The Physiology of the Ascent of Sap

For experiments lasting a few hours the rate of the complete revolution of the drum is adjusted to once in an hour, the length of the recording surface being 300 mm. Successive marks when recorded by drops falling at intervals of, say, six seconds, are a little over i mm. apart. It is more convenient to have the successive dots marked for every three drops, in which case the dots are 3 mm. apart ; the adjustment of the counterpoise on the Tilter allows this to be done with great precision. The weight of the three accumulated drops causes a sudden tilt of the lever by which the water is completely emptied into the vessel v, after which the Tilter assumes its normal horizontal position. The following precautions should be observed for accurate work : the plant . should be so placed that the drops fall

near the fulcrum of the lever, to avoid upset of the lever by the momentum of the falhng drop. Another pre- caution is to prevent the adhesion of any remnant of the exuded drop to the spoon-shaped receptacle : the main- tenance of a very clean and even surface removes this source of error. In long-continued experiments, say, for the determin- ation of the diurnal periodicity, the Electromagnetic Writer is arranged to subside vertically through lo cm., which is the height of the drum, in the course of twenty-four hours. The record is in the form of a spiral, its total length being 720 cm., too long for reproduction in a book. To obviate this difficulty, the drum is adjusted to a speed of revolution of once in twenty-four hours. The period of successive tilting of the lever is also appropriately modified by moving the sliding counterpoise, so that successive electric con- tacts are made on the exudation of every 5 to 10 c.c. of sap. The Tilter empties the exuded Hquid into the vessel placed underneath ; this serves as an independent check for the total quantity of exudation during twenty- four hours.

I have carried out experiments on exudation with numerous plants ; those with Cucurhita and Zea Mays may be regarded as typical. In Cucurhita the root-system is very extended and, generally speaking, buried deep in the soil : in Zea Mays, on the other hand, the root is nearer the surface and does not cover a large area. The sap exuded contains inorganic and organic sub- stances in solution. I shall give later the composition of the exuded sap of Palm trees, which contains a large quantity of sugar. The following is the result of the analysis of the exudate from Cucurhita and Zea Mays.

The normal rate of exudation was found to remain constant under uniform external conditions, lor the record obtained with a root-stock cut close to the ground shows that the exudation continued practically unifonn from hour to hour, and this for several days. Allowance must be made, however, for the general physiological depression caused by the decapitation of the plant. But this change is continuous and not marked by any diurnal variation. This will be seen from the reproductions of portions of a continuous record taken for thirty-six hours ; the portions of the record are for four hours at intervals of twelve hours, i.e. from 9 a.m. to i p.m., from 9 p.m. to i a.m., and from 9 a.m. to I P.M. from February 11 to 13 (fig. 43). The intervals between successive dots in each row are practically the same ; but the gradual slowing down of the rate is seen in counting the number of dots at the beginning, the middle and the end of the rows, in which each dot repre- sents I c.c. of exuded sap. It will be noted that on the first day the exudation for four hours was 43 c.c. ; after an interval of twelve hours it had fallen to 30 c.c. ; and after

a further period of twelve hours to 21 c.c. ; the total for these twelve hours was thus 94 c.c. The sap collected for twenty-four hours was 180 c.c, which is practically double the quantity recorded for twelve hours. The notable fact is that the decline was continuous, there being no variation due to the alternation of day and night. The reason for this is to be found in the uniform conditions in which the deep-seated root-system was maintained. It was protected from light, and there was no great range in the variation of temperature of the subsoil in February. The external variations, moreover, did not affect the root in the soil.

Having thus found that exudation remains constant under uniform external conditions, we can proceed to study the effects of physiological variations on the rate. Having seen how the cellular activity underlying the ascent of sap is depressed by drought, we may expect that exudation will be found to be similarly affected. Thus, in a particular experiment with Cucurbita, the root-stock under drought did not exhibit any exudation ; but applica- tion of water near the root-stock induced it, though in a spasmodic manner. On digging up the plant it was found that the water had only reached a limited portion of the

root, most of the root-system being still in dry soil ; the spasmodic exudation was thus due to the irrigation of a restricted portion of the root. After extensive irrigation in a second experiment, the arrested exudation was found to be renewed, and this at a very uniform rate. The following experiments show the effect of mechanical and electric stimulation on the rate of exudation. Mechanical Stimulus. — Experiments were carried out with a field-specimen of Cncurhita to ascertain the effect of increasing intensity of stimulus on the rate of exudation. In the first experiment a small lateral root was dug up, taking care that it remained coated with moist clay. A portion of this root was then cut off with a pair of scissors ; the effect of the moderate stimulus of this cut was to reduce the rate from the normal 52 c.mm. to 42 c.mm. per minute. The normal rate was restored after seventeen minutes. A larger lateral root was next cut off, which caused a more intense stimulation, and induced a depression of the rate of exudation from 52 c.mm. to 7 c.mm. per minute, or about one-eighth the normal ; the period of recovery was now found prolonged to 50 minutes. Pricks were then ad- ministered to the main root, inducing a diminution of the rate to 4 c.mm. per minute, or to one-thirteenth ; the period of recovery from this intense stimulation was prolonged to three hours. Thus an increase in the intensity of the stimulus gave rise to an increased depression in the rate of exudation, and a corresponding prolongation of the period of recovery. In the first two experiments described above, the diminished rate of exudation could not have been due to the loss of a portion of the root, which was quite negligible compared to the very extensive root- system ; the subsequent recovery to the normal rate shows, moreover, that the induced diminution was undoubtedly due to the retarding action of stimulus.

Electric Stimulus. — After the normal rate of exudation of another root-stock had been recorded, two mctalHc prongs were buried in the soil on opposite sides of the plant to act as electrodes, and the root was stimulated by the passage of an induction-shock of moderate intensity Fig. 44. The Effect of Meclianical and Electrical Stimulus in Retardation of Exudation M, the effect of mechanical, e, that of electrical, stimulus. applied for a minute. The normal rate of exudation of this specimen was 80 c.mm. per minute. After the passage of the shock the rate of exudation was greatly depressed, the rate being now 6 c.mm. per minute ; complete recovery was only attained after several hours. I give two records in fig. 44, in which m represents the effect of mechanical, and E that of electrical, stimulus on the rate of exudation.

Table XX. — Showing the Immediate Effect of Stimulus on Exudation and Subsequent Recovery The successive dots are relatively close at the beginning ; but after the application of the stimulus the intervals become widened, showing a depression of the rate of exudation. The slow recovery is seen in the gradual approximation of the successive dots to each other. The rest of the record is not long enough to exhibit the complete recovery. In Table XX. (p. 139) are given the quantitative results of a different experiment, showing the immediate effect of stimulus in retardation of exudation and the gradual recovery towards normal.

The effect of a poisonous solution of formaldehyde was next observed. The normal rate of exudation of the particular specimen of Cucurhita was 28 c.mm. per minute. Application of the poisonous solution caused a marked depression, the rate of exudation being now reduced to 0'35 c.mm. per minute. Continued action of the poison produced complete arrest. Wieler found that exudation ceased when the roots of seedlings, or of older plants from water-culture, were placed in a dilute solution of chloroform. Pfeffer's criticism of these results is ' that the experiments are not always conclusive, since if the chloroform is too strong, the plant is readily injured, or may be killed.'

In order to meet this objection, a definite ph3'siological test was employed which is free from the uncertainty arising from the possibility of a fatal effect caused by the anaesthetic. We found that, in the case of the stem, the preliminary action of chloroform was to induce an enhance- ment of the rate of ascent ; this being followed, under con- tinued action, by a depression or abohtion of the ascent. In regard to exudation, results have been obtained in every way similar to the above. Thus, with a given root-stock of Cucurhita, the normal rate was 52 c.mm. per minute. The preliminary effect of the application of chloroform was to enhance it to 68 c.mm. ; after this, the depressing effect set in, the maximum depression reducing the rate

of exudation to 36 c.mm. per minute. Owing to the dissipa- tion of the chloroform-vapour in the soil, the plant exhibited a recovery after two hours. I then applied chloroform for a second time, when the phenomenon of accommodation, or acquired immunity, was shown in a very interesting manner ; for the dose had to be increased to obtain the previous effects. According to the generally accepted theory, the exudation from the cut end of the stock is due to filtration under pressure, the active force being the root-pressure set up by some specific activity of the cells in the root.

There is no ground for the assumption that the activity of the root-cells is in any way specifically different from that of the cells of the stem. We have found that the effects of variation of temperature, of anaesthetics, and of poisons are the same in the one case as in the other. The active pressure which causes the expulsion of sap from the cut surface of a stem is not generated by the root alone ; the stem also contributes. Thus stems of Grasses, with their cut ends placed in moist sand, exhibit exudation under pressure, just as do specimens with roots. This propulsive force therefore exists not only in the root but also in every portion of the stem. It is moreover not strictly true that the sap is forced through a passive layer of tissue at the cut end by filtration under pressure exerted by the distant root. The root and every section of the stem exerts pressure ; the total pressure is the sum of their additive effects.

Effect of Local Application of Chloroform. — The following experiment will show that the terminal layer also takes an active part in the outflow of sap from the cut end. We have seen that the application of dilute chloroform to the root caused a transient enhancement of exudation. Taking another root-stock of Cucurbita, 1 applied dilute chloroform to the cut end itself. The specimen was young and its rate of exudation was one drop in 220 seconds. On the application of chloroform, the rate of exudation was con- tinuously increased till after twenty-five minutes the rate had become one drop in fifty seconds, that is to say, more than four times as rapid. The plant recovered its normal rate in the course of an hour.

Effect of Variation of Temperature. — It has been shown that the rate of ascent of sap is increased when the root or the cut end of the stem is raised in temperature (p. 58). Exudation is likewise enhanced by raising the temperature of the stem-portion of a root-stock. In the case of a root-stock of Cucurhita cut close to the ground, there was, we saw, no change in the rate of exudation though the external temperature underwent a diurnal variation. This was because the roots were buried in the soil, and moderate variation of the atmospheric temper- ature did not affect the temperature of the root. When, instead of cutting the stem close to the ground, a short length was allowed to remain above ground, the exudation was then found to undergo an increase with a rise of temper- ature of the outside air. The temperature outside was at its maximum at 2 p.m., and the exudation from the cut surface of the stock was also found to attain its maximum rate at that hour. This will be studied in greater detail in the next chapter, where it will be shown that, under conditions described above, the pressure exerted by the sap also undergoes a parallel increase. In other words, exudation and root-pressure are brought about not merely by the activity of the root, but also by that of the shoot.

In conclusion, the expulsion of sap by living cells may be generally considered. Electric stimulus has been shown to induce a diminution of the rate of ascent of sap in the stem, and a diminution of the rate of exudation from the root-stock. It is possible to arrive at a definite explanation of these effects by reference to the responsive action common to all living cells, rhythmic or ordinary. This identity is made evident by a comparison of the curves of mechanical response and recovery of Mimosa leaf, and of responsive diminution of exudation and subsequent recovery in the root-stock of Cucurhita. In the former, stimulus causes the physiological change in the cells which we call contraction, resulting in a diminution of turgor and the fall of the leaf ; gradual recovery of the cells restores the normal turgor and the expanded position of the leaf. A similar physiological change, in response to stimulus,

Fig. 45. Curves showing Similarity of Response to the Action of Stimulus and Recovery of Mimosa (left figure) to that of Exudation in Ciicurbita (right-hand figure) must undoubtedly occur in the active cells that propel the sap ; which, affecting, as it does, the conducting channels, causes a concomitant diminution of the rate of flow. The identical character of the two responses will be seen in the records given (fig. 45) of the response of the leaf of Mimosa and that of the root-stock of Cucurhita, in which the ordinate represents the diminution (negative variation) of turgor and of exudation respectively. In both, the height of response is increased with the increasing intensity of stimulus, with corresponding prolongation of the period of recovery.

of the pulvinus of Mimosa, on stimulation, is an essential part of its motile mechanism, and this applies also to the pulvinule of the leaflet of Desmodium in its ' spon- taneous ' oscillation. In the preceding pages, evidence has been accumulated which demonstrates that the active expulsion of sap by living cells is an essential part not only of the mechanisms of movement, but also of the mechanisms for the distribution of liquid throughout the plant. Experi- ment has shown that the ascent of sap in the stem, and the excretion (transpiration) of water by the leaves, are mani- festations of this cellular activity ; and now exudation from the cut surface of the root-stock has been shown to be traceable to the same cause. In herbaceous plants, where the wood-vascular tissue is but slightly developed, exudation takes place mainly from the cortex of the cut surface ; whereas in trees, ' bleeding ' comes from the vascular tissue which has become charged with liquid by the pumping activity of the cortex.

What may be precisely the mechanism of the process of expulsion in the individual cell is not yet clear. It may be (i) an active contraction of the lining layer of protoplasm ; or (2) an increased permeability of this layer, permitting the escape of cell-sap under the elastic pressure of the stretched cell-wall ; or probably the co-operation of both these factors, and perhaps others. In any case it is a manifestation of that contractility which is one of the fundamental properties of living protoplasm.

The exudation from the cut end of the root-stock is depressed or arrested by drought, and renewed after irrigation. Mechanical or electric stimulus induces a retardation or arrest of exudation ; this is followed, on the cessation of the stimulus, by recovery, which becomes protracted if the stimulus has been strong. Dilute chloroform applied at the cut surface enhances the rate of exudation, showing that the terminal layer at the surface also takes an active part in the exudation.

The resultant exudation is thus due to the co-ordinated action of active cells throughout the whole length of the root-stock. The responsive action of the cells concerned in exuda- tion is fundamentally similar to the expulsion of water by the cells in the pulvinus of Mimosa when stimulated. General considerations — Diurnal periodicity of root-pressure — The recording apparatus — Relation between temperature and pressure — Diurnal variation of pressure in deciduous trees — Diurnal variation of exudation — The effect of light — Summary.

The mechanism of the propulsion of sap in a plant has been compared with the action of a pump, and the analogy holds good even in many details. Let us imagine a tubular well supplied with water from the deep soil ; the water is raised by a pump, the activity of which may be gauged in two different ways, dynamic and static. The activity of the pump may thus be found from the rate of the outflow, or from the height of the balancing column of water. In the corresponding phenomenon in plants, the activity of the root-stock may be measured, either by the rate of exudation, or by the root-pressure, i.e., the column of liquid which that pressure can sustain. As the exudation and the root-pressure are different expressions of an identical cellular activity, a relation might be expected to exist between the two, under similar conditions, i.e. :

(i) Plants with high root-pressure should also exhibit a high rate of exudation, and vice versa. in exudation. The above conclusions would follow as necessary conse- quences of the theory of cellular activity in the ascent of sap. In practice, however, we are confronted with numerous anomalies, which are given below in order of increasing complexity and difficulty of explanation. {a) Apparent independence of exudation and pressure. — It is often found that a plant with low root-pressure exudes a large quantity of sap ; conversely, other plants with high root-pressure exhibit feeble exudation.

(6) Irregular distribution of pressure. — Manometers attached to the tree at different heights should exhibit a decrease of pressure from below upwards ; but this is seldom the case. (c) Eccentricities in the diurnal variation of exudation. — Exudation from plants often exhibits an erratic diurnal variation. Thus while some may exhibit a minimum exudation of sap in the forenoon and a maximum exudation in the afternoon, other plants exhibit precisely the converse results. These eccentricities are so inexphcable that, according to Jost, ' there can be no hesitation in concluding that we are still far from having reached a satisfactory explanation of the phenomenon.' ^

{d) Exudation in Palms. — The widest divergence in the relation between pressure and exudation presents itself in certain Palms. In many deciduous trees, in the absence of transpiration from the leaves, there is a con- siderable intra-vascular pressure ; so that when a hole is drilled in the trunk, a copious exudation of sap under pressure follows. In Palms, however, the phenomenon is very different. The Palmyra Palm [Borassus fiahellifer) , which attains a height of lOO feet (30 metres), grows very slowly and is said to live for a couple of centuries. Under appropriate conditions, as described in Chap. XIII., the exudation of sap from it is very copious. Disregarding the great resistance offered by the tissue to the ascent of sap, a pressure of three atmospheres would be necessary to raise water to this height. But Palms do not exhibit any root-pressure ; this has been found to be the case by Molisch in Arenga saccharifera, and I find it to be equally

the fact in Phccnix sylvestris. The driUing of auger-holes in the trunk is not followed by any exudation, though the outflow of sap is copious after the surface of the trunk has been subjected to repeated injury. Here we have an apparently inexplicable phenomenon of exudation without any pressure to enforce it. In approaching this complex problem, we must realise that the ascent of sap and its diverse manifestations are due to the activity not of any single part of the tree, but of all its parts. The different regions, the root, the shoot, and the transpiring leaves, are subjected, as already stated, to external variations which affect them unequally, and the resultant effect is, to a great extent, due to the algebraical summation of the partial effects. The physiological activity is modified by the state of turgor of the tree, and this is determined by the relative gain or loss of water. As regards the gain, the organ of absorption, the root, is completely shielded from light, and to a great extent from the diurnal variation of temperature. The conducting stem is, on the other hand, subjected to physiological changes in the diurnal variation of temperature, and the alternation of light and darkness. In the case of herbaceous plants, light acts as a stimulus on the cortical tissue concerned in the transport of sap, and lowers the power of conduction. But in the case of trees, the thick bark is impervious to light ; hence the incidence of sunlight on the trunk would raise the temperature and enhance the velocity of the ascent.

Turning next to the negative factor — the loss of water by transpiration — we have diverse influences which, by their physical and physiological actions, enhance or depress the rate of loss. Among these may be mentioned the effects of diurnal variation of temperature and of light, the action of wind, the varying hygrometric condition of the air, and so on. It will thus be seen that the internal pressure and the exudation, which depend on the relative gain or loss of

water by the plant, are modified by numerous factors, some of which are concordant and others in conflict ; these vary in different degrees according to the changing external conditions. It is therefore not at all surprising that the observed results should have appeared to be so capricious. It may, however, be possible to unravel the complexities by the process of isolation. We shall therefore take up in this chapter the question of the relation between pres- sure and exudation in a root-stock which bears no side- branch with transpiring leaves to complicate the phenomena.

In discussing the relation between root-pressure and exudation, especially the case of high root-pressure with minimum exudation and vice versa, we have to take full account of two factors, namely, the resistance offered by the tissue and the area of supply of water. In the action of a pump in a tubular well, it is evident that the outflow will be diminished when the pipe is choked with sand ; again, too quick an outflow from the pump may dry up the well unless the subterraneous supply of water is adequate. Similarly, in the exudation of sap, tissues of different plants will offer unequal resistance to the flow of water. The modifying influence of the area of the absorbing root is shown in experiments with two specimens of Cucurbita, one grown in a pot, and the other under field-conditions. In the former, though the exudation was at first moderate, it slowed down on account of the limited area of its ab- sorbing root-system. The Cucurbita grown under field- conditions gave, on the other hand, a copious exudation which remained practically constant day after day. From the facts described above it will be seen that a fair com- parison between pressure and exudation can only be made when the two factors of resistance and source of supply remain constant, as in experiments carried out with one and the same plant. When this condition is fulfilled, the relation between exudation and pressure will be found to be very definite, as will be seen in the experiments detailed in the next chapter.

As a gauge of cellular activity, the statical method of balancing root-pressure by a column of water or mercury is perhaps more satisfactory than the rate of exudation ; for in the former the variable factors of conductivity and the area of supply do not enter into the question. They may prolong the period for the attainment of balance, but do not modify the balancing pressure itself. The explanation of the irregular variation of pressure will be dealt with in the next chapter.