EPA-600/2-76-241 October 1976 Environmental Protection Technology Series AOUTE TOXIC EFFECTS OF PETROLEUM REFINERY WASEWAIERS ON Robert S. Kerr Environmental Research Laboratory ifftee «l Researcfc ------- RESEARCH REPORTING SERIES Research reports of the Office of Research and Development, U.S. Environmental Protection Agency, have been grouped into five series. These five broad categories were established to facilitate further development and application of environmental technology. Elimination of traditional grouping was consciously planned to foster technology transfer and1 a maximum interface in related fields. The five series are: 1. Environmental Health Effects Research 2. Environmental Protection Technology 3. Ecological Research 4. Environmental Monitoring 5. Socioeconomic Environmental Studies This report has been assigned to the ENVIRONMENTAL PROTECTION TECHNOLOGY series. This series describes research performed to develop and demonstrate instrumentation, equipment, and methodology to repair or prevent environmental degradation from point and non-point sources of pollution. This work provides the new or improved technology required for the control and treatment of pollution sources to meet environmental quality standards. This document is available to the public through the National Technical Informa- tion Service, Springfield, Virginia 22161. ------- EPA-600/2-76-241 October 1976 ACUTE TOXIC EFFECTS OF PETROLEUM REFINERY WASTEWATERS ON REDEAR SUNFISH by John E. Matthews Leon H. Myers Robert S. Kerr Environmental Research Laboratory Ada, Oklahoma 74820 ROBERT S. KERR ENVIRONMENTAL RESEARCH LABORATORY OFFICE OF RESEARCH AND DEVELOPMENT U.S. ENVIRONMENTAL PROTECTION AGENCY ADA, OKLAHOMA 74820 , • ->; , T /8 F J /'i ..,^'.,,,, iioy/ii iwyw ------- DISCLAIMER This report has been reviewed by the Robert S. Kerr Environmental Research Laboratory, U.S. Environmental Protection Agency, and approved for publication. Mention of trade names or commercial products does not constitute endorsement or recommendation for use. 11 ------- ABSTRACT Static bioassays of 24 hours' duration were performed on samples of wastewaters provided by 22 domestic petroleum refiners. These waste- waters! represent three types of water discharges prevalent to this industry: process wastewaters prior to dilution with other streams; API separator effluents which are a conjugate of various streams within a refinery; and wastewaters following treatment by activated sludge systems. Bioassays were performed using redear sunfish (Lepomis microlophus) as test organisms. Selection of this species was due to its being a member of the important freshwater family, Centrarchidae, its wide- spread distribution, and the availability of a plentiful supply with a controlled background. Twenty-four hour 50 percent tolerance limits (TL ) of the various wastewaters are compared with results of chemical analyses performed during the same study. Toxicity varied considerably both between refineries and for waste streams from within a single refinery. Results of these analyses and observed behavioral symptoms of distressed fish revealed that ammonia, sulfides, and phenolics, alone or in combination, were major contributors to toxicity exerted in most samples. Three refineries had samples which were more toxic than anticipated based on results of chemical analyses, indicating the presence of other toxic compounds in unknown quantities; e.g., various hydrocarbons. Results of bioassays of samples from various wastewater treatment stages of five refineries with activated sludge treatment systems revealed a decrease in toxicity as the degree of treatment increased indicating a reduction in concentration of contributing biodegradeable toxicants. This report was submitted in fulfillment of ROAP 21 AZP, Task 050 by the Robert S. Kerr Environmental Research Laboratory of the U. S. Environmental Protection Agency. Work was completed as of January 1974. 111 ------- CONTENTS Sections Page I Conclusions 1 II Recommendations 3 III Introduction 4 IV Methods 7 V Results 11 VI Discussion 20 VII References 29 VIII Appendices 31 ------- ACKNOWLEDGMENTS The U. S. Environmental Protection Agency, Robert S. Kerr Environmental Research Laboratory (RSKERL), gratefully acknowledges the cooperation of the Bureau of Sports Fisheries and Wildlife, National Fish Hatchery, Tishomingo, Oklahoma, in providing experimental organisms for use in this project. The following people at the RSKERL are recognized for their assistance in conducting these experiments: Tommy Redman, James McNabb, Bob Newport, Kenneth Jackson, and Roger Cosby. Particular appreciation is extended to Dr. William A. Brungs, National Water Quality Laboratory, Duluth, Minnesota; Mr. David N. Peters, Region VI, Dallas, Texas; and Mr. Marvin Wood, RSKERL, for their consultation and suggestions during the study and review of the experimental results. VI ------- SECTION I CONCLUSIONS Short-term static bioassays of 24-hours1 duration can be an effective tool for screening industrial process wastewaters to locate sources of toxic agents; these tests can also be used to evaluate effectiveness of industrial waste treatment processes. Static bioassays cannot be used to obtain reliable 50 percent tolerance limit (TL™) values for low toxicity wastes that exert a high oxygen O \J demand although a range can often be established by an experienced observer. Dissolved oxygen (DO) usually becomes critical at about 12 hours; low DO plays a major role in a mortallity after this time. Activated sludge treatment processes tend to reduce the oxygen demand beyond the critical stage. Raw wastewaters from different oil refineries vary greatly in their toxic characteristics; wastewaters from different processes within a single refinery also vary greatly in their toxic characteristics. Toxicity of oil refinery wastewaters varies considerably at different treatment stages in the activated sludge process; toxicity appears to decrease following each stage of treatment. Toxicity of oil refinery wastewaters cannot always be predicted from results of chemical analyses; the toxic effect of the waste is dependent on the synergistic or antagonistic activity of toxicants present. The most common toxic constituents of untreated oil refinery waste- waters are: ammonia, sulfides, phenolic compounds, and cyanides. Raw wastewaters also may contain other toxic compounds including various hydrocarbons. ------- Due to the volatile and unstable nature of some toxic components of oil refinery wastewaters which may have led to a reduction in concen- trations during sample transportation, storage, and handling, TL,_0 values obtained during these tests may be higher than the actual value. Acute toxic effects of raw wastewaters from oil refineries are generally exerted within the first 12 hours of the static test; therefore, a 24-hour test will provide good positive results under static conditions. If samples containing toxic compounds, other than those mentioned above, toxic effects may be exerted over an extended period. Tests should then be continued for at least 48 hours to obtain more positive results. Toxicity of the final clarifier effluent from oil refineries with activated sludge treatment systems is dependent on toxic constituents present in the influent and their concentrations. Results of chemical analyses conducted during these tests indicate that concentrations of most toxicants other than ammonia are reduced by activated sludge treat- ment systems and at least a four-fold decrease in toxicity can be ex- pected after treatment. Although activated sludge treatment systems have very little effect on ammonia concentrations, the toxic effect of ammonia is lessened due to a decrease in pH of the treated effluent. ------- SECTION II RECOMMENDATIONS With the many and varied processes employed by oil refineries in the refining of crude oil, the results of this study and others indicate the need to further document the effect of treated refinery wastewaters and individual wastewater constituents on fish and other aquatic life in receiving waters under a wide range of environmental conditions. This project was conducted to determine short-term acute toxic effects of refinery wastewaters on a single fish species. A research study to investigate long-term cumulative effects of treated refinery waste- waters on fish and other aquatic life should be conducted. The oil refining industry should be encouraged to use short-term static bioassays to assist in their pollution control effort. Some of the ways in which this type of bioassays can be used are: 1. Acute toxicity of final effluents can be determined. 2. Toxicity of individual process wastewaters can be determined. These effluents, if highly toxic, may be modified, eliminated, or treated. 3. Location of processes which contribute high toxicity waste- waters make possible alternative methods of controlling the potential problem. The smaller quantities of wastewater may be pretreated to reduce toxicity or stored with regulated release into the refinery wastewater treatment system. 4. Leaks, spills, or other losses of toxic compounds into waste streams can be detected, located, and remedied. 5. The effectiveness of treatment systems on removing toxic compounds can be established. ------- SECTION III INTRODUCTION Industrial wastes may be detrimental to aquatic life by direct toxic action or by altering natural qualities of the receiving water. Petro- leum refinery wastewaters can be characterized as having a high bio- chemical oxygen demand (BOD) and significant toxicity.1 Adverse effects from acute concentrations of refinery effluents during several days of 2 exposure have been reported. Experiments with five species of fish have shown that the emulsified effluent from American Petroleum Insti- tute's (API) gravity oil separators is toxic to fish unless diluted or otherwise treated. Treatment of refinery effluents to reduce tox- icity also results in improvement in other objectionable characteristics including oil content, oxygen demand, chemical composition, turbidity, 4 and odor. products and wastes from oil refineries are extremely complex in their composition and their effects on water quality. While it is recognized that each particular oil refinery poses its individual problems, certain pollutants are common to all refineries. Process wastewaters are high in phenolics, ammonia, and hydrogen sulfide and pose the primary water pollution problem. All of these compounds are relatively high in toxicity to aquatic life. In addition to these common constituents, process wastewaters may contain various hydrocarbons toxic to aquatic life. ' Findings during a cooperative American Petroleum Institute/ Environmental Protection Agency (API/EPA) waste characterization study of petroleum refinery wastewaters conducted at the Robert S. Kerr Environmental Research Laboratory (RSKERL) concurrently with this ------- project indicate that these wastewaters may contain a large number of substances which if discharged without treatment into receiving g waters could pose a serious threat to fish and other aquatic life. It was also found, however, that refinery treatment processes sub- stantially reduce the acute toxic effect of these substances. Refinery wastewaters often contain a number of these different toxi- cants which, when mixed or under the influence of other characteristics of the effluent or receiving water, may produce an entirely different toxicity level from that of the pure components. Results of chemical analyses for possible toxic components compared with toxicity levels reported in the literature for each particular component can not always be expected to produce satisfactory answers. The concentration of these pollutants in a particular waste stream is dependent on the process from whichT the stream flows as well as in- process procedures for reduction of a potential troublesome pollutant. The concentration in the final effluent is dependent on the type and efficiency of treatment to which the wastewater is subjected. The use of some type of bioassay to determine the toxicity of a complex wastewater can be the most effective and accurate method of assessing potential danger. The bioassay provides a direct satisfactory method for evaluating acute toxicity of chemically complex wastes. Toxicity determinations rather than complete reliance on BOD and chemical determinations are necessary for the safe disposal of complex wastes. Two phases of a study of oil refinery wastewaters were conducted at the RSKERL during 1972. During" Phase I of the study, samples were collected by EPA and API personnel from in-plant wastewaters at 17 selected refineries located throughout the United States and shipped to the RSKERL for analysis of 28 parameters. A single set of samples was received for each of the refineries. Samples were preserved in tightly capped cubitainers and refrigerated at 4° C until tested. The ------- number of sample points tested for each refinery ranged from 1 to 13. During Phase II of the study, samples were collected from various stages of the activated sludge treatment process at five selected oil refineries. Samples were collected for 14 consecutive days at each refinery. On March 28, 1972, a series of 24-hour static bioassays were started at the RSKERL to screen samples collected during Phase I of the study. The purpose of these tests were (1) to evaluate and compare toxicity of in-plant wastewaters of selected oil refineries throughout the United States; (2) to compare actual toxicity results with anticipated toxic action derived from results of chemical analyses of the samples for potential toxicants normally contained in process wastewaters; (3) to check for the presence of unknown toxic compounds; and (4) to observe and catalog behavioral activities of fish exposed to these wastewaters. On May 10, 1972, a series of 24-hour static bioassays were started on samples collected during Phase II of the study. Two or three bioassays were conducted on composites of the 14 samples collected at each loca- tion. The purpose of these tests were (1) to evaluate and compare toxicity at different stages of treatment of selected refineries with activated sludge treatment systems; and (2) to evaluate the efficiency of activated sludge treatment for removal of toxic constituents from refinery wastewaters. ------- SECTION IV METHODS Bioassay procedures used were modified from those described in Standard 9 Methods for the Examination of Water and Wastewater, 13th Edition. Modifications were necessary because of the small volume of sample available for use, the number of samples to be tested, and the resources available. Test procedures outlined in this paper can be expected to measure short-term effects only. The long-term effect is not measured. Thus, there may be a cumulative effect of the waste over a longer period of time. A series of three to five dilutions and a control were set up for each sample or composite tested. Test concentrations were prepared on a percent-by-volume basis in the diluent water. Two, one-gallon glass jars containing three and one-half liters in each or one, five-gallon glass jar containing ten liters were used per dilution. Five test organisms were placed in each jar after the necessary volume of sample was mixed with the diluent water. This gave either ten fish in seven liters per dilution or five fish in ten liters per dilution. The latter was used to try to alleviate problems of low oxygen in some of the tests and to ensure that stress from crowded conditions did not affect the test results. Test organisms used were redear sunfish (Lepomis microlophus) that were 20-50 millimeters in length and weighing 0.5-1.5 gram each. Selection of this species was due to its being a member of the important family Centrarchidae, its widespread distribution, and the availability of a plentiful supply with a controlled background. This species was easily maintained under laboratory conditions and ------- proved highly satisfactory for conducting short-term static bioassays. Fish were obtained from the National Fish Hatchery, Tishomingo, Oklahoma, as needed. Fish were held in stainless steel holding tanks at a constant temperature and saturated dissolved oxygen 16vel until used. Only fish that appeared to be in excellent condition were used in the testing. The dilution range selected for each sample was based on chemical data for that sample compared with reported toxicity values for measured chemical constituents. If fish died in all dilutions for a test, it was assumed that unknown toxic agents were present in the sample and lower series of dilutions were tested until a TL^ value was obtained. Logrithmic or geometric series of dilutions were used in all tests. Dilution water in all tests was tap water from the Ada, Oklahoma, water supply system (pH 8.3-8.7, Hardness 300-350 mg/1 as CaCO_). O Use of a common dilution water made it possible to compare toxicity levels of samples from different refineries. Dilution water in each test container was aerated to saturation prior to adding the sample and test organisms. Aeration was not used during the test period because of the volatile nature of many of the constituents of the wastewaters. Preliminary tests indicated that most toxicity of oil refinery wastes in static tests is exerted during the first 12 hours. This may be due to the volatile nature of many of the toxic constituents. Per- sonal communication with other scientist who have tested similar wastes confirmed this. ' Therefore, 24 hours was chosen as the test period. The 24-hour test period tended to eliminate low DO as a contributor to death in latter stages of the tests during Phase I. Low DO during the latter stages was still a problem during Phase II because of the higher waste volumes used in the tests. ------- Bioassay record sheets were maintained for each sample tested. Obser- vations and selected analysis were made initially at six hours, twelve hours and twenty-four hours. Survival percentage was recorded at each dilution. The 24-hour survival percentage for each dilution was plotted on semi-log paper and a straight line connected between successive dilution percentages at which greater than and less than 50 percent survival occurred (Figure 1). The 24-hour TL^ value (percentage dilution at which 50 percent of test organisms survived for 24 hours under conditions of the test) was then interpolated from the straight line. All TL^ values were calculated in terms of the initial percentage volume of wastewater; undoubtedly, effective concen- trations of the various toxic constituents became lower with time. If more than 10 percent of the control organisms had died during a test, results for that test would have been invalidated. Within test precision was checked by plotting survival percentages and establishing a TL5Q individually for the two series of test containers used for each test (Table 1 and Figure 1). Table 1. WITHIN TEST CHECK OF PRECISION Dilution Test Containers A B Total 10% 5 5 10 5.6% 5 5 10 3.2% 5 5 10 1.8% 5 5 10 1.0% 5 5 10 Control 5 5 10 ------- 1000 800 600 400 200 100(5) 80 60 40 20 10 0 Log Scale 1000 Sample No. 6992:RW-5 Starting Date: 5-29-72 Hour: 9:00 a.m. Final Results: rL50 Time Intervals A-24 0.042 B-24 0.045 24 hr 0.043 320 Concentrations expressed as percent 0.1% -> 0.01% 180 0.100 0.056 0.018 OOIO 50 Percent Survival Test species: Redear Sunfish Temperature range: 21.5°-25°C Dilution water source: Ada City Water Notes: WilH t;wi mmi no and loss of equilibrium noted at 5 minutes in 0.1% dilution. All fish dead in 50 minutes. Eight fish dead in 2 hours in in 0.056% dilution. Two live fish in obvious distress. Tests continued for 48 hours with no change in TL value. John E. Matthews 100 Bioassay Concentrations A-Front row of test containers B-Back row of test containers Figure 1. Bioassay record sheet 10 ------- SECTION V RESULTS PHASE I Bioassays were conducted on 50 samples of in-plant wastewaters from 17 oil refineries during this phase of the study. The 24-hour TL5Q results are presented in Table 2. Because only a limited amount of each sample was available for use, 24-hour TL5Q values were not obtained for all samples tested. The 24-hour TLg for those samples exhibiting greater than 50 percent survival in all dilutions tested was reported as greater than the highest dilution tested. A 24-hour TL™ ranging from 0.04%-13.5% was established for 20 samples from eight refineries under the conditions of these tests. Bioassay data for the toxic samples are presented in Appendix A. No 24-hour TL™ could be established for the remaining 30 samples. Nine refineries had no samples which exhibited toxicity under these conditions at the dilutions tested. Three refineries had one or more samples toxic at <0.3%. These same refineries also had at least one other sample toxic at <5%. Three additional refineries had one or more samples toxic at <5%. The remaining two refineries for which a 24-hour TL,.., was established had single in-plant wastewater streams with a TL,-n of 7.5% and 13.5%. Three refineries had one or more samples which were considerably more toxic than indicated by results of chemical analyses. Toxicity of these samples was most likely due to the presence of toxic hydrocarbon compounds. The TL,_n for one sample was attributed to low DO and one sample to low pH. 11 ------- Table 2. BIOASSAY RESULTS - PHASE I Sample 6992 RW-1 11 RW-2 " RW-3 11 RW-4 11 RW-5 11 RW-6 11 RW-7 11 RW-8 " RW-9 " RW-10 11 RW-11 " RW-12 " RW-13 59991 RW-1 69991 RW-1 693 RW-1 " RW-2 " RW-3 69995 RW-1 11 RW-2 22 RW-1 " RW-2 69994 RW-1 " RW-2 6993 RW-1 24-hour TL5Q (% by volume) Comment >10 0.042 Same at 48 hours 0.043 Same at 48 hours >10 >10 >18 >15 >10 >18 0.05 Same at 48 hours 3.8 3.2% at 48 hours >10 >10 7.5 13.5 0.23a .08% at 36 hours 1.8 7.5 Low DO >10 >10 >18 >18 4.5 1.9 Low pH >10 12 ------- Table 2 (Continued). BIOASSAY RESULTS - PHASE I Sample 595 it 6995 ti it u II II II It II It 92 u 59 59993 u 106 56 67 6996 u u u ti RW-1 RW-2 RW-1 RW-2 RW-3 RW-4 RW-5 RW-6 RW-7 RW-8 RW-9 RW-10 RW-1 RW-2 RW-1 RW-1 RW-2 RW-1 RW-1 RW-1 RW-1 RW-2 RW-3 RW-4 RW-5 24-hour TL (% by volume) Comment 10 18 6.5 18 13 Distress 13,5 Distress 4.2 7.2 14 16 15 18 10 2.4 10 10 10 18 10 10 10 10 4.2 2.4% at 48 hours 4.2 0.05 Toxicity greater than indicated by results of chemical analyses. 13 ------- Estimated concentrations at the TL^ value were made for common toxic constituents found in refinery wastewaters for purposes of identifying the most likely toxic agent(s) in each sample (Table 3). These esti- mations were made using results of analyses performed on corresponding water samples. Results upon which these estimations are based are presented in Appendix C. These estimations cannot be assumed to be actual concentrations present at the measured TL • however, analyses of water samples from some of the bioassay test containers after 24 hours indicate that the estimated values are close enough approximations of the real values to use for the purposes intended. These projected data along with the actual measurements confirm that ammonia, sulfides, and phenolic compounds are the major contributing toxic constituents present in oil refinery wastewaters. Fourteen of the samples for which a TL,_n was established during this study con- tained these constituents individually, or in combination, at concen- trations reported in the literature to be toxic to fish and aquatic life. ' ' Individual or contributing toxic levels of ammonia was estimated in 12 samples, sulfide in nine samples, and phenolics in eight samples. Five samples did not contain sufficient concentra- tions of these three constituents, individually or in combination, to account for the toxic level found. It is assumed, therefore, that toxicity of these five samples was due to the presence of unknown toxic compounds, e.g., hydrocarbons. The behavior of distressed and dying fish is often an indication of the type or category of the agent(s) responsible for death. ' ' ' Continuous observations were made during these tests and the exhibited symptoms noted. Observed symptom were compared with behavioral symptoms reported in the literature for constituents suspected to be present. During this series of tests, distressed and dying fish exhibited four different symptom patterns: 14 ------- Table 3. ESTIMATED INITIAL CONCENTRATIONS OF COMMON TOXICANTS AT 24-HOUR TL PERCENTAGE VOLUME (mg/1) O \J Refinery/Sample No. 6992 it ii ii 59991 69991 693 it 69994 69995 ii ii it ii 92 6996 ii ii RW-2 RW-3 RW-10 RW-11 RW-1 RW-1 RW-1 RW-2 RW-1 RW-1 RW-3 RW-4 RW-5 RW-6 RW-2 RW-3 RW-4 RW-5 Ammonia 1.50 0.10 1.68 3.61 8.85 5.40 0.22 3.58 5.63 5.27 0.17 0.20 5.04 0.06 2.04 0.22 14.7 1.35 Sulfide 1.67 0.02 1.92 4.90 6.30 1.89 0.12 0.20 0.86 7.15 0.05 0.12 3.28 0.01 0.08 0.06 - 1.50 Phenol Cyanide 0.22 0.003 6.24 0.007 0.25 0.001 0.05 0.013 0.57 0.056 0.39 0.19 0.15 0.001 0.01 0.003 0.20 0.017 0.35 0.07 0.07 2.60 0.004 0.03 3.36 0.13 - 0.21 0.061 pH 8.6 8.6 8.7 8.5 8.7 8.2 8.4 9.1 8.9 9.0 8.6 8.6 8.9 8.4 9.0 7.8 8.3 8.6 TL50 % by Vol 0.042 0.043 0.05 3.8 7.5 13.5 0.23 1.8 4.5 6.5 13 13.5 4.2 7.2 2.4 4.2 4.2 0.05 Measured Concentrations 6992 693 it 69994 69995 92 RW-2 RW-1 RW-2 RW-1 RW-1 RW-2 2.40 3.55 5.40 5.85 3.95 1.00 0.20 2.71 15 ------- 1. Wild erratic swimming; loss of equilibrium with periodic spurts; death very shortly with gills swollen (ammonia, low pH,- some phenolic compounds). 2. Wild erratic swimming; loss of equilibrium; lethargic condi- tion; highly irritable; death after long period (petrochemicals, phenolics). 3. Some hyperactivity prior to loss of equilibrium; no irritation; state of anaesthesia for long period prior to death; gills swollen (cyanides, phenols, sulfides). 4. Slow circular swimming at surface; irritable; quick spurts from lethargic condition; state of anaesthesia on bottom for long period prior to death; gills swollen (petrochemicals, phenols, cyanides, sulfides). The two most common patterns were those in numbers 1 and 4. PHASE II During Phase II, 30 bioassays were conducted on composite samples collected from various stages of the treatment systems of five oil refineries (Figure 2). A total of 12 samples were tested. Two or three bioassays were run on each sample. Results of these toxicity tests are tabulated in Table 4. Low dissolved oxygen in four of the tests prevented establishment of reliable 24-hour TL-,, values for DU those samples; hence, 12-hour TL5Q values for each sample have also been included in Table 4. Bioassay data for each sample are presented in Appendix B. Results of chemical analyses of samples collected from the same sample points concurrent with bioassay samples are pre- sented in Appendix C. Toxicity values remained relatively constant for different composites from the same sample location. API separator effluent samples from the five refineries exhibited some toxicity. The 24-hour TL_0 of these samples from the five refineries varied from 4%-50%. Activated 16 ------- Refinery No. 9973 Process Water API _(S) Separator Equalization Aeration Basin Final (S) Clarifier Refinery No. 2115 Oily ^_ API (S)^ Primary Water Separator Clarifier Trickling (S) Aeration Filter Basin Final (S) Clarifier Refinery No. 0288 Oily Water' API Separator Air (S) Aeration Floatation Basin Final (S) Clarifier Refinery No. 6512 All Waste Water API _ Equalization ^ . Chemical (S) Final (S) Separator M Coagulation riai-i-p-ioT. „, .... Clarifier Refinery No. 6693 All Waste Water API (S) Aeration Separator Basin Final (S) Clarifier (S) Denotes sample point Figure 2. Sample points from activated sludge refinery wastewater systems chosen for toxicity tests ------- Table 4. BIOASSAY RESULTS - PHASE II Composite Samples 24-hr. TL5Q Sample Location Refinery 9973 API Separator Final Clarifier ABC (% Dilution) h h 29° 19° >56 >88 h 14° 100 12-hr. TL5Q ABC (% Dilution) 42 34 41 Estimated 24- Estimated NH hr. TL5Q Range Cone, at TL5Qa (% Dilution) (mg/1) 34-42 3.9 (5.1-3.1) 100 12.5 No mortality or stress Refinery 2115 API Separator Trickling Filter Final Clarifier Refinery 6512 Equalization Chem. Coag. Final Clarifier No Refinery 0288 Air Floatation Final Clarifier No Refinery 6693 API Separator Final Clarifier No 4.2 4.2 12.5 16 18 19.5 h h 12.5? 18? n h 12.5° 28° 100 100 mortality or h 10.5 50 >48 100 mortality or 24 18 >41 100 mortality or 3.5 11.5 20 - - - stress - - stress 22.5 - stress 4.2 13.5 34 >56 100 Stress h 34b >32 4.2 3.7 16 13 35 >28 V, 42b 60 >56 Stress _ 41 22.5 3.3-4.2 4.7 (6.4-3.8) 11-16 12.8 (20.0-10.0) 18-20 19.8 (33.0-9.5) 32-56 42-60 100 24-56 100 18-24 100 values in parenthesis are measured concentrations (at 24 hrs) in test containers bracketing the interpolated TL,.n value. i Low dissolved oxygen contributed to deaths in lower dilutions. ------- sludge systems of all refineries samples reduced toxicity. Final clarifier effluents from four refineries illicited no mortality or stress symptoms during the test period. The API separator effluent from refinery 2115 exerted a 24-hour TL of 4.2%-3.5% for three composite samples; corresponding values for the final clarifier effluent were 18%, 19.5%, and 20%. The major contributing toxic agent in this refinery is ammonia. Since the concentration of ammonia was essentially the same at both locations, the decrease in toxicity is most likely due to removal of contributing biodegradeable toxicants as well as a decrease in pH of the wastewater as it passed through the treatment system. Observations were made of the behavioral patterns of distressed fish. These observations revealed the same four sypmtom patterns as those noted in Phase I of this study. 19 ------- SECTION VI DISCUSSION PHASE I Bioassay tests discussed herein were made using untreated process waste- waters from 17 selected oil refineries. Toxic characteristics of samples tested varied greatly between the 17 refineries. There is also a wide variation in toxicity of samples from different processes within an individual refinery. Results of the tests conducted and observations made during the tests for each individual refinery are discussed below. Refinery 6992 Thirteen samples from this refinery were tested. Four of the samples exerted a 24-hour TL5Q at dilutions tested: 0.042%, 0.043%, 0.05%, and 3.8%. Tests were extended over 48 hours with no change in the first three values and a reduction to 3.2% in the other test. The DO did not drop below 4.0 mg/1 during the first 24 hours in any of the tests; therefore, toxicity was due to wastewater constituents. Most of the toxicity exerted by these samples is due to high concen- trations of constituents normally found in oil refinery wastewaters (ammonia, phenolic compounds, sulfide, cyanide) and was predictable from results of chemical analyses. Symptoms exhibited by distressed fish were either those described in numbers 1 or 3 in Section V. These symptoms include those reported for the most common toxic constituents of oil refinery wastewaters. Refinery 59991 The sample tested from this refinery exerted a 24-hour TL^n of 7.5%. *?u The DO remained greater than 5.0 mg/1 throughout the test period; 20 ------- therefore, toxicity was attributed to constituents in the wastewater. Symptoms of distressed fish were intermediate between numbers 1 and 4 in Section V, suggesting that the toxic agents were most likely ammonia and sulfide in combination. The TL<-n value was predictable from results of chemical analysis. Refinery 69991 This sample exerted a 24-hour TLt_n of 13.5%. The DO remained greater than 3.5 mg/1 throughout the test period; therefore, toxicity was attributed to wastewater constituents. Symptoms of dying fish were those listed in number 4 of Section V, suggesting that the toxic agents were most likely ammonia, sulfide, and cyanide in combination. The TL value was predictable from results of chemical analyses. Refinery 693 Each of the three samples analyzed from this refinery exerted a 24-hour TL_ of <10% although toxicity in sample RW-3 was most likely due to low DO. Sample RW-1 was much more toxic than predictable from results of chemical analyses. Dilutions were first set up from 10%-1%. Fish in all five dilutions became distressed immediately and were dead in less than two hours. Symptoms were those stated in number 2 of Section V. The sample had a definite creosote smell; therefore, the most likely toxic agents was one of the highly toxic phenolic compounds. Additional dilution series of 1%-0.1% and 0.1%-0.01% were set up to establish a 24-hour TL . In the 1%-0.1% series all test fish were in distress within four hours with the same symptoms. A 24-hour TL of 0.24% O \J was established. Live fish in the remaining dilutions were in obvious distress. Fish in 0.1% dilution were removed at the end of test period and placed in clean aerated water. Symptoms appeared to be gone after six hours. Aeration was stopped and fish observed for 24 hours. All fish were alive and appeared healthy at the end of 24 hours. 21 ------- Observation time was extended in 0.1%-0.01% dilution series to 36 hours. All fish were alive but in obvious distress at the end of 24 hours. Dissolved oxygen was greater than 4 mg/1 at 24 hours but had dropped to 2.5 mg/1 at 36 hours and tests were discontinued. The 36-hour TL of 0.05% may have been influenced by the low DO concen- tration. All live fish were in distress at the end of 36 hours, but fish at .018% and .010% did not become irritated as they did at higher dilutions. Sample RW-2 exerted a 24-hour TL5Q between 1.35%-2.40%. Insufficient sample was available to set up a 1.8% dilution. Symptoms were those listed in number 1 of Section V. The TL5 was predictable from the high ammonia values in the raw sample. Refinery 69995 A 24-hour TL for these two sample could not be established from the dilutions tested and was reported as >10%. Some mortality and distress we-re noted at 10% dilution for sample RW-1; however, low DO definitely contributed to the mortality. Distress was observed prior to the DO drop. All fish in RW-2 dilutions were alive and healthy at end of test period. Refinery 22 A 24-hour TL for these two samples could not be established from the dilutions tested and was reported as >18%. Fish at 18% for sample RW-2 were in distress at 12 hours, suggesting a 24-hour TL of 18%- O \J 32%. Dissolved oxygen was >4 mg/1 at end of test. Refinery 69994 Toxicity in sample RW-1 was predictable from results of chemical analyses. All fish in 10% dilution were dead within 30 minutes with symptoms described in number 1 of Section V, suggesting ammonia as 22 ------- the major toxic agent. Eight fish were dead in 5.6% dilution within six hours; the two live fish remained in distress throughout the test. Dissolved oxygen was >4.0 mg/1 at end of test. Toxicity for sample RW-2 was mostly due to low pH (raw sample pH 0.9). All fish in 10% dilution were dead in five minutes with visable bleeding from gills (pH 2.1). All fish in 5.6% dilution were dead in 25 minutes and seven fish in 3.2% dilution were dead in 30 minutes (pH 2.6 and 3.25) One fish was alive in 3.2% dilution at end of test (pH 6.2 at 24 hours). All fish were alive and in no distress in 1% dilution (pH 7.1). Since lack of sufficient sample prohibited a set-up at 1.8% dilution, the 24-hour TL_n was reported to be between 1.35%- 2. 50%. ^u Refinery 6993 No toxicity was exerted or distress observed at dilutions tested (TL5 Refinery 595 No toxicity exerted or distress symptoms observed during test period at dilutions tested for sample RW-1 (TL5Q >10%) . Sample RW-2 exerted no toxicity during 24 hours (TL >18%) . Fish in 18% dilution were in obvious distress at surface throughout the test and in 10% dilution after 12 hours, suggesting a 24-hour TL for 18%-32%. Dissolved oxygen was low at 24 hours . Refinery 6995 A 24-hour TL was established for five of 10 samples tested. Three of the toxic samples, RW-3, RW-4, and RW-6, exerted toxicity greater than that predictable from results of chemical analyses. All live fish for these three samples were in distress at the end of 24 hours. Symptoms for all three samples were those listed in number 4 of Section V. Toxicity of these samples was most likely due to toxic hydrocarbon compounds. A plasticizer smell was noted. 23 ------- Mortality for samples RW-1 and RW-5 was predictable from results of chemical analyses. Symptoms exhibited were those listed in numbers 1 and 4 of Section V. The most likely toxic agents were combinations of ammonia and sulfide in RW-1 and ammonia, sulfide, and phenolics in RW-5. Low DO may have contributed to mortality in 5.6% dilution for sample RW-1. Fish in samples RW-2, RW-7, RW-9, and RW-10 exhibited no stress during the test period at the dilutions tested. Two fish died in samples RW-8 in 18% dilution and remaining fish were in obvious distress after 12 hours. Since DO remained >4.5 mg/1 throughout the test, a TL n of J\J between 18%-32% could be predicted for this sample. Refinery 92 Samples RW-1 exerted no toxicity during the test period at the dilutions tested. Fish in 10% and 5.6% dilutions were in distress at surface at end of test due primarily to low DO. Sample RW-2 exerted a TL5Q of 2.4% for the test period with the DO remaining greater than 5.0 mg/1. Distress was noted in 1.8% dilution after 12 hours. Symptoms of distressed fish were those listed in number 1 in Section V. Toxicity in this sample was predictable from results of chemical analyses (phenol 144 mg/1; NH_ 85 mg/1). J Refinery 59 Although a TL™ could not be established at the dilutions tested (TL^f, >10%), all fish exhibited an abnormal behavior after 12 hours. Dissolved oxygen remained adequate throughout the test. Refinery 59995 No toxicity was exerted or distress symptoms exhibited at dilutions tested during the test period for these two samples (TL >10%). 24 ------- Refinery 106 No toxicity was exerted during test period at dilutions tested. Fish in 18% and 10% dilutions exhibited distress symptoms at end of test period primarily due to low DO (TL,.,, >18%) . Refinery 56 No toxicity was exerted during the test period at the dilutions tested; however, fish at 10% and 5.6% dilutions exhibited distress symptoms after six hours (TL5Q >10%). Refinery 67 No toxicity was exerted or distress symptoms exhibited during test period at dilutions tested (TL5Q >10%). Refinery 6996 Three of the five samples tested exerted a 24-hour TL,.,. at dilutions tested. Sample RW-3 exerted toxicity much greater than indicated by results of chemical analyses. No toxicity was exerted or distress symptoms exhibited in sample RW-1 at dilutions tested (TL5_ >10%). Sample RW-2 exerted no toxicity at dilutions tested (TV >10%); however, fish in the 10% dilution were in distress at surface at 12 hours with DO >5.0 mg/1 throughout the test period. A 24-hour TL5Q of 10%-18% is indicated. Sample RW-3 exhibited symptoms described in number 4 of Section V. Fish in 3.2% and 1.8% dilutions were in obvious distress at end of 24-hour period; no distress was observed in 1.0% dilution. Test was extended through 48 hours with some mortality in 3.2% and 1.8%; fish in 1.0% were not in distress at 48 hours. Dissolved oxygen remained sufficiently high throughout the 48-hour period to have no effect on results. The toxic agent is unknown but a hydrocarbon compound is suspected. 25 ------- Fish in sample RW-4 exhibited symptoms described in number 3 of Section V, while those in sample RW-5 exhibited symptoms described in number 1. Dissolved oxygen remained >4.0 mg/1 in both tests throughout the test period. Toxicity in both samples was predictable from results of chemical analyses (RW-4: ammonia 355 mg/1; RW-5: ammonia 3,500 mg/1, phenol 400 mg/1, sulfide 3,000 mg/1, cyanide 128 mg/1). With the ammonia value of 355 mg/1, RW-4 predicted to be more toxic than the bioassay indicated. PHASE II Bioassay tests discussed herein were conducted using samples from various treatment stages of activated sludge treatment systems at five selected oil refineries. Location of sample points in these refineries are noted in Figure 2. Toxic characteristics of the samples tested varied considerably between different oil refineries and between different treatment stages within individual refineries. Toxicity decreased at all selected refineries as the degree of treatment increased. Results of the tests conducted and observations made during the tests for each individual refinery are discussed below. Refinery 9973 Effluent samples from the API separator and final clarifier were tested from the treatment system of this refinery. Although low DO after 12 hours prevented establishment of a reliable 24-hour TL^_ for the separator effluent, some toxicity was exerted. Twelve-hour TL 's of 42%, 34%, and 41% for the three composites tested were obtained prior to the DO drop. A rerun of a composite of all samples using five fish in ten liters gave no mortality at 40%; therefore, the 24-hour TL for this sample is estimated to be approximately the Ov/ same as the 12-hour value. Symptoms of dying fish were those listed in number 2 in Section V. Death was most likely due to the combined toxic action of constituents usually found in oil refinery wastewaters 26 ------- (ammonia, phenolics, sulfides, cyanides). No mortality was exerted or distress symptoms observed in 24 hours in a 100% composite sample of the final clarifier effluent. Refinery 2115 Samples from effluents of all three stages of treatment exerted a definite 24-hour TL™. Three composite samples for each stage of treatment were tested with toxicity decreasing as the stage of treat- ment progressed from approximately 4% for the API separator effluent to approximately 20% for the final clarifier effluent. The high ammonia content at all stages of treatment was most likely the major contributing toxic agent, although symptoms of dying fish indicate some other toxic agent or agents were present. Stress symptoms were those listed in numbers 1 or 3 of Section V. Higher concentrations of ammonia seemed to be necessary for the same level of toxicity as the level of treatment progressed. A corresponding decrease in pH as treatment progresses most likely influenced ammonia toxicity in these tests since toxicity of ammonia increases with an increase in PH.5' 10 Refinery 6512 Tests were run on two composite samples of effluents from each of three stages of treatment. Effluent samples from the equalization and chemi- cal coagulation units exhibited a demand that depleted the oxygen to a level where test results are considered unreliable. Five fish were used in ten liters of solution for the test of composite B. This did not decrease the effect of low DO enough to make the results reliable. Based on stress symptoms of test fish and early mortality during the tests, the 24-hour TL,_0 is estimated to be between 32%-56% for equal- ization unit samples and 42%-60% for chemical coagulation unit samples discounting effects of low DO. Stress symptoms were those listed in 27 ------- number 4 of Section V. No mortality occurred or stress symptoms were observed during the test period for the final clarifier effluent sample in a 100% solution. Refinery 0289 Tests were run on two composite samples for each of two stages of treat- ment. Low DO in composite A for the air flotation unit sample prevented establishment of a reliable TL . By using five fish in ten liters of solution for composite B, a TL,-n of 50% was obtained. Since the final DO was 2.5 mg/1 a range of 24%-56% is estimated for this sample. No mortality occurred or stress symptoms were observed in a 100% sample solution of the final clarifier effluent. Refinery 6695 Tests were run on three composite samples of the API separator effluent and two composites of the final clarifier effluent. The 24-hour TL^_ of approximately 21% (18%, 22.5%, and 24%) for the API separator effluent was reduced to where no mortality occurred or stress symptoms were observed in a 100% solution of the final clarifier effluent. Stress symptoms in the API tests were a combination of those listed in numbers 1 and 3 of Section V. 28 ------- SECTION VII REFERENCES 1. Katz, M. The Effects of Pollution Upon Aquatic Life. In: Water and Water Pollution Handbook, Volume I (L. L. Ciaccio, Editor). Marcel Dekker, Inc., New York, N.Y. pp. 297-328. 1971. 2. Graham, R. J. and T. C. Dorris. Long-Term Toxicity Bioassay of Oil Refinery Effluents. Water Research, Pergamon Press (London). 2_:643-663. 1968. 3. Clemens, H. P. and P. Summers. Will Refinery Wastes Kill Fish? Petroleum Refining. 32_(8): 145-147. 1953. 4. Dorris, T. C., W. Gould, and C. R. Jenkins. Toxicity Bioassay of Oil Refinery Effluents in Oklahoma. In: Biological Problems in Water Pollution, 1959 Seminar Transactions. Robert A. Taft Sanitary Engineering Center, Cincinnati, Ohio. Public Health Service Technical Report W60-3. pp. 276-285. 1960. 5. McKinney, R. E. Biological Treatment Systems for Refinery Wastes. Journal Water Pollution Control Federation. _39_(3) :346-359. 1967. 6. McKee, J. E. and H. W. Wolf (Editors). Water Quality Criteria. California State Water Quality Control Board, Sacramento. Publi- cation No. 3-A. pp. 229-232, 237-240. 1963. 7. Wilber C. G. Oil Pollution. In: The Biological Aspects of Water Pollution. Thomas Publishing Co., Springfield, Illinois, pp. 73-92. 1969. 8. Petroleum-Organic Chemicals Wastes Section. National Petroleum Refining Wastewater Characterization Studies. Robert S. Kerr Environmental Research Laboratory, U. S. Environmental Protection Agency, Ada, Oklahoma. (to be published) 29 ------- 9. American Public Health Assocation. Standard Methods for the Examination of Water and Wastewater, 13th Edition. American Public Health Assocation, New York, N.Y. pp. 562-577. 1971. 10. Brungs, W. A. Personal Communication. National Water Quality Laboratory, U. S. Environmental Protection Agency, Duluth, Minnesota. 1972. 11. Burks, S. L. Personal Communication. Zoology Department, Oklahoma State University, Stillwater. 1972. 12. Jones, J. R. E. Fish and River Pollution. Butterworths Inc., London, England, pp. 83-106, 142-152. 1964. 13. Turnbull, H., J. G. DeMann, and R. F. Weaton. Toxicity of Various Refinery Materials to Fresh Water Fish. Industrial and Engineering Chemistry. 4i6(2)-.324-333. 1954. 14. Pickering, Q. H., and C. Henderson. Acute Toxicity of Some Impor- tant Petrochemicals to Fish. Journal Water Pollution Control Federation. 38(9):1419-1429. 1966. ------- SECTION VIII APPENDICES Page A. Raw Data for Toxic Samples - Phase I 32 B. Raw Data - Phase II 40 C. Analytical Data - Phase I and Phase II 53 31 ------- APPENDIX A RAW DATA FOR TOXIC SAMPLES - PHASE I 32 ------- Sample 6992:RW-2 TL5() . 0.042% Cone . of Waste (% by vol.) 0.1 0.056 0.032 0.018 0.010 Control Sample 6992: Cone, of Waste (% by vol.) 0.1 0.056 0.032 0.018 0.010 Control Sample 6992: Cone . of Waste (% by vol.) 0.1 0.056 0.032 0.018 0.010 Control No. of Test No. Animals at 10 10 10 10 10 10 RW-3 TL5() - 0 No. of Test No. Animals at 10 10 10 10 10 10 RW-10 TL5() - No. of Test No. Animals at 10 10 10 10 10 10 Alive 24 hrs 0 2 8 9 10 10 .043 Alive 24 hrs 0 1 10 9 9 10 0.05% Alive 24 hrs 0 4 9 8 9 10 D0a (mg/1) 7.0 6.3 4.3 - - 3.5 DO (mg/1) 8.6 - 4.3 - - 3.7 DO (mg/1) 5.8 5.2 2.2 - - 3.8 Temp. (° C) 23 23 23 23 23 23 Temp. (° C) 22 - 22 - 23 22 Temp. (° C) 23 24 22 - - 23 pH Sulfide Start (mg/1) 8.6 1.2 8.6 1.1 8.3 0.9 8.4 8.6 8.6 pH Start - - 8.6 - - 8.65 PH Start 8.7 - - - - 8.6 aDO, temperature, sulfide, ammonia, and phenol measurements were taken at 24 hours or at the death of all 10 fish. 33 ------- Sample 6992:RW-11 TL - 3.8% O \J Cone, of Waste (% by vol.) 5.6 3.2 1.8 1.0 X).56 Control Sample 59991 Cone . of Waste (% by vol.) 10 5.6 3.2 1.8 1.0 Control Sample 69991 Cone, of Waste (% by vol.) 18 10 5.6 3.2 1.8 Control No. of Test Animals 10 10 10 10 10 10 : RW- 1 TL No. of Test Animals 10 10 10 10 10 10 :RW-1 TL No. of Test Animals 10 10 10 10 10 10 No. Alive at 24 hrs 3 6 10 9 9 10 - 7.5% No. Alive at 24 hrs 0 10 10 10 10 10 5Q - 13.5% No. Alive at 24 hrs 0 10 10 10 10 10 D0a (mg/1) 6.0 - - _ _ 3.7 DO (mg/1) 7.2 4.0 _ _ _ 4.6 DO (mg/1) 4.0 2.5 2.8 _ _ 4.0 Temp.a (° C) 23 _ _ _ _ 23 Temp. (° C) 23.5 24 _ _ _ 24 Temp. (° C) 24.5 24 24 _ _ 24.5 pH Start 8.5 8.5 _ _ _ 8.7 pH Start 8.7 8.6 _ _ _, 8.6 pH Start 8.2 8.4 8.4 _ _ 8.3 DO, temperature, sulfide, ammonia, and phenol measurements were taken at 24 hours or at the death of all 10 fish. 34 ------- Sample 693:RW-1 TL5() - 0.23% Cone, of Waste (% by vol.) 1.0 0.56 0.32 0.18 0.10 Control No. of Test Animals 10 10 10 10 10 10 Sample 693:RW-2 TL5Q Cone, of Waste (% by vol.) 10 5.6 3.2 1.8 1..0 Control Sample 693 Cone, of Waste (% by vol.) 10.0 5.6 3.2 1.8 1.0 Control No. of Test Animals 10 10 10 No set up 10 10 :RW-3 TL5() No. of Test Animals 10 10 10 10 10 10 No. Alive at 24 hrs 0 0 0 9 10 10 -1.8% No. Alive at 24 hrs 0 0 0 X 10 10 - 7.5% No. Alive at 24 hrs 0 10 10 10 10 10 D0a (mg/1) 8.0 8.0 7.5 4.1 - 3.7 DO (mg/1) 8.7 8.8 7.5 X 4.3 4.6 DO (mg/1) 0.5 4.6 - - - 4.3 Temp.a (° C) 21.5 22 23.5 24 24 24 Temp. (° C) 22.5 23 24 X 24 24 Temp. (° C) 25 25 - - - 25 pH Phenol Start (mg/1) 8.5 0.89 8.4 - _ 0.07 - - pH Ammonia Start (mg/1) 9.5 16.5 9.2 9.0 9.1 4.8 X X 8.6 2.3 - pH Start 8.4 - - - - - aDO, temperature, sulfide, ammonia, and phenol measurements were taken at 24 hours or at the death of all 10 fish. 35 ------- Sample 69994:RW-1 - 4.5% Cone . of Waste (% by vol.) 10 5.6 3.2 1.8 1.0 Control Sample 69994 Cone, of Waste (% by vol.) 10 5.6 3.2 1.8 1.0 Control Sample 6995 Cone, of Waste (% by vol.) 10 5.6 3.2 1.8 1.0 Control No. of Test No. Animals at 10 10 10 10 10 10 :RW-2 TL - No. of Test No. Animals at 10 10 10 No set up 10 10 :RW-1 TL5() - No. of Test No. Animals at 10 10 10 10 10 10 Alive 24 hrs 0 2 10 10 10 10 1.9% Alive 24 hrs 0 0 1 X 10 10 5.6% Alive 24 hrs 0 7 9 10 10 10 D0a (mg/1) 8.5 5.0 4.1 - - 4.2 DO (mg/1) 7.8 - 7.2 X 4.5 4.5 DO (mg/1) 7.5 1.6 4.1 - - 4.5 Temp . a (° C) 23 25 23.5 - - 24 Temp. (° C) 22 - 24.5 X 24.5 24 Temp. C C) 23.5 25 24.5 - - 25 pH Ammoniaa Start (mg/1) 9.0 12.0 6.5 4.0 - - - PH Start 2.1 2.6 3.25 X 7.2 8.55 pH Ammonia Start (mg/1) 9.0 9.0 8.7 5.4 3.3 - - - DO, temperature, sulfide, ammonia, and phenol measurements were taken at 24 hours or at the death of all 10 fish. 36 ------- Sample 6995:RW-3 - 13% Cone, of Waste (% by vol.) 18 10 5.6 3.2 No. of Test Animals 10 10 10 10 1.8 No set up Control Sample 6995 Cone . of Waste (% by vol.) 18 10 5.6 3.2 1.8 Control 10 :RW-4 TL5Q No. of Test Animals 10 10 10 10 No set up 10 Sample 6995:RW-5 TL5Q Cone, of Waste (% by vol.) 5.6 3.2 1.8 1.0 0.56 Control No. of Test Animals 10 10 10 10 10 10 No. Alive at 24 hrs 0 9 10 10 X 10 - 13.5% No. Alive at 24 hrs 2 8 10 10 X 10 - 4.2% No. Alive at 24 hrs 0 10 10 10 10 10 D0a (mg/1) 7.3 4.5 - - X 4.7 DO (mg/1) 4.5 4.8 - - X 4.0 DO (mg/1) 4.7 3.4 - - - 2.6 a Temp. (° C) 24 24 - - X 24 Temp. (° C) 24 24 - - X 24 Temp. (° C) 24 24 - - - 23.5 pH Start 8.6 - - - X ~ pH Start 8.55 - - - X — PH Start 8.85 - - - - - aDO, temperature, sulfide, ammonia, and phenol measurements were taken at 24 hours or at the death of all 10 fish. 37 ------- Sample 6995:RW-6 TL - 7.2% Cone, of Waste (% by vol 18 10 5.6 3.2 1.8 Control Sample 92 Cone, of Waste (% by vol 10 5.6 3.2 1.8 1.0 Control No. of Test . ) Animals 10 10 10 10 No set up 10 :RW-2 TL5Q No. of Test .) Animals 10 10 10 10 10 10 Sample 6996 :RW- 3 TL Cone, of Waste (% by vol 10 5.6 3.2 1.8 1.0 Control No. of Test . ) Animals 10 10 10 10 10 10 No. Alive at 24 hrs 0 0 9 10 X 10 - 2.4% No. Alive at 24 hrs ( 0 0 0 10 10 10 50 - 4-2% No. Alive at 24 hrs 0 0 10 10 10 10 D0a Temp.a (mg/1) (° C) 7.0 22 7.0 23.5 5.6 23 - X X 24 DO Temp . pH mg/1) (° C) Start 6.5 22.0 9.2 5.7 23.0 9.1 5.3 23.5 9.0 2.5 23.5 _ 4.6 23.5 DO Temp . (mg/1) (° C) 6.9 22.5 7.0 22 4.8 21.5 5.6 21.5 - 6.0 21.5 pH Start _ 8.4 8.4 - X - Ammonia (mg/1) 14.2 '8.1 4.6 3.3 2.0 0.7 PH Start 7.7 - - - - 8.4 Phenol' (mg/1) 11.7 6.9 4.0 - - - a. DO, temperature, sulfide, ammonia, and phenol measurements were taken at 24 hours or at the death of all 10 fish. 38 ------- Sample 6996:RW-4 TL - 4.2% Cone, of Waste (% by vol.) 5.6 3.2 1.8 1.0 0.56 Control No. of Test Animals 10 10 10 10 10 10 No. Alive at 24 hrs 0 10 9 10 10 10 D0a (mg/1) 6.5 3.8 - - - 3.7 Temp . a (° C) 23 22 - - - 22 pH Start 8.25 8.50 8.50 - - 8.40 Sample 6996:RW-5 TL - 0.05% oU Cone, of Waste (% by vol.) 0.1 0.056 0.032 0.018 0.010 Control No. of Test Animals 10 10 10 10 10 10 No. Alive at 24 hrs 0 4 10 9 10 10 DO (mg/1) 7.3 5.6 0 - - 4.4 Temp. (° C) 22.5 22 23 - - 22.5 PH Start 8.65 8.45 - - - 8.45 DO, temperature, sulfide, ammonia, and phenol measurements were taken at 24 hours or at the death of all 10 fish. 39 ------- APPENDIX B RAW DATA - PHASE II 40 ------- SAMPLE REFINERY 9973: API SEPARATOR EFFLUENT Composite A 12-hour TL5 - 42% Cone . of Waste (% by vol) 56 32 18 10 5.6 Control Composite B Cone . of Waste (% by vol) 48 24 12 6 3 Control Composite C Cone . of Waste (% by vol) 48 36 25 16.8 12.6 Control No. of Test No. Animals at 10 10 10 10 10 10 12-hour TL No. of Test No. Animals at 10 10 10 10 10 10 12-hour TL No. of Test No. Animals at 10 10 10 10 10 10 alive No. 12 hrs at 3 10 10 10 10 10 50 - 34% alive No. 12 hrs at 0 10 10 10 10 10 50 - 41% alive No. 12 hrs at 0 10 10 10 10 10 alive 24 hrs 0 4 10 10 10 10 alive 24 hrs 0 2 10 10 10 10 alive 24 hrs 0 0 2 3 10 10 D0a (mg/1) 12 hr 24 hr 3.9 3.8 4.0 0.5 4.0 1.2 1.3 - 5.2 3.1 DO (mg/1) 12 hr 24 hr 3.8 2.0 1.0 1.5 1.5 - 4.8 2.5 DO (mg/1) 12 hr 24 hr 3.6 0.5 3.2 0.6 3.5 1.0 1.2 1.3 4.9 3.1 Temp. (° C) 22 21.5 21.5 _ _ 21.5 Temp. (° C) 22 22 22 22 _ 22 Temp. C C) 22 22 22 22 22 22 pH Start 8.9 8.8 - _ _ 8.7 pH Start 8.6 8.5 - _ _ 8.4 PH Start 8.9 8.8 8.7 8.7 8.6 8.6 Ammonia (mg/D Start 5.1 3.1 2.0 _ _ - DO readings at 12 hours, 24 hours, or at death of all 10 fish. Temperature readings at 24 hours or at death of all 10 fish. 41 ------- SAMPLE REFINERY 9973: FINAL CLARIFIER EFFLUENT Composite A 24-hour TL5Q - >56% Cone, of Waste (% by vol) 56 32 18 10 5.6 Control Composite B Cone . of Waste (% by vol) 88 44 22 11 Control Composite C Cone, of Waste (% by vol) 100 50 25 12.5 6.25 Control No. of Test No. Animals at 10 10 10 10 10 10 24-hour TL No. of Test No. Animals at 10 10 10 10 10 alive No 12 hrs at 10 10 10 10 10 10 50 - >88% alive No 12 hrs at 10 10 10 10 10 24-hour TL5Q - 100% No. of Test No. Animals at 10 10 10 10 10 10 alive No 12 hrs at 10 10 10 10 10 10 . alive 24 hrs 10 10 10 10 10 10 . alive 24 hrs 10 10 10 10 10 D0a Og/D 12 hr 24 hr 6.0 3.7 - _ - - 5.1 3.0 DO Og/D 12 hr 24 hr 6.1 3.1 5.5 2.7 - - 3.8 1.5 Temp. (° C) 22 - - - - 22 Temp. (° C) 22.5 22 - - 22.5 pH Start 8.3 - - - - 8.2 pH Start 8.0 8.3 - - 8.4 Ammonia (mg/1) Start 7.0 4.0 _ - - - (no mortality or observed stress) . alive 24 hrs 10 10 10 10 10 10 DO Og/D 12 hr 24 hr 7.2 4.0 6.2 3.5 - _ - 4.0 2.0 Temp. (° C) 22 22 - - - 22 pH Start 7.6 8.3 - - - 8.6 DO readings at 12 hours, 24 hours, or at death of all 10 fish. 'Temperature readings at 24 hours or at death of all 10 fish. ------- SAMPLE REFINERY 2115: API SEPARATOR EFFLUENT Composite A 24-hour TL - 4.2% Cone . of Waste (% by vol) 10 5.6 3.2 1.8 1.0 Control Composite B Cone, of Waste (% by vol) 10 5.6 3.2 1.8 1.0 Control Composite C Cone . of Waste (% by vol) 7.5 5.6 4.2 3.2 2.4 Control No. of Test No. Animals at 10 10 10 10 10 10 alive 12 hrs 0 0 10 10 10 10 24-hour TL - 4 50 No. of Test No. Animals at, 10 10 10 10 10 10 24-hour TL No. of Test No. Animals at 10 10 10 10 10 10 alive 12 hrs 0 0 10 10 10 10 50 " 3 alive 12 hrs 0 0 0 10 10 10 No." alive at 24 hrs 0 0 10 10 10 10 .2% No. alive at 24 hrs 0 0 10 10 10 10 .5% No. alive at 24 hrs 0 0 0 8 10 10 D0a (mg/1) 12 hr 24 hr 7.6 7.7 5.7 2.7 - _ 5.2 3.0 DO (mg/1) 12 hr 24 hr 7.8 7.6 4.8 2.5 - _ 4.5 2.5 DO (mg/1) 12 hr 24 hr 8.1 8.1 7.5 5.1 2.2 5.5 1.7 4.5 1.7 Temp. (° C) 21 21.5 22 - - 22 Temp. (° C) 21 21.5 22 - - 22 Temp. (° Q 20 20 22 22 22 22 PH Start 9.4 9.0 8.9 8.6 - 8.4 PH Start 9.3 9.1 8.8 - _ 8.6 PH Start 9.3 9.1 9.0 8.8 8.7 8.6 Ammonia (mg/1) Start 11.1 6.4 3.8 2.25 1.1 0.2 DO readings at 12 hours, 24 hours, or at death of all 10 fish. Temperature readings at 24 hours or at death of all 10 fish. 43 ------- SAMPLE REFINERY 2115: TRICKLING FILTER EFFLUENT Composite A 24-hour TL - 12.5% Cone . of Waste (% by vol) 32 18 10 5.6 3.2 Control Composite B Cone, of Waste (% by vol) 24 12 6 3 1.5 Control Composite C Cone, of Waste (% by vol) 24 18 13.5 10.0 7.5 Control No. of Test No. Animals at 10 10 10 10 10 10 2 4 -hour TL No. of Test No. Animals at 10 10 10 10 10 10 alive 12 hrs 0 0 10 10 10 10 50 ' 16! alive 12 hrs 0 10 10 10 10 10 24-hour TL - 11 oU No. of Test No. Animals at 10 10 10 10 10 10 alive 12 hrs 0 0 4 10 10 10 No. alive at 24 hrs 0 0 8 10 10 10 \ No. alive at 24 hrs 0 10 10 10 10 10 .5% No. alive at 24 hrs 0 0 0 10 10 10 D0a (mg/1) 12 hr 24 hr 7.6 - 5.3 2.0 5.3 2.5 - 5.8 2.6 DO (mg/1) . 12 hr 24 hr 7.0 4.7 1.5 5.3 3.0 - - 4.7 2.5 DO (mg/1) 12 hr 24 hr 8.0 6.9 5.9 5.5 5.8 3.3 3.6 5.7 4.0 Temp. (° C) 18 - 22 22 - 22 Temp. 20 22 22 - - 22 Temp. 18 21 22 22 22 22 pH Start 8.8 - 8.6 8.6 - 8.7 PH Start 8.4 8.3 8.3 - - 8.5 pH Start 8.6 8.6 - - - 8.6 Ammonia (mg/1) Start 33.0 20.0 10.0 6.2 - - *DO readings at 12 hours, 24 hours, or at death of all 10 fish. 'Temperature readings at 24 hours or at death of all 10 fish. 44 ------- SAMPLE REFINERY 2115: FINAL CLARIFIER EFFLUENT Composite A 24-hour TL - 18% Cone. of Waste ( % by 32 18 10 5. 3. vol) 6 2 Control No. of Test Animals 10 10 10 10 10 10 D0a No. alive at 12 hrs 0 10 10 10 10 10 No. alive at 24 hrs 0 5 10 10 10 10 (mg/1) 12 6. 5. 5. - - 6. hr 0 6 8 3 24 2 2 3 hr - .1 .2 - - .1 Temp. (° C) 22 22 22 - - - 22 Ammonia pH (mg/1) Start Start 8.5 33.0 19.8 9.5 5.8 - 8.7 Composite B 24-hour TL 50 Cone, of Waste (% by vol) 48 24 12 6 3 Control No. of Test Animals 10 10 , 10 10 10 10 DO No. alive at 12 hrs 1 10 10 10 10 10 No. alive at 24 hrs 0 3 10 10 10 10 (mi 12 hr 5.3 5.1 - - - 5.0 J/D 24 hr 5.2 2.5 3.2 - - 3.1 Temp. (° C) 22 22 22 - - 22 pH Start 8.2 8.4 - - - 8.6 Composite C 24-hour TL5Q - 20% Cone. ( of Waste % by vol) 28 21 15 11 8 .5 .5 .7 Control No. of Test Animals 10 10 10 10 10 10 DO No. alive at 12 hrs 10 10 10 10 10 10 No. alive at 24 hrs 1 4 10 10 10 10 (mg/1) 12 hr 5.2 5.3 5.5 - - 5.5 24 4 3 2 4 hr .0 .0 .8 - - .0 Te C° 22 22 22 - 22 mp. .5 .5 .5 - .5 pH Start 8.2 8.4 - - - 8.5 DO readings at 12 hours, 24 hours, or at death of all 10 fish. Temperature readings at 24 hours or at death of all 10 fish. 45 ------- SAMPLE REFINERY 6512: EQUALIZATION UNIT EFFLUENT Composite A Estimated 24-hoUr TL - 32%-56% Cone, of c Waste % by vol) 56 32 18 10 5.6 Control No. of Test Animals 10 10 10 10 10 10 D0a No. alive at 12 hrs 10 10 10 10 10 10 No. alive at 24 hrs 3 2 1 8 10 10 Ong/l) 12 hr 2.6 2.7 1.6 - - 2.5 24 0 0 0 0 1 1 hr .4 .5 .5 .8 .0 .1 Temp. pH (° 22. 22. 22. 23 23 23 C) 5 5 5 Start 8. 8. 8. - - 8. 1 3 5 6 Composite B Estimated 24-hour TL_Q - 32%-56% Cone, of Waste (% by vol) 56 32 18 10 5.6 Control No. of Test No. Animals at 5 5 No set up 5 5 5 DO , alive 12 hrs 0 5 X 5 5 5 No. alive at 24 hrs 0 0 X 5 5 5 (mg/D 12 hr 1.0 2.2 X 4.2 5.5 5.5 24 hr - 1.0 X 2.1 3.0 3.3 Temp. C° C) 22.5 23.5 X 23.5 23.5 23.5 pH Start 8.0 8.3 X 8.6 - 8.7 DO readings at 12 hours, 24 hours, or at death of all 10 (5) fish. Temperature readings at 24 hours or at death of all 10 (5) fish. 46 ------- SAMPLE REFINERY 6512: CHEMICAL COAGULATION UNIT EFFLUENT Composite A Estimated 24-hour TL5Q - 42%-60% Cone, of Waste (% by vol) 100 56 32 18 10 Control No. of Test Animals 10 10 10 10 10 10 D0a No. alive at 12 hrs 10 10 10 10 10 10 No. alive at 24 hrs 1 0 2 2 7 10 (mg/D 12 hr 2.2 2.0 2.0 - - 2.7 24 hr 0.3 0.3 0.3 0.5 0.8 1.1 V, Temp . (° C) 22.5 22.5 22.5 22.5 22.5 23 PH Start 7.6 8.0 8.4 - - 8.7 Composite B Estimated 24-hour TL5Q - 42%-60% Cone, of Waste (% by vol) 67 40 20 10 5 Control No. of Test Animals 5 5 5 5 5 5 DO No. alive at 12 hrs 2 4 5 5 5 5 No. alive at 24 hrs 0 0 5 5 5 5 Og/1) 12 hr 2.8 3.2 4.4 - - 6.4 24 hr 0.5 0.6 1.5 - - 5.5 Temp. 22.5 22.5 23 _ - 23 pH Start 7.7 8.0 - _ - - aDO readings at 12 hours, 24 hours, or at death of all 10 (5) fish. Temperature readings at 24 hours or at death of all 10 (5) fish. 47 ------- SAMPLE REFINERY 6512: FINAL CLARIFIER EFFLUENT Composite A 24-hour TL,_n - 100% (no mortality or observed stress) Cone, of Waste (% by vo-1) 100 56 32 18 10 Control No. of Test Animals 10 10 10 10 10? 10 No. alive at 12 hrs 10 10 10 10 10 10 No. alive at 24 hrs 10 10 10 10 10 10 D0a (mg/D 12 hr 6.2 5.5 - _ - 2.2 24 hr 2.4 2.5 2.7 _ _ 1.2 Temp.b C° C) 22 22 22 _ - 22.5 pH Start 7.4 8.3 - _ - 8.7 Composite B 24-hour TL - 100% (observed stree and 40% mortality) Cone . of Waste (% by vol) 100 50 25 12.5 Control No. of Test Animals 5 5 5 5 5 DO No. alive at 12 hrs 5 5 5 5 5 No. alive at 24 hrs 5 5 5 5 5 (mg/1) 12 hr 6.5 6.2 6.0 _ 6.3 24 hr 3.7 3.5 4.0 _ 4.5 Temp. 23 23 23 _ 23 pH Start 7.4 7.9 - _ - DO readings at 12 hours, 24 hours, or at death of all 10 (5) fish. Temperature readings at 24 hours or at death of all 10 (5) fish. ------- SAMPLE REFINERY 0289: AIR FLOATATION Composite A Estimated 24-hour TL5Q - 32%-48% Cone, of Waste (% by vol) 48 24 12 6 3 Control No. of Test Animals 10 10 10 10 10 10 No. alive at 12 hrs 0 10 10 10 10 10 No. alive at 24 hrs 0 2 4 10 10 9 D0a (mg/lD 12 hr 1.2 2.2 1.5 - - 1.2 24 hr _ 0.9 0.8 1.1 1.1 1.1 Temp . pH (° C) Start 23 23 23 23 23 23 8.4 8.6 8.7 - - 8.7 Composite B Estimated 24-hour TL - 50% Cone, of Waste (% by vol) 56 32 18 10 5.6 Control No. of Test Animals 5 5 5 5 5 5 DO No. alive at 12 hrs 5 5 5 5 5 5 No. alive at 24 hrs 2 5 5 5 5 5 (mg/1) 12 hr 4.2 4.7 5.0 - - 5.3 24 hr 2.0 2.2 2.9 - _ 3.8 Temp. 23 23 23 - _ 23 PH Start 8.0 8.4 8.5 - _ 8.7 aDO readings at 12 hours, 24 hours, or at death of all 10 (5) fish. Temperature readings at 24 hours or at death of all 10 (5) fish. 49 ------- SAMPLE REFINERY 0289: FINAL CLARIFIER EFFLUENT Composite A 24-hour TL5Q - >48% Cone, of Waste (% by vol) 48 24 12 6 Control No. of Test Animals 10 10 10 10 10 D0a No. alive at 12 hrs 10 10 10 10 10 No. alive at 24 hrs 9 9 10 10 10 (mg/1) 12 hr 3.4 2.5 2.5 - 2.7 24 hr 1.2 1.2 1.4 - 1.9 h Temp. (° C) 23 23 23 - 23.5 pH Start 8.6 8.7 - - 8.7 Composite B 24-hour TL_n - 100% (no mortality or observed stress) Cone, of Waste (% by vol) 100 50 25 12.5 Control No. of Test Animals 5 5 5 5 5 DO No. alive at 12 hrs 5 5 5 5 5 No. alive at 24 hrs 5 5 5 5 5 (mg 12 hr 7.0 6.6 6.3 - 6.3 A) 24 hf 5.1 5.5 4.5 - 4.6 Temp. (° C) 23 23 23 - 23 pH Start 7.7 8.1 - - - DO readings at 12 hours, 24 hours, or at death of all 10 (5) fish. temperature readings at 24 hours or at death of all 10 (5) fish. 50 ------- SAMPLE REFINERY 6693: API SEPARATOR EFFLUENT Composite A 24-hour TL5Q - 24% Cone . of Waste (% by vol) 32 18 10 5.6 3.2 Control Composite B Cone . of Waste (% by vol) 56 32 18 10 5.6 Control Composite C Cone, of Waste (% by vol) 56 32 18 10 5.6 Control No. of Test No. Animals at 10 10 10 10 10 10 24-hour TL No. of Test No. Animals at 10 10 10 10 10 10 24-hour TL No. of Test No. Animals at 10 10 10 10 10 10 alive 12 hrs 9 10 10 10 10 10 50 ' 18 alive 12 hrs 0 1 10 10 10 10 50 " 22 alive 12 hrs 0 0 8 10 10 10 No. alive at 24 hrs 0 10 10 10 10 10 % No. alive at 24 hrs 0 0 5 9 10 10 .s, No. alive at 24 hrs 0 0 8 10 10 10 D0a (mg/1) 12 hr 24 hr 5.9 4.2 6.4 4.8 - - _ 6.4 4.1 DO (mg/1) 12 hr 24 hr 8.5 6.6 6.6 2.0 - - 5.8 3.7 DO (mg/1) 12 hr 24 hr 7.7 7.0 6.0 2.8 - _ 6.4 4.8 h Temp . (° C) 24 24 - - - 24 Temp. (° C) 22.5 24 - - - 24.5 Temp. C° C) 20 23 23 - _ 23 pH Start 8.8 8.5 - - - 8.4 pH Start 8.9 8.8 - - - 8.8 PH Start 9.0 8.9 8.8 _ _ - DO readings at 12 hours, 24 hours, or at death of all 10 fish. ^Temperature readings at 24 hours or at death of all 10 fish. 51 ------- SAMPLE REFINERY 6693: FINAL CLARIFIER EFFLUENT Composite A 24-hour TL™ - >41% Cone. of Waste No. of Test No. alive No. alive (mg/l) Temp. pH by vol) Animals at 12 hrs at 24 hrs 12 hr 24 hr (° C) Start 41 32 18 10 Control 10 10 10 10 10 10 10 10 10 10 10 10 10 10 10 7.6 - - - 7.5 5.9 - - - 6.6 24 - - - 24 8.1 - - - 8.7 Composite B 24-hour TL™ - 100% (no mortality or distress observed) Cone, of No. of DO Waste Test No. alive No. alive (mg/l) Temp. pH (% by vol) Animals at 12 hrs at 24 hrs 12 hr 24 hr (° C) Start 100 .50 25 12.5 Control 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 7.0 5.7 4.4 3.8 24.5 3.0 25 7.7 8.4 8.7 DO readings at 12 hours, 24 hours, or at death of all 10 (5) fish. Temperature readings at 24 hours or at death of all 10 (5) fish. 52 ------- APPENDIX C ANALYTICAL DATA - PHASE I AND PHASE II 53 ------- RESULTS OF CHEMICAL ANALYSES OF PHASE I SAMPLES COLLECTED CONCURRENT WITH BIOASSAY SAMPLES Chemical Parameter 0 Sample No. 6992 RW-2 6992 RW-3 6992 RW-10 6992 RW-11 5991 RW-1 69991 RW-1 693 RW-1 693 RW-2 69994 RW-1 6995 RW-1 6995 RW-3 6995 RW-4 6995 RW-5 6995 RW-6 92 RW-2 6996 RW-3 6996 RW-4 6996 RW-5 Ammonia (mg/1) 3575 225 3350 95 118 40 95 199 125 81 1.3 1.5 120 0.85 85 5.2 350 2700 Phenolics (mg/1) 525 14,500 500 1.4 7.6 2.9 64 0.52 4.5 5.4 0.52 0.52 62 0.38 140 3.1 <0.01 410 Sulfide (mg/1) 3975 37 3846 129 84 14 51 11 19 110 0.4 0.9 78 0.2 3.40 1.4 1.0 3000 Cyanide (mg/1) 0.6 17.0 1.4 0.34 0.74 1.4 0.22 0.14 0.38 0.03 <0.03 <0.03 0.1 <0.03 0.08 <0.03 <0.03 122 Samples for which definite TL,_n was established. 54 ------- RESULTS OF CHEMICAL ANALYSES OF PHASE II SAMPLES COLLECTED CONCURRENT WITH BIOASSAY SAMPLES Chemical Parameter Sample Location Refinery 9973 API Separator Final Clarifier Refinery 2115 API Separator Trickling Filter Final Clarifier Refinery 6512 Equal . Chamber Chem. Coag. Unit Final Clarifier Refinery 0288 Air Float. Unit Final Clarifier Refinery 6693 API Separator Final Clarifier Ammonia (mg/1) 9.25 6.4 127 108.5 106 6.9 6.75 0.375 12.0 11.25 28.4 26.25 Phenolics (mg/D 11.0 0.01 10.45 0.99 0.035 0.61 0.405 0.01 0.02 0.01 3.4 0.01 Sulfide (mg/1) 2.88 0.15 24.3 0.75 0.275 1.0 1.5 0.425 0.7 0.5 32.5 0.2 Cyanide (mg/1) 0.135 0.10 1.50 0.245 0.07 0.22 0.20 0.14 0.06 0.02 0.29 0.185 The median value from results of analyses of samples collected on 14 consecutive days at each location is presented. 55 ------- TECHNICAL REPORT DATA (Please read Instructions on the reverse before completing) . REPORT NO. EPA-600/2-76-241 3. RECIPIENT'S ACCESSION NO. 4. TITLE AND SUBTITLE ACUTE TOXIC EFFECTS OF PETROLEUM REFINERY WASTEWATERS ON REDEAR SUNFISH 5. REPORT DATE October 1976 (IssuingJJate) 6. PERFORMING ORGANIZATION CODE . AUTHOR(S) John E. Matthews and Leon H. Myers 8. PERFORMING ORGANIZATION REPORT NO. . PERFORMING ORGANIZATION NAME AND ADDRESS Robert S. Kerr Environmental Research Laboratory Office of Research and Development U.S. Environmental Protection Agency Ada, Oklahoma 74820 10. PROGRAM ELEMENT NO. 1BB036 11. CONTRACT/GRANT NO. 12. SPONSORING AGENCY NAME AND ADDRESS Same as above 13. TYPE OF REPORT AND PERIOD COVERED Interim 14. SPONSORING AGENCY CODE EPA-ORD 15. SUPPLEMENTARY NOTES 16. ABSTRACT Static bioassays of 24 hours' duration were performed on samples of wastewaters provided by 22 domestic petroleum refiners. These wastewaters represent three types of water discharges prevalent to this industry: process wastewaters prior to dilu- tion with other streams; API separator effluents which are a conjugate of various streams within a refinery; and wastewaters following treatment by activated sludge systems. Bioassays were performed using redear sunfish (Lepomis microlophus) as test organisms. Twenty-four hour 50 percent tolerance limits (TL^) of the various waste- waters are compared with results of chemical analyses performed during the same study. Toxicity varied considerably both between refineries and for waste streams from with- in a single refinery. Results of these analyses and observed behavioral symptoms of distressed fish revealed that ammonia, sulfides, and phenolics, alone or in combina- tion, were major contributors to toxicity exerted in most samples. Three refineries had samples which were more toxic than anticipated based on results of chemical analyses, indicating the presence of other toxic compounds in unknown quantities; e.g., various hydrocarbons. Results of bioassays of samples from various wastewater treatment stages of five refineries with activated sludge treatment systems revealed a decrease in. toxicity as the degree of treatment increased indicating a reduction in concentration of -' - - ntri hiit i nor hi r. f nv-i f>gTTf- 17. KEY WORDS AND DOCUMENT ANALYSIS DESCRIPTORS b.lDENTIFIERS/OPEN ENDED TERMS COS AT I Field/Group Bioassay Freshwater fishes Toxicity Redear sunfish Acute toxicity 06A 06T 18. DISTRIBUTION STATEMENT Release Unlimited 19. SECURITY CLASS (ThisReport)' 21. NO. OF PAGES 62 20. SECURITY CLASS (Thispage) 22. PRICE EPA Form 2220-1 (9-73) 56 U S. GOVERNMENT PRINTING OFFICE-. 1976-757-056/5^2 Region No. 5-11 ------- |