In vitro reduction of manganese dioxide by a ferrireductase system from the white-rot fungus <Emphasis Type="Italic">Phanerochaete sordida</Emphasis> YK-624

Reduction of manganese dioxide is demonstrated for an in vitro ferrireductase system that includes NADPH-dependent ferrireductase and the iron-binding compound (IBC) isolated from the white-rot fungus Phanerochaete sordida YK-624. The Fe(II)–IBC complex was more effective in reducing manganese dioxide to Mn(II) than were complexes of Fe(II) and organic acids of low molecular weight such as nitrilotriacetate, although IBC also reduced manganese dioxide to Mn(II) in the absence of Fe(II). The generated Fe(III)–IBC complex was a better substrate for NADPH-dependent ferrireductase than were other ferric chelates, suggesting that the Fe(III)–IBC complex is reduced to an Fe(II) complex by NADPH-dependent ferrireductase. Moreover, production of the Fe(III)–IBC complex by the reduction of manganese dioxide in a reaction system containing Fe(II) and IBC was observed to be coupled to reduction of the Fe(III)–IBC complex by NADPH-dependent ferrireductase. These results indicate that the ferrireductase system of P. sordida YK-624 plays an important role in the reduction of manganese dioxide, which is necessary for the production and function of manganese peroxidase.     

Volatile organic compounds from wood and their influences on museum artifact materials I. Differences in wood species and analyses of causal substances of deterioration

Wood is generally used as the interior material in museum storage rooms. Recently, however, the effects of volatile organic compounds (VOCs) from wood on artifacts has become a topic of great concern. The VOCs from four species of wood (western red cedar, spruce, kiri, and sugi) and their effects on artifact materials (two types of metal, seven types of pigment) were investigated using a deterioration-accelerating test, gas chromatography-mass spectrometry, X-ray diffraction analysis, and X-ray photoelectron spectroscopy. The results suggested that the influences on artifact materials varied greatly with wood species, and depended on specific components such as hinokitiol or acetic acid rather than the amount of total volatile organic compounds (TVOCs). It is a very serious problem that of the four species of wood, western red cedar (rich in hinokitiol), which has been recommended as an interior material for museum storage rooms, showed the heaviest deterioration on metal samples, and only this type of wood discolored enpaku (white lead) and rokushou (malachite, verdigris). In such storage rooms, museum artifacts should be carefully monitored. When selecting an interior material for a storage room or studying methods of preventing deterioration, it is very important to consider fully the characteristics of wood VOCs, not only the amount of TVOC.Part of this paper was presented at the 24th (Tokyo, June 2002) and 25th (Kyoto, June 2003) Annual Meeting of the Japan Society for the Conservation of Cultural Property     

In vitro screening of angiotensin I-converting enzyme inhibitors from Japanese cedar (Cryptomeria japonica)

Screening and isolation of angiotensin I-converting enzyme (ACE) inhibitors from Japanese cedar (Cryptomeria japonica) based on the in vitro ACE inhibitory assay were attempted. The ethanol extract from outer bark showed the highest inhibitory activity (IC₅₀ is 16g/ml) among 24 extracts prepared from roots, leaves, heartwood, sapwood, inner bark, and outer bark by successive extraction with four solvents. The fractionation of the outer bark ethanol extract followed by the bioassay resulted in the isolation of two strong ACE inhibitors, catechin and dimeric procyanidin B3. The bioassay of three flavan-3-ols including (+)-catechin and six flavones revealed that most of these compounds have high ACE inhibitory activity. The results suggest that the phenolic hydroxyl group at the C7 position and heterocyclic oxygen atom of these compounds are important for expressing the inhibitory activity.     

Structural changes of residual lignin in softwood kraft pulp treated with manganese peroxidase

Structural changes of residual lignin in unbleached softwood kraft pulp (SWKP) during manganese peroxidase (MnP) treatment were investigated to obtain some understanding of the biobleaching action of SWKP with MnP treatment. Alkaline-extracted lignin from darkened SWKP by MnP showed more intense color and contained moreo-quinone than that from control SWKP. However, no difference in the conjugated-carbonyl was observed between the lignins from MnP-treated and control SWKP. The nitrobenzene oxidation analysis revealed that oxidative condensation of non-condensed lignin in SWKP occurs during an early stage of MnP treatment. These observations were supported by the model experiment in which the lignin prepared from control SWKP was subjected to MnP treatments three times, and the changes of color and functional groups in the lignin were determined after each treatment. These results suggested that an increase ino-quinone and the condensation reaction of non-condensed lignin in SWKP are responsible for the characteristic darkening of SWKP during MnP treatment. It was also ascertained that darkened lignin was degraded and brightened by repeated MnP treatments.     

Production of transgenic cloned pigs expressing the far-red fluorescent protein monomeric Plum

Monomeric Plum (Plum), a far-red fluorescent protein with photostability and photopermeability, is potentially suitable for in vivo imaging and detection of fluorescence in body tissues. The aim of this study was to generate transgenic cloned pigs exhibiting systemic expression of Plum using somatic cell nuclear transfer (SCNT) technology. Nuclear donor cells for SCNT were obtained by introducing a Plum-expression vector driven by a combination of the cytomegalovirus early enhancer and chicken beta-actin promoter into porcine fetal fibroblasts (PFFs). The cleavage and blastocyst formation rates of reconstructed SCNT embryos were 81.0% (34/42) and 78.6% (33/42), respectively. At 36–37 days of gestation, three fetuses systemically expressing Plum were obtained from one recipient to which 103 SCNT embryos were transferred (3/103, 2.9%). For generation of offspring expressing Plum, rejuvenated PFFs were established from one cloned fetus and used as nuclear donor cells. Four cloned offspring and one stillborn cloned offspring were produced from one recipient to which 117 SCNT embryos were transferred (5/117, 4.3%). All offspring exhibited high levels of Plum fluorescence in blood cells, such as lymphocytes, monocytes and granulocytes. In addition, the skin, heart, kidney, pancreas, liver and spleen also exhibited Plum expression. These observations demonstrated that transfer of the Plum gene did not interfere with the development of porcine SCNT embryos and resulted in the successful generation of transgenic cloned pigs that systemically expressed Plum. This is the first report of the generation and characterization of transgenic cloned pigs expressing the far-red fluorescent protein Plum.     

Polyethylene degradation by lignin-degrading fungi and manganese peroxidase

Degradation of high-molecular-weight polyethylene membrane by lignin-degrading fungi, IZU-154, Phanerochaete chrysosporium, and Trametes versicolor, was investigated under various nutritional conditions. IZU-154 showed the most significant polyethylene degradation among the three lignin-degrading fungi under nitrogen- or carbon-limited culture conditions. Furthermore, for T. versicolor and P. chrysosporium, the addition of Mn(II) into nitrogen- or carbon-limited culture medium enhanced polyethylene degradation. These results suggest that polyethylene degradation is related to ligninolytic activity of lignin-degrading fungi. Treatment of polyethylene membrane with partially purified manganese peroxidase (MnP) caused significant degradation in the presence of Tween 80, Mn(II), and Mn(III) chelator. This result demonstrates that MnP is the key enzyme in polyethylene degradation by lignin-degrading fungi.This study was presented in part at the 9th International Symposium on Wood and Pulping Chemistry, Montreal, Canada, June 9–12, 1997     

Characterization of Antioxidant Activity of Heated Mycosporine-like Amino Acids from Red Alga Dulse Palmaria palmata in Japan

We recently demonstrated the monthly variation and antioxidant activity of mycosporine-like amino acids (MAAs) from red alga dulse in Japan. The antioxidant activity of MAAs in acidic conditions was low compared to that in neutral and alkali conditions, but we found strong antioxidant activity from the heated crude MAA fraction in acidic conditions. In this study, we identified and characterized the key compounds involved in the antioxidant activity of this fraction. We first isolated two MAAs, palythine, and porphyra-334, from the fraction and evaluated the activities of the two MAAs when heated. MAAs possess absorption maxima at around 330 nm, while the heated MAAs lost this absorption. The heated MAAs showed a high ABTS radical scavenging activity at pH 5.8–8.0. We then determined the structure of heated palythine via ESI-MS and NMR analyses and speculated about the putative antioxidant mechanism. Finally, a suitable production condition of the heated compounds was determined at 120 °C for 30 min at pH 8.0. We revealed compounds from red algae with antioxidant activities at a wide range of pH values, and this information will be useful for the functional processing of food.     

Myricanol and myricanone biosynthesis in <Emphasis Type="Italic">Myrica rubra</Emphasis>: incorporation of two molecules of 4-coumaric acid

After feeding experiments of Myrica rubra young shoots with 4-[8,9-¹³C₂]coumaric acid, mass spectrometric analyses revealed that the cyclic diarylheptanoids, myricanol and myricanone, were derived from two molecules of 4-coumaric acid. ¹³C Nuclear magnetic resonance analysis of myricanol isolated after administration of 4-[8,9-¹³C₂]coumaric acid demonstrated that the C-8 and C-9 atoms of 4-coumaric acid are incorporated into C-8, C-9, C-11, and C-12 of the corresponding myricanol. Part of this report was presented at the 56th Annual Meeting of the Japan Wood Research Society, Akita, August 2006     

Population structure of the Japanese eel Anguilla japonica as examined by mitochondrial DNA sequencing

SUMMARY: To examine the population structure of the Japanese eel Anguilla japonica , mitochondrial DNA sequence analysis was made for various samples of glass eels that might reflect genetic characteristics of spawning aggregates. The mitochondrial DNA from a total of 51 glass eels collected at Tanegashima Island, Kanagawa and Ibaraki in different periods within an inshore migrating season was sequenced for a 615 base-pair fragment from the tRNA^Thr gene to the central part of the control region. The DNA region was so variable that no individual was found to possess the same sequence, although average sequence differences within samples (1.07–1.63) were not large. Average sequence differences between samples (1.22–1.57) were comparable to those within samples, suggesting no genetic heterogeneity among samples. Tree analysis of the sequences showed neither a geographical nor temporal structure of population. Furthermore, although all DNA sequences from the present study were different from one another, two sequences were found to be the same with those reported from individuals collected in different localities in different years. Altogether, the present mitochondrial DNA sequence analysis of glass eels did not provide any evidence for genetic subdivision of the Japanese eel population.