Oxygen and Hydrogen Isotope Ratios of Water from Photosynthetic Tissues of CAM and C(3) Plants

Water samples from photosynthetic tissues of C₃ and Crassulacean acid metabolism (CAM) plants that grew together in the field were extracted and the stable oxygen and hydrogen isotope ratios determined. During the day, ¹⁸O/¹⁶O and deuterium/hydrogen (D/H) ratios of water from CAM plants were lower than those observed in water from C₃ plants. The patterns of diurnal variation (or lack thereof) in isotope ratios of plant water are consistent with the gross anatomical and physiological characteristics of the plants studied here. Our observations support the previously advanced hypothesis that high D/H ratios in cellulose nitrate prepared from CAM plants relative to those for C₃ plants are not caused by greater deuterium enrichment in the water in CAM plants, but rather by isotopic fractionations associated with different biochemical reactions in the two types of plants.     

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.     

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.     

兼性CAM植物长叶景天叶片在C3和CAM型时的光氧化作用

通过停止浇水产生水分胁迫使兼性CAM植物长叶景天(Sedum spectabile Boreau)叶片光合途径由C3型转为CAM型.干旱15 d时观察到典型的CAM生理特征,且叶片的δ13C值与含水量成线性相关.水分胁迫改变了叶绿素荧光参数和抗氧化能力,ΦPSⅡ和qP降低50%和34%,NPQ提高约180%,SOD活性和清除DPPH@ 自由基能力也明显下降, 但膜半透性变化不大.当将处于C3(浇水)和诱导为CAM(缺水)型的叶圆片用外源甲基紫精(MV)和强光作光氧化处理后,与C3型叶片相比,诱导CAM型叶片的NPQ不能提高,qP和ΦPSⅡ降至很低水平,光系统处于高还原态,光能供给与消耗失衡(1-qP=0.86和(1-qP)/NPQ>1),膜系统几乎失去完整性.这种严重的光氧化损伤表明,与我们以前报告的专性CAM植物不同,以兼性CAM植物诱导表达的CAM型未能显示比C3型较强的耐光氧化优势.讨论了出现这种光氧化敏感性差别的可能原因.     

Epidermal Micromorphology and Mesophyll Structure of Populus euphratica Heteromorphic Leaves at Different Development Stages

Leaf epidermal micromorphology and mesophyll structure during the development of Populus euphratica heteromorphic leaves, including linear, lanceolate, ovate, dentate ovate, dentate rhombic, dentate broad-ovate and dentate fan-shaped leaves, were studied by using electron and light microscopy. During development of heteromorphic leaves, epidermal appendages (wax crystals and trichomes) and special cells (mucilage cells and crystal idioblasts) increased in all leaf types while chloroplast ultrastructure and stomatal characters show maximum photosynthetic activity in dentate ovate and rhombic leaves. Also, functional analysis by subordinate function values shows that the maximum adaptability to adverse stress was exhibited in the broad type of mature leaves. The 12 heteromorphic leaf types are classified into three major groups by hierarchical cluster analysis: young, developing and mature leaves. Mature leaves can effectively obtain the highest stress resistance by combining the protection of xerophytic anatomy from drought stress, regulation of water uptake in micro-environment by mucilage and crystal idioblasts, and assistant defense of transpiration reduction through leaf epidermal appendages, which improves photosynthetic activity under arid desert conditions. Our data confirms that the main leaf function is differentiated during the developing process of heteromorphic leaves.     

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.     

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)     

Stable Carbon Isotope Ratios of Lipid Biomarkers of Sulfate-Reducing Bacteria

We examined the potential use of natural-abundance stable carbon isotope ratios of lipids for determining substrate usage by sulfate-reducing bacteria (SRB). Four SRB were grown under autotrophic, mixotrophic, or heterotrophic growth conditions, and the δ¹³C values of their individual fatty acids (FA) were determined. The FA were usually ¹³C depleted in relation to biomass, with Δδ¹³C_{(FA − biomass)} of −4 to −17‰; the greatest depletion occurred during heterotrophic growth. The exception was Desulfotomaculum acetoxidans, for which substrate limitation resulted in biomass and FA becoming isotopically heavier than the acetate substrate. The δ¹³C values of FA in Desulfotomaculum acetoxidans varied with the position of the double bond in the monounsaturated C₁₆ and C₁₈ FA, with FA becoming progressively more ¹³C depleted as the double bond approached the methyl end. Mixotrophic growth of Desulfovibrio desulfuricans resulted in little depletion of the i17:1 biomarker relative to biomass or acetate, whereas growth with lactate resulted in a higher proportion of i17:1 with a greater depletion in ¹³C. The relative abundances of 10Me16:0 in Desulfobacter hydrogenophilus and Desulfobacterium autotrophicum were not affected by growth conditions, yet the Δδ¹³C_{(FA − substrate)} values of 10Me16:0 were considerably greater during autotrophic growth. These experiments indicate that FA δ¹³C values can be useful for interpreting carbon utilization by SRB in natural environments.     

Isolation of Mesophyll Cells from Sedum telephium Leaves

A technique is described for mechanically isolating metabolically active individual spongy mesophyll cells from the Crassulacean acid metabolism plant, Sedum telephium. Mature leaves are selected at about 2 PM when acidity is low, and three different media are used to reduce the problem of leaf acidity and to maintain isotonic conditions. The upper and lower epidermis is peeled from chilled leaves and the leaves are suspended in a buffered “soaking medium,” then gently ground with a mortar and pestle. Cells and debris are separated using a “washing medium,” with cells being filtered through a 136 micron net and collected on an 80 micron net. Cells then are suspended in a “cell suspension medium” and concentrated by centrifugation. Approximately 2 hours are required for the isolation procedure, and activity in CO₂ fixation is constant for up to 4 hours after isolation. Microscopic examination shows about 65% of the isolated cells appear intact and unplasmolyzed and are similar to leaf msophyll cells. The yield of cells on a leaf chlorophyll basis is about 1%. A light micrograph of the isolated cells is given.     

Effects of Light Quality on Photosynthetic Carbon Metabolism in C4 and C3 Plants: Rapid Movements of Photosynthetic Intermediates Between Mesophyll and Bundle Sheath Cells

The influence of varying light intensity and quality on thecarbon labelling patterns in Rumex vesicarius (a C₃ plant),Setaria italica (a malate-formingC₄ plant), and Amaranthus paniculatus(an aspartate-forming C₄ plant) was studied. In A. paniculatusand B. vesicarius blue light decreased the transfer of radioactivityto sugars and starch but in S. italica only slightly decreasedradioactivity in sugar phosphates, sucrose, and insolubles.Negligible transfer was observed from the C₄ acids to sugarphosphates, sucrose, and starch under dim blue-green and blue-yellowlights in S. italica and A. paniculatus. Blue light favouredthe formation of malate, aspartate, and alanine in all threeplants. The differential effect of blue and red light suggesteda variation in the mechanisms of C₄-photosynthesis in Setariaand Amaranthus. Leaves of S. italica and A. paniculatus were allowed to photosynthesizein ¹⁴CO₂ for 5 s and then the distribution of the labelled productsbetween the mesophyll and the bundle sheath cells was determinedduring subsequent photosynthesis in ¹²CO₂. Malate and aspartatewhich appeared initially in the mesophyll layer moved rapidlyinto the bundle sheath cells. Phosphoglyceric acid originatingin the bundle sheath moved swiftly to the mesophyll layer. Sugarphosphates were recovered from both the mesophyll and the bundlesheath cells. Most of the starch was found in the bundle sheathcells while sucrose and alanine were localized in the mesophyllcells.     

Relationships among Mercury Concentration,and Stable Isotope Ratios of Carbon and Nitrogen in the Scalp Hair of Residents from Seven Countries: Effects of Marine Fish and C4 Plants Consumption

We analyzed the Hg concentration, and δ¹³C and δ¹⁵N values in the scalp hair of residents from seven countries; Vietnam, New Zealand, Spain, the USA, South Korea, Brazil and Japan. Relationships among the data in each country and among the seven countries were then examined. The highest Hg concentration as well as the highest or higher δ¹⁵N value in each country was found in the hair of a heavy marine fish-eater, whereas the lowest Hg concentration and δ¹⁵N value were found in the hair of a vegetarian or non (marginal)-fish eater. Hg concentrations were positively correlated with the δ¹⁵N values in each country, and increased markedly in samples with δ¹⁵N values exceeding 9.0 ‰, probably due to fish consumption. The highest Hg concentration could be found in sample, with a δ¹³C value between -19 and -18‰, probably reflecting the δ¹³C value of the marine food web.     

Photosynthetic Characteristics of the Palisade Mesophyll and Spongy Mesophyll in the CAM/C4 Intermediate Plant,Peperomia camptotricha*

Several photosynthetic parameters were examined in the different tissue layers of leaves from the recently characterized crassulacean acid metabolism/C₄ intermediate plant, Peperomia camptotricha (Nishio and Ting, 1987). Light appears to control the development of certain photosynthetic characteristics within the tissue layers, while factors other than light seem to dictate others. Analysis of the chlorophyll content (including P700) and chlorophyll-proteins indicated that more light harvesting chlorophylls were associated with reaction centers in the tissues that were shaded by overlying tissue. Electron transport activity and chlorophyll-protein analysis indicated that the ratio of photosystem I to photosystem II in the spongy mesophyll (the abaxial tissue type and hence most shaded) was elevated relative to the overlying chlorophyll-rich median palisade mesophyll. The elevated photosystem I relative to photosystem II in the spongy mesophyll of Peperomia camptotricha may be related to C₄ metabolism and an increased requirement for ATP.     

Plants Regenerated from Mesophyll Protoplasts of Cottonwood New Clone

Poplar NL-80106 (Populus deltoides×P, simonii) mesophyll protoplasts were isolated from leaves of 30 days-old sterile shoot, with 4 × 107/g fr. wt of protoplast yield after purification. The protoplasts were cultured in KM8p and MS liquid media containing 2 mg/L 2, 4-D, 0. 5 mg/L NAA and 0.5 mg/L KT. Higher plating density and lower osmatic pressure (0.45 mol/L) were proved to be favourable to division of protoplast-derived cells. The first division initiated 5 days after culture, and the division frequency reached 4.5 % on the 10th day. A number'of cell colonies and microcalli was formed in 12 weeks. Using organic nitrates and glucose in protoplast culture medium was beneficial to increase division frequency and plating efficiency. The calli were allowed to grow to 4--6 mm in height with red colour and compact structure on the gelrite-sohdified NLZ1 proliferation medium in 3 weeks and were transferred onto NLF differentiation medium where the frequency of shoot formation could reach 100%. The 3 cm high shoots were then cut off from the callus and rooted on 1/2 MS medium.     

Proton/Pyruvate Cotransport into Mesophyll Chloroplasts of C4 Plants

Light-enhanced active pyruvate uptake into mesophyll chloroplastsof C₄ plants was reported to be mimicked by either of the twotypes of cation jump: H⁺-jump in maize and phylogenically relatedspecies (H⁺-type) and Na⁺-jump in all the other C₄ species tested(Na⁺-type) [Aoki, N., Ohnishi, J. and Kanai, R. (1992) PlantCell Physiol. 33: 805]. In this study, medium and stromal pH was monitored in the suspensionof C₄ mesophyll chloroplasts. Medium alkalization lasting for5 to 10 seconds after pyruvate addition was detected by a pHelectrode and observed only in the light and only in mesophyllchloroplasts from H⁺-type species, Zea mays L. and Coix lacryma-jobiL., but not in those from Na⁺-type species Panicum miliaceumL., Setaria italica (L.) Beauv. and Panicum maximum Jacq. Theinitial rate of H⁺ consumption showed good correlation with[¹⁴C]pyruvate uptake measured by silicone oil filtering centrifugation,both being inhibited by N-ethylmaleimide and 7-chloro-4-nitrobenzo-2-oxa-l,3-diazole to the same degree. The ratio of the rate of H⁺ uptaketo that of pyruvate uptake was always about 1. Pyruvate-inducedacidification of the stroma was observed in maize mesophyllchloroplasts. These results show one to one cotransport of H⁺and pyruvate anion into mesophyll chloroplasts of H⁺-type C₄species in the light. (Received January 5, 1994; Accepted May 6, 1994)     

Distribution of Nitrate-assimilating Enzymes between Mesophyll Protoplasts and Bundle Sheath Cells in Leaves of Three Groups of C(4) Plants

Intercellular distribution of enzymes involved in amino nitrogen synthesis was studied in leaves of species representing three C₄ groups, i.e. Sorghum bicolor, Zea mays, Digitaria sanguinalis (NADP malic enzyme type); Panicum miliaceum (NAD malic enzyme type); and Panicum maximum (phosphoenolpyruvate carboxykinase type). Nitrate reductase, nitrite reductase, glutamine synthetase, and glutamate synthase were predominantly localized in mesophyll cells of all the species, except in P. maximum where nitrite reductase had similar activity on a chlorophyll basis, in both mesophyll and bundle sheath cells. NADH-glutamate dehydrogenase was concentrated in the bundle sheath cells, while NADPH-glutamate dehydrogenase was localized in both mesophyll and bundle sheath cells. The activities of nitrate-assimilating enzymes, except for nitrate reductase, were high enough to account for the proposed in vivo rates of nitrate assimilation.     

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.     

Effect of Wound Shock on Protein Synthesis in Leaves of C3 and C4 Plants

Wound stress causes an initial decrease in the uptake and subsequentincorporation of radiolabelled methionine in leaves of C₃ (peanutand soybean) and C₄ (sorghum and maize) plants. A four-hourincubation of excised leaves at 25C permits them to recoverfrom wounding and thereby facilitates monitoring of changesin protein synthesis caused by heat shock or other types ofenvironmental stress. (Received December 7, 1992; Accepted February 16, 1993)     

Isolation of Mesophyll Cells from Mulberry Leaves for Photosynthetic Studies

A procedure was developed for isolating mesophyll cells frommulberry leaves that have consistently high photosynthetic activity.To obtain photosynthetically active cells from some plants,their petioles, immersed in water, had to be kept in the darkfor 3–4 days prior to cell isolation. (Received May 6, 1983; Accepted August 15, 1983)     

中国北方干湿气候区C_3草本植物δ~（13）C值及其与湿润指数的关系

通过对中国北方C3草本植物稳定性碳同位素的测定以及有关该区植被碳同位素资料的收集,共获取了47个样点的地理位置、气候因子和325个植物样品的碳同位素数据;计算了中国北方不同气候分区的湿润指数,分析了C3草本植物δ13C值的空间特征以及与湿润指数等环境因子之间的关系。在所调查的范围内,中国北方地区C3草本植物δ13C值的分布区间为-29.9‰--25.4‰,平均值为-27.3‰。C3草本植物δ13C的平均值从半湿润地区到半干旱地区再到干旱地区显著变重;3个气候分区植物δ13C值的变化范围分别是-29.9‰--26.7‰（半湿润区）、-28.4‰--25.6‰（半干旱区）和-28.0‰--25.4‰（干旱区）。一元回归分析表明,各气候分区C3草本植物δ13C值与湿润指数的关系存在差异,在半干旱区、半湿润区和整个北方地区,C3草本植物δ13C值与湿润指数均呈显著线性负相关（P〈0.05）,随着湿润指数的增加,C3植物δ13C平均值均变轻,但下降幅度不同。而在北方干旱气候区内,C3草本植物δ13C与湿润指数呈显著正相关（P〈0.05）,湿润指数每升高0.1,植物δ13C平均值增加1.3‰。年均温度可能是决定该区内各样点湿润指数和C3植物对13C分馏能力差别的主要原因。