Arch. Environm.Contam.Toxicol.

7, 325-337

Archives of

Environmental Contamination and Toxicology

Acute Toxicity of Hydrogen Cyanide to Freshwater Fishes 1'2
Lloyd L. Smith, Jr., Steven J. Broderius, Donavon M. Oseid, Gary L. Kimball, and Walter M. Koenst Departmentof Entomology,Fisheries,and Wildlife,Universityof Minnesota,1980FolweUAvenue, St. Paul, Minnesota55108

Abstract. Acute toxicity of hydrogen cyanide was determined at various
temperatures from 4~ to 30~ and oxygen concentrations of 3.36 to 9.26 mg/L on different life history stages of five species of fish: fathead minnow, Pimephales promelas Refinesque; bluegill, Lepomis macrochirus Rafinesque; yellow perch, Perca flavescens (MitchiU); brook trout, Salvelinus fontinalis (Mitchill); and rainbow trout, Salmo gairdneri Richardson. Median lethal threshold concentrations and 96-hr LC50's were established by flowthrough type bioassays. Acute toxicity varied from 57 /zg/L for juvenile rainbow trout to 191 /zg/L for field stocks of juvenile fathead minnows. Juvenile fish were more sensitive at lower temperatures and at oxygen levels below 5 mg/L. For most species juveniles were most sensitive and eggs more resistant. Compounds containing the cyanide group are present in many industrial and municipal effluents, including those from iron and steel mills, oil refineries, and plating plants, and constitute a significant source of toxicants introduced into aquatic ecosystems. In aqueous solution the cyanide radical from simple alkali cyanides such as NaCN hydrolyzes to form free cyanide (CN- ion and molecular HCN). The molecular (un-ionized) component predominates at pH values (6.08.0) found in most natural waters, with less than 6% free cyanide occurring in the ionic form below pH 8 at 25~ As the pH of aqueous simple cyanide solutions is increased the percentage of free cyanide present as the CN- ion is increased to satisfy the equilibrium reaction of HCN ~ H § + CN-. The world literature on the toxicity to fish of various cyanides was reviewed by Doudoroff (1976). Wuhrmann and Woker (1948), Bridges (1958), and Doudoroff et al. (1966) have concluded that HCN is the principal toxic cyanide form. However, Broderius et al. (1977) have demonstrated that even though molecular HCN is more toxic than the CN- ion, the anion does contribute to the total toxicity and to a greater degree as the test pH increases. Since HCN is highly toxic to fish and most invertebrates, and further since some relatively nontoxic

1 Paper No. 9954, ScientificJournal Series, Minnesota Agricultural Experiment Station, St. Paul, Minnesota. Research supported by the U.S. Environmental Protection Agency, EnvironmentalResearch Laboratory, Duluth,Minnesota,under Grant No. R802914. 0090-4341/78/0007-0325 $02.60 @ 1978 Springer-Verlag New York Inc.

iron-cyartide compounds photodecompose.S. A comprehensive analysis of the water was reported by Smith et al. . The experimental water had a total hardness and alkalinity of approximately 220 and 235 mg/L as CaCO3. cyanide constitutes a hazard in certain waste receiving water. During rearing and maintenance. Water flow to each test chamber was 1. Sodium cyanide stock solutions were adjusted to pH 11 with NaOH. respectively. Test chambers for juveniles and trout swim-up fry were glass aquaria 50 x 24 x 20 cm high filled to 20 L of test solution. juvenile perch were collected in the field. Bluegills were obtained from wild stock with eggs spawned and fry hatched in the laboratory. Fathead minnows were cultured in our laboratory from brood stock originally obtained from the U. Jr." Metaframe. Environmental Protection Agency's Environmental Research Laboratory in Duluth. Salmo gairdneri Richardson. fish were fed different foods including (1) mashed hard-boiled egg yolk. Yellow perch eggs were also collected in the field and hatched in the laboratory for fry tests.326 L . Yellow perch eggs were obtained from wild adult stock which spawned in the laboratory. Juveniles were held at test conditions for seven days before being placed in test chambers. et al. The present paper reports on 337 acute toxicity tests designed to determine the 96-hr median lethal toxicant concentration (LC50) of HCN and the median lethal threshold concentration (LTC) (Sprague 1969) at various temperatures and dissolved oxygen concentrations for three life history stages of four species of fish and for juveniles of one other. condition of test fish.5 hr. Perca flavescens (Mitchill). and juveniles. Pimephales prornelas Rafinesque.3 L at each 3min cycle assuring 99% displacement in 3. Inc. Smith. L . Salvelinus fontinalis (MitchiU). (2) a mixture of ground hardboiled egg and lettuce. Fish used in the tests were the fathead minnow. Temperature acclimation for eggs and fry was at the rate of llYC/hr and for juveniles 2*C/hr. Minnesota. and test conditions led to the present series of tests conducted with a single water source and using uniform experimental procedures. and (5) "Glencoe" dry pellets. The toxicant was delivered to test chambers from intermittent-flow diluters modified from those of Mount and Brungs (1967). or were reared in the laboratory. Materials and Methods Different species used for acute tests were secured as eggs. Brook and rainbow trout were obtained as newly hardened eggs or as 24-hr fry from state hatcheries. releasing HCN. Water supply for the laboratory tests was from a deep well and was transmitted to experiments through polyvinyl chloride pipe. Juvenile bluegills were collected from local waters. Juvenile wild-stock fatheads were collected from Como Lake in St. Paul. Lepornis macrochirus Rafinesque.3). Eggs and fry were randomly placed in test chambers as soon as test conditions were reached. and rainbow trout. fry. yellow perch. newly hatched brine shrimp (Artemia salina) ("San Francisco Bay Brand. A wide range of reported lethal concentrations to freshwater fishes supposedly attributable to difference in species. Juvenile fish brought from the field were given prophylactic treatment with neomycin and tetracycline at 20 mg/L for 4-hr periods on three consecutive days. bluegill. Eggs and fry were tested in screen bottomed acrylic cylinders each covered with a bakelite lid and held in a 20-L chamber so that a portion of the water from each cycle flowed upward through the screen to the outlet. (3) live. Minnesota. brook trout. (4) frozen mature brine shrimp. Test water adjusted to appropriate temperature and oxygen was conveyed by gravity from elevated head tanks to the diluter apparatus. All tests were conducted under two fluorescent lamps (Luxor 3 Use of product or trade names mentioned throughout the text does not constitute endorsement. (1976).

their 95% confidence Table 1.Toxicity of HCN to Fish 327 Inc. K.0~K_z>(1/b)(x2/K-2)l/~(N'/2) -~j~for heterogeneous data. 10 10-20 10-20 10 10 Juveniles Length (ram) 26-45 13-28 48-62 40-68 40-68 . The freshwater fish used for testing and their life history stage. Length (ram) 5-6 4 4 14-16 No. but following temperature acclimation juveniles were held in test chambers for three days prior to toxicant exposure. "Vita-Lite") providing an intensity of 55 to 78 footcandles at the water surface and a photoperiod of12 hr light. The symbol. Upper and lower 95% confidence limits for the 96-hr median lethal tolerance (LC50) and median lethal threshold concentrations (LTC) were calculated from the equations log LCS0 • 1. S. If this value is greater than the tabulated (Chi) z with K-2 dr. pH. Only identified live eggs were tested.96(1/b)(N'/2) -1~ for homogeneous data or log LC50 • t. 03. The symbol. 1971) with calculated HCN concentrations based on corresponding pH and temperature measurements and using dissociation constants of molecular HCN as defined by Izatt et al. When the data were such that probit analysis by computer was not possible. (1962). Results A c u t e t o x i c i t y o f H C N v a r i e d f o r d i f f e r e n t fish life h i s t o r y s t a g e s w i t h e g g s b e i n g most resistant and newly hatched fry and juveniles the most sensitive. eggs 25-50 25 20-70 25 . refers to the number of test organisms expected to die within the mortality interval of 16 to 84%. Observations on mortality of test organisms exposed to known cyanide concentrations were made daily. Free cyanide concentrations in each chamber were determined daily by the Epstein colorimetric method (American Public Health Association et al. The above formulas were derived from Litchfield and Wilcoxon (1949) and Firmey (1971). N'. All reported concentrations of toxicant. 25 10-50 25 10 . The reciprocal of the log-probit line's slope (l/b) is equivalent to the standard deviation of the logarithm of the population's tolerance frequency distribution (&) or the logarithm of Litchfield and Wilcoxon's (1949) slope function. size. The median lethal threshold concentrations were determined at the end of a time interval when no mortality in any test treatment had occurred for at least 48 hr. and density in the bioassay chambers are presented in Table 1. Median lethal concentrations for 96-hr and lethal threshold periods. Eggs and fry were tested immediately on introduction to the test chambers. . calculations were made graphically. refers to the number of treatment levels excluding controls. Juveniles were fasted during the last 24 hr of acclimation to the bioassay systems and during the first 96 hr of toxicant exposure. and temperature were based on analyses made on water from the test chambers. Length (mm) 5-6 4 4 9-10 . The concentration-percant mortality data were analyzed with a logarithmic-probability (log-probi0 program (Dixon 1973). Number of organisms per test chamber and range in total length at different life history stages Sac fry Species Fathead minnow Bluegill Yellow perch Brook trout Rainbow trout No. and • to the (Chi) z value calculated for goodness of fit of the regression line to the data points. the data are significantly heterogeneous. 25 10-50 25 10 . Swim-up fry No. No.

At the hatching LC50 values essentially all the newly hatched sac fry were premature and deformed.9~ and comparable DO levels the LTC values increase progressively from 61. Fathead Minnow Tests with different life history stages of the fathead minnow were conducted at about 15~ to 25~ and 4 to 7 mg/L dissolved oxygen. 20~. respectively. and 17 days. Bluegill Bluegill eggs incubated at about 25~ and DO ranging from 6. et al. No change in sensitivity related to test temperature was observed for juveniles (Table 4).2*C resulted in some increased sensitivity at hatching. There was little difference between the 96-hr and LTC values for juvenile perch.14 mg/L DO an increase in sensitivity was apparent (Table 3). However.0~ to 21.0 ~ to 15. .4~ to 24.0~ and at comparable DO levels the LTC values were determined to increase progressively from 87. At comparable Do levels it appears that a reduction in temperature from 25.56 mg/L DO and 21oc with LTC values decreasing from 107 to 75. There was no clear trend associated with temperature for fry tests.48 to 6. In juvenile tests at 8. Smith. but for tests at about 25~ and below 5.328 L. For fry and juvenile tests there was little difference in the 96-hr and median lethal threshold concentrations. Fry were considerably more sensitive to cyanide then eggs. and log-probit regression analysis of the concentration-percent mortality curves for each exposure period are detailed in Tables 2 through 13. 10.6 to 101 tzg/L HCN (Table 8). For juvenile tests at 15. Delayed embryological development at low temperature and low DO resulted in no apparent difference in resistance with decreasing DO but an increase in resistance with decreasing temperature for 96-hr egg tests (Table 2).39 mg/L showed no correlation in sensitivity at hatching to high concentrations of HCN (Table 5).7 to 120 txg/L HCN (Table 6). Median lethal HCN concentrations at hatching were markedly reduced for tests at about 25~ and when the DO was less than 5.L.90 mg/ L DO and 25~ Yellow Perch Results from yellow perch egg and fry experiments conducted at about 10~ to 18~ and 4 to 7 mg/L dissolved oxygen are so variable that specified median lethal concentrations cannot be considered definitive (Table 7).90 to 3. only a slight reduction in sensitivity was observed for juveniles tested at 3. showing at 25~ a substantial decrease in 7-day LTC values with decrease in DO.10 to 3.52 mg/L. limits. and 15~ was 6. An increase in sensitivity was observed for juveniles tested at 7. Jr. The time required to hatching for eggs at about 25~.0 /zg/L HCN.

5.77 5.g/L 95% Confidence limits p.488 -13.743 -13.88 11 7.840 6.288 .5.14 3.6 24.1 -81.34 7.96 8.973 4.989 .621 7.6.99 1 8.89 6 7.9 8 5 40 36 5 8 6 6 9 I0 12 10 -- .0--154 -122-266 -135-193 -- a A probit o f 4.6 90. a n d 6.g/L 95% Confidence limits 15.507 9.0 24.2-93.2.0 c o r r e s p o n d s to 16.p r o b i t regression analysis ~ 6.02 3 2 9 7.25 7.9-111 -71.0 20.291 .314 122 -99.T a b l e 2.95 2 7.9 24.612 .0 24.159 3. 50.13 3.2-93.419 .475 8.508 2.3-142 -86.563 4.0 25.72 2 7.376 .52 6.411 .494 6.2 ~r 20.90 3 7.-14.8.0 ~D 24.8. of tests b p.6-I lO 71.940 9.8--113 -83.84 7.36 6.38 6.0.46 5. r e s p e c t i v e l y .8.6.721 5.5. w h e n ~ l is the m a x i m u m likelihood probit v a l u e o f p e r c e n t mortality and Xt is log H C N c o n c e n t r a t i o n in p.7.3-130 96.6-157 -83.00 5 7.063 .6 108 113 -92.g/L No.491 6.3-190 -115-293 -140-274 --- -126 -118 -116 -113 -180 -162 187 -90.51 4.194 8.145 .471 6. A c u t e t o x i c i t y o f H C N to fathead minnow swim-up fry expressed as 96-hr LC50 and median lethal threshold concentrations 96-hr L C 5 0 LTC M e a n test conditions L o g . 5.971 8.86 6 DO mg/L pH Treatments a No.9.765 1.8 25.909 4.1 81.6 90.5-133 -102 -96.14 6.033 -15.86 2 DO mg/L pH Treatments b a b p.411 4. and 84% mortality.5-133 .9-174 -97.6 108 I13 104-143 -88.170 .9 13 15 21 13 lO 8 28 -12.7 24.3-130 96.262 .g/L for the r e g r e s s i o n equation r = a + b(Iog Xl) b N u m b e r o f H C N c o n c e n t r a t i o n s in the r e g r e s s i o n analysis T a b l e 3.992 -- 352 -273 -202 -121 -184 -196 -202 274-453 -162-463 -130-314 -77.6.143 5.17 7.0.863 -- 3.358 .4.175 .g/L p. A c u t e t o x i c i t y o f H C N o L o g probit r e g r e s s i o n analysis a 96-hr L C 5 0 Hatch LC50 to f a t h e a d m i n n o w eggs expressed as 96-hr LC50 and median lethal concentrations at hatching M e a n test conditions ~ 6. of tests 95% Confidence limits 95% Confidence limits 15.

8.01 6.86 4 15 15 II II -27.99 6.92 7.20 7.146 121 -128 -116-125 -109-149 --119 -123 -115-123 -105-143 DO mg/L pH Treatments a b IcJL pg/L NO.58 7.75 7.8 20.978 131 124-138 131 106 119 129 120 124-138 87.415 82. Acute toxicity of HCN to fathead minnow juveniles expressed as 96-hr LC50 and median lethal threshold concentrations.883 5.512 .78 4 8 9 13 -21.580 -18.g20 3.2.0 25.2 3.256 3.9-129 I I I .924 15. Mean test conditions Log-probit regression analysis 96-hr L C 5 0 LTC *C 15.455 16.538 15.4.9 24~8 tI II 4 10 7 22 28 9 .902 -12.58 5.417 7.80 3 4 7.90 2 2 2 2 6 6 8 6 .0 6.446 I 1.565 . o f tests ~. .9-129 I 11-129 124-133 113-128 7.185 --------690 .9 -116-132 122-153 20.80 2 4 -24.611 9.81 7.9-120 -117-189 -193-247 -i 10-340 c~ .2.630 -13.292 -20.355 365 --232 -276 -271 -- 188-709 --147-366 -241-316 -200-368 -- ~ 205 109 -149 ~ 218 -194 -156-270 99.973 5.756 -18.70 4 6.791 22.9 24.4 76.83 7.0 20.485 12.08 6.8.117 .415 106 119 129 120 87.13 7.032 -27.176 6.8.1.18.9.68 5.872 7.09 6.2 25.8 25.0 3.90 4 9 -28.07 7.99 3.0 20.13 7.167 -28.502 -20.381 .4 76.1 25.9 82.0 25.I l.93 8 7.72 7.0 Fry 20.90 4 7.388 -40.0 5.933 125 -137 117-133 -122-153 ~ 123 137 7.4-88.349 .5 . of tests 95% Confidence limits 95% Confidence limits 19.802 2.535 693 580 461-1033 240-1192 572-841 343-980 2.494 .157 3.630 6.9 24.222 .722 11.04 7.0 24.19.592 13.07 7.4-88.433 7.868 15.a.08 6.069 11.96 2 2 8 2 7 7 28 6 .1 25.818 15.90 7.3.59 5.79 7. Acute toxicity of HCN to bluegill eggs and sw/m-up fry expressed as 96-hr LCS0 and median lethal threshold or hatching concentrations Mean test conditions Log-probit regression analysis 96-hr L C 5 0 L T C o r hatch L C 5 0 ~ DO mg/L pH Treatments a b No.279 10.086 .Table 4.g/L 95% Confidence limits ~g/L 95% Confidence limits Egg__2s 25.129 124-133 113-128 Table 5.616 11.98 7.39 4.236 3.91 4 4.70 7.89 6 24.618 .494 -23.

92 7.7 15.1 .955 -25.734 --- 288 ->329 >317 >389 >276 >357 295 337 >395 >338 270-307 ------256-339 202-561 --- -130 159 125 147 <109 < 170 --<179 <165 131-194 92..190 -17.0 99.87 7.5.1 18.11.261 --44.0 24.2 -81.7 113 120 125 ---81.97 6.006 8.003 11.0 3.340 --6.099 .249 2.2 -103-112 -86.431 .0 108 -99.0-308 .813 2. of tests b /zg/L 95% Confidence limits p.20 6.15.36 4.692 I 1.671 -19.1 75.593 13.0 14.072 ---10.485 -30.0 14.982 --31.1 -m m 108 99.90 7.8 66.173 -21.1 14.4 8 9.0 25.48 5.94 7..06 3.160 83.7 -87.71 7.86 6 6 5 6 7.0 -<92.92 8.849 4.08 8.05 6.910 --- 6.86 1 3 2 1 6 7.32 7.1.8-227 109-133 115-135 -61.9 24.0-93.1 25.u p fry e x p r e s s e d as 96-hr LC50 and median lethal threshold or hatching concentrations Mean test conditions Log-probit regression analysis 96-hr LC50 L T C or hatch LC50 ~ DO mg/L pH Treatments a No.84 5.70 7.050 -14. A c u t e t o x i c i t y o f H C N to bluegill j u v e n i l e s e x p r e s s e d as 96-hr L C 5 0 a n d m e d i a n lethal t h r e s h o l d c o n c e n t r a t i o n s Log-probit regression analysis 96-hr LC50 LTC .90 7.7 113 120 125 -104-112 86.80 2 DO mg/L pH Treatments a b #g/L p.0 15.7-170 70.76 7.04 7.8 -- 95% Confidence limRs ('3 ~z 8.72 7.T a b l e 6.1 17.0 Fry 10.283 -25.9 -5 -6 3 -10 9 12 15 16 12 --40.0 87.< Mean test conditions ID ~ 6.03 7.78 7.17 7.491 .17 6.66 2 8 3 6 6 8 -- 14.87 7.0 14.04 3. of tests 95% Confidence limits -59.83 7.g/L No.44 7.90 2 2 2 2 7.00 7.96 6.0-116 55.0-116 55.968 -14. A c u t e t o x i c i t y o f H C N to y e l l o w p e r c h e g g s a n d s w i m .0 18.90 7.3-64.12 7.839 -16.8 20.609 10.07 7.847 .160 -32.g/L 95% Confidence limits Eggs 14.766 17.797 18.0-85.0 14.0-93.900 9.8-227 109-133 115-135 T a b l e 7.0.94 2 2 2 4 2 -8 8 --7.35 6.

13 7.7-101 -97.711 -31.03 7.5 -68.78 2 7.037 12. o f tests gg/L 95% Confidence fimits ttg/L 95% Confidence limits Eggs 7.74 5 7.82 6 7.149 -5.0 10.0-95.05 3.964 -30.993 3.19.69 5 7.2 -91.0 4 11 6 6 7 7 8 -2.453 4.8--104 -982-106 -102-115 -- -87.502 5.790 -21.88 7.0 21.538 18.96 8.0 63.214 12.2 -88.8 -95.768 -19.56 5.9 -102 -108 -- 85.80 7.1-150 290-423 -403-665 -193-342 -- --106-270 -107-401 -143-185 .329 -30. A c u t e t o x i c i t y o f H C N t o b r o o k t r o u t e g g s a n d s a c f r y e x p r e s s e d a s 9 6 .650 13.10 6.6-105 -101-114 T a b l e 9.266 14.367 -28.8-96.6 -93.98 3.4-257 -43.868 12.172 12.0 10.0 5.4-92.0 -93.5 ~ 101 -107 -83.68 7.10 7.0 <71 -169 -- 207 -163 13.069 90.114 -9.496 3.891 5. of tests 95% Confidence limits 95% Confidence limits 15.082 17.0 10.935 -21.1 13.030 3.83 6 7.0 18.09 6. A c u t e t o x i c i t y o f H C N Mean test conditions Log-probit regression analysis 96-hr LC50 LTC to y e l l o w p e r c h j u v e n i l e s e x p r e s s e d as 96-hr L C 5 0 a n d m e d i a n lethal t h r e s h o l d c o n c e n t r a t i o n s *C 6.82 6 DO mg/L pH Treatments a b t~g/L /~g/L t~ No.00 7.71 7.50 6.13 5.8-95.h r L C 5 0 a n d m e d i a n l e t h a l t h r e s h o l d o r h a t c h i n g concentrations Mean test conditions Log-probit regression analysis 96-hr LC50 LTC or hatch LC50 ~ DO mg/L pH Treatments a b No.327 108 350 -518 -257 -- 77.512 11.0 -75.2 -75.84 2 7.9-219 O'3 Sac f~ 10.1 20 20 22 22 13 11 13 13 20 21 17 18 -17.4 -94.T a b l e 8.77 2 2 3 3 1 6 -10 -4 -2.367 >212 >232 >228 >242 >232 ------ 118 <98 155 ~ 98.062 13.96 3.2 -69.4 21.939 -3.573 -19.842 -4.8-82.0 -86.575 17.79 7.095 -3.518 6.0 21.0 21.1 10.478 -8.0-222 -93.82 6 7.206 .78 2 4 7.96 6.843 -19.735 -2.695 3.215 18.743 --I.64 5.0 -91.989 -7.7-82.0 10.225 --3.323 -19.

To test the response of fathead minnows.96 mg/L DO demonstrated a marked influence of different oxygen concentrations on HCN toxicity. The median lethal concentrations at hatching for tests conducted at about 6 mg/L DO varied from 98. When DO levels were 4. field stock was collected when lake temperatures were close to the target test temperatures. At the intermediate level of 6.82 mg/L. and rainbow trout juveniles was approximately threefold. while at 7. the slope for a specific species is smallest for egg bioassays and then increases with successive advances in development to that determined for juveniles.90. Sac fry tests at 10. The 95% confidence limit for this LC50 value is 55. respectively.9 tzg/L I-ICN.7 t~g/L.02. and 8.7-58.96 mg/L DO they were 518 and 207/zg/L HCN. and 96. respectively. but did change with life history stage.50 to 7.9. Toxicity tests with juvenile brook trout demonstrated little difference between the 96-hr and LTC values and a positive correlation between the increase in resistance to HCN and an increase in either temperature or dissolved oxygen (Table 11). The 96-hr LC50 and LTC at 3. respectively (Table 9). Eggs of all species tested were the most resistant life history stage with fry and juveniles demonstrating similar sensitivities. and 13~ the LTC values at comparable DO levels of about 6 mg/L were 73. Since the tests reported above were run at different times and on different lots of fish.7. respectively.3.03 mg/L DO the corresponding values were 350 and 169 tzg/L HCN.0 to 155/zg/L HCN with no correlation to test temperature observed (Table 9). 8. The difference in median lethal concentration between field stock fathead minnows. and 97.0~ were determined to be 69. 6.5 tzg/L HCN. Swim-up brook trout fry showed a response similar as sac fry to DO with no correlation being observed with test temperature. and 103 tzg/L HCN. and 8.8 mg/L DO were the most sensitive of all species tested with a 96-hr LC50 of 57. it seems desirable to eliminate to whatever extent possible these variables.9. 6. respectively (Table 10).00. the LTC values at 10.8.& C and 3.9 ~ 10. This procedure was followed in an attempt to evaluate the toxicity of HCN to fish subjected naturally to various temperatures at different times of the year. Fish were tested at 5~ . There was little difference between the 96-hr and LTC values. the most resistant species tested. 83.2/zg/L HCN. Slopes of the log-probit toxicity curves were similar for comparable stages of all species tested.04.06. The LTC values at 3. When the test temperature was 6.50 mg/L DO were 108 and less than 71/zg/L HCN. Rainbow trout juveniles tested at 10~ and 8. the 96hr LC50 values were all greater than 212/zg/L HCN. In general.8.00.Toxicity of HCN to Fish 333 Brook Trout In brook trout egg tests conducted at about 7~to 13~ and 4 to 8 mg/L DO.02 mg/L DO were 53. 88. Effect of Temperature on Acute Toxicity A number of tests with different species seemed to indicate a marked positive correlation between resistance to HCN and temperature rather than a negative one as might be anticipated. 83. 83.4.

634 21.2 -71.02 6.7.0 10. o f tests 95% Confidence limits 95% Confidence limits 7.5-82.04 20 9.5-96.0 10.9 -83. A c u t e t o x i c i t y o f H C N t o b r o o k t r o u t s w i m .137 6.377 !1.8-62.7 -85.2 7.290 32.8-106 -- -53.1 -77.944 8.0 10 10 37 39 14 15 19 21 -10.82 6 7.4 -76.435 29.1-116 78.8 -86.663 9.00 8.916 -- 75.7 -97.80 14 3.01 9.5-105 -94.898 .891 -35.695 86.0 71.7 13.986 16.0--88.8 -88.3 70.9 -83.000 28.2-103 -91.507 24.962 .042 -42.90 7.5 -93.914 -12.851 -50.996 10.0 13.524 7.613 14.85 6 7.0 10.180 .0 6.0 10.4-104 --- DO mg/L pH Treatments a b /alL p. o f tests /tg/L 95% Confidence limits /zg/L 95% Confidence limits 6.1-103 -95.6-76.06 8.g/L No.0 13.195 55.8 -85.4 -103 -87.T a b l e 10.5 -82.7 -81.153 23.346 .0 -77.0 -92.6 -106 -92.883 26.84 4 DO mg/L pH Treatments a b No. A c u t e t o x i c i t y o f H C N Log-probit regression analysis 96-hr LC50 to brook trout juveniles expressed as 96-hr LC50 and median lethal threshold concentrations LTC Mean test conditions ~ 6.04 8.8--94.9 -96.582 --82.709 -12.061 -47.7 -100--112 -79.74 6 8.0 10.2-80.585 27.4 -84.9-101 -- .79 8 7.04 8.4 -93.04 7.793 -19.3 -83.872 -17.1-92.9-92.017 -39.8.659 11.000 -46.9-102 --- -73.9 -88.19.866 -31.0-90.8 -69.0-86.2-92.954 6.5 112 -62.82 6.896 -53.06 2 7.2-98.26 4.817 -28.1 8 7 6 6 10 10 14 9 !! 10 5 5 8 8 -- -41.910 -36.0 -75.5 ~ t12 71.08 1 7.08 2 7.02 6.u p f r y e x p r e s s e d as 9 6 .2 -- 46.h r L C S 0 a n d m e d i a n l e t h a l t h r e s h o l d c o n c e n t r a t i o n s Log-probit regression analysis t~ 96-hr LC50 LTC Mean test conditions ~ 6.995 -21.11.874 8.9 6.73 6 10.504 20.8 ~ 88.0 -46.698 12.303 -59.372 7.2-90.8-66.40 8.9 T a b l e 11.84 4 8.83 6 14 7.8 10.569 -- 24.90 6 7.486 19.7 -100 -98.6 -62.

except for rainbow trout which are appreciably more sensitive than brook trout or other species used. but their response is often so variable that definite assignment of an acute toxic concentration of HCN cannot be made with assurance. a positive relationship between temperature from 4. there was approximately a twofold increase in tolerance between 8. Therefore. The increase in sensitivity of field stock fathead minnows at temperatures above 15~ a relationship not observed for laboratory reared individuals. When acutely lethal concentrations are used to estimate "safe levels" of HCN.0 mg/L (Table 6). In the tests run with bluegills at dissolved oxygen concentrations of about 6. The range of acutely toxic concentrations of HCN between species is not great in the juvenile stage.0 mg/L (Table 12). The tolerance increased at a relatively uniform rate with temperature increases to 12~ but between 12~ to 18~ the tolerance increased rapidly with successive 30C intervals. In a series of tests run during a two-month period with brook trout from a single hatchery lot.4 ~ and 24. With field stock fathead minnows sensitivity also increased at temperatures of 20~ and above. A slight increase in tolerance to HCN was observed with a decrease in test temperature from 30~ to 20~ The corresponding 96-hr LC50 values.0 ~ to 18. The implication of lowered oxygen and lower temperatures during cold weather seasons in systems receiving cyanide wastes should be carefully evaluated in the development of effluent and stream standards set by regulatory agencies. Temperature has a marked effect on acute toxicity of HCN with juvenile fish in general becoming more sensitive at lower temperatures. respectively (Table 13).0~ and tolerance to HCN was more clearly apparent than for fathead minnows. was probably a result of increased environmental stresses imposed on field stock individuals during the hot summer months.0/zg/L at 18~ and the LTC from 51. respectively.0/zg/L at 4.&C to 143. All tests were at a dissolved oxygen concentration of about 6.9~ Discussion Determined 96-hr values for most species and their life history stages occur at a somewhat higher HCN concentration than when acute toxicity is measured at the median lethal threshold concentration.Toxicity of HCN to Fish 335 intervals from 30"C in August to 5~ in December 1975.0/~g/L. threshold tolerance also markedly decreased so at 5~ with a 96-hr LC50 greater than 167/zg/L the LTC reached after 35 days of exposure was 91. increased from 157 to 174/xg/L HCN. Oxygen levels below 5 mg/ L result in increased sensitivity for all juveniles tested." LTC values should be the basis of calculation. it appears that if application factors (Mount 1977) are applied to acutely lethal results to establish "safe levels. The 96-hr LC50 varied from 53. which were equal to the respective LTC's. Among the species tested at high oxygen levels the acute toxicity of HCN to juveniles varied from 57 tzg/L for rainbow trout to 191 tzg/L for field stock fathead minnows. it should be noted that chronic tests indicate "no effect" levels to be . As the test temperature decreased in the fall.8 to 138.5/zg/L. Fish eggs are the most tolerant life history stage examined. The maximum tolerance to HCN was observed in October and at 15~ with the 96-hr LC50 and LTC being 191 and 184 /zg/L HCN.

88 20.0 -96.16 5.0 7.673 -31.24 7.259 23.94 8.0 7.474 -26.h r L C 5 0 a n d m e d i a n l e t h a l t h r e s h o l d concentrations Log-probit regression analysis 96-hr LC50 LTC Mean test conditions *C 2 2 2 2 2 2 DO mg/L Treatment a b tLg/L /.0 -61.87 7.827 -18.83 9.626 21.3 -55.0 10.66 8.840 15.04 7.0 6.737 18.5 -135 -184 174 160 157 52. of tests t~g/L 95% Confidence limits /.331 16.99 8. A c u t e t o x i c i t y o f H C N concentrations Log-probit regression analysis 96-hr LC50 to b r o o k t r o u t j u v e n i l e s at t e m p e r a t u r e s f r o m 4 .160 11.~g/L 95% Confidence limits 4.24 7.5 -143 49.Lg/L pH No.82 5.06 8.5 97.0 30.17.8 -61.0 6.849 -32.3-137 --- -51.0 15.5-55.07 6.T a b l e 12.00 5 4 -4 -4 -5 5 -- -28.0 7.15 6 7 5 8 8 8 6 6 -12.7-71.957 >167 165 -191 -174 160 157 -140-194 -178-205 -158-192 140-182 126-194 91.82 15.09 7.757 -17.02 7. of tests 95% Confidence limits 95% Confidence limits 5.483 -32.301 --25.282 17.30 7.7 -66.532 -44.02 19.135 --32.8 -68.92 6.799 -20.809 .903 --31.h r L C 5 0 a n d m e d i a n l e t h a l t h r e s h o l d Mean test conditions LTC ~ 2 2 2 2 2 2 DO mg/L Treatments a b pH No.6 138 -48.0 25.3-57.2 --79.8 -59.995 11.242 9. 0 ~ to 18~ e x p r e s s e d as 9 6 .19 7.3 72.0-64.88 12.0-161 -129-141 -173-196 158-192 140--182 126--194 T a b l e 13.3 -72.3-118 -- .02 7.0 ------69.804 -21.0 7.128 9. A c u t e t o x i c i t y o f H C N t o wild s t o c k f a t h e a d m i n n o w j u v e n i l e s at 5~ i n t e r v a l s f r o m 5 ~ to 30~ e x p r e s s e d a s 9 6 .234 -- 53.0 6.0 7.374 9.403 -19.903 -15.

262 (1977).. Jr. Broderius: Chronic effects of hydrogen cyanide on the fathead minnow. Chem. 1977. p. Mount. J. Amer. but the possibility of increased sensitivity and adverse effects from this factor should not be ignored. Bioassay methods for acute toxicity.. No chronic tests have been conducted incorporating low temperature cycles. 99 (1949). Bridges.. References American Public Health Association. W. 823 (1962). 253 (1958). 105. D. in relation to concentrations of molecular hydrocyanic acid.: Sodium cyanide as a fish poison. G. 11. American Water Works Association. Fish. and S. L. Amer. In press (1978). Berkeley: University of California Press (1973). Smith. II. Experimental investigations of ammonia and hydrocyanic acid poisoning). and Water Pollution Control Federation: Standard methods for the examination of water and wastewater. 1977) and between 5 and 11 /zg/L for brook trout (Koenst et al. L. M. 3 ed. L. Woker: Beitrage zur toxicologic der fishche. Smith. Technol.. and S. and D. Exp.. R. Tubb (ed. D. Inorg. L. J. U. Res. In press (1977). M. Environ. T. J. 1. Jr. Fish.). R. Corvallis Environ. Doudoroff. G. J. J.. D. E1-Kandelgy: Toxicity of hydrogen sulfide to various life history stages of bluegill (Lepomis macrochirus). Izatt. Water Pollut. Ecol. Smith. Wuhrmann. D. J.S. J. Trans. Sci.. Dixon.I. Doudoroff.. and S. EPA600/3-76-038. and AS* values as a function of temperature for hydrocyanic acid dissociation in aqueous solution. Water Res. Fish. Minnesota: U.. Pharmacol. J. Soc. Leduc. T. Duluth. 3 ed. J. Bench: Thermodynamics of metal-cyanide coordination. J. and W. accepted December 3. Sci. 6 (1966). Ser. A review. Jr. Jr. Soc.: An assessment of application factors in aquatic toxicology. Corvallis. D. Schwiz. In Richard A.. 1977). London: Cambridge University Press (1971). Kimball. 13 ed. No. Ecol. Cyanide levels as low as 5/zg/L HCN inhibited spawning by bluegills (Kimball et al. L. Fish. Oseid. and F. Trans. Ther. Hydrol. J. Water Res. Environmental Protection Agency (1976). Schneider: Acute toxicity offish to solutions containing complex metal cyanides. 1977 . Broderius.: BMD biomedical computer programs. Kimball. S. 95. (Contributions to fish toxicology. Environmental Protection Agency (1977). II. R.: Probit analysis. 883 (1977). W. Jr. Brungs: A simplified dosing apparatus for fish toxicology studies. pk. Smith. T. Broderius: Chronic toxicity of hydrogen cyanide to bluegills.. Board Can. 1.. 210 (1948). Mount.. Oregon: U. Sprague. 96. Res. Manuscript received August I0. I. a symposium. Koenst. 11. 21 (1967). 793 (1969). K. Res. Fish and Wildlife Service. in press). Lind: Relative toxicity of free cyanide and dissolved sulfide forms to the fathead minnow (Pimephalespromelas Rafinesque). I. Ser. Experimentelle untersuchungen fiber die ammoniak. L. Environmental Research Laboratory-Duluth. and H. EPA-600/3-77-085. I.S. Z.S. L. M.. Litchfield. J. J. Pack. Lab. L. G. and S. Toxicity to fish of cyanides and related compounds. Broderius: Effect of chronic exposure of brook trout to sublethal concentrations of hydrogen cyanide.und blaus~turevergiftung. 183. and C. B. Amer. Rep. A.Toxicity of HCN to Fish 337 between 12 and 19/zg/L for fatheads (Lind e t at.. Smith. Control Fed. Recent advances in fish toxicology. G. P. Lind. Finney. W. Soc. L. 3. and R. Washington: American Public Health Association (1971). AH*. 49. R.. Wilcoxon: A simplified method of evaluating dose-effect experiments. P. L.: Measurement of pollutant toxicity to fish. Fish. L. Spec. Res. Trans. T. L. Jr. 442 (1976). Christenson..

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