The isolation of leaf protoplasts from the submerged aquatic angiosperm Potamogeton lucens L.

Abstract. A method for isolating protoplasts from leaves of the submerged aquatic angiosperm Potamogeton lucens L. is described. The protoplasts are produced enzymatically from leaf strips using 1.5% (w/v) Cellulysin, 0.3% (w/v) Macerozyme R10, and 2.5% (v/v) β-glucuronidase at 27°C in the dark. Subsequently the protoplasts are purified on a discontinuous Ficoll gradient. The yield obtained is approximately 20% of the starting material on a chlorophyll basis. The viability is high, namely more than 90% as estimated with Evans Blue. Cells of the intact leaves of P. lucens can use HCO₃⁻ for photosynthesis. ¹⁴CO₂ fixation experiments at pH 6.0 and 8.5 suggest that these isolated protoplasts can also use bicarbonate as a carbon source.     

Coupling of Proton Fluxes in the Polar Leaves of Potamogeton lucens L

An attempt has been made to quantify the light-induced H⁺ effluxand influx observed in polar leaves of Potamogeton lucens.Theseproton fluxes are spatially separated. The H⁺ efflux, mediatedby a plasmalemma bound H⁺ –ATPase, occurs across theplasmamembrane at the morphological lower epidermis and is accompaniedby an H⁺ influx (or OH^– efflux) at the upper side oftheleaf. As a result, these leaves exhibit a remarkable pH–polarityin the light. The pH near the lower epidermis may drop to avalueas low as 3.5, while a pH of about 10.5 can be observed at theupper epidermis. Obviously this phenomenon requires theco–ordinationof transport processes in the different cell layers of the leaftissue. These observations led to quantitative studies oftherelation between the H⁺ fluxes at either plasmalemma. Thesefluxes were calculated from the pH values recorded at twodistancesfrom the leaf surface. Although the H⁺ influx always exceededthe efflux, a coupling between the transport processesacrosseither plasma membrane became evident from the time–coursesof the two fluxes. Key words: Potamogeton lucens, proton flux, flux coupling, pH–;polarity     

Translocation of 14Carbon in Tobacco following Assimilation of 14Carbon Dioxide by a Single Leaf

¹⁴CO₂ was assimilated by single leaves (presentation leaves)of tobacco plants for periods of 2–3 hours. The plantswere then kept in air in continuous light and the redistributionof radioactivity determined at various times up to 96 hours.There was a complete turnover of sucrose in the presentationleaf in about 24 hours without change in amount. Starch turnedover more slowly and simultaneously increased in amount. 20–30per cent. of the radioactivity appeared to be irreversibly incorporatedinto the presentation leaf. Of the material exported from thepresentation leaf some 3 per cent. reached the upper leavesand stem apex. Import into leaves above the presentation leafwas completed in about 6 hours. No activity appeared in leavesbelow the presentation leaf, therefore the balance of the exportedactivity was retained in the stem and roots. The distribution of radioactivity in the leaves followed a well-definedpattern determined by the vascular interconnexions. Radioautographs of stem sections provided some information concerningdistribution of radioactivity in the stem.     

Short-Term Fixation of 14Carbon by the Submerged Aquatic Angiosperm Potamogeton pectinatus

In ¹⁴C fixation experiments, 3-phosphoglyceric acid was thefirst product of carbon assimilation in the light in Potamogetonpectinatus. The pattern of early ¹⁴C-labelled compounds wasthe same over a range of pH values of the medium from 3.5 to8.1. Rates of ¹⁴C incorporation declined with increasing pHof the medium indicating that free CO₂ is the major exogenoussource of carbon for photosynthesis in Potamogeton pectinatus.     

Photosynthetic Assimilation of 14CO2 by Mature Brown Stems of the Tea Plant (Camellia sinensis L.)

The fixation of ¹⁴CO₂ by mature brown stems of the tea plantwas studied by supplying ¹⁴CO₂ to selected stems of pruned andintact plants for 24 h under field conditions. Utilization of¹⁴C assimilates for the production of new shoots was also examined.The photosynthetic nature of the fixation of ¹⁴CO₂ is demonstrated.The efficiency of this fixation was very low compared with thattaking place in leaves. The movement of labelled assimilatesfrom stem bark to the roots was inappreciable, whereas newlyemerged shoots on the pruned frame drew labelled assimilatesfrom stem bark.     

Isotope Fractionation in Photosynthetic Bacteria during Carbon Dioxide Assimilation

The δ _PDB¹³C values have been determined for the cellular constituents and metabolic intermediates of autotrophically grown Chromatium vinosum. The isotopic composition of the HCO₃^- in the medium and the carbon isotopic composition of the bacterial cells change with the growth of the culture. The δ _PDB¹³C value of the HCO₃^- in the media changes from an initial value of −6.6‰ to +8.1‰ after 10 days of bacterial growth and the δ _PDB¹³C value of the bacterial cells change from −37.5‰ to −29.2‰ in the same period. The amount of carbon isotope fractionation during the synthesis of hexoses by the photoassimilation of CO₂ has a range of −15.5‰ at time zero to −22.0‰ after 10 days. This range of fractionation compares to the range of carbon isotope fractionation for the synthesis of sugars from CO₂ by ribulose 1,5-diphosphate carboxylase and the Calvin cycle.     

Genetic diversity of an endangered aquatic plant,Potamogeton lucens subspecies sinicus

The genetic diversity of the species, Potamogeton lucens subsp. sinicus var. teganumensis, which is critically endangered in Japan, was investigated. This species now occurs in only two known localities in Japan. One is a native population (Oitoike population), but the other (Teganuma-Okahotto population) is found in a small artificial pond that was dug in 1998. It is considered that the Teganuma-Okahotto population grew from a soil seed bank. Based on RAPD variation, we compared the genetic diversity of the two populations of P. lucens var. teganumensis in Japan and one population of P. lucens subsp. sinicus var. sinicus in China. The Teganuma-Okahotto population showed RAPD variation, suggesting that it may be derived from more than one seed buried in old sediments. This population also had the highest value of Shannon's Information Index among the three study populations. This finding suggests that seeds buried in sediments can contain genetic variability, and may be used to conserve the genetic diversity of rare and endangered plants.     

Assimilation of 14CO2 by the Inflorescence of Poa annua L. and Lolium perenne L.

The relative assimilatory activity of the inflorescence, itsindividual components, and the leaves of flowering tillers ofPoa annua L. and Lolium perenneL. was determined over the periodfrom inflorescence emergence to seed shedding. The pattern of¹⁴CO₂ fixation was similar for both species and the inflorescencewas by far the most important assimilatory organ of the reproductivetiller, particularly over the latter period of seed developmentas leaf senescence progressed. With the exception of the seedsall parts of the inflorescence showed significant assimilatoryactivity and the lemmas and paleas accounted for 40–50per cent of the total ¹⁴C fixed by the inflorescence in bothspecies. The importance of the grass inflorescence as a photosyntheticstructure is discussed in relation to similar studies on cereals. Poa annua, Lolium perenne, carbon dioxide assimilation, inflorescence     

Photosynthetic Fixation of 14Carbon by Internodal Cells of Chara corallina

Maximum fixation rates of 120 and 60 pmol cm^–2 s ^–1wereobtained when exogenous carbon was supplied as ¹CO₂ and H¹⁴CO₃^–respectively. These values are considerably higher than thosepreviously reported for this species. A kinetic analysis wasperformed on this data. Substrate saturation in the concentrationrange 1.0–1.5 mM was observed for both CO₂ and HCO₃^– In the presence of exogenous CO², a linear relationship wasobserved between light intensity and fixation while the HCO₃^–relationship was slightly sigmoidal. Fixation saturated at intensitiesof 15–20 W m^–2 and 13–15 W m^–2 for exogenous¹⁴CO² and H¹⁴CO₃^–respectively. The presence, in this species, of an extremely active HCO₃^–transport system, situated in the plasmalemma, demonstratesthat when alkaline solutions are employed the involvement ofthis ion cannot be ignored during electrical studies on thismembrane. The maximum H¹⁴CO₃^– influxes obtained duringthis study are the largest ionic fluxes measured for any Characeanspecies. It was demonstrated that CO² for fixation can be supplied simultaneouslyby gaseous diffusion and HCO₃^– transport (cf. Raven, 1968).Inhibition of H¹⁴CO₃^– influx was observed in the presenceof Tris, Tricine, and borate buffers, and CO₃^{2 –} alsoappeared to act as a strong inhibitor. The possible mechanism(s)by which this inhibition occurs is discussed.     

ent-Labdane glycosides from the aquatic plant Potamogeton lucens and analytical evaluation of the lipophilic extract constituents of various Potamogeton species

Two new ent-labdane glycosides, one known furano-ent-labdane and a new hydroxylated fatty acid were isolated from the dichloromethane extract of the freshwater aquatic plant Potamogeton lucens. The new compounds were assigned the structures of beta-d-glucopyranosyl-8(17),13-ent-labdadien-16,15-olid-18-oate, 18-beta-d-glucopyranosyloxy-8(17),13-ent-labdadien-16,15-olide and 13(R)-hydroxy-octadeca-(9Z,11E,15Z)-trien-oic acid by spectroscopic means. The algicidal activity of these compounds was tested against Raphidocelis subcapitata. Based on our previous study of Potamogeton pectinatus, other constituents were identified in P. lucens by LC-UV-MS, LC-NMR and GC-MS. The lipophilic extract profiles of both species are presented. Two other species, Potamogeton perfoliatus and P. crispus, were also investigated by analytical comparison of their non-polar extracts. The distribution of ent-labdanes characterized in Potamogeton is summarized.     

Photosynthetic plasticity in Potamogeton pectinatus L. from Argentina: strategies to survive adverse light conditions

Argentine Potamogeton pectinatus L. was grown in The Netherlands under laboratory conditions at four light intensities (50, 100, 150 and 200 µE m^–2 s^–1), and photosynthetic performance was evaluated after about 1, 2 and 3 months of growth. At these moments, chlorophyll-a and -b and tissue N and P content were also determined. During the growing period, plant lengths and number of secondary shoots were measured. In the field in Argentina, photosynthetic performance of P. pectinatus was also measured at different light intensities created by artificial shading at various times during the growing season. Field and laboratory photosynthetic results were in good agreement. P. pectinatus showed a significant plasticityin its photosynthesis, rather than in morphology. A fairly constant maximum photosynthetic rate with reduced light enabled the plants to maintain netproduction rates rather unaffected at low light intensities. Still, it can be predicted that increasing turbidity from 1–2 m^–1 at present to 3 m^–1 could lead to a strongly light-limited growth which should reduce the present weed problem considerably. Such a turbidity increase might be achieved by the introduction of a fairly dense bottom-feeding fish population like Common carp (Cyprinus carpio L.).     

Characterization of the Type of Photosynthetic Carbon Dioxide Fixation in Jute (Corchorus olitorius L.)

Activities of photosynthetic and photorespiratory enzymes viz.,ribulose bisphosphate carboxylase, phosphoenol pyruvate carboxylaseand glycolate oxidase from jute (Corchorus olitorius L.; cv.JRO 632) leaves were compared with those from maize (C₄) andsunflower (C₃) leaves. The photosynthetic CO₂ fixation products,the release of ¹⁴CO₂ in light and dark following photosynthesisin ¹⁴CO₂, chlorophyll a: b ratio, gross leaf photosyntheticrate and dry matter production rate were also studied. The resultsshow that jute is a C₃ plant. Key words: Jute, Corchorus olitorius, C₃ photosynthesis     

石斛属植物光合碳同化途径地理分异研究进展

系统总结了我国石斛属植物光合作用现状,结合石斛属植物光合作用碳同化途径研究,概括了兼性景天酸代谢植物的碳同化路径特征,分析了景天酸代谢表达程度与生态环境的关系,进而归纳出石斛属植物在光合作用碳同化途径方面存在地理分异。最后指出了石斛属植物系统发育研究领域存在的问题,提出从系统发育视角,采用分子系统学技术来研究石斛属植物光合碳同化途径地理分异的生物学机制。     

Photosynthetic Units and Carbon Assimilation in Leaves of Grain Sorghum under Different Light Intensities

The Cyt f and P700 contents in leaves of three Sorghum, varietieswere measured, in relation to their carbon assimilation, underdifferent light intensities during growth. At the maximum irradiationused (1,800 µE m^–2 s^–1) the ratio of P700to Cyt f was close to unity, whereas under low irradiation (450µE m^–2 s^–1) the ratio of P700 to Cyt f rangedfrom two to three. A strikingly positive correlation existedbetween the P700 contents of the leaves and their rates of carbondioxide fixation, dry matter production and Cyt f contents,only when the plants were grown under high light intensities.The P700 content of the leaves in plants grown under low irradiationwas unrelated to the contents of Cyt f. Thus, at a high lightintensity there is a close relationship between the Cyt f andP700 levels, but at low intensities the amounts of electroncarriers and the reaction centre are independent. (Received March 7, 1983; Accepted August 24, 1983)     

Role of Potassium in Carbon Dioxide Assimilation in Medicago sativa L

Alfalfa was grown hydroponically in 0, 0.6, and 4.8 millimolar K in order to determine the influence of tissue level of K on photosynthesis, dark respiration, photorespiration, stomatal and mesophyll resistance to CO₂, photosystem I and II activity, and synthesis and activity of ribulose 1,5-bisphosphate carboxylase (RuBPc).     

Physiological Site of Ethylene Effects on Carbon Dioxide Assimilation in Glycine max L. Merr

The physiological site of ethylene action on CO₂ assimilation was investigated in intact plants of Glycine max L., using a whole-plant, open exposure system equipped witha remotely operated single-leaf cuvette. The objective of the study was met by investigating in control and ethylene-treated plants the (a) synchrony in response of CO₂ assimilation, stomatal conductance to water vapor, and substomatal CO₂ partial pressure; (b) response of CO₂ assimilation as a function of a range of substomatal CO₂ partial pressures; and (c) response of CO₂ assimilation as a function of a range of photon flux densities. After exposure to 410 micromoles per cubic meter of ethylene for 2.0 hours, CO₂ assimilation and stomatal conductance declined in synchrony, while substomatal CO₂ partial pressure remained unchanged until exposure times equaled and exceeded 3.0 hours. Because incipient changes in CO₂ assimilation occurred without a change in the CO₂ partial pressure in the leaf interior, it is concluded that both stomatal physiology and the chloroplast's CO₂ assimilatory capacity were initial sites of ethylene action. After 3.5 hours the effect of ethylene on stomatal conductance and CO₂ assimilation exhibited saturation kinetics, and the effect was substantially more pronounced for stomatal conductance than for CO₂ assimilation. Based on the response of CO₂ assimilation to a range of substomatal CO₂ partial pressures, ethylene did not affect either the CO₂ compensation point or carboxylation efficiency at subsaturating CO₂ partial pressures. Above-ambient supplies of CO₂ did not alleviate the diminished rates of CO₂ assimilation. In partitioning the limitations imposed on CO₂ assimilation in control and ethylene-treated plants, the stomatal component accounted for only 16 and 4%, respectively. The response of CO₂ assimilation to a range of photon flux densities suggests that ethylene reduced apparent quantum yield by nearly 50%. Thus, the pronounced decline in net photosynthetic CO₂ assimilation in the presence of ethylene was due more to a loss in the mesophyll tissue's intrinsic capacity to assimilate CO₂ than to a reduction in stomatal conductance.