Formation of soluble microbial products (SMP) by activated sludge at various salinities

Soluble microbial products (SMP) present a significant component of effluent organic matter from biological wastewater treatment reactors, and can affect the membrane fouling and formation of disinfection by-products. Thus, SMP have attracted increasing concerns in wastewater treatment and reclamation. In this work, the formation of SMP by activated sludge at various NaCl concentrations is investigated by using fluorescence excitation–emission matrix (EEM) spectroscopy with parallel factor analysis (PARAFAC) and fluorescence regional integration (FRI). The results show that a high level of salinity decreases substrate removal efficiency and leads to an accumulation of SMP, especially proteins. Three components of SMP, one protein-like and two humic-acid-like components, are identified by PARAFAC, which exhibit different trends with the variation of NaCl concentration. FRI analysis reveals that the majority of protein fluorescence is attributed to tryptophan and tryptophan-like proteins, rather than tyrosine and tyrosine-like proteins. With an increase in NaCl concentration, the normalized volume percentages of tyrosine and tryptophan region increase, while those of humic- and fulvic-acid-like region decrease significantly. This work demonstrates that salinity affects the formation of SMP, and that EEM with PARAFAC in combination with FRI analysis is a useful tool to get insight into the formation of SMP by activated sludge.     

Biodegradation behavior of natural organic matter (NOM) in a biological aerated filter (BAF) as a pretreatment for ultrafiltration (UF) of river water

In this study, biodegradation of natural organic matter (NOM) in a biological aerated filter (BAF) as pretreatment of UF treating river water was investigated. Photometric measurement, three-dimensional excitation–emission matrix (EEM) fluorescence spectroscopy and liquid chromatography with online organic carbon detector (LC-OCD) were used to investigate the fate of NOM fractions in the BAF + UF process. Results showed that the BAF process could effectively remove particles and parts of dissolved organic matter, which led to a lower NOM loading in the UF system, but different NOM fractions showed different biodegradation potentials. Further biodegradation batch experiments confirmed this observation and identified that polysaccharides and proteins (quantified using photometric methods) contained a large proportion of readily biodegradable matter while humic substances were mainly composed of inert organic substances. According to EEM measurements, it is evident that protein-like substances were more readily eliminated by microorganisms than humic-like substances. LC-OCD data also supported the phenomena that the polysaccharides and large-size proteins were more degradable than humic substances.     

Fluorescence characteristics and biodegradability of dissolved organic matter in forest and wetland soils from coastal temperate watersheds in southeast Alaska

Understanding how the concentration and chemical quality of dissolved organic matter (DOM) varies in soils is critical because DOM influences an array of biological, chemical, and physical processes. We used PARAFAC modeling of excitation–emission fluorescence spectroscopy, specific UV absorbance (SUVA₂₅₄) and biodegradable dissolved organic carbon (BDOC) incubations to investigate the chemical quality of DOM in soil water collected from 25 cm piezometers in four different wetland and forest soils: bog, forested wetland, fen and upland forest. There were significant differences in soil solution concentrations of dissolved organic C, N, and P, DOC:DON ratios, SUVA₂₅₄ and BDOC among the four soil types. Throughout the sampling period, average DOC concentrations in the four soil types ranged from 9–32 mg C l⁻¹ and between 23–42% of the DOC was biodegradable. Seasonal patterns in dissolved nutrient concentrations and BDOC were observed in the three wetland types suggesting strong biotic controls over DOM concentrations in wetland soils. PARAFAC modeling of excitation–emission fluorescence spectroscopy showed that protein-like fluorescence was positively correlated (r ² = 0.82; P < 0.001) with BDOC for all soil types taken together. This finding indicates that PARAFAC modeling may substantially improve the ability to predict BDOC in natural environments. Coincident measurements of DOM concentrations, BDOC and PARAFAC modeling confirmed that the four soil types contain DOM with distinct chemical properties and have unique fluorescent fingerprints. DOM inputs to streams from the four soil types therefore have the potential to alter stream biogeochemical processes differently by influencing temporal patterns in stream heterotrophic productivity.     

Characterization of extracellular polymeric substances from biofilm in the process of starting-up a partial nitrification process under salt stress

In this study, the characteristics of extracellular polymeric substance (EPS) fractions of biofilm during the process of establishing a partial nitrification under salt stress were analyzed in terms of concentrations, molecular weight distribution, and three-dimensional excitation–emission matrix (EEM) fluorescence spectroscopy. A partial nitrification was formed successfully with a salinity of 1%. Results indicated that the amount of total EPS increased from 54.2 mg g⁻¹ VSS⁻¹ on day 1 to 99.6 mg g⁻¹ VSS⁻¹ on day 55 due to the NaCl concentration changed from 0 to 10.0 g L⁻¹ in a biofilm reactor. The changes of loosely bound EPS (LB-EPS) compounds under different salt concentrations appeared to be more significant than those of the tightly bound EPS. A clear release of polysaccharides in the LB-EPS fraction was detected during the enhancement of salinity. This was considered as a protective response of bacteria to the salinity. Three fluorescence peaks were identified in the EEM fluorescence spectra of the EPS fraction samples. Two peaks were assigned to the protein-like fluorophores, and the third peak was located at the excitation/emission wavelengths of 275 nm/425–435 nm of the spectra of EPS fractions till the salinity maintained constant at 1%. This information is valuable for understanding the characteristics of EPS isolated from biomass in a saline nitrogen removal system.     

Characterization of Nanming River (southwestern China) sewerage-impacted pollution using an excitation-emission matrix and PARAFAC

Nanming River, the largest urban river in Guizhou Province, southwestern China plateau, has been severely polluted for decades. This study characterizes the organic materials and their sources in the upstream and downstream waters by dissolved organic carbon (DOC), excitation emission matrix (EEM) spectroscopy, parallel factor (PARAFAC) analysis and photo-microbial experiments. DOC concentrations were low (47–120 μM C) upstream and relatively high (146–462 μM C) downstream. The PARAFAC studies on the sample EEM spectra demonstrated that the upstream dissolved organic matter (DOM) was mostly composed of one component that had a fulvic acid-like substance; downstream DOM was composed of two components with mixtures of tryptophan-like and fulvic acid-like substances. From the results of the sewerage drainage samples collected along the bank of the river, it is evident that both household detergent-like and protein-like or tryptophan-like substances are predominantly present, indicating that untreated sewerage effluents are the major sources of organic matter pollution in Nanming River. The degradation experiments conducted on river, sewerage drainage and commercial detergent samples demonstrated that the detergent-like and tryptophan-like substances are both photochemically and microbiologically more decomposable than fulvic acid-like materials under sunlight and dark incubations. These results suggest that the input of the untreated sewerage effluents along the streams is the major pollution source in Nanming River, and the fluorescent DOM was efficiently affected by both photochemical and microbial processes.     

Characterizing the photochemical degradation of aquatic humic substances from a dystrophic lake using excitation-emission matrix fluorescence spectroscopy and parallel factor analysis

Three-dimensional excitation-emission matrix (EEM) fluorescence and parallel factor analysis (PARAFAC) were used to monitor composition and reactivity changes caused by the photochemical degradation of aquatic humic substances (AHS) from a dystrophic lake in Kushiro Wetland, Japan. AHS-rich lake water was exposed to three treatments in summer and winter 2014: radiation with the full solar wavelength range, radiation with the >320-nm solar wavelength range, and no solar radiation. Irradiation caused AHS-like peaks to shift to shorter wavelengths in the EEM contour plots, implying that AHS photodegradation caused the formation of lower-molecular-weight fractions or more simply structured components. Three components were identified from PARAFAC analyses: AHS-1 (excitation/emission wavelengths of maxima: <252 and 315 nm/426 nm), AHS-2 (360 and 261 nm/489 nm), and AHS-3 (276 nm/403 nm). These components had different photosensitivities. AHS-1 was most sensitive to full solar radiation, while AHS-2 was most sensitive to >320-nm radiation. More photodegradation of these components occurred in the summer than in the winter, indicating that photodegradation depended on light intensity. AHS-3 was photoresistant. The different characteristics of the components reflected the in situ dynamics of the components. The AHS-3 fluorescence intensity was positively correlated with the dissolved organic carbon concentration but the AHS-1 and AHS-2 fluorescence intensities were not. The EEM–PARAFAC method was found to be a good tool for tracing AHS-like materials in situ and in the laboratory.     

Return of Salmon-Derived Nutrients from the Riparian Zone to the Stream during a Storm in Southeastern Alaska

Spawning salmon deliver nutrients (salmon-derived nutrients, SDN) to natal watersheds that can be incorporated into terrestrial and aquatic food webs, potentially increasing ecosystem productivity. Peterson Creek, a coastal watershed in southeast Alaska that supports several species of anadromous fish, was sampled over the course of a storm during September 2006 to test the hypothesis that stormflows re-introduce stored SDN into the stream. We used stable isotopes and PARAFAC modeling of fluorescence excitation–emission spectroscopy to detect flushing of DOM from salmon carcasses in the riparian zone back into a spawning stream. During the early storm hydrograph, streamwater concentrations of NH₄–N and total dissolved phosphorus (TDP), the fluorescent protein tyrosine and the δ¹⁵N content of DOM peaked, followed by a rapid decrease during maximum stormflow. Although δ¹⁵N has previously been used to track SDN in riparian zones, the use of fluorescence spectroscopy provides an independent indicator that SDN are being returned from the riparian zone to the stream after a period of intermediate storage outside the stream channel. Our findings further demonstrate the utility of using both δ¹⁵N of streamwater DOM and fluorescence spectroscopy with PARAFAC modeling to monitor how the pool of streamwater DOM changes in spawning salmon streams.     

Fluorescence spectroscopy as a potential metabonomic tool for early detection of colorectal cancer

Fluorescence spectroscopy Excitation Emission Matrix (EEM) measurements were applied on human blood plasma samples from a case control study on colorectal cancer. Samples were collected before large bowel endoscopy and included patients with colorectal cancer or with adenomas, and from individuals with other non malignant findings or no findings (N = 308). The objective of the study was to explore the possibilities for applying fluorescence spectroscopy as a tool for detection of colorectal cancer. Parallel Factor Analysis (PARAFAC) was applied to decompose the fluorescence EEMs into estimates of the underlying fluorophores in the sample. Both the pooled score matrix from PARAFAC, holding the relative concentrations of the derived components, and the raw unfolded spectra were used as basis for discrimination models between cancer and the various controls. Both methods gave test set validated sensitivity and specificity values around 0.75 between cancer and controls, and poor discriminations between the various controls. The PARAFAC solution gave better options for analyzing the chemical mechanisms behind the discrimination, and revealed a blue shift in tryptophan emission in the cancer patients, a result that supports previous findings. The present findings show how fluorescence spectroscopy and chemometrics can help in cancer diagnostics, and with PARAFAC fluorescence spectroscopy can be a potential metabonomic tool.     

Erratum to: Fluorescence spectroscopy as a potential metabonomic tool for early detection of colorectal cancer

Fluorescence spectroscopy Excitation Emission Matrix (EEM) measurements were applied on human blood plasma samples from a case control study on colorectal cancer. Samples were collected before large bowel endoscopy and included patients with colorectal cancer or with adenomas, and from individuals with other non malignant findings or no findings (N = 308). The objective of the study was to explore the possibilities for applying fluorescence spectroscopy as a tool for detection of colorectal cancer. Parallel Factor Analysis (PARAFAC) was applied to decompose the fluorescence EEMs into estimates of the underlying fluorophores in the sample. Both the pooled score matrix from PARAFAC, holding the relative concentrations of the derived components, and the raw unfolded spectra were used as basis for discrimination models between cancer and the various controls. Both methods gave test set validated sensitivity and specificity values around 0.75 between cancer and controls, and poor discriminations between the various controls. The PARAFAC solution gave better options for analyzing the chemical mechanisms behind the discrimination, and revealed a blue shift in tryptophan emission in the cancer patients, a result that supports previous findings. The present findings show how fluorescence spectroscopy and chemometrics can help in cancer diagnostics, and with PARAFAC fluorescence spectroscopy can be a potential metabonomic tool.

     

Distribution of dissolved organic carbon and dissolved fulvic acid in mesotrophic Lake Biwa, Japan

The dissolved organic carbon (DOC) concentrations in mesotrophic Lake Biwa were determined by a total organic carbon (TOC) analyzer, and DOC molecular size distributions were determined by size exclusion chromatography (SEC) using a fluorescence detector at excitation/emission (Ex/Em) levels of 300/425 nm with the eluent at pH 9.7. The fluorescence wavelengths for detection were chosen from the result of excitation–emission matrix spectrometry (EEM) analysis for dissolved fulvic acid (DFA) extracted from Ado River (peak A, Ex/Em = 260–270/430–440 nm; peak B, Ex/Em = 300–310/420–430 nm). Ado River DFA was eluted with a retention time (RT) of 7.4–8.9 min and the apparent molecular weight was estimated at 22–87 kDa based on the elution curve for the spherical protein molecular weight standard. A DFA peak eluted at the same retention time as Ado River DFA also appeared in all the samples of Lake Biwa water. From the linear relationship between the peak areas with an RT of 7.4–8.9 min by SEC analysis and DOC values of DFA by TOC analysis of a series of DFA samples (r² = 0.9995), the concentrations of DFA in the lake water were roughly calculated. DFA was distributed within the range 0.25–0.43 mg C l⁻¹ and accounted for 15%–41% of DOC, with the highest ratios observed at a depth of 70 m in August and the lowest at 2.5 m in May.     

Study of Gallium Arsenide Etching in a DC Discharge in Low-Pressure HCl-Containing Mixtures

Halogen-containing plasmas are often used to form topological structures on semiconductor surfaces; therefore, spectral monitoring of the etching process is an important diagnostic tool in modern electronics. In this work, the emission spectra of gas discharges in mixtures of hydrogen chloride with argon, chlorine, and hydrogen in the presence of a semiconducting gallium arsenide plate were studied. Spectral lines and bands of the GaAs etching products appropriate for monitoring the etching rate were determined. It is shown that the emission intensity of the etching products is proportional to the GaAs etching rate in plasmas of HCl mixtures with Ar and Cl₂, which makes it possible to monitor the etching process in real time by means of spectral methods.     

Modelling Förster resonance energy transfer (FRET) using anisotropy resolved multi-dimensional emission spectroscopy (ARMES)

BackgroundFörster Resonance Energy Transfer (FRET) is widely used to study the structure and dynamics of biomolecular systems and also causes the non-linear fluorescence response observed in multi-fluorophore proteins. Accurate FRET analysis, in terms of measuring changes in donor and acceptor spectra and energy transfer efficiency is therefore critical.MethodsWe demonstrate a novel quantitative FRET analysis using anisotropy resolved multidimensional emission spectroscopy (ARMES) in a Human Serum Albumin (HSA) and 1,8-anilinonaphathalene sulfonate (ANS) model. ARMES combines 4D measurement of polarized excitation emission matrices (pEEM) with multivariate data analysis to spectrally resolve contributing fluorophores. Multivariate analysis (Parallel Factor, PARAFAC and restricted Tucker3) was used to resolve fluorophore contributions and for modelling the quenching of HSA emission and the HSA-ANS interactions.ResultspEEM spectra were modelled using Tucker3 which accommodates non-linearities introduced by FRET and a priori chemical knowledge was used to optimise the solution, thus resolving three components: HSA emission, ANS emission from indirect FRET excitation, and ANS emission from direct excitation. Perpendicular emission measurements were more sensitive to indirectly excited acceptor emission. PARAFAC modelling of HSA, donor emission, separated ANS FRET interacting (Tryptophan) and non-interacting (Tyrosine) components. This enabled a new way of calculating quenching constants using the multi-dimensional emission of individual donor fluorophores.ConclusionsFRET efficiency could be calculated using the multi-dimensional, resolved emission of the interacting donor fluorophores only which yielded higher ET efficiencies compared to conventional methods.General significanceShows the potential of multidimensional fluorescence measurements and data analysis for more accurate FRET modelling in proteins.     

Dissolved Organic Matter Characteristics Across a Subtropical Wetland’s Landscape: Application of Optical Properties in the Assessment of Environmental Dynamics

Wetlands are known to be important sources of dissolved organic matter (DOM) to rivers and coastal environments. However, the environmental dynamics of DOM within wetlands have not been well documented on large spatial scales. To better assess DOM dynamics within large wetlands, we determined high resolution spatial distributions of dissolved organic carbon (DOC) concentrations and DOM quality by excitation–emission matrix spectroscopy combined with parallel factor analysis (EEM–PARAFAC) in a subtropical freshwater wetland, the Everglades, Florida, USA. DOC concentrations decreased from north to south along the general water flow path and were linearly correlated with chloride concentration, a tracer of water derived from the Everglades Agricultural Area (EAA), suggesting that agricultural activities are directly or indirectly a major source of DOM in the Everglades. The optical properties of DOM, however, also changed successively along the water flow path from high molecular weight, peat-soil and highly oxidized agricultural soil-derived DOM to the north, to lower molecular weight, biologically produced DOM to the south. These results suggest that even though DOC concentration seems to be distributed conservatively, DOM sources and diagenetic processing can be dynamic throughout wetland landscapes. As such, EEM–PARAFAC clearly revealed that humic-enriched DOM from the EAA is gradually replaced by microbial- and plant-derived DOM along the general water flow path, while additional humic-like contributions are added from marsh soils. Results presented here indicate that both hydrology and primary productivity are important drivers controlling DOM dynamics in large wetlands. The biogeochemical processes controlling the DOM composition are complex and merit further investigation.     

Effects of aeration intensity on formation of phenol-fed aerobic granules and extracellular polymeric substances

Effect of air aeration intensities on granule formation and extracellular polymeric substances content in three identical sequential batch reactors were investigated. The excitation–emission–matrix spectra and multiple staining and confocal laser scanning microscope revealed proteins, polysaccharides, lipids, and humic substances in the sludge and granule samples. Seed sludge flocs were compacted at low aeration rate, with produced extracellular polymeric substances of 50.2–76.7 mg g⁻¹ of proteins, 50.2–77.3 mg g⁻¹ carbohydrates and 74 mg g⁻¹ humic substances. High aeration rate accelerated formation of 1.0–1.5 mm granules with smooth outer surface. The corresponding quantities of extracellular polymeric substances were 309–537 mg g⁻¹ of proteins, 61–109 mg g⁻¹ carbohydrates, 49–92 mg g⁻¹ humic substances, and 49–68 mg g⁻¹ lipids. Intermediate aeration rate produced 3.0–3.5 mm granules with surface filaments. Reactor failure occurred with overgrowth of filaments, probably owing to the deficiency of nutrient in liquid phase. No correlation was noted between extracellular polymeric substances composition and the proliferation of filamentous microorganisms on granule surface.     

Tracking variations in fluorescent-dissolved organic matter in an aerobic submerged membrane bioreactor using excitation–emission matrix spectra combined with parallel factor analysis

In this study, the variations in the fluorescent components of dissolved organic matter (DOM) were tracked for an aerobic submerged membrane bioreactor (MBR) at three different operation stages (cake layer formation, condensation, and after cleaning). The fluorescent DOM was characterized using excitation–emission matrix (EEM) spectroscopy combined with parallel factor analysis (PARAFAC). Non-aromatic carbon structures appear to be actively involved in the membrane fouling for the cake layer formation stage as revealed by much higher UV-absorbing DOM per organic carbon found in the effluent versus those inside the reactor. Four fluorescent components were successfully identified from the reactor and the effluent DOMs by EEM-PARAFAC modeling. Among those in the reactor, microbial humic-like fluorescence was the most abundant component at the cake layer formation stage and tryptophan-like fluorescence at the condensation stage. In contrast to the reactor, relatively similar composition of the PARAFAC components was exhibited for the effluent at all three stages. Tryptophan-like fluorescence displayed the largest difference between the reactor and the effluent, suggesting that this component could be a good tracer for membrane fouling. It appears that the fluorescent DOM was involved in membrane fouling by cake layer formation rather than by internal pore adsorption because its difference between the reactor and the effluent was the highest among all the four components, even after the membrane cleaning. Our study provided an insight into the fate and the behavior fluorescent DOM components for an MBR system, which could be an indicator of the membrane fouling.     

Photochemical degradation of chromophoric-dissolved organic matter exposed to simulated UV-B and natural solar radiation

Photochemical degradation of chromophoric-dissolved organic matter (CDOM) by UV-B radiation decreases CDOM absorption in the UV region and fluorescence intensity, and alters CDOM composition. CDOM absorption, fluorescence, and the spectral slope indicating the CDOM composition were studied using 0.22-μm-filtered samples of Meiliang Bay water from Lake Taihu that were exposed to short-term (0–12 h) simulated UV-B radiation and long-term (0–12 days) natural solar radiation in summer. CDOM absorption coefficient and fluorescence decreased with increasing exposure time, which relates to the amounts of absorbed light energy. The decreases of CDOM absorption and normalized fluorescence corresponded to first order kinetics reactions. Different decreases of CDOM absorption and fluorescence at different wavelengths suggested that the composition of CDOM changed when it absorbed ultraviolet radiation. Photochemical degradation increased the spectral slope during 275–295 nm region (S _275–295) but decreased the spectral slope during 275–295 nm region (S _350–400). The slope ratio S _R (S _275–295:S _350–400) increased in the photochemical process, which could be used as an indicator of photobleaching and composition change of CDOM. Our results show that photochemical degradation is important in the cycling of CDOM, which indicated change in the composition of CDOM. Handling editor: Luigi Naselli-Flores     

Light-induced quenching of chlorophyll fluorescence at 77 K in leaves,chloroplasts and Photosystem II particles

The light-induced chlorophyll (Chl) fluorescence decline at 77 K was investigated in segments of leaves, isolated thylakoids or Photosystem (PS) II particles. The intensity of chlorophyll fluorescence declines by about 40% upon 16 min of irradiation with 1000 μmol m⁻² s⁻¹ of white light. The decline follows biphasic kinetics, which can be fitted by two exponentials with amplitudes of approximately 20 and 22% and decay times of 0.42 and 4.6 min, respectively. The decline is stable at 77 K, however, it is reversed by warming of samples up to 270 K. This proves that the decline is caused by quenching of fluorescence and not by pigment photodegradation. The quantum yield for the induction of the fluorescence decline is by four to five orders lower than the quantum yield of Q_A reduction. Fluorescence quenching is only slightly affected by addition of ferricyanide or dithionite which are known to prevent or stimulate the light-induced accumulation of reduced pheophytin (Pheo). The normalised spectrum of the fluorescence quenching has two maxima at 685 and 695 nm for PS II emission and a plateau for PS I emission showing that the major quenching occurs within PS II. ‘Light-minus-dark’ difference absorbance spectra in the blue spectral region show an electrochromic shift for all samples. No absorbance change indicating Chl oxidation or Pheo reduction is observed in the blue (410–600 nm) and near infrared (730–900 nm) spectral regions. Absorbance change in the red spectral region shows a broad-band decrease at approximately 680 nm for thylakoids or two narrow bands at 677 and 670–672 nm for PS II particles, likely resulting also from electrochromism. These absorbance changes follow the slow component of the fluorescence decline. No absorbance changes corresponding to the fast component are found between 410 and 900 nm. This proves that the two components of the fluorescence decline reflect the formation of two different quenchers. The slow component of the light-induced fluorescence decline at 77 K is related to charge accumulation on a non-pigment molecule of the PS II complex. This revised version was published online in June 2006 with corrections to the Cover Date.     

Controls of dissolved organic matter quality: evidence from a large‐scale boreal lake survey

Inland waters transport large amounts of dissolved organic matter (DOM) from terrestrial environments to the oceans, but DOM also reacts en route, with substantial water column losses by mineralization and sedimentation. For DOM transformations along the aquatic continuum, lakes play an important role as they retain waters in the landscape allowing for more time to alter DOM. We know DOM losses are significant at the global scale, yet little is known about how the reactivity of DOM varies across landscapes and climates. DOM reactivity is inherently linked to its chemical composition. We used fluorescence spectroscopy to explore DOM quality from 560 lakes distributed across Sweden and encompassed a wide climatic gradient typical of the boreal ecozone. Six fluorescence components were identified using parallel factor analysis (PARAFAC). The intensity and relative abundance of these components were analyzed in relation to lake chemistry, catchment, and climate characteristics. Land cover, particularly the percentage of water in the catchment, was a primary factor explaining variability in PARAFAC components. Likewise, lake water retention time influenced DOM quality. These results suggest that processes occurring in upstream water bodies, in addition to the lake itself, have a dominant influence on DOM quality. PARAFAC components with longer emission wavelengths, or red‐shifted components, were most reactive. In contrast, protein‐like components were most persistent within lakes. Generalized characteristics of PARAFAC components based on emission wavelength could ease future interpretation of fluorescence spectra. An important secondary influence on DOM quality was mean annual temperature, which ranged between ?6.2 and +7.5 °C. These results suggest that DOM reactivity depends more heavily on the duration of time taken to pass through the landscape, rather than temperature. Projected increases in runoff in the boreal region may force lake DOM toward a higher overall amount and proportion of humic‐like substances.     

Red antenna states of photosystem I from Synechococcus sp. PCC 7002

Absorption, fluorescence and single-molecule spectroscopy at low temperatures were used to elucidate spectral properties, heterogeneities and dynamics of the red-shifted chlorophyll a (Chla) molecules responsible for the fluorescence in photosystem I (PSI) from the cyanobacterium Synechoccocus sp. PCC 7002. The 77 K absorption spectrum indicates the presence of 2–3 red-shifted Chla’s absorbing at about 708 nm. The fluorescence emission spectrum is dominated by a broad band at 714 nm. The emission spectra of single PSI complexes show zero-phonon lines (ZPLs) as well as a broad intensity distribution without ZPLs. The spectral region below 710 nm often shows ZPLs, they form a spectral band with a maximum at 698 nm (F698). The region above 710 nm is dominated by broad intensity distributions and the observation of ZPLs is less frequent. The broad distributions are due to the emission of the C708 Chla’s and the emission from F698 stems from a Chla species absorbing at the blue side of P700. The properties of these two emissions show a close relation to those of the C708 and C719 pools observed in T. elongatus. Therefore an assignment of F698 and C708 to Chla-species with similarities to C708 and C719 in T. elongatus is proposed.     

Red region excitation spectra of protochlorophyllide in dark-grown leaves from plant species with different proportions of its spectral forms

Etiolated leaves of three different species, maize, wheat, and pea, as well as a pea mutant (lip1) were used to compare the excitation spectra of protochlorophyllide (Pchlide) in the red region. The species used have different composition of short-wavelength and long-wavelength Pchlide forms. The relation between different forms was furthermore changed through incubating the leaves in 5-aminolevulinic acid (ALA), which caused an accumulation of short-wavelength Pchlide forms, as shown by changes in absorption and fluorescence spectra. This is the first time a comprehensive comparison is made between excitation spectra from different species covering an emission wavelength range of 675–750 nm using fluorescence equipment with electronic compensation for the variations in excitation irradiance. The different forms of Pchlide having excitations peaks at 628, 632, 637, 650, and 672 nm could be best measured at 675, 700, 710, 725, and 750 nm, respectively. Measuring emission at wavelengths between 675– 710 nm gave an exaggeration of the short-wavelength forms and measuring at longer wavelengths gave for the pea leaves an exaggeration of the 672 nm peak. In general, an energy transfer from short-wavelength Pchlide forms to long-wavelength Pchlide forms occurred, but such an energy transfer sometimes seemed to be limited as a result of a discrete location of the Pchlide spectral forms. The excitation spectra resembling the absorption spectrum most were measured at an emission wavelength of 740 nm. Measuring the excitation at 710 nm gave higher intensity of the spectra but the short-wavelength forms were accentuated.