Ping Jei Lin

Kinetic characteristics of bacterial azo-dye decolorization by Pseudomonas luteola

A Pseudomonas luteola strain expressing azoreductase activity was utilized to remove the color of an azo dye (reactive red 22) from contaminated solutions. The effects of substrate concentrations, medium compositions, and operation parameters (e.g., pH, temperature, dissolved oxygen, etc.) on decolorization of the azo dye by a P. luteola strain were systematically investigated to reveal the key factors that dominate the performance of azo-dye decolorization. The metabolites resulting from bacterial decolorization were analyzed by high-performance liquid chromatography (HPLC) and mass spectrometery (MS). The results show that the dissolved oxygen and glucose concentration retarded decolorization of reactive red 22 by P. luteola. The optimal azo-dye decolorization occurred at 37 degrees C, while more rapid decolorization took place over pH 7-9. Yeast extract and tryptone strongly enhanced the decolorization. The Michaelis-Menten model can satisfactorily describe the dependence of specific decolorization rate on the concentration of substrate (reactive red 22 or yeast extract). Decolorization of the azo dye by intact cells of P. luteola was essentially independent of the growth phase, whereas the azoreductase activity of the cell-free extract decreased in the order of late-stationary phase > early-stationary phase > mid-log phase. This suggests that mass transfer of the azo dye across the cell membrane may be the rate-limiting step. The HPLC and MS analyses suggest that both partial reduction and complete cleavage of the azo bond could contribute to decolorization of reactive red 22 by P. luteola.

Microbial hydrogen production with immobilized sewage sludge

Municipal sewage sludge was immobilized to produce hydrogen gas under anaerobic conditions. Cell immobilization was essentially achieved by gel entrapment approaches, which were physically or chemically modified by addition of activated carbon (AC), polyurethane (PU), and acrylic latex plus silicone (ALSC). The performance of hydrogen fermentation with a variety of immobilized‐cell systems was assessed to identify the optimal type of immobilized cells for practical uses. With sucrose as the limiting carbon source, hydrogen production was more efficient with the immobilized‐cell system than with the suspended‐cell system, and in both cases the predominant soluble metabolites were butyric acid and acetic acid. Addition of activated carbon into alginate gel (denoted as CA/AC cells) enhanced the hydrogen production rate ( vH2) and substrate‐based yield ( YH2/sucrose) by 70% and 52%, respectively, over the conventional alginate‐immobilized cells. Further supplementation of polyurethane or acrylic latex/silicone increased the mechanical strength and operation stability of the immobilized cells but caused a decrease in the hydrogen production rate. Kinetic studies show that the dependence of specific hydrogen production rates on the concentration of limiting substrate (sucrose) can be described by Michaelis‐Menten model with good agreement. The kinetic analysis suggests that CA/AC cells may contain higher concentration of active biocatalysts for hydrogen production, while PU and ALSC cells had better affinity to the substrate. Acclimation of the immobilized cells led to a remarkable enhancement in vH2 with a 25‐fold increase for CA/AC and ca. 10‐ to 15‐fold increases for PU and ALSC cells. However, the ALSC cells were found to have better durability than PU and CA/AC cells as they allowed stable hydrogen production for over 24 repeated runs.

High-Efficiency Hydrogen Production by an Anaerobic, Thermophilic Enrichment Culture From an Icelandic Hot Spring

Dark fermentative hydrogen production from glucose by a thermophilic culture (33HL), enriched from an Icelandic hot spring sediment sample, was studied in two continuous‐flow, completely stirred tank reactors (CSTR1, CSTR2) and in one semi‐continuous, anaerobic sequencing batch reactor (ASBR) at 58°C. The 33HL produced H2 yield (HY) of up to 3.2 mol‐H2/mol‐glucose along with acetate in batch assay. In the CSTR1 with 33HL inoculum, H2 production was unstable. In the ASBR, maintained with 33HL, the H2 production enhanced after the addition of 6 mg/L of FeSO4 · 7H2O resulting in HY up to 2.51 mol‐H2/mol‐glucose (H2 production rate (HPR) of 7.85 mmol/h/L). The H2 production increase was associated with an increase in butyrate production. In the CSTR2, with ASBR inoculum and FeSO4 supplementation, stable, high‐rate H2 production was obtained with HPR up to 45.8 mmol/h/L (1.1 L/h/L) and HY of 1.54 mol‐H2/mol‐glucose. The 33HL batch enrichment was dominated by bacterial strains closely affiliated with Thermobrachium celere (99.8–100%). T. celere affiliated strains, however, did not thrive in the three open system bioreactors. Instead, Thermoanaerobacterium aotearoense (98.5–99.6%) affiliated strains, producing H2 along with butyrate and acetate, dominated the reactor cultures. This culture had higher H2 production efficiency (HY and specific HPR) than reported for mesophilic mixed cultures. Further, the thermophilic culture readily formed granules in CSTR and ASBR systems. In summary, the thermophilic culture as characterized by high H2 production efficiency and ready granulation is considered very promising for H2 fermentation from carbohydrates. Biotechnol. Bioeng. 2008;101: 665–678.

Bioreactors configured with distributors and carriers enhance the performance of continuous dark hydrogen fermentation.

Anaerobic granular sludge bed (AnGSB) bioreactors were supplemented with activated carbon carriers and combined with distributors (e.g., acrylic resin board, stainless steel net and plastic net) installed at different locations to investigate the effect of distributor/carrier on biohydrogen production efficiency. The results show that plastic net stimulated the substrate/microorganisms contact and sludge granulation, thereby leading to a much better H(2) production performance when compared with those obtained from traditional CSTR. The highest H(2) production rate (7.89 L/h/L) and yield (3.42 mol H(2)/mol sucrose) were obtained when two pieces of plastic nets were installed at both 4 cm and 16 cm from the bottom of AnGSB without carrier addition and the bioreactor was operated at a HRT of 0.5h. For the AnGSB installed with two pieces of plastic net distributors, addition of carriers led to significant improvement on the H(2) production efficiency at a longer HRT (1-4h) when compared with the carrier-absent system.

Detoxification of mercury by immobilized mercuric reductase

Mercuric reductase which originated from a recombinant Escherichia coli PWS1 was purified and immobilized on a chemically modified diatomaceous earth support. The mercury reduction kinetics, pH dependence, storage stability, and reusability of the immobilized enzyme were investigated. Four dyes were examined for their electron transfer efficiency with the soluble and bound mercuric reductase. Continuous mercury detoxification by the immobilized mercuric reductase was also performed in fixed-bed processes. The effects of bed-length, mercury loading rate, and electron donor on the performance of the fixed beds were assessed. Immobilized mercuric reductase exhibited substrate-inhibition-type kinetics with a maximal activity (1.2 nmol Hg mg−1 protein s−1) occurring at an initial Hg2+ concentration of 50 µmol dm−3. The optimal pH was 7.0 for the soluble and immobilized mercuric reductase, but the immobilized enzyme maintained higher relative activity for less favorable pH values. Immobilization of the enzyme appeared to significantly enhance its storage stability and reusability. Of four artificial electron donors tested, azure A (5 mmol dm−3) demonstrated the highest relative activity (78%) for soluble mercuric reductase. For the immobilized enzyme, neutral red (5 mmol dm−3) gave a relative activity of nearly 82%. With a fixed-bed, the mercury-reducing efficiency of using neutral red was only 30–40% of that obtained using NADPH. Fixed-bed operations also showed that increased bed length facilitated mercury reduction rate, and the optimal performance of the beds was achieved at a flow rate of approximately 100–200 cm3 h−1. © 1999 Society of Chemical Industry