- 1 CHAPTER V
EFFECT OF ARSENIC COMPOUNDS
- 1.0.1 I. Presence of Arsenic in Plants.
- 1.0.2 II. Effect of Arsenic on the Growth of Higher Plants.
- 18.104.22.168 1. Toxic effect.
- 22.214.171.124 2. Effect of arsenic compounds on germination.
- 126.96.36.199 3. Do arsenic compounds stimulate higher plants?
- 1.0.3 III. Effect of Arsenic Compounds on Certain of the Lower Plants.
- 1.0.4 Conclusion.
EFFECT OF ARSENIC COMPOUNDS
I. Presence of Arsenic in Plants.
The occurrence of arsenic as an occasional constituent of plants has been recognised for many years. Chatin (1845)found that if a plant were supplied with arsenical compounds at the roots arsenic was absorbed, but that it was distributed unequally to the various tissues. The greatest accumulation of the element was in the floral receptacle and the leaves, while it was scarce in the fruits, seeds, stems, roots and petals. E. Davy (1859) commented on the presence of arsenic in plants cultivated for food. He grew peas in pots and watered them for a short time with a saturated aqueous solution of arsenious acid, the application being then discontinued. The plants, apparently uninjured by the treatment, flowered and formed seeds. On analysis arsenic was readily detected in all parts of the plant, including the seeds. Other analyses revealed the presence of the element in cabbage plants (from pots) and turnips (from field), both of which had been manured with superphosphate containing some amount of arsenic. This absorption of arsenic by the roots of plants was further established by Phillips (1882).
Various physiological workers have pointed out that this element is frequently or usually present in animal tissues.Cerný (1901) reached the general conclusion that minimal traces of arsenic can occur in animal organisms, but that these play no part in the organism and indeed are not constant in their occurrence. Bertrand (1902) established its presence in minute quantities in the thyroid glands of the ox and pig, hair and nails of the dog, and the feathers of the goose. Gautier and Clausmann (1904) realised the constant presence of arsenic in human tissues and recognised that it must inevitably be introduced into the body with the food. This led them to estimate the arsenic present in various animal and vegetable foods, some of their results being given in the following table.
Arsenic per 100 parts fresh substance in µ gr. (= thousandth part of a milligram).
|„||(from Franche Comté)||·85|
|Whole green cabbage||·2|
|Outside leaves of cabbage||·0||(absent)|
Arsenic was also found in wine and beer and in considerable quantities in sea water and various kinds of salt. Since it cannot be found in some things even in the least traces, the authors conclude that it is incorrect to say that the element is always present or that it is essential to all living cells.
S. H. Collins (1902) found that barley is able to absorb relatively large quantities of arsenic. The plants were grown in pots on soil which originally contained a certain amount of the substance, and various combinations of arsenic acid, arsenious acid and superphosphate were added. Particulars and details are not given by the author, except that arsenic was detected by Reinsch’s test in the grains from all the experimental pots, and in one case (not specified) in the upper and lower halves of the straw and in the threshed ears. The analyses of the soil at the close of the experiments showed the presence of 7–22 parts arsenious acid per million.
Wehmer (1911) quotes references to the occurrence of arsenic in Vitis vinifera. The element was detected in the ash of the must and its presence was attributed to treatment of the plants with arsenical compounds. In this connection it is interesting to note the observation of Swain and Harkins (1908), who, while acknowledging the absorption of arsenic from the soil by many plants, yet indicate that in the case of those plants which are exposed to smelter smoke the arsenic is deposited on the vegetation, and is not absorbed by the latter from the soil.
II. Effect of Arsenic on the Growth of Higher Plants.
1. Toxic effect.
(a) Toxic action of arsenic compounds in water cultures in the presence of nutrients.
The poisonous action of arsenic on plants has long been recognised. Chatin (1845) gave accounts of tissues poisoned by strong arsenical solutions. Nobbe, Baessler and Will (1884) carried on water culture experiments with buckwheat, oats, maize and alder, and found that arsenic was a particularly strong poison for these plants. When small quantities of arsenious acid (As2O3) were added to the food solutions, growth was measurably hindered by a concentration of 1/1,000,000 As (reckoned as As). The element only appears in plants in very small quantity and can never be detected in notable quantities. The aerial organs show the effect of arsenical poisoning by intense withering, interrupted by periods of recovery, but eventually followed by death. It was also found that if plant roots were exposed to the action of arsenical solutions for a short period, say ten minutes, and then were transferred to normal food solutions, the action of the poison was delayed, but eventually hindering of growth or death occurred, according to the strength of the poison used in the first solution.
At the same time that Nobbe, Baessler and Will were establishing the great toxicity of the lower oxide of arsenic, Knop (1884) was carrying the matter a step further by comparing the action of arsenious and arsenic acid and their derivatives on plant growth. He established the fact that while arsenious acid is a strong poison for maize plants, arsenic acid in small quantities is not toxic to the roots and that the plants can produce flowers and fruit in its presence. Arsenic acid applied as potassium arsenate proved to be harmful to young maize seedlings if the solutions contained ·05–·1 gm. arsenic acid per litre (= 1/–2/20,000 arsenic acid). If however the plants were allowed to form 10–15 leaves in a pure food solution and then when strongly rooted were transferred to a solution of ·05 gm. arsenic acid per litre, they were found to grow strongly and develope big healthy leaves. Careful measurements indicated that the development is unchecked by the addition of the poison, though arsenic was determined in the ash of the treated plants.
Stoklasa (1896, 1898) tested the effect of arsenic compounds on plant growth with special attention to their comparative relation to phosphoric acid. He corroborated Knop’s statement as to the greater toxicity of arsenious acid and arsenites in comparison with arsenic acid and arsenates, stating that 1/100,000 mol. wt. arsenious acid per litre causes definite trouble in plants, while with arsenic acid 1/1000 mol. wt. per litre first shows a noticeable toxicity. Water culture experiments were made with and without phosphoric acid, in each case with and without the addition of arsenic and arsenious acid. It was found that the arsenic acid was unable to replace the phosphoric acid, the plants decaying in the flower in the absence of the latter. In the complete absence of phosphoric acid, arsenic acid causes a strong production of organic substances up to the flowering time. The following figures were obtained with maize:—
|·002 gm.||As2O3||with P2O5||2·84 gm. dry wt.|
Comparative experiments with the two arsenical oxides showed that varying times were required to kill different plants. Young seedlings were brought into solutions containing 1/10,000 mol. wt. arsenious acid (= ·019 gm. As2O3 per litre) and the plants died in a very short time.
|Hordeum distichum||46 hours|
With ten times the strength of arsenic acid (1/1000 mol. wt. = ·23 gm. per litre) the plants took much longer to kill.
|Hordeum distichum||24·5 days|
Various experiments have been carried on at Rothamsted with peas and barley. With arsenious acid on barley a depressing influence is manifest even at a concentration of 1/10,000,000, while no growth at all is possible with 1/10,000 and upwards. Apparently the toxic action on the root ceases at a higher strength than on the shoot, as with 1/1,000,000 and less the dry weight of the root remains practically constant. At this same strength the shoots look better than the controls, but this is not apparent in the dry weights (Figs. 9 and 10). With peas the depression is again evident to 1/10,000,000, but the plants are more sensitive to the higher concentrations, as no growth can take place in the presence of 1/250,000 arsenious acid (Fig. 11). A striking difference is observed with arsenic acid on barley, as apparently this does not act as a toxic even with such comparatively great concentrations as 1/100,000, though possibly the shoot is slightly depressed by this strength (Fig. 12).
Fig. 9. Photograph showing the action of arsenious acid on barley in the presence of nutrient salts. (March 16th–May 9th, 1911.)
Fig. 10. Curve showing the mean value of the dry weights of ten series of barley plants grown in the presence of arsenious acid and nutrient salts. (March 16th–May 9th, 1911.)
Fig. 11. Curve showing the mean value of the dry weights of ten series of pea plants grown in the presence of arsenious acid and nutrient salts. (June 8th–July 21st, 1910.)
Fig. 12. Curve showing the mean value of the dry weights of ten series of barley plants grown in the presence of arsenic acid and nutrient salts. (Feb. 28th–April 24th, 1911.)
With sodium arsenite the dilutions were carried further, to 1/250,000,000, but this still depressed barley to some extent (Fig. 13). With peas the results vary somewhat in the different tests, the depression with 1/2,500,000 and less being usually slight, though occasionally it is much more strongly marked (Fig. 14). In a single series with sodium arsenate barley was apparently unaffected by a concentration of 1/1,000,000, but from this point down to 1/250,000,000 a constant depression showed itself, which was paralleled by a similar depression in the sodium arsenite series from 1/25,000,000 to 1/250,000,000, the curves grading downwards instead of up towards the normal. With peas sodium arsenate has little or no action, though it is just possible that the rather irregular curves indicate a very slight depression below the normal throughout.
Fig. 13. Curve showing the mean value of the dry weights of ten series of barley plants grown in the presence of sodium arsenite and nutrient salts. (Feb. 10th–April 18th, 1913.)
Fig. 14. Curve showing the mean value of the dry weights of ten series of pea plants grown in the presence of sodium arsenite and nutrient salts. (June 27th–Aug. 10th, 1911.)
(b) Toxic effect of arsenic compounds in sand cultures.
Comparatively few tests seem to have been made as to the action of arsenical solutions in sand cultures. Stoklasa (1898) repeated his water culture work, using sand as a medium, and found analogous results by the two methods, i.e. that arsenites are far more toxic than arsenates, and also that the degree of toxicity of a salt varies with the plant to which it is applied, as was shown by the fact that different plants lived for varying times when treated with similar strengths of solution.
(c) Toxic effect of arsenic when applied to soil cultures.
Daubeny (1862) watered barley plants with a solution of arsenious acid, 1 ounce in 10 gallons, five times in succession, and found that the crop arrived at maturity about a fortnight earlier than the untreated part of the crop, though the amount harvested was rather less. With turnips four waterings had no effect upon the time of maturity, but again the crop was slightly decreased. The analyses made indicated that no arsenic was taken into the tissues, but that it merely adhered to the external surfaces.
Gorup-Besanez (1863) mixed 30 grams arsenious acid with 30·7 litres soil, growing two plants on this quantity of earth. Most of his experimental plants (Polygonum Fagopyrum, Pisum sativum, and Secale cereale) developed normally, butPanicum italicum died soon after the plants appeared above the surface, the leaves being very badly coloured. Analyses by Marsh’s test showed no trace of arsenic in 20 grams dry matter from Secale cereale, but in 148 grams Polygonum Fagopyrum the presence of arsenic was evident, though the mirror formed was weak. With such a large proportion of arsenious acid in the soil it seems hardly conceivable that the plants were not injured to some extent, and also it is probable that with more careful analyses arsenic would have been detected in those instances in which its presence was denied. Yet it must be remembered that Davy (1859) had treated pea plants in pots with a saturated solution of arsenious acid for a short time and had stated that the plants were uninjured. Thus both Gorup-Besanez and Davy concur in the opinion that Pisum sativum is indifferent to relatively large quantities of arsenious acid when presented in the soil, whereas the Rothamsted experiments show that in water cultures the plant is extremely sensitive even to minute traces of the substance. It is possible that the arsenic in the solution added to the soil enters into combination with other substances, forming insoluble compounds, thus being removed from the sphere of action and rendered unable to affect plant life. If this be so, the apparent immunity of certain plants to arsenious acid is explained. F. C. Phillips (1882), in his experiments on various flowering plants, such as geraniums, coleas and pansies, found that compounds of arsenic in the soil exercised a distinct poisoning influence, tending, when present in large amount, to check the formation of roots, so that the vitality of the plant was so far reduced as to interfere with nutrition and growth, or even to kill it outright. He also stated that traces of arsenic were found in all the plants grown upon the poisoned soil.
In this connection it is interesting to note that a certain proportion of arsenic is frequently present in the superphosphate used as manure. In view of the known toxicity of arsenical compounds to plant life the question arose as to whether superphosphate manuring would exercise a detrimental influence on account of its arsenic content. Experiments carried out by Stoklasa (1898), however, indicate that there is not sufficient arsenic in maximum doses of superphosphate to exercise a toxic action in the field.
(d) Physiological considerations.
The physiological action of arsenic compounds on plant life early attracted the attention of investigators. Chatin (1845)put forward some rather curious and unexpected considerations with regard to this action. He stated that the effect of arsenic on plant growth is determined more by the constitution and temperament of individual plants than by their age, and that apparently difference in the sex of plants is of no significance. The chief determining agent, however, is the species, and Chatin found that as a general rule Cryptogams are more sensitive than Phanerogams, and Monocotyledons than Dicotyledons, as is shown by the fact that under treatment the former perish first. Some extreme exceptions exist, though, asMucor mucedo and Penicillium glaucum will grow on moist arsenious acid, whereas leguminous plants are killed by an arsenical solution in a few hours. Chatin held the view that elimination of the poison succeeded its absorption, and that this elimination is complete if the plant lives long enough. Here again the species exerts a great influence on the excretory functions of the plants. Lupins and Phaseolus are presumably able to eliminate in six weeks all the arsenious acid they can absorb without dying. Most Dicotyledons need 3–5 months, while Monocotyledons retain traces of poison for six months after its absorption. Lichens are said to eliminate it more slowly still. Again, woody species are longer in freeing themselves than herbaceous, and young plants carry out the elimination more easily than old plants. The excretory function is influenced by other physiological factors such as dryness and season. The toxic effects and elimination are supposed to act inversely and parallel, the absorbed arsenious acid combining with alkaline bases, making a very soluble salt which is excreted by the roots. Calcium chloride is given as the antidote to arsenious acid, all soluble acid being “neutralised” by it. This view of the elimination of arsenic apparently did not gain much support, as no further references to the matter have so far come to light. In view of the work of some modern investigators (Wilfarth, Römer and Wimmer) on the excretion of salts by plant roots, the idea may prove of fresh interest. Chatin also found that moving or still air influenced the working of the poison, indicating that the external physical conditions affect the toxic action considerably. Nearly forty years later Nobbe, Baessler and Will found that, if transpiration were hindered by placing plants in a dark or moist room, it was possible to keep the plants turgescent in arsenic solutions for a long time without thereby increasing the toxic effect later on. The poisonous action proceeds from the roots, of which the protoplasm is disorganised and the osmotic action hindered. Finally, in the presence of sufficient of the poison, the root dies without growth.
Stoklasa (1896, 1898) again found that phanerogamic plants can withstand arsenic poisoning for some time in the dark or in CO2-free air, provided that glucose is given in the food solution. The arsenic poisoning is at its maximum during carbon assimilation by means of chlorophyll. The toxic action of arsenious and arsenic acids, especially in phanerogams, is due to injury to the chlorophyll activity. The destruction of the living molecule is far more rapid in the chlorophyll apparatus than in the protoplasm of the plant cell.
Thus it seems that the physiological cause of the toxicity of arsenic is partly a direct action on the root protoplasm, whereby its osmotic action is hindered, and partly a detrimental action upon those functions which are directly concerned with the elaboration processes of nutrition.
2. Effect of arsenic compounds on germination.
In view of the great toxicity of arsenic to plants in their various stages of development, one would naturally expect to find a similar action with regard to the germination of the seeds. Davy (1859) casually mentioned cases in which watering with arsenical solutions or dipping seeds in arseniated water prevented germination. Heckel (1875) found that arsenious acid checks germination and kills the embryo at relatively feeble doses, ·25 gm. to 90 gm. water. Guthrie and Helms (1903–4–5) carried out a systematic series of experiments to test the effect of arsenic compounds upon different farm crops. Various amounts of arsenious acid were added to soil in pot experiments, and the seeds of the several crops were then sown. With barley, wheat and rye 0·10% arsenious acid had little or no effect on germination, while an increase in the poison exercised a retarding action. Maize could withstand 0·40% arsenious acid without retardation being perceptible. The aftergrowth with the different crops varied considerably. The wheat plants with 0·10% arsenious acid grew all right at first, but later on they developed weakly. The toxic action increased rapidly as the strength of the poison rose in the different pots. Barley proved even more sensitive than wheat, for even 0·05% arsenious acid affected the growth adversely. After a time the plants with 0·05–0·06% recovered and grew strongly, though not so well as the controls, but those with 0·10% practically died off. Rye behaved in the reverse way from wheat. The plants with 0·10% were slightly checked at first but later recovered and made growth quite equal to the check plants. Growth was stunted with 0·20% arsenious acid, and the plants were killed with 0·30%, so that rye is far less sensitive than barley. With maize the growth was slightly affected with 0·05% As2O3, and increasingly so with greater quantities. It was also found that the action of 0·8% As2O3 was strongly adverse to the germination of all plants, and that above this strength germination was altogether prevented.
The results show very clearly how impossible it is to draw any general conclusions with regard to the action of arsenic compounds on plants, as they emphasise the strong individuality of the species in their reaction.
3. Do arsenic compounds stimulate higher plants?
The question of stimulation due to arsenic does not seem to have engaged the attention of investigators to any extent. Water culture experiments at Rothamsted have so far yielded negative results, and no stimulation has yet been obtained with any plant, with the possible exception of white lupin with sodium arsenite. In a single series a stimulus was suggested, beginning to make itself felt at 1/500,000, rising to an optimum at 1/10,000,000. No stress can be laid on this result, as it is never safe to draw any certain conclusions without several repetitions of the same experiment. With arsenic acid on barley a possible stimulus is sometimes indicated to the eye, the plants being fine and of a particularly healthy dark colour, but this is not corroborated by the dry weights. Additional tests were made with peas and barley, treated with sodium arsenite and arsenate, the dilutions being carried down to 1/250,000,000, but no evidence of stimulus was obtained, so that it hardly seems possible that arsenic can act as a stimulative agent for these two plants when grown in water cultures. It had been thought that the failure to find a stimulation point hitherto might be due to the too great concentration of the toxic substance rather than to the actual inability of the poison to stimulate, but this hypothesis must now be dismissed so far as these plants are concerned.
III. Effect of Arsenic Compounds on Certain of the Lower Plants.
Loew (1883) was sceptical concerning the specific toxicity of arsenic for plant protoplasm. He was convinced that arsenic and arsenious acid were poisonous to algae, not because of their specific character as arsenical compounds, but because of their acid nature, algae being peculiarly sensitive to any acid, and he maintained that these substances were not more poisonous than vinegar or citric acid. He placed various species of Spirogyra in solutions of ·2 gm. potassium arsenate per litre water (1/5000), and found that the algae grew well without making any abnormal growth in a fortnight, showing hardly one dead thread. Some of this alga was then transferred to a 1/1000 solution of potassium arsenate. This suited it excellently and it increased and the appearance under the microscope was very fresh and strong, which was attributed more to the potash than to the arsenic acid. Loew maintained that for the lower animals and for many of the lower plants arsenic in the form of neutral salts is not a poison. When the differentiation of the protoplasm into certain organs reaches a specific degree in the higher plants, then the poisonous action of the arsenic compounds comes into play.
Knop (1884) found that certain unicellular green algae grew luxuriantly in a neutral solution supplied with potassium arsenate. Bouilhac (1894) concerned himself chiefly with the possibility of the replacement of phosphates by arsenates. He recognised that the influence of arsenic is not the same on all species of plants, so he confined his attention to certain of the algae. Stichococcus bacillaris Naegeli was found to live and reproduce itself in a mineral solution containing arsenic acid. Even in the presence of phosphoric acid the arsenic acid favours growth, the best dose being about 1/1000. The arsenic acid is capable of partly replacing phosphoric acid. Other species of algae, Protococcus infusionum, Ulothrix tenerrima, andPhormidium Valderianum invaded the original culture of Stichococcus from the atmosphere, but with no arsenic or phosphoric acid their development was poor. The jars with arsenic compounds were invaded by still more species which grew strongly. Under these conditions it is evident that these algae are capable of assimilating arsenic, and the addition of arsenic acid to a solution free from phosphoric acid is sufficient to enable these algae to live satisfactorily, the arsenates in this case replacing the phosphates. Ono (1900) found that algae are favourably influenced by small doses of poisons, the optimal quantity for algae being lower than that for fungi. Protococcus showed a possible stimulus when grown in concentrations of potassium arsenate varying from ·00002–·0005%. This possible stimulus is interesting in view of the failure to observe stimulation in higher plants by minute traces of arsenic.
The effect of arsenic on fungi is of special interest in that it has a direct bearing upon hygienic and commercial interests. Gosio (1892, 1897, 1901) found that certain of the fungi, Mucor mucedo and Aspergillus glaucum, will grow on various arsenic compounds and exercise a reducing influence on them. These moulds attack all oxygen compounds of arsenic including copper arsenite, and develope arsenical gases. Sulphur compounds of arsenic are not influenced by these fungi. The same moulds would, if cultivated in soil containing arsenic, develope hydrogen arsenide. Penicillium glaucumhas such a strong and definite action on arsenic compounds that he states that there is no doubt of the possibility of poisoning by arsenical gas in a room hung with paper containing arsenic. The compounds are so extraordinarily potent that if a mouse is placed in a vessel in which the mould is strongly developed in the presence of arsenic, it dies in a few seconds.Penicillium brevicaule uses the element in its development as a food substance. If material containing arsenic is placed in contact with dead fungi no reaction occurs. The life activity of the mould is evidently necessary for the reaction by which the arsenic-containing gases are liberated. Csapodi (1894) put forward the earlier results of Gosio and noted that the so-called arsenical fungicides do not only fail to kill the mould fungi but actually favour their development. This action explains why wallpaper containing arsenic is so disadvantageous in a room. Abba (1898) severely tested Gosio’s method of detecting arsenic by means of growths of Penicillium brevicaule, whereby arsenic gases are liberated, vindicating the method completely, and establishing the test as an exceptionally delicate one. Segale (1904) applied the same method to the detection of the presence of arsenic in animal tissues.
Ono (1900) grew Penicillium cultures with solutions of potassium arsenate and found no important differences either of depression or stimulation. Orlowski (1902–3) stated that small doses of arsenic (1/1000–1/100% Sodium arsen—) stimulate the growth of Aspergillus niger, larger doses up to 1/8% retard growth, while 1/6% kills. Spores of the fungus taken from soil containing arsenic are said to possess an immunity against arsenic, in that they germinate in the presence of an arsenic content which rapidly kills control fungi. This immunity is not specific for arsenic, but extends also to other poisons. The chemical composition and water content are not altered.
The toxic effect of arsenic upon higher plants is much more marked with arsenious acid and its compounds than with arsenic acid and its derivatives. No definite evidence of stimulation has yet been obtained with any arsenic compound, however great the dilution at which it is applied. With certain algae a stimulus may occur, and it is possible that arsenic acid is capable of replacing phosphoric acid to some extent under certain conditions. With fungi the toxic effect of great concentrations is marked with certain species, but there are others which are capable of living happily on arsenical compounds and of liberating highly poisonous arsenic gas.