TA10 Microbial Remediation of Acid Mine Drainage Wastes
D135-136
8:00 AM - 12:00 PM, Tuesday
() Use of Respirometry to Determine Metal Toxicity Through Measurement of Hydrogen Consumption by Sulfate Reducing Bacteria in the Presence of Heavy Metals.
Tabak, H1, Holder, E1, Kupferle, M1, Haines, J1, 1 U.S. EPA
ABSTRACT- Acid mine drainage (AMD), an acidic metal-bearing wastewater poses a severe pollution problem attributed to post-mining activities. The metals (metal sulfates) encountered in AMD and considered of concern for risk assessment are: arsenic, cadmium, aluminum, manganese, iron, lead,, zinc and copper and the inorganic contaminant in the acidic waste is sulfate. A unique method was developed for the biotreatment of acid mine water present in the Berkeley Pit, Butte, Montana and other AMD containing pits throughout USA and abroad and for resource recovery of metals from such acidic mine wastes, The biotreatment of AMD and metal biorecovery and recycle process is based on complete separation of the biological sulfate reduction step due to the hydrogen consuming sulfate reducing bacteria (SRB) activity and the metal precipitation step. Hydrogen sulfide produced in the SRB membrane bioreactor systems is used in the metal sequential separation and precipitation step to form insoluble metal sulfides and hydroxides. USEPA research studies have shown that the heavy metals as ions or metal sulfides in the acidic wastes are toxic and inhibitory to SRB. It is thus essential to develop methods for determination of metal concentrations that are toxic and/or inhibitory to SRB which are active in sulfate bioconversion resulting in the productiun of hydrogen sulfide that is used in the metal separation and precipitation step for their recovery and recycle._CR__LF__CR__LF_A respirometric procedure was developed to determine the toxic and inhibitory effects of the heavy metals on the hydrogen consuming SRB by_CR__LF_quantitatively measuring the hydrogen utilization by SRB in the_CR__LF_presence of heavy metals encountered in AMD An enrichment culture of SRB was developed that is mixotrophic, utilizing C02 and acetate as the carbon source and H2 as the electron donor for the bioconversion of sulfate to hydrogen sulfide. Respirometers (NCON Systems, Inc.) were_CR__LF_adapted to measure H2 uptake by SRB in presence of various_CR__LF_concentrations of the metals. Hydrogen uptake by SRB is similar to the uptake of oxygen in aerobic cultures in that it causes the production of another gas, hydrogen sulfide under sulfate reducing condition. instead of carbon dioxide under aerobic conditions. The hydrogen sulfide gas produced by the sulfate reduction activity is removed with a zinc_CR__LF_acetate trap, analogous to an alkaline trap in the classical_CR__LF_respirometry tests, but allowing for the retension of CO2 in the_CR__LF_headspace of the culture flask. Respirometry can measure both_CR__LF_suppression of hydrogen uptake (toxicity) and/or increased lag time before hydrogen uptake begins (inhibition), relative to control system flasks without the target metal. Most bactriological media contain_CR__LF_components which form metal complexes and thus reducing metal_CR__LF_bioavailability to SRB. The culture media formulated by Sani et al (Advances in Environmental Research 5, )p.269-276.(2001) were modified_CR__LF_for use in the respirometric studies to quantitate the metal_CR__LF_toxicity/inhibition to H2 consuming SRB cultures. Metal complexation was significantly reduced by replacing orthophosphate with tryptone and PIPES for pH control._CR__LF__CR__LF_Respirometric studies were undertaken to measure toxicity and inhibition_CR__LF_of heavy metals, zinc, copper and iron to H2 consuming SRB._CR__LF_Resprirometric data on H2 uptake, analytical data on sulfate_CR__LF_bioconversion, and hydrogen sulfide production as well as biomass measurements indicate that zinc has an inhibitory effect on SRB between 10 and 25 ppm and that copper is inhibitory at 17 ppm. Concentration levels of these metals above those showing inhibitory effects to SRB, were shown to be toxic to SRB as indicated by complete suppression of H2 uptake in the respirometric culture systems. Iron , as ferrous or ferric species, was found not to be toxic at concentrations up to 4-00 ppm. In experiments combining Fe (200 ppm) with Cu , the sensitivity of the SRB to Cu was decreased so that inhibition occurred at 7ppm instead of 4ppm. Similar experiments combining Fe with Zn also showed a decrease of sensitivity of the SRB to Zn in that inhibition occurred at 15 ppm instead of 10 ppm._CR__LF__CR__LF_ Experiments with varied dilutions of AMD were also undertaken to determine at what dilutions the concentration levels of the heavy metals in AMD show inhibition to SRB. Studies with 1-100 dilution of AMD showed inhibition to SRB at 6 ppm Zn, 5 ppm Fe, 3 ppm Al, 2 ppm Cu and 2 ppm Mn concentration levels of the primary heavy metal components._CR__LF_
Key words: respirometry, metal toxicity, inhibition measurements