Isolation and Structure of a New Victoxinine Derivative Produced by Helminthosporium victoriae

The structures of alkyl radicals generated in several methyl esters of fatty acids by irradiation with UV light were studied by the spin trapping technique. A spin trap, deuterated nitrosodurene, traps alkyl radicals in both saturated and unsaturated esters at the ambient temperature. The trapped radicals and their hyperfine splitting constants from several esters were as follows: pentadienyl radicals (a_N= 13.8 ~ 14.0 G, a_H = 5.9 ~ 6.0 G) from methyl linoleate, linolenate and docosahexaenoate; allyl radicals (a_N = 13.9 G, a_H = 6.8 G) and α-carbon radicals (a_N = 13.3 G, a_H = 10.0 G) from methyl oleate and elaidate; α-carbon radicals (a_N = 13.3 ~ 13.4 G, a_H = 9.6 ~ 10.0 G) and secondary alkyl radicals (a_N = 13.9 G, a_H = 6.8 ~ 7.2 G) from saturated esters.     

Effects of Alloxan Diabetes and Hypophysectomy on Growth,and Plasma and Liver Free Amino Acids of Rats Fed Excess Glycine Diets

Three chitinases (EC 3.2.1.14) were purified from yam, Dioscorea opposita THUMB, by fractionation with ammonium sulfate, chromatographies on DEAE-Cellulose and DEAE-Sephadex A-50, chromatofocusing and gel filtration on Bio-Gel P-60. The purified enzymes (E-l, E-2 and E-3) showed single bands on sodium dodecylsulfate polyacrylamide gel electrophoresis, and the molecular weights were estimated to be 33,500. The pIs were 4.05 (E-l), 4.0 (E-2) and 3.8 (E-3). All enzymes were glycoproteins and the neutral sugar contents were 3.6% (E-l), 3.6 (E-2) and 0.9% (E-3). The N-terminal amino acids of E-l and E-3 were the same and determined to be histidine. All enzymes hydrolyzed glycolchitin, but not p-nitrophenyl-2-acetamido-2-deoxy-β-d-glucopyranoside or Micrococcus lysodeikticus cell walls. E-l and E-3 were stable in the pH range of 5 ~ 11, and below 60°C. These enzymes showed two optimum pHs around 3.5 and 8.0 or 8.5 with glycolchitin as substrate.     

Isolation and Characterization of PDE-I and II,the Inhibitors of Cyclic Adenosine-3′,5′-monophosphate Phosphodiesterase

In the screening for inhibitors of cyclic adenosine-3′,5′-monophosphate phosphodiesterase, two compounds, PDE-I (C₁₃H₁₃N₃O₅) and PDE-II (C₁₄H₁₄N₂O₅), were isolated from culture filtrates of a Streptomyces. Concentrations for 50% inhibitions of PDE-I and PDE-II against the high Km enzyme were 15 µm and 13 µm, and those against the low Km enzyme were 65 µm and 130 µm, respectively. Production, isolation and characterization of these compounds are described.     

Supplemental Effects of Arginine and Methionine on Growth,and on Formations of Urea and Creatine of Adrenalectomized Rats Fed High Glycine Diets

The effects of adrenalectomy on growth, some enzyme activities in the liver and kidney, and urinary excretion of urea, creatinine and creatine were investigated in rats fed the 10% casein diets containing 7% glycine with or without l-arginine and l-methionine (10C, 10C7G and 10C7ArgMet).

Body weight gains of the intact 10C and 10C7GArgMet groups were almost same as the corresponding adrenalectomized groups. The body weight of the adrenalectomized 10C7G group was extremely decreased though that of the intact 10C7G group was maintained almost constant; but the decrease was recovered by the administration of hydrocortisone. The activities of liver arginase and carbamylphosphate synthetase were not affected by those diets. Liver serine dehydratase and ornithine δ-aminotransferase activities were increased in the intact 10C7G and 10C7GArgMet groups, but these increases were depressed by adrenalectomy. Glutamate-pyruvate transminase activities in the liver of intact 10C7G and 10C7GArgMet groups were also enhanced, but were extremely decreased in the corresponding adrenalectomized groups. Kidney transamidinase activity was not affected by adrenalectomy. The amount of urinary excreted urea was almost unchanged by adrenalectomy, but was increased by hydrocortisone administration. The amounts of excreted creatine of the adrenalectomized groups were generally larger than the corresponding intact groups, but slightly decreased by the administration of hydrocortisone. The amount of excreted creatinine was not generally affected by adrenalectomy.     

Glycine-U-14C Metabolism in Young Rats Fed the 10% Casein Diets Containing Excess Glycine

Nine hours after rats fed ad libitum for 14 days a 10% caein diet (10C), a 10% casein diet containing 7% glycine (10C7G) and a 10% casein diet containing 7% glycine with 1.4% l-arginine HCI and 0.9% l-methionine (10C7GArgMet) were force-fed 10 ml of each diet suspension containing 5 μCi of glycine-U-¹⁴C per 100 g of body weight, the radioactivity recoveries of ¹⁴C in expired CO₂, tissue components and urine were determined.

The radioactivity recovery of ¹⁴C in the expired C0₂ of the 10C7G group was generally higher than that of the 10C7GArgMet group, and those of both groups would have been much higher than that of the 10C group unless the isotope had been diluted. The amount of expiratory ¹⁴C of rats fed a 25 % casein diet containing 7% glycine was not different from that of the 10C7G group. The recovery of ¹⁴C in the trichloroacetic acid (TCA) soluble fraction of muscle of the 10C7G and the 10C7GArgMet groups were greater than that of the 10C group, but there was no difference between the 10C7G and the 10C7GArgMet groups. The recoveries of ¹⁴C in the TCA soluble fraction and protein of plasma and liver, and the muscle protein were negligible in all the groups. The amount of glycine-¹⁴C incorporated into the carcass lipids of the 10C7GArgMet group was larger than that of other groups. Those in the carcass lipids of the 10C7G and the 10C7GArgMet groups would have been much higher than that of the 10C group unless the dilution of the isotope had taken place. The recoveries of ¹⁴C in the liver and muscle glycogen, and liver lipids were remarkably small in all the groups. From the above results, it was suggested that the degradation of glycine-¹⁴C to expiratory CO₂ was not accelerated, but the rate of incorporation of the isotope into carcass lipids was increased by the supplementation of l-arginine and l-methionine to the 10C7G diet as compared with that of rats fed the 10C7G diet.     

Studies on Glyoxalase Inhibitor Isolation of a New Active Agent,MS-3, from a Mushroom Culture

Cultured broths were screened by measuring the inhibition of glyoxalase, using 105,000 g supernatant of rat liver homogenates as a crude enzyme. An active agent, MS–3 (C₂₁H₂₄O₇), was isolated from a cultured mushroom. ID₅₀ value of MS–3 against the crude glyoxalase was 12 mcg/ml. MS–3 has low toxicity and inhibited the growth of Yoshida rat sarcoma cells in cell culture. The production, isolation, and properties of MS–3 are described.     

The Reaction Mechanism of Rat Liver Glyoxalase I and Its Inhibition by MS-3

Glyoxalase I from rat liver was purified about 25-fold by acetone fractionation and ion-exchange chromatography on CM-Sephadex and DEAE-cellulose columns. The kinetic study of the enzymatic reaction supported the one-substrate mechanism : the hemimercaptal adduct produced nonenzymatically from methylglyoxal and glutathione is the substrate. The Km value determined was 0.1 mm and similar to that of porcine erythrocytes enzyme but differed significantly from that of yeast enzyme. It was inhibited by free glutathione competitively (Ki 1.2 mm). Kinetic studies on inhibition of glyoxalase I by MS–3 which was obtained from a cultured mushroom, Stereum hirsutum, indicated the inhibition type was competitive with the hemimercaptal adduct (Ki 4.6 × 10^?6 m). By the graphical study of the multiple inhibition kinetics free glutathione and MS–3 were shown to bind at the same sites of the enzyme.     

Lactobacillus helveticus SBT2171 alleviates allergic symptoms in a murine model for pollen allergy

ABSTRACT

Lactic acid bacteria are known to have various health-promoting effects and are highly expected to find applications in anti-allergic food materials. In this study, we focused on Lactobacillus helveticus SBT2171 (LH2171), which reportedly modifies some unique immune responses and ameliorated symptoms of patients allergic to mites and house dust in the previous studies. We examined the effect of LH2171 on cytokine production by antigen-stimulated murine naïve splenocytes in vitro and demonstrated that it inhibited IL-4 and IL-13 production while enhancing IFN-γ and IL-10 production. Then, we examined the anti-allergic effect of LH2171 in vivo using a murine model of pollen allergy and found that LH2171 reduced the sneezing frequency when orally administered to mice. We successfully confirmed the immune modulatory activity of LH2171 and its anti-allergic activity against inhaled antigens. These evidences would contribute to identifying the anti-allergic mechanism of LH2171.

Abbreviations: ALDH: aldehyde dehydrogenase; EGCG: epigallocatechin gallate; LAB: lactic acid bacteria; LH2171: Lactobacillus helveticus SBT2171; NALT: nasal-associated lymphoid tissue; OVA: ovalbumin     

Transmission ratio distortion of molecular markers in a doubled haploid population originated from a natural hybrid between Osmunda japonica and O. lancea

In ferns, intra-gametophytic selfing occurs as a mode of reproduction where two gametes from the same gametophyte form a completely homozygous sporophyte. Intra-gametophytic selfing is considered to be prevented by lethal or deleterious recessive genes in several diploid species. In order to investigate the modes and tempo of selection acting different developmental stages, doubled haploids obtained from intra-gametophytic selfing within isolated gametophytes of a putative F1 hybrid between Osmunda japonica and O. lancea were analyzed with EST_derived molecular markers, and the distribution pattern of transmission ratio distortion (TRD) along linkage map was clarified. As the results, the markers with skewness were clustered in two linkage groups. For the two highly distorted regions, gametophytes and F2 population were also examined. The markers skewed towards O. japonica on a linkage group (LG_2) showed skewness also in gametophytes, and the TRD was generated in the process of spore formation or growth of gametophytes. Also, selection appeared to be operating in the gametophytic stage. The markers on other linkage group (LG_11) showed highest skewness towards O. lancea in doubled haploids, and it was suggested that the segregation of LG_11 were influenced by zygotic lethality or genotypic evaluation and that some deleterious recessive genes exist in LG_11 and reduce the viability of homozygotes with O. japonica alleles. It is very likely that a region of LG_11were responsible for the low frequencies of intra-gametophytic selfing in O. japonica.