Aquatic Moss Fontinalis Antipyretica Research Articles

Influence of storage method on the content of photosynthetic pigments of the aquatic moss Fontinalis antipyretica

Aquatic bryophytes are frequently used as bioindicators of water quality in rivers. Poor water quality increases physiological stress in moss, and stress levels can be estimated by measuring the concentrations of photosynthetic pigments and by calculating pigmentary indices. However, studies collecting many samples in a short time may need to store them until analysis. In the meantime, the pigments may suffer some degradation. Given the lack of studies on this problem for aquatic bryophytes, this one investigates the effect of three storage methods (refrigerated at 6 °C, frozen at − 20 °C and dried at room temperature) on the concentrations of chlorophyll a, chlorophyll b, pheophytin a and the following ratios: chlorophyll a/chlorophyll b, D430/D410, D665/D665a, D430/D665 and D480/D665 (where Dx is the absorbance at x nm) in the aquatic moss Fontinalis antipyretica. The results showed that refrigeration was the most suitable method, freezing might be suitable for some parameters, and drying was inadequate for this purpose. We recommend that all studies on photosynthetic pigments detail the time elapsed from collection to sample analysis, as well as the storage method used.

Unravelling the metal uptake process in mosses: Comparison of aquatic and terrestrial species as air pollution biomonitors

Transplanted mosses have been widely shown to be excellent tools for biomonitoring air pollution; however, it is not clear how the functional groups present on their surfaces affect the uptake of metal cations. In the present study, we examined differences in trace metal accumulation in two terrestrial and one aquatic moss species, and investigated whether the differences depended on their physico-chemical characteristics. In the laboratory, we determined C, N and H contents in their tissues and obtained the ATR-FTIR spectra (to identify the presence of functional groups). We also conducted surface acid-base titrations and metal adsorption assays with Cd, Cu and Pb. In the field, we exposed transplants of each species near different air-polluting industries, and determined the mosses enrichment of Al, Cd, Co, Cr, Cu, Fe, Ni, Pb and V. Laboratory results demonstrated higher metal uptake capacity in the terrestrial mosses Sphagnum palustre and Pseudoscleropodium purum, compared to that in the aquatic moss Fontinalis antipyretica, which can be attributed to a greater abundance of acidic functional groups (i.e. negatively charged binding sites) on the surface of the terrestrial mosses. The affinity of moss for certain elements depends on the abundance and nature of surface functional groups. Accordingly, the metal concentrations generally reached higher levels in S. palustre transplants compared to the other species, except for the uptake of Hg, which was higher in F. antipyretica. However, the findings also suggest an interaction between the type of environment (terrestrial or aquatic) and the moss characteristics that may influence the abovementioned trend. Thus, irrespective of the physico-chemical characteristics, metal uptake varied depending on the environment of origin of the mosses “i.e. atmospheric or aquatic”. In other words, the findings suggest that species that accumulate more metals in terrestrial environments will accumulate lower amounts of metals in aquatic environments and vice versa.

Influence of the storage procedure on the trace element content measured in the aquatic moss Fontinalis antipyretica Hedw

Aquatic bryophytes have been used as pollution biomonitors for decades. Despite this, sample collection and preparation methods have not been standardized, which makes it difficult to compare the results of different studies. Most times, the samples have to be stored before processing, for example, when many of them are collected in a short time, as occurs in extensive pollution studies. Storage must be done in a way that does not change the pollutant concentrations in the samples. We studied whether the concentrations of Al, As, Ba, Cd, Cr, Cu, Fe, Hg, Mn, Ni, Pb, and Zn in the aquatic moss Fontinalis antipyretica were affected by three storage procedures: dry at room temperature, fresh (in refrigerator at 6°C), and frozen at -20°C. In addition, we evaluated whether the subsequent washing of the samples affected the concentrations of these elements differently depending on the storage method. Our results showed that the three methods were, in general, adequate since the concentrations did not change, and we did not observe differences between washed and unwashed samples either. Since the simplest method is refrigeration, we concluded that this is the best of them. However, the concentrations of Hg increased steadily over time in the fresh material, probably because of redistribution after volatilization from the basal parts of the mosses. We believe that the respiration of the plants lowered the concentrations of oxygen inside the hermetically sealed bags containing the samples, thus promoting the reduction of the Hg and its posterior volatilization and redistribution. We did not observe interactions between the storage method and the posterior washing of the samples. PRACTITIONER POINTS: We studied the influence of storage procedure on element content in aquatic mosses. The procedures were as follows: dried at room temperature, stored in refrigerator, and frozen. The procedures did not change the element contents, except for Hg in refrigerator. All the procedures seemed suitable, but refrigeration was the simplest one.

Toxic effects of a mixture of five pharmaceutical drugs assessed using Fontinalis antipyretica Hedw.

The potential health risks associated with the pharmaceuticals released into the environment through effluents from sewage treatment plants have become a major cause for concern. Owing to the lack of effective indicators, monitoring the concentration of these pollutants in the aquatic environment is challenging. The aim of this study was to assess the toxicity of a mixture of five pharmaceutical drugs (paracetamol, carbamazepine, diclofenac, irbesartan, and naproxen) using the aquatic moss Fontinalis antipyretica as a bioindicator and bioaccumulator. We examined the effects of the drug mixture on the cellular antioxidant system, chlorophyll content, and morphological traits of F. antipyretica. The plant was exposed for 5 months to three concentrations of the mixture, including the environmental concentration (MX1), and 10- (MX10) and 100-times (MX100) the environmental concentration. The results showed that only carbamazepine and irbesartan were accumulated by the species. The bioconcentration level increased with exposure time, with the maximum uptake at the 4th month of exposure. The increase in bioaccumulation with exposure time was more evident in plants exposed to MX100. Analysis of the activity of antioxidant enzymes showed that superoxide dismutase (SOD, EC 1.15.1.1.) and catalase (EC 1.11.1.6.) were highly sensitive to the drug mixture. The activity of the enzymes was significantly higher in plants exposed to MX100; however, the activity of guaiacol peroxidase (GPX, EC 1.11.1.7.) was not significantly affected. Plants exposed to MX10 and MX100 had significantly lower total chlorophyll content and chlorophyll a/b ratio compared with those of plants in the control group; however, photosynthetic activity was restored after 5 months of exposure. The morphological characteristics of F. antipyretica were less sensitive to the treatment conditions.

Effect of Current Velocity on Cd Accumulation in the Aquatic Moss Fontinalis antipyretica

With the aim of further standardizing biomonitoring techniques with aquatic mosses, the relationship between the velocity of water flow and cadmium (Cd) accumulation in transplants of the moss Fontinalis antipyretica was investigated. For this purpose, moss transplants were exposed in a controlled aquatic environment to different concentrations of Cd (0, 4, 16 and 36 ng g−1) and different water velocities (10, 30, 50, 70 and 90 cm s−1). The Cd concentrations in the moss transplants mainly depended on the Cd concentration in the water, but a small fraction of the variance was explained by water velocity. The Cd concentrations in moss were standardized to remove the effect of the concentration in the water so all the data could be analyzed together. The regression model for the standardized concentrations explained 23% of the variance in Cd accumulation in F. antipyretica and water velocity proved to be a significant predictor of Cd accumulation.

Optimization of the Biomonitoring Technique with the Aquatic Moss Fontinalis antipyretica Hedw.: Selection of Shoot Segment Length for Determining Trace Element Concentrations

Bryophytes, including Fontinalis antipyretica Hedw., are widely used for biomonitoring aquatic environments. However, some methodological aspects, such as the fraction of shoot analysed, have not yet been standardized. The main objective of this study was to establish the length of segments of F. antipyretica shoots that should be used for trace element determination in monitoring studies. For this purpose, we determined the concentrations of 12 different trace elements (11 metals and one metalloid) in five successive 1 cm-long segments of the shoots of F. antipyretica collected from four different locations. We found that the concentrations of most elements increased steadily towards the basal part of the shoots without becoming stable. Sometimes, these increasing trends were concealed by another underlying trend of increasing dry/fresh weight ratios, caused by morphological changes (leaf density, shoot thickness) related with shoot development. The concentration trends were due to factors such as the deposition of Mn oxides on the moss surface. This element showed the largest increases in concentration towards the basal part of the shoot and its concentrations were closely correlated with those of most of the other elements. Considering the results obtained, we recommend the use of 3 cm-long apical segments for determining trace element concentrations in F. antipyretica. This recommendation is based on the following: (1) The change in dry weight beyond this length of segment is small; (2) the workload and the quantity of material to collect in the river is acceptable; (3) Most previous studies have used shoot segments of similar lengths.

Long-established and new active biomonitors jointly reveal potentially toxic element gradients across spatial scales in freshwater ecosystems

At the core of the adaptive ecosystem management paradigm, environmental monitoring, especially using biomonitors, mandates the search for tools and approaches to overcome its current limitations, both in terms of novel species and analytical techniques to extract information from the data. In this context, a large field biomonitoring study was performed in order to validate a candidate active biomonitor, the macrophytic alga Chara gymnophylla, using a long-established biomonitor (the aquatic moss Fontinalis antipyretica) as reference, innovative spatial analysis approaches and the area of two of the main freshwater ecosystems of the “Cilento Vallo di Diano e Alburni” National Park as experimental setting. Specifically, the concentrations of 19 potentially toxic elements (Al, As, Ca, Cd, Co, Cr, Cu, Fe, K, Mg, Mn, Na, Ni, P, Pb, S, Si, V, Zn) in 3 alga and 3 moss bags placed in each of 41 sites along the Bussento and Calore Salernitano rivers were analyzed. The coherence of element gradients between biomonitors, the spatial heterogeneity in element distribution and their spatial scales were evaluated using robust multivariate approaches, random models and spectral decomposition of the spatial gradients. Results demonstrate the substantial uniformity in responses between the biomonitors, both showing similar element spatial distributions, within and between-site variability, the same spatial connectivity and spatial variations at similar scales. On the one hand, the approach adopted and the findings obtained allow definitely validating Ch. gymnophylla as a novel active biomonitor of potentially toxic elements in freshwater ecosystems and, on the other hand, deriving clear contamination scenarios, with indication of the alleged sources.

Developing a Biotechnological Tool for Monitoring Water Quality: In Vitro Clone Culture of the Aquatic Moss Fontinalis Antipyretica

One of the main factors limiting active biomonitoring with aquatic mosses is the lack of sufficient material. A laboratory culture of the moss would solve this problem and thus convert the technique into a valuable biotechnological tool for monitoring water quality. With this aim, we first established small and large scale axenic in vitro culture systems for the aquatic moss Fontinalis antipyretica. We then attempted to enhance the growth rate of the cultures by modifying temperature, photoperiod and medium composition (N:P ratio, P concentration, CO2 supply, NH4NO3 supply and sucrose supply). None of these modifications greatly increased the in vitro growth rate. However, the growth rates were sufficiently high (relative to the initial weight of the cultures) in both systems (45 and 6 mg·day−1·g−1 for flasks and bioreactors respectively) to enable the production of large amounts of material. The ability to culture the material will substantially improve the applicability of the moss bag technique.

Biomonitoring freshwater FISH farms by measuring nitrogen concentrations and the δ15N signal in living and devitalized moss transplants

The trophic balance of freshwater aquaculture activities has traditionally been monitored by chemical analysis of water; however, the parameters measured are usually characterized by high temporal variability. Aquatic mosses can be used as biomonitors as they integrate both continuous and episodic contamination events. Here we report, for the first time, a method for monitoring N enrichment in the surroundings of fish farms by measuring the N content and isotopic signal (δ15N) of transplanted living and devitalized specimens of the aquatic moss Fontinalis antipyretica. For this purpose, moss samples (“moss bags”) were exposed at increasing distances (10, 100, 300 and 1000 m) up- and downstream of the effluent discharge points of four trout farms, for 10 and 30 days. The low natural (background) variability in δ15N in upstream samples enabled detection of outlier values, caused by aquaculture discharges, at distances of 10 and 100 m downstream, especially in devitalized moss and after 10 days of exposure. However, the unexpectedly low N contents of moss samples exposed close to the discharge points complicates interpretation of the high levels of N forms detected by conventional physicochemical analysis of water. Although the mechanisms that modify N parameters in moss tissues were not clear, measurement of the isotopic signal δ15N in devitalized moss exposed for 10 days proved useful for monitoring the N pollution associated with intensive freshwater aquaculture.

Using devitalized moss for active biomonitoring of water pollution

This paper presents the results of an experiment carried out for the first time in situ to select a treatment to devitalize mosses for use in active biomonitoring of water pollution. Three devitalizing treatments for the aquatic moss Fontinalis antipyretica were tested (i.e. oven-drying at 100 °C, oven-drying with a 50-80-100 °C temperature ramp, and boiling in water), and the effects of these on loss of material during exposure of the transplants and on the accumulation of different heavy metals and metalloids were determined. The suitability of using devitalized samples of the terrestrial moss Sphagnum denticulatum to biomonitor aquatic environments was also tested. The structure of mosses was altered in different ways by the devitalizing treatments. Devitalization by boiling water led to significantly less loss of material (p < 0.01) than the oven-drying treatments. However, devitalization by oven-drying with a temperature ramp yielded more stable results in relation to both loss of material and accumulation of elements. With the aim of standardizing the moss bag technique, the use of F. antipyretica devitalized by oven-drying with a temperature ramp is recommended, rather than other devitalization treatments or use of S. denticulatum.

Kinetic modelling of cadmium and lead removal by aquatic mosses

Because biosorption is a low cost and effective method for treating metal-bearing wastewaters, understanding the process kinetics is relevant for design purposes. In the present study, the performance of the aquatic moss Fontinalis antipyretica for removing cadmium and lead from simulated wastewaters has been evaluated. Five kinetic models (first-order, pseudo-first-order, Elovich, modified Ritchie second-order and pseudo-second-order) were fitted to the experimental data and compared. Previously, the effect of parameters such as the initial solution pH, contact time, and initial metal ion concentration on biosorption was investigated. The initial pH of the solution was found to have an optimum value in the range of 4.0-6.0. The equilibrium sorption capacity of cadmium and lead by Fontinalis antipyretica increased with the initial metal concentration. For an initial metal concentration of 10 mg L-1, the uptake capacity of the moss, at equilibrium, is the same for both metals (4.8 mg g-1). Nevertheless, when the initial concentration increases up to 100 mg L-1, the uptake of Pb(II) was higher than 78%. The pseudo-second order biosorption kinetics provided the better correlation with the experimental data (R² ≥ 0.999).

Physiological Effects of Exposure to Arsenic, Mercury, Antimony and Selenium in the Aquatic Moss Fontinalis antipyretica Hedw.

Laboratory experiments were carried out to determine the effects of exposure to different concentrations of As, Hg, Sb and Se on photosynthetic and respiratory rates and on photosynthetic efficiency in the aquatic bryophyte Fontinalis antipyretica Hedw. Specimens of the moss, collected from a clean site, were incubated in solutions of As, Hg, Sb and Se (at concentrations ranging from 0.1 μg l−1 to 10,000 μg l−1) for up to 22 days. The photosynthetic and respiratory rates were then determined by the light/dark bottle technique, and the photosynthetic efficiency was measured by the saturation pulse method. Although different responses were observed in relation to the concentration of the elements, clear responses in net photosynthesis and photosynthetic efficiency were generally only observed in the moss exposed to the highest concentrations of these elements in solution. Mercury was apparently the most toxic of the elements studied. Net photosynthesis and photosynthetic efficiency were also related to tissue concentrations of these elements in the moss. Despite the higher toxicity of Hg, this element can be accumulated at high concentrations in moss, probably at extracellular sites. For Sb, the same tissue concentration had very different physiological effects depending on the initial concentration to which the moss was exposed in solution. Temporal trends in chlorophyll fluorescence were more stable than trends in net photosynthesis. The respiratory rate was very variable and was not clearly related to the concentration of elements in solution or in moss tissues.

Biomonitoring urban fluvial contamination on the basis of physiological stress induced in transplants of the aquatic moss Fontinalis antipyretica Hedw.

We examined the effects of urban-derived fluvial contamination on aquatic bryophytes with the aim of improving use of these organisms for active biomonitoring of this type of contamination. Samples of the aquatic moss Fontinalis antipyretica were transplanted to five sites of a river affected by different levels of urban contamination. The aquatic contamination was characterized by a series of physical and chemical parameters, which were then related to the physiological responses determined in transplants of the bryophyte. Comparison of the data revealed the physiological parameters that best differentiated between levels of contamination. The active biomonitoring technique proved to be a clear and rapid method of determining the existence of a contamination gradient on the basis of the degree of pigmentary and photosynthetic stress in the moss transplants. Of the parameters considered, the concentration of phaeopigments and the net photosynthetic rate provided the clearest responses.

Fractionation of trace metals in a contaminated freshwater stream using membrane filtration, ultrafiltration, DGT and transplanted aquatic moss

Four metal speciation and fractionation techniques – DGT (diffusive gradients in thin films), 1-kDa ultrafiltration, 0.22-µm membrane filtration and aquatic moss – were simultaneously applied to a small, contaminated freshwater stream in northern Sweden to investigate differences and similarities between the methods regarding trace metal speciation and their dependence on geochemical water properties. The investigated metals comprise Al, Cd, Co, Cu, Fe, Mn, Ni, and Zn. The normal DGT devices with Chelex cation exchanger were used. Shoots from the aquatic moss Fontinalis antipyretica L ex Hedw. were collected in a non-polluted brook and transplanted to the sampling site for exposure. It was evident that 0.22-µm membrane filtration, 1-kDa ultrafiltration and DGT generally measured different metal fractions where &lt;1-kDa ultrafiltered concentrations were lower than DGT labile concentrations which in turn were lower than &lt;0.22-µm concentrations. The differences between DGT and &lt;1-kDa concentrations indicate the occurrence of labile colloids discriminated by ultrafiltration. Strong correlations between DGT and &lt;1 kDa concentrations were found for Al, Cu, Cd, Co and Zn. Despite a rigorous sample cleaning, retention of particulate matter on the moss samples was revealed by a significant correlation between metal concentrations in moss and particulate Fe. Generally, elevated trace metal concentrations were found in moss exposed at the sampling site compared to reference moss from the non-polluted brook. No significant correlations were found between DGT-labile concentrations and moss concentrations. Supplementary material: DGT – labile concentrations of Al, Cd, Co, Cu, Fe, Mn, Ni and Zn from deployments in Gråbergsbäcken Stream is available online at http://www.geolsoc.org.uk/SUP18566 . A hard copy can be obtained from the Society Library.

Selenium and its species in the aquatic moss Fontinalis antipyretica

The ability of the widely distributed aquatic moss Fontinalis antipyretica to take up Se from water was studied. Nine locations in the Notranjska region (Slovenia) with different land use in the catchment were sampled for water and moss in the year 2010 in spring, summer and autumn. The concentrations of Se in water at all locations did not exceed 0.2ngmL−1. F. antipyretica took up Se in the range between 345 and 2250ngg−1. All results for Se are expressed on dry matter basis. The Se content varied depending on the location and season. The highest concentration (2250±170ngg−1) of Se was found in the Žerovniščica stream that flows through an agricultural area with dairy farming. The fraction of insoluble Se compounds in the residue after enzymatic hydrolysis using protease (XIV) was around 75%. Soluble Se compounds in the enzymatic extract of F. antipyretica were separated and measured using HPLC coupled to ICP-MS. Se(IV) and Se(VI) were found but no organic Se compounds were detected, even at the highest concentration.

The impact of dehydration rate on the production and cellular location of reactive oxygen species in an aquatic moss.

The aquatic moss Fontinalis antipyretica requires a slow rate of dehydration to survive a desiccation event. The present work examined whether differences in the dehydration rate resulted in corresponding differences in the production of reactive oxygen species (ROS) and therefore in the amount of cell damage. Intracellular ROS production by the aquatic moss was assessed with confocal laser microscopy and the ROS-specific chemical probe 2,7-dichlorodihydrofluorescein diacetate. The production of hydrogen peroxide was also quantified and its cellular location was assessed. The rehydration of slowly dried cells was associated with lower ROS production, thereby reducing the amount of cellular damage and increasing cell survival. A high oxygen consumption burst accompanied the initial stages of rehydration, perhaps due to the burst of ROS production. A slow dehydration rate may induce cell protection mechanisms that serve to limit ROS production and reduce the oxidative burst, decreasing the number of damaged and dead cells due upon rehydration.

Uptake Kinetics of As, Hg, Sb, and Se in the Aquatic Moss Fontinalis antipyretica Hedw

Laboratory experiments were carried out to study the uptake kinetics of selected metals and metalloids in the aquatic moss Fontinalis antipyretica. For this purpose, moss specimens from a clean site were exposed to concentrations of As, Hg, Sb, and Se ranging from 0.1 to 10,000 μg l−1, for incubation times of between 1 and 22 days, and the tissue concentrations of the metals in the moss specimens were then measured. Uptake kinetics followed different patterns in relation to exposure time, although the most common was Michaelis–Menten kinetics. On the contrary, the contamination factors followed very similar patterns in relation to the exposure concentrations in all cases, with a good fit to logarithmic equations. The bioconcentration factors tended to decrease as exposure concentration increased. The bioconcentration factors for Hg were extremely high, even at the lowest concentration in water and for the shortest incubation time, which implies that F. antipyretica has a high capacity to magnify Hg levels in water, which is an important characteristic in a good biomonitor. According to the time to reach equilibrium, the minimum exposure time recommended for use in active biomonitoring by means of transplants is very variable, although high levels of the elements, except Sb, were found in the moss tissues within a few days. We do not recommend the use of this moss species to biomonitor low concentrations of Sb in water. The differences in maximum contamination factors and lowest bioconcentration factors suggest that As and Se were the most toxic of the elements under study.

Biochemical fractionation and cellular distribution of americium and plutonium in the biomass of freshwater macrophytes

Accumulation of americium (241Am) and plutonium (238,242Pu) and their distribution in cell compartments and biochemical components of the biomass of freshwater aquatic plants Elodea canadensis,Ceratophyllum demersum and Myrioplyllum spicatum and aquatic moss Fontinalis antipyretica have been investigated in laboratory experiments. Americium and plutonium taken up from water by Elodea canadensis apical shoots were mainly absorbed by structural components of plant cells (90% for 241Am; 89% for 238Pu and 82–87% for 242Pu). About 10–18% of isotope activity was recorded in the cytosol fraction. The major concentration (76–92%) of americium was bound to cell wall cellulose-like polysaccharides of Elodea canadensis, Myriophyllum spicatum, Ceratophyllum demersum and Fontinalis antipyretica, 8–24% of americium activity was registered in the fraction of proteins and carbohydrates, and just a minor concentration (<1%) in the lipid fraction. The distribution of plutonium in the biomass fractions of Elodea was similar to that of americium. Hence, americium and plutonium had the highest affinity to cellulose-like polysaccharides of cell walls of freshwater submerged macrophytes.

Determination of antimicrobial and antiproliferative activities of the aquatic moss Fontinalis antipyretica Hedw.

The purpose of the current study was to investigate the possible antimicrobial and anticancer properties of the aquatic moss Fontinalis antipyretica Hedw. (Fontinalaceae). We first obtained 8 different extracts (methanol; chloroform; acetone; ethyl acetate A, B, C, and D) by two different extraction processes. The antimicrobial activity of these extracts was then assessed by using the well diffusion method against 8 bacterial and 7 fungal strains. The results of antimicrobial studies showed that extracts of chloroform, acetone, and ethyl acetate A and C were active against almost all the species tested. The most effective extract was C, which was then screened through a preparative TLC (bioautography) for its active components. Spot 4 of extract C exhibited the highest antimicrobial activity against 5 bacterial strains (B. cereus, B. subtilis, E. faecalis, E. aerogenes, and Y. enterocolitica) and 3 fungal strains (A. flavus, F. solani, and F. graminearum), with MIC values of 93.8-375.0 µg/mL and 187.5-375.0 µg/mL, respectively. In addition, in vitro toxicity of the active component of extract C at the concentrations of 0.16, 1.6, 16, 80, and 160 µg/mL was tested. Depending on dose and time, only 80 and 160 µg/mL decreased rat glioma (C6) cell viability after 24 or 48 h. The present study suggests the possibility that Fontinalis antipyretica possesses antimicrobial and anticancer agent(s).

Intracellular and extracellular ammonium (NH4 +) uptake and its toxic effects on the aquatic biomonitor Fontinalis antipyretica

The objective of this work is to validate the use of the aquatic moss Fontinalis antipyretica as biomonitor of NH(4)(+) aquatic pollution. In order to achieve this objective we needed to understand the pattern of uptake of NH(4)(+) by the moss and evaluate the impact of high concentrations on its physiological performance. The cellular location of NH(4)(+) in the moss is crucial for understanding its monitoring capacity. We were able to show that a sequential elution technique, based on the use of NiCl(2) as an efficient displacing agent, allowed the quantification of the cellular location of NH(4)(+). This was done along a concentration gradient and time of exposure. The extracellular and intracellular NH(4)(+) concentrations that caused significant physiological impact in membrane permeability of F. antipyretica were the same that caused significant decreasing in the photosynthetic capacity of the same moss. The former NH(4)(+) concentration thresholds were shown to decrease with increasing exposure time. These results are important since under natural conditions lower concentration of NH(4)(+) are present in waters but for very long periods of time. The importance of applying this knowledge in biomonitoring studies to fulfil the requirements of the Water Framework Directive is discussed.