174 research outputs found
Spectral and Temporal Laser Fluorescence Analysis Such as for Natural Aquatic Environments
An Advanced Laser Fluorometer (ALF) can combine spectrally and temporally resolved measurements of laser-stimulated emission (LSE) for characterization of dissolved and particulate matter, including fluorescence constituents, in liquids. Spectral deconvolution (SDC) analysis of LSE spectral measurements can accurately retrieve information about individual fluorescent bands, such as can be attributed to chlorophyll-a (Chl-a), phycobiliprotein (PBP) pigments, or chromophoric dissolved organic matter (CDOM), among others. Improved physiological assessments of photosynthesizing organisms can use SDC analysis and temporal LSE measurements to assess variable fluorescence corrected for SDC-retrieved background fluorescence. Fluorescence assessments of Chl-a concentration based on LSE spectral measurements can be improved using photo-physiological information from temporal measurements. Quantitative assessments of PBP pigments, CDOM, and other fluorescent constituents, as well as basic structural characterizations of photosynthesizing populations, can be performed using SDC analysis of LSE spectral measurements
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Analysis of spectral excitation for measurements of fluorescence constituents in natural waters
Field measurements of chlorophyll-a (Chl), phycoerythrin (PE), chromophoric dissolved organic matter (CDOM), and variable fluorescence (Fv/Fm) in diverse waters of the California Current, Mediterranean Sea and Gulf of Mexico using 375, 405, 510 and 532 nm laser excitation wavelengths (EW) are analyzed. EW = 375 and 405 nm were found more suitable for Chl assessment in high-Chl (> 10 μg/l) waters. Both EW = 532 and 510 nm can be used to efficiently stimulate PE fluorescence for structural characterization of phytoplankton communities. EW = 375 nm and 405 nm can provide best results for CDOM assessments in offshore oceanic waters; the green EWs can be also used for CDOM measurements in fresh and estuarine water types in conjunction with spectral discrimination between CDOM and PE fluorescence. Both EW = 405 and 510 are suitable for photo-physiological Fv/Fm assessments, though using EW = 405 nm may result in underestimation of PE-containing phytoplankton groups present in mixed phytoplankton assemblages
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Next generation Advanced Laser Fluorometry (ALF) for characterization of natural aquatic environments: new instruments
The new optical design allows single- or multi-wavelength excitation of laser-stimulated emission (LSE), provides optimized LSE optical collection for spectral and temporal analyses, and incorporates swappable modules for flow-through and small-volume sample measurements. The basic instrument configuration uses 510 nm laser excitation for assessments of chlorophyll-a, phycobiliprotein pigments, variable fluorescence (F_v/F_m) and chromophoric dissolved organic matter (CDOM) in CDOM-rich waters. The three-laser instrument configuration (375, 405, and 510 nm excitation) provides additional Fv/Fm measurements with 405 nm excitation, CDOM assessments in a broad concentration range, and potential for spectral discrimination between oil and CDOM fluorescence. The new measurement protocols, analytical algorithms and examples of laboratory and field measurements are discussed
Advanced laser fluorometry of natural aquatic environments
The Advanced Laser Fluorometer (ALF) provides spectral deconvolution (SDC) analysis of the laser-stimulated emission (LSE) excited at 405 or 532 nm for assessment of chlorophyll a, phycoerythrin, and chromophoric dissolved organic matter. Three spectral types of phycoerythrin are discriminated for characterization of cyanobacteria and cryptophytes in mixed phototrophic populations. The SDC analysis is integrated with measurements of variable fluorescence, Fv/Fm, corrected for the SDC-retrieved background fluorescence, BNC, for improved photophysiological assessments of phytoplankton. The ALF deployments in the Atlantic and Pacific Oceans, and Chesapeake, Delaware, and Monterey Bays revealed significant spectral complexity of LSE. Considerable variability in chlorophyll a fluorescence peak, 673-685 nm, was detected. High correlation (R2 = 0.93) was observed in diverse water types between chlorophyll a concentration and fluorescence normalized to water Raman scattering. Three unidentified red bands, peaking at 625, 644, and 662 nm, were detected in the LSE excited at 405 nm. Significant variability in the BNC/chlorophyll a ratio was observed in diverse waters. Examples of the ALF spectral correction of Fv/Fm, underway shipboard measurements of horizontal variability, and vertical distributions compiled from the discrete samples analyses are presented. The field deployments have demonstrated the utility of the ALF technique as an integrated tool for research and observations
Photo-physiological variability in phytoplankton chlorophyll fluorescence and assessment of chlorophyll concentration
Photo-physiological variability of in vivo chlorophyll fluorescence (CF) per unit of chlorophyll concentration (CC) is analyzed using a biophysical model to improve the accuracy of CC assessments. Field measurements of CF and photosystem II (PSII) photochemical yield (PY) with the Advanced Laser Fluorometer (ALF) in the Delaware and Chesapeake Bays are analyzed vs. high-performance liquid chromatography (HPLC) CC retrievals. It is shown that isolation from ambient light, PSII saturating excitation, optimized phytoplankton exposure to excitation, and phytoplankton dark adaptation may provide accurate in vivo CC fluorescence measurements (R2 = 0.90-0.95 vs. HPLC retrievals). For in situ or flow-through measurements that do not allow for dark adaptation, concurrent PY measurements can be used to adjust for CF non-photochemical quenching (NPQ) and improve the accuracy of CC fluorescence assessments. Field evaluation has shown the NPQ-invariance of CF/PY and CF(PY-1-1) parameters and their high correlation with HPLC CC retrievals (R2 = 0.74-0.96), while the NPQ-affected CF measurements correlated poorly with CC (R2 = -0.22)
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Pump-During-Probe Fluorometry of Phytoplankton: Group-Specific Photosynthetic Characteristics from Individual Cell Analysis
Saturating-flash fluorescence techniques are used to monitor the state of the photosynthetic apparatus in phytoplankton under natural conditions. At present these are bulk water measurements, which produce estimates of average properties of all the fluorescent particles present in a sample. Here we describe an improved approach for single-cell measurements of phytoplankton. We have combined individual- cell 'pump-during-probe' measurements of chlorophyll fluorescence induction on the time scale of 30 to 100 microseconds with flow cytometric characterization of each cell, to obtain population-specific photosynthetic characteristics. The results provide information about the potential quantum yield of photochemistry, the fraction of functional reaction centers, and the functional absorption cross section for photosystem 2
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Airborne Test of Laser Pump-And-Probe Technique for Assessment of Phytoplankton Photochemical Characteristics
Initial results of the airborne LIDAR measurement of photochemical quantum yield, ΦPo, and functional absorption cross-section, σPS II, of Photosystem II (PS II) are reported. NASA's AOL3 LIDAR was modified to implement short-pulse pump-and-probe (SP-P P) LIDAR measurement protocol. The prototype system is capable of measuring a pump-induced increase in probe-stimulated chlorophyll fluorescence, ΔF/Fsat, along with the acquisition of conventional LIDAR-fluorosensor products from an operational altitude of 150 m. The use of a PS II sub-saturating probe pulse increases the response signal but also results in excessive energy quenching (EEQ) affecting the ΔF/Fsat magnitude. The airborne data indicated up to a 3-fold EEQ-caused decline in ΔF/Fsat, and 2-fold variability in the EEQ rate constant over a spatial scale a few hundred kilometers. Therefore, continuous monitoring of EEQ parameters must be incorporated in the operational SP-P P protocol to provide data correction for the EEQ effect. Simultaneous airborne LIDAR measurements of ΦPo and σPS II with EEQ correction were shown to be feasible and optimal laser excitation parameters were determined. Strong daytime ΔF/Fsat decline under ambient light was found in the near-surface water layer over large aquatic areas. An example of SP-P P LIDAR measurement of phytoplankton photochemical and fluorescent characteristics in the Chesapeake Bay mouth is presented. Prospects for future SP-P P development and related problems are discussed
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Short-Pulse Pump-And-Probe Technique for Airborne Laser Assessment of Photosystem II Photochemical Characteristics
The development of a technique for laser measurement of fPhotosystem II (PS II) photochemical characteristics of phytoplankton and terrestrial vegetation from an airborne platform is described. Results of theoretical analysis and experimental study of pump-and-probe measurement of the PS II functional absorption cross-section and photochemical quantum yield are presented. The use of 10 ns probe pulses of PS II sub-saturating intensity provides a significant, up to 150-fold, increase in the fluorescence signal compared to conventional 'weak-probe' protocol. Little effect on the fluorescence yield from the probe-induced closure of PS II reaction centers is expected over the short pulse duration, and thus a relatively intense probe pulse can be used. On the other hand, a correction must be made for the probe-induced carotenoid triplet quenching and singlet-singlet annihilation. A Stern-Volmer model developed for this correction assumes a linear dependence of the quenching rate on the laser pulse fluence, which was experimentally validated. The PS II saturating pump pulse fluence (532 nm excitation) was found to be 10 and 40 mumol quanta m(-2) for phytoplankton samples and leaves of higher plants, respectively. Thirty mus was determined as the optimal delay in the pump-probe pair. Our results indicate that the short-pulse pump-and-probe measurement of PS II photochemical characteristics can be implemented from an airborne platform using existing laser and LIDAR technologies
Aquatic laser fluorescence analyzer: field evaluation in the northern Gulf of Mexico
The new Aquatic Laser Fluorescence Analyzer (ALFA) provides spectral and temporal measurements of laser-stimulated emission (LSE) for assessment of phytoplankton pigments, community structure, photochemical efficiency (PY), and chromophoric dissolved organic matter (CDOM). The instrument was deployed in the Northern Gulf of Mexico to evaluate the ALFA analytical capabilities across the estuarine-marine gradient. The robust relationships between the pigment fluorescence and independent pigment measurements were used to validate the ALFA analytical algorithms and calibrate the instrument. The maximal PY magnitudes, PYm = PY(1-1.35·10−4PAR)−1, were estimated using the underway measurements of PY and photosynthetically active radiation (PAR). The chlorophyll (Chl) spatial patterns were calculated using the ratio of Chl fluorescence to PY to eliminate the effect of non-photochemical quenching on the underway Chl assessments. These measurements have provided rich information about spatial distributions of Chl, PYm, CDOM, and phytoplankton community structure, and demonstrated the utility of the ALFA instrument for oceanographic studies and bio-environmental surveys. The data suggest that the fluorescence measurements with 514 nm excitation can provide informative data for characterization of the CDOM-rich fresh, estuarine, and coastal aquatic environments
Laser fluorescence analysis of phytoplankton across a frontal zone in the California Current ecosystem
Spatial variability of chlorophyll, phycobiliproteins, chromophoric dissolved organic matter and variable fluorescence (Fv/Fm) was analyzed across a deep-water density front in the Southern California Current Ecosystem using an Advanced Laser Fluorometer (ALF) calibrated to assess chlorophyll concentration (Cchl), total autotrophic carbon (AC) and Synechococcus carbon biomass (SYN). Three distinct autotrophic assemblages were identified. Fluorescence was found to be three to four times higher in cooler mesotrophic waters north of the front than in warm oligotrophic waters to the south. Northern waters were distinguished by a shallow pigment maximum dominated by a blue-water type of Synechococcus and by the presence of green-water Synechococcus and cryptophytes; only blue-water Synechococcus were detected at lower concentration south of the front. The highest Cchl and AC values, accompanied by elevated Fv/Fm and chlorophyll fluorescence per unit of Cchl, and minimal Synechococcus abundance, were found directly at the front in a 20–40 m deep layer dominated by diatoms. The covariation of Fv/Fm with nitrate concentration in this layer, along with the structural changes in the phytoplankton community, suggest that it had been generated by in situ processes rather than advection. Strong structural responses to the local hydrography were also revealed by high-frequency underway ALF surface sampling, which detected an abrupt transition from low to high SYN on the northern side of a sharp salinity gradient at the front. Synechococcus-specific phycoerythrin fluorescence (FPE12) and SYN were highly correlated in surface waters (R2= 0.95), while FPE12:SYN gradually increased with depth. Strong relationships were found for chlorophyll fluorescence versus Cchl (R2= 0.95) and AC (R2= 0.79)
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