Carbon Isotope Ratios of Epidermal and Mesophyll Tissues from Leaves of C3 and CAM Plants

The ¹³C values for epidermal and mesophyll tissues of two C₃plants, Commelina communis and Tulipa gesneriana, and a CAMplant, Kalancho daigremontiana, were measured. The values forthe tissues of both C3 plants were similar. In young leavesof Kalancho, the epidermis and the mesophyll showed S13C valueswhich were nearly identical, and similar to those found in C3plants. However, markedly more negative values for epidermalcompared to mesophyll tissue, were obtained in the mature Kalancholeaf. This is consistent with the facts that the epidermis ina CAM leaf is formed when leaves engage in C₃ photosynthesisand that subsequent dark CO₂ fixation in guard cells or mesophyllcells makes only a small contribution to total epidermal carbon. (Received January 27, 1981; Accepted May 14, 1981)     

不同城市居民头发中氧、氢稳定同位素检验和特征初探

目的 人体组织的稳定同位素组成与其生长期间的个体饮食情况、所处环境及代谢状况有关。人头发一经长出便不再与身体进行物质交换，化学性质稳定，易于采集，是研究人体组织稳定同位素组成的良好对象。构成人体的氧、氢元素主要来自于所摄入的水和食物，其中氧、氢稳定同位素组成会通过角蛋白的形式被记录于头发当中。不同地区居民头发中氧、氢稳定同位素组成差异可被用于推断人的饮食情况、生活地域和活动轨迹信息，在法庭科学等研究领域具有重要意义。方法 本研究利用元素分析仪-稳定同位素比质谱仪（EA-IRMS）对国内不同地区常住居民头发样本进行氧、氢稳定同位素比值检测和分析。结果 部分城市间居民头发δ¹⁸O和δ²H存在显著性差异，整体δ¹⁸O和δ²H存在显著正相关性。对所得稳定同位素数据进行判别分析推断头发的地域来源，其交互验证整体判别准确率为63.9%，结合碳、氮稳定同位素数据后，其判别准确率大幅提升，交互验证的整体判别准确率达到76.0%。随着判别分析中使用的稳定同位素种类的增加，判别函数模型的判别能力明显增强。结论 利用4种元素稳定同位素数据建立的多层感知器神经网络模型的整体判别准确率为82.8%，径向基函数神经网络模型整体判别准确率为78.8%，3种溯源推断数学模型中，多层感知器神经网络模型的判别准确率最高。     

The Identification of Photosynthetic Subpathways of Some C4 and CAM Plants

The photosynthetic subpathways of five C4 plants and one CAM plant were distinguished according to their chemical, physiological and cytological characteristics. Based on C4 acid decarboxylation enzymes, four C4 plants of Setaria glauca, Sporobolus indicus, Zoysia tenuifolia and Leptochloa chinensis all exhibited the functional high activities of PEP carboxykinase and aspartate aminotransferase as seen in the known PEP-CK subtype. The δ13C value of –12.43% in leaves of L. chinensis was also consistent with that range among PEP-CK subtype. So, these species were classified into PEP-CK subtype. However, their chloroplasts in bundle sheath cells were evenly distributed, not as that displayed centrifugally or centripetally in three typical subtypes. The even arrangement of chloroplasts in bundle sheath cells was likely to be an evolutional intermediate from centripetal (NAD ME type) to centrifugal types (NADP-ME and most PEP-CK types). The high activities of NAD-malic enzyme and aspartate aminotransferase, accompanied with the centripetally located chloroplasts, 0.057 of quantum yield and tile δ13C value of –15.3% in leaves of C4 dicot Euphobia hirta indicated characteristics of NAD-ME subtype. Moreover, CAM plant Aloe vera clearly fell into PEP-CK sybtype because of its high activity of PEP-CK both in whole leaf and green tissue.     

Quantum Yields of CAM Plants Measured by Photosynthetic O(2) Exchange

The quantum yield of photosynthetic O₂ exchange was measured in eight species of leaf succulents representative of both malic enzyme type and phosphoenolpyruvate carboxykinase type CAM plants. Measurements were made at 25°C and CO₂ saturation using a leaf disc O₂ electrode system, either during or after deacidification. The mean quantum yield was 0.095 ± 0.012 (sd) moles O₂ per mole quanta, which compared with 0.094 ± 0.006 (sd) moles O₂ per mole quanta for spinach leaf discs measured under the same conditions. There were no consistent differences in quantum yield between decarboxylation types or during different phases of CAM metabolism. On the basis of current notions of compartmentation of CAM biochemistry, our observations are interpreted to indicate that CO₂ refixation is energetically independent of gluconeogenesis during deacidification.     

Temperature Dependence of Carbon Isotope Fractionation in CAM Plants

The carbon isotope fractionation associated with nocturnal malic acid synthesis in Kalanchoë daigremontiana and Bryophyllum tubiflorum was calculated from the isotopic composition of carbon-4 of malic acid, after appropriate corrections. In the lowest temperature treatment (17°C nights, 23°C days), the isotope fractionation for both plants is −4‰ (that is, malate is enriched in ¹³C relative to the atmosphere). For K. daigremontiana, the isotope fractionation decreases with increasing temperature, becoming approximately 0‰ at 27°C/33°C. Detailed analysis of temperature effects on the isotope fractionation indicates that stomatal aperture decreases with increasing temperature and carboxylation capacity increases. For B. tubiflorum, the temperature dependence of the isotope fractionation is smaller and is principally attributed to the normal temperature dependences of the rates of diffusion and carboxylation steps. The small change in the isotopic composition of remaining malic acid in both species which is observed during deacidification indicates that malate release, rather than decarboxylation, is rate limiting in the deacidification process.     

Diffusional Contribution to Carbon Isotope Fractionation during Dark CO(2) Fixation in CAM Plants

A mathematical model is developed which can be used to predict in vivo carbon isotope fractionations associated with carbon fixation in plants in terms of diffusion, CO₂ hydration, and carboxylation components. This model also permits calculation of internal CO₂ concentration for comparison with results of gas-exchange experiments. The isotope fractionations associated with carbon fixation in Kalanchoë daigremontiana and Bryophyllum tubiflorum have been measured by isolation of malic acid following dark fixation and enzymic determination of the isotopic composition of carbon-4 of this material. Corrections are made for residual malic acid, fumarase activity, and respiration. Comparison of these data with calculations from the model indicates that the rate of carbon fixation is limited principally by diffusion, rather than by carboxylation. Processes subsequent to the initial carboxylation also contribute to the over-all isotopic composition of the plant.     

Hydrogen and carbon isotopic fractionations of lipid biosynthesis among terrestrial (C3, C4 and CAM) and aquatic plants

Compound-specific hydrogen and carbon isotopic compositions in n-alkanoic acids, phytol and sterols were determined for various plant classes (terrestrial C3-angiosperm; C3-gymnosperm; C4; crassulacean acid metabolism (CAM); and aquatic C3 plants) in order to investigate isotopic fractionations among various plant classes. In all plants, lipid biomolecules are depleted in both D (up to 324 per thousand ) and 13C (up to 14.7 per thousand ) relative to ambient water and bulk tissue, respectively. In addition, the magnitude of D- and 13C-depletion of lipid biomolecules is distinctive depending on plant classes. For example, C3 angiosperm n-alkanoic acids are less depleted in D (95+/-23 per thousand ) and 13C (4.3 +/- 2.5 per thousand ) relative to ambient water and bulk tissue, respectively, while C4 plant n-alkanoic acids are more depleted in D (119 +/- 15 per thousand ) and 13C (10.2 +/- 2.0 per thousand ). On the other hand, C3 angiosperm phytol and sterols are much more depleted in D (306 +/-12 per thousand for phytol, 211+/-15 per thousand for sterol) with less depletion in 13C (4.1 +/- 1.1 per thousand for phytol, 1.3 +/- 0.9 per thousand for sterol) relative to ambient water and bulk tissue, respectively, while C4 plant phytol and sterols are less depleted in D (254 +/- 7 per thousand for phytol, 186 +/- 13 per thousand for sterols) with much more depletion in 13C (9.0 +/- 1.2 per thousand for phytol, 5.0 +/- 1.1 per thousand for sterols). Among various plant classes, there is a positive correlation between the D- and 13C-depletion for n-alkanoic acids, while a negative correlation was found for phytol and sterols from the same plants.     

Oxygen and Hydrogen Stable Isotope Ratios of Bulk Needles Reveal the Geographic Origin of Norway Spruce in the European Alps

Background

Tracking timber is necessary in order to prevent illegal logging and protect local timber production, but there is as yet no suitable analytical traceability method. Stable isotope ratios in plants are known to reflect geographical variations. In this study we analysed four stable isotope ratios in order to develop a model able to identify the geographic origin of Norway spruce in the European Alps.

Methodology and Principal Findings

δ¹⁸O, δ²H, δ¹³C and δ¹⁵N were measured in bulk needles of Picea abies sampled in 20 sites in and around the European Alps. Environmental and spatial variables were found to be related to the measured isotope ratios. An ordinary least squares regression was used to identify the most important factor in stable isotope variability in bulk needles. Spatial autocorrelation was tested for all isotope ratios by means of Moran’s I. δ¹⁸O, δ²H and δ¹⁵N values differed significantly between sites. Distance from the coast had the greatest influence on δ²H, while latitude and longitude were strongly related to δ¹⁸O. δ¹³C values did not appear to have any relationship with geographical position, while δ¹⁵N values were influenced by distance from the motorway. The regression model improved the explanatory power of the spatial and environmental variables. Positive spatial autocorrelations were found for δ¹⁸O and δ²H values.

Conclusions

The δ ¹⁸O, δ²H and δ¹⁵N values in P. abies bulk needles are a suitable proxy to identify geographic origin as they vary according to geographical position. Although the regression model showed the explanatory variables to have significant power and stability, we conclude that our model might be improved by multivariate spatial interpolation of the δ ¹⁸O and δ²H values.     

Polypeptide Composition of Envelope Membranes Isolated from Chloroplasts of C3, C4, and CAM Plants

Chloroplast envelopes were isolated from chloroplasts purifiedfrom Spinacea oleracea L. (C₃), Panicum miliaceum L. (NAD-malicenzyme-type C₁), Digitaria sanguinalis (L.) Scop. (NADP-malicenzyme-type C₄), Kalanchoe daigremontiana Hamet et Perrier (constitutiveCAM), and from Mesembryanthemum crystallinum L. (inducible CAM)performing either C₃ photosynthesis or Crassulacean acid metabolism(CAM). For each species, methods were developed to isolate chloroplastenvelopes free of thylakoid contamination. The polypeptidesof ribulose bisphosphate (RuBP) carboxylase which has been consistentlyreported in envelope preparations of spinach were not foundin envelope preparations of C₄ mesophyll chloroplasts. Silverstaining of envelope polypeptides resolved electrophoreticallyon sodium dodecylsulfate polyacrylamide gradient slab gels produceda more complex profile than did Coomassie staining which haspreviously been used with C₃ envelope preparations, even thoughsilver reacted poorly with polypeptides corresponding to thesubunits of RuBP carboxylase. All of the plants examined possesseda major polypeptide of 27 to 29 kilodaltons (kD) which was previouslysuggested to be the phosphate translocator in spinach. WithC₃ M. crystallinum, the 29 kD polypeptide stained most intensely.After induction of CAM, a 32 kD polypeptide also stained intensely,giving a profile similar to that obtained with the constitutiveCAM species. A 32 kD polypeptide was also prominent in C₄ envelopepreparations, suggesting that a 32 kD polypeptide may be a translocatorprotein which is required in Crassulacean acid metabolism andC4 photosynthesis, but not in C₃ photosynthesis. (Received April 25, 1983; Accepted July 9, 1983)     

C4光合关键酶基因转化C3植物

文章介绍C4光合关键酶和转运蛋白基因转化C3植物的研究进展。     

A Comparison of Dark Respiration between C(3) and C(4) Plants

Lower respiratory costs were hypothesized as providing an additional benefit in C₄ plants compared to C₃ plants due to less investment in proteins in C₄ leaves. Therefore, photosynthesis and dark respiration of mature leaves were compared between a number of C₄ and C₃ species. Although photosynthetic rates were generally greater in C₄ when compared to C₃ species, no differences were found in dark respiration rates of individual leaves at either the beginning or after 16 h of the dark period. The effects of nitrogen on photosynthesis and respiration of individual leaves and whole plants were also investigated in two species that occupy similar habitats, Amaranthus retroflexus (C₄) and Chenopodium album (C₃). For mature leaves of both species, there was no relationship between leaf nitrogen and leaf respiration, with leaves of both species exhibiting a similar rate of decline after 16 h of darkness. In contrast, leaf photosynthesis increased with increasing leaf nitrogen in both species, with the C₄ species displaying a greater photosynthetic response to leaf nitrogen. For whole plants of both species grown at different nitrogen levels, there was a clear linear relationship between net CO₂ uptake and CO₂ efflux in the dark. The dependence of nightly CO₂ efflux on CO₂ uptake was similar for both species, although the response of CO₂ uptake to leaf nitrogen was much steeper in the C₄ species, Amaranthus retroflexus. Rates of growth and maintenance respiration by whole plants of both species were similar, with both species displaying higher rates at higher leaf nitrogen. There were no significant differences in leaf or whole plant maintenance respiration between species at any temperature between 18 and 42°C. The data suggest no obvious differences in respiratory costs in C₄ and C₃ plants.     

Photosynthetic Characteristics of Segregates from Hybrids between Flaveria brownii (C4 Like) and Flaveria linearis (C3-C4)

Characteristics related to C4 photosynthesis were studied in reciprocal F1 hybrids and F2 plants from Flaveria brownii (C4 like) and Flaveria linearis (C3-C4). The reciprocal F1 plants differed in 13C/12C ratios of leaves and the percentage of 14C initially incorporated into C4 acids, being more like the pollen parents in these traits. They did not differ in apparent photosynthesis or in O2 inhibition of apparent photosynthesis and differed only slightly in CO2 compensation concentration at 175 [mu]mol quanta m-2 s-1 and 400 mL L-1 O2. The 13C/12C ratios of 78 F2 progeny from the two F1 plants exhibited a normal distribution centered between those of the parents, with a few values slightly higher and lower than the parents. Apparent photosynthesis at 130 [mu]L L-1 CO2 and inhibition of photosynthesis by O2 was nearly normally distributed in the F2 population, but no values for F2 plants approached those for F. brownii (15.4 [mu]mol m-2 s-1 and 7.8%, respectively). Distribution of the CO2 compensation concentration measured at 1000 [mu]mol quanta m-2 s-1 and 400 mL L-1 of O2 in the F2 population was skewed toward F. brownii with 72% of the progeny having values <9 [mu]L of CO2 L-1 compared to 1.5 and 27.2 [mu]L L-1 for F. brownii and F. linearis, respectively. Correlations among traits of F2 plants were low (coefficients of 0.30 to -0.49), indicating that the C4- related traits are not closely linked in segregating populations. Plants in the F2 population selected for high or low apparent photosynthesis at 130 [mu]L of CO2 L-1 (six each) did not rank consistently high or low for 13C/12C ratios, O2 inhibition of apparent photosynthesis, CO2 compensation concentration, or activities of phosphoenolpyruvate carboxylase or NADP-malic enzyme. This study confirms results of earlier work that indicates independent segregation of C4 traits and also shows that the C4-like parental type can be recovered, at least for some characteristics (13C/12C ratio), in segregating populations. Recovery of fully functional C4 plants awaits further experimentation with C4 x C3 or C4 x C3-C4 hybrid plants that produce fertile progeny.     

Multielement Isotope Ratios of Vegetables from Integrated and Organic Production

In this paper we discuss the use of isotope ratios as indicators of organic production. Few studies have investigated the influence of plant nutrition on the isotopic signatures of plants. As plant nutrition is often significantly different between integrated and organic production systems the isotope ratios in the plants may reflect this. Plant samples from a 2-year field-experiment were analyzed for ¹⁵N, ¹³C and ³⁴S content of the bulk-material and ¹⁸O-content of the leaf water. In this experiment cabbages (Brassica oleracea v. capitata f. alba cv. Rolly), onions (Allium cepa cv. Alisa Craig), lettuces (Lactuca sativa v. capitata cv. Ponchito) and Chinese cabbage (Brassica pekinesis cv. Parkin) were cultivated according to good agricultural practices for integrated and organic production. No differences in the δ ³⁴S and δ ¹⁸O values of the plants grown under the two production systems were observed. The organically produced vegetables were significantly enriched in ¹⁵N and depleted in ¹³C compared to those grown under the integrated system.     

Phosphoenolpyruvate carboxylase genes in C3, crassulacean acid metabolism (CAM) and C3/CAM intermediate species of the genus Clusia: rapid reversible C3/CAM switches are based on the C3 housekeeping gene

The genus Clusia includes species that exhibit either the C3 or crassulacean acid metabolism (CAM) mode of photosynthesis, or those that are able to switch between both modes according to water availability. In order to screen for species-specific genetic variability, we investigated the key carboxylase for CAM, phosphoenolpyruvate carboxylase (PEPC). Sequence analysis of DNA isolated from the obligate CAM species, Clusia hilariana, the obligate C3 species, Clusia multiflora, and an intermediate species that can switch between C3 and CAM photosynthesis, Clusia minor, revealed three different isoforms for C. hilariana and one each for the other two species. Sequence alignments indicated that PEPC from the intermediate species had high homology with the C3 protein and with one of CAM plant proteins. These were assumed to constitute 'housekeeping' proteins, which can also support CAM in intermediate species. The other two isoforms of the CAM plant C. hilariana were either CAM-specific or showed homologies with PEPC from roots. Phylogenetic trees derived from neighbour-joining analysis of amino acid sequences from 13 different Clusia species resulted in two distinct groups of plants with either 'housekeeping' PEPC only, or additionally CAM-related isoforms. Only C. hilariana showed the third, probably root-specific isoform. The high homology of the PEPC from the intermediate species with the C3 protein indicates that for the reversible transition from the C3 to CAM mode of photosynthesis, the C3 type of PEPC is sufficient. Its expression, however, is strongly increased under CAM-inducing conditions. The use of the C3 isoform could have facilitated the evolution of CAM within the genus, which occurred independently for several times.     

Oxygen and Hydrogen Isotope Fractionation during Cellulose Metabolism in Lemna gibba L

Lemna gibba L. B3 was grown under heterotrophic, photoheterotrophic, and autotrophic conditions in water having a variety of hydrogen and oxygen isotopic compositions. The slopes of the linear regression lines between the isotopic composition of water and leaf cellulose indicated that under the three growth conditions about 40, 70, and 100% of oxygens and carbon-bound hydrogens of cellulose exchanged with those of water prior to cellulose formation. Using the equations of the linear relationships, we estimated the overall fractionation factors between water and the exchanged oxygen and carbon bound-hydrogen of cellulose. At least two very different isotope effects must determine the hydrogen isotopic composition of Lemna cellulose. One reflects the photosynthetic reduction of NADP, while the second reflects exchange reactions that occur subsequent to NADP reduction. Oxygen isotopic composition of cellulose apparently is determined by a single type of exchange reaction with water. Under different growth conditions, variations in metabolic fluxes affect the hydrogen isotopic composition of cellulose by influencing the extent to which the two isotope effects mentioned above are recorded. The oxygen isotopic composition of cellulose is not affected by such changes in growth conditions.     

Carbon and Hydrogen Isotope Ratios in Bromeliads Growing under Different Light Environments in Natural Conditions

Wild Ananas species in northern South America occur in shady environments and appear to be relatively intolerant to droughts associated with growth under full sun exposure. This behaviour contrasts with the higher productivity of commercial varieties of Ananas comosus when grown under full sun exposure. Such differentiation within the genus offers an opportunity to study the process of adaptation of apparently high light avoiding species into true sun plants. As a first approximation, the analysis of nitrogen content and carbon and hydrogen isotope ratios of bromeliads growing under natural conditions was undertaken to test the following hypotheses: 1. Leaf nitrogen content of plants grown under partial shade is higher than that of the same species in the same habitat growing under full sun exposure, due to the higher availability to nitrogen in the under-canopy, but also to the lower proportion of structural carbohydrates in shade leaves; 2. δ¹³C values are usually more negative in CAM bromeliads growing under partial shade because of the lower contribution of CAM to total carbon gain, and the probable fixation of CO₂ originating from soil respiration; 3. δD values of CAM bromeliads are less negative than those of C3 bromeliads, but CAM bromeliads grown in shady habitats tend to have more negative δD values because of the lower relative accumulation of deuterium in leaf tissue water, and also because of their relatively lower CAM activity. The results show a clear differentiation between CAM and C3 bromeliads based on δ¹³C values, and in general δD values are less negative in CAM bromeliads. However, in several species overlapping δD values between C3 and CAM bromeliads were observed. The analysis of paired samples of the same species grown under contrasting light intensity usually conformed with the expectations. A number of deviations from the hypotheses were observed which appeared to be related to particular environmental conditions. The interpretation of δD values obtained from total organic matter is made difficult by the local variation of hydrogen/deuterium ratios in water available to the plant.     

The Carbon Isotope Ratios and Centents of Mineral Elements in Leaves of Chinese Medi cinal Plants

Leaf carbon isotope ratios and 13 kinds of mineral elements were measured on 36 species of common Chinese medicinal plants in a subtropical monsoon forest of Ding Hu Shan in Guangdong Province. The δ13C value were from –26.4 to –32.6%, indicating that all of the species belonged the photosynthetic C3 types. The relative lower value of δ13C was observed in the life form of shrubs. The contents of 7 elements (N, P, K, Ca, Na Mg, Si) were dependent upon the species, life form, medicinal function and medicinal part. Herb type medicine and the used medicinal part of leaves or whole plant showed higher levels of above elements than the others. Among the nine groups with different medicinal functions, it was found that more nitrogen was in the leaves of medicinal plants for hemophthisis, hypertension and stomachic troubles, more phosphorus and potassium were in the leaves for cancer and snake bite medicines, but more calcium and magnesium were in the leaves for curing rheumatics. Ferric, aluminium and manganese were the main composition of microelements in leaves. There were higher content of ferric in leaves for hemophthisis medicine, higher zinc in leaves for cold and hypertension medicine, and higer cuprum in leaves of stomachic medicine. It was suggested that the pattern of mineral elements in leaves of Chinese medicinal plants reflected the different properties of absorption and accumulation. Some additional effect due to the high content of certain element might be associated with the main function of that medicine.     

Photosynthetic Characteristics of the C(3)-C(4) Intermediate Parthenium hysterophorus

The weedy species Parthenium hysterophorus (Asteraceae) possesses a Kranz-like leaf anatomy. The bundle sheath cells are thick-walled and contain numerous granal chloroplasts, prominent mitochondria, and peroxisomes, all largely arranged in a centripetal position. Both mesophyll and bundle sheath chloroplasts accumulate starch. P. hysterophorus exhibits reduced photorespiration as indicated by a moderately low CO₂ compensation concentration (20-25 microliters per liter at 30°C and 21% O₂) and by a reduced sensitivity of net photosynthesis to 21% O₂. In contrast, the related C₃ species P. incanum and P. argentatum (guayule) lack Kranz anatomy, have higher CO₂ compensation concentrations (about 55 microliters per liter), and show a greater inhibition of photosynthesis by 21% O₂. Furthermore, in P. hysterophorus the CO₂ compensation concentration is relatively less sensitive to changes in O₂ concentrations and shows a biphasic response to changing O₂, with a transition point at about 11% O₂. Based on these results, P. hysterophorus is classified as a C₃-C₄ intermediate. The activities of diagnostic enzymes of C₄ photosynthesis in P. hysterophorus were very low, comparable to those observed in the C₃ species P. incanum (e.g. phosphoenolpyruvate carboxylase activity of 10-29 micromoles per milligram of chlorophyll per hour). Exposures of leaves of each species to ¹⁴CO₂ (for 8 seconds) in the light resulted in 3-phosphoglycerate and sugar phosphates being the predominant initial ¹⁴C products (77-84%), with ≤4% of the ¹⁴C-label in malate plus aspartate. These results indicate that in the C₃-C₄ intermediate P. hysterophorus, the reduction in leaf photorespiration cannot be attributed to C₄ photosynthesis.     

Unsolved Problems of Carbonic Anhydrases Functioning in Photosynthetic Cells of Higher C3 Plants

Biochemistry (Moscow) - The review presents current data on carbonic anhydrases found in various compartments of photosynthetic cells of higher plants. The available data on expression of genes...