This paper records some observations on the reactions of minnows to phenolic substances. The fish are placed in a horizontal tube, half of which is filled with flowing water and half with flowing solution of the concentration to be tested. Water and solution are very sharply differentiated.

Solutions of phenol, ortho- and para-cresol are highly toxic and appear to have some specific effect on the nervous system of fish, causing them to lose their sense of balance and capability of co-ordinated movement. The time the solutions take to produce this effect is much shorter than the survival time; in a 0.01% solution of para-cresol, for example, a minnow loses its sense of balance in about 70 sec. but takes about 30 min. to die.

Over the concentration range 0.04-0.0004% minnows appear to have little or no power of discriminating between phenol and water, swim into the solution and become intoxicated.

At concentrations of 0.03-0.04% minnows will avoid para-cresol and ortho-cresol solutions immediately and escape intoxication. At somewhat lower concentrations the fish venture into the solution, and this quickly destroys their capability of recognizing and avoiding it. The situation is markedly different from that observed in the case of lead and zinc salts; here the time the fish takes to establish an avoiding reaction to the solution is very much shorter than the time this takes to destroy its capability of doing so.

Phenolic substances form an important type of polluting chemical in industrial wastes; they are present in certain gheep dips, in coal tar and synthetic tar and in effluents from gas works, coke ovens and factories producing disinfectants. Shelford (1917) studied the effect of phenol on fishes and found that 0·1% solutions were quickly fatal; below this concentration the toxicity fell off rapidly. Ortho-cresol was rather more toxic than phenol; para-cresol and meta-cresol less toxic. Shelford also tested the reactions of fishes to these substances in the gradient tank and concluded that they were generally ‘indifferent’ or ‘positive’ to the solutions. Mason-Jones (1930) investigated the toxicity to fish of a wide range of substances found in tar, using perch, yearling trout and trout fry and describes the symptoms produced by phenol and the cresols. At the higher concentrations there is a very characteristic rapid loss of the sense of balance; the fish swims with a wild, dashing movement and turns on its side; the gill covers, at first widely opened, are tightly closed; the respiratory movements become irregular and feeble, and before dying the fish may turn turtle. After death in strong solutions of phenol the gill tissues may be collapsed, but this symptom is not evident in the case of the cresols. Alexander, Southgate & Bassindale (1935, pp. 107–8) measured the toxicity of para-cresol and phenol to rainbow trout and also observed this rapid effect on the equilibrium of the animal, concluding that their experiments confirmed the opinion of other workers that phenolic substances act as specific poisons on the nervous system of fish. Ellis (1937, pp. 416–17) tested the toxicity of phenol to the goldfish and also states that it produces a paralysis of neuromuscular mechanisms. A detailed discussion of the pharmacology of phenol and the cresols is given by Edmunds & Gunn (1936, pp. 734–40); their action on fishes is not discussed but it is stated that in the frog phenol causes fibrillary twitching in the muscles followed by tonic convulsions and then a complete paralysis of the central nervous system.

In papers recently published in this Journal the writer (Jones, 1947, 1948) has described an apparatus for testing the reactions of fish to toxic solutions; this differs from the Shelford gradient tank in that water and solution are sharply differentiated so that the fish is presented with a definite concentration step, not a gradient. The substances tested included salts of lead, zinc, copper and mercury, hydrogen sulphide and ammonia. The present paper describes similar experiments with phenol, para-cresol and ortho-cresol.

The apparatus was essentially similar to that used for the experiments with sulphide solutions (Jones, 1948, pp. 23–4). One 10 1. aspirator with inflow and outflow supplied water to the right or left side of the experiment tube, and solution was supplied to left or right from a 15 1. aspirator fitted with an air intake tube. All solutions were made up immediately before use with tap water from the same supply as that connected to the water-supply aspirator, and the arrangement for automatically making up fresh solution which was used for the sulphide solutions was not necessary. The tube in which the fish were placed was a little larger than that used in the previous studies, measuring 34 mm. in internal diameter; this permitted the use of fish 28–32 mm. in length. With this larger tube a little difficulty was experienced in obtaining a sharp, vertical separation of water and solution in the centre; this was overcome by fitting small glass baffles in front of the internal apertures of the inlet tubes.

The method of recording the results follows that devised by Shelford (1917). The space between two parallel lines represents the length of the tube, minutes are marked off on a descending vertical scale and the movements of the fish are copied. In all the experiments only one fish was placed in the tube; as one of the chief symptoms in the case of phenolic substances is very rapid and erratic swimming, the use of more than one is impracticable.

Survival curves for 25–30 mm. minnows in phenol, para-cresol and ortho-cresol solutions are drawn in Fig. 1. The concentration is expressed in percentage (g. phenol, etc., per 100 ml. solution). On this basis phenol is distinctly the most toxic of the three and ortho-cresol the least toxic. The behaviour of the fish conformed well to the descriptions given by other workers. In the stronger solutions the sense of balance is lost almost at once, the fish falling over on to its side; there are wild, dashing movements at first, later only feeble attempts at swimming and shallow, flickering opercular movement. In the weaker solutions the sense of balance is also affected very quickly, the fish falls over but may regain its equilibrium temporarily a number of times; finally, however, it loses all power of co-ordinated movement and remains on its side with occasional attempts at swimming and respiratory movements which are almost imperceptible. This helpless condition prevails for the rest of the survival time, the death point being rather ill-defined.

Fig. 1.

Survival times for minnows in solutions of phenol, ortho-cresol and para-cresol. The points are means of three determinations. Temp. 17° C. pH of solutions 6·6-6·8.

Fig. 1.

Survival times for minnows in solutions of phenol, ortho-cresol and para-cresol. The points are means of three determinations. Temp. 17° C. pH of solutions 6·6-6·8.

Twelve reaction experiments were carried out with phenol at concentrations of 0·04·0·0003%, and four representative results are given in Fig. 2. The minnows showed little or no capacity for recognizing and avoiding the solution at any concentration. In the 0·04% experiment the phenol was admitted on the right when the fish was in the left half of the tube; however, it entered the solution at once, there it gulped and moved very jerkily, visited the water zone and returned twice, at 2 min. lost its sense of balance, swam wildly up and down the tube about six times, and finally rested on its side almost motionless. In the 0·01 % experiment the phenol was run in on the side occupied by the minnow; for over a minute the fish displayed no sign of irritation, floating gently on the current towards the water zone. At 4 min. symptoms of intoxication suddenly developed with wild swimming up and down, and at 6 min. the sense of equilibrium was quite lost; the fish now found its way into the water where it kept up feeble movement; at 8 min. it had recovered to a considerable extent and was swimming upright once more. At 9–10 min. the water-phenol flow was reversed, but again the minnow showed no power of discriminating between water and solution and at 13 min. lost its sense of balance for the second time. It recovered quickly on being placed in an aquarium.

Fig. 2.

The reactions of minnows to 0·04, 0·01, 0·004 and 0·0004% solutions of phenol. pH of water 6·8, of solutions 6·6-6·8. Temp. 16·5-17·5° C. Survival times at these concentrations (fish continuously immersed in the solution) 4 min., 8 min., 24 min., 40-50 hr.

Fig. 2.

The reactions of minnows to 0·04, 0·01, 0·004 and 0·0004% solutions of phenol. pH of water 6·8, of solutions 6·6-6·8. Temp. 16·5-17·5° C. Survival times at these concentrations (fish continuously immersed in the solution) 4 min., 8 min., 24 min., 40-50 hr.

The same result is evident at 0·004%; the fish visited the solution several times, and this resulted in intoxication and collapse. At 0·002% minnows would enter the solution half of the tube, remain almost motionless for 1–2 min. and then fall over. A 0·001 % solution is much less rapid in its effect, and a spell of several minutes in the phenol does not produce anything worse than a slight uneasiness and irregularity in respiratory movement. The 0·0004% experiment included in Fig. 2 was run for 13 min., during which time the fish crossed the water-solution junction at least 30 times, but no avoiding action or symptoms of intoxication developed. This concentration is near the threshold of toxicity for phenol.

Twelve experiments were run with para-cresol covering the concentration range 0·03–0·002 %. Four records are given in Fig. 3; this includes two results for 0·03 % to illustrate the good measure of agreement observed in different experiments. At 0·03%, in marked contrast to what is seen in the case of phenol, a most definite avoiding action was evident. The minnows would not enter the solution at all; if they swam to the water-solution junction they would halt, make some gobbling respiratory movements and retreat. Para-cresol is so toxic at this concentration-that even these momentary contacts were sufficient to produce some measure of intoxication, and the fish appeared somewhat distressed at the end of the experiment but had not lost their sense of balance. At somewhat greater dilutions the animal’s capability of detecting and avoiding the solution seems to diminish very sharply. A fanmeasure of avoiding action was evident at 0·02%, but at 0·01% (Fig. 3) the fish penetrated the cresol about six times, and this resulted in a sudden attack of furious swimming up and down ending in collapse. Recovery began at 8 min. At still greater dilution the water-solution junction is usually crossed with no hesitation; in experiments at a concentration of 0·004 and 0·003 % the fish appeared to be less happy in the solution than in the water, and an extremely vague negative reaction might be observed. In the fourth record in Fig. 3 this vague tendency to prefer the water zone was shown at the beginning of the experiment; at 12 min., however, the minnow swam far into the solution and remained motionless for nearly half a minute. At 0·003 % the intoxicating power of para-cresol solutions is declining, and at the end of the experiment the fish was swimming more or less normally.

Fig. 3.

The reactions of minnows to 0·03 (two records), 0·01 and 0·003% solutions of para-cresol. pH of water and solutions 6·8. Temp. 17·5-18° C. Survival times at these concentrations (fish continuously immersed in the solution) 7 min., 27 min., 4-5 hr.

Fig. 3.

The reactions of minnows to 0·03 (two records), 0·01 and 0·003% solutions of para-cresol. pH of water and solutions 6·8. Temp. 17·5-18° C. Survival times at these concentrations (fish continuously immersed in the solution) 7 min., 27 min., 4-5 hr.

The writer has shown (1948) that the minnow will display a definite avoiding reaction to very dilute lead nitrate solutions. This avoiding reaction is quickly developed; at 10−3 N it seems immediate and at 10−5 N appears in about 5 min. These solutions take a considerable time to produce any marked toxic effect; the survival time of minnows in 10−3 N-lead nitrate is about 4 hr., and during the greater part of this time the fish retains its sense of balance and swims more or less normally. Gasterosteos aculeatus, similarly, will establish an avoiding reaction to 10−4 N-lead nitrate in about 5 min.; at this concentration the survival time is about 10 hr., and the only toxic symptom which develops in the first hour is a gradual rise in the rate of the respiratory movements. The general position in the case of zinc salts is similar. In an experiment with Pygosteus pungitius (Jones, 1947, p. 117) it was found that after 36 min. immersion in a 0·01 N-zinc sulphate solution (fatal in about 100 min.) the fish would quickly select the water zone of the apparatus. Lead and zinc salts kill fish by impairing the respiratory efficiency of the gills and appear to have no specific effect on the nervous system; the time taken by the animal to establish an avoiding reaction to the toxic solution is a small fraction of the time this takes to destroy its capability of doing so. The general situation in the case of para-cresol is very different; here it would appear that the solution has two distinct effects, an irritating effect which provokes the avoiding reaction and an intoxicating effect which destroy’s the animal’s power of swimming in a co-ordinated manner. The irritating effect is probably predominant at high concentrations, so that an avoiding reaction is immediately or very quickly established, but at lower concentrations the solution rapidly becomes less effective as an irritant but little less effective in intoxicating power; the time the fish takes to avoid the solution and the time this takes to destroy its capability of doing so thus come to be separated by the narrowest of margins. At 0·03 % paracresol the sense of balance begins to fail in about 20 sec. (see the data set out in Fig. 4), but as the fish will avoid the para-cresol zone almost immediately it escapes intoxication. At 0·01 % para-cresol the fish does not appear to be irritated by the solution but persists in venturing into it; the paralysing effect at this concentration is little less rapid than at 0·03 %, and so an avoiding reaction is never established. At 0·003 % it is probable that the solution has a sufficient irritating effect to induce some measure of preference for the water zone, but at this dilution only about 4 min. immersion in the solution is needed to disturb the animal’s equilibrium and the situation is still critical.

Fig. 4.

The effect of para-cresol solutions on the minnow’s sense of balance; the lower points are the times at which the fish begin to have difficulty in swimming upright, and the upper points are the times at which the fish appear to lose completely their sense of balance and ability to swim in a co-ordinated manner. All times are very approximate. Temp. 18° C.

Fig. 4.

The effect of para-cresol solutions on the minnow’s sense of balance; the lower points are the times at which the fish begin to have difficulty in swimming upright, and the upper points are the times at which the fish appear to lose completely their sense of balance and ability to swim in a co-ordinated manner. All times are very approximate. Temp. 18° C.

With ortho-cresol eight experiments were run over the concentration range 0·04–0·003 %, and four are recorded in Fig. 5. The results are very similar to those obtained with para-cresol; at 0·04% the fish avoided the solution very well, but at greater dilutions the animal’s capability for recognizing the solution seems to decline rapidly, for at 0·02% the fish persisted in entering it with the usual result, in the 0·01% experiment the minnow became intoxicated very quickly, and after the usual series of wild rushes up and down stopped on its side in the water zone. Here it regained its equilibrium in about 3 min., at 10 min. swam into the solution, had a second fit of intoxication, settled in the water and at 16 min. was recovering again. At 0·003 % the same inability to recognize the cresol was evident, but the symptoms of intoxication took several minutes to develop.

Fig. 5.

The reactions of minnows to 0·04, 0·02, 0·01 and 0·003% ortho-cresol. pH of water and solutions 6·8. Temp. 17-18° C. Survival times at these concentrations (fish continuously immersed in the solution) 13 min., 15 min., 43 min., 22 hr.

Fig. 5.

The reactions of minnows to 0·04, 0·02, 0·01 and 0·003% ortho-cresol. pH of water and solutions 6·8. Temp. 17-18° C. Survival times at these concentrations (fish continuously immersed in the solution) 13 min., 15 min., 43 min., 22 hr.

In addition to the foregoing experiments with pure solutions of phenol and cresol a number of trials were made with various dilutions of a phenolic effluent supplied by a firm of manufacturing chemists. The neat effluent was a clear solution of pH 6·8 and strong ‘carbolic ‘odour. The total concentration of phenolic substances, including phenol, cresols and a very small amount of xylenol, was stated to be approximately 0·0762% w/v, the chief other substances present were sodium sulphite, sodium chloride and sodium sulphate. The behaviour of the minnows was essentially similar to that observed in the experiments with pure solutions of cresols. To a 1 : 13 dilution they reacted very definitely, refusing to enter the solution at all and displaying obvious signs of great irritation on encountering it; continuously immersed in the solution they lost their sense of balance almost at once and died in about 10 min. A 1 : 6 dilution also produced an avoiding reaction but of rather less definite form. A 1 : 10 dilution of the effluent, fatal in 35 min., did not appear to be recognized, the fish entering it with no hesitation, and 1 : 20, 1 : 40 and 1 : 100 dilutions similarly produced no response.

In the case of phenol there appears to be little chance of fish avoiding a solution of any concentration, and while high concentrations of ortho- and para-cresol may be detected and avoided the animal does not immediately reject solutions of greater dilution whose power of upsetting its sense of balance is little inferior. Once the power of co-ordinated movement is lost there can be little possibility of developing any definite negative reaction. Phenol and the cresols are therefore highly dangerous polluting substances, but their toxicity falls off very quickly with decrease in concentration, and it is probable that at great dilution they are not as stable as lead, copper and zinc salts. Hence it is possible that a slight degree of pollution may do no harm, provided that the effluent is immediately diluted to the maximum degree possible and not allowed to run into pools where it may mix very slowly with the stream water to form solutions of comparatively high concentration into which the fish may venture.

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