Abstract-A new, mechanistically based approach is presented for the quantitative determination of the uncoupling and inhibitory activity of compounds interfering with energy transduction. Time-resolved spectroscopy of single-turnover events in the photosystem of the photosynthetic bacterium Rhodobacter sphaeroides can quantitatively distinguish uncoupling from inhibition. The decay kinetics of the membrane potential after a single turnover flash are used as a measure of uncoupling activity, and the redox kinetics of several components of the electron transfer chain are used as indicators of specific inhibition at various potential inhibitory sites. Results are presented for 21 nitrated and chlorinated phenols, some reference uncouplers, and some anisoles. Inhibition was exclusively detected at one specific quinone binding site, the quinone reductase site Q i . For most phenols, uncoupling was observed at lower concentrations than inhibition with the exception of alkylated 2,6-dinitrophenols and 2,4,6-trichlorophenol, where both effects occurred in the same concentration range. No direct correlation was observed between the uncoupling and inhibitory activity of a given compound. The data obtained with this new method correlate well with data from various bioassays on energy-transducing systems, indicating that this method may also be well suited as a screening tool for compounds suspected to interfere with energy transduction.

The anaerobic biodegradation of hydrocarbons at mineral oil contaminated sites has gathered increasing interest as a naturally occurring remediation process. The aim of this study was to investigate biodegradation of hydrocarbons in laboratory aquifer columns in the absence of O 2 and NO 3 y , and to calculate a mass balance of the anaerobic biodegradation processes. The laboratory columns contained aquifer material from a diesel fuel contaminated aquifer. They were operated at 258C for 65 days with artificial groundwater that contained only SO 2y and CO 42 as externally supplied oxidants. After 31 days of column operation, stable concentration profiles were found for most of the measured dissolved species. Within 14 h residence time, about 0.24 mM SO 4 2y were consumed and dissolved Fe II Ž.Žup to 0.012 mM ,.Mn II Ž.Žup to 0.06 mM ,.and CH 4 Žup to 0.38 mM were produced. The alkalinity and the dissolved inorganic carbon .ŽDIC concentration . increased and the DIC became enriched in 13 C. In the column, n-alkanes were selectively removed while branched alkanes persisted, suggesting a biological degradation. Furthermore, based on changes of concentrations of aromatic compounds with similar physical-chemical properties in the effluent, it was concluded that toluene, p-xylene and naphthalene were degraded. A carbon mass balance revealed that 65% of the hydrocarbons removed from the column were recovered as DIC, 20% were recovered as CH 4 , and 15% were eluted from the column. The calculations indicated that hydrocarbon mineralization coupled to SO 4 2y reduction and methanogenesis contributed in equal proportions to the hydrocarbon removal. Hydrocarbon mineralization coupled to ) Corresponding author. Tel.: q41-1-633-6042; fax: q41-1-633-1122; e-mail: hoehener@ito.umnw.ethz.ch 1 Present address: BMG Engineering AG, CH-8952 Schlieren, Switzerland.Published in Journal of Contaminant Hydrology 32, issue 1-2, 1998 which should be used for any reference to this work Fe III Ž.and MnŽIV.reduction was of minor importance. DIC, alkalinity, and stable carbon isotope balances were shown to be a useful tool to verify hydrocarbon mineralization.

This study presents a stepwise concept to assess the in situ microbial mineralization of petroleum hydrocarbons (PHC) in aquifers. A new graphical method based on stable carbon isotope ratios (delta 13C) was developed to verify the origin of dissolved inorganic carbon (DIC). The concept and the isotope method were applied to an aquifer in Student, Switzerland, in which more than 34,000 liters of heating oil were accidentally released. Chemical analyses of ground water revealed that in this aquifer locally, anaerobic conditions prevailed, and that PHC mineralization was linked to the consumption of oxidants such as O2, NO3-, and SO4(2-) and the production of reduced species such as Fe2+, Mn2+, H2S and CH4. However, alkalinity and DIC balances showed a quantitative disagreement in the link between oxidant consumption and DIC production, indicating that chemical data alone may not be a reliable assessment tool. delta 13C ratios in DIC have been used before for bioremediation assessment, but results were reported to be negatively influenced by methanogenesis. Using the new graphical method to display delta 13C data, it was possible to identify anomalies found in methanogenic monitoring wells. It could be shown that 88% of the DIC produced in the contaminated aquifer originated from microbial PHC mineralization. Thus, the new graphical method to display delta 13C ratios appears to be a useful tool for the assessment of microbial hydrocarbon mineralization in a complex environment.

A new, mechanistically based approach is presented for the quantitative determination of the uncoupling and inhibitory activity of compounds interfering with energy transduction. Time‐resolved spectroscopy of single‐turnover events in the photosystem of the photosynthetic bacterium Rhodobacter sphaeroides can quantitatively distinguish uncoupling from inhibition. The decay kinetics of the membrane potential after a single turnover flash are used as a measure of uncoupling activity, and the redox kinetics of several components of the electron transfer chain are used as indicators of specific inhibition at various potential inhibitory sites. Results are presented for 21 nitrated and chlorinated phenols, some reference uncouplers, and some anisoles. Inhibition was exclusively detected at one specific quinone binding site, the quinone reductase site Qi. For most phenols, uncoupling was observed at lower concentrations than inhibition with the exception of alkylated 2,6‐dinitrophenols and 2,4,6‐trichlorophenol, where both effects occurred in the same concentration range. No direct correlation was observed between the uncoupling and inhibitory activity of a given compound. The data obtained with this new method correlate well with data from various bioassays on energy‐transducing systems, indicating that this method may also be well suited as a screening tool for compounds suspected to interfere with energy transduction.