An automatic system to study the responses of ... - Springer Link

AN AUTOMATIC SYSTEM MERGED MACROPHYTES F. Hugh DAWSON, Freshwater

Biological

TO STUDY THE RESPONSES OF RESPIRATION TO ENVIRONMENTAL VARIABLES

Derek F. WESTLAKE Association,

& Gordon

River Laboratory,

AND PHOTOSYNTHESIS

BY SUB-

I. WILLIAMS Wareham,

Dorset, BHzo 6BB, England

Received February 28, 1980 Keywords: metabolic chamber, submerged aquatic macrophytes, Ranunculus,

Abstract An apparatus to measure the rates of respiration and photosynthesis of aquatic plants in water at velocities of up to zoo.mm SC’ in a closed water-flow system with partial recirculation, is described. The temperature, the light regime and the concentration of dissolved oxygen are controlled automatically. Typical results are given for Ranunculus penicillatus var. calcareus which were repeatable between the same season in different years and compared with published data.

Introduction Studies of the metabolic activities of submerged macrophytes of flowing waters, in relation to their behaviour in the field, require the use of water velocities similar to those typical of rivers; such velocities were not attained in earlier experiments (Gessner & Pannier, 1958; Owens & Maris, 1964; Westlake, 1967; McDonnell & Weeter, 1971; Glanzer, 1974). The apparatus was essentially similar to that described by Westlake (1967) but adapted formuch higher water velocities around the plant, together with automatic selection and improved control of the water temperature, dissolved oxygen concentration and light regime. The higher velocities used meant that changes in oxygen concentration would not be detectable in a through-flow system, so most of the water was recirculated. The oxygen concentration of the circulating water was maintained at a preset level by the exchange of some of the circulating water with water of known concentration, higher for respiration experiments and lower for photosynthesis. The rates of the metabolic oxygen changes were determined from the differences in oxygen concentration between the added water and the circulating water, and the volumes of water that were exchanged to maintain the internal concentration. The ranges of conditions used cover the prevailing conditions of the river which were also being mon-

respiration, photosynthesis

itored and the data obtained were to be incorporated temporal growth model for the plant (to be published where).

in a else-

The Apparatus

The experimental chamber was a shallow oblong clear Perspex box, (310 x 170 x 31 mm internally; Fig. I, MC). The inlet and outlet were shaped to given ‘piston’ or ‘plug’ water flow through the chamber. The plant was held spread out by nylon nets (I 3 mm mesh) at IO mm from the upper and lower faces of the chamber; this reduced selfshading and helped to distribute the flow evenly. The inside of the lid was slightly domed and vented to aid in the removal of air bubbles if necessary. The internal water temperature was monitored with a thermistor probe. Water was circulated by a centrifugal pump (P, Stuart Turner No. 18) which could give velocities of up to 200 mm s-’ in the metabolic chamber. The water flowed past a dissolved oxygen probe (02), housed in a Perspex case, through the experimental chamber (EC) to a mixing vessel before returning through a heat exchanger (H3) of stainless steel tube. The experimental chamber was submerged under 100 mm of water in a constant temperature water bath (WB) together with the mixing vessel and the heat exchangers. The temperature of the stirred water bath was maintained by a heater, controlled by another thermistor and a switching circuit (Fig. za), which worked in opposition to a continuous cooling system. The bath itself was thermally insulated inside a light-proof box. The cooling system circulated water at 3°C from a refrigerated storage tank through a long heat exchanger coil in the water bath. Either of two rates of flow could be selected by solenoid valves. The water in the circulating system was pumped from a side channel of the R. Frome at East Stoke. This water was typical of chalk streams, being very hard (2.7-4. I meq I-’ total alkalinity) and relatively rich in most plant nu277

Hydrobiologia 77. 277-285 (1981). ooIa-a1ja/at/o773-ozjy$or.ao. o Dr. W. Junk b.v. Publishers, The Hague. Printed in the Netherlands.

constantWater Head

CT

GL

- ---

I

_--_

H3 ,-- _________-_ _.- -_--- -

Fig. I. A diagramatic representation in vertical section of the metabolic chamber and water supply and circulation system. (CH constant head; FI and Fz filters; Heat exchangecoils HI, H2 and H3; water conditioning tube CT, with gas-lift counter-current circulation GL, and oxygen probe 01; the illumination system L; metabolic chamber MC, in vertical section and below, in surface view; recirculation pump P and control oxygen probe 02 and release valve V, mixing vessel MV and insulated water bath WB,

including area within dashed lines). trients (1-4 mg I-’ nitrate-nitrogen, 0.02-0.16 mg 1-l phosphate-phosphorus and 1-3 mg 1-l potassium). The pH was generally between 7.5-8.5 which gave c. 200 mg 1-l HCOsand c. I-IO mg 1-r free CO*. The water was filtered three times through filters of progressively decreasing pore size; firstly through a coarse glass-filter, before reaching the constant head device (CH, Fig. I), secondly through a prefilter (FI, Geldman 12533 submicron Acroflow cartridge) and finally through a tine filter (F2, Gelman 12571 0.45 pm pore size). The filtered river water was conditioned before use by bubbling either oxygen or nitrogen into the water contained in a tall narrow thermally-insulated tube (CT, I .7 x o. I m). (Other gas mixtures could be used to increase the sensitivity of the apparatus in special studies e.g. 50 : 50 278

oxygen/nitrogen for respiration or IO : 90 for photosynthesis). Carbon dioxide at the partial pressure of normal air (0.03%) was included in both gases to maintain the pH of the water at air equilibrium. The water in the tube was brought to the water bath temperature bypassing through a heat exchanger in the inlet (HI) and a counter-current circulation through another heat exchanger (Hz), which was induced by a gas lift pump effect (GL) in a tube adjacent to the conditioning tube. Both the dissolved oxygen concentration and the temperature of this conditioned supply water were recorded; the oxygen sensor housing was modified using a concentric ring device to increase the water velocity past the membrane surface (Dawson & Henville, in prep.). The variables,irradiance, watertemperature and oxygen

concentration in the chamber, and oxygen concentration in the conditioned water could each be held at various levels. Any combination of sixteen possible internal levels could be set up to be selected in any repeatable sequence over a 48 hr period, together with the appropriate gas equilibrium in the conditioned water, each combination lasting for 4 hr. Each internal variable had a matrix board with columns for six levels and rows for the twelve times. These matrix boards were constructed from brass strips mounted horizontally and vertically and separated by insulating plastic. Each selection column was connected to a pre-set circuit which could be put into electrical contact with a timing row, by inserting a screw. This either directly switched a circuit by a low voltage relay (e.g. for the lighting system) or allowed a control circuit to operate at the pre-determined balance point (e.g. oxygen control system). The timing rows of each matrix board were connected to a uniselector switch (6 bank 12 way) which was advanced automatically by four hourly pulses from a time switch. The lights were 0.61 m (2 foot) 40 W fluorescent tubes (Osram ‘North light’) arranged in two staggered banks, each with eight tubes 75 mm apart (L). An alternative and simpler system using up to four Altas colour-arc MBIF bulbs (400 watt) supplemented by 12 (60 watt) tungsten bulbs and filtered using a sheet of cinamoid ‘steel blue’ (Strand Electric and Engineering Co Ltd, London) to simulate natural daylight, has subsequently been used successfully. The lid of the light box was lined with reflective foil and fitted with an extractor fan(F) which operated when the lights were on. Four levels of irradiance were available, which could be preset, from zero rising to c. 44 J m-* s-’ (c. 10,000 lux) as the maximum down-welling photosynthetically available irradiance. The up-welling irradiance was enhanced by another reflective sheet: 1, hich increased the total PAR by c. 0.5. When a level column was activated by a timing row in the selection matrix an appropriate dimmer circuit was switched in. When zero irradiance was selected, oxygen was also selected as the conditioning gas; otherwise nitrogen was selected. Temperature was controlled by putting the bath thermistor as one arm of a Wheatstone bridge circuit. Another arm was connected to a set of five variable resistors, corresponding to a series of temperatures (normally intervals of 5°C between 5”and 3o”C), each of which was connected to a level column on the temperature matrix board (Fig. 2a). The one-way imbalance between the resistance of the measuring thermistor and the pre-set variable resistor selected was detected by an amplifier operating in an onoff mode and indirectly switching a power triac through a

zero voltage circuit; the latter reduces the electrical interference on other parts of the apparatus. Activation of a particular temperature column also switched an appropriate cooling level. A small flow was always maintained to partially compensate for general uncontrolled heat gains and this could be augmented by a partially open solenoid valve or by a fully open valve. The oxygen concentration in the circulating water was controlled by comparing the voltage from the oxygen sensor with the preset reference voltages selected by the oxygen matrix board (cf. temperature). These usually corresponded to 25% saturation intervals of air saturation (c. 2-3 mg 1-l) between 50 and 200% (4-20 mg 1-l). If out of balance, i.e. below during respiration or above during photosynthesis, a solenoid valve operated (V, Fig. I), and pulse unit (5 counts s-‘) and counter were started (Fig. 2b). This valve released water at a suitable rate from the system thus allowing conditioned water to enter and, because a constant head was maintained between the inlet and outlet, the time for which the valve was open could be used as a measure of water input and thus the quantity of oxygen consumed or produced could be calculated. The open time was printed at regular intervals (6 min). The output of the oxygen sensor was temperature compensated, using two thermistors to give an equal output at air saturation over the experimental range of temperatures. Oxygen concentrations were also determined chemically by the Winkler method at regular intervals and used to interpolate the changes in conditioned water, circulating water and oxygen concentration after correcting for barometric pressure changes. The linearity of response was checked by an additional calibration using water in equilibrium with an oxygen enriched mixture (40 : 60 oxygen/nitrogen) (Dawson & Henville, in prep.). The total delay between the detection of imbalance and the detection of balance, partially caused by the sensor, and partially by the need for the conditioned water to mix into the circulating water and reach the oxygen sensor, lead initially to fluctuations in the oxygen concentration. These were reduced by increasing the sensitivity and decreasing the response time of the oxygen sensor by using a very thin polyethylene membrane (0.04 mm) to cover the electrode. The 90% response time was then about 30 seconds which was sufficient to maintain the oxygen concentration in the circulating water within * I % of that required. Experimen.tal

Design

and Procedure

The plant material was always taken from the same established clones growing naturally at an average depth of 0.5 279

TI T2 T3 T4 T5

UNISELECTOR BANK I 24OVIL) -

--c--------

TEMPERATURE COMPENSATED D.O. PROBE & AMPLIFIER

N 240~

280

50 Hz

Fig. t. Circuit diagrams (b) oxygen concentration synthetic or respiration list).

for (a) water bath temperature selector; selector with reversal circuit for photostudies. (See Appendix 1 for component

50Hz

APPENDIX

I

Component

List

Temperature RI R2 R3 R4 R5 R6 R7 R8 RVI-6 Cl c2 DI ICI IC2 RLA SI

Control

Circuit

Fig. 2a

ZDr

Resistor, 5.6 kohm, %W Resistor, 5.6 kohm, %W Resistor, 1.0 kohm, %w Resistor, 4.7 kohm, %W Resistor, 4.7 kohm, %W Resistor, 47 ohm, %W Resistor, 20 kohm, 4W Resistor, 8.2 kohm, %W Potentionmeter, 5 kohm, IW (lo-turn Cermet) Capacitor, 0.22 PF Capacitor, IOO pF, 15V Diode IN4148 Integrated Circuit Type 741 Integrated Circuit Type CA 3059 (R.C.A. Ltd) Reed Relay, 12V (R-S Components 348-986) Uniselector, 6 Bank, 12 Way(A.E.1. T33653A-Service Trading Co., London W4 5BB) Thermistor TH-B12 (R-S Components 151-029) Triac, 2N5574 (R-S Components 261-558) Zener Diode BZY 88, 3.6 V

Dissolved

Oxygen

RI-R6 R7 R8 Rg RIO RVI-RV6 RV7 RV8 RV9 CI c2 C3 DI MI ICI ICI RLB SI sv

Resistor, IOO kohm, %W Resistor, 1.5 kohm, %W Resistor 1.0 kohm, %w Resistor, 3g ohm, %W Resistor, IOO ohm, %W Potentiometer, 5 kohm, IW (lo-turn Cermet) Potentiometer, 5 kohm, I W (2o-turn Cermet) Potentiometer, 500 ohm, IW (2o-turn Cermet) Potentiometer, IOO kohm, I W (zo-turn Cermet) Capacitor, 0.22 PF Capacitor, 250 pF, 15V Capacitor, 0.47 PF Diode 1N4148 Meter 0-500 PA, 150 ohm (or Elmes Recorder 1002) Integrated Circuit Type 5A-4 (Ancom Ltd) Integrated Circuit Type 741 Transistorised Relay (R-S Components 349-254) Uniselector (See Temp. Control Component List) Solenoid valve, 24oV, 50 Hz Thermistor CL22 (ITT Electronic Services-Harlow) Thermistor GL53 (ITT Electronic Services-Harlow) Zener Diode, BZY 88, 4.7V Zener Diode, BXY 88, 3.OV

THI

QI

ZDI ZD2

River Frome and 0.3 m in the Bere Stream at Holly Bush, Bere Regis. It was normally collected as complete shoots, includingflowers, roots or moribund material if required to represent the typical stand, and was cleaned rapidly of as much attached detritus, animals and algae as possible, using water from the collection site at the same temperature. The plant material was introduced as quickly as possible to the chamber and the water circulation started. The initial temperature and oxygen concentration previously established, were as close as possible to those at the collection site. The typical experiment of 40-48 h duration started with a period of equilibrium near the collection temperature and oxygen concentration before ranging both over that encountered by the plant in the field. Changes in conditions were however chosen to cause the minimum stress to the plant and it was normal practice to return to the initial conditions near the end of the experimental sequence. Experiments on thermal tolerance were undertaken at the end of runs. The pla’nts and the system required up to 2 h of the 4 h period in order to stabilize to the new conditions selected, mainly to allow the conditioned water to stabilize to the new temperature or oxygenconcentration. Inphotosynthetic studies plants were given at least two consecutive dark periods during the normal night and in respiration studies at least two light periods were given during the normal day. At the end of an experimental run the plant material was removed from the apparatus, dried at 105°C overnight and weighed after cooling in a desiccator. The chamber was immediately resealed and an estimate of metabolism due to microbial build up undertaken. This was the error of sampling the oxygen concentration i.e. 0.2 mg, normally very low (< 2% of the respiration rate) and the measuring system itself (3.5 I) was stable to within 0~ (l.h)-’ (see later) when operated on tap water. Diffusion of oxygen between the mixing vessel and the circulating system was negligible. After use the apparatus was flushed with a small quantity of hypochlorous acid and drained; the apparatus was flushed with filtered river water before reuse.

m in the

Control

Cricuit

Fig.

2b

Results and discussion The apparatus was used to determine the rates of respiration and photosynthesis of Ranunculus penicillatus var. calcareus (R. W. Butcher) C. D. K. Cook under conditions typical of the site of collection. The annual range of water temperature was 3-26°C and dissolved oxygen concentrations varied from c. 50-170% of air saturation. 281

Errors The errors involved in the determination of metabolic rates using this apparatus can be assigned to three groups: (I) the variability of biological material; (2) the physical and chemical errors involved especially with the balancing of the recirculating system, and: (3) the analytical errors of the dissolved oxygen determinations. The first group was minimised by undertaking comparisons on the same clones of materials. The results for the effect of temperature on the plant typify the overall error range shown by shoots of such plant material (i 0.2 mg (g.h)-‘, Fig. 4). The second error group for balancing of the dissolved oxygen in the recirculating water is self-compensating with a variation in the electronic control of less than I% of the set oxygen concentration. The output of the system was sampled by continually allowing the released water to flow through the sample bottle used for the chemical Winkler determination of oxygen and the concentration in the water was periodically determined during steady state conditions in the chamber. The overall error of sampling this water and determining its oxygen concentration was about * 0.2 mg 1-l at. IO mg I-’ and * 0.4 at 50 mg 1-l and compares favourably with the variation in the oxygen equilibrium, saturation values expected from the various tables available. Respiration

of R. calcareus

Water velocity The specific respiration rate increased with velocity up to c. 200 mm s-’ but above 50 mm s-l, the increases were not significant (Fig. 3). These experiments were undertaken at air saturation and 15°C. Westlake (I 967) found no significant increase in the rate of respiration above 0.4 mm s-’ but however the maximum velocity reported was only 5 mm s-l and his technique meant the variance increased rapidly at the higher velocities. Over this low range of velocities his data for respiration and photosynthesis fitted to a relationship with the square root of the velocity (Westlake 1980) but with the wider range used in these experiments a relationship with the inverse of the square root gave a better fit (Fig. 3), especially at the higher velocities. The initial and final respiration rates were insignificantly different when measured at 50 mm s-‘. A standard velocity of between 50-70 mms-‘, i.e. on the plateau, was therefore chosen for most experiments; the errors attributable to velocity fluctuations in this region were insignificant by comparison with other errors. For example, a variation in 282

1.5 1

,t-

+

t--t

IO- /t Respiration mg Q.(gJ$

rate ~5 t

0’ 0

50 Water

Velocity

100 mm ~71

Fig.

3. The effect of water velocity on the respiration rate of R. the R. Frome. The fitted line is based on the relationship of hourly mean dark respiration rate to the reciprocal of the square root of the velocity (I/(V)‘.~) and has a highly significant correlation (r = 0.96***, n = see text). Ranges of values determined are indicated by lengt of individual lines. Rates are based on dry weight of plant material.

calcareus from

respiration of + 0.05 at 50 mm s-’ would cover a range of 30-100 mm s-’ if this error was wholly attributable to velocity fluctuation. Temperature The specific respiration rate for complete plant shoots increased with temperature from 5” to 25°C (Fig. 4). At temperatures above 25”C, which are rarely reached in the river, permanent damage (falling rates of metabolism) occurred to an increasing extent. Below this region the temperature coefficient was c. 2.5 when expressed as a QIO; this is in general agreement with the low velocity experiments

Respiration rate w Oz.(g.h)-'

< Ij !&g: 1:

1

0.5022 0 Fig. 4. The of summer R. Frome I0 mg I-‘.

:* : 10

20 Temperature 'C

30

effect of water temperature on the rate of respiration (0) and late autumn(e) shoots ofR. calcareus from the at East Stoke at a dissolved oxygen concentration of The mean values are from five similar experiments

obtained in

1974

and in 1976 (1974 dots of set displaced to the left,

1976 to the right). The mean value which is plotted is based on the six minute estimates of water released over the final stable z hours of each temperature period and whose individual 95% confidence limits were + 0.19 mg (g.h)-‘.

of both Owens & Maris (1964) and Westlake (1967) for this species. At any particular temperature the respiration in autumn was higher than in the summer in both 1974 and 1976 for plants from the R. Frome; this was probably mainly due to the higher ‘proportion of active apical parts in the biomass at the time, but there may have been some biochemical adaptation to the lower river temperatures. The seasonal changes in the rate of respiration were studied in more detail in plants of R. calcareus from the Bere Stream at Hollybush, Bere Regis (Fig. 5). This site is on a spring-fed stream with clearer shallower water and it has a narrower annual temperature range (5.5-155°C) than the R. Frome at East Stoke (3-26”(Z). At Hollybush the daily temperature fluctuations were greater than variations in the monthly means, which varied little from 10°C whereas in the R. Frome daily variations were smaller and progressive changes extending over several days occurred. At Hollybush seasonal variations in the respiration rate at 10°C were closely related to the seasonal rates of increase of biomass and thus to the seasonal growth pattern of the plants (Dawson 1976). In addition to the seasonal changes in the respiration rate shown at particular temperature, there were changes in the temperature at which permanent

damage started to occur, indicating that there was some seasonal temperature adaptation. Comparisons between the sites on the Bere Stream and R. Frome were difficult because of the non-coincidence of growth cycles and the occasional near absence of plants at one or other of the sites. Also the majority of the Hollybush determinations were made at very low constant water velocities (c. 0.32 mm s-‘) using a simpler version of the apparatus with only through water Row. Oxygen concentration There were almost linear increases in the respiration rate with increases in dissolved oxygen in the circulating water over the range 50-150% of air saturation and at temperatures over the range IO-25’C (Fig. 6). This linear relationship differs from the data of Owens & Maris (1964) but their experiments were conducted in static and agitated water and dissolved oxygen was allowed to decline to low levels as a result of respiration. Photosynthesis Carbon

Mean growth rate,

When net photosynthesis was determined in this apparatus, starting with river water in equilibrium with air, and using nitrogen with 0.03% carbon dioxide to condition the water replaced while controlling the oxygen, photosynthesis declined rapidly after the first hour. After about four

so-

biomass increase 9 w~~~$2 25 -

Respiration rate

0

sources

I

I

I

1

Respiration fate Temperature ‘C Monthly range

mg O&g. h?

_

2

0 “J

FM’AMJ’JAS’OND’J Month

FM’

Fig, 5. The seasonal course of (a) the increase of biomass of R. calcareus from the Bere Stream at Hollybush, Bere Regis; (b) the plant respiration at IO”C, IO mg 1.’ and 0.32 mm s-’ determined on complete plant shoots from the above site and the probable changes during the growth cycle (dashed line) and(c) the monthly ranges of water temperature at the collection site.

0 Oxygen

10 concentration

mg

20

i1

Fig. 6. The effect of dissolved oxygen concentration on the rate of respiration of complete summer shoots of R. culcareus from the R. Frome at a range of temperatures. For clarity the 95% confidence limits are not included but were an average * 0.22 mg (g.h)-’ for all temperatures.

283

hours the net oxygen output was about zero. The initial rate could be restored by flushing the system with fresh river water in equilibrium with air but photosynthesis again declined after an hour. Results for photosynthesis experiments were obtained either using the stable period after flushing or using water conditioned by nitrogen containing 3% carbon dioxide, which stabilised photosyntheses for long periods. The addition of sodium bicarbonate or hydrochloric acid did not restore the initial rate of photosynthesis. The pH of the initial conditioned waters was about 8.38.6, whereas the pH of the water in equilibrium with 3% carbon dioxide was about 7,2-7.4, which is the lower end of the range of values normally found in the river (c. 7.28.9) Oxygen concentration Net photosynthesis at 44 J rn-’ s-’ irradiance decreased when the dissolved oxygen in the circulating water was increased both at 0.03% and at 3% carbon dioxide levels. Temperature An increase in temperature in these experiments decreased the rate of net photosynthesis (e.g. Fig. 7, pH 8.4), which is not unexpected if the temperature coefficient of photosynthesis is less than that of respiration. Under the conditions of this particular experiment, using.material gathered in early August, net photosynthesis would have become negative at about 17Oc, only 2O above the mean water temperature at that time. At the lower pH a much larger temperature change would be needed. This stresses that hight light, cool water and supersaturation with carbon dioxide are favourable for good growth of Ranunculus, as already observed in the field, and that the seasonal adaptations of the respiration rate to the mean water temperature are important. Most of the decrease in net photosynthesis shown when either oxygen concentration or temperature are increased can be accounted for by the effect of these factors on dark respiration.

Summary

Apparatus to measure the rates of respiration and photosynthesis of aquatic macrophytes from rivers, under ex284

Net 2 Photosynthesis mg

0&O-C’

1

-1

-1 Oxygen

10 ‘“’ 20’ I*) 30 concentration mg O#

Fig. 7. The effect of dissolved oxygen concentration on the rate of net photosynthesis of complete shoots from the R. Frome at two temperatures and at two pH 7.2 and 8.4 (or carbon dioxide equilibrium with 0.03% and 0.3% in the atmosphere). The rates at pH 7.2 could be maintained for the normal experimental period (4 h) whereas the initial rate at 0.03% for the first hour (see text).

and pH 8.4 was only maintained

perimental conditions close to those of their natural environment, was developed. The temperature, light regime and concentration of dissolved oxygen could be selected and controlled automatically and flow rates up to 200 mm s-l could be set. The water was partially recirculated and changes in the oxygen concentration were measured by monitoring the additions and removals of water of known oxygen concentrations that were necessary to maintain the concentration constant. The dark respiration rate of complete shoots of Ranunculus peniciliatus var. calcareus responded to water velocity, reaching a plateau above about 50 mm s-‘. Respiration rates increased with temperature up to 25’,C, with a Q,o of about 2.5, but at higher temperatures permanent damage occurred. There was some evidence ofadaptation to higher summer temperatures. At any particular temperature the seasonal changes in specific respiration were related to seasonal changes in the growth rate of the stands. Respiration rates also increased with increases in dissolved oxygen concentration. The rate of net photosynthesis by similar shoots was decreased by increases in both temperature and oxygen. Most of this effect could be accounted for by the predicted increases in dark respiration. With summer shoots, in water with low free carbon dioxide, net photosynthesis became zero at only a few degrees above the mean river temperature. The rate of net photosynthesis declined rapidly after one hour in water about pH 8.5, but could be maintained indefinitely in water about pH 7.3. This did not appear to be simply related to the free carbon dioxide and bicarbonate equilibrium.

Acknowledgements We wish

to thank

design

of

the

oxygen

probe

W. H.

Moore

temperature and

for permission

compensation

its recorder

output.

to use the circuit

The

for

apparatus

the was

by H. Zschorn, J. Morgan and S. Shinn while Miss W. Purchase, a student from Bath University, P.

constructed Henville, able

T. Ware

assistance

This

work

Agriculture,

and

Dr

at various was

supported,

F. B. Michaelis

provided

valu-

times. in part,

by the

Fisheries and Food (Commission

Ministry

of

6. I I).

References Dawson, F. H. 1976. The annual production of the aquatic macrophyte Ranunculus penicillatus var. calcareus (R. W. Butcher) C. D. K. Cook. Aquatic Botany 2: 51-73. Dawson, F. H. & Henville, P. in prep. Some of the characteristics of Mackereth-type dissolved oxygen sensors and the optimisation of their output in field applications. Gessner, F. & Pannier, F. 1958. Der Sauerstoffverbrauch der Wasserpflanzen bei verschiedenen Sauerstoffspannungen. Hydrobiologia IO: 323-35 I. GlBnzer, U. 1974. Experimentelle Untersuchungen iiber das Verhalten submerser Macrophyten bei NHA’-Belastung. Verhandlungen der Gesellschaft Gkologie, Saarbriicken 1973: 175-179. Hough, R. A. 1974. Photorespiration and productivity in submersed aquatic vascular plants. Limnology & Oceanography

19: 912-927. Hartman, R. T. &Brown, D. L. 1967. Changesininternalatmosphere of submersed vascular hydrophytes in relation to photosynthesis. Ecology 48: 252-258. McDonnell, A. .I. & Weeter, D. W. 1971. Respiration of aquatic macrophytes in eutrophic ecosystems. Research Publication No. 67, Institute for Research into Land and Water Resources, Pennsylvania State University. Owens, M. & Maris, P. J. 1964. Some factors affecting the respiration of some aquatic plants. Hydrobioiogia 23: 533-543. Westlake, D. F. 1967. Some effects of low-velocity currents on the metabolism of aquatic macrophytes. Journal of Experimental Botany 18: 187-205. Westlake, D. F. 1978. Rapid exchange of between plant and water. Verhandlungen der Internationalen Vereiningung fiir theoretische und angewandte limnologie 20: 2363-2367. Westlake, D. F. 1980. Primary Production -.Macrophytes In: Le Cren, E. D. & Lowe-McConnell, R. H. (Eds.)The Functioning of Freshwater Ecosystems, pp. 177-182. Cambridge: Cambridge University Press.

285