James M. Gossett

Biodegradation of cis-Dichloroethene as the Sole Carbon Source by a β-Proteobacterium

ABSTRACT An aerobic bacterium capable of growth on cis-dichloroethene (cDCE) as a sole carbon and energy source was isolated by enrichment culture. The 16S ribosomal DNA sequence of the isolate (strain JS666) had 97.9% identity to the sequence from Polaromonas vacuolata, indicating that the isolate was a β-proteobacterium. At 20°C, strain JS666 grew on cDCE with a minimum doubling time of 73 ± 7 h and a growth yield of 6.1 g of protein/mol of cDCE. Chloride analysis indicated that complete dechlorination of cDCE occurred during growth. The half-velocity constant for cDCE transformation was 1.6 ± 0.2 μM, and the maximum specific substrate utilization rate ranged from 12.6 to 16.8 nmol/min/mg of protein. Resting cells grown on cDCE could transform cDCE, ethene, vinyl chloride, trans-dichloroethene, trichloroethene, and 1,2-dichloroethane. Epoxyethane was produced from ethene by cDCE-grown cells, suggesting that an epoxidation reaction is the first step in cDCE degradation.

Phylogenetic and Kinetic Diversity of Aerobic Vinyl Chloride-Assimilating Bacteria from Contaminated Sites

ABSTRACT Aerobic bacteria that grow on vinyl chloride (VC) have been isolated previously, but their diversity and distribution are largely unknown. It is also unclear whether such bacteria contribute to the natural attenuation of VC at chlorinated-ethene-contaminated sites. We detected aerobic VC biodegradation in 23 of 37 microcosms and enrichments inoculated with samples from various sites. Twelve different bacteria (11 Mycobacterium strains and 1 Nocardioides strain) capable of growth on VC as the sole carbon source were isolated, and 5 representative strains were examined further. All the isolates grew on ethene in addition to VC and contained VC-inducible ethene monooxygenase activity. The Mycobacterium strains (JS60, JS61, JS616, and JS617) all had similar growth yields (5.4 to 6.6 g of protein/mol), maximum specific growth rates (0.17 to 0.23 day−1), and maximum specific substrate utilization rates (9 to 16 nmol/min/mg of protein) with VC. The Nocardioides strain (JS614) had a higher growth yield (10.3 g of protein/mol), growth rate (0.71 day−1), and substrate utilization rate (43 nmol/min/mg of protein) with VC but was much more sensitive to VC starvation. Half-velocity constant (Ks) values for VC were between 0.5 and 3.2 μM, while Ks values for oxygen ranged from 0.03 to 0.3 mg/liter. Our results indicate that aerobic VC-degrading microorganisms (predominantly Mycobacterium strains) are widely distributed at sites contaminated with chlorinated solvents and are likely to be responsible for the natural attenuation of VC.

Reductive dehalogenation of chlorinated ethenes and halogenated ethanes by a high-rate anaerobic enrichment culture.

An anaerobic enrichment culture, using CH 3 OH as an electron donor, dechlorinated tetrachloroethene (PCN, 55 μmol added/100 mL of culture) nearly stoichiometrically to vinyl chloride (VC) in 20 h with negligible buildup of other intermediates and at a maximum rate of 4.6±0.4 μmol of PCN transformed/mg of volatile suspended solids per day. Appreciable conversion of VC to NTH occurred only after the PCN was nearly depleted, suggesting the inhibition of VC dechlorination by PCN. PCN, trichloroethene, cis-1,2-dichloroethene (DCN), and 1,1-DCN were all rapidly metabolized to VC with near zero-order kinetics and apparently inhibited subsequent VC dechlorination

Sustained aerobic oxidation of vinyl chloride at low oxygen concentrations.

One possible explanation for unexplained disappearance of vinyl chloride (VC) from what are thought to be anaerobic subsurface environments is that the environments are, in fact, not anaerobic. Rather, they might be subject to low, steady influx of oxygen, and aerobic oxidation could be occurring at extremely low oxygen concentrations. Studies were conducted with VC-oxidizing transfer cultures derived from two chloroethene-contaminated sites, as well as with microcosms constructed from sediment and groundwater from one of these sites. Oxygen was steadily delivered to the experimental systems using permeation tubes to maintain low dissolved oxygen throughout the time-course of investigation. VC oxidation was sustained at dissolved oxygen concentrations below 0.02 mg/L in the two transfer cultures, and below 0.1 mg/L in the microcosms. This supports the possibility that-at least at some sites-apparent loss of VC from what are thought to be anaerobic zones might, in fact, be due to aerobic pathways occurring under conditions of low oxygen flux (e.g., via diffusion from surrounding aerobic regions and/or from recharge events).

Using FTIR to predict saccharification from enzymatic hydrolysis of alkali‐pretreated biomasses

Fourier transform infrared, attenuated total reflectance (FTIR‐ATR) spectroscopy combined with partial least squares (PLS) regression accurately predicted 72‐h glucose and xylose conversions (g sugars/100 g potential sugars) and yields (g sugars/100 g dry solids) from cellulase‐mediated hydrolysis of alkali‐pretreated lignocellulose. Six plant biomasses that represent a variety of potential biofuel feedstocks—two switchgrass cultivars, big bluestem grass, a low‐impact, high‐diversity mixture of 32 species of prairie biomasses, mixed hardwood, and corn stover—were subjected to four levels of low‐temperature NaOH pretreatment to produce 24 samples with a wide range of potential digestibility. PLS models were constructed by correlating FTIR spectra of pretreated samples to measured values of gluose and xylose conversions and yields. Variable selection, based on 90% confidence intervals of regression‐coefficient matrices, improved the predictive ability of the models, while simplifying them considerably. Final models predicted sugar conversions with coefficient of determination for cross‐validation (Q2) values of 0.90 for glucose and 0.89 for xylose, and sugar yields with Q2 values of 0.92 for glucose and 0.91 for xylose. The sugar‐yield models are noteworthy for their ability to predict enzymatic saccharification per mass dry solids without a priori knowledge of the composition of the solids. All peaks retained in the final regression coefficient matrices were previously assigned to chemical bonds and functional groups in lignocellulose, demonstrating that the models were based on real chemical information. This study demonstrates that FTIR spectroscopy combined with PLS regression can be used to rapidly estimate sugar conversions and yields from enzymatic hydrolysis of pretreated plant biomass. Biotechnol. Bioeng. 2012; 109:353–362.

Assessment of commercial hemicellulases for saccharification of alkaline pretreated perennial biomass.

The objective of this research was to measure the effects of different cellulase and hemicellulase mixtures on fermentable sugar production from two different perennial biomasses--switchgrass and a low-impact, high-diversity prairie biomass mixture (LIHD). Each was subjected to NaOH pretreatment, followed by hydrolysis with a commercial cellulase and β-glucosidase mixture [CB] supplemented with either of two hemicellulases. For both biomasses, there was little gain in sugar yield when using CB alone beyond 20-25 mg/g TS; further gain in yield was possible only through hemicellulase supplementation. An equation that modeled CB and hemicellulase effects as occurring independently fit the data reasonably well, except at the lowest of cellulase loadings with hemicellulase, where synergistic interactions were evident. Examination of the marginal effectiveness of enzyme loadings (incremental grams sugar per incremental mg enzyme) over a broad range of loadings suggests that there is no need to customize enzymatic hydrolysis for NaOH-pretreated switchgrass and LIHD.

High-solids aerobic decomposition: pilot-scale reactor development and experimentation

Pilot-scale reactors have been constructed to mimic the central core of an aerated static bed or in-vessel composting process. The 770 litre reactors were instrumented to measure temporal and spatial variations in temperature, oxygen and moisture content. Experiments were performed with a synthetic food waste (SFW) and digested biosolids using four different aeration rates and two initial moisture contents. An analysis of the temporal and spatial temperature and oxygen profiles has shown the systems replicate well and represent a process with one-dimensional spatial variation. An analysis of oxygen gradients has shown that cumulative oxygen depletion and oxygen depletion rates within the bed increased with increasing aeration rate in the SFW experiments, however, they decreased with increasing aeration rate in the biosolids experiments. The SFW studies showed that a 10% variation in initial moisture content had little influence on cumulative O2 consumed, but had a significant influence on the location of maximum biological activity within the bed. Maximum temperatures varied from 58 to 74°C in the SFW experiments and from 43 to 60°C in the biosolids experiments. In all experiments the maximum temperatures and the positions where they occurred varied with initial moisture content and aeration rate. In the SFW experiments maximum axial temperature differences coincided with significant axial differences in moisture content, while in the biosolids experiments maximum axial temperature differences coincided with minimal axial differences in moisture content.

Effect of chemical coagulation on anaerobic digestibility of organic materials

Abstract Previous studies have indicated that coagulants used in wastewater treatment (principally alum or ferric chloride) cause resulting sludges to be less biodegradable in subsequent anaerobic digestion. The objectives of this research were to determine the types of organic materials whose digestibility is most affected by coagulants. Substrates studied included: raw wastewater, activated sludge, three proteins, glycine, cellulose, glucose, butyric acid, and palmitic acid. In general, those substrates whose digestibilities were most affected were those that are insoluble in water and/or are known to form complexes with iron or aluminum. Thus, amino acids, proteins, and long-chain fatty acids were particularly affected, while glucose and butyric acid were not. Cellulose was moderately affected, suggesting that physical enmeshment as well as chemical interactions may be important in determining extent of coagulant effects.

THE ROLE OF PERIODIC AGITATION AND WATER ADDITION IN MANAGING MOISTURE LIMITATIONS DURING HIGH-SOLIDS AEROBIC DECOMPOSITION

Abstract The results of this study were obtained from a pilot-scale experimental system designed to mimic an agitated bed composting process. This choice of scale represents a compromise between a better controlled and reproducible bench-scale system-and a reality-based full-scale system. Two substrates were studied: a synthetic food waste (dry dog food) and anaerobically digested, polymer–dewatered biosolids. The goal was to evaluate the degree to which periodic agitation: (1) lessened spatial gradients in moisture and temperature; and (2) thereby improved sustained degradation rates and cumulative extent of biodegradation. Coefficient of variation for cumulative O 2 measurements in the three replications of the 55% dog food and wood chips experiment with an aeration rate of 100 l min −1 were of the order of 25–30% which was consistent with other bench and pilot-scale studies. Drying occurred in both static-bed and agitated-bed studies. However, moisture gradients in the agitated-bed were smaller than in the static-bed experiments. Also, drying of the solids matrix increased with increased aeration rates for all the experiments. For the 45% dog food and wood chips experiments, a moisture content of 30% was reached in 180–300 h for aeration rates of 50–100 l min −1 , respectively; while for the 55% dog food and wood chips experiments, 30% was reached in 320–480 h for aeration rates of 50–100 l min −1 . The results of the water addition study showed that adding water three times per week resulted in a cumulative O 2 uptake at 622 g O 2 kg −1 of TS at 496 h, a 48% increase over no water addition for an additional 96 h of decomposition, and a 26% increase over adding water once a week.

The Determination of Henry’s Constant for Volatile Organics by Equilibrium Partitioning in Closed Systems

Equilibrium Partitioning in Closed Systems (EPICS) is a simple technique proposed for determining the Henry’s constants of volatile organics in water. It requires no special apparatus, and obtains results from the measurement of gas concentration ratios so that the preparation of standard curves for determining exact concentrations is not required. Henry’s constants for five priority pollutants at 10 to 30°C were measured with this technique and compared with literature values and with Henry’s constants measured in a bubble column. The results indicate that the technique is accurate. In addition, the technique is unencumbered by equilibration problems which limit the utility of other methods for determining Henry’s constant.