Journal of Experimental Botany, Vol. 33, No. 133, pp. 241-250, April 1982 The Dynamics of Carbon Supply from Leaves of Barley Plants Grown in Long or Short Days A. J. GORDON, G. J. A. RYLE, D. F. MITCHELL AND C. E. POWELL The Grassland Research Institute, Hurley, Maidenhead, Berkshire, SL6 5LR, UJC. Received 26 October 1981 ABSTRACT The role of the mature leaf in supplying carbon for growth in other parts of the plant was examined using a steady-rate 14 CO 2 labelling technique. The pattern of events occurring in the leaf during one complete 24 h cycle was compared in plants grown in, and adapted to long and short photoperiods. The rates of leaf photosynthesis, night respiration and daytime loss of carbon from the growing regions of the plant Were similar in long or short photoperiods. As a percentage of the total carbon fixed during the photoperiod, total respiration was c. 50% for short day plants but only 25% for long day plants. Thirty to forty per cent of the carbon fixed during the photoperiod was retained in the leaf for export during darkness the rest was exported immediately. In leaves of short day plants sucrose and starch were the main form of the stored carbon. By the end of the dark period these compounds had been almost completely depleted. In leaves of long day plants there were much larger basal levels of sucrose and starch, upon which the diurnal variations were superimposed. These leaves also accumulated fructosans. The delay in starch remobilization previously found in leaves of short day plants was also evident in leaves of long day plants even though large concentrations of sucrose and fructosans were present This suggests the presence of distinct pools of sucrose in the leaf. INTRODUCTION Recent publications have examined the patterns of carbon supply from leaves of uniculm barley plants during light and dark periods. Using a steady state U C-labelling technique (Gordon, Ryle, Powell, and Mitchell, 1980a) we have calculated rates of carbon export and accumulation as well as that of respiration. In other experiments carbohydrate levels were measured and were reported in a subsequent paper (Gordon, Ryle, and Webb, 19806). These plants were grown in short photoperiods (8 8-5 h) in a controlled environment and displayed remarkable adaption to this environment in the way that carbon was metered out for use in the growing regions of the plant. We report here experiments to observe the effect on the plant of a photoperiod of 16 h. The results are compared with those obtained for plants adapted to short photoperiods. Apart from observations of general trends in the growth of the plants, we were particularly interested in the mature leaf: its photosynthetic rate, the accumulation and export of carbon and the chemical nature of compounds stored and remobilized during light and dark periods. In the leaves of barley plants grown in short days we have proposed that starch remobilization during darkness occurs only when the sucrose concentration falls to a low level. Since we expected that in long days the levels of carbohydrates would be higher than in short days, we were also interested in the effect that this might have on sucrose and starch metabolism.
242 Gordon et al. Carbon Supply from Barley Leaves MATERIALS AND METHODS Plant material and growth conditions Plants of uniculm barley (Kindred Uniculm 97) were grown as described previously (Gordon, Ryle, and Powell, 1977, 1979) in Saxcil cabinets with a day/night temperature regime of 23/18 C and a photoperiod of 16 h (long day plants). A mixture of fluorescent and incandescent lamps provided a quantum flux density of 817 //E m~ 2 s~'. Uniform plants were used as soon as the second leaf was fully expanded. Short day plants (8-5 h photoperiods), from a separate experiment, were grown in conditions, which, in all respects except the daylength, were identical to those for long days. Some of the information concerning short day plants has been published previously (Gordon et al., 1980a) and a portion of this is reproduced here to aid comparison between long and short day plants (Table 1). Steady state "C-labelling The terminal 13 cm of the attached second leaf of 20-22 plants was enclosed in a Perspex assimilation chamber situated in a Saxcil growth cabinet which provided identical light, temperature and humidity conditions to those in which the plants were grown. Throughout the photoperiod the leaves in the assimilation chamber were exposed to a stream of outside air supplemented with 14 CO 2 to give a I4 CO 2 / 12 CO 2 mixture of low, constant specific activity (Gordon et al., 1980a). The air was supplied to the leaves at a constant rate set to ensure that CO 2 depletion by the photosynthesizing leaves, monitored continuously with an infrared gas analyser, did not exceed 10%. During the first 0-5 0-75 h of the photoperiod CO 2 fixation rose from zero to maximum and then remained constant for the entire photoperiod. At intervals during the photoperiod, whqe steady state 14 CO 2 fixation continued, and during the subsequent dark period, leaves were removed from the assimilation chamber. The fed leaf (the portion of the second leaf exposed to "CO^) was cut off immediately, its length and area measured and then immersed in boiling 80% (v/v) ethanol. The remainder of the exposed leaf (rest of leaf blade plus sheath), the terminal meristem (including all younger leaves and stem which was 1-3 mm long), the roots, and the old leaves were either dried, weighed and oxidised to determine the l4 C content or were killed in boiling 80% ethanol. The plants parts killed in ethanol were stored at 2 C for subsequent extraction and analysis as described before (Gordon et al., 1977, 1979). Carbohydrate analysis The absolute quantities of glucose, fructose, sucrose, oligofmctosans and starch were determined as described previously (Gordon et al., 1977, 1979). Radioactivity in neutral sugars was determined by initial separation by descending paper chromatography (Whatman No. 1, solvent :propan-l-ol; ethyl acetate: water 7:1:2 by vol.). Each sugar spot was then cut out, combusted in a Packard sample oxidiser and the 14 C content determined by scintillation counting. Theory and calculations based on steady state labelling data In an earlier publication (Gordon et al., 1980a) we discussed the theory behind the technique of steady state labelling of attached leaves. Here we made no attempt to measure gross photosynthesis (Ludwig and Canvin, 1971) but have used the method to determine the respiratory loss of U C from the whole plant as labelled materials are exported from the fed leaf and are metabolized at their destinations (Gordon et al, 1980a). This loss of carbon can also be estimated from the difference between the calculated rate of 14 C accumulation by the leaf (determined from the known photosynthetic CO 2 depletion and from the known specific activity of the labelled CO^) and the rate of 14 C accumulation measured over several hours. When comparing the data from long and short day plants, radioactivity has been translated into mg C per unit leaf area. Expressed as such the data refer to events occurring in 1 dm 2 of fed leaf area or to the carbon derived from 1 dm 2 of leaf area. The factor used for converting radioactivity to mg C dm" 2 for short day plants was 1-253 kbq mg C" 1 (the rate of 14 C accumulation, 111 kbq h" 1 leaf" 1, divided by the photosynthetic rate, 8-86 mg C dm" 2 h" 1 ), and for long day plants, 1-396 kbq mg C" 1 (11-8 kbq h" 1 leaf"' divided by 8-45 mg C dm" 2 h" 1 )- RESULTS The I4 C content of long day plants and their parts during the 16 h steady state labelling of the young mature second leaf, and during the following 8 h dark period is shown in Fig. 1. The rate of 14 C accumulation by the whole plant during the photoperiod was constant and similar
Gordon et al. Carbon Supply from Barley Leaves 243 150 125 s 100 S 75 g o OS 50 25 0 0 10 15 20 25 Time after beginning of photoperiod /h Fio. 1. Accumulation of 14 C in long day plants and their parts during the 16 h steady state labelling of the young mature second leaf, and during the following 8 h dark period. Whole plant (#), fed leaf (O). roots ( ), terminal meristem (D). The unshaded and shaded areas on the time axis represent light and dark periods. Curves are fitted by eye. to that found for short day plants (Table 1). This is to be expected since both leaf photosynthesis and the daytime rate of loss of U C as respiration from long day plants were similar to their respective values in short day plants (Table 1). Further respiratory loss of 14 C from the long day plant during darkness was difficult to measure precisely because of variation in the data (Fig. 1), but was estimated to be about 9% of the total 14 C fixed during the photoperiod. Thus, of the total 14 C fixed by long day plants during one photoperiod, approximately 25% was lost in respiration within 24 h. Accumulation of l4 C in plant parts Fed leaf. During the photoperiod the leaf simultaneously exported and stored photosynthetically fixed carbon (Fig. 1, Table 1; Gordon et al., 1980a). The rate of 14 C accumulation in the fed leaf of long day plants tended to decline as time progressed (Fig. 1) but by the end of the photoperiod the carbon retained in the leaf amounted to 32% of the total fixed or 38% of that unrespired (Table 1). During darkness the 14 C content of the leaf declined at a constant rate (Fig. 1) but about 16% of the total 14 C fixed during the photoperiod still remained in the leaf after one complete diurnal cycle. Roots and terminal meristem. In these young long day plants of uniculm barley the main destination of 14 C exported from the leaf was the roots, where approximately 34% of the total C fixed by the leaf was accumulated by 24 h (47 mg C from 1 dm 2 of leaf). In contrast only 14% was found in the terminal meristem (19 mg C from 1 dm 2 of leaf). In short day plants
244 Gordon et al. Carbon Supply from Barley Leaves TABLE 1. Carbon supply from a young mature leaf of uniculm barley adapted to either long or short days Carbon fluxes calculated from the 8-5 h photoperiod" use of U C as a quantitative 16 h photoperiod tracer Total mgc Rate dm" 1 mg C dm" 2 h" 1 % Total mg C dm" 2 Rate mg C dm" 2 h -' % During photoperiod Photosynthesis Respiration Accumulation by whole plant Accumulation by leaf Accumulation by TM* Accumulation by roots Export from leaf 75-3 15-3 60-0 29-9 9-3 12-5 45-4 8-86 1-77 709 3-52 109 1-47 5-34 100 20 80 40 12 17 60 135-2 21-8 113-4 43-5 15-4 40-7 91-7 8-45 1-36 709 2-72 0-94 2-54 5-73 100 16 84 32 11 30 68 During darkness Respiration Accumulation by TM* Accumulation by roots Export from leaf 20-9 3-8 5-3 26-0 1-35 0-25 0-34 1-68 28 5 7 35 12-2 40 5-9 22-0 1-53 0-50 0-74 2-75 9 3 4 16 During complete 24 h Respiration Accumulation by whole plant Accumulation by leaf Accumulation by TM' Accumulation by roots Export from leaf 36-2 39-1 3-9 131 17-8 71-4 48 52 5 17 24 95 34 101-2 21-5 19-1 46-6 113-7 25 75 16 14 34 84 Some of the data under the heading '8-5 h photoperiod' are taken from Gordon et al., 1980a. * TM = Terminal meristem. the amounts accumulated by the roots and terminal meristem were 18 mg and 13 mg C from 1 dm 2 of leaf per day respectively (equivalent to 24% and 17% of the total 14 C fixed respectively). This points to a substantial difference in the allocation of resources which favours root growth in long day plants. However this was not at the expense of the terminal meristem since more carbon accumulated in the terminal meristem of long day plants compared with the same organ in short day plants (19 mg C versus 13 mg C from 1 dm~ 2 leaf)- In long day plants a further 14 mg C dm" 2 were found in the rest of the fed leaf and old leaves (4 mg C in short day plants). To substantiate the trends derived from the U C data, which have been outlined above, groups of 10 plants were harvested at intervals, dissected into the appropriate parts, then dried and weighed (Table 2). It is clear that the plants in 16 h photoperiods increased in weight at more than twice the rate of short day plants; also the root:shoot ratio and the specific leaf weight were higher in long day plants. Chemical distribution of 14 C and the carbohydrate status of the fed leaf During the 16 h photoperiod the leaves of long day plants exported about 5-7 mg carbon dm" 2 h" 1 while simultaneously accumulating 2-7 mg C dm" 2 h" 1 (Table 1). The rate of export
Gordon et al. Carbon Supply from Barley Leaves 245 TABLE 2. Growth characteristics of plants accustomed to long (16 h) or short (8-5 h) photoperiods Photoperiod Days after sowing Top Dry weights (mg) Root Total Root Top Specific leaf wt mg dm~ 2 Rate of weight increase 8-5 h 18 19 19 19 20 21 22 248 283 325 276 384 406 445 113 113 132 98 157 170 177 361 396 457 374 541 576 622 mean 0-46" 0-40 0-41 0-36 0-41 0-42 0-40 0-41. > 304 75mgd- 1 16 h 16 19 21 21 226 590 662 686 139 341 542 487 365 931 1204 1173 mean 0-621 0-58 0-82 0-71 0-68 J - 527 168 mg d~' was slightly less and the rate of storage in the leaf slightly more in short day plants than in long day plants (Table 1). In the dark period the rate of export from the leaf of long day plants was somewhat greater (2-75 mg C dm~ 2 h" 1 ) than that from short day leaves (1-68 mg C dm" 2 h~') but, since in short days the nights were longer, the total amount exported during darkness was similar in both long and short day leaves (22 mg versus 26 mg C). In long day plants the 14 C stored in the leaf was mainly in the form of neutral sugars (77% at 16 h) (Fig. 2) with a further 16% in the warm water-soluble (mainly high molecular weight fructosan, some starch and some sugar phosphates) and starch fractions. During the dark period the neutral sugars were rapidly exported from long day leaves, but some 46% of the 14 C in this fraction when the lights went off remained in the leaf at the end of the night. Thus a substantial fraction of the carbon accumulated by the leaf was carried over from one day to the next. This may be the reason why the specific leaf weight was so much higher in long day plants than in short day plants (Table 2). The neutral sugar fractions from the exposed leaves of both long and short day plants were examined by paper chromatography (Fig. 3A and B). It is clear that in both cases the rate of increase in sucrose concentration declined with time during the photoperiod and that during the dark period there was an exponential depletion of sucrose from the leaf. However, whereas in the short day plant, sucrose was almost completely depleted during darkness, in the long day plants, although the initial rate of depletion was almost double that in short days, the amount of 14 C remaining as sucrose at the end of the night was about 25% of that present 8 h earlier. Also in long day plants a substantial amount of 14 C was found as hexoses and oligofructosans reaching about 42% of the U C in neutral sugars at the end of the photoperiod. However, apart from a decline in the amount of U C in the trisaccharide (Fig. 3A) of long day leaves there was no evidence to suggest that carbohydrate was remobilized from the fructosan pool during darkness. Only very small amounts of 14 C were found in hexoses and fructosans in the leaves of short day plants. The amount of 14 C found in the warm water and starch fraction of leaves at the end of the photoperiod was approximately equal in the two light treatments (Fig. 2; Gordon et al.,
246 Gordon et al. Carbon Supply from Barley Leaves 50 00 20 10 0 0 5 10 15 20 25 Time after beginning of photoperiod / h Fio. 2. Diurnal variations in the amount of n4 C associated with the neutral sugar (0) and starch plus water soluble (O) fractions of the leaf supplied with 14 CO 2 during the photoperiod. Curves are fitted by eye. o o o o ->O O o O 10 15 20 25 0 5 10 15 Time after beginning of photoperiod /h 20 25 Fio. 3. Components of the neutral sugar fraction of the fed leaf from long (A) and short (B) day plants. Neutral sugars were separated by paper chromatography, individual spots cut out, combusted in a Packard sample ozidizer and the radioactivity determined by scintillation counting. Sucrose ( ), glucose plus fructose (O). trisaccharide ( ), oligosaccharides (A). Curves are fitted by eye.
Gordon et al. Carbon Supply from Barley Leaves 247 1980a), which suggests that the rate of accumulation of 14 C in this fraction in leaves of long day plants is only half that for short day plants. Absolute quantities of some leaf carbohydrates from long and short day plants during the diurnal period are shown in Fig. 4. The most striking feature of these results is that leaves of plants grown in long days contained much higher basal levels of particularly sucrose and starch, than similar leaves of short day plants. The data in Fig. 4 are expressed as mg carbohydrate per 100 mg structural dry weight (ethanol insoluble dry weight minus the 351.f D " 10 15 20 25 10 15 20 25 Time after beginning of photopenod / b Fio. 4. Diurnal variations in the leaf carbohydrate content of plants adapted to long (#) or short (O) photoperiods. A, sucrose; B, starch; c, glucose; D, ethanol insoluble fructosan. The data are expressed as mg sugar per 100 mg structural dry weight (ethanol insoluble dry weight minus the weight of starch). Vertical lines indicate the end of the photopenod. Curves are fitted by eye. weight of starch) but when expressed in terms of leaf area (short days 157-6 mg structural dry wt dm~ 2 ; long days 232-1 mg dm" 2 ) the differences in absolute quantities between long and short day leaves are even greater, rising from 3 mg sucrose dm~ 2 at lights-on to 35 mg dm~ 2 at the end of the photoperiod in short day plants and from 33 mg to 75 mg dm~ 2 during the 16 h photoperiod in long day plants. Thus the mean rates of sucrose accumulation in leaves during the photoperiod of long and short day plants were 2-63 and 3-76 mg h" 1 dm" 2 respectively. During darkness sucrose concentration rapidly declined in leaves of both long and short day plants, the initial rate of depletion being greater in leaves of long day plants. We have reported previously (Gordon et al., 1980a, b) that during darkness the starch level in barley leaves initially remains constant for several hours before remobilization begins. This pattern is also evident here. Both the I4 C content of starch in long day plants (Fig. 2) and the absolute quantity of starch in short day plants (Fig. 4B) suggest that, during the early part of the dark period, starch remains unmetabolized. Following this period of stasis, degradation begins and the starch level falls dramatically until the next photoperiod begins.
248 Gordon et al. Carbon Supply from Barley Leaves The variability in the data for starch levels in leaves of long day plants (Fig. 4B) restricts us to the conclusion, in this case, that starch increases during the day and decreases during darkness. The amounts of glucose and high molecular weight fructosan (insoluble in 80% ethanol but soluble in water) were small compared with sucrose and starch (about 5 mg per 100 mg structural dry wt at their peaks in long day leaves, negligible in short day leaves). The fructosan content of the neutral sugar fraction (soluble in 80% ethanol) was not measured. DISCUSSION Plants grown in, and adapted to long photoperiods (a) gained weight faster, (b) directed a much larger portion of the day's assimilate to the roots, and (c) had leaves that were much heavier per unit area, than plants grown in short days. The net photosynthetic rate of the youngest fully expanded leaf was the same in both long and short day plants and remained constant from early in the photoperiod until its end. Neither the generally higher carbohydrate concentrations in the long day plants, nor the accumulation during the photoperiod caused any reduction in photosynthetic rate (Chatterton, 1973; Upmeyer and Koller, 1973; Thome and Roller, 1974; Milford and Pearman, 1975; Chatterton and Silvius, 1979; Mauney, Guinn, Fry, and Hesketh, 1979; Potter and Breen, 1980). The overall respiratory losses were similar in both sets of plants (c. 35 mg C from 1 dm 2 leaf)- In relative terms this represented nearly half the carbon fixed by the short day plants, but only a quarter in the long day plants. In 24 h, therefore, long day plants accumulated (from 1 dm 2 leaf) c. 100 g carbon compared with only 40 mg in short day plants a difference which was reflected in their greater rate of dry weight increase (Table 2). Similarly, the patterns and rates of 14 C export, import and accumulation in the various organs of the long and short day plants (Table 1) were reflected in the rates of dry weight increase of these organs and were consistent with the contrasting root/shoot ratios and specific leaf weights that were observed (Table 2). Diurnal variations in carbohydrate levels Sucrose and starch were the predominant carbohydrates found in leaves of both long and short day barley plants. Hexoses and fructosans were present in almost negligible amounts in short day leaves and were only a small fraction of the total leaf carbohydrate content of long day plants. The most striking difference was the much greater basal levels of both sucrose and starch in leaves of long day plants. Superimposed on these basal levels, the diurnal fluctuations in sucrose and starch of the two sets of plants were similar. Generally sucrose accumulated at a decreasing rate during the light period. This decreasing rate may be correlated with an increase in the synthesis of starch, after a lag in the first part of the photoperiod. The rate of carbon export during the photoperiod from leaves of long day plants was estimated from the 14 C data to be c. 5-7 mg C dm~ 2 h~'. Since these leaves contained a large pool of unlabelled sucrose it is possible that the true export rate is somewhat greater. To test this possibility we calculated the specific activity of sucrose from the curves of Figs 3 A and 4 and compared the increase in specific activity during the photoperiod (0-0-338 kbq mg sucrose" 1 Fig. 5) with the specific activity calculated by assuming that the initial sucrose level of 33 mg dm" 2 remained unchanged. To this was added an increment calculated from the specific activity of fixed carbon (1-396 kbq mg C" 1 ) and the quantity of M C found as sucrose at any particular time (Fig. 3A). The fact that there is little difference between these estimates of specific activity suggests that unlabelled sucrose remaining in the leaf from
Gordon et al. Carbon Supply from Barley Leaves 249 0.0 0 5 10 15 20 25 Time after beginning of photoperiod/h Fio. 5. Specific activity of sucrose in leaves of long day plants calculated from data of Figs 3A and 4A (O) compared with that calculated from an estimate of sucrose concentration (#) assuming that the initial unlabelled sucrose concentration was 33 mg drrr 2 to which was added an increment based on the specific activity of fixed 14 CO 2 and the amount of 14 C found as sucrose (Fig. 3 A). previous days is not exported during the photoperiod. This seems likely if the unlabelled sucrose is mostly confined to a storage pool, for example the vacuole (Fisher and Outlaw, 1979). During steady state labelling in the photoperiod it can be imagined that labelled sucrose in the cytoplasm may be used both for immediate translocation and for storage in the vacuole. In the dark period the specific activity of the leaf sucrose declined suggesting that labelled sucrose is exported first. This, too, can be envisaged by assuming at least two pools of sucrose one of which (e.g. the cytoplasm) has a high specific activity derived from current photosynthate, and the other has a low specific activity (e.g. the vacuole) derived from a combination of cold sucrose from previous days and stored labelled sucrose from current photosynthate. We assume that sucrose for export in the dark would initially be derived from the cytoplasm (high specific activity) and would be replenished from the lower specific activity sucrose from the vacuole. Control of starch degradation It appeared from earlier work that starch degradation during darkness may be controlled by the concentration of sucrose or other intermediary metabolites outside the chloroplast (Gordon et al., 1980a, b). This proposal fits in well with current views about the potential control, not only of starch metabolism but of chloroplast metabolism generally (including photosynthesis) by a feedback mechanism from the sink regions of the plant (Herold, 1980). Our earlier reported findings with short day plants (Gordon et al., 1980a, b) indicated that starch remobilization was delayed until the sucrose level declined to about 20 mg dm~ 2. The same was true of the short day plants reported here; starch degradation began when the sucrose level was c. 15 mg dm~ 2. In the long day plants, leaf sucrose and starch levels were considerably higher but exhibited a similar diurnal fluctuation. Starch degradation appeared to be delayed in these leaves, also (Fig. 2), but began when the sucrose level was c. 36 mg dm" 2. Thus, it is clearly not total sucrose level which is controlling starch metabolism, but could be the sucrose level of the cytoplasm (see discussion in Gordon et al., 19806). However,
250 Gordon et al. Carbon Supply from Barley Leaves the link between sucrose level and starch metabolism must be sought by a different experimental approach. It is essential to establish the relationship between chloroplast, cytoplasm and vacuole as well as that between the mesophyll cells and phloem. ACKNOWLEDGEMENTS We are grateful to the staff of the Controlled Environment Facility for assistance and to Miss Jane Woledge, Dr. M. J. Robson and Dr. E. L. Leafe for constructive criticism of an earlier version of this manuscript The Grassland Research Institute is financed through the Agricultural Research Council. LITERATURE CITED CHATTERTON, N. J., 1973. Product inhibition of photosynthesis in alfalfa leaves as related to specific leaf weight Crop Set. 13,284-5. and Sn-vius, J. E., 1979. Photosynthate partitioning into starch in soybean leaves. I. Effects of photoperiod versus photosynthetic period duration. PI. Physiol., Lancaster, 64, 749-53. FISHER, D. B., and OUTLAW, W. H., 1979. Sucrose compartmentation in the palisade parenchyma of Viciafaba L. Ibid. 64,481-3. GORDON, A. J., RYLE, G. J. A., and POWELL, C. E., 1977. The strategy of carbon utilization in uniculm barley. I. The chemical fate of photosynthetically assimilated I4 C. /. exp. Bot. 28,1258-69. 1979. The strategy of carbon utilization in uniculm barley. II. The effect of continuous light and continuous dark treatments. Ibid. 30,589-99. and MITCHELL, D., 1980a. Export, mobilization and respiration of assimilates in uniculm barley during light and darkness. Ibid. 31,461-73. and WEBB, G., 19806. The relationship between sucrose and starch during 'dark' export from leaves of uniculm barley. Ibid. 31,845 50. HEROLD, A., 1980. Regulation of photosynthesis by sink activity the missing link. New Phytol. 86, 131^14. LUDWIO, L. J., and CANVDM, D. T., 1971. An open gas-exchange system for the simultaneous measurement ofthec0 2 and u CO 2 fluxesfrom leaves. Can. J. Bot. 49, 1299-313. MAUNEY, J. R., GUINN, G., FRY, K. E., and HESKETH, J. D., 1979. Correlation of photosynthetic carbon dioxide uptake and carbohydrate accumulation in cotton, soybean, sunflower and sorghum. Photosynthetica, 13,260-6. MILFORD, G. F. J., and PEARMAN, I., 1975. The relationship between photosynthesis and the concentrations of carbohydrates in the leaves of sugar beet. Ibid. 9(1), 78-83. POTTER, J. R., and BREEN, P. J., 1980. Maintenance of high photosynthetic rates during the accumulation of high leaf starch levels in sunflower and soybean. PI. Physiol., Lancaster, 66, 528-31. THORNE, J. H., and ROLLER, H. R., 1974. Influence of assimilate demand on photosynthesis, diffusive resistances, translocation and carbohydrate levels of soybean leaves. Ibid. 54,201-7. UPMEYER, D. J., and KOLLER, H. R., 1973. Diurnal trends in net photosynthetic rate and carbohydrate levels of soybean leaves. Ibid. 51,871 4.