When N availability increases, N may be taken up by plants in surplus and accumulated rather than being used in biomass production. At a cellular level, if protein is not synthesized, NH4+ is assimilated into specific N metabolites of which free amino acids are particularly important. Thus accumulation of amino acids could potentially be used to monitor N impacts on vegetation. Concentrations of a few specific amino acids, (which are species specific), are particularly well correlated with N deposition. Species typical of N- poor habitats accumulate arginine, whereas those more typical of N-rich habitats tend to accumulate asparagine (Nordin and Nasholm 1997). Changes in the composition of the amino acid pool may also occur in response to enhanced N deposition (Nordin et al. 1998; Pitcairn et al. in press).
- In early studies in the Netherlands, damaged trees were found to contain high concentration of the free amino acid arginine (van Dijk and Roelofs 1988).
- Several experiments have confirmed the link between enhanced N deposition and arginine concentrations. For example, when N deposition to the forest floor was reduced to 1-2 kg N ha-1 y-1 in the NITREX plot in Speuld forest, arginine concentrations in coniferous needles declined 3 fold within 4 years (Boxman et al. 1995). Another example is the very large reduction in arginine concentrations with increasing distance from a fur farm in Finland (Pietila et al. 1991).
- Nasholm et al. (1994) found that exposure to large concentrations of N increased the accumulation of amino acid N (as arginine, aspargine, glutamine and others) in a range of forest plants from trees to bryophytes, and suggested that concentrations of specific amino acids in a range of plants may be used to indicate atmospheric N deposition.
- Baxter et al. (1992) showed a rapid response in amino acid concentrations in two populations of Sphagnum cuspidatum following addition of 0.1 and 1.0 mM NH4+ in the laboratory.
- Downwind of a poultry farm in Scotland, UK, Pitcairn et al (2003) determined amino acid concentrations in 3 moss species (Rhytidiadelphus triquetrus, Brachythecium rutabulum and Pseudoscleropodium purum) in late autumn and showed a strong relationship with distance from the poultry buildings and hence with NH3 concentrations. Arginine was the dominant amino acid at high concentrations close to the buildings in all 3 species The linear regression of arginine concentration with log distance from the poultry farm gave values for r2 of > 0.95 for the 3 species demonstrating the potential for arginine accumulation in moss as an indicator of enhanced N deposition.
Suitability to indicate atmospheric concentrations:
There is insufficient evidence to determine whether this method is suitable to indicate atmospheric N concentrations, although amino acid content of 'pool' Sphagna responded to increased concentrations of ammonium in the laboratory.
Suitability to indicate atmospheric depositions:
Although the positive relationship between to amino acid concentrations and N deposition is well proved, the fact that different species may accumulate different amino acids in response to enhanced N deposition reduces the robustness of the method. It is therefore necessary to assess the changes of each of the main amino acids. There is insufficient evidence to determine whether this method is more suited to a wide range of N deposition levels or to low or very high levels. There is evidence to suggest that addition of NH4+-N may lead to larger increases in amino acid N (and also total N) than with NO3- -N.
Suitability to indicate environmental impacts:
Although there is good evidence of a relationship between amino acid concentrations and impacts of N deposition, the fact that different species may accumulate different amino acids in response to enhanced N deposition reduces the robustness of the method.
Sensitivity to other factors:
- Concentrations of amino acids tend to change seasonally. Late autumn is the best season for sampling for this method for most species
- Accumulation of amino acids can occur in response to growth limiting conditions where nutrients other than N are restricting growth e.g. amino acids can accumulate in response to SO2 pollution. Thus amino acid accumulation might be regarded as a non-specific indicator of perturbation of nutrient availability, which does affect the robustness of the method. Changes in the composition of the amino acid pool may overcome this factor, but there is insufficient data available at present to evaluate this.
- Concentrations of free amino acids are affected by several environmental factors including nutrient deficiencies and the form of the nitrogen deposition and the sites of storage also vary between species.
As amino acids are a storage compound, with elevated values reflecting surplus N, the method will reflect directly the time constant of excess N in the plants. Typically, this may result from gradual N accumulation in the ecosystem over several years, however, experimental work (e.g. Baxter et al. 1992) shows that the effects of new exposure can be determined in a few days.
The method is not limited to habitat or species. However, storage amino acids tend to be species dependent and not all species respond to enhance N by storage in the same amino acid. Species typical of N- poor habitats accumulate arginine, whereas those more typical of N-rich habitats tend to accumulate asparagine.
Large quantities of material are not required. Approximately 3 g fresh weight of material should be collected to provide a minimum of 0.5 g of dried, ground material for chemical analysis.
Expertise in field:
Specific species sampling requires training in species identification particularly of bryophytes or lichens, which many Agency staff may have already received. When age or growth phase of the plant or foliage is important, these must be identifiable by staff. Ideally, samples should be collected, cleaned (removal of litter, other species etc) and frozen and freeze-dried before being transported to an analytical laboratory by post, and specific conditions for storage and time between collection and freeze drying must be observed. When suitable equipment is not available for freeze-drying, samples can be frozen and delivered to the analytical laboratory by car in a ?cool?box. Alternatively they could be transported in the fresh state as quickly as possible (by courier or special delivery) for sample preparation at the analytical laboratory.
Expertise in laboratory:
Determination of amino acids requires extraction of the freeze dried sample and subsequent analysis using specialist expensive laboratory equipment, HPLC (prone to break-down) and skilled operators. Data analysis and interpretation also require experience.
Cost (per unit sample): £10-100
Cost Comment: For 3 species, 3 replicates per species and 3 locations per site.
Total: £21 Per 27 samples (£567).