A Critical Assessment of Relating Resazurin–Resorufin Experiments to Reach‐Scale Metabolism in Lowland Streams

Abstract

The reactive tracer resazurin is frequently used to investigate metabolic processes and hyporheic exchange in streams, because its transformation to resorufin is considered a function of oxygen turnover in the presence of living cells and thus of ecosystem respiration. This has been investigated and confirmed in a number of laboratory and batch studies but so far not unanimously been verified in the field. In this study we conducted 13 tracer tests with resazurin in different small, low-gradient streams typical of lowland regions. We determined tracer processing rates and ratios based on metrics commonly used to assess reactive-tracer tests with resazurin and compared them to flow characteristics and respiration rates determined from diel-oxygen recordings following a two-station approach. We found no statistical relationship between tracer processing and parameters of transport, indicating that resazurin-to-resorufin transformation cannot be explained by hydrological parameters alone. The statistical relationship between resazurin processing and respiration rates based on diel-oxygen profiles was strongest if resazurin-to-resorufin transformation ratios were used, and uptake lengths were also found to be a good proxy. However, the relationships were not linear, and weak for individual streams, contrary to what was expected based on previous laboratory and batch studies. We attribute this finding to a lack of process understanding of resazurin reactions under field conditions, and generally insignificant hyporheic exchange fluxes in lowland streams. Our findings highlight important limitations of the tracer technique under field conditions. Plain Language SummaryResazurin is a compound that can be used to track the flow of water and metabolic processes in rivers. This has been tested in laboratory experiments before, but until now the method has not been fully verified in the field. In this study we apply resazurin to 13 different small rivers and measure its transformation. We compare our results to other, independent methods that are often used to determine metabolic processes. We find good relationships between the two different methods for all rivers together, but not for individual rivers. Also, the relationships are not linear, which makes predicting metabolic rates from the transformation of resazurin difficult.

Solute Transport and Transformation in an Intermittent, Headwater Mountain Stream with Diurnal Discharge Fluctuations

“…Vegetation in the basin is primarily red alder in the valley bottom, with stands of Douglas fir of varied ages and shrubs on the hillslopes [24]. Stream discharge varied due to evapotranspiration in the basin [5,[22][23][24]95,[103][104][105][106][107][108], but contiguous surface flow along the reach was never observed (i.e., the reach was spatially intermittent for the entire study period). We monitored the maximum and minimum extent of surface water during the study, flagging the most expansive and contracted conditions within the study reach and mapping their positions with a topographic survey ( Figure 1).…”

Spatiotemporal Variability in Transport and Reactive Processes Across a First‐ to Fifth‐Order Fluvial Network

“…In the presence of mildly reducing conditions, Raz undergoes an irreversible reduction to its daughter product resorufin, Rru, and experimental work with microbial communities has shown that (1) the transformation of Raz to Rru is proportional to oxygen consumption (González‐Pinzón et al, 2012) and (2) the mass balance of Raz and Rru closes over time scales relevant for field experiments (Dallan et al, 2020). Therefore, the transformation from Raz to Rru has been used as a proxy for in situ aerobic metabolism and water‐sediment interactions in the last decade (Argerich et al, 2011; González‐Pinzón et al, 2014, 2016; González‐Pinzón et al, 2015b; Knapp & Cirpka, 2018; see review by Knapp et al, 2018).…”

“…(2) the mass balance of Raz and Rru closes over time scales relevant for field experiments (Dallan et al, 2020). Therefore, the transformation from Raz to Rru has been used as a proxy for in situ aerobic metabolism and water-sediment interactions in the last decade (Argerich et al, 2011;González-Pinzón et al, 2014González-Pinzón et al, 2015b;Knapp & Cirpka, 2018; see review by .…”

“…These tests were done in five different stream orders along the Jemez River, New Mexico (Figure 2a), during two different flow regimes (i.e., summer baseflow and spring higher flow). The Raz‐Rru system was chosen for this study because the irreversible transformation of Raz to Rru provides an in situ estimation of hyporheic zone processing as a fraction of riverine processing through an estimation of metabolic activity (Dallan et al, 2020 ; Haggerty et al, 2008; González‐Pinzón et al, 2012, 2014, 2016; González‐Pinzón et al, 2015b; Knapp & Cirpka, 2018). While not all the parameters describing reactive transport across the Jemez River network varied consistently with stream order or discharge, we found a trend toward more reaction‐limited conditions with increasing discharge across all stream orders, as Damköhler numbers consistently decreased along the fluvial network in response to decreasing processing rate coefficients.…”

Modeling the Fate of Pharmaceuticals in a Fourth‐Order River Under Competing Assumptions of Transient Storage

“…A key problem is that the most successful studies on hyporheic exchange and the reactivity of the hyporheic zone were performed in small, comparably steep, low-order streams whereas waste-water treatment plants are typically located at somewhat larger, less steep, higher-order rivers with little hyporheic exchange. Also, the transferability of resazurin reactivity to aerobic metabolism is limited (Knapp & Cirpka, 2018). Finally, there is no experience and reasonable way to relate or transfer any such rates to the turnover rate of emerging pollutants.…”