A modification of Mover's method has been found satisfactory for the quantitative measurement of resazurin reduction in biological systems. For convenience, the reducing systems were studied separately and the contribution of each system to total dye reduction was determined. The rapid reduction of resazurin in milk due to sun-light was found to be a photochemical reaction initiated by the excitation of the dye by light of a wavelength of approximately 600 mμ. It was found that the main reducing system in normal raw milk was ascorbic acid dependent, whereas in mastitic milk the main reducing system was leucocytes. Physical fractionation of mastitic milk, sonic treatment, and the use of metabolic inhibitors strongly indicated that there was an intimate relationship between the presence of leucocytes and the reducing activities of mastitic milk. Studies with cell-free extracts of leucocytes and bacteria showed that the reducing systems were essentially the same, and involved reduced pyridine nucleotides as electron donors. The mechanisms of resazurin reduction have been confirmed by studies involving the use of metabolic inhibitors, respirometry, variation of the energy source and kinetic studies. Finally, the successful reconstruction of several artificial resazurin reducing systems further confirmed the validity of this mechanism. / Land and Food Systems, Faculty of / Graduate

Thesis (M.S.P.H.)--University of Michigan, 1938.

Thesis (M.S.P.H.)--University of Michigan, 1938.

A criopreservação do sêmen é de muita importância para a produção de equinos pois permite o amplo comércio internacional de sêmen e a redução dos custos com transporte de animais. Entretanto, o sêmen criopreservado apresenta reduzida longevidade e integridade funcional, um obstáculo para a expansão dessa técnica. Recentes descobertas sugerem que os maiores danos durante a criopreservação de sêmen de garanhões ocorrem devido ao desequilíbrio osmótico e às espécies reativas de oxigênio. Um método para combater os efeitos deletérios da congelação é utilizar substâncias que possam amenizar os danos subletais. O objetivo desse experimento foi avaliar o efeito da adição de duas substâncias com efeito protetor celular (mio-inositol e ácido ferúlico) ao meio de congelação, na qualidade do sêmen criopreservado de garanhões. Para isso foram realizadas 5 colheitas de sêmen de 5 garanhões. O sêmen foi processado para criopreservação e antes do envase foi dividido em 3 tratamentos: controle, mio-inositol (30mM) e ácido ferúlico (160 µM). As palhetas foram descongeladas e analisadas quanto a motilidade computadorizada (CASA), integridade de membrana, estado metabólico e produção de espécies reativas de oxigênio (EROs) em microscopia de epifluorescência nos tempos 0, 2 e 4h de incubação a 37°C. Previamente foram realizadas validações da técnica de mensuração simultânea da membrana plasmática e estado metabólico (sondas PI, H33342 e resazurina) e correlação entre duas técnicas de mensuração de EROs (DCFH-DA e DCFH-DA com H33342). Os dados obtidos foram avaliados no programa SAS, versão 9,3 (SAS, 2011) utilizando ANOVA e medidas repetidas no tempo, para as validações foram realizadas regressões lineares simples. A adição de mio-inositol no diluidor de congelação resultou em maior motilidade total no tempo 2h e 4h e maior motilidade progressiva nos tempos 0 e 2h de incubação pós-descongelação em relação aos outros grupos. O tratamento com mio-inositol resultou em maior quantidade de células com membrana plasmática íntegra nos tempo 0h (53,3±1,4%); 2h (50,0±1,0%) e 4h (37,9±1,5%) de incubação que o grupo controle: 0h (48,0±1,0%); 2h (44,9±1,1%); e 4h (31,5±1,7%). O ácido ferúlico prejudicou as características de motilidade (CASA) e a integridade da membrana plasmática, entretanto melhorou o estado metabólico em todos os tempos. Conclui-se que o mio-inositol melhora as características de motilidade e a integridade de membranas espermáticas do sêmen criopreservado de equinos e que o ácido ferúlico apesar de prejudicar essas características promove melhor metabolismo espermático, mensurado pela técnica de redução da resazurina. / Sperm cryopreservation is of great importance for equine production, it assists in the international semen trade and reduce costs with animal transportation. However, frozen/thawed semen has reduced longevity and functional integrity and these is an obstacle to the expansion of this technique. The improvement in quality of cryopreserved semen is very important, especially to help become available more widely new technologies, such as sperm sexing. Recent findings suggest that the greatest damage during cryopreservation of stallion semen occur due to osmotic imbalance and the reactive oxygen species (ROS). One method to mitigate the deleterious effects of freezing is using substances that might lessen the sublethal damage. The objective of this experiment was to evaluate the quality of frozen/thawed stallion sperm after using two substances that have protective effects in the freezing medium: myo-inositol and ferulic acid. Were performed five sperm collection of 5 stallions. The semen was processed and before cryopreservation it was divided into three treatments: control, myo-inositol (30 mM) and ferulic acid (160 mM). The straws were thawed and analyzed at 0, 2 and 4 h of incubation time at 37°C. It was performed computerized motility (CASA), membrane integrity, metabolic status and ROS production using an epifluorescence microscope. Before beginning of the experiment were carried out validations of the technique of simultaneous assessment of plasma membrane and metabolic state (PI probes, H33342 and resazurin) and correlation between two measurement techniques ROS (DCFH-DA and DCFH-DA with H33342). The data were analyzed using the SAS version 9.3 (SAS, 2011) with ANOVA and repeated measures. For the probes validation were used linear regressions. The addition of myo-inositol in the freezing extender resulted in higher total motility in time 2h and 4h and increased progressive cells at 0 and 2 h of incubation post-thaw compared to the other groups. Treatment with myo-inositol resulted in a higher number of cells with intact plasma membrane in time 0h (53.3 ± 1.4%); 2h (50.0 ± 1.0%) and 4h (37.9 ± 1.5%) of incubation than the control group: 0h (48.0 ± 1.0%), 2h (44.9 ± 1.1%), and 4h (31.5 ± 1.7%). The use of ferulic acid resulted in the worst characteristics of motility (CASA) and plasma membrane integrity, however improved metabolic status at all incubation times. It is concluded that myo-inositol improves sperm motility characteristics and preserves membrane integrity of equine cryopreserved semen. Ferulic acid although impairing these characteristics, promotes better sperm metabolism, measured by resazurin test.

A criopreservação do sêmen é de muita importância para a produção de equinos pois permite o amplo comércio internacional de sêmen e a redução dos custos com transporte de animais. Entretanto, o sêmen criopreservado apresenta reduzida longevidade e integridade funcional, um obstáculo para a expansão dessa técnica. Recentes descobertas sugerem que os maiores danos durante a criopreservação de sêmen de garanhões ocorrem devido ao desequilíbrio osmótico e às espécies reativas de oxigênio. Um método para combater os efeitos deletérios da congelação é utilizar substâncias que possam amenizar os danos subletais. O objetivo desse experimento foi avaliar o efeito da adição de duas substâncias com efeito protetor celular (mio-inositol e ácido ferúlico) ao meio de congelação, na qualidade do sêmen criopreservado de garanhões. Para isso foram realizadas 5 colheitas de sêmen de 5 garanhões. O sêmen foi processado para criopreservação e antes do envase foi dividido em 3 tratamentos: controle, mio-inositol (30mM) e ácido ferúlico (160 µM). As palhetas foram descongeladas e analisadas quanto a motilidade computadorizada (CASA), integridade de membrana, estado metabólico e produção de espécies reativas de oxigênio (EROs) em microscopia de epifluorescência nos tempos 0, 2 e 4h de incubação a 37°C. Previamente foram realizadas validações da técnica de mensuração simultânea da membrana plasmática e estado metabólico (sondas PI, H33342 e resazurina) e correlação entre duas técnicas de mensuração de EROs (DCFH-DA e DCFH-DA com H33342). Os dados obtidos foram avaliados no programa SAS, versão 9,3 (SAS, 2011) utilizando ANOVA e medidas repetidas no tempo, para as validações foram realizadas regressões lineares simples. A adição de mio-inositol no diluidor de congelação resultou em maior motilidade total no tempo 2h e 4h e maior motilidade progressiva nos tempos 0 e 2h de incubação pós-descongelação em relação aos outros grupos. O tratamento com mio-inositol resultou em maior quantidade de células com membrana plasmática íntegra nos tempo 0h (53,3±1,4%); 2h (50,0±1,0%) e 4h (37,9±1,5%) de incubação que o grupo controle: 0h (48,0±1,0%); 2h (44,9±1,1%); e 4h (31,5±1,7%). O ácido ferúlico prejudicou as características de motilidade (CASA) e a integridade da membrana plasmática, entretanto melhorou o estado metabólico em todos os tempos. Conclui-se que o mio-inositol melhora as características de motilidade e a integridade de membranas espermáticas do sêmen criopreservado de equinos e que o ácido ferúlico apesar de prejudicar essas características promove melhor metabolismo espermático, mensurado pela técnica de redução da resazurina. / Sperm cryopreservation is of great importance for equine production, it assists in the international semen trade and reduce costs with animal transportation. However, frozen/thawed semen has reduced longevity and functional integrity and these is an obstacle to the expansion of this technique. The improvement in quality of cryopreserved semen is very important, especially to help become available more widely new technologies, such as sperm sexing. Recent findings suggest that the greatest damage during cryopreservation of stallion semen occur due to osmotic imbalance and the reactive oxygen species (ROS). One method to mitigate the deleterious effects of freezing is using substances that might lessen the sublethal damage. The objective of this experiment was to evaluate the quality of frozen/thawed stallion sperm after using two substances that have protective effects in the freezing medium: myo-inositol and ferulic acid. Were performed five sperm collection of 5 stallions. The semen was processed and before cryopreservation it was divided into three treatments: control, myo-inositol (30 mM) and ferulic acid (160 mM). The straws were thawed and analyzed at 0, 2 and 4 h of incubation time at 37°C. It was performed computerized motility (CASA), membrane integrity, metabolic status and ROS production using an epifluorescence microscope. Before beginning of the experiment were carried out validations of the technique of simultaneous assessment of plasma membrane and metabolic state (PI probes, H33342 and resazurin) and correlation between two measurement techniques ROS (DCFH-DA and DCFH-DA with H33342). The data were analyzed using the SAS version 9.3 (SAS, 2011) with ANOVA and repeated measures. For the probes validation were used linear regressions. The addition of myo-inositol in the freezing extender resulted in higher total motility in time 2h and 4h and increased progressive cells at 0 and 2 h of incubation post-thaw compared to the other groups. Treatment with myo-inositol resulted in a higher number of cells with intact plasma membrane in time 0h (53.3 ± 1.4%); 2h (50.0 ± 1.0%) and 4h (37.9 ± 1.5%) of incubation than the control group: 0h (48.0 ± 1.0%), 2h (44.9 ± 1.1%), and 4h (31.5 ± 1.7%). The use of ferulic acid resulted in the worst characteristics of motility (CASA) and plasma membrane integrity, however improved metabolic status at all incubation times. It is concluded that myo-inositol improves sperm motility characteristics and preserves membrane integrity of equine cryopreserved semen. Ferulic acid although impairing these characteristics, promotes better sperm metabolism, measured by resazurin test.

Cancer, the second leading cause of death worldwide, is characterized by uncontrolled and abnormal cell growth. Even though researchers have made significant progress in its treatment over the past several decades, innovative therapeutic approaches that both improve patient survival and lessen the many debilitating side effects of conventional cancer treatments are vital. Accordingly, we first investigated the mechanism of interaction of a bimetallic complex, Ru(II)-Rh(III), with DNA. Non-covalent binding of Ru(II)-Rh(III) is strong and involves electrostatic and, potentially, groove binding interactions. Ru(II)-Rh(III) photobinds and photocleaves DNA through an O2-independent, metal-center mediated mechanism that could be beneficial in hypoxic tumors. Furthermore, the extent of covalent binding and cleavage of DNA, which inhibit PCR amplification, is dependent upon the strength of the non-covalent interactions. These results suggest that the toxicity of Ru(II)-Rh(III) could be selectively generated in tissues irradiated with light (e.g., a tumor). Secondly, we identified protein combinations selectively present in melanoma, which could be utilized in heteromultivalency. Heteromultivalent scaffolds display higher affinity towards cells that express a protein combination in comparison to those with only one of the proteins, which facilitates cell discrimination. Using an empirically-optimized threshold-based screening method and expression profiles of melanoma patients and normal tissues, we identified surface proteins and protein combinations that are selectively found in melanoma patients and not in normal tissues. After a preliminary validation process using the scientific literature, we used immunofluorescence to confirm differential expression of some of these combinations in established melanoma cell lines in comparison to immortalized keratinocytes controls. Finally, we investigated the resazurin assay, a method used for the evaluation of proliferation and cytotoxicity in more than 2,000 publications. We found that only ~14% of these utilized validated assay conditions, while ~40% failed to report essential analytical parameters needed for their replication. We evaluated assay parameters needed for accurate estimation of cell number in eight cell lines, and found that these are highly variable and independent of tissue type, growth kinetics, and energetic parameters. Furthermore, we obtained some insights into the biochemical reduction of resazurin and proposed minimum reporting standards, along with a sample protocol for assay validation. / PHD / Cancer, a group of diseases characterized by uncontrolled and abnormal cell growth, is the second-leading cause of death worldwide. Even though researchers have made significant progress in its treatment over the past several decades, innovative therapeutic approaches that both improve survival outcome and lessen the many debilitating side-effects of conventional cancer treatments are vital. First, we investigated the mechanism of interaction of a particular molecule, Ru(II)- Rh(III), with DNA. We found that Ru(II)-Rh(III) is strongly attracted to DNA due to its charge and an interaction with the indentations along its helix. Upon light activation only, Ru(II)-Rh(III) binds to and cleaves DNA without the need for molecular oxygen, which is scarce in tumors and can limit the activity of other drugs, and to an extent that is affected by the concentration of ions in the solution. Thus, the cytotoxic effect of Ru(II)-Rh(III) might be selectively activated in those tissues that are irradiated with light (e.g., a tumor). Secondly, we identified protein combinations selectively present in melanoma, which could be utilized in heteromultivalency. Heteromultivalent scaffolds bind strongly to cells that express a combination of proteins rather than one protein at a time, making them excellent candidates for delivering a payload in a selective manner. Using expression profiles of melanoma and normal tissues, we identified surface proteins and protein combinations that are selectively found in melanoma patients and not in normal tissues. After a preliminary validation process using the scientific literature, we used confirmed differences in the expression intensities of some of these combinations in melanoma cell lines in comparison to normal skin controls. Finally, we investigated the resazurin assay, a method used for the evaluation of cell growth and drug candidates in more than 2,000 publications. We found that only ~14% of these utilized validated assay conditions, while ~40% failed to report essential analytical parameters needed for their replication. We evaluated assay conditions for eight cell lines, and found that these are highly variable and independent of tissue type and some metabolic parameters. Furthermore, we obtained insights into the mechanism through which cells react with resazurin and proposed minimum reporting standards for publications, along with a protocol for assay validation.

Ökosystem-Ingenieure sind Organismen, deren Auswirkung auf die Funktion von Ökosystemen im Vergleich zu ihrer Anzahl und Biomasse überproportional groß ist. Ein klassisches Beispiel für Ökosystem-Ingenieure sind grabende Organismen, deren Aktivitäten (Bioturbation) sowohl die Sedimentmatrix als auch das Porenwasser in aquatischen Sedimenten beeinflussen. Solche Tiere wirken auf eine große Anzahl von biogeochemischen Prozessen in benthischen Ökosystemen ein, unter anderem auf die aerobe Atmung (Respiration). Die Respiration aquatischer Sedimente umfasst häufig über 50 % der gesamten Respiration von aquatischen Systemen und spielt eine große Rolle im globalen Kohlenstoffkreislauf. Die vorliegende Doktorarbeit beschäftigt sich mit den Auswirkungen der physikalischen Umwelt (Sedimenteigenschaften, am meistens hydraulischer Leitfähigkeit) auf die mikrobielle Respiration von Sedimenten, in denen Bioturbation durch Chironomidenlarven stattfindet. Um die Auswirkungen von Bioturbation auf Respiration zu messen und zu identifizieren, wurde eine neue Messmethode entwickelt (Kapitel 4.1). Kapitel 4.2 zeigt, dass der Einfluss von Bioturbation auf die Respiration des Sediments mit zunehmender Temperatur ansteigt. Kapitel 4.3 belegt, dass Resazurin auch für die Messung der Respiration in marinen Sedimenten geeignet ist. Kapitel 4.4 vergleicht und begutachtet die große Anzahl und Vielfalt hydrologischer, biogeochemischer und ökologischer Tracer einschließlich Resazurin. Die physikalische Umwelt (Sedimentmatrix) kontrolliert wie stark die Auswirkungen der Bioturbation auf die Respiration des Sedimentes sind. Dementsprechend liefert diese Doktorarbeit die Basis für das Verständnis der Auswirkungen benthischer Bioturbation auf Respiration und Kohlenstoffumsatz in limnischen und marinen Sedimenten. / Ecosystem engineers are organisms, whose impact on ecosystem functioning is disproportionally large compared to their abundance and biomass. A classic example of ecosystem engineers are burrowing organisms whose activities (bioturbation) affect the sediment matrix and pore solutes in aquatic sediments. Bioturbating animals are impacting on a number of biogeochemical processes in benthic ecosystems, including, among others, aerobic respiration. Respiration of aquatic sediments often comprises over 50% of the total respiration of aquatic systems, and plays a tremendous role in the global carbon cycle. The present thesis deals with the impacts of the physical environment (sediment characteristics, mainly hydraulic conductivity and grain fractions) on the (microbial) respiration of bioturbated sediments. In order to disentangle the effects of bioturbation on respiration, a novel measurement method has been developed (Chapter 4.1). Chapter 4.2 reveals that the impact of bioturbation on sediment respiration increases with increasing temperature. Chapter 4.3 shows that resazurin can also be used for the measurement of respiration in bioirrigated marine sediments. Chapter 4.4 reviews the large number and diversity of hydrological, biogeochemical and ecological tracers including resazurin. The present thesis shows that in sediments with low hydraulic conductivity (diffusion-dominated sediments) (Chapters 4.1,4.2) bioturbation is altering sediment respiration to a larger extent than in sediments with high hydraulic conductivity (advection-dominated sediment) (Chapter 4.3). The physical environment (sediment matrix) controls the intensity of the impacts of bioturbation on sediment respiration. Thus, this thesis provides a basis for understanding the impact of benthic bioturbators on respiration and carbon sequestering in freshwater and marine sediments.

A leishmaniose é uma infecção causada pelo protozoário do gênero Leishmania, que causa um impacto social e econômico elevado, sendo a segunda maior incidência mundial de doença parasitária. Com o intuito de encontrar novas substâncias ou medicamentos de menor toxicidade e de custos inferiores que poderiam ser utilizados nos tratamentos de casos de infecção e, principalmente em situações de resistência parasitária, tem sido averiguada a possibilidade de utilização de substâncias de origem natural, tanto animal como vegetal, e substâncias sintéticas. Entre as que recentemente apresentaram propriedades biológicas úteis, estão alguns derivados de lignanas dibenzilbutirolactônicas que apresentam significativa ação tripanocida. Desse modo, propusemos avaliar a atividade biológica dessas lignanas em sistema in vitro, sobre formas promastigotas e amastigotas de Leishmania amazonensis. Os compostos utilizados foram (1) 7-O-N,N-dimetiletilamino cubebina, (3) cubebina, (D) 6-6´dinitroinoquinina e (H) hinoquinina. Para avaliação da atividade das substâncias foram utilizadas duas metodologias colorimétricas, MTT e Alamar Blue® e a contagem microscópica em hemocitômetro. Pelos resultados obtidos verificamos que pela contagem microscópica das formas promastigotas, as substâncias (1) 7-O-N,N-dimetiletilamino cubebina, (3) cubebina e (D) 6-6´dinitroinoquinina apresentaram potencial leishmanicida, sendo encontrados os seguintes valores de IC50: (1) = 19,4M; (3) 3,6M; (D) = 15,5M. Entretanto, os resultados obtidos com as mesmas substâncias em formas amastigotas do parasita não demonstraram atividade leishmanicida. Os métodos colorimétricos não apresentaram correspondência com a contagem microscópica, mostrando-se ineficazes para essa classe de substância. / Leishmaniasis is an infection caused by a protozoan parasite of the genus Leishmania, being the second great incidence of parasitic diseases in the world. In the search for new drugs or substances with low costs and low toxicity wich could be used in case of infection and situations of parasite resistance, being evaluated the possibility to use natural or synthetic substances. Dibenzylbutirolactonic lignans are substances with useful biological properties presenting significant trypanocidal effect. In this way we proposed to evaluate the in vitro biological activity of the lignans in of Leishmania amazonensis promastigote and amastigote forms . The compounds used were: (1) 7-O-N,N-dimethylamine cubebine, (3) cubebine, (D) 6-6´dinitroinokinine e (H) hinokinin. For the evaluation of these substances, two colorimetric methods were empoyed: MTT and Alamar Blue® , followed by microscopic counting in haemocytometer. After the results obtained after the couting of promastigote forms, we could verify that the substances (1) 7-O-N,N-dimethylamine cubebine, (3) cubebine, (D) 6-6´dinitroinokinine, and (H) hinokinin showed leishmanicidal potential, being fouded the followed values of IC50: (1) = 19.4M; (3) 3.6M; (D) = 15.5M. On the other hand the results obtained of the same substances in the parasite amastigote forms showed no leishmanicidal effect. The colorimetric methods showed no correspondence with microscopic couting demonstrating inefficacy of this substance class.

A leishmaniose é uma infecção causada pelo protozoário do gênero Leishmania, que causa um impacto social e econômico elevado, sendo a segunda maior incidência mundial de doença parasitária. Com o intuito de encontrar novas substâncias ou medicamentos de menor toxicidade e de custos inferiores que poderiam ser utilizados nos tratamentos de casos de infecção e, principalmente em situações de resistência parasitária, tem sido averiguada a possibilidade de utilização de substâncias de origem natural, tanto animal como vegetal, e substâncias sintéticas. Entre as que recentemente apresentaram propriedades biológicas úteis, estão alguns derivados de lignanas dibenzilbutirolactônicas que apresentam significativa ação tripanocida. Desse modo, propusemos avaliar a atividade biológica dessas lignanas em sistema in vitro, sobre formas promastigotas e amastigotas de Leishmania amazonensis. Os compostos utilizados foram (1) 7-O-N,N-dimetiletilamino cubebina, (3) cubebina, (D) 6-6´dinitroinoquinina e (H) hinoquinina. Para avaliação da atividade das substâncias foram utilizadas duas metodologias colorimétricas, MTT e Alamar Blue® e a contagem microscópica em hemocitômetro. Pelos resultados obtidos verificamos que pela contagem microscópica das formas promastigotas, as substâncias (1) 7-O-N,N-dimetiletilamino cubebina, (3) cubebina e (D) 6-6´dinitroinoquinina apresentaram potencial leishmanicida, sendo encontrados os seguintes valores de IC50: (1) = 19,4M; (3) 3,6M; (D) = 15,5M. Entretanto, os resultados obtidos com as mesmas substâncias em formas amastigotas do parasita não demonstraram atividade leishmanicida. Os métodos colorimétricos não apresentaram correspondência com a contagem microscópica, mostrando-se ineficazes para essa classe de substância. / Leishmaniasis is an infection caused by a protozoan parasite of the genus Leishmania, being the second great incidence of parasitic diseases in the world. In the search for new drugs or substances with low costs and low toxicity wich could be used in case of infection and situations of parasite resistance, being evaluated the possibility to use natural or synthetic substances. Dibenzylbutirolactonic lignans are substances with useful biological properties presenting significant trypanocidal effect. In this way we proposed to evaluate the in vitro biological activity of the lignans in of Leishmania amazonensis promastigote and amastigote forms . The compounds used were: (1) 7-O-N,N-dimethylamine cubebine, (3) cubebine, (D) 6-6´dinitroinokinine e (H) hinokinin. For the evaluation of these substances, two colorimetric methods were empoyed: MTT and Alamar Blue® , followed by microscopic counting in haemocytometer. After the results obtained after the couting of promastigote forms, we could verify that the substances (1) 7-O-N,N-dimethylamine cubebine, (3) cubebine, (D) 6-6´dinitroinokinine, and (H) hinokinin showed leishmanicidal potential, being fouded the followed values of IC50: (1) = 19.4M; (3) 3.6M; (D) = 15.5M. On the other hand the results obtained of the same substances in the parasite amastigote forms showed no leishmanicidal effect. The colorimetric methods showed no correspondence with microscopic couting demonstrating inefficacy of this substance class.

The importance of dissolved organic carbon (DOC) to aquatic environments is well established in the scientific community. In boreal landscapes, small streams receive water from headwater lakes, mires, and discrete flow paths that drain riparian soils. The goal of this study was to investigate the importance of these discrete riparian inputs (DRIPs) as sources of DOC and to explore whether quantity and quality of DOC from DRIPs differs from other sources in the landscape, including groundwaters that are not as hydrologically connected to streams. To do this, I collected water from already established riparian groundwater wells installed at the Krycklan Catchment Study (KCS) in northern Sweden, as well as from an adjacent lake, stream, and mire. Microbial activity (respiration) was analyzed in 24-hour laboratory incubations using a metabolically active dye, resazurin (Raz) which in the presence of aerobic respiration transforms into resorufin (Rru). Rru is easily measured in the lab, and its production can serve as a proxy for rates of microbial respiration. DOC concentration was also measured at each location, along with specific absorbance at 254 nm (SUVA254) and the absorbance ratio (254/365 nm) as indices of DOC quality. The results show a large variation in DOC concentration among potential water sources to the stream. Furthermore, there was a strong correlation (R2=0.96) between Rru production and DOC concentration among these sources, but no significant difference (p=0.067) in median Rru production between DRIPs and non-DRIPs. Overall, these results highlight important spatial variability in DOC from different water sources in the landscape, which likely have important consequences for patterns of microbial respiration in streams.