Bitterness of Phenylalanine- and Tyrosine-containing Peptides

In order to investigate the role of phenylalanine and tyrosine residues in the bitter taste of peptides, some oligopeptides containing phenylalanine or tyrosine were synthesized and their taste was evaluated. The hydrophobicity of the phenylalanine or tyrosine molecule markedly caused the bitter taste in peptide. The bitterness was more intense when phenylalanine was located at the C- terminus and when the content of phenylalanine or tyrosine was increased in peptides. The hydrophobic residue in peptides functioned as a bitter taste determinant site. The experimental results suggest the existence of an additional site for the bitter taste of peptides.     

Epidemiological and Ecological Studies of Japanese Encephalitis in Okinawa,Subtropical Area in Japan. I. Investigations on Antibody Levels to Japanese Encephalitis Virus in Swine Sera and Vector Mosquito in Okinawa,Miyako and Ishigaki Islands

From 1985 to 1989, serum specimens of swine raised in the northern, central and southern areas in Okinawa island were examined for Japanese encephalitis (JE) virus antibody by ELISA and hemagglutination-inhibition test. The antibody positive rate was found to be higher in the north and central than in the south. The 2-mercaptoethanol sensitive antibody to JE was detected mostly in June and July, and occasionally in other months except February and March. There was no month when all specimens from three areas turned antibody-negative simultaneously, indicating that JE virus transmission to swine lasted longer in Okinawa island than in other temperate areas in Japan. From 1986 to 1991, the vector mosquitoes (Culex tritaeniorhynchus) were collected in a pig farm in the south of Okinawa island. A total of 153 strains of JE virus was isolated from the vector mosquitoes mainly in June. In Miyako and Ishigaki islands, the antibody positive rate in swine sera was found to be extremely low, compared with that in Okinawa island. In Miyako island, where no paddy rice field is cultivated, a few adults as well as larvae of the vector mosquito were collected, while in Ishigaki island, where there are many watered rice fields, a lot of adults as well as larvae were collected. Although the environmental situation is quite different between the two islands, JE virus transmission appeared to be very low in both islands.     

Assessment of Poliovirus Eradication in Japan: Genomic Analysis of Polioviruses Isolated from River Water and Sewage in Toyama Prefecture

Seventy-eight poliovirus strains isolated from river water and sewage in Toyama Prefecture, Japan, during 1993 to 1995 were characterized by the PCR-restriction fragment length polymorphism (RFLP) method and by partially sequencing the VP3 and VP1 regions of the viral genome. Of these isolates, 36 were identified as Sabin vaccine strains, and 42 were identified as vaccine variant strains that had less than 1.4% nucleotide divergence from the Sabin strains, including 7 isolates with patterns different from those of Sabin strains as determined by PCR-RFLP analysis. These findings suggest that wild-type poliovirus was not circulating in Toyama Prefecture.     

Cenomanian-Turonian carbon isotope events recorded in terrestrial organic matter from northern Japan

Carbon isotopic fluctuations recorded in terrestrial organic matter from the Upper Cretaceous Cenomanian-Turonian succession of Hokkaido, Japan show a remarkably similar pattern to isotopic fluctuation in carbonate carbon reported from southern England and the Italian Apennines. Chronological correlation for the Cenomanian-Turonian section of Japan is based on planktonic foraminiferal biostratigraphy with some additional data from macrofossils, allowing a detailed comparison of isotope stratigraphies for Japan and Europe. Three distinct features of the isotope profile, namely a positive spike near the Cenomanian/Turonian boundary, a shoulder in the lower-middle Turonian and a level segment in the middle-upper Turonian are observed in Japan. These features are biostratigraphically comparable and correlative with isotope profiles reported from Europe. The single factor influencing isotopic composition of both terrestrial organic carbon and marine carbonate carbon is isotopic change in the global CO₂ reservoir of the ocean-atmosphere system. If these three carbon isotope features are synchronous, they may be inferred as having been caused by global phenomena and as providing global high-resolution chemo-stratigraphic markers. Carbon isotope stratigraphy could be an important bridge between terrestrial inland sequences and marine sequences.     

Effect of a newly developed ketolide antibiotic, telithromycin, on metabolism of theophylline and expression of cytochrome P450 in rats

The effects of a newly-developed ketolide antibiotic, telithromycin, on the metabolism of theophylline and the expression of hepatic cytochrome P450 (CYP) 1A2 and CYP3A2 were investigated in rats. Telithromycin at a high dose (100 mg/kg of body weight) was injected intraperitoneally once a day for 3 days. Twenty-four hours (day 4) after the final administration of telithromycin, theophylline (10 mg/kg) was administered intravenously. The presence of telithromycin significantly delayed the disappearance of theophylline from plasma. Parameters related to the pharmacokinetic interaction between theophylline and telithromycin were examined by noncompartmental methods. A significant decrease in the systemic clearance of theophylline was observed in the presence of telithromycin. Pretreatment with telithromycin significantly decreased the metabolic clearance of the major metabolites, 1-methyluric acid and 1,3-dimethyluric acid, with no change in the renal clearance of theophylline, suggesting that the decreased systemic clearance of theophylline by telithromycin is due to reduction of their metabolic clearance. Pretreatment with telithromycin significantly decreased the activity of 7-ethoxyresorufin O-deethylation and testosterone 6 beta-hydroxylation, suggesting that telithromycin decreases the activity of hepatic CYP1A2 and CYP3A2. Western blot analysis revealed that telithromycin significantly decreased the protein levels of CYP1A2 and CYP3A2 in the liver, which could explain the observed decreases in the systemic clearance of theophylline and metabolic clearance of 1-methyluric acid and 1,3-dimethyluric acid. The present study suggests that telithromycin at the dose used in this study alters the pharmacokinetics and metabolism of theophylline, due to reductions in the activity and expression of hepatic CYP1A2 and CYP3A2.     

A 3-hydroxy β-end group in xanthophylls is preferentially oxidized to a 3-oxo ε-end group in mammals

We previously found that mice fed lutein accumulated its oxidative metabolites (3′-hydroxy-ε,ε-caroten-3-one and ε,ε-carotene-3,3′-dione) as major carotenoids, suggesting that mammals can convert xanthophylls to keto-carotenoids by the oxidation of hydroxyl groups. Here we elucidated the metabolic activities of mouse liver for several xanthophylls. When lutein was incubated with liver postmitochondrial fraction in the presence of NAD⁺, (3′R,6′R)-3′-hydroxy-β,ε-caroten-3-one and (6RS,3′R,6′R)-3′-hydroxy-ε,ε-caroten-3-one were produced as major oxidation products. The former accumulated only at the early stage and was assumed to be an intermediate, followed by isomerization to the latter. The configuration at the C3′ and C6′ of the ε-end group in lutein was retained in the two oxidation products. These results indicate that the 3-hydroxy β-end group in lutein was preferentially oxidized to a 3-oxo ε-end group via a 3-oxo β-end group. Other xanthophylls such as β-cryptoxanthin and zeaxanthin, which have a 3-hydroxy β-end group, were also oxidized in the same manner as lutein. These keto-carotenoids, derived from dietary xanthophylls, were confirmed to be present in plasma of normal human subjects, and β,ε-caroten-3′-one was significantly increased by the ingestion of β-cryptoxanthin. Thus, humans as well as mice have oxidative activity to convert the 3-hydroxy β-end group of xanthophylls to a 3-oxo ε-end group.     

Determination of timiperone in rat plasma by high-performance liquid chromatography with electrochemical detection

We report a sensitive new method for the determination of timiperone in rat plasma by using high-performance liquid chromatography with electrochemical detection. The method involves extraction of plasma samples with heptane-isoamyl alcohol at pH>8, followed by back-extraction into dilute acetic acid. Separation was accomplished by reversed-phase high-performance liquid chromatography on an ODS column with the mobile phase consisting of 0.1 M phosphate buffer (pH 3.5)-acetonitrile-methanol (65:20:15, v/v). Recovery was greater than 80%. Calibration curve was linear over the concentration range 0.5–50.0 ng/ml. The limit of quantitation of timiperone was 0.5 ng/ml plasma.     

Drug-specific T cells derived from patients with drug-induced allergic hepatitis.

Drug-induced allergic hepatitis is a tissue-specific inflammatory disease caused by hypersensitivity to a particular drug. Although the frequency of drug-induced allergic hepatitis appears to increase in proportion to the medicine, the mechanism by which tissue specificity is determined is still to be elucidated. In this study, we established CD4+ T cell clones specific for particular drugs from patients with drug-induced allergic hepatitis accompanied with mild blood eosinophilia and analyzed the possible role of liver protein as a directing factor of liver-specific inflammatory reactions. All CD4+ T cell clones obtained from two patients with this disease proliferated in response to a combination of the particular drug plus liver specific protein (LSP), which consists of over 30 proteins. Some T cell clones were responsive to an antigenic conformation consisting of the 200-kDa glycoprotein (partly purified LSP), a component of LSP, plus the causal drug. In contrast, all CD4+ T cell clones from a patient with simple drug-induced eosinophilia responded to the causal drug in the absence of LSP and partly purified LSP. These data suggested that LSP or partly purified LSP of the appropriate Ag is the target that leads to liver-specific inflammation in drug-induced allergic hepatitis. Furthermore, T cell lines derived from patients with drug-induced allergic hepatitis and simple drug-induced eosinophilia produced large amounts of IL-5 after the appropriate antigenic stimulation, whereas CD4+ T cell clones from donors with a normal amount of peripheral blood eosinophils secreted a much less IL-5. Taken together, these results indicate that overproduction of IL-5 by the allergen-sensitized T cells may result in blood eosinophilia.     

Efficient synthesis of (R)-3-hydroxypentanenitrile in high enantiomeric excess by enzymatic reduction of 3-oxopentanenitrile

(R)-3-Hydroxypentanenitrile (HPN) is an important intermediate in the synthesis of an immunosuppressive inosine 5′-monophosphate dehydrogenase inhibitor. An efficient enzymatic procedure for the synthesis of (R)-HPN with over 99 % enantiomeric excess using a novel acetoacetyl-CoA reductase (AdKR) from Achromobacter denitrificans was successfully established. Many microorganisms are known to reduce 3-oxopentannitrile (KPN) to (R)-HPN. An enzyme from A. denitrificans partially purified using ion exchange chromatography reduced KPN to (R)-HPN with high enantioselectivity. The AdKR gene was cloned and sequenced and found to comprise 738 bp and encode a polypeptide of 26,399 Da. The deduced amino acid sequence showed a high degree of similarity to those of other putative acetoacetyl-CoA reductases and putative 3-ketoacyl-ACP reductases. The AdKR gene was singly expressed and coexpressed together with a glucose dehydrogenase (GDH) as a coenzyme regenerator in Escherichia coli under the control of the lac promoter. (R)-HPN was synthesized with over 99 % e.e. using a cell-free extract of recombinant E. coli cells coexpressing AdKR and GDH.