A fluorescence-based bioassay for aquatic macrophytes and its suitability for effect analysis of non-photosystem II inhibitors

Background, Goals and Scope During the last years the miniaturization of toxicity test systems for rapid and parallel measurements of large quantities of samples has often been discussed. For unicellular algae as well as for aquatic macrophytes, fluorescence-based miniaturized test systems have been introduced to analyze photosystem II (PSII) inhibitors. Nevertheless, high-throughput screening should also guarantee the effect detection of a broad range of toxicants in order to ensure routinely applicable, high-throughput measuring device experiments which can cover a broad range of toxicants and modes of action others than PSII inhibition. Thus, the aim of this study was to establish a fast and reproducible measuring system for non-PSII inhibitors for aquatic macrophyte species to overcome major limitations for use. Methods A newly developed imaging pulse-amplitude-modulated chlorophyll fluorometer (I-PAM) was applied as an effect detector in short-term bioassays with the aquatic macrophyte species Lemna minor. This multiwell-plate based measuring device enabled the incubation and measurement of up to 24 samples in parallel. The chemicals paraquat-dichloride, alizarine and triclosan were chosen as representatives for the toxicant groups of non-PSII herbicides, polycyclic aromatic hydrocarbons (PAHs) and pharmaceuticals and personal care products (PPCPs), which are often detected in the aquatic environment. The I-PAM was used (i) to establish and validate the sensitivity of the test system to the three non-PSII inhibitors, (ii) to compare the test systems with standardized and established biotests for aquatic macrophytes, and (iii) to define necessary time scales in aquatic macrophyte testing. For validation of the fluorescence-based assay, the standard growth test with L. minor (ISO/DIS 20079) was performed in parallel for each chemical. Results The results revealed that fluorescence-based measurements with the I-PAM allow rapid and parallel analysis of large amounts of aquatic macrophyte samples. The I-PAM enabled the recording of concentration-effect-curves with L. minor samples on a 24-well plate with single measurements. Fluorescence-based concentration-effect-curves could be detected for all three chemicals after only 1 h of incubation. After 4–5 h incubation time, the maximum inhibition of fluorescence showed an 80–100% effect for the chemicals tested. The EC50 after 24 h incubation were estimated to be 0.06 mg/L, 0.84 mg/L and 1.69 mg/L for paraquatdichloride, alizarine and triclosan, respectively. Discussion The results obtained with the I-PAM after 24 h for the herbicide paraquat-dichloride and the polycyclic aromatic hydrocarbon alizarine were in good accordance with median effective concentrations (EC50s) obtained by the standardized growth test for L. minor after 7 d incubation (0.09 mg/L and 0.79 mg/L for paraquat-dichloride and alizarine, respectively). Those results were in accordance with literature findings for the two chemicals. In contrast, fluorescence-based EC50 of the antimicrobial agent triclosan proved to be two orders of magnitude greater when compared to the standard growth test with 7 d incubation time (0.026 mg/L) as well as with literature findings. Conclusion Typically, aquatic macrophyte testing is very time consuming and relies on laborious experimental set-ups. The I-PAM measuring device enabled fast effect screening for the three chemicals tested. While established test systems for aquatic macrophytes need incubation times of ≥ 7 d, the I-PAM can detect inhibitory effects much earlier (24 h), even if inhibition of chemicals is not specifically associated with PSII. Thus, the fluorescence-based bioassay with the I-PAM offers a promising approach for the miniaturization and high-throughput testing of chemicals with aquatic macrophytes. For the chemical triclosan, however, the short-term effect prediction with the I-PAM has been shown to be less sensitive than with long-term bioassays, which might be due to physicochemical substance properties such as lipophilicity. Recommendations and Perspectives The results of this study show that the I-PAM represents a promising tool for decreasing the incubation times of aquatic macrophyte toxicity testing to about 24 h as a supplement to existing test batteries. The applicability of this I-PAM bioassay on emergent and submerged aquatic macrophyte species should be investigated in further studies. Regarding considerations that physicochemical properties of the tested substances might play an important role in microplate bioassays, the I-PAM bioassay should either be accompanied by evaluating physicochemical properties modeled from structural information prior to an experimental investigation, or by intensified chemical analyses to identify and determine nominal concentrations of the toxicants tested. The chemicals paraquat-dichloride, alizarine and triclosan were chosen as representatives for the toxicant groups of non-PSII herbicides, PAHs and PPCPs which are often detected in the aquatic environment. Nevertheless, in order to ensure a routinely applicable measuring device, experiments with a broader range of toxicants and samples of surface and/or waste waters are necessary. ESS-Submission Editor: Dr. Markus Hecker (MHecker@Entrix.com)     

Energy,food, and land — The ecological traps of humankind

Humans’ superiority over all other organisms on earth rests on five main foundations: command of fire requiring fuel; controlled production of food and other biotic substances; utilization of metals and other non-living materials for construction and appliances; technically determined, urban-oriented living standard; economically and culturally regulated societal organization. The young discipline of ecology has revealed that the progress of civilization and technology attained, and being further pursued by humankind, and generally taken for granted and permanent, is leading into ecological traps. This metaphor circumscribes ecological situations where finite resources are being exhausted or rendered non-utilizable without a realistic prospect of restitution. Energy, food and land are the principal, closely interrelated traps; but the absolutely decisive resource in question is land whose increasing scarcity is totally underrated. Land is needed for fulfilling growing food demands, for producing renewable energy in the post-fossil and post-nuclear era, for maintaining other ecosystem services, for urban-industrial uses, transport, material extraction, refuse deposition, but also for leisure, recreation, and nature conservation. All these needs compete for land, food and non-food biomass production moreover for good soils that are scarcer than ever. We are preoccupied with fighting climate change and loss of biodiversity; but these are minor problems we could adapt to, albeit painfully, and their solution will fail if we are caught in the interrelated traps of energy, food, and land scarcity. Land and soils, finite and irreproducible resources, are the key issues we have to devote our work to, based on careful ecological information, planning and design for proper uses and purposes. The article concludes with a short reflection on economy and competition as general driving forces, and on the role and reputation of today’s ecology. Updated version of the keynote lecture presented at the EcoSummit 2007 in Beijing, China, May 24. The article is gratefully dedicated to the memory of my late colleague and friend Frank B. Golley.     

Distribution of elements in needles of Pinus massoniana (Lamb.) was uneven and affected by needle age

Macronutrients (P, S, K, Na, Mg, Ca), heavy metals (Fe, Zn, Mn, Cu, Pb, Cr, Ni, Cd,) and Al concentrations as well as values of Ca/Al in the tip, middle and base sections, and sheaths of current year and previous year needles of Pinus massoniana from Xiqiao Mountain were analyzed and the distribution patterns of those elements were compared. The results indicated that many elements were unevenly distributed among the different components of needles. Possible deficiency of P, K, Ca, Mn and Al toxicity occurred in needles under air pollution. Heavy metals may threaten the health of Masson pine. Needle sheaths were good places to look for particulate pollutants, in this case including Fe, Cu, Zn, Pb, Cr, Cd and Al.     

催化湿式氧化技术处理山梨酸生产废水的研究

采用催化湿式氧化技术处理生产山梨酸过程中产生的高浓度有机废水,对催化剂组分进行优选.对反应温度、O2分压(Po2)和废水pH等工艺条件进行考察.实验结果表明:采用自制CuO-Cr2O3-La2O3/TiO2为复合催化剂处理该有机废水时表现出较好的催化活性;在190℃、Po2=2.1 MPa、pH=6.1、COD=10 030.0 mg/L时,反应90 min的COD去除率达到98.3%,而在相同条件下未加催化剂的湿式氧化的COD去除率只有60.1%.     

Perfluorinated compounds in the Pearl River and Yangtze River of China

A total of 14 perfluorinated compounds (PFCs) were quantified in river water samples collected from tributaries of the Pearl River (Guangzhou Province, south China) and the Yangtze River (central China). Among the PFCs analyzed, perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) were the two compounds with the highest concentrations. PFOS concentrations ranged from 0.90 to 99 ng/l and <0.01–14 ng/l in samples from the Pearl River and Yangtze River, respectively; whereas those for PFOA ranged from 0.85 to 13 ng/l and 2.0–260 ng/l. Lower concentrations were measured for perfluorobutane sulfonate (PFBS), perfluorohexane sulfonate (PFHxS), perfluorooctanesulfoamide (PFOSA), perfluorohexanoic acid (PFHxA), perfluoroheptanoic acid (PFHpA), perfluorononaoic acid (PFNA), perfluorodecanoic acid (PFDA), and perfluoroundecanoic acid (PFUnDA). Concentrations of several perfluorocarboxylic acids, including perfluorododecanoic acid (PFDoDA), perfluorotetradecanoic acid (PFTeDA), perfluorohexadecanoic acid (PFHxDA) and perfluorooctadecanoic acid (PFOcDA) were lower than the limits of quantification in all the samples analyzed. The highest concentrations of most PFCs were observed in water samples from the Yangtze River near Shanghai, the major industrial and financial centre in China. In addition, sampling locations in the lower reaches of the Yangtze River with a reduced flow rate might serve as a final sink for contaminants from the upstream river runoffs. Generally, PFOS was the dominant PFC found in samples from the Pearl River, while PFOA was the predominant PFC in water from the Yangtze River. Specifically, a considerable amount of PFBS (22.9–26.1% of total PFC analyzed) was measured in water collected near Nanjing, which indicates the presence of potential sources of PFBS in this part of China. Completely different PFC composition profiles were observed for samples from the Pearl River and the Yangtze River. This indicates the presence of dissimilar sources in these two regions.     

A study of climate change and anthropogenic impacts in West Africa

BACKGROUND, AIM AND SCOPE: During the last decades ecological conditions in West Africa have dramatically changed. Very evident is the climate change, which has resulted in a southward shift of the climate zones, e.g. a spread of the desert (Sahara) into the Sahelian zone. After the drought period of the early 1970s and 1980s, livestock density increased resulting in an intensification of grazing pressure. This anthropogenous phenomenon leads to similar landscape changes as those caused by the climate. Only very few investigations exist on vegetation dynamics, climate changes and land use changes for the Sudanian zone. The paper presents data on changes of precipitation, of land use, of the geographical range of species, and of the composition of the flora, which have to be regarded as proofs of the sahelisation of large areas of the Sudanian zone. MATERIALS AND METHODS: Area of investigation: Burkina Faso. Precipitation data analysis: precipitation data from 67 stations; time series analysis and geo-statistical spatial interpolation. Analysis of land use change: Landsat satellite MSS and ETM+ data, acquired for two different dates between 1972 and 2001 analyzed by the software ERDAS/IMAGINE version 8.6 and ArcView 3.2 with the Spatial Analyst extension. Intensive ground truthing (160 training areas). Inventory of the flora: based on the data of the Herbarium Senckenbergianum (FR) in Frankfurt, Germany, and of the herbarium of the university of Ouagadougou (OUA), Burkina Faso, as well as on various investigations on the vegetation of Burkina Faso carried out in the years 1990 to 2005 by the team of the senior author. Life form analysis of the flora: based on the inventory of permanent plots. RESULTS AND DISCUSSION: Precipitation: Remarkable latitudinal shift of isohyets towards the South translates to a general reduction of average rainfall in great parts of the country. The last decade (1990-1999) shows some improvement, however, the more humid conditions of the 1950's and 1960's are not yet established again. Landcover change: In the study region the extent of arable fields and young fallows increased during the last 30 years from 580 km(2) in 1972 to 2870 km(2) in 2001. This means an average land cover conversion rate of 0.9% per year for the 6 departments considered. Change of the distribution of Sahelian and Sudanian species: Several species, mentioned in older literature as strictly Sahelian, today also occur in the Sudanian zone. Parallel to the spread of former strictly Sahelian species into the Sudanian zone, some former Sahelo-Sudanian species have withdrawn from the Sahel. Changes of the life form spectra of the flora: Considering their life form spectra, the flora of heavily grazed and of protected areas in the Sudanian zone show great differences. On areas intensively grazed the percentage of therophytes is evidently higher than on protected areas. Just the opposite is true for the phanerophytes. Their percentage is higher on the protected area than on the grazed zones. At the first glance, it is obvious to link the changes in flora and vegetation with the climate changes that have occurred during the last five decades (decrease of annual precipitation). However, not only climatic conditions have changed, but also population has increased, the percentage of land intensively used for agriculture and pasturing has increased and the time for soil regeneration today is much shorter than it was some decades ago. Thus, the landscape of the Sudanian zone has become a more Sahelian character. A comparison of the flora of an intensively used area of the Sudanian zone with that of a protected area shows a remarkable change in the life form spectra. The spectrum of the intensively used area is almost identical with that of the typical Sahelian flora. This comparison shows that the anthropogenic influence plays a greater role in the sahelisation of the Sudanian zone than the climate change. CONCLUSION: Climate change and anthropogenic influence both, lead to a sahelisation of landscape and flora. Thus in many parts of the Sudanian zone of West Africa sahelisation phenomena will remain and even increase independently from the reestablishment of the more humid climate conditions of the 1950ies. RECOMMENDATIONS AND PERSPECTIVES: In order to maintain some parts of the characteristic Sudanian landscape with its characteristic flora and vegetation, the number and size of protected areas should be augmented. For all protected areas it has to be ensured, that protection is reality, i.e. respected an understood by local people, not only fiction. As long as the enlargement of intensively used areas continues the sahelisation of flora, vegetation and landscape will continue too.     

Degradation of pentachlorophenol by pure and mixed cultures in two different soils

GOAL, SCOPE AND BACKGROUND: Pentachlorophenol (PCP) is the second highest volume pesticide used in the United States. It is a mutagenic compound whose exposure poses significant health effects, One of the most desirable, environmentally friendly treatment methods is bioremediation. For soil-based contamination, the effectiveness of bioremediation will also be affected by the presence of an active indigenous population, sorption of the contaminant onto the soil, and environmental parameters. METHODS: Two pure strains and their mixed culture were used to evaluate PCP biodegradation in two different field soils, Columbia (CO) and New Mexico (NM). Biostimulation of the indigenous microbes was evaluated by adding nutrients. The efficiency of adding bacteria strains (bioaugmentation) for degrading PCP was determined with Arthrobacter sp., Flavobacterium sp. and a 50:50 mixture of the two bacteria strains. RESULTS: In CO soil, only 24%, 12% and 25% of the initial PCP concentration were degraded by Flavobacterium sp., Arthrobacter sp. and mixed culture, respectively. Arthrobacter sp. was used in NM soil with two initial concentrations and achieved degradation efficiencies of 57% and 61% for 361 and 95 mg kg- concentrations, respectively. Discussion. Analysis via statistical methods showed that the bacteria had different efficiencies on PCP degradation in each soil. 2 CONCLUSIONS: All bacteria catalyzed a higher PCP degradation when present in NM soil. Second, Flavobacterium sp. degraded more PCP than Arthrobacter sp. in CO soil. The mixed culture achieved the highest degradation efficiency regardless of the initial concentration or soil origin. RECOMMENDATIONS AND PERSPECTIVES: The effect of the soil properties, such as the soil organic matter (SOM) on PCP biodegradation should be investigated. Future work can also investigate the effect of aging time on biodegradation.     

Bioremediation process on Brazil shoreline

GOAL, SCOPE AND BACKGROUND: Bioremediation technique can be considered a promising alternative to clean oil spills using microbial processes to reduce the concentration and/or the toxicity of pollutants. To understand the importance of this work we must know that there is only little research performed to date using bioremediation techniques to clean oil spills in tropical countries. So, the main objective of this work is to analyze the behavior of a laboratory's bioremediation test using nutrients on coastal sediments. METHODS: The bioremediation process is followed through geochemical analysis during the tests. This organic material is analyzed by medium pressure liquid chromatography (MPLC), gas chromatography/flame ionization detection (GC/FID) and gas chromatography/ mass spectrometry. By microbial counting, the number of total bacteria and degrading bacteria is determined during the experiments, in order to confirm the effectiveness of the bioremediation process. The seawater obtained throughout the bioremediation process is analyzed for nutrients grade (phosphate and ammonium ions) and also for its toxicity (Microtox tests) due the presence of hydrocarbons and fertilizer. RESULTS: The results from the geochemical analyses of the oil show a relative decrease in the saturated hydrocarbon fraction that is compensated by a relative enrichment on polar compounds. It's confirmed by the fingerprint evaluation where it is possible to see a complete reduction of the normal alkanes followed by isoprenoids. Seawater analysis done by toxicity and nutrients analysis, such as microbial counting (total and degrading bacteria), confirm the fertilizer effectiveness during the bioremediation process. DISCUSSION: Results from simulating test using NPK, a low-price plant fertilizer, suggest that it's able to stimulate the degradation process. Results from medium pressure liquid chromatography (MPLC), done at two different depths (surface and subsurface), show different behavior during the biodegradation process where the later is seen to be more susceptible to microbial attack. Data from bioremediation unit shows a bigger reduction of the saturated fraction, followed by some smaller reduction of aromatic fractions, compensated by a relative increase from polar compounds (NSO). n-C17/pristane, n-C18/ fitane and pristane/fitane rates show constant values for the unity control, different from bioremediation samples which have a significant reduction, especially on subsurface areas, where a strong fall in the rates, seen to be reduced to zero over twenty days, had occurred during the first ten days. However, sample surfaces are reduced to zero in thirty days of experiments, proving that biodegradation is better on subsurfaces. Gaseous chromatography/mass spectrometry (CG/MS) analysis shows constant values to cyclic biomarker rates and aromatic compounds, suggesting that the biodegradation process is not strong enough to reduce these composites. Microbial analysis shows a reduction on heterotrophic (total bacteria) number from control unit, probably because the bacteria uses the spill oil like carbon source and energy. However, the number increases on bioremediation unit, because it uses NPK like a biostimulator. The hydrocarbonoclastic number isn't enough on the first moment, but it's detected after 30 days and quantified in all units, showing big values especially in bioremediation. Toxicity tests confirm that NPK fertilizer does not intoxicate the shoreline during the application of the bioremediation technique. Some nutrient concentration shows high values of ammonium and phosphate per bioremediation unit, reducing by the end of the experiment. CONCLUSIONS: Results reached the goal, finding a proper nutrient (NPK fertilizer) to stimulate the biodegradation process, growing bacteria responsible for reducing impact-contaminated coast ambient by oil spills. Chemical analysis of oil shows a reduction in the saturated fraction with a relative enrichment in polar composites (NSO) and the aromatic fraction from oil remaining constant. Subsurface samples show more biodegradation than surface samples, probably because the first one has higher humidity. Linear alcanes are more biodegraded than isoprenoids, confirming the biodegradation susceptibility order. Saturated cyclic biomarkers and aromatic compounds show constant behavior maybe because the nutrients or time was not enough for microorganismic attack. Fertilizer does not demonstrate any toxic effects in local biota so that it does not compromise the technique applicability and the environment is not saturated by nutrients during the simulation, especially since the coastal environment is an open system affected daily by tides. Therefore, bioremediation tests can be classified as moderate, reaching level 5 in the classification scale by Peters & Moldowan (1993). RECOMMENDATIONS AND PERSPECTIVES: The use of marine environment by the petroleum industry on exploration, production and transportation operation, transform this oil to become the most important pollutant in the oceans. Bioremediation is an important technique used to clean spilled oil impacting on shorelines, accelerating the biodegradation process by using fertilizer growing the microorganisms responsible for decontaminating the environment. We recommend confirming the efficiency of NPK nutrient used on bioremediation simulating experiments on beaches, while monitoring the chemical changes long-term. NPK fertilizer can be used to stimulate the biodegradation process on shoreline impacted by spilled oil.     

Effects of molecular weight and concentration of carboxymethyl cellulose on morphology of hydroxyapatite nanoparticles as prepared with one-step wet chemical method

Nano-sized apatite particles (nAP) synthesized with carboxymethyl cellulose (CMC) have shown great application potentials in in situ heavy metal remediation. However, differences in CMC’s properties effects on the size of nAP produced are not well understood. In this paper, two types of CMC, with respective molecular weights (MW) of ～120000 and ～240000 Dalton or respective polymerization degrees of 500 (CMC-500) and 1050 (CMC-1050), were studied in a concentration range of 0.05%–0.5% (w/w) for nAP synthesis. Morphology of the particles was characterized with transmission electron microscopy (TEM). Results showed that 0.05% CMC-500 solution gave an average particle size of 148.7±134.9 nm, 0.25% CMC-500 solution produced particles of 21.8±20.4 nm, and, 0.5% CMC-500 solution contained particles of 15.8±7.7 nm. In comparison, 0.05% CMC-1050 solution produced nanoparticles of 6.8±3.2 nm, 0.25% CMC-1050 produced smaller nAP of 4.3±3.2 nm, and 0.5% CMC-1050 synthesized the smallest nanoparticles in this study, with an average diameter of 3.0±2.1 nm. Chemical composition of the products was identified with X-ray diffraction (XRD) as pure hydroxyapatite. Interactions between nAP and CMC were discussed with help of attenuated total reflection Fourier transform infrared (ATRFTIR) spectroscopic data. This study showed that CMC at higher concentration as well as higher MW facilitated to produce finer nanoparticles, showing that nAP size could be manipulated by selecting appropriate CMC MW and/or applying appropriate CMC concentration.