Identification of bis-quaternary ammonium compounds using mild ionization mass spectrometry

Ethonium, an antimicrobial chemotherapeutic agent, was investigated by mass spectrometry (MS) under various ionization conditions: electron impact, field ionization, field desorption (FD) and fast atom ionization. FDMS was found to be the most suitable procedure for ethonium identification. Relation of the ED mass spectra to the distance between the nitrogen atoms in bis-quaternary ammonium compounds is discussed. It was shown that the most intensive ions with m/z 499, 315 in the FD mass spectra corresponded to the ethonium specific fragmentation and their occurrence in the spectra could serve as a sufficient criterion useful in qualitative and quantitative assay of the drug in the sample.     

Synthesis of a quaternary polyprenyl ammonium salt

Reaction of primary C(55)-allylic alcohol moraprenol (WT(3)C(7-9)-OH, a polyprenol from mulberry leaves) with triethylamine in the presence of phosphorus oxychloride leads to a quaternary ammonium chloride with a good yield (72%) and high cis-stereoselectivity of the terminal isoprene unit. Cationic polyprenyl derivatives may be useful for transfection and immunological studies.     

Production of nitrate spikes in a model of ammonium biodegradation

Nitrification at the site of a contaminant ammonium plume from a former coal carbonisation plant can be modelled with three competing bacterial populations of Nitrosomonas, Nitrobacter, and Brocadia anammoxidans. Oscillations of chemical species at the site can be explained by a reduced model of ammonium competition between Nitrosomonas and B. anammoxidans which effectively acts as an activator-inhibitor system. Stable oscillations occur in conditions of low nutrient (ammonium) supply and this causes a spatial travelling wave in a borehole profile when diffusion is introduced.     

Enhanced biodegradation of phenanthrene in a biphasic culture system

Degradation of phenanthrene byPseudomonas aeruginosa AK1 was examined in (i) an aqueous mineral salts medium to which phenanthrene particles of varying size (i.e. diameter) were added, and (ii) an aqueous/organic biphasic culture system consisting of mineral salts medium supplemented with 2,2,4,4,6,8,8-heptamethylnonane (HMN) as the phenanthrene-carrying organic phase. In both systems, the rate of phenanthrene biodegradation could be significantly enhanced by manipulations leading to improved phenanthrene mass transfer into the aqueous phase. With crystalline phenanthrene, the rate of biodegradation was found to be directly correlated to the particle surface area, whereas in the biphasic system the rate of biodegradation of the dissolved phenanthrene was mainly governed by the HMN/water interface area. In the latter system, exponential growth with a doubling time t_d of 6–8 hours has been achieved under conditions of intensive agitation of the medium indicating that phenanthrene degradation by strain AK1 is limited mainly by physicochemical parameters. Addition of selected surfactants to the culture medium was found to accelerate phenanthrene degradation by strain AK1 only under conditions of low agitation (in the presence of HMN) and after pretreatment of phenanthrene crystals by ultrasonication (in the absence of HMN). Evidence is presented that the stimulating effect of the surfactants was primarily due to improved dispersion of phenanthrene particle agglomerates (in the aqueous mineral salts medium supplemented with phenanthrene crystals) or of the phenanthrene-carrying lipophilic solvent drops (in the aqueous/organic biphasic culture system) whereas the solubilizing activity towards phenanthrene was neglectible. Under conditions of intensive mixing of the culture medium (i.e. if a high particle surface area or HMN/water interface area, respectively, is provided), the addition of surfactants did not enhance phenanthrene biodegradation.     

Enhanced hydrocarbon biodegradation by a newly isolated Bacillus subtilis strain

The relation between hydrocarbon degradation and biosurfactant (rhamnolipid) production by a new Bacillus subtilis 22BN strain was investigated. The strain was isolated for its capacity to utilize n-hexadecane and naphthalene and at the same time to produce surface-active compound at high concentrations (1.5 - 2.0 g l(-1)). Biosurfactant production was detected by surface tension lowering and emulsifying activity. The strain is a good degrader of both hydrocarbons used with degradability of 98.3 +/- 1% and 75 +/- 2% for n-hexadecane and naphthalene, respectively. Measurement of cell hydrophobicity showed that the combination of slightly soluble substrate and rhamnolipid developed higher hydrophobicity correlated with increased utilization of both hydrocarbon substrates. To our knowledge, this is the first report of Bacillus subtilis strain that degrades hydrophobic compounds and at the same time produces rhamnolipid biosurfactant.     

Primary biodegradation of amine oxide and quaternary ammonium amphiphiles

Biodegradation of two amphiphilic “soft” antimicrobially active derivatives of lauric (dodecanoic) acid, a quaternary ammonium salt and an amine oxide bearing an amide or ester group, was followed using microorganisms from activated sludge. Primary biodegradation was determined by ion-selective electrodes, total biodegradation as the chemical oxygen demand. Though organic ammonium salts quickly undergo primary biodegradation, the rest of the molecule is difficult to destroy. In contrast, amine oxides are easily biodegradable.     

Mutants of Saccharomyces cerevisiae resistant to a quaternary ammonium salt

Three quaternary ammonium salts were compared in respect of their ability to select resistant mutants of S. cerevisiae. The mutants tolerating slightly higher IM compound concentration were analysed. They appeared to be the products of nuclear gene mutation segregating monogenically but strongly influenced by genetic background. The mutant IMR when transformed to rho degrees lost resistance below the level of minimal inhibitory concentration of original strain. Possible hypothesis explaining this phenomenon is presented.     

Enhanced biodegradation of phenanthrene by a marine bacterium in presence of a synthetic surfactant

The biodegradation of phenanthrene by the marine strain Sphingomonas sp. 2MPII (DSMZ 11572) was enhanced by the solubilizating properties of the nonionic surfactant Tween 80. After 197 h of incubation, 85 +/- 4% of the initial amount of phenanthrene (0.4 g l-1) was biodegraded in presence of Tween 80 (0.5 g l-1) as opposed to 52 +/- 5% without this synthetic surfactant. These results confirm that the activity of the strain 2MPII is limited by the bioavailability of the polycyclic aromatic hydrocarbon (PAH) substrate in the aqueous phase. Tween 80 appears to be efficient in increasing the bioavailability of hydrophobic compounds such as PAHs.     

Enhanced dimethyl phthalate biodegradation by accelerating phthalic acid di-oxygenation

The aerobic biodegradation of dimethyl phthalate (DMP) is initiated with two hydrolysis reactions that generate an intermediate, phthalic acid (PA), that is further biodegraded through a two-step di-oxygenation reaction. DMP biodegradation is inhibited when PA accumulates, but DMP’s biodegradation can be enhanced by adding an exogenous electron donor. We evaluated the effect of adding succinate, acetate, or formate as an exogenous electron donor. PA removal rates were increased by 15 and 30% for initial PA concentrations of 0.3 and 0.6 mM when 0.15 and 0.30 mM succinate, respectively, were added as exogenous electron donor. The same electron-equivalent additions of acetate and formate had the same acceleration impacts on PA removal. Consequently, the DMP-removal rate, even PA coexisting with DMP simultaneously, was accelerated by 37% by simultaneous addition of 0.3 mM succinate. Thus, lowering the accumulation of PA by addition of an electron increased the rate of DMP biodegradation.     

Enhanced octadecane dispersion and biodegradation by a Pseudomonas rhamnolipid surfactant (biosurfactant).

A microbial surfactant (biosurfactant) was investigated for its potential to enhance bioavailability and, hence, the biodegradation of octadecane. The rhamnolipid biosurfactant used in this study was extracted from culture supernatants after growth of Pseudomonas aeruginosa ATCC 9027 in phosphate-limited proteose peptone-glucose-ammonium salts medium. Dispersion of octadecane in aqueous solutions was dramatically enhanced by 300 mg of the rhamnolipid biosurfactant per liter, increasing by a factor of more than 4 orders of magnitude, from 0.009 to > 250 mg/liter. The relative enhancement of octadecane dispersion was much greater at low rhamnolipid concentrations than at high concentrations. Rhamnolipid-enhanced octadecane dispersion was found to be dependent on pH and shaking speed. Biodegradation experiments done with an initial octadecane concentration of 1,500 mg/liter showed that 20% of the octadecane was mineralized in 84 h in the presence of 300 mg of rhamnolipid per liter, compared with only 5% octadecane mineralization when no surfactant was present. These results indicate that rhamnolipids may have potential for facilitating the bioremediation of sites contaminated with hydrocarbons having limited water solubility.     

Enhanced anaerobic biodegradation of benzene-toluene-ethylbenzene-xylene-ethanol mixtures in bioaugmented aquifer columns

Methanogenic flowthrough aquifer columns were used to investigate the potential of bioaugmentation to enhance anaerobic benzene-toluene-ethylbenzene-xylene (BTEX) degradation in groundwater contaminated with ethanol-blended gasoline. Two different methanogenic consortia (enriched with benzene or toluene and o-xylene) were used as inocula. Toluene was the only hydrocarbon degraded within 3 years in columns that were not bioaugmented, although anaerobic toluene degradation was observed after only 2 years of acclimation. Significant benzene biodegradation (up to 88%) was observed only in a column bioaugmented with the benzene-enriched methanogenic consortium, and this removal efficiency was sustained for 1 year with no significant decrease in permeability due to bioaugmentation. Benzene removal was hindered by the presence of toluene, which is a more labile substrate under anaerobic conditions. Real-time quantitative PCR analysis showed that the highest numbers of bssA gene copies (coding for benzylsuccinate synthase) occurred in aquifer samples exhibiting the highest rate of toluene degradation, which suggests that this gene could be a useful biomarker for environmental forensic analysis of anaerobic toluene bioremediation potential. bssA continued to be detected in the columns 1 year after column feeding ceased, indicating the robustness of the added catabolic potential. Overall, these results suggest that anaerobic bioaugmentation might enhance the natural attenuation of BTEX in groundwater contaminated with ethanol-blended gasoline, although field trials would be needed to demonstrate its feasibility. This approach may be especially attractive for removing benzene, which is the most toxic and commonly the most persistent BTEX compound under anaerobic conditions.     

Minor groove binding of a bis-quaternary ammonium compound: the crystal structure of SN 7167 bound to d(CGCGAATTCGCG)2.

The X-ray crystal structure of the complex between the synthetic antitumour and antiviral DNA binding ligand SN 7167 and the DNA oligonucleotide d(CGCGAATTCGCG)2 has been determined to an R factor of 18.3% at 2.6 A resolution. The ligand is located within the minor groove and covers almost 6 bp with the 1-methylpyridinium ring extending as far as the C9-G16 base pair and the 1-methylquinolinium ring lying between the G4-C21 and A5-T20 base pairs. The ligand interacts only weakly with the DNA, as evidenced by long range contacts and shallow penetration into the groove. This structure is compared with that of the complex between the parent compound SN 6999 and the alkylated DNA sequence d(CGC[e6G]AATTCGCG)2. There are significant differences between the two structures in the extent of DNA bending, ligand conformation and groove binding.     

Enhanced oily sludge biodegradation by a tensio-active agent isolated from Pseudomonas aeruginosa USB-CS1

The biodegradation of an oily sludge is facilitated by a microbial tensio-active agent isolated from Pseudomonas aeruginosa USB-CS1. The optimal oil-in-water dispersion conditions are as follows: pH 6.5, temperature 30 °C, agitation 150 rev/min. The total hydrocarbon content shows that the biodegradation of the oily substrate mediated by the biosurfactant or by the biosurfactant–P. aeruginosa USB-CS1 complex is significantly higher after 30 days of incubation than that in other experimental conditions, by a mean of 70%. Substrate fractionation by column chromatography reveals that, if biosurfactant is present, saturated and aromatic compounds are more susceptible to microbial degradation than they are in other biodegradation systems by an average of 55% and 40% respectively. These results suggest that the stimulatory effects of the biosurfactant on the biodegradation of the oily substrate are limited over time by the loss of surface activity of the biosurfactant after 30 days of incubation. Received : 7 August 1996 / Received revision : 6 December 1996 / Accepted : 4 January 1997     

Enhanced biodegradation of methoxychlor in soil under sequential environmental conditions.

Ring-U-[14C]methoxychlor [1,1-bis(p-methoxyphenyl)-2,2,2-trichloroethane] was incubated in soil under aerobic and anaerobic conditions. Primary degradation of methoxychlor occurred under anaerobic conditions, but not under aerobic conditions, after 3 months of incubation. Analysis of soil extracts, using gas chromatography, demonstrated that only 10% of the compound remained at initial concentrations of 10 and 100 ppm (wt/wt) of methoxychlor. Evidence is presented that a dechlorination reaction was responsible for primary degradation of methoxychlor. Analysis of soils treated with 100 ppm of methoxychlor in the presence of 2% HgCl2 showed that 100% of the compound remained after 3 months, indicating that degradation in the unpoisoned flasks was biologically mediated. Methanogenic organisms, however, are probably not involved, as strong inhibition of methane production was observed in all soils treated with methoxychlor. During the 3-month incubation period, little or no evaluation of 14CO2 or 14CH4 occurred under either aerobic or anaerobic conditions. Cometabolic processes may be responsible for the extensive molecular changes which occurred with methoxychlor because the rate of its disappearance from soil was observed to level off after exhaustion of soil organic matter. After this incubation period, soils previously incubated under anaerobic conditions were converted to aerobic conditions. The rates of 14CO2 evolution from soils exposed to anaerobic and aerobic sequences of environments ranged from 10- to 70-fold greater than that observed for soils exposed solely to an aerobic environment.     

Effect of salt on aerobic biodegradation of petroleum hydrocarbons in contaminated groundwater

Hydrocarbon-contaminated soil and groundwater at oil and gas production sites may be additionally impacted by salts due to release of produced waters. However, little is known about the effect of salt on the in-situ biodegradation of hydrocarbons by terrestrial microbes, especially at low temperatures. To study this effect, we prepared a groundwater-soil slurry from two sites in Canada: a former flare pit site contaminated with flare pit residue (Site A), and a natural gas processing facility contaminated with natural gas condensate (Site B). The slurry with its indigenous microbes was amended with radiolabeled hydrocarbons dissolved in free product plus nutrients and/or NaCl, and incubated in aerobic biometer flasks with gyrotory shaking at either 25 or 10°C for up to 5 weeks. Cumulative production of ¹⁴CO₂ was measured and the lag time, rate and extent of mineralization were calculated. For Site A, concentrations of NaCl ≥1% (w/v) delayed the onset of mineralization of both ¹⁴C-hexadecane and ¹⁴C-phenanthrene under nutrient-amended conditions, but once biodegradation began the degradation rates were similar over the range of salt concentrations tested (0–5% NaCl). For Site B, increasing concentrations of NaCl ≥1% (w/v) increased the lag time and decreased the rate and extent of mineralization of aliphatic and aromatic substrates. Of particular interest is the observation that low concentrations of salt (≤1% NaCl) slightly stimulated mineralization in some cases.     

Enhanced biodegradation of methylhydrazine and hydrazine contaminated NASA wastewater in fixed-film bioreactor

The aerobic biodegradation of National Aeronautics and Space Administration (NASA) wastewater that contains mixtures of highly concentrated methylhydrazine/hydrazine, citric acid and their reaction product was studied on a laboratory-scale fixed film trickle-bed reactor. The degrading organisms, Achromobacter sp., Rhodococcus B30 and Rhodococcus J10, were immobilized on coarse sand grains used as support-media in the columns. Under continuous flow operation, Rhodococcus sp. degraded the methylhydrazine content of the wastewater from a concentration of 10 to 2.5 mg/mL within 12 days and the hydrazine from 0.8 to 0.1 mg/mL in 7 days. The Achromobacter sp. was equally efficient in degrading the organics present in the wastewater, reducing the concentration of the methylhydrazine from 10 to 5 mg/mL within 12 days and that of the hydrazine from 0.8 to 0.2 mg/mL in 7 days. The pseudo first-order rate constants of 0.137 day^-1 and 0.232 day^-1 were obtained for the removal of methylhydrazine and hydrazine, respectively, in wastewater in the reactor column. In the batch cultures, rate constants for the degradation were 0.046 and 0.079 day^-1 for methylhydrazine and hydrazine respectively. These results demonstrate that the continuous flow bioreactor afford greater degradation efficiencies than those obtained when the wastewater was incubated with the microbes in growth-limited batch experiments. They also show that wastewater containing hydrazine is more amenable to microbial degradation than one that is predominant in methylhydrazine, in spite of the longer lag period observed for hydrazine containing wastewater. The influence of substrate concentration and recycle rate on the degradation efficiency is reported. The major advantages of the trickle-bed reactor over the batch system include very high substrate volumetric rate of turnover, higher rates of degradation and tolerance of the 100% concentrated NASA wastewater. The results of the present laboratory scale study will be of great importance in the design and operation of an industrial immobilized biofilm reactor for the treatment of methylhydrazine and hydrazine contaminated NASA wastewater.     

Amino acid auxotrophy increases sensitivity of Saccharomyces cerevisiae to a quaternary ammonium salt IM.

Relationships between amino acid auxotrophy and N-dodecyloxy-carboxy-methyl-N-N-N-trimethyl ammonium chloride (IM) sensitivity have been investigated in isogenic yeast strains Saccharomyces cerevisiae and their meiotic segregants. It has been found, that auxotrophy increases the level of sensitivity to this salt markedly. A gene conferring resistance to that drug cancels the auxotrophy-dependent sensitivity.