PAIN with and without PAR: variants for third-spin assisted heteronuclear polarization transfer

In this article, we describe third-spin assisted heteronuclear recoupling experiments, which play an increasingly important role in measuring long-range heteronuclear couplings, in particular ¹⁵N–¹³C, in proteins. In the proton-assisted insensitive nuclei cross polarization (PAIN-CP) experiment (de Paëpe et al. in J Chem Phys 134:095101, 2011), heteronuclear polarization transfer is always accompanied by homonuclear transfer of the proton-assisted recoupling (PAR) type. We present a phase-alternating experiment that promotes heteronuclear (e.g. ¹⁵N → ¹³C) polarization transfer while simultaneously minimizing homonuclear (e.g.¹³C → ¹³C) transfer (PAIN without PAR). This minimization of homonuclear polarization transfer is based on the principle of the resonant second-order transfer (RESORT) recoupling scheme where the passive proton spins are irradiated by a phase-alternating sequence and the modulation frequency is matched to an integer multiple of the spinning frequency. The similarities and differences between the PAIN-CP and this het-RESORT experiment are discussed here.     

Model parameterization to simulate and compare the PAR absorption potential of two competing plant species

Mountain pastures dominated by the pasture grass Setaria sphacelata in the Andes of southern Ecuador are heavily infested by southern bracken (Pteridium arachnoideum), a major problem for pasture management. Field observations suggest that bracken might outcompete the grass due to its competitive strength with regard to the absorption of photosynthetically active radiation (PAR). To understand the PAR absorption potential of both species, the aims of the current paper are to (1) parameterize a radiation scheme of a two-big-leaf model by deriving structural (LAI, leaf angle parameter) and optical (leaf albedo, transmittance) plant traits for average individuals from field surveys, (2) to initialize the properly parameterized radiation scheme with realistic global irradiation conditions of the Rio San Francisco Valley in the Andes of southern Ecuador, and (3) to compare the PAR absorption capabilities of both species under typical local weather conditions. Field data show that bracken reveals a slightly higher average leaf area index (LAI) and more horizontally oriented leaves in comparison to Setaria. Spectrometer measurements reveal that bracken and Setaria are characterized by a similar average leaf absorptance. Simulations with the average diurnal course of incoming solar radiation (1998–2005) and the mean leaf–sun geometry reveal that PAR absorption is fairly equal for both species. However, the comparison of typical clear and overcast days show that two parameters, (1) the relation of incoming diffuse and direct irradiance, and (2) the leaf–sun geometry play a major role for PAR absorption in the two-big-leaf approach: Under cloudy sky conditions (mainly diffuse irradiance), PAR absorption is slightly higher for Setaria while under clear sky conditions (mainly direct irradiance), the average bracken individual is characterized by a higher PAR absorption potential. (∼74 MJ m⁻² year⁻¹). The latter situation which occurs if the maximum daily irradiance exceeds 615 W m⁻² is mainly due to the nearly orthogonal incidence of the direct solar beam onto the horizontally oriented frond area which implies a high amount of direct PAR absorption during the noon maximum of direct irradiance. Such situations of solar irradiance favoring a higher PAR absorptance of bracken occur in ∼36% of the observation period (1998–2005). By considering the annual course of PAR irradiance in the San Francisco Valley, the clear advantage of bracken on clear days (36% of all days) is completely compensated by the slight but more frequent advantage of Setaria under overcast conditions (64% of all days). This means that neither bracken nor Setaria show a distinct advantage in PAR absorption capability under the current climatic conditions of the study area.     

Net primary productivity estimation and its relationship with tree diversity for tropical dry deciduous forests of central India

Present study aims at estimation and validation of net primary productivity (NPP) using production efficiency model (PEM), and its possible relationship with tree diversity. The PEM estimates NPP, based on light use efficiency (LUE) and intercepted photosynthetically active radiation (IPAR). Weighted average LUE varied between 0.02 gC/μmol/m² of PAR (Mixed forest (miscellaneous)) to 0.08 gC/μmol/m² of PAR (Acacia forest), in growing phase (GP), and 0.0008 gC/μmol/m² of PAR (Boswellia mixed forest) to 0.023 gC/μmol/m² of PAR (Acacia forest) during the senescent phase (SP). The average weighted LUE for tropical dry and Moist deciduous forest (MDF) in GP were 0.05 gC/μmol/m² of PAR and 0.03 gC/μmol/m² of PAR, respectively. The average IPAR for different forest types was 2079.58 μmol/m²/s during GP and 1510.58 μmol/m²/s during SP. The PEM based NPP varied between 0.58–275.78 gC/m²/year during GP and 0.43–74.34 gC/m²/year during SP. The PEM based NPP and conventional (ground based) NPP were related with R ² of 0.55. The tree diversity and NPP relationship was observed with R ² of 0.55 at the level of both plot and forest types.     

Developmental phase-dependent photosynthetic responses to ultraviolet-B radiation: damage, defence, and adaptation of primary leaves of wheat seedlings

Alterations in photosynthetic capacity of primary leaves of wheat seedlings in response to ultraviolet-B (UV-B; 280–320 nm; 60 μmol m⁻² s⁻¹) exposure alone and in combination with photosynthetically active radiation (PAR; 400–800 nm; 200 μmol m⁻² s⁻¹) during different phases of leaf growth and development were assessed. UV-B exposure resulted in a phase-dependent differential loss in photosynthetic pigments, photochemical potential, photosystem 2 (PS2) quantum yield, and in vivo O₂ evolution. UV-B exposure induced maximum damage to the photosynthetic apparatus during senescence phase of development. The damages were partially alleviated when UV-B exposure was accompanied by PAR. UV-B induced an enhancement in accumulation of flavonoids during all phases of development while it caused a decline in anthocyanin content during senescence. The differential changes in these parameters demonstrated the adaptation ability of leaves to UV-B stress during all phases of development and the ability was modified in UV-B+ PAR exposed samples.     

<Superscript>13</Superscript>C-detected NMR experiments for measuring chemical shifts and coupling constants in nucleic acid bases

The paper presents a set of two-dimensional experiments that utilize direct ¹³C detection to provide proton–carbon, carbon–carbon and carbon–nitrogen correlations in the bases of nucleic acids. The set includes a ¹³C-detected proton–carbon correlation experiment for the measurement of ¹³C–¹³C couplings, the CaCb experiment for correlating two quaternary carbons, the HCaCb experiment for the ¹³C–¹³C correlations in cases where one of the carbons has a proton attached, the HCC-TOCSY experiment for correlating a proton with a network of coupled carbons, and a ¹³C-detected ¹³C–¹⁵N correlation experiment for detecting the nitrogen nuclei that cannot be detected via protons. The IPAP procedure is used for extracting the carbon–carbon couplings and/or carbon decoupling in the direct dimension, while the S³E procedure is preferred in the indirect dimension of the carbon–nitrogen experiment to obtain the value of the coupling constant. The experiments supply accurate values of ¹³C and ¹⁵N chemical shifts and carbon–carbon and carbon–nitrogen coupling constants. These values can help to reveal structural features of nucleic acids either directly or via induced changes when the sample is dissolved in oriented media. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Measurement of imino <Superscript>1</Superscript>H–<Superscript>1</Superscript>H residual dipolar couplings in RNA

Imino ¹H–¹⁵N residual dipolar couplings (RDCs) provide additional structural information that complements standard ¹H–¹H NOEs leading to improvements in both the local and global structure of RNAs. Here, we report measurement of imino ¹H–¹H RDCs for the Iron Responsive Element (IRE) RNA and native E. coli tRNA^Val using a BEST-Jcomp-HMQC2 experiment. ¹H–¹H RDCs are observed between the imino protons in G–U wobble base pairs and between imino protons on neighboring base pairs in both RNAs. These imino ¹H–¹H RDCs complement standard ¹H–¹⁵N RDCs because the ¹H–¹H vectors generally point along the helical axis, roughly perpendicular to ¹H–¹⁵N RDCs. The use of longitudinal relaxation enhancement increased the signal-to-noise of the spectra by ~3.5-fold over the standard experiment. The ability to measure imino ¹H–¹H RDCs offers a new restraint, which can be used in NMR domain orientation and structural studies of RNAs. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Characteristics of net ecosystem carbon dioxide exchange (NEE) from August to October of Alpine meadow on the Tibetan Plateau, China

The Alpine meadow is one of the vegetation types widely distributed on the Tibetan Plateau in China with an area of about 1.2 million square kilometers. The Damxung rangeland station, located in the hinterland of the Tibetan Plateau, is covered with an typical vegetation. The continuous carbon flux data (from August to middle October, 2003) measured with the open-path eddy covariance system was used to analyze the diurnal variation pattern of net ecosystem carbon dioxide exchange (NEE) and its relationship with the environmental factors, such as photosynthetically active radiation (PAR), precipitation, and temperature. Results showed that NEE presented obvious diurnal variation pattern with single-peak of diurnal maximum carbon assimilation at 11: 00–12: 00 (local time) with an average of −0.268 mg CO₂·m⁻²·s⁻¹, i.e., −6.08 μmol CO₂·m⁻²·s⁻¹. During the daytime, NEE fitted fairly well with PAR in a rectangular hyperbola function with the apparent quantum yield (0.020 3 μmol CO₂ μmol⁻¹ PAR) and maximum ecosystem assimilation (9.741 1 μmol CO₂·m⁻²·s⁻¹). During the night-time, NEE showed a good exponential relation with the soil temperature at 5 cm depth. __________ Translated from Acta Ecologica Sinica 2005, 25(8): 1948–1952 [译自: 生态学报, 2005, 25(8): 1948–1952]     

Five decades of rapid forest spread in the Pieria Mountains (N. Greece) reconstructed by means of high-resolution pollen analysis and aerial photographs

The rapid spread of pine forests on the western side of the Pieria Mountains (N. Greece) began in 1945 and was facilitated by the recession of human activity and the siliceous geological substrate of the area. The pine expansion is visible not only in the vegetation maps for the years 1945, 1960 and 1992 but is also reflected in pollen data acquired from the high resolution analysis of a peat sequence covering the period 1945–1997, according to ²¹⁰Pb and ¹³⁷Cs dating. A possible inflation of the calculated PAR values due to discrepancies in the dating model is suspected and indirectly traced from the comparison of the Pinus mean PAR values for the most recent eight years of the monolith with those of a Tauber trap placed in the same mire. The presence of a high pollen producer such as pine enables the calculation of reliable PAR values, despite any discrepancies in the age-depth model. The Pinus PAR values “quantify” better the expansion of this timberline species, while pollen percentages reflect the same event in a “qualitative” way, i.e. as part of the general vegetation changes that take place in the area.     

SOFAST-HMQC Experiments for Recording Two-dimensional Deteronuclear Correlation Spectra of Proteins within a Few Seconds

Fast multidimensional NMR with a time resolution of a few seconds provides a new tool for high throughput screening and site-resolved real-time studies of kinetic molecular processes by NMR. Recently we have demonstrated the feasibility to record protein ¹H–¹⁵N correlation spectra in a few seconds of acquisition time using a new SOFAST-HMQC experiment (Schanda and Brutscher (2005) J. Am. Chem. Soc. 127, 8014). Here, we investigate in detail the performance of SOFAST-HMQC to record ¹H–¹⁵N and ¹H−¹³C correlation spectra of proteins of different size and at different magnetic field strengths. Compared to standard ¹H–¹⁵N correlation experiments SOFAST-HMQC provides a significant gain in sensitivity, especially for fast repetition rates. Guidelines are provided on how to set up SOFAST-HMQC experiments for a given protein sample. In addition, an alternative pulse scheme, IPAP-SOFAST-HMQC is presented that allows application on NMR spectrometers equipped with cryogenic probes, and fast measurement of one-bond ¹H–¹³C and ¹H–¹⁵N scalar and residual dipolar coupling constants.     

Spectral editing: selection of methyl groups in multidimensional solid-state magic-angle spinning NMR

A simple spectroscopic filtering technique is presented that may aid the assignment of ¹³C and ¹⁵N resonances of methyl-containing amino-acids in solid-state magic-angle spinning (MAS) NMR. A filtering block that selects methyl resonances is introduced in two-dimensional (2D) ¹³C-homonuclear and ¹⁵N–¹³C heteronuclear correlation experiments. The 2D ¹³C–¹³C correlation spectra are recorded with the methyl filter implemented prior to a ¹³C–¹³C mixing step. It is shown that these methyl-filtered ¹³C-homonuclear correlation spectra are instrumental in the assignment of C_δ resonances of leucines by suppression of C_γ–C_δ cross peaks. Further, a methyl filter is implemented prior to a ¹⁵N–¹³C transferred-echo double resonance (TEDOR) exchange scheme to obtain 2D ¹⁵N–¹³C heteronuclear correlation spectra. These experiments provide correlations between methyl groups and backbone amides. Some of the observed sequential ¹⁵N–¹³C correlations form the basis for initial sequence-specific assignments of backbone signals of the outer-membrane protein G.     

Optical properties and optical constituents of the Faroe Islands shelf and off-shelf waters,North East Atlantic

The study comprises a data set of CTD, optical properties—K ₀(PAR), c _p, a(PAR), b(PAR)—and optical constituents—Chl a, SPM, CDOM—from 72 shelf and off-shelf stations in the Faroe Islands (62°N, 7°W) North East Atlantic, in early spring 2005. Results showed that shelf waters surrounding the islands were cold and low saline, whereas off-shelf waters were warmer (~1°C) and more saline (~0.05) PSU. A pronounced oceanographic front separated the two waters, and diffuse light attenuation K ₀(PAR), beam attenuation c _p, Chl a, absorption a(PAR), and scattering coefficient b(PAR) were all significantly higher on the shelf. Analyses showed that off-shelf light attenuation K ₀(PAR) was governed by Chl a, shown by a high (r ² = 0.64) Chl a–K ₀(PAR) correlation, whereas light attenuation on the shelf was governed by both Chl a, SPM, and CDOM in combination. A Chl a specific diffuse attenuation coefficient K₀^* ( \textPAR ) K_{0}^{*} \left( {\text{PAR}} \right) of 0.056 (m² mg⁻¹ Chl a) and a Chl a specific beam attenuation ( c_\textp^* c_{\text{p}}^{*} ) of 0.27 (m² mg⁻¹ Chl a) coefficients were derived for the off-shelf. It is pointed out that Chl a is the single variable that changes over time as no rivers with high SPM and CDOM enter the shelf area. Data were obtained in early spring, and Chl a concentrations were low ~0.5 mg Chl a m⁻³. Spring bloom Chl a are about 10 mg Chl a m⁻³ and estimations showed that shelf K ₀(PAR) will increase about 5 times and beam attenuation about 10 times. The Faroe Islands shelf–off-shelf waters is a clear example where physical conditions maintain some clear differences in optical properties and optical constituents. The complete data set is enclosed.     

Stress-Induced Changes in Optical Properties,Pigment and Fatty Acid Content of <Emphasis Type="Italic">Nannochloropsis</Emphasis> sp.: Implications for Non-destructive Assay of Total Fatty Acids

In order to develop a practical approach for fast and non-destructive assay of total fatty acid (TFA) and pigments in the biomass of the marine microalga Nannochloropsis sp. changes in TFA, chlorophyll, and carotenoid contents were monitored in parallel with the cell suspension absorbance. The experiments were conducted with the cultures grown under normal (complete nutrient f/2 medium at 75 μmol PAR photons/(m² s)) or stressful (nitrogen-lacking media at 350 μmol PAR photons/(m² s)) conditions. The reliable measurement of the cell suspension absorbance using a spectrophotometer without integrating sphere was achieved by deposition of cells on glass–fiber filters in the chlorophyll content range of 3–13 mg/L. Under stressful conditions, a 30–50% decline in biomass and chlorophyll, retention of carotenoids and a build-up of TFA (15–45 % of dry weight) were recorded. Spectral regions sensitive to widely ranging changes in carotenoid-to-chlorophyll ratio and correlated changes of TFA content were revealed. Employing the tight inter-correlation of stress-induced changes in lipid metabolism and rearrangement of the pigment apparatus, the spectral indices were constructed for non-destructive assessment of carotenoid-to-chlorophyll ratio (range 0.3–0.6; root mean square error (RMSE) = 0.03; r ² = 0.93) as well as TFA content of Nannochloropsis sp. biomass (range 5.0–45%; RMSE = 3.23 %; r ² = 0.89) in the broad band 400–550 nm normalized to that in chlorophyll absorption band (centered at 678 nm). The findings are discussed in the context of real-time monitoring of the TFA accumulation by Nannochloropsis cultures under stressful conditions.     

Predicting the light attenuation coefficient through Secchi disk depth and beam attenuation coefficient in a large, shallow, freshwater lake

The diffuse attenuation coefficient of photosynthetically active radiation (PAR) (400–700 nm) (K _d(PAR)) is one of the most important optical properties of water. Our purpose was to create K _d(PAR) prediction models from the Secchi disk depth (SDD) and beam attenuation coefficient of particulate and dissolved organic matter (C _t−w(PAR), excluding pure water) in the PAR range. We compare their performance and prediction precision by using the determination coefficient (r ²), relative root mean square error (RRMSE), and mean relative error (MRE). Our dataset comprised 1,067 measurements, including K _d(PAR), SDD, and C _t−w(PAR) taken in shallow, eutrophic, Lake Taihu, China, from 2005 to 2010. The prediction models of K _d(PAR) were based on the linear model with an intercept of zero, using the inverse SDD, and the nonlinear model using SDD. The linear model generated a slope of 1.369, which was not significantly different from 1.7, the index used worldwide, but significantly lower than the value of 2.26. The nonlinear model gave a slightly more reliable prediction of K _d(PAR) with a r ² of 0.804. Compared to the SDD, C _t−w(PAR) was more significantly correlated to K _d(PAR) based on the linear model, with a significantly higher r ² and lower RMSE and RE. Considering the measurement simplicity of C _t−w(PAR) and data acquisition feasibility from high-frequency autonomous buoys and satellites, our results demonstrated that this prediction model reliably estimates K _d(PAR), and could be used to significantly expand optical observations in an environment where the conditions for underwater PAR measurement are limited.     

Estimating absolute pollen productivity for some European Tertiary-relict taxa

Tertiary-relict plants are survivors from the pre-Quaternary periods. Today, most European Tertiary relicts are confined to small, isolated stands distributed in the Mediterranean and Black Sea regions. In the past, however, the fossil record indicates that these species were probably distributed over large parts of the European continent and may have been important constituents of the vegetation. Little is known about their pollen representation, which limits our ability to reconstruct this past vegetation with any accuracy. This paper draws on the results of pollen trapping experiments in Bulgaria and Georgia, where relict stands of Aesculus hippocastanum, Cercis siliquastrum, Fagus orientalis, Juglans regia and Pterocarya fraxinifolia are still in existence. We compared average pollen accumulation rates (PAR) to vegetation data from around the trapping locations to derive estimates of absolute pollen productivity using various pollen dispersal functions. Composite dispersal functions that model pollen components carried above the vegetation canopy and falling as rain provided better relationships between PAR and plant abundance than functions that consider only a single component or the ‘trunk-space’ component carried under the canopy. A composite dispersal function with a simple model for regional pollen and the best overall correlation statistics gave the following estimates of absolute pollen productivity (grains cm⁻² yr⁻¹ with 1 SE intervals): Carpinus betulus 19,000–28,700; Fagus orientalis 15,600–20,400; Juglans regia 27,200–36,200; Pterocarya fraxinifolia 182,000–192,600; Quercus spp. 21,700–24,800; Tilia begoniifolia 51,600–68,300; and T. tomentosa 14,700–18,200. These estimates were applied to fossil data from the Black Sea coast to reconstruct palaeovegetation using absolute and relative methods.     

Photosynthesis and water-use characteristics in Indian mangroves

Photosynthesis and water efflux were measured in different PAR and stomatal conductance in members of Avicenniaceae and Rhizophoraceae. Trend of leaf temperature with irradiance and its effect on photosynthesis were also estimated. In most of the studied species, photosynthesis and stomatal conductance followed similar trends with increase in irradiance. The rate of net photosynthesis and stomatal conductance were higher in members of Avicenniaceae than in Rhizophoraceae. In Avicenniaceae, the optimum PAR for maximum photosynthesis ranged between 1340–1685 (μmol m^-2s^-1, which was also higher than that of Rhizophoraceae (840-1557 μmol m^-2s^-1). Almost in all the studied taxa, transpiration and stomatal conductance followed similar trends and reached the maximal peaks at the same PAR value. The range of breakeven leaf temperature was almost the same in both the families (34-36°C in Avicenniaceae and 33.5-36.3°C in Rhizophoraceae), beyond which assimilation rate declined.     

13C, 15N Resonance Assignment of Parts of the HET-s Prion Protein in its Amyloid Form

The partial ¹⁵N and ¹³C solid-state NMR resonance assignment of the HET-s prion protein fragment 218–289 in its amyloid form is presented. It is based on experiments measured at MAS frequencies in the range of 20–40 kHz using exclusively adiabatic polarization-transfer schemes. The resonance assignment within each residue is based on two-dimensional ¹³C––¹³C correlation spectra utilizing the DREAM mixing scheme. The sequential linking of the assigned residues used a set of two- and three-dimensional ¹⁵N––¹³C correlation experiments. Almost all cross peaks visible in the spectra are assigned, but only resonances from 43 of the 78 amino-acid residues could be detected. The missing residues are thought to be highly disordered and/or highly dynamic giving rise to broad resonance lines that escaped detection in the experiments applied. The line widths of the observed resonances are narrow and comparable to line widths observed in micro-crystalline samples. The 43 assigned residues are located in two fragments of about 20 residues. Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

Thallus morphology and optical characteristics affect growth and DNA damage by UV radiation in juvenile Arctic Laminaria sporophytes

Growth of young sporophytes of the brown algae Laminaria digitata, L. saccharina and L. solidungula from Spitsbergen were measured in the laboratory after being exposed for 21 days to either photosynthetically active radiation (PAR=P) or to full light spectrum (PAR + UV-A + UV-B=PAB) using of cutoff glass filters. The plants were grown at 8±2°C and 16 h light : 8 h dark cycles with 6 h additional ultraviolet radiation (UVR) exposure in the middle of the light period. Growth was measured every 10 min using growth chambers with online video measuring technique. Tissue morphology and absorption spectra were measured in untreated young sporophytes while chlorophyll (Chl) a content and DNA damage were measured in treated thalli at the end of the experiment. In all species, growth rates were significantly higher in sporophytes exposed to P alone compared to sporophytes exposed to PAB. Tissue DNA damage is dependent on thallus thickness and absorption spectra characteristics of pigments and UV-absorbing compounds. In sporophytes exposed to UVR, energy demands for repair of DNA damage and synthesis of UV-absorbing compounds for protection effectively diverts photosynthate at the expense of growth. Photosynthetic pigment was not significantly different between treatments suggesting a capacity for acclimation to moderate UVR fluence. The general growth pattern in sporophytes exposed to P alone showed an increasing growth rate from the onset of light (0500–0900 hours) to a peak at the middle of the light phase (0900–1500 hours), a decline towards the end of the light phase (1500–2100 hours) and a minimum “low” growth in the dark (2100–0500 hours) relative to growth during the entire light phase. Under PAB, different growth patterns were observed such as growth compensation at night in L. digitata, delayed growth recovery in L. saccharina and minimal but continuous growth in L. solidungula. Growth as an integrative parameter of all physiological processes showed that the effect of UVR is correlated to the depth distribution of these species.     

Sucrose unloading in the hypocotyl of the Ricinus communis L. seedling measured by 13C-nuclear magnetic resonance spectroscopy in vivo

The unloading of sucrose in the apical part of the hypocotyl of Ricinus communis L. seedlings was measured by ¹³C-nuclear magnetic resonance (NMR) spectroscopy. The cotyledons of the seedling were immersed in 5 mM Mes buffer containing 100 mM ¹³C-labeled sucrose. At intervals of 70–90 min, ¹³C-NMR spectra with broadband decoupling and nuclear Overhauser enhancement were acquired in vivo. The spectra showed growing ¹³C-resonances of the labeled positions in the sucrose molecule reaching steady-state labeling within 7–8 h. The specific ¹³C labeling of sucrose in the G₁-position changed from 0.38 in the supplied sucrose solution to 0.16 in the sucrose extracted from the hypocotyl piece at the end of the experiment (13 h). Labeling of starch (and other insolubles) in the hypocotyl piece was ca. 0.10. It is proposed that the decreased specific labeling of unloaded sucrose is mostly due to the separate local pools of sucrose in the cortex and pith parenchyma, respectively, and less to continuous starch degradation and conversion to sucrose. The report gives an example of the application of ¹³C-NMR spectroscopy in assimilate allocation studies. Received: 10 October 1998 / Accepted: 31 December 1998     

Peak fitting in 2D <Superscript>1</Superscript>H–<Superscript>13</Superscript>C HSQC NMR spectra for metabolomic studies

A modified Lorentzian distribution function is used to model peaks in two-dimensional (2D) ¹H–¹³C heteronuclear single quantum coherence (HSQC) nuclear magnetic resonance (NMR) spectra. The model fit is used to determine accurate chemical shifts from genuine signals in complex metabolite mixtures such as blood. The algorithm can be used to extract features from a set of spectra from different samples for exploratory metabolomics. First a reference spectrum is created in which the peak intensities are given by the median value over all samples at each point in the 2D spectra so that ¹H–¹³C correlations in any spectra are accounted for. The mathematical model provides a footprint for each peak in the reference spectrum, which can be used to bin the ¹H–¹³C correlations in each HSQC spectrum. The binned intensities are then used as variables in multivariate analyses and those found to be discriminatory are rapidly identified by cross referencing the chemical shifts of the bins with a database of ¹³C and ¹H chemical shift correlations from known metabolites.     

In situ Responses of Phytoplankton from the Subtropical Lake La Angostura (Tucumán, Argentina) in Relation to Solar Ultraviolet Radiation Exposure and Mixing Conditions

In situ experiments were conducted at various depths in the water column to determine the effects of solar ultraviolet radiation (UVR, 280–400 nm) on photosynthesis of natural phytoplankton assemblages from the subtropical Lake La Angostura (Argentina, 26°45′ S; 65°37° W, 1980 m asl.). Water samples were taken daily and incubated under three radiation treatments: (a) Samples exposed to UVR + Photosynthetic Available Radiation (PAR) – PAB treatment (280–700 nm); (b) Samples exposed to ultraviolet-A radiation (UV-A) + PAR – PA treatment (320–700 nm), and, (c) Samples exposed to PAR only – P treatment (400–700 nm). Additionally, depth profiles were done to determine different physical (i.e., temperature and underwater radiation field) and biological characteristics of the water column – photosynthetic pigments, UV-absorbing compounds, cell concentration, deoxyribonucleic acid (DNA) and cyclobutane pyrimidine dimers (CPDs). The effects of UVR on natural phytoplankton assemblages were significant only in the first 50 cm of the water column, causing a decrease in photosynthetic rates of 36 and 20% due to UV-A and ultraviolet-B radiation (UV-B), respectively; below this depth, however, there were no significant differences between radiation treatments. Concentration of CPDs per mega base of DNA in natural phytoplankton was low, <27 CPDs MB⁻¹ between 0 and 4 m. Data on net DNA damage, together with that on mixing conditions of the water column, suggest that mixing can favour phytoplankton by allowing cells to be transported to depths where active repair can take place. This mechanism to reduce UVR-induced DNA damage would be of great advantage for these assemblages dominated by small cyanobacteria and chlorophytes where UV-absorbing compounds that could act as sunscreens are virtually absent.