Association between brown sugar intake and decreased risk of cancer in the Amami islands region, Japan.

BACKGROUND AND OBJECTIVES: Although excess white sugar intake imposes various health burdens, brown sugar is high in minerals, polyphenols, and polycosanol. However, few epidemiological studies have assessed brown sugar intake for health benefit. People in the Amami islands region, with a relatively high proportion of individuals with longevity, consume brown sugar as a type of refreshment. This cohort study was conducted in Amami to clarify the association of brown sugar intake with mortality risk and cancer incidence. METHODS AND STUDY DESIGN: Participants were recruited from the general population of Amami as part of the Japan Multi-Institutional Collaborative Cohort Study. The number of eligible participants was 5004 (2057 men and 2947 women). During the median follow-up period of 13.4 years, 274 deaths and 338 cases of cancer were observed. HRs and 95% CIs were estimated using the Cox proportional hazard model, after adjusting for sugar-related and other variables. RESULTS: After adjusting for their related confounding factors, brown sugar intake was associated with decreased HRs and a decreasing trend for all-site and stomach cancer incidence (p = 0.001 and 0.017, respectively) in women and men, and for breast cancer incidence (p = 0.034) in women. Additionally, a decreasing trend in the HRs for lung cancer incidence was observed among never and ex-smokers (p = 0.039). Decreased HRs for overall death, cancer, and cardiovascular disease were not apparent. CONCLUSIONS: Brown sugar intake was associated with decreased risk of all-site, stomach, and breast cancer incidences in the Amami population.

Intake of local vegetables and decreased risk of mortality and cancer incidence in Amami island regions, Japan.

BACKGROUND AND OBJECTIVES: There is emerging scientific evidence of the health benefits of traditional food plants at both molecular and folk remedy levels; however, epidemiological observations are limited. The Amami island region of Japan has a variety of unique traditions conserved till today, where a cohort study was conducted in 2005. The objective of this study was to investigate the associations between the intake of common and local vegetables and the risk of mortality and cancer incidence in Amami. METHODS AND STUDY DESIGN: Participants were enrolled from the general population of Amami as part of the Japan Multi-institutional Collaborative Cohort (J-MICC) Study. In total, 5,015 participants (2,053 men and 2,962 women) aged 35-69 years were enrolled in this study. They were followed up to obtain information on movement, death, and cancer incidence. The hazard ratios (HRs) and 95% CIs were estimated using the Cox proportional hazard model after adjusting for potential confounding factors. RESULTS: A significant inverse association was observed between cabbage intake and the HRs for overall mortality (p for trend=0.046) and lung cancer incidence (p=0.016). Intake of handama and togan as local vegetables was associated with decreased HRs for overall mortality (p=0.019 and 0.036, respectively). CONCLUSIONS: While the molecular and biochemical reasoning and residual confounding factors behind this association remain unclear, the findings of this study suggest that the dietary lifestyle in Amami has a positive impact on the residents, which can significantly decrease mortality risk.

Characterization of the denitrifying bacterial community in a full-scale rockwool biofilter for compost waste-gas treatment.

The potential denitrification activity and the composition of the denitrifying bacterial community in a full-scale rockwool biofilter used for treating livestock manure composting emissions were analyzed. Packing material sampled from the rockwool biofilter was anoxically batch-incubated with 15N-labeled nitrate in the presence of different electron donors (compost extract, ammonium, hydrogen sulfide, propionate, and acetate), and responses were compared with those of activated sludge from a livestock wastewater treatment facility. Overnight batch-incubation showed that potential denitrification activity for the rockwool samples was higher with added compost extract than with other potential electron donors. The number of 16S rRNA and nosZ genes in the rockwool samples were in the range of 1.64-3.27 × 109 and 0.28-2.27 × 108 copies/g dry, respectively. Denaturing gradient gel electrophoresis analysis targeting nirK, nirS, and nosZ genes indicated that the distribution of nir genes was spread in a vertical direction and the distribution of nosZ genes was spread horizontally within the biofilter. The corresponding denitrifying enzymes were mainly related to those from Phyllobacteriaceae, Bradyrhizobiaceae, and Alcaligenaceae bacteria and to environmental clones retrieved from agricultural soil, activated sludge, freshwater environments, and guts of earthworms or other invertebrates. A nosZ gene fragment having 99% nucleotide sequence identity with that of Oligotropha carboxidovorans was also detected. Some nirK fragments were related to NirK from micro-aerobic environments. Thus, denitrification in this full-scale rockwool biofilter might be achieved by a consortium of denitrifying bacteria adapted to the intensely aerated ecosystem and utilizing mainly organic matter supplied by the livestock manure composting waste-gas stream.

Utilization of Bacillus sp. strain TAT105 as a biological additive to reduce ammonia emissions during composting of swine feces.

Bacillus sp. strain TAT105 is a thermophilic, ammonium-tolerant bacterium that grows assimilating ammonium nitrogen and reduces ammonia emission during composting of swine feces. To develop a practical use of TAT105, a dried solid culture of TAT105 (5.3 × 10(9) CFU/g of dry matter) was prepared as an additive. It could be stored for one year without significant reduction of TAT105. Laboratory-scale composting of swine feces was conducted by mixing the additive. When the additive, mixed with an equal weight of water one day before use, was added to obtain a TAT105 concentration of above 10(7) CFU/g of dry matter in the initial material, the ammonia concentration emitted was lower and nitrogen loss was approximately 22% lower in the treatment with the additive than in the control treatment without the additive. The colony formation on an agar medium containing high ammonium could be used for enumeration of TAT105 in the composted materials.

Effect of pH on phosphorus, copper, and zinc elution from swine wastewater activated sludge.

With the goal of reducing the amounts of phosphorus (P), copper (Cu), and zinc (Zn) discharged from swine wastewater activated sludge treatment facilities, we studied the elution of these elements from activated sludge at various pH values. Sludge samples with neutral pH collected from three farms were incubated at pH values ranging from 3 to 10. The soluble concentrations of these elements changed dramatically with pH and were highest at pH 3. We assumed that P present in the sludge under neutral and alkaline conditions was in insoluble form bound up with magnesium (Mg) and calcium (Ca), because Ca and Mg also eluted from the sludge at low pH. To clarify forms of Zn and Cu in the sludge, we performed a sequential extraction analysis. Zinc in adsorbed, organically bound, and sulfide fractions made up a large proportion of the total Zn. Copper in organically bound, carbonate, and sulfide fractions made up a large proportion of the total Cu. The soluble P concentrations were lowest at pH 9 or 10 (11-36 mg/L), the soluble Zn concentrations were lowest at pH 8 or 9 (0.07-0.15 mg/L), and the soluble Cu concentrations were lowest at pH 6-9 (0.2 mg/L, the detection limit).

Sulfation of chlorotyrosine and nitrotyrosine by human lung endothelial and epithelial cells: role of the human SULT1A3.

During inflammation, potent reactive oxidants formed may cause chlorination and nitration of both free and protein-bound tyrosine. In addition to serving as biomarkers of inflammation-mediated oxidative stress, elevated levels of chlorotyrosine and nitrotyrosine have been linked to the pathogenesis of lung and vascular disorders. The current study was designed to investigate whether the lung cells are equipped with mechanisms for counteracting these tyrosine derivatives. By metabolic labeling, chlorotyrosine O-[³5S]sulfate and nitrotyrosine O-[³5S]sulfate were found to be generated and released into the labeling media of human lung endothelial and epithelial cells labeled with [³5S]sulfate in the presence of added chlorotyrosine and nitrotyrosine. Enzymatic assays using the eleven known human cytosolic sulfotransferases (SULTs) revealed SULT1A3 as the enzyme responsible for catalyzing the sulfation of chlorotyrosine and nitrotyrosine. Reverse transcription-polymerase chain reaction (RT-PCR) analysis demonstrated the expression of SULT1A3 in the lung endothelial and epithelial cells used in this study. Kinetic constants of the sulfation of chlorotyrosine and nitrotyrosine by SULT1A3 were determined. Collectively, these results suggest that sulfation by SULT1A3 in lung endothelial and epithelial cells may play a role in the inactivation and/or disposal of excess chlorotyrosine and nitrotyrosine generated during inflammation.

Inhibitory effects of nitrative stress on the sulfation of 17ß-estradiol and 4-methoxyestradiol by human MCF 10A mammary epithelial cells.

Prolonged exposure to high level of estrogen is a known risk factor for breast carcinogenesis. It has been suggested recently that nitrative stress may be an etiologic factor for breast carcinogenesis. Since sulfation plays a major role in the homeostasis of estrogens and their metabolites, we attempted in the present study to find out whether nitrative stress may affect the homeostasis of estrogens through sulfation. Metabolic labeling experiments revealed that the amount of sulfated 17beta-estradiol or 4-methoxyestradiol decreased dramatically in MCF-10A mammary epithelial cells incubated in the presence of 3-morpholinosydnonimine (SIN-1) or diethylenetriamine NONOate (DETA NONOate), two nitric oxide donors commonly used to simulate nitrative stress conditions. In searching for the mechanism underlying the decrease of the sulfation of 17beta-estradiol and 4-methoxyestradiol, we demonstrated in an in vitro nitration experiment, that the human cytosolic sulfotransferase isoform 1E1 (SULT1E1), a major estrogen-sulfating enzyme, lost its estrogen-sulfating activity proportionately to the degree of nitration on tyrosine residues. Moreover, cell lysates prepared from MCF-10A cells treated with SIN-1 or DETA NONOate also showed much lower 4-methoxyestradiol-sulfating activities, compared with those determined with cell lysate prepared from control MCF-10A cells.

Tolerance of anammox reactor packed with zeolite to partial supply of nitrite or ammonium using purified livestock wastewater.

Anammox reaction requires nitrite and ammonium in the ratio of 1.1-1.3. However, controlling a partial nitrification process prior to the anammox process to maintain this ratio in an influent to the anammox reactor is not easy. In this study, the effect of zeolite on anammox reaction was investigated to determine a method of ammonium preservation in case of partial supply of nitrite or ammonium. Up-flow column type anammox reactors, filled with either zeolite or non-woven fabric, were operated in two-week intervals with purified livestock wastewater containing either ammonium or nitrite. The zeolite reactor showed significantly higher nitrogen removal rates than the non-woven fabric reactor for both influents. When the influent contained ammonium, it was adsorbed onto zeolite, while anammox tolerated starvation for two weeks. In a subsequent reaction cycle, when the influent contained nitrite, anammox used the nitrite and the ammonium desorbed from zeolite. The highest nitrogen removal rates were 0.71 and 0.29 gN/L/day, observed in the zeolite reactor, with the ammonium and nitrite influents, respectively. The limiting factor for reactor performance was zeolite saturation level when the influent contained ammonium and anammox reaction rate when the influent contained nitrite. This study demonstrated that zeolite can buffer the unbalance of the nitrite to ammonium ratio in an anammox reaction, and showed the scopes for improvement under each influent.

Effects of thiosulfate addition on ammonia and nitrogen removal in biofilters packed with Oyaishi (pumice tuff).

Ammonia removal is achieved partly by absorption and nitrification in biofilters, resulting in the accumulation of nitrogen and the necessity of treating the effluent water. We investigated the effects of thiosulfate addition to a biofilter containing pumice tuff for ammonia and nitrogen removal in a laboratory-scale experiment. The addition of thiosulfate to the circulating water led to a decreased nitrate and nitrite along with an increase of sulfate. The inorganic nitrogen in the circulating water decreased by up to 44% with thiosulfate addition compared to without thiosulfate. Batch experiments revealed that denitrification activity decreased exponentially along with increases in dissolved oxygen; however, approximately 30% of denitrification activity was maintained at dissolved oxygen concentration of 3.3 mg/L. Metabarcoding of 16S rRNA genes indicated that the genus Thiobacillus had a relative abundance of 0.002%-0.016% of total bacteria in the biofilter packing material. The circulating water pH was decreased below 5 with sulfur oxidation, and ammonium was accumulated without pH control resulting in a decrease in the relative abundance of the family Nitrosomonadaceae. Its relative abundance increased with control of pH to near neutral, indicating that ammonia-oxidizing activity could be maintained by adjusting pH. Thiosulfate addition could stimulate nitrogen removal by sulfur-dependent denitrification in biofiltration systems.

Glucose enhances collectrin protein expression in insulin-producing MIN6 beta cells.

Collectrin is a novel target gene of hepatocyte nuclear factor-1alpha in pancreatic beta-cells and controls insulin exocytosis. Although glucose is known to stimulate the expression of genes of the insulin secretory pathway, there is no information on how glucose regulates collectrin expression. We investigated the effects of glucose on the expression of collectrin in MIN6 beta-cell line. Glucose, in a dose-dependent manner, increased collectrin protein levels without changing collectrin mRNA levels and protein stability, indicating that glucose stimulation of collectrin protein expression is primarily mediated at a translational level. Although mannose and pyruvate also increased collectrin protein expression level, neither 2-deoxyglucose, mitochondrial fuels leucine and glutamate, sulphonylurea nor Ca(2+) channel blockers, mimicked the effects of glucose. These data indicate the involvement of mitochondrial TCA cycle intermediates, distal to pyruvate, in the regulation of collectrin protein expression in beta-cells.

Nitrogen removal by co-occurring methane oxidation, denitrification, aerobic ammonium oxidation, and anammox.

The pathway for removing NO(3)(-) and NH(4)(+) from wastewater in the presence of both CH(4) and O(2) was clarified by studying microbial activity and community. Batch incubation tests were performed to characterize the microbial activity of the sludge, which was acclimatized in a bioreactor in which O(2) and CH(4) were supplied to treat wastewater containing NO(3)(-) and NH(4)(+) . The tests showed that the sludge removed significant amounts of NO(3)(-) and NH(4)(+) in the presence of CH(4) and O(2), and the presence of the activity of methane oxidation, denitrification, nitrification, and anammox in the sludge. It was estimated that the total inorganic nitrogen removal was attributed to denitrification associated with methane oxidation as 53.4%, microbial assimilation as 37.9%, and anammox as 8.7%. Nitrification also contributed to NH(4)(+) decrease as 34.5% and anammox as 6.4%. Anammox activity was unambiguously demonstrated by (29)N(2) production in anaerobic batch incubation with (15)N-labeled inorganic nitrogen compounds. The presence of methane-oxidizing bacteria and candidate denitrifiers in the sludge was shown by denaturing gradient gel electrophoresis of 16S rRNA gene fragments. Clone library analysis of the PCR-amplified 16S rRNA gene fragment using specific primers for aerobic ammonium oxidizer and anammox revealed the presence of these bacteria. The results reveal that complex nitrogen-removal processes occur in the presence of CH(4) and O(2) by methanotroph, denitrifier, aerobic ammonium oxidizer, and anammox.

Characterization and ontogenic study of novel steroid-sulfating SULT3 sulfotransferases from zebrafish.

In vertebrates, sulfation as catalyzed by members of the cytosolic sulfotransferase (SULT) family has been suggested to be involved in the homeostasis of steroids. To establish the zebrafish as a model for investigating how sulfation functions to regulate steroid metabolism during the developmental process, we have embarked on the identification of steroid-sulfating SULTs in zebrafish. By searching the GenBank database, we identified two putative cytosolic SULT sequences from zebrafish, designated SULT3 ST1 and ST2. The recombinant proteins of these two zebrafish SULT3 STs were expressed in and purified from BL21 (DE3) cells transformed with the pGEX-2TK expression vector harboring SULT3 ST1 or ST2 cDNA. Upon enzymatic characterization, purified SULT3 ST1 displayed the strongest sulfating activity toward 17beta-estradiol among the endogenous substrates tested, while SULT3 ST2 exhibited substrate specificity toward hydroxysteroids, particularly dehydroepiandrosterone (DHEA). The pH-dependence and kinetic constants of these two enzymes with 17beta-estradiol and DHEA were determined. A developmental expression study revealed distinct patterns of the expression of SULT3 ST1 and ST2 during embryonic development and throughout the larval stage onto maturity. Collectively, these results imply that these two steroid-sulfating SULT3 STs may play differential roles in the metabolism and regulation of steroids during zebrafish development and in adulthood.

Identification and characterization of two novel cytosolic sulfotransferases, SULT1 ST7 and SULT1 ST8, from zebrafish.

Cytosolic sulfotransferases (SULTs) constitute a family of Phase II detoxification enzymes that are involved in the protection against potentially harmful xenobiotics as well as the regulation and homeostasis of endogenous compounds. Compared with humans and rodents, the zebrafish serves as an excellent model for studying the role of SULTs in the detoxification of environmental pollutants including environmental estrogens. By searching the expressed sequence tag database, two zebrafish cDNAs encoding putative SULTs were identified. Sequence analysis indicated that these two putative zebrafish SULTs belong to the SULT1 gene family. The recombinant form of these two novel zebrafish SULTs, designated SULT1 ST7 and SULT1 ST8, were expressed using the pGEX-2TK glutathione S-transferase (GST) gene fusion system and purified from transformed BL21 (DE3) cells. Purified GST-fusion protein form of SULT1 ST7 and SULT1 ST8 exhibited strong sulfating activities toward environmental estrogens, particularly hydroxylated polychlorinated biphenyls (PCBs), among various endogenous and xenobiotic compounds tested as substrates. pH-dependence experiments showed that SULT1 ST7 and SULT1 ST8 displayed pH optima at 6.5 and 8.0, respectively. Kinetic parameters of the two enzymes in catalyzing the sulfation of catechin and chlorogenic acid as well as 3-chloro-4-biphenylol were determined. Developmental expression experiments revealed distinct patterns of expression of SULT1 ST7 and SULT1 ST8 during embryonic development and throughout the larval stage onto maturity.

Treatment of swine wastewater in continuous activated sludge systems under different dissolved oxygen conditions: Reactor operation and evaluation using modelling.

Swine wastewater was treated in two continuously aerated activated sludge (AS) systems at high (AS1: 1.7-2.6 mg/L) and low (AS2: 0.04-0.08 mg/L) dissolved oxygen (DO), and at three temperatures (10, 20, and 30 °C). Biochemical oxygen demand (BOD) removal was >94.8%. Meanwhile, total nitrogen (N) removal was significantly higher in AS2, at 64, 89, and 88%, than in AS1, at 12, 24, and 46%, for 10, 20, and 30 °C, respectively. The experimental data were considered in a simulation study using an AS model for BOD and N removal, which also included nitrite, free ammonia, free nitrous acid, and temperature. Simulations at high-DO showed that ammonium was partly oxidized into nitrate but not removed, whereas at low-DO ammonium was removed mainly through the nitrite shortcut in simultaneous nitrification-denitrification. This study demonstrates that treatment at low-DO is an effective method for removing N, and modelling a helpful tool for its optimization.

Removal and recovery of phosphorous from swine wastewater by demonstration crystallization reactor and struvite accumulation device.

A demonstration crystallization reactor and struvite accumulation device for the removal and recovery of phosphorous was constructed and their performance was evaluated using actual swine wastewater for 3.5 years. The wastewater pH was increased by aeration, and the concentrations of total P and soluble PO(4)-P were reduced by a struvite crystallization reaction induced under a high pH condition. A 30% MgCl(2) addition was effective in enhancing the struvite crystallization reaction. The concentrations of suspended solids, total Zn and total Cu, were also decreased by the settling function of the reactor. On removing the efficiencies of these components, no noticeable seasonal fluctuation in performance was observed during the 3.5-year operation. In terms of maximum yield, 171g struvite was obtained from 1m(3) swine wastewater by the demonstration accumulation device for struvite recovery. The recovered struvite needed only air-drying before use since it was approximately 95% pure even without washing.

Anammox biofilm in activated sludge swine wastewater treatment plants.

We investigated anammox with a focus on biofilm in 10 wastewater treatment plants (WWTPs) that use activated sludge treatment of swine wastewater. In three plants, we found red biofilms in aeration tanks or final sedimentation tanks. The biofilm had higher anammox 16S rRNA gene copy numbers (up to 1.35 × 1012 copies/g-VSS) and higher anammox activity (up to 295 µmoL/g-ignition loss/h) than suspended solids in the same tank. Pyrosequencing analysis revealed that Planctomycetes accounted for up to 17.7% of total reads in the biofilm. Most of them were related to Candidatus Brocadia or Ca. Jettenia. The highest copy number and the highest proportion of Planctomycetes were comparable to those of enriched anammox sludge. Thus, swine WWTPs that use activated sludge treatment can fortuitously acquire anammox biofilm. Thus, concentrated anammox can be detected by focusing on red biofilm.

Effect of electron donors on anammox coupling with nitrate reduction for removing nitrogen from nitrate and ammonium.

Anammox coupling with nitrate reduction under various electron donors was studied using sludge acclimatized to have anammox and denitrification activities. Due to a deficiency in electron donors for NO(3)(-) reduction, anammox activity in an inorganic medium containing NO(3)(-) and NH(4)(+) was lower than that in NO(2)(-) and NH(4)(+). Anammox could use NO(2)(-) competitively against denitrifiers under a very limited NO(2)(-) concentration, and additions of swine wastewater or acetate stimulated anammox activity in an inorganic medium containing NO(3)(-) and NH(4)(+) with no inhibition effects. However, a high concentration of swine wastewater caused an exponential increase in denitrification activity. The addition of hydrogen and iron stimulated anammox activity in an inorganic medium containing NO(3)(-) and NH(4)(+), but iron showed an inhibitory effect on anammox in a medium containing NO(2)(-) and NH(4)(+). Hydrogen was shown to be advantageous since it did not increase denitrification even when its addition was increased.

Effects of struvite formation and nitratation promotion on nitrogenous emissions such as NH3, N2O and NO during swine manure composting.

To reduce nitrogenous emissions from composting, two different countermeasures were applied simultaneously in swine manure composting. One was forming struvite by adding Mg and P at the start of composting, and the other was to promote nitratation (nitrite being oxidized nitrate) by adding nitrite-oxidizing bacteria after the thermophilic phase of composting. In the laboratory- and mid-scale composting experiments, 25-43% of NH3, 52-80% of N2O and 96-99% of NO emissions were reduced. From the nitrogen balance, it was revealed that the struvite formation reduced not only NH3, but also other nitrogenous emissions except N2O. The amount of total nitrogen losses was reduced by 60% by the two combined countermeasures, against 51% by the struvite formation alone. However, the nitratation promotion dissolved struvite crystals due to the pH decline, diminishing the effect of struvite as a slow-release fertilizer.

Evidence of exponential growth of an anammox population in an anaerobic batch culture.

Twenty-five replicates of growth medium for anaerobic ammonium oxidation (anammox) containing (15)N-labeled ammonium and non-labeled nitrite were inoculated into an anammox enrichment culture at low density, and anaerobically incubated batchwise. In the headspace, (29)N(2) partial pressure linearly increased via anammox in 25 vials, confirming that anammox populations were viable in all subcultures. On prolonged incubation, exponential increases in (29)N(2) were not observed in all but 13 subcultures, suggesting that the anammox population may not proliferate unless all conditions for growth are satisfied. The estimated first-order rate coefficients in those 13 subcultures varied from 0.0029 to 0.0048 h(-1).

Characteristics of the microbial community associated with ammonia oxidation in a full-scale rockwool biofilter treating malodors from livestock manure composting.

The relationship between the activity and community structure of microbes associated with the oxidation of ammonia in a full-scale rockwool biofilter was examined by kinetic, denaturing gradient gel electrophoresis (DGGE), and sequence analyses. The packing materials were sampled from two different depths at 3 sites. Estimated K(m) values were similar among depths at same sampling sites, while V(max) differed in the mid-point sample. The lower depth of this site had the highest V(max). A correspondence analysis showed the DGGE profile of ammonia-oxidizing bacterial amoA of the lower depth of the mid-point sample to be distinguishable from the others. Banding patterns at other sites were similar among depths. Banding patterns of ammonia-oxidizing archaeal amoA of the mid-point sample were also similar among depths. The results suggested an association between the ammonia-oxidizing bacterial community's composition and ammonium oxidation kinetics in samples. Sequence analysis indicated that the ammonia-oxidizing bacterial community mainly belonged to the Nitrosomonas europaea lineage and Nitrosospira cluster 3. The ammonia-oxidizing archaeal amoA-like sequences were related to those belonging to soil and sediment groups, including one with 84% nucleotide similarity with Nitrosopumilus maritimus.