II. Effect of Boron on the Growth of Higher Plants.

1. Toxic effect.

(a) Toxic action of boron compounds in water cultures.

Excessive quantities of boric acid are decidedly poisonous to plants, the action being well marked in water cultures. [Knop (1884)] found that free boric acid was poisonous in neutral food solutions when present at the rate of ·5 gram per litre, but he was not able to detect boron in the ash of the roots of the experimental plants. [Archangeli (1885)] placed seedlings of maize, white lupins, Vicia sativa and Triticum vulgare in solutions of boric acid varying in concentration from 1–·05%, with controls in spring water. In the latter case the development was normal, with 1% boric acid the plants were killed, while it was found that the weaker the solution (within the indicated limits) the stronger the root and shoot growth.

[Hotter (1890)] stated that it was known that 1/20,000 boric acid by weight was harmful to soy beans in nutritive solutions. He experimented with peas and maize, placing the seedlings first in distilled water, later in nutritive solutions. When the peas were nineteen days old they were transferred to nutritive solutions containing 1/1000–1/100,000 boric acid by weight per litre, and within three days the plants with 1/1000 showed signs of injury. Two days later all the plants showed signs of poisoning in that, even with the weakest strengths, the lower leaves were flecked with brown, especially at the edges, while with the greater strengths the lower leaves were dead and the flecking had extended to the upper leaves. In eleven days from the start the plants with 1/1000 boric acid were completely dead, while the other plants showed more or less signs of poisoning. The dry matter and ash decreased steadily with the increase in the boric acid, while the boric acid per 100,000 parts of dry matter increased steadily from 8 to 557 parts. Similar experiments were carried on with potassium borate and with borax; the results showed that, weight for weight, borax is less toxic than potassium borate, which in turn is less toxic than boric acid, while at a strength of 1/100,000 there is little to choose between the three poisons. Similar results were obtained with maize; plants treated with boric acid or potassium borate yielded about 2300 parts boric acid in 100,000 parts dry matter. The general conclusion arrived at by Hotter was that the effect is not so much that of a general poisoning as of a bleaching of parts of the leaf, mere traces of boron being harmless. The cause of injury is local inhibition of assimilation and killing of roots in stronger concentrations. Increase of the strength of boron raises the toxicity until 1/1000 practically inhibits increase in dry substance. The boron was found to be fairly evenly distributed through sound and affected organs.

[Kahlenberg and True (1896)] worked with seedlings of Lupinus albus L., limiting their experiments to those of 15–24 hours in duration. Various combinations of boron and other substances were tested. With boric acid alone 2⁄25 gram molecule per litre killed the plants, with 1⁄25 they were apparently just alive, while 1/100 and less had no injurious effect. Boromannitic acid was possibly more poisonous than the boric acid, while a combination of boric acid and cane sugar proved slightly less toxic. The short duration of these experiments limited their scope considerably, as with certain concentrations the toxic action would not become evident within the prescribed limits of time.

[Agulhon (1910 a)] worked with sterile nutrient solutions, and found that the higher strengths of boric acid hindered growth, 200 mg. boric acid per litre rendering growth impossible. He supported Hotter’s idea that the toxic action affects the roots and the formation of chlorophyll, and he stated that the plants are less green as the dose of boron increases, plants growing in doses of above 10 mg. per litre being yellowish. In other experiments he found that at 100 mg. boric acid per litre life seems impossible for the plant. The roots seem to be more adversely affected by toxic doses than do the shoots. In control plants Agulhon determined the stem/root ratio as 6, with a little boron as 7, while the ratio rose to 13 as the dose of the poison increased to 50–100 mg. boron per litre.

The Rothamsted experiments show that boric acid is definitely poisonous to barley down to a strength of 1/250,000 ([Fig. 15]), the depressing effect frequently being evident at much smaller concentrations, while peas can withstand far more of the poison, the limit of toxicity being about 1/25–1/50 thousand ([Fig. 16]). With the greater strengths of poison the lower leaves of both barley and peas are badly damaged. In barley the leaves turn yellow with big brown spots, giving the leaves a curious, mottled appearance, while with peas the poisoning seems to begin at the tip and edge of the leaves, spreading inwards, without, however, showing the large spots as in barley. So far as chemical tests go at present, it is very probable that boron is deposited in the leaves in the same way as manganese, and that this is the cause of the degeneration. As with manganese, the lower leaves are attacked first, and the trouble spreads upwards, one leaf after another being involved. These observations fit in very well with those of Hotter, and the hypothesis of direct boron poisoning gains support from the fact that in dilutions which produce stimulation of the shoot the leaves show hardly any sign of dying off, even after prolonged growth in the solutions. With barley the effects of boron can be seen in the leaves in concentrations as low as 1/2,500,000, and it may be significant that this is the point at which the depressant action of boric acid entirely ceases in many cases.

Fig. 15. Curve showing the mean value of the dry weights of ten series of barley plants grown in the presence of boric acid and nutrient salts. (May 1st–June 20th, 1911.)

Fig. 16. Photograph showing the action of boric acid on pea plants in the presence of nutrient salts. (Sept. 30th–Dec. 20th, 1912.)

1. Control.
2.1/5,000boricacid.
3.1/10,000„„
4.1/25,000„„
5.1/50,000„„
6.1/100,000„„
7.1/250,000„„
8.1/500,000„„
9.1/1,000,000„„

Tests with white lupins gave no conclusive results, as for some reason it proved very difficult to get satisfactory plants in water cultures. When they are grown under such conditions the roots always tend to get more or less diseased and covered with slime, probably fungal in nature. In the presence of much boric acid the roots remain in a much healthier condition, which suggests that the acid has in this case a strong antiseptic action, and protects the roots. With high concentrations the lower leaves of the plant are badly affected, just as with peas and barley, turning brown and withering at an early date. Various experiments have been made with yellow lupins, but these again are very difficult to grow well in water cultures, as they are apt to drop their leaves for no apparent reason. Generally speaking, the evidence goes to prove that boric acid is toxic down to a concentration of about 500 parts in 25 million. It is difficult to get a true control with which to make comparisons as the plants without boric acid are encumbered with the slime on their roots, which naturally interferes with normal growth, while the plants in the presence of boric acid have the unfair advantage due to the probable antiseptic action of the boron. The effect of the boron poisoning is again evident in the dying off of the lower leaves, which become flaccid and drooping and finally drop off. The lupins grown with boron are very active in the putting forth of lateral roots, so much so that the cortex of the roots is split along the line of emergence of the laterals, which are very numerous and crowded.

(b) Toxic action of boron compounds in sand cultures.

[Agulhon (1910 a)] moistened 2 kgm. pure sand with 500 c.c. nutritive solution for each pot, and boron was added at the rate of 0, 0·1, 1, 10, and 50 mg. boric acid per litre of nutritive solution. Twenty wheat seeds were sown in each pot, and after twelve days the healthy plants in the first four pots were 6–8 cm. high, but those with the maximum amount of boron showed yellowish leaves only 3 cm. long. After three months’ growth the plants were harvested, when those with most boron were found to have died after making about 10 cm. growth. The toxic doses in sand proved to be weaker than those in water cultures, probably because evaporation from the surface of the sand caused concentration of the poisonous liquid.

(c) Toxic action of boron compounds in soil experiments.

Long before any experimental work was done with boron in water cultures, the poisonous properties of the substance were recognised with regard to plants growing in soil. [Peligot (1876)] grew haricots in porous earthenware pots, the plants being watered by rain and by solutions, each containing about 2 grams per litre of such substances as borax, borate of potassium, and boric acid, other pots receiving various fertilisers, as potassium nitrate, sodium nitrate, &c. This quantity of boron completely killed off the plants receiving it, whether it was applied as free or combined boric acid, while the fertilised plants completed their development well. On this account the deleterious action was attributed to the boric acid and not to the sodium or potassium base supplied. Peligot hinted at the improbability of a substance like boron, which is so poisonous to plants, being really innocuous to human beings when it is used as a preservative for foods.

[Nakamura (1903)] also found that borax is harmful in pot cultures if present in large quantities, 50 mg. borax per kgm. of soil exerting a very injurious influence, while even 10 mg. per kgm. did some damage. [Agulhon (1910 c)] found that the toxic doses of boric acid in soil cultures approached those in nutritive solutions rather than in sand cultures, a phenomenon that he attributed to the fact that the boric acid was fixed by the soil, probably as insoluble borate of calcium, so that the surface concentration obtained with sand cultures was avoided. He found that the ash of plants grown with excess of boron contained more than the normal amount of boron, while the weight of ash per 100 dry matter was also increased. He concluded that the plant thus suffers an over-mineralisation and in consequence an augmentation of its hold on water, so that the fresh weight of the plant may indicate a more favourable action of the boric acid than does the dry weight. Other investigators ([Fliche and Grandeau 1874]) had found the same increase in the proportion of ash in chestnut trees grown on too calcareous soil, so Agulhon concluded that one is here dealing with a general reaction of plants to an excess of a useful element.

Other experiments were carried on in the open field, maize being grown on control plots and on plots receiving 2 gm. boron per square metre. At first the latter plants were behind, the dose being too strong. Eventually, however, they pulled up level and the dry weights from the two plots proved to be nearly the same, the fresh weights being identical. Maize is evidently far less sensitive to boron poisoning than are peas and oats, for with these one-half the original amount of boron (= 1 gm. per sq. metre) proved toxic.

Interesting results were obtained ([Agulhon 1910 a]) by repeated experiments with the same soil containing boron. It was found that sand or soil containing a proportion of boron which is lethal or toxic to a first culture will allow much better growth with a second and subsequent crops. Repeated experiments on the same soil may show the change from a lethal dose to a toxic one, thence to an indifferent and finally to an optimum concentration. Furthermore ([Agulhon 1910 b]) the very plants may accustom themselves to greater quantities of boron, the increased power of resistance being transmitted. He concluded from his experiments that the progeny of the second generation of maize were able to withstand quantities of boron that were toxic to control plants[13]. Agulhon once again emphasised the fact that for toxic doses of boron the first symptom is the more or less marked disappearance of chlorophyll, though the aerial parts are not affected so soon as the roots.

2. Effect of boron compounds on germination.

One of the first indications that boron compounds affect the germination of seeds was given by [Heckel (1875)] who found that germination was retarded for 1–3 days by weak solutions of borates (·25 gm. to 20 gm. water), and was stopped altogether by stronger solutions (·60 gm. to 20 gm. water). [Archangeli (1885)] tested the germination of a variety of seeds of Leguminosae, Gramineae, and of Cannabis, Iberis, Raphanus, Collinsia, and Linum in the presence of boric acid. The seeds were placed in bowls with solutions of ·25, ·5, and 1% boric acid at temperatures ranging from 16°–23° C. The bowls were covered with glass plates to prevent evaporation and consequent increase of concentration, controls in spring water being dealt with under similar conditions. 1% boric acid was found to check germination altogether, and the weaker the concentration the less was the process hindered. [Morel] soaked seeds of haricots and wheat in various solutions of boric acid, and found that germination was generally hindered or inhibited. The deleterious action diminishes as the strength of the solution or the time of contact diminishes, but solutions of the same concentration do not act equally on all seeds. Boric acid and borax proved to be similar in their action qualitatively.

The deleterious effect of strong doses of boric acid on germination was confirmed by [Agulhon (1910 a)], the higher quantities (above 10 mg. boric acid per litre) retarding germination of wheat.

3. Does boron stimulate higher plants?

Of recent years a few investigators have thrown out hints as to the stimulant action exerted by boron compounds on plants. [Roxas] indicated that M/100,000 (M = molecular weight) of boric acid exercised a favourable action on rice. [Nakamura (1903)] tested the point by means of pot cultures. Peas and spinach plants were grown in soil which received 1 and 5 mg. borax per kgm. With peas the 1 mg. exerted evident stimulant action, as determined by the increase in height of the shoot over that of the control, 5 mg. seeming to be slightly depressant in action. With spinach a stimulation was observed both in weight and height with a dose of 5 mg. borax per kgm.

Average weight Average length of leaves
5 mg. borax 10·3538·2
Control7·234·0

Agulhon ([1910 c] and [d]) took the matter up still more definitely and made many tests of various kinds, in water, sand and pot cultures.

(a) Water cultures.

His water cultures were made under sterile conditions, the seeds when possible being sterilised with corrosive sublimate, the germinating apparatus being also sterilised. With wheat a stimulant action was evident, maximum growth being obtained with between 2·5 and 10 mg. boric acid per litre, though the dry weight increase did not quite keep pace with that of the fresh weight, a fact to which previous reference has been made. The chief improvement is in the root, the stem/root ratio falling to 5, as against 6 in the control series. Visual observation indicated that the roots of plants receiving 5–10 mg. boric acid per litre are longer than the others, though they are less rich in adventitious roots. The increased dry weight due to boron may amount to as much as 30%.

(b) Sand cultures.

Agulhon again observed stimulation in this case. 2 kgm. of sand were moistened with 500 c.c. nutritive solution, varying quantities of boric acid being added in addition. ·1 mg. boric acid per litre of N.S. (·05 mg. per pot) gave an increase of 25% fresh weight, and 7·5% dry weight. The stimulating doses seem to be weaker than in the experiments with liquid media, probably because the evaporation from the sand increases the concentration of the boric acid at the surface. It was also noticed that the increase of weight varied in experiments made at different times. With oats the stimulating influence is greater than with wheat, showing that some plants are more sensitive than others to the influence of boron. With radish 1 mg. boric acid per litre exercised a stimulating effect, the enormous average increase of 61% in fresh weight occurring with this strength, though this only represented an average increase of 9·6% dry weight.

(c) Soil cultures.

Here again the stimulating action was evident with higher concentrations than in sand cultures, and Agulhon obtained good results with strengths that are toxic in sand. The evaporation from earth is not so rapid as from sand, so that the concentration is not increased, and also some of the boric acid is withdrawn from the solution by interaction with the soil, so that the stimulating concentration rises in the scale.

In field experiments Agulhon found that peas were more sensitive to the toxic action of boric acid than is maize. A strength of boric acid (= 1 gm. B per sq. metre) that poisoned peas, gave an increase of 61% fresh weight and 39% dry weight with maize; half the strength proved to be indifferent for peas, the improvement with maize equalling 56% increase fresh and 50% increase dry. Curiously enough, judging by appearances in the first experiment, an unfavourable influence was at work, though in reality a great stimulation was being caused. Colza gave a good increase with similar strengths, but with turnips 1 gm. B per sq. metre only favoured the aerial parts, while ·5 gm. B per sq. metre only increased root development. Agulhon concluded that it is as yet impossible to determine with any precision the exact part that boron plays in the plant economy. He suggests that boron is a “particulier” element characteristic of a certain group of individuals or of life under particular conditions. In his summary he argues that each series of individuals adapted to different environments has doubtless need of particular elements, and that perhaps chemical causes and morphological differences are very closely connected. Boron may be of this “particulier élément” type in the higher plants of the vegetable kingdom, and it may be useful commercially as a manurial agent, the “catalytic manure” of Bertrand and Agulhon.

While the higher concentrations of boric acid proved definitely toxic to both peas and barley in the Rothamsted water cultures, some evidence of stimulation was obtained with the lower strengths. With barley the question of stimulation is still an open one, as below the toxic limit growth seems fairly level in most of the experimental series. The lower limit of toxicity varies from 40–4 parts boric acid per 10,000,000 according to circumstances. Below this critical concentration the boric acid has apparently no action, either depressant or stimulant, unless the stimulation should prove to begin at a dilution of 1/50,000,000, but the evidence on this point is not sufficiently well marked or consistent to be conclusive. This failure to detect stimulation was somewhat unexpected, as when judged by the eye the plants treated with the lower concentrations of boric acid seemed better than the controls, and also exhibited a particularly healthy green colouration.

Peas on the other hand are definitely stimulated with traces of boric acid, concentrations of 1/100,000 and less causing an improvement in growth, while under some experimental conditions even higher amounts of boric acid were beneficial. All the stimulated plants showed the characteristic dark green colour which seems to be associated with the presence of minute traces of boron in the nutritive solution. An interesting morphological feature was the strong development of small side shoots from the base of the plants in the presence of medium amounts of boric acid, from 1 part in 100,000 downwards. This gave rise to a certain bushiness of growth, which was less evident as the concentration of the stimulant decreased. The general outcome of the tests seems to be that boric acid needs to be supplied in relatively great strength to be fatal to pea plants, and that the toxic action gives place to a stimulative one high up in the scale of concentration. As far as experiments have already gone it seems as though the stimulation is not a progressive one, as the effect of 1/100,000 boric acid is as good as that of 1/20,000,000, a flat curve connecting the two. This, however, needs confirmation.

Yellow lupins also give some evidence of stimulation with concentrations of about 1/50,000 boric acid, the improvement being far more strongly marked in some sets of experiments than in others.