal fluid of dogs with encephalitis

Many cases of encephalitic disease with unknown etiologies have been reported in specific breeds of small dogs. High mobility group box 1 (HMGB1) in neuronal cells was recently found to be a novel cytokine-like mediator that is a marker of neuronal necrosis and inflammation. The aim of this study was to determine whether HMGB1 levels are elevated in the cerebrospinal fluid (CSF) of dogs suspected of having encephalitis. CSF was obtained from 31 dogs that were diagnosed with an encephalitic disease by clinical examinations and magnetic resonance image (MRI) scanning. The CSF samples were analyzed via western blotting (WB) and an enzyme-linked immunosorbent assay (ELISA) with a polyclonal antibody against HMGB1. The mean HMGB1 concentration was significantly higher in the encephalitic dogs than that in the healthy controls. The concentrations of HMGB1 were correlated with the cell counts and total protein concentrations, which are known CSF indicators of the neuronal inflammation associated with encephalitis. These results suggest that HMGB1 protein in CSF confirms the presence of necrosis and inflammation in most cases of canine encephalitis and that HMGB1 will be a new indicator of encephalitis.     

Immunohistochemical examination of PDGF-AB, TGF-beta and their receptors in the hemocytes of a tick, Ornithodoros moubata (Acari: Argasidae)

Growth factors, Platelet Derived Growth Factor (PDGF) and Transforming Growth Factor (TGF)-beta, were demonstrated in vertebrate and invertebrate immmunocytes. It is generally known that the growth factors are important in various biological processes, such as the regulation of cell differentiation, development and wound healing. In the present study, the presence of TGF-beta1 and PDGF-receptor-alpha in plasmatocytes and PDGF-AB in granulocytes of a soft tick, Ornithodoros moubata, was confirmed immunohistochemically. The tick midgut might be damaged by intracellular digestion and penetration of protozoa. Therefore, it is considered that PDGF from granulocytes may affect the PDGF-receptor-alpha in plasmatocytes and TGF-beta from plasmatocytes may function to repair the midgut. The results obtained here add to the elucidation of the functions of tick hemocytes.     

Lysine decarboxylase-negative Salmonella enterica serovar enteritidis: antibiotic susceptibility, phage and PFGE typing

One hundred twenty Salmonella Enteritidis isolates collected from 1992 to 2005 in Nagasaki prefecture (65 isolates from 40 outbreak cases, 44 from sporadic diarrhea patients, and 11 from chicken-related products) were investigated by their antibiotic susceptibility profiles, phage typing, and pulsed-field gel electrophoresis (PFGE) typing. Out of them, 18 were identified as lysine decarboxylase (LDC)-negative isolates, and 15 showed resistance toward streptomycin. Based on the PFGE typing, the isolates were classified into five clusters by UPGMA clustering method. Three LDC-negative isolates belonged to cluster A and were of phage type (PT) 4 and isolated between 2000 and 2004. Other 15 LDC-negative isolates belonged to cluster E. They were PT1, reacted but did not conform (RDNC), or untypable and were isolated between 2001 and 2004. LDC-negative isolates of the cluster A differed from LDC-negative isolates of the cluster E in antibiotic susceptibility profiles, phage typing, and PFGE typing. LDC-negative isolates of the cluster E were isolated after 2001 in Nagasaki prefecture.     

Putative host cell receptor for fowl adenovirus detected in gizzard

This work was done to identify a fowl adenovirus (FAV) binding protein in the gizzard, a known target organ for certain strains of FAV serotype 1. By using a virus overlay protein binding assay (VOPBA), a putative FAV binding protein of approximately 200 kDa expressed in the gizzard was detected.     

Dynamic aspects of N.P.K. uptake and O2 secretion in relation to the metabolic pathways within the plant roots

As previously reported^1)-4) the inhibition of nutrients uptake by rice plant with such respiration inhibitors as hydrogen sulfide, butyric acid, cyanide, and azide differed strikingly from element to element. The order of suppression, however, was similar in these respiratory inhibitors, and the Ce I Itlon coefficients as revealed in terms of percents of decrease were grouped in the order of K²O·P²O⁵>SiO²·SO³·Br>MnO·NH⁴-N·H²O2> MgO·CaO. This established order attracted the authors interest to elucidate the essential mechanism of the uptake of each element, possibly linked specifically with metabolic pathways, particularly of respiration, within the plant roots. In view of the prime importance of three major fertilizer elements N.P.K., the specific metabolic linkages of these three elements were subjected to a energetic study.     

Diagnosis of equine piroplasmosis in Xinjiang province of China by the enzyme-linked immunosorbent assays using recombinant antigens

The prevalence of equine piroplasmosis in Xinjiang province, China, was examined by enzyme-linked immunosorbent assays (ELISAs). A total of 70 serum samples were taken from horses pastured on three farms in western Xinjiang, and examined for diagnosis of equine Babesia equi (B. equi) infection and B. caballi infection by ELISAs using recombinant equi merozoite antigen 1 (EMA-1) and recombinant P48 antigen, respectively. Of the 70 samples, 28 (40.0%) and 17 (24.3%) samples were positive for B. equi infection and B. caballi infection, respectively. In addition, 11 (15.7%) samples were positive for both B. equi and B. caballi infections. These results indicate that equine piroplasmosis is widespread and therefore a cause for serious concern in western Xinjiang. To our knowledge, this is the first report describing a survey on equine piroplasmosis in Xinjiang province, China.     

Micropores and mesopores in the cell wall of dry wood

To investigate micropores and mesopores in the cell walls of dry wood, CO₂ gas and N₂ gas adsorption onto dry wood were measured at ice-water temperature (273 K) and liquid nitrogen temperature (77 K). CO₂ gas adsorption isotherms obtained were used for determining micropore volumes smaller than 0.6 nm by the HK method (Horvath-Kawazoe method), and N₂ gas adsorption isotherms obtained were used for determining the mesopore volume between 2 nm and 50 nm by the Barrett-Joyner-Halenda (BJH) method. Micropores and mesopores existed in cell walls of dry wood, and the cumulative pore volume was much larger for micropores than for mesopores. Micropores in the cell wall of dry wood decreased with elevating heat treatment temperature, and the decreased micropore was reproducible by wetting and drying. Mesopores did not decrease so much with elevating heat treatment temperature. Micropore volumes for the softwood Hinoki and the hardwood Buna were compared. A larger amount of micropores existed in hardwood Buna than in softwood Hinoki, and this relationship was considered to correspond to the difference in thermal softening properties for lignin in water-swollen Hinoki and Buna. This result probably indicates that micropores in the cell walls of dry wood relate to the structure of lignin.