Excitation Energy-Transfer Dynamics of Brown Algal Photosynthetic Antennas.

Fucoxanthin-chlorophyll-a/c protein (FCP) complexes from brown algae Cladosiphon okamuranus TOKIDA (Okinawa Mozuku in Japanese) contain the only species of carbonyl carotenoid, fucoxanthin, which exhibits spectral characteristics attributed to an intramolecular charge-transfer (ICT) property that arises in polar environments due to the presence of the carbonyl group in its polyene backbone. Here, we investigated the role of the ICT property of fucoxanthin in ultrafast energy transfer to chlorophyll-a/c in brown algal photosynthesis using femtosecond pump-probe spectroscopic measurements. The observed excited-state dynamics show that the ICT character of fucoxanthin in FCP extends its absorption band to longer wavelengths and enhances its electronic interaction with chlorophyll-a molecules, leading to efficient energy transfer from fucoxanthin to chlorophyll-a.

Aflatoxins and ochratoxin A in tea prepared from naturally contaminated powdered ginger.

The migration of several major mycotoxins, aflatoxins B(1) (AFB(1)), B(2), G(1), and G(2) (AFT, total of the aflatoxins) and ochratoxin A (OTA), from naturally contaminated powdered ginger to surrounding liquid (tea) was investigated. The toxins are commonly found in cereal grains and are toxic, carcinogenic and thermostable. Ginger root is widely used for digestive problems. Powdered ginger (2 g) found to contain AFT and OTA was placed in an empty heat sealable tea bag. The tea bag was heat-sealed and used to prepare tea under different conditions: temperature (50 and 100 degrees C), time (5 and 10 min) and volume (100 and 200 ml). The tea bag was placed in hot water and stirred every 1 min for 5 s during the first 5 min of steeping. After steeping, the tea bag was removed and the tea and ginger residue (in the tea bag) were analysed separately for AFT and OTA. After extraction and immunoaffinity column (IAC) clean-up, the isolated AFT and OTA were separated by reversed-phase liquid chromatography and quantified using a fluorescence detector. At 100 degrees C, approximately 30-40% of AFB(1) and AFT and 20-30% of OTA in the contaminated ginger were found in the ginger tea; the total amounts of AFT and OTA in tea varied less than 5% under the three conditions of preparation. At 50 degrees C, about 10% of OTA and AFT were found in tea. This is the first study on the migration of AFT from botanicals to tea. It is also the first to study the distribution of AFT and OTA from powdered ginger to tea and ginger residue.

Effect of processing on ochratoxin A content in dried beans.

Dried pink beans naturally contaminated with ochratoxin A (OTA) and dried carioca beans artificially contaminated with OTA by inoculation with Aspergillus ochraceus (ATCC 22947) were tested for ochratoxin A levels as follows: dried beans were washed with water for 2, 60 or 120 min, soaked in water for 60, 120 min or 10 h, and cooked for 60 or 120 min. At each step, test water and beans were separated. Test water, raw beans and cooked beans were analyzed for OTA. The amount of OTA partitioned into water and in residual beans was determined by methanol-sodium bicarbonate extraction, buffer dilution, immunoaffinity column cleanup, liquid chromatographic separation and fluorescence detection. The results demonstrated that the distribution of OTA in processing water and beans depends on the method of preparation. All treatments (washing, soaking and cooking) when applied individually reduced the amounts of OTA retained in bean flour and whole beans. Higher amounts of OTA remained in whole beans than in bean flour after removing the processing water. The combination of the three treatments eliminated about 50% of the toxin from whole beans. This study provides evidence that discarding the washing, soaking and cooking water leads to a significant reduction in OTA contamination in dried beans.

Intracellular changes of metal elements by fucoidan extracted from brown seaweed (Cladosiphon okamuranus).

This study was undertaken to elucidate the intracellular changes of metal elements after the administration of fucoidan extracted from Cladosiphon okamuranus. TRL1215 cells (normal rat liver cell line) were treated with 0, 0.1, or 1.0 mg/ml fucoidan and incubated in 5% CO2 at 37 degrees C. The cellular levels of Mg, Al, Fe, and Zn were significantly increased in the 1.0 mg/ml fucoidan-treated cells compared to those of the 0.1 mg/ml fucoidan-treated cells and the control. Next, TRL1215 cells were cultured on Mylar film overnight. At 24 h after 5-bromo-2'-deoxyuridine dosing, 0, 0.1, or 1.0 mg/ml fucoidan was treated for 9 h. The cellular distribution of elements was analyzed using in-air micro-micro-particle induced X-ray emission. The X-ray spectra showed that yields of Al, Mg, and Zn were high in order of the 1.0 mg/ml fucoidan-treated sample, the 0.1 mg/ml fucoidan-treated sample, and the control. Fe yield was mildly increased by fucoidan administration. In fucoidan-treated cells, the focal accumulation of Br was correlated spatially with phosphorous-rich region, suggesting that Br was localized within the nucleus. Al distribution provided a spatial association with Br map. These data suggest that fucoidan increases the accumulations of Al, Mg, Fe, and Zn in normal rat hepatocytes, and fucoidan-binding Al is postulated to be transferred into the nucleus.

Adsorption of di-2-pyridyl ketone salicyloylhydrazone on Amberlite XAD-2 and XAD-7 resins: characteristics and isotherms.

The adsorption of DPKSH onto Amberlite XAD-2 (styrene resin) and XAD-7 (acrylic ester resin) has been investigated, at (25+/-1) degrees C and pH 4.7. The experimental equilibrium data were fitted to the Langmuir, Freundlich, and Dubinin-Radushkevich (D-R) models. These three models provide a very good fit for both resins and the respective constants KL, KF, and KDR were calculated. For the same DPKSH concentration interval, the minimum time of contact for adsorption maximum at XAD-7 was smaller than at XAD-2 and the maximum amount of DPKSH adsorbed per gram of XAD-2 is smaller than at XAD-7. The investigation indicates that the mean sorption energy (E) characterizes a physical adsorption and the surfaces of both resins are energetically heterogeneous. The constants obtained in these studied systems were correlated and compared with those obtained for the silica gel/DPKSH system.

Kinetic modeling of adsorption of di-2-pyridylketone salicyloylhydrazone on silica gel.

The kinetics of DPKSH (di-2-pyridylketone salicyloylhydrazone) adsorption onto silica gel has been investigated at (25+/-1) degrees C and pH 1 and 4.7. The kinetics of adsorption of DPKSH is discussed using three kinetic models, the first-order Lagergren model, the pseudo-second-order model, and the intraparticle diffusion model. The adsorption of DPKSH, at pH 1 and 4.7, onto silica gel proceeds according to the pseudo-second-order model and the correlation coefficients were very close to 1. The intraparticle diffusion of DPKSH molecules within the silica gel particles was identified as the rate-limiting step. The parameters of the pseudo-second-order model are q(max,calc)=1.02 x 10(-4) and 1.5 x 10(-4) g DPKSH/g silica; k(2)=3.01 x 10(4) and 9.67 x 10(4) h(-1)g silica/g DPKSH, respectively, for pH 1 and 4.7.

Adsorption of di-2-pyridyl ketone salicyloylhydrazone on silica gel: characteristics and isotherms.

The adsorption of DPKSH onto silica gel was investigated, at 25+/-1 degrees C and pH 1, 4.7 and 12. For the same DPKSH concentration interval, the minimum required time of contact for adsorption maximum at pH 4.7 was smaller than at pH 1 and the maximum amount of DPKSH adsorbed per gram of silica at pH 1 is smaller than at pH 4.7. At pH 12 the DPKSH adsorption onto silica gel was not significant. The adsorption data followed Langmuir, Freundlich and Dubinin-Radushkevich (D-R) isotherms. The maximum amount of solute adsorbed (m(ads)(max)) and the adsorption constant, K(L), were derived from Langmuir isotherm. The Freundlich constants 1/n and K(F) related, respectively, to the energetic heterogeneity of adsorption sites and an empirical constant were evaluated. The mean sorption free energy (E) of DPKSH adsorption onto silica gel was calculated from D-R isotherm indicating a physical adsorption mode. Finally, conductimetric titrations showed the silica particle basicity and acidity as 0.002 and 0.3mmolg(-1), respectively.

Occurrence of aflatoxins M(1) and M(2) in milk commercialized in Ribeirão Preto-SP, Brazil.

Aflatoxins are toxic metabolites found in foods and feeds. When ruminants eat foodstuffs containing aflatoxins B(1) and B(2), these toxins are metabolized and excreted as aflatoxin M(1) and M(2) in milk. The aim was to determine the incidence of these aflatoxins in commercial milk collected from supermarkets in Ribeirão Preto-SP, Brazil, and consisting of 60 ultrahigh temperature (UHT) milk samples and 79 pasteurized milk samples. The milk samples were analysed according to method 986.16 of AOAC International. None of the milk samples analysed were contaminated with aflatoxin M(2), and aflatoxin M(1) was detected in 29 (20.9%) of samples in the range 50-240 ng l(-1). The results show that despite a high occurrence of aflatoxin M(1) in commercial pasteurized and UHT milk sold in Ribeirão Preto in 1999 and 2000, the contamination level of these toxins could not be considered a serious public health problem according to MERCOSUR Technical Regulations. However, levels in 20.9% of the milk samples exceeded the concentration of 50 ng l(-1) permitted by the European Union. Although it is not necessary to continue monitoring the incidence and levels of aflatoxins M(1) and M(2) in milk samples, surveillance could be appropriate.

Properties of fucoidan from Cladosiphon okamuranus tokida in gastric mucosal protection.

To elucidate the anti-ulcer potential of Cladosiphon fucoidan, anti-peptic activity, bFGF stabilizing activity and inflammatory properties of this and related substances were investigated. Anti-peptic activity was observed with this and other sulfated polysaccharides such as dextran sulfate, carrageenan, and Fucus fucoidan. However, non-sulfated polysaccharides such as mannan and dextran did not exert the anti-peptic activity. The loss of bFGF bioactivity was prevented by all sulfated polysaccharides tested except chondroitin sulfate, at pH 7.4 and at pH 4.0. At pH 2.0, only heparin protected the bFGF activity. The generation of superoxide by macrophages and PMNs was stimulated by dextran sulfate, carrageenan, and Fucus fucoidan, whereas Cladosiphon fucoidan, heparin and chondroitin did not. Dextran sulfate, carrageenan, and Fucus fucoidan also stimulated the secretion of TNFalpha from macrophages, while Cladosiphon fucoidan did not. Thus, Cladosiphon fucoidan is a sulfated polysaccharide without inflammatory action. These results suggest that Cladosiphon fucoidan is a safe substance with potential for gastric protection.

HLA genes and haplotypes in Ryukyuans suggest recent gene flow to the Okinawa Islands.

Polymorphism of HLA genes was investigated in a population sample of Ryukyuans living on the main island of Okinawa (n = 197), in the southwestern islands of Japan. Serological typing was applied to class I loci (HLA-A, -B, and -C) and to HLA-DRB1; nucleotide sequence-level typing was performed using PCR microtiter plate hybridization and PCR single-strand conformation polymorphism methods. Ryukyuans showed a higher frequency of DRB1*0405 and lower frequencies of DRB1*1502 and DRB1*1302 compared with Hondo Japanese living on main islands. Principal components and phylogenetic analyses of 12 East Asian populations, including Ryukyuans, were performed based on the allele frequencies of HLA-A, -B, and -DRB1. In the principal components analysis 3 Japanese populations (Ryukyuans, Hondo Japanese, and Ainu) formed a cluster and showed the highest affinity to 2 Korean populations. In the phylogenetic tree Ryukyuans and Ainu were neighbors, but the genetic distance between them was larger than the distances between Ryukyuans and Hondo Japanese and between Ryukyuans and Korean populations. The geographic cline of the predominant haplotype in Ryukyuans, A*24-B*54-DRB1*0405, suggests that an ancestral population possessing A*24-B*54-DRB1*0405 moved into the Okinawa Islands after the divergence of Ryukyuans from the Ainu. Such a recent gene flow, probably from South China to the Okinawa Islands, is considered the major cause of difference in genetic characteristics between Ryukyuans and the Ainu.

Study of complex formation in the manganese(II)/azide system.

The complex formation between Mn(II) cations and N(3)(-) anions was studied in aqueous medium at 25 degrees C and ionic strength 2.0 M (NaClO(4)). Data of average ligand number, n (Bjerrum's function), were obtained from pH measurements on the Mn(II)/N(3)(-)/HN(3) system followed by integration to obtain Leden's function, F(0)(L). Graphical treatment of data and a matrix solution of simultaneous equations have given the following overall formation constants of mononuclear stepwise complexes: beta(1)=4.15+/-0.02 M(-1), beta(2)=6.61+/-0.04 M(-2), beta(3)=3.33+/-0.02 M(-3), beta(4)=0.63+/-0.01 M(-4). A linear plot of log K(n) vs. (n-1) shows no change in the configuration during complex formation. Slow spontaneous oxidation of solutions to Mn(III) occurs when the N(3)(-) concentration is greater than 1.0 M.

The primary structure of a hemorrhagic factor, HR2b, from the venom of Okinawa habu (Trimeresurus flavoviridis).

The complete amino acid sequence of a hemorrhagic factor, HR2b, from the venom of Okinawa habu was determined. The hemorrhagic factor was fragmented by CNBr cleavage, trypsin, staphylococcal protease V8 and lysyl endopeptidase digestions. The resulting peptides were purified on high performance chromatography, and sequenced by Edman degradation. HR2b was composed of 204 amino acids with pyroglutamyl residue at the amino terminus, and the calculated mol. wt based on the amino acid composition was 23,335. There are three disulfide linkages in the primary structure. The consensus sequence (His-Glu-Xaa-Xaa-His) for zinc-binding site of zinc-requiring metalloproteinases was found in the structure. The primary structure of HR2b shows a significant similarity with that of HR2a of Amami habu venom; 98.5% identity.

Helicobacter pylori infection and chronic atrophic gastritis among Japanese blood donors: a cross-sectional study.

To evaluate an association between Helicobacter pylori (H. pylori) infection and chronic atrophic gastritis (CAG), an established precursor of gastric cancer, we performed a cross-sectional study using IgG antibody against H. pylori and pepsinogens of blood donors in four prefectures in Japan. Although a geographic correlation between the age-adjusted prevalence rates for H. pylori infection and those for CAG was not seen, the age-adjusted odds ratios (OR) of H. pylori infection for CAG were high in each area (around five for men and from four to 12.6 for women). The association between them weakened with advancing age; the ORs in the youngest age group (16-29 yrs) and in the oldest age group (50-64 yrs) were 12.5 and 2.8 for men, and 11.5 and 5.2 for women, respectively. These findings suggest that H. pylori infection is strongly associated with CAG, while there are some other factors interacting in the development of CAG. A prospective cohort study in which CAG and H. pylori infection are taken into account will be necessary to assess the risks of gastric cancer.

Correlation between the prevalence of gastritis and gastric cancer in Japan.

To verify the hypothesis that chronic atrophic gastritis (CAG) is a precancerous lesion for gastric cancer, we investigated the correlation between the prevalence rates for CAG determined by levels of serum pepsinogens (I and II) of randomly selected, healthy, blood donors and the mortality rates for gastric cancer among four prefectures (Akita, Iwate, Miyagi, and Okinawa) in Japan. The prevalence rates for CAG according to a criterion that the level of pepsinogen I is below 70 ng/ml and the pepsinogen I to pepsinogen II ratio is below 3.0, were the highest in Akita for both sexes (38.8 percent in males, 37.8 percent in females) and the lowest in Okinawa (13.9 percent in males, 12.3 percent in females), where mortality rates for gastric cancer are the highest and the lowest in Japan, respectively. The correlation between the prevalence rates for CAG and the standardized mortality ratios for gastric cancer among these four prefectures was statistically significant in males (r = 0.97, P = 0.03), but less significant in females (r = 0.92, P = 0.08). These data strongly support the hypothesis that CAG is a precancerous lesion. The limitations of our cross-sectional study and the advantages of measuring the levels of serum pepsinogens in epidemiologic studies and in mass-screening programs for gastric cancer are discussed.

Characterization of three hemorrhagic factors from the venom of Okinawa habu (Trimeresurus flavoviridis).

Three hemorrhagic factors, HR1, HR2a and HR2b, of Okinawa habu venom were characterized in terms of their subunit structure, amino acid composition, metal content and immunological properties. HR1 is a dimer (mol. wt 90,000) consisting of two identical subunits at 25 degrees C, but polymerizes to form a tetramer at 4 degrees C. Two peaks corresponding to the dimer and the tetramer were observed upon ultracentrifugation analysis at 20 degrees C. HR2a and HR2b are monomers (mol. wt 24,000 and 19,000, respectively). HR1, HR2a and HR2b contain 407, 203 and 161 amino acids, respectively and the respective mol. wt based on the amino acid composition are 45,988, 23,075 and 18,457. The hemorrhagic factors contain Zn2+, Ca2+ and Mg2+, and were irreversibly inhibited by incubation with chelating reagents. The three hemorrhagic factors were immunologically distinguished from each other, and the hemorrhagic activities were inhibited by the respective antiserum. The activity of HR2a was also inhibited by the antiserum against HR2b.

Orosomucoid (ORM) typing by isoelectric focusing: an analysis of ORM haplotypes.

A new separator isoelectric focusing method for typing of orosomucoid (ORM) was developed. This method provided a superior resolution of ORM patterns: two close bands of ORM1*5.2 products were clearly separated. A total of 364 subjects from Okinawa (Japan) were classified into 21 ORM phenotypes determined by 6 ORM1 and 7 ORM2 alleles including a polymorphic silent allele, ORM2*QO, and 2 new rare variants, ORM2*18 and ORM2*19. These phenotypes were also explained by 12 ORM haplotypes, half of which were polymorphic.

Purification and crystallization of hemorrhagic factor, HR2b, from the venom of Trimeresurus flavoviridis (habu).

The hemorrhagic factor, HR2b, was purified from the venom of Trimeresurus flavoviridis (habu) by a combination of gel filtration, cation exchange column chromatography and high performance liquid chromatography. The purified HR2b was homogeneous by the criteria of ultracentrifugation and SDS-disc electrophoresis. The mol. wt of HR2b was 18,000 and 18,500 by gel filtration on Sephadex G-50 and by SDS-disc electrophoresis, respectively, indicating a monomer structure for the hemorrhagic factor. Crystals of HR2b, taking the form of thin plates, were obtained in the presence of ammonium sulfate.

Factor I (C3b inactivator) polymorphism among five populations in Eurasia.

Factor I (C3b inactivator) polymorphism in the Japanese (in Western and Southern Japan), Taiwanese, Nepalese and French was studied using isoelectric focusing on polyacrylamide gels. The exposure of passively blotted nitrocellulose membranes to glutaraldehyde vapor facilitated the subsequent immunodetection of a low concentration of factor I and permitted the reliable identification of the three phenotypes determined by two codominant alleles FI*A and FI*B. The data indicated a west-to-east genocline, ranging from France to Western Japan, in which FI*A changed from 0.006 to 0.120.