Analysis of a bio-electrochemical reactor containing carbon fiber textiles for the anaerobic digestion of tomato plant residues.

A bio-electrochemical system packed with supporting material can promote anaerobic digestion for several types of organic waste. To expand the target organic matters of a BES, tomato plant residues (TPRs), generated year-round as agricultural and cellulosic waste, were treated using three methanogenic reactors: a continuous stirred tank reactor (CSTR), a carbon fiber textile (CFT) reactor, and a bio-electrochemical reactor (BER) including CFT with electrochemical regulation (BER + CFT). CFT had positive effects on methane fermentation and methanogen abundance. The microbial population stimulated by electrochemical regulation, including hydrogenotrophic methanogens, cellulose-degrading bacteria, and acetate-degrading bacteria, suppressed acetate accumulation, as evidenced by the low acetate concentration in the suspended fraction in the BER + CFT. These results indicated that the microbial community in the BER + CFT facilitated the efficient decomposition of TPR and its intermediates such as acetate to methane.

Electrochemically applied potentials induce growth and metabolic shift changes in the hyperthermophilic bacterium Thermotoga maritima MSB8.

Bioelectrochemical systems are an attractive technology for regulating microbial activity. The effect of an applied potential on hydrolysis of starch in Thermotoga maritima as a model bacterium was investigated in this study. A cathodic potential (-0.6 and -0.8 V) induced 5-h earlier growth initiation of T. maritima with starch as the polymeric substrate than that without electrochemical regulation. Moreover, metabolic patterns of starch consumption were altered by the cathodic potential. While acetate, H2, and CO2 were the major products of starch consumption in the control experiment without electrolysis, lactate accumulation was detected rather than decreased acetate and H2 levels in the bioelectrochemical system experiments with the cathodic potential. These results indicate that the applied potential could control microbial activities related to the hydrolysis of polymeric organic substances and shift carbon and electron flux to a lactate-producing reaction in T. maritima.

Label-free impedimetric immunoassay for trace levels of polychlorinated biphenyls in insulating oil.

A rapid, ultrasensitive, and practical label-free impedimetric immunoassay for measuring trace levels of total polychlorinated biphenyls (PCBs) in insulating oil was developed. First, we developed a novel monoclonal antibody (RU6F9) for PCBs by using a designed immunogen and characterized its binding affinity for a commercial mixtures of PCBs and its main congeners. A micro comblike gold electrode was fabricated, and the antibody was covalently immobilized on the electrode through a self-assembled monolayer formed by dithiobis-N-succinimidyl propionate. The antigen-binding event on the surface of the functionalized electrode was determined as the change in charge transfer resistance by using electrochemical impedance spectroscopy. The resulting impedimetric immunoassay in aqueous solution achieved a wide determination range (0.01-10 µg/L) and a low detection limit (LOD) of 0.001 µg/L, which was 100-fold more sensitive than a conventional flow-based immunoassay for PCBs. By combining the impedimetric immunoassay with a cleanup procedure for insulating oil utilizing a multilayer cleanup column followed by DMSO partitioning, an LOD of 0.052 mg/kg-oil was achieved, which satisfied the Japanese regulation criterion of 0.5 mg/kg-oil. Finally, the immunoassay was employed to determine total PCB levels in actual used insulating oils (n = 33) sampled from a used transformer containing trace levels of PCBs, and the results agreed well with the Japanese official method (HRGC/HRMS).

Operation of a cylindrical bioelectrochemical reactor containing carbon fiber fabric for efficient methane fermentation from thickened sewage sludge.

A bioelectrochemical reactor (BER) containing carbon fiber fabric (CFF) (BER+CFF) enabled efficient methane fermentation from thickened sewage sludge. A cylindrical BER+CFF was proposed and scaled-up to a volume of 4.0-L. Thickened sewage sludge was treated using three types of methanogenic reactors. The working electrode potential in the BER+CFF was regulated at -0.8 V (vs. Ag/AgCl). BER+CFF showed gas production of 3.57 L L(-1) day(-1) at a hydraulic retention time (HRT) of 4.0 days; however, non-BER+CFF showed a lower gas production rate (0.83 L L(-1) day(-1)) at this HRT, suggesting positive effects of electrochemical regulation. A stirred tank reactor (without CFF) deteriorated at an HRT of 10 days, suggesting positive effects of CFF. 16S rRNA gene analysis showed that the BER+CFF included 3 kinds of hydrogenotrophic methanogens and 1 aceticlastic methanogen. These results demonstrate the effectiveness of the BER+CFF for scale-up and flexibility of this technology.

Trace-level mercury ion (Hg2+) analysis in aqueous sample based on solid-phase extraction followed by microfluidic immunoassay.

Mercury is considered the most important heavy-metal pollutant, because of the likelihood of bioaccumulation and toxicity. Monitoring widespread ionic mercury (Hg(2+)) contamination requires high-throughput and cost-effective methods to screen large numbers of environmental samples. In this study, we developed a simple and sensitive analysis for Hg(2+) in environmental aqueous samples by combining a microfluidic immunoassay and solid-phase extraction (SPE). Using a microfluidic platform, an ultrasensitive Hg(2+) immunoassay, which yields results within only 10 min and with a lower detection limit (LOD) of 0.13 µg/L, was developed. To allow application of the developed immunoassay to actual environmental aqueous samples, we developed an ion-exchange resin (IER)-based SPE for selective Hg(2+) extraction from an ion mixture. When using optimized SPE conditions, followed by the microfluidic immunoassay, the LOD of the assay was 0.83 µg/L, which satisfied the guideline values for drinking water suggested by the United States Environmental Protection Agency (USEPA) (2 µg/L; total mercury), and the World Health Organisation (WHO) (6 µg/L; inorganic mercury). Actual water samples, including tap water, mineral water, and river water, which had been spiked with trace levels of Hg(2+), were well-analyzed by SPE, followed by microfluidic Hg(2+) immunoassay, and the results agreed with those obtained from reduction vaporizing-atomic adsorption spectroscopy.

Construction of hydrogen fermentation from garbage slurry using the membrane free bioelectrochemical system.

The aim of this study was to show the effectiveness of the membrane free bioelectrochemical system (BES) using three electrodes on inhibition of methanogenesis and construction of hydrogen fermentation from the artificial garbage slurry. The electrical redox-potential on the working electrode was adjusted to -1.0V (vs. Ag/AgCl) that has positive effect on methanogenesis. The redox-potential on the counter electrode was measured to be 1.6V. The pH in the effluents was 5.5-6.4. Hydrogen production rate at the cathode side was similar to that at the anode side and much higher than that calculated from current, and reached a maximum of 2445±815 (average±standard deviation) mL L(-1) d(-1) at an organic loading rate of 58.7g dichromate chemical oxygen demand per L d(-1). Methane production was negligible throughout the experiment. Acetate and butyrate were the main products of the fermentation using a BES; these offered favorable conditions for hydrogen production. The bacterial community in the bioelectrochemical hydrogen fermentor differed from that in the methanogenic seed sludge and included hitherto unknown species. These results show that high redox-potential on the anodic electrode and acidic pH in the membrane free BES can be utilized for hydrogen fermentation from the artificial garbage slurry by avoiding methanogenesis.

Bioelectrochemical regulation accelerates facultatively syntrophic proteolysis.

Bioelectrochemical systems can affect microbial metabolism by controlling the redox potential. We constructed bioelectrochemical cultures of the proteolytic bacterium, Coprothermobacter proteolyticus strain CT-1, both as a single-culture and as a co-culture with the hydrogenotrophic methanogen, Methanothermobacter thermautotrophicus strain ∆H, to investigate the influences of bioelectrochemical regulation on facultatively syntrophic proteolysis. The co-culture and single-culture were cultivated at 55°C with an anaerobic medium containing casein as the carbon source. The working electrode potential of the bioelectrochemical system was controlled at -0.8V (vs. Ag/AgCl) for bioelectrochemical cultures and was not controlled for non-bioelectrochemical cultures. The cell densities of hydrogenotrophic methanogen and methane production in the bioelectrochemical co-culture were 3.6 and 1.5 times higher than those in the non-bioelectrochemical co-culture after 7 days of cultivation, respectively. Contrastingly, the cell density of Coprothermobacter sp. in the bioelectrochemical co-culture was only 1.3 times higher than that in the non-bioelectrochemical co-culture. The protein decomposition rates were nearly proportional to the cell density of Coprothermobacter sp. in the all types of cultures. These results indicate that bioelectrochemical regulation, particularly, affected the carbon fixation of the hydrogenotrophic methanogen and that facultatively syntrophic proteolysis was accelerated as a result of hydrogen consumption by the methanogens growing well in bioelectrochemical co-cultures.

Microfluidic heavy metal immunoassay based on absorbance measurement.

A simple and rapid flow-based multioperation immunoassay for heavy metals using a microfluidic device was developed. The antigen-immobilized microparticles in a sub-channel were introduced as the solid phase into a main channel structures through a channel flow mechanism and packed into a detection area enclosed by dam-like structures in the microfluidic device. A mixture of a heavy metal and a gold nanoparticle-labeled antibody was made to flow toward the corresponding metal through the main channel and make brief contact with the solid phase. A small portion of the free antibody was captured and accumulated on the packed solid phase. The measured absorbance of the gold label was proportional to the free antibody portion and, thus, to the metal concentration. Each of the monoclonal antibodies specific for cadmium-EDTA, chromium-EDTA, or lead-DTPA was applied to the single-channel microfluidic device. Under optimized conditions of flow rate, volume, and antibody concentration, the theoretical (antibody K(d)-limited) detection levels of the three heavy metal species were achieved within only 7 min. The dynamic range for cadmium, chromium, and lead was 0.57-60.06 ppb, 0.03-0.97 ppb, and 0.04-5.28 ppb, respectively. An integrated microchannel device for simultaneous multiflow was also successfully developed and evaluated. The multiplex cadmium immunoassay of four samples was completed within 8 min for a dynamic range of 0.42-37.48 ppb. Present microfluidic heavy metal immunoassays satisfied the Japanese environmental standard for cadmium, chromium and, lead, which provided in the soil contamination countermeasures act.

Syntrophic degradation of proteinaceous materials by the thermophilic strains Coprothermobacter proteolyticus and Methanothermobacter thermautotrophicus.

Protein is a major component of organic solid wastes, and therefore, it is necessary to further elucidate thermophilic protein degradation process. The effects of hydrogenotrophic methanogens on protein degradation were investigated using the proteolytic bacterial strain CT-1 that was isolated from a methanogenic thermophilic (55°C) packed-bed reactor degrading artificial garbage slurry. Strain CT-1 was closely related to Coprothermobacter proteolyticus, which is frequently found in methanogenic reactors degrading organic solid wastes. Strain CT-1 was cultivated in the absence or presence of Methanothermobacter thermautotrophicus by using 3 kinds of proteinaceous substrates. Degradation rates of casein, gelatin, and bovine serum albumin were higher in co-cultures than in monocultures. Strain CT-1 showed faster growth in co-cultures than in monocultures. M. thermautotrophicus comprised 5.5-6.0% of the total cells in co-culture. Increased production of ammonia and acetate was observed in co-cultures than in monocultures, suggesting that addition of M. thermautotrophicus increases the products of protein degradation. Hydrogen produced in the monocultures was converted to methane in co-cultures. These results suggest that thermophilic proteolytic bacteria find it favorable to syntrophically degrade protein in a methanogenic environment, and that it is important to retain hydrogen-scavenging methanogens within the reactor.

A bioelectrochemical reactor containing carbon fiber textiles enables efficient methane fermentation from garbage slurry.

A packed-bed system includes supporting materials to retain microorganisms and a bioelectrochemical system influences the microbial metabolism. In our study, carbon fiber textiles (CFT) as a supporting material was attached onto a carbon working electrode in a bioelectrochemical reactor (BER) that degrades garbage slurry to methane, in order to investigate the effect of combining electrochemical regulation and packing CFT. The potential on the working electrode in the BER containing CFT was set to -1.0 V or -0.8 V (vs. Ag/AgCl). BERs containing CFT exhibited higher methane production, elimination of dichromate chemical oxygen demand, and the ratio of methanogens in the suspended fraction than reactors containing CFT without electrochemical regulation at an organic loading rate (OLR) of 27.8 gCODcr/L/day. In addition, BERs containing CFT exhibited higher reactor performances than BERs without CFT at this OLR. Our results revealed that the new design that combined electrochemical regulation and packing CFT was effective.

Methanogenic communities on the electrodes of bioelectrochemical reactors without membranes.

Methane fermentation was successfully carried out in bioelectrochemical reactors without membranes under a working potential of -0.6 or -0.8 V (vs. Ag/AgCl) and neutral pH conditions. The hydrogenotrophic methanogens that dominated on the anodic and cathodic electrodes differed from those found on the electrodes in the control reactors without electrochemical reactions.

Bioelectrochemical system accelerates microbial growth and degradation of filter paper.

Bioelectrochemical reactors (BERs) with a cathodic working potential of -0.6 or -0.8 V more efficiently degraded cellulosic material, i.e., filter paper (57.4-74.1% in 3 days and 95.9-96.3% in 7 days) than did control reactors without giving exogenous potential (15.4% in 3 days and 64.2% in 7 days). At the same time, resultant conversions to methane and carbon dioxide in cathodic working chamber of BERs by application of electrochemical reduction in 3 days of operation were larger than control reactors. However, cumulative methane production in cathodic BERs was similar to those in control reactors after 7 days of operation. Microscopic observation and 16S rRNA gene analysis showed that microbial growth in the entire consortium was higher after 2 days of operation of cathodic BERs as compared with the control reactors. In addition, the number of methanogenic 16S rRNA gene copies in cathodic BERs was higher than in control reactors. Moreover, archaeal community structures constructed in cathodic BERs consisted of hydrogenotrophic methanogen-related organisms and differed from those in control reactors after 2 days of operation. Specifically, the amount of Methanothermobacter species in cathodic BERs was higher within archaeal communities than in those control reactors after 2 days of operation. Electrochemical reduction may be effective for accelerating microbial growth in the start-up period and thereby increasing microbial treatment of cellulosic waste and methane production.

Efficient treatment of garbage slurry in methanogenic bioreactor packed by fibrous sponge with high porosity.

Adding a supporting material to a methanogenic bioreactor treating garbage slurry can improve efficiency of methane production. However, little is known on how characteristics (e.g., porosity and hydrophobicity) of the supporting material affect the bioreactor degrading garbage slurry. We describe the reactor performances and microbial communities in bioreactors containing hydrophilic or hydrophobic sheets, or fibrous hydrophilic or hydrophobic sponges. The porosity affected the efficiency of methane production and solid waste removal more than the hydrophilic or hydrophobic nature of the supporting material. When the terminal restriction fragment length polymorphism technique was used at a lower organic loading rate (OLR), microbial diversities in the suspended fraction were retained on the hydrophobic, but not the hydrophilic, sheets. Moreover, real-time quantitative polymerase chain reaction (PCR) performed at a higher OLR revealed that the excellent performance of reactors containing fibrous sponges with high porosity (98%) was supported by a clear increase in the numbers of methanogens on these sponges, resulting in larger total numbers of methanogens in the reactors. In addition, the bacterial communities in fractions retained on both the hydrophobic and hydrophilic fibrous sponges differed from those in the suspended fraction, thus increasing bacterial diversity in the reactor. Thus, higher porosity of the supporting material improves the bioreactor performance by increasing the amount of methanogens and bacterial diversity; surface hydrophobicity contributes to maintaining the suspended microbial community.

Bioelectrochemical system stabilizes methane fermentation from garbage slurry.

Methanogenic bioreactors, which are packed with supporting material, have attracted attention as an efficient means of degrading garbage. We aimed to increase bioreactor performance by using an electrochemical system to regulate the electrical potential on supporting material. At an organic loading rate of 26.9g dichromate chemical oxygen demand (CODcr)/L/day, reactors with a potential of -0.6 or -0.8V, generated by a cathodic electrochemical reaction, showed greater removal of CODcr and methanogenesis than reactors with a potential of 0.0 or -0.3V, generated by anodic reaction, or control reactors without electrochemical regulation. 16S rRNA gene analysis revealed that the same methanogens were present in all our reactors, but quantitative real-time polymerase chain reaction showed that higher prokaryotic and methanogenic copy numbers were present on cathodic electrodes than on anodic or control electrodes. These results indicate that cathodic electrochemical regulation can support methane fermentation from garbage.

Facilitation of voluntary swallowing by chemical stimulation of the posterior tongue and pharyngeal region in humans.

In this study, we investigated the functional difference between chemical stimulations of the posterior tongue (PT) and pharyngeal region (PR) for facilitation of voluntary swallowing in humans. The PT or PR stimulation consisted of infusion of water (distilled water), 0.3 M NaCl solution or olive oil (non-chemical stimulant) into the PT or the PR through a fine tube at a very slow infusion rate (0.2 ml/min). Water was used as a stimulant of water receptors. A solution of 0.3 M NaCl was used as an inhibitor of the response of water receptors and as a stimulant of salt taste receptors. Excitation of the mucosal receptors would facilitate voluntary swallowing and diminution of sensory inputs from the oral mucosa would induce difficulty in swallowing. Swallowing intervals (SIs) during voluntary swallowing were measured by submental electromyographic activity. Infusion of water into the PR shortened SI (facilitation of swallowing) and infusion of 0.3 M NaCl or olive oil into the same region prolonged it (difficulty in swallowing). On the other hand, infusion of water into the PT prolonged SI and infusion of 0.3 M NaCl into the same region shortened it. The results suggest that water receptors are localized in the PR and that salt taste receptors are almost absent in the PR and present in the PT. With diminution of sensory inputs from the oral mucosa, central inputs would play a dominant role in initiating swallowing voluntarily, and SI would be prolonged. With weak stimulation (infusion of 0.3 M NaCl into the PR or infusion of water into the PT), SI was prolonged and inter-individual variation in SI was pronounced, suggesting that the ability of the central regulation of swallowing to perform repetitive voluntary swallowing varies among subjects. With stimulation of water receptors or salt taste receptors, SI was shortened and inter-individual variation in SI was moderate, suggesting that sensory inputs are important for performing voluntary swallowing smoothly and that the sensory inputs compensate for the difficulty in performing swallowing caused by the central mechanism.

Case study of electrochemical metal removal from actual sediment, sludge, sewage and scallop organs and subsequent pH adjustment of sediment for agricultural use.

In this paper we propose a method for chemical-free removal of metal from lake sediment, and its subsequent pH adjustment, based on electrochemical migration and precipitation. Such a method would enable the utilization of sediment as composting material. Sediment was placed in the anode side of a dual-bath electrochemical reactor separated by a thimble-shape cellulose filter from the cathode side, which was filled with pure water. When voltage was applied, contaminant metals in the sediment on the anode side migrated toward the cathode side, and precipitated due to the alkaline conditions caused by the cathodic reaction. After 10 days of electrolysis with 400 mA of constant current of 150 g wet lake sediment, the removal ratios of 13 kinds of elements after the electrochemical treatment were measured. Cd and Zn, the elements for which agricultural standards apply, showed 98% and 86% removal, respectively. The type of metal removed changed over time, and the order of removal was roughly from light metals to heavy metals. The acidified lake sediment after electrolysis could be neutralized without significant recontamination with Zn and Cd by using the alkaline cathode solution collected during electrolysis under a condition of tap water overflow at a rate of 1.5 L/h. The electrochemical metal removal method was effective not only for lake sediment, but also for municipal sludge cake, human sewage, and contaminated scallop organs. Cathode overflow during electrolysis tended to increase metal removal and decrease required voltage.

Effect of electrical stimulation of the internal capsule on nociceptive neurons responding to orofacial stimuli in the medullary dorsal horn of the rat.

Internal capsule (IC) stimulation has been used clinically to alleviate central pain. However, the neuronal mechanisms underlying pain relief by IC stimulation are poorly understood. In order to elucidate the analgesic mechanism, the effect of IC conditioning stimulation on nociceptive neurons in the rat medullary dorsal horn was investigated in the present study. Rats were anaesthetized with N(2)O-O(2) (2:1) and 0.5% halothane and were immobilized with pancuronium bromide. Two kinds of nociceptive neurons, wide dynamic range (WDR) and nociceptive specific (NS) neurons, responding to noxious stimulations of the face and oral structures were recorded in the trigeminal caudal nucleus and the medial reticular subnuclei. A test stimulus with a single rectangular pulse (2ms in duration, 5-70V) was applied to the centre of the receptive field. Responses in 55.9% of the WDR neurons and in 60% of the NS neurons were inhibited by conditioning stimuli to the ipsilateral IC with trains of 33 pulses (100-300microA) at 330Hz. The percents of peak inhibitory effects on WDR neurons and NS neurons were 78.1+/-25.0% (n=19) and 89.0+/-13.6% (n=3), respectively. The inhibitory effect continued for conditioning-test intervals of up to 500ms. Effective sites for conditioning stimulation were concentrated in the lateral side of the IC. These findings suggest that modulation of nociceptive transmission by IC stimulation occurs at second-order neurons via a presynaptic phenomenon by corticofugal fibers in the IC.

Respiratory isozyme, two types of rusticyanin of Acidithiobacillus ferrooxidans.

Among the members of the copper protein superfamily, the type I enzyme rusticyanin, which is found as an electron carrier in the oxidative respiratory chain of Acidithiobacillus ferrooxidans, is the only one to have both a high redox potential and acid stability. Here we report that two forms of the rusticyanin gene (rus) are present in the genomes of some strains of A. ferrooxidans. The more common form of rus (type-A) was found to be present in all six strains studied, including those harboring only a single copy of the gene. In addition a less common form (type-B) occurred in strains harboring multiple copies of the gene. The two genes were expressed as rusticyanin isozymes with differing surface charges due to differences in their amino acid composition. Still, the copper coordination sites were completely conserved, thereby maintaining the high redox potential necessary for an electron carrier.

Responses of diencephalic nociceptive neurones to orofacial stimuli and effects of internal capsule stimulation in the rat.

The effect of conditioning stimulation of the internal capsule on nociceptive neurones in the rat diencephalon was investigated. The animals were anaesthetised with N(2)O/O(2) (2:1) and 0.5% halothane, and immobilised with pancuronium bromide. Nociceptive neurones responding to noxious stimulation of the face and oral structures were recorded in the ventral posteromedial nucleus, posterior group and zona incerta. These neurones were classified into wide dynamic range and nociceptive-specific types. Functional segregation of these nociceptive neurones was not apparent within the nucleus or between nuclei. A test stimulus with a single rectangular pulse (5-70 V) was applied to the centre of the receptive field; the nociceptive neurones exhibited short- and/or long-latency responses. Both responses in about 45% of the nociceptive neurones were inhibited by conditioning stimuli to the contralateral internal capsule with trains of 33 pulses (300 microA) at 330 Hz. The percent inhibitory effects on the nociceptive neurones of each area were 68.0+/-14.8% (n = 6) in the ventral posteromedial nucleus, 72.8+/-12.4% (n = 4) in the posterior group and 61.5+/-7.5% (n = 4) in the zona incerta. Effective sites for conditioning stimulation were concentrated in the lateral side of the internal capsule, through which the corticofugal fibres from the somatosensory cortex pass. These findings suggest that the transmission of nociceptive information to the diencephalon is modulated by stimulation of the internal capsule at the level of the trigeminal sensory complex in the brainstem. They might provide a novel way to elucidate the neurophysiological basis for antinociception by stimulation of the internal capsule observed in clinical studies.

Anaerobic respiration using Fe(3+), S(0), and H(2) in the chemolithoautotrophic bacterium Acidithiobacillus ferrooxidans.

The chemolithoautotrophic bacterium Acidithiobacillus ferrooxidans has been known as an aerobe that respires on iron and sulfur. Here we show that the bacterium could chemolithoautotrophically grow not only on H(2)/O(2) under aerobic conditions but also on H(2)/Fe(3+), H(2)/S(0), or S(0)/Fe(3+) under anaerobic conditions. Anaerobic respiration using Fe(3+) or S(0) as an electron acceptor and H(2) or S(0) as an electron donor serves as a primary energy source of the bacterium. Anaerobic respiration based on reduction of Fe(3+) induced the bacterium to synthesize significant amounts of a c-type cytochrome that was purified as an acid-stable and soluble 28-kDa monomer. The purified cytochrome in the oxidized form was reduced in the presence of the crude extract, and the reduced cytochrome was reoxidized by Fe(3+). Respiration based on reduction of Fe(3+) coupled to oxidation of a c-type cytochrome may be involved in the primary mechanism of energy production in the bacterium on anaerobic iron respiration.