Espécies nitrogenadas vem sendo adicionadas ao meio ambiente de forma intensa desde o desenvolvimento do processo Haber-Bosch para transformação de N2 em NH3, alterando significativamente o ciclo biogeoquímico do nitrogênio no ambiente. Uma vez na atmosfera, o nitrogênio reativo é depositado de volta à superfície da Terra por processos de deposição úmida e seca. A importância de levar em conta as formas orgânicas de nitrogênio para estimar a deposição de nitrogênio atmosférico pela chuva (úmida) já é conhecida, no entanto, ainda há poucos trabalhos que avaliam essa fração devido às dificuldades e incertezas impostas pelos métodos analíticos disponíveis. Este trabalho traz o desenvolvimento de um novo método, simples e de baixo custo, para a determinação de nitrogênio orgânico (N-org) em água de chuva utilizando o processo foto-Fenton e um foto-reator construído de forma artesanal. Por meio da adição de solução de Fenton (50 µmol L-1 Fe2+ e 2 mmol L-1 H2O2) e 90 min de radiação UV (85 °C) foi possível obter em média 106 ± 8% de recuperação de nitrogênio para soluções contendo 50 µmol N L-1 das moléculas modelos: ureia, serina, glicina e histidina. No caso da arginina, 90 min de radiação foi suficiente para degradar soluções contendo 10 µmol N L-1. O reator comercial se mostrou mais eficiente na degradação dos compostos testados (30 min), no entanto, com um tempo de 75 min, o reator artesanal atingiu os mesmos resultados. Com o uso de 0,2 g L-1 TiO2 e 120 min de radiação UV, também foi possível obter resultados satisfatórios. Porém, esse método possuiu valores de branco elevados, havendo necessidade de filtrar as amostras irradiadas antes da análise, adicionando tempo e custo ao procedimento analítico. O método desenvolvido utilizando foto-Fenton foi aplicado para a determinação de N-org em amostras de água de chuva coletadas na cidade de Ribeirão Preto (SP) de 2013 a 2017. A concentração de N-org variou de 3,5 a 195 µmol N L-1 com concentração média ponderada pelo volume (MPV) de 17,7 ± 1,0 µmol N L-1 (n=236). Essa média foi maior que aquelas obtidas em água de chuva de várias partes do mundo, podendo ser atribuída a elevada queima de biomassa na região de estudo. As concentrações de aminoácidos livres dissolvidos (AA) representaram em média 15 ± 12% (n=144) em relação à fração orgânica de nitrogênio na chuva, enquanto as concentrações de ureia foram próximas ou inferiores ao limite de quantificação do método (0,5 µmol N L-1). Considerando toda série temporal iniciada no mesmo sítio amostral desde 2005, as concentrações MPV calculadas para os íons NH4+ foi de 22,2 ± 1,1 µmol L-1 (n=460), NO3- de 13,3 ± 0,6 µmol L-1 (n=466), sendo que a concentração de NO2- foi irrelevante. Foram obtidas concentrações significativamente mais elevadas (teste-t, P=0,05) de NH4+, NO3-, N-org e AA no período de safra da cana (seco) com relação à entressafra (chuvoso), para todos os anos avaliados. Apesar da colheita manual da cana ter sido drasticamente reduzida, o fato de manter a mesma tendência sazonal desde 2005, demonstra que a prática da queima de biomassa ainda é intensa na região. O uso de fogo para manejo na área rural ainda é comum, além de haver grandes áreas queimadas de forma acidental. A deposição úmida de nitrogênio (N-org + NH4+ + NO3-) para Ribeirão Preto foi de 10,5 kg (N) ha-1 ano-1, sendo que a fração orgânica representou 33% dessa deposição, demonstrando a importância de se determinar N-org para melhor estimar os fluxos atmosféricos de deposição. A massa estimada de nitrogênio depositada pela chuva é cerca de 16% do nitrogênio aplicado por meio de fertilizantes em culturas de cana. Somando as deposições de nitrogênio pelo material particulado, pela fase gasosa, e úmida, esta última representa 71% do fluxo atmosférico de nitrogênio. Nesse contexto, a fração orgânica corresponde a 24% da deposição total, sendo que este valor ainda pode estar sendo subestimado, pois as concentrações de N-org no material particulado não foram determinadas. A simplicidade e exatidão do método aqui proposto pode facilitar a aquisição de dados de N-org na chuva de outras partes do mundo, melhorando assim o conhecimento sobre o ciclo biogeoquímico global do nitrogênio. / Nitrogen species have been intensively added to the environment since the development of Haber-Bosch process for N2 transformation to NH3, significantly altering the biogeochemical cycle of nitrogen. Once in the atmosphere, reactive nitrogen is deposited back to the Earth\'s surface by wet and dry deposition processes. The importance of taking into account the organic forms of nitrogen to estimate atmospheric nitrogen deposition by rain is already known. However, few studies evaluated this fraction due to the difficulties and uncertainties imposed by the available analytical methods. This work presents the development of a new, simple and low-cost method for the determination of organic nitrogen (N-org) in rainwater using photo-Fenton process and a homemade photo-reactor. By adding Fenton solution (50 mol L-1 Fe2+ and 2 mmol L-1 H2O2) and keeping the reaction under UV radiation for 90 min (85 °C), it was possible to obtain an average of 106 ± 8% nitrogen recovery for solutions containing 50 mol N L-1 of the model molecules: urea, serine, glycine and histidine. In the case of arginine, 90 min of radiation was sufficient to degrade solutions containing 10 mol N L-1. A commercial reactor showed to be more efficient in degradation of the tested compounds (30 min). However, with a time of 75 min, the homemade reactor achieved the same results. With 0.2 g L-1 TiO2 and 120 min of UV radiation, it was also possible to obtain satisfactory results. Nevertheless, this method had high blank values, and filtration of irradiated samples before the analysis was necessary, which increased time and cost to the analytical procedure. The developed method using photo-Fenton was applied to determine N-org in rainwater samples collected in Ribeirão Preto city (SP) from 2013 to 2017. N-org concentrations ranged from 3.5 to 195 mol N L-1 with a volume-weighted mean concentration (VWM) of 17.7 ± 1.0 mol N L-1 (n = 236). This value was higher than those reported for rainwater from different parts of the world, and this fact can be attributed to the high biomass burning in the study region. Dissolved free amino acids (AA) mean concentrations represented 15 ± 12% (n = 144) of the organic nitrogen fraction in rain, while urea concentrations were close to or below the limit of quantification of the method (0.5 mol N L-1). Considering all temporal series, initiated in 2005 at the same sampling site, the VWM concentration calculated for NH4+ ions was 22.2 ± 1.1 mol L-1 (n = 460) and for NO3- was 13.3 ± 0.6 mol L-1 (n = 466), while NO2- mean concentration was irrelevant. Significantly higher concentrations (t-test, P = 0.05) of NH4+, NO3-, N-org and AA were obtained during the harvest period (dry season) in relation to the non-harvest one (rainy season), for all evaluated years. Although manual harvesting was drastically reduced, the fact that the same seasonal trend has been maintained since 2005 demonstrates that the practice of biomass burning is still intense in the region. In rural area, using fire for land management is still common, in addition to large areas burned by unintentional fires. Nitrogen deposition (N-org + NH4+ + NO3-) in Ribeirão Preto was 10.5 kg (N) ha-1 year-1, and the organic fraction represented 33% of this deposition, demonstrating the importance of determining N-org to better estimate the atmospheric deposition fluxes. The estimated mass of nitrogen deposited by rain represents approximately 16% of the nitrogen introduced by fertilizers in sugarcane crops. Summing up the nitrogen deposition by particulate matter, by gas phase, and by rain, the latter represents 71% of the atmospheric nitrogen flux. In this context, the organic fraction corresponds to 24% of the total deposition, and this value may still be underestimated, since N-org concentrations in particulate matter were not determined. The simplicity and accuracy of the method proposed here may facilitate acquisition of N-org data in rainwater from other parts of the world, thus improving the knowledge on the global biogeochemical cycle of nitrogen.

The following research has focused on the utility of Ulva as an indicator of seawater nitrogen loading. Evaluation was made in three ways: 1) Observation of large-scale geographic variation in nitrogen status in natural populations around New Zealand in summer and winter, 2) Laboratory-based experimental assessment of the biochemical responses of N-indices in Ulva to nitrogen enrichment, and 3) Culturing standardized test-Ulva under low nutrient conditions which could be deployed into a variety of field situations. Seawater inorganic nutrient (nitrate, nitrite, ammonium and phosphate) concentrations and nitrogen (N)-indices (free amino acids, chlorophyll and total tissue nitrogen) in natural Ulva populations from 32 sites around New Zealand were compared. Sites were divided into 6 environmental categories: sheltered rural, exposed rural, rock pools, sheltered urban, exposed urban, and nitrogen-enriched urban sites. Seawater nutrient concentrations were highly variable between all sites in summer and winter. However, in the summer enriched urban sites had the highest mean total inorganic nitrogen concentrations and Ulva with the highest mean levels of all N-indices compared with any other environmental category. In the winter, Ulva contained more nitrogen (reflected in all N-indices) compared with Ulva in the summer, particularly in populations growing in colder southern seawater on more exposed coasts. The increase in Ulva N-status was not explained by increased seawater inorganic nitrogen concentrations. With univariate and multivariate statistical approaches it was shown that there was a significant effect of seawater temperature and site exposure on N-status in Ulva. Compared with other N-indices, stable nitrogen isotopes (δ15N) from Ulva growing in enriched urban sites had the widest range (4.77 ± 0.04 ‰ to 15.16 ± 0.03 ‰) of values compared with all other categories in both summer and winter. Conversely, Ulva from exposed rural sites had the lowest range of δ15N values compared with any other category (6.7 ± 0.1 to 8.8 ± 0.1 ‰) and showed no seasonal change in mean values (7.8 ‰ and 7.6 ‰ for summer and winter, respectively). In addition, δ15N values in Ulva were the only N-index that showed a significant difference between urban and rural categories. To test the relationship between inorganic nitrogen concentration in seawater and the responses of biochemical nitrogen indices in Ulva pertusa, several experiments were conducted in an outdoor, flow-through culture apparatus, in summer and winter. In this apparatus effects of ammonium concentration, nitrogen source (nitrate and ammonium), light and seawater motion were investigated. Of the same three N-indices examined in natural Ulva populations (free amino acids, chlorophyll and total tissue nitrogen), increases in free amino acids, particularly asparagine, provided the strongest indicator of increases in nitrogen availability. In addition, while tissue nitrogen and chlorophyll also increased with seawater nitrogen concentration, it was apparent that these indices were also strongly influenced by light, and probably season. Rates of ammonium assimilation provided no overall measure of the availability of nitrogen in seawater and were clearly affected by season. Similarly, growth rates in Ulva only showed a response to nitrogen addition in summer months. Stable isotopes of nitrogen (δ15N) in Ulva provided a clear distinction between natural and synthetic nitrogen sources, but more importantly, showed only minor fractionation (ranging from 1.3 ‰ to -1.9 ‰) of 15N supplied from synthetic nitrate and ammonium under both light-saturating and light-limiting conditions. To further develop Ulva as a standardized test-organism it was cultured in low-nutrient (non-polluted) seawater to deplete internal storage pools of nitrogen. Each month the resulting test-Ulva was then placed in surface-moored growth enclosures at a range of coastal sites around Auckland and then monitored for one year. In winter there were increases in seawater inorganic nitrogen concentrations and concomitant increases in free amino acid content. However, tissue nitrogen and chlorophyll content in test-Ulva showed similar increases (possibly saturating) across all sites suggesting that seasonal increases in these N-indices were also due to other seasonal factors (e.g., surface irradiance and / or seawater temperature). On the other hand, the total free amino acid pool showed strong differences between a low-nitrogen reference site and the other study sites all year round. It was probable that test-Ulva was integrating differences in tidally-averaged nitrogen loading that were not reliably detected in instantaneous seawater samples. In addition to N-indices in test-Ulva, levels of tissue heavy metals and stable isotopes of nitrogen showed strong differences with higher values of both typically found in urban environments compared with values found in non-polluted reference sites. It is concluded that several abiotic and biotic factors affect nitrogen status in Ulva, but the average nitrogen concentration in seawater, and the physical factors of temperature, light and water motion, appear to be the overarching determinants. It is further suggested that in combination with Ulva tissue δ15N values, tissue nitrogen and the free amino acid pool, as quantitative biochemical measures of nitrogen availability, are likely to provide useful information on both the amount and composition of nitrogen entering coastal environments. / Foundation for Research, Science and Technology. Auckland Regional Council.

The following research has focused on the utility of Ulva as an indicator of seawater nitrogen loading. Evaluation was made in three ways: 1) Observation of large-scale geographic variation in nitrogen status in natural populations around New Zealand in summer and winter, 2) Laboratory-based experimental assessment of the biochemical responses of N-indices in Ulva to nitrogen enrichment, and 3) Culturing standardized test-Ulva under low nutrient conditions which could be deployed into a variety of field situations. Seawater inorganic nutrient (nitrate, nitrite, ammonium and phosphate) concentrations and nitrogen (N)-indices (free amino acids, chlorophyll and total tissue nitrogen) in natural Ulva populations from 32 sites around New Zealand were compared. Sites were divided into 6 environmental categories: sheltered rural, exposed rural, rock pools, sheltered urban, exposed urban, and nitrogen-enriched urban sites. Seawater nutrient concentrations were highly variable between all sites in summer and winter. However, in the summer enriched urban sites had the highest mean total inorganic nitrogen concentrations and Ulva with the highest mean levels of all N-indices compared with any other environmental category. In the winter, Ulva contained more nitrogen (reflected in all N-indices) compared with Ulva in the summer, particularly in populations growing in colder southern seawater on more exposed coasts. The increase in Ulva N-status was not explained by increased seawater inorganic nitrogen concentrations. With univariate and multivariate statistical approaches it was shown that there was a significant effect of seawater temperature and site exposure on N-status in Ulva. Compared with other N-indices, stable nitrogen isotopes (δ15N) from Ulva growing in enriched urban sites had the widest range (4.77 ± 0.04 ‰ to 15.16 ± 0.03 ‰) of values compared with all other categories in both summer and winter. Conversely, Ulva from exposed rural sites had the lowest range of δ15N values compared with any other category (6.7 ± 0.1 to 8.8 ± 0.1 ‰) and showed no seasonal change in mean values (7.8 ‰ and 7.6 ‰ for summer and winter, respectively). In addition, δ15N values in Ulva were the only N-index that showed a significant difference between urban and rural categories. To test the relationship between inorganic nitrogen concentration in seawater and the responses of biochemical nitrogen indices in Ulva pertusa, several experiments were conducted in an outdoor, flow-through culture apparatus, in summer and winter. In this apparatus effects of ammonium concentration, nitrogen source (nitrate and ammonium), light and seawater motion were investigated. Of the same three N-indices examined in natural Ulva populations (free amino acids, chlorophyll and total tissue nitrogen), increases in free amino acids, particularly asparagine, provided the strongest indicator of increases in nitrogen availability. In addition, while tissue nitrogen and chlorophyll also increased with seawater nitrogen concentration, it was apparent that these indices were also strongly influenced by light, and probably season. Rates of ammonium assimilation provided no overall measure of the availability of nitrogen in seawater and were clearly affected by season. Similarly, growth rates in Ulva only showed a response to nitrogen addition in summer months. Stable isotopes of nitrogen (δ15N) in Ulva provided a clear distinction between natural and synthetic nitrogen sources, but more importantly, showed only minor fractionation (ranging from 1.3 ‰ to -1.9 ‰) of 15N supplied from synthetic nitrate and ammonium under both light-saturating and light-limiting conditions. To further develop Ulva as a standardized test-organism it was cultured in low-nutrient (non-polluted) seawater to deplete internal storage pools of nitrogen. Each month the resulting test-Ulva was then placed in surface-moored growth enclosures at a range of coastal sites around Auckland and then monitored for one year. In winter there were increases in seawater inorganic nitrogen concentrations and concomitant increases in free amino acid content. However, tissue nitrogen and chlorophyll content in test-Ulva showed similar increases (possibly saturating) across all sites suggesting that seasonal increases in these N-indices were also due to other seasonal factors (e.g., surface irradiance and / or seawater temperature). On the other hand, the total free amino acid pool showed strong differences between a low-nitrogen reference site and the other study sites all year round. It was probable that test-Ulva was integrating differences in tidally-averaged nitrogen loading that were not reliably detected in instantaneous seawater samples. In addition to N-indices in test-Ulva, levels of tissue heavy metals and stable isotopes of nitrogen showed strong differences with higher values of both typically found in urban environments compared with values found in non-polluted reference sites. It is concluded that several abiotic and biotic factors affect nitrogen status in Ulva, but the average nitrogen concentration in seawater, and the physical factors of temperature, light and water motion, appear to be the overarching determinants. It is further suggested that in combination with Ulva tissue δ15N values, tissue nitrogen and the free amino acid pool, as quantitative biochemical measures of nitrogen availability, are likely to provide useful information on both the amount and composition of nitrogen entering coastal environments. / Foundation for Research, Science and Technology. Auckland Regional Council.

The following research has focused on the utility of Ulva as an indicator of seawater nitrogen loading. Evaluation was made in three ways: 1) Observation of large-scale geographic variation in nitrogen status in natural populations around New Zealand in summer and winter, 2) Laboratory-based experimental assessment of the biochemical responses of N-indices in Ulva to nitrogen enrichment, and 3) Culturing standardized test-Ulva under low nutrient conditions which could be deployed into a variety of field situations. Seawater inorganic nutrient (nitrate, nitrite, ammonium and phosphate) concentrations and nitrogen (N)-indices (free amino acids, chlorophyll and total tissue nitrogen) in natural Ulva populations from 32 sites around New Zealand were compared. Sites were divided into 6 environmental categories: sheltered rural, exposed rural, rock pools, sheltered urban, exposed urban, and nitrogen-enriched urban sites. Seawater nutrient concentrations were highly variable between all sites in summer and winter. However, in the summer enriched urban sites had the highest mean total inorganic nitrogen concentrations and Ulva with the highest mean levels of all N-indices compared with any other environmental category. In the winter, Ulva contained more nitrogen (reflected in all N-indices) compared with Ulva in the summer, particularly in populations growing in colder southern seawater on more exposed coasts. The increase in Ulva N-status was not explained by increased seawater inorganic nitrogen concentrations. With univariate and multivariate statistical approaches it was shown that there was a significant effect of seawater temperature and site exposure on N-status in Ulva. Compared with other N-indices, stable nitrogen isotopes (δ15N) from Ulva growing in enriched urban sites had the widest range (4.77 ± 0.04 ‰ to 15.16 ± 0.03 ‰) of values compared with all other categories in both summer and winter. Conversely, Ulva from exposed rural sites had the lowest range of δ15N values compared with any other category (6.7 ± 0.1 to 8.8 ± 0.1 ‰) and showed no seasonal change in mean values (7.8 ‰ and 7.6 ‰ for summer and winter, respectively). In addition, δ15N values in Ulva were the only N-index that showed a significant difference between urban and rural categories. To test the relationship between inorganic nitrogen concentration in seawater and the responses of biochemical nitrogen indices in Ulva pertusa, several experiments were conducted in an outdoor, flow-through culture apparatus, in summer and winter. In this apparatus effects of ammonium concentration, nitrogen source (nitrate and ammonium), light and seawater motion were investigated. Of the same three N-indices examined in natural Ulva populations (free amino acids, chlorophyll and total tissue nitrogen), increases in free amino acids, particularly asparagine, provided the strongest indicator of increases in nitrogen availability. In addition, while tissue nitrogen and chlorophyll also increased with seawater nitrogen concentration, it was apparent that these indices were also strongly influenced by light, and probably season. Rates of ammonium assimilation provided no overall measure of the availability of nitrogen in seawater and were clearly affected by season. Similarly, growth rates in Ulva only showed a response to nitrogen addition in summer months. Stable isotopes of nitrogen (δ15N) in Ulva provided a clear distinction between natural and synthetic nitrogen sources, but more importantly, showed only minor fractionation (ranging from 1.3 ‰ to -1.9 ‰) of 15N supplied from synthetic nitrate and ammonium under both light-saturating and light-limiting conditions. To further develop Ulva as a standardized test-organism it was cultured in low-nutrient (non-polluted) seawater to deplete internal storage pools of nitrogen. Each month the resulting test-Ulva was then placed in surface-moored growth enclosures at a range of coastal sites around Auckland and then monitored for one year. In winter there were increases in seawater inorganic nitrogen concentrations and concomitant increases in free amino acid content. However, tissue nitrogen and chlorophyll content in test-Ulva showed similar increases (possibly saturating) across all sites suggesting that seasonal increases in these N-indices were also due to other seasonal factors (e.g., surface irradiance and / or seawater temperature). On the other hand, the total free amino acid pool showed strong differences between a low-nitrogen reference site and the other study sites all year round. It was probable that test-Ulva was integrating differences in tidally-averaged nitrogen loading that were not reliably detected in instantaneous seawater samples. In addition to N-indices in test-Ulva, levels of tissue heavy metals and stable isotopes of nitrogen showed strong differences with higher values of both typically found in urban environments compared with values found in non-polluted reference sites. It is concluded that several abiotic and biotic factors affect nitrogen status in Ulva, but the average nitrogen concentration in seawater, and the physical factors of temperature, light and water motion, appear to be the overarching determinants. It is further suggested that in combination with Ulva tissue δ15N values, tissue nitrogen and the free amino acid pool, as quantitative biochemical measures of nitrogen availability, are likely to provide useful information on both the amount and composition of nitrogen entering coastal environments. / Foundation for Research, Science and Technology. Auckland Regional Council.

The following research has focused on the utility of Ulva as an indicator of seawater nitrogen loading. Evaluation was made in three ways: 1) Observation of large-scale geographic variation in nitrogen status in natural populations around New Zealand in summer and winter, 2) Laboratory-based experimental assessment of the biochemical responses of N-indices in Ulva to nitrogen enrichment, and 3) Culturing standardized test-Ulva under low nutrient conditions which could be deployed into a variety of field situations. Seawater inorganic nutrient (nitrate, nitrite, ammonium and phosphate) concentrations and nitrogen (N)-indices (free amino acids, chlorophyll and total tissue nitrogen) in natural Ulva populations from 32 sites around New Zealand were compared. Sites were divided into 6 environmental categories: sheltered rural, exposed rural, rock pools, sheltered urban, exposed urban, and nitrogen-enriched urban sites. Seawater nutrient concentrations were highly variable between all sites in summer and winter. However, in the summer enriched urban sites had the highest mean total inorganic nitrogen concentrations and Ulva with the highest mean levels of all N-indices compared with any other environmental category. In the winter, Ulva contained more nitrogen (reflected in all N-indices) compared with Ulva in the summer, particularly in populations growing in colder southern seawater on more exposed coasts. The increase in Ulva N-status was not explained by increased seawater inorganic nitrogen concentrations. With univariate and multivariate statistical approaches it was shown that there was a significant effect of seawater temperature and site exposure on N-status in Ulva. Compared with other N-indices, stable nitrogen isotopes (δ15N) from Ulva growing in enriched urban sites had the widest range (4.77 ± 0.04 ‰ to 15.16 ± 0.03 ‰) of values compared with all other categories in both summer and winter. Conversely, Ulva from exposed rural sites had the lowest range of δ15N values compared with any other category (6.7 ± 0.1 to 8.8 ± 0.1 ‰) and showed no seasonal change in mean values (7.8 ‰ and 7.6 ‰ for summer and winter, respectively). In addition, δ15N values in Ulva were the only N-index that showed a significant difference between urban and rural categories. To test the relationship between inorganic nitrogen concentration in seawater and the responses of biochemical nitrogen indices in Ulva pertusa, several experiments were conducted in an outdoor, flow-through culture apparatus, in summer and winter. In this apparatus effects of ammonium concentration, nitrogen source (nitrate and ammonium), light and seawater motion were investigated. Of the same three N-indices examined in natural Ulva populations (free amino acids, chlorophyll and total tissue nitrogen), increases in free amino acids, particularly asparagine, provided the strongest indicator of increases in nitrogen availability. In addition, while tissue nitrogen and chlorophyll also increased with seawater nitrogen concentration, it was apparent that these indices were also strongly influenced by light, and probably season. Rates of ammonium assimilation provided no overall measure of the availability of nitrogen in seawater and were clearly affected by season. Similarly, growth rates in Ulva only showed a response to nitrogen addition in summer months. Stable isotopes of nitrogen (δ15N) in Ulva provided a clear distinction between natural and synthetic nitrogen sources, but more importantly, showed only minor fractionation (ranging from 1.3 ‰ to -1.9 ‰) of 15N supplied from synthetic nitrate and ammonium under both light-saturating and light-limiting conditions. To further develop Ulva as a standardized test-organism it was cultured in low-nutrient (non-polluted) seawater to deplete internal storage pools of nitrogen. Each month the resulting test-Ulva was then placed in surface-moored growth enclosures at a range of coastal sites around Auckland and then monitored for one year. In winter there were increases in seawater inorganic nitrogen concentrations and concomitant increases in free amino acid content. However, tissue nitrogen and chlorophyll content in test-Ulva showed similar increases (possibly saturating) across all sites suggesting that seasonal increases in these N-indices were also due to other seasonal factors (e.g., surface irradiance and / or seawater temperature). On the other hand, the total free amino acid pool showed strong differences between a low-nitrogen reference site and the other study sites all year round. It was probable that test-Ulva was integrating differences in tidally-averaged nitrogen loading that were not reliably detected in instantaneous seawater samples. In addition to N-indices in test-Ulva, levels of tissue heavy metals and stable isotopes of nitrogen showed strong differences with higher values of both typically found in urban environments compared with values found in non-polluted reference sites. It is concluded that several abiotic and biotic factors affect nitrogen status in Ulva, but the average nitrogen concentration in seawater, and the physical factors of temperature, light and water motion, appear to be the overarching determinants. It is further suggested that in combination with Ulva tissue δ15N values, tissue nitrogen and the free amino acid pool, as quantitative biochemical measures of nitrogen availability, are likely to provide useful information on both the amount and composition of nitrogen entering coastal environments. / Foundation for Research, Science and Technology. Auckland Regional Council.

This study deals with an effect of fertilization on amount and quality of free amino acids in soil from wet meadows affected by nutrient loading. Free amino acids were analysed in the soil from the field experiment situated at wet meadows, where an increased nutrient input is simulated (fertilizer NPK). The experiment was established on two sites with different types of soil {--} Záblatí with organic soil and Hamr with mineral soil, both areas in the South Bohemia region. The soil was repeatedly sampled for more than two years and year. One part of this study is focused on testing of ninhydrine-method and HPLC-method for measuring free amino acids and efficiency of different extractants.

Espécies nitrogenadas vem sendo adicionadas ao meio ambiente de forma intensa desde o desenvolvimento do processo Haber-Bosch para transformação de N2 em NH3, alterando significativamente o ciclo biogeoquímico do nitrogênio no ambiente. Uma vez na atmosfera, o nitrogênio reativo é depositado de volta à superfície da Terra por processos de deposição úmida e seca. A importância de levar em conta as formas orgânicas de nitrogênio para estimar a deposição de nitrogênio atmosférico pela chuva (úmida) já é conhecida, no entanto, ainda há poucos trabalhos que avaliam essa fração devido às dificuldades e incertezas impostas pelos métodos analíticos disponíveis. Este trabalho traz o desenvolvimento de um novo método, simples e de baixo custo, para a determinação de nitrogênio orgânico (N-org) em água de chuva utilizando o processo foto-Fenton e um foto-reator construído de forma artesanal. Por meio da adição de solução de Fenton (50 µmol L-1 Fe2+ e 2 mmol L-1 H2O2) e 90 min de radiação UV (85 °C) foi possível obter em média 106 ± 8% de recuperação de nitrogênio para soluções contendo 50 µmol N L-1 das moléculas modelos: ureia, serina, glicina e histidina. No caso da arginina, 90 min de radiação foi suficiente para degradar soluções contendo 10 µmol N L-1. O reator comercial se mostrou mais eficiente na degradação dos compostos testados (30 min), no entanto, com um tempo de 75 min, o reator artesanal atingiu os mesmos resultados. Com o uso de 0,2 g L-1 TiO2 e 120 min de radiação UV, também foi possível obter resultados satisfatórios. Porém, esse método possuiu valores de branco elevados, havendo necessidade de filtrar as amostras irradiadas antes da análise, adicionando tempo e custo ao procedimento analítico. O método desenvolvido utilizando foto-Fenton foi aplicado para a determinação de N-org em amostras de água de chuva coletadas na cidade de Ribeirão Preto (SP) de 2013 a 2017. A concentração de N-org variou de 3,5 a 195 µmol N L-1 com concentração média ponderada pelo volume (MPV) de 17,7 ± 1,0 µmol N L-1 (n=236). Essa média foi maior que aquelas obtidas em água de chuva de várias partes do mundo, podendo ser atribuída a elevada queima de biomassa na região de estudo. As concentrações de aminoácidos livres dissolvidos (AA) representaram em média 15 ± 12% (n=144) em relação à fração orgânica de nitrogênio na chuva, enquanto as concentrações de ureia foram próximas ou inferiores ao limite de quantificação do método (0,5 µmol N L-1). Considerando toda série temporal iniciada no mesmo sítio amostral desde 2005, as concentrações MPV calculadas para os íons NH4+ foi de 22,2 ± 1,1 µmol L-1 (n=460), NO3- de 13,3 ± 0,6 µmol L-1 (n=466), sendo que a concentração de NO2- foi irrelevante. Foram obtidas concentrações significativamente mais elevadas (teste-t, P=0,05) de NH4+, NO3-, N-org e AA no período de safra da cana (seco) com relação à entressafra (chuvoso), para todos os anos avaliados. Apesar da colheita manual da cana ter sido drasticamente reduzida, o fato de manter a mesma tendência sazonal desde 2005, demonstra que a prática da queima de biomassa ainda é intensa na região. O uso de fogo para manejo na área rural ainda é comum, além de haver grandes áreas queimadas de forma acidental. A deposição úmida de nitrogênio (N-org + NH4+ + NO3-) para Ribeirão Preto foi de 10,5 kg (N) ha-1 ano-1, sendo que a fração orgânica representou 33% dessa deposição, demonstrando a importância de se determinar N-org para melhor estimar os fluxos atmosféricos de deposição. A massa estimada de nitrogênio depositada pela chuva é cerca de 16% do nitrogênio aplicado por meio de fertilizantes em culturas de cana. Somando as deposições de nitrogênio pelo material particulado, pela fase gasosa, e úmida, esta última representa 71% do fluxo atmosférico de nitrogênio. Nesse contexto, a fração orgânica corresponde a 24% da deposição total, sendo que este valor ainda pode estar sendo subestimado, pois as concentrações de N-org no material particulado não foram determinadas. A simplicidade e exatidão do método aqui proposto pode facilitar a aquisição de dados de N-org na chuva de outras partes do mundo, melhorando assim o conhecimento sobre o ciclo biogeoquímico global do nitrogênio. / Nitrogen species have been intensively added to the environment since the development of Haber-Bosch process for N2 transformation to NH3, significantly altering the biogeochemical cycle of nitrogen. Once in the atmosphere, reactive nitrogen is deposited back to the Earth\'s surface by wet and dry deposition processes. The importance of taking into account the organic forms of nitrogen to estimate atmospheric nitrogen deposition by rain is already known. However, few studies evaluated this fraction due to the difficulties and uncertainties imposed by the available analytical methods. This work presents the development of a new, simple and low-cost method for the determination of organic nitrogen (N-org) in rainwater using photo-Fenton process and a homemade photo-reactor. By adding Fenton solution (50 mol L-1 Fe2+ and 2 mmol L-1 H2O2) and keeping the reaction under UV radiation for 90 min (85 °C), it was possible to obtain an average of 106 ± 8% nitrogen recovery for solutions containing 50 mol N L-1 of the model molecules: urea, serine, glycine and histidine. In the case of arginine, 90 min of radiation was sufficient to degrade solutions containing 10 mol N L-1. A commercial reactor showed to be more efficient in degradation of the tested compounds (30 min). However, with a time of 75 min, the homemade reactor achieved the same results. With 0.2 g L-1 TiO2 and 120 min of UV radiation, it was also possible to obtain satisfactory results. Nevertheless, this method had high blank values, and filtration of irradiated samples before the analysis was necessary, which increased time and cost to the analytical procedure. The developed method using photo-Fenton was applied to determine N-org in rainwater samples collected in Ribeirão Preto city (SP) from 2013 to 2017. N-org concentrations ranged from 3.5 to 195 mol N L-1 with a volume-weighted mean concentration (VWM) of 17.7 ± 1.0 mol N L-1 (n = 236). This value was higher than those reported for rainwater from different parts of the world, and this fact can be attributed to the high biomass burning in the study region. Dissolved free amino acids (AA) mean concentrations represented 15 ± 12% (n = 144) of the organic nitrogen fraction in rain, while urea concentrations were close to or below the limit of quantification of the method (0.5 mol N L-1). Considering all temporal series, initiated in 2005 at the same sampling site, the VWM concentration calculated for NH4+ ions was 22.2 ± 1.1 mol L-1 (n = 460) and for NO3- was 13.3 ± 0.6 mol L-1 (n = 466), while NO2- mean concentration was irrelevant. Significantly higher concentrations (t-test, P = 0.05) of NH4+, NO3-, N-org and AA were obtained during the harvest period (dry season) in relation to the non-harvest one (rainy season), for all evaluated years. Although manual harvesting was drastically reduced, the fact that the same seasonal trend has been maintained since 2005 demonstrates that the practice of biomass burning is still intense in the region. In rural area, using fire for land management is still common, in addition to large areas burned by unintentional fires. Nitrogen deposition (N-org + NH4+ + NO3-) in Ribeirão Preto was 10.5 kg (N) ha-1 year-1, and the organic fraction represented 33% of this deposition, demonstrating the importance of determining N-org to better estimate the atmospheric deposition fluxes. The estimated mass of nitrogen deposited by rain represents approximately 16% of the nitrogen introduced by fertilizers in sugarcane crops. Summing up the nitrogen deposition by particulate matter, by gas phase, and by rain, the latter represents 71% of the atmospheric nitrogen flux. In this context, the organic fraction corresponds to 24% of the total deposition, and this value may still be underestimated, since N-org concentrations in particulate matter were not determined. The simplicity and accuracy of the method proposed here may facilitate acquisition of N-org data in rainwater from other parts of the world, thus improving the knowledge on the global biogeochemical cycle of nitrogen.

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