Methods

I. Stream chemistry sample collection methods: core-sediment core was taken from the benthic zone of the streamgeopump-geopump used to pump stream water into collection bottlegrab-collection bottle filled with stream water by hand and filtered in the fieldgrabfilter- stream water collected by hand and filtered in field. Unfiltered and filtered samples placed in separate collection bottles.isco- sample collected by use of an ISCO automated samplerpoint- sampled collected by method outlined in patent US8337121sedimentgrab- sediment sample taken in field by hand and placed in collection bottlesyringe- sample collected from stream by syringe and placed in collection bottlesyringe_filter- sample collected from stream by syringe filter. Unfiltered and filtered samples placed in separate collection bottles. II. Stream chemistry analytical methods: All water samples were kept on ice and in the dark following collection, then were either acidified (TN/TP, TDN/TDP) or frozen until analysis (all other analytes).no32_2- This is NO_3-N which is operationally defined as nitrate nitrogen + nitrite nitrogen. Determined by flow injection analysis on Astoria Pacific Instruments Autoanalyzer (APIA).nh4_n, tn1, tp1, tdn, tdp- All analytes measured by flow injection analysis on Astoria Pacific Instruments Autoanalyzer (APIA).srp- measured colorometrically using the molybdate blue method [APHA 1995] and a Beckman spectrophotometer.doc- measured using a Shimadzu carbon analyzer.doc_qual- the goal in doing this analysis is to determine the source of dissolved organic carbon (doc) measured in a particular riverine ecosystem. This was achieved by UV absorbance which provides an estimate of the aromaticity of the doc in a sample, and by extension, the potential source of the doc.cl, no2, no3, br, and so4- all measured by ion chromatography. See http://www.nemi.gov; method number 4110C. Detection limits for method number 4110C: cl-20&micro;g/l, no2-15&micro;g/l, no3-17&micro;g/l, br-75&micro;g/l, and so4-75&micro;g/l.ysi_cond, do, ph_field, wtemp- all measured by use of a standard YSI meter.tss- measured by standard methods. A thoroughly mixed sample is filtered and dried at 103-105 degreesCelcius. The obtained residue represents the amount of solids suspended in the sample solution. See http://www.nemi.giv; method number D5907.tot_om- measured by standard methods. The residue obtained from the tss procedure is ignited at 550 degreesCelcius and weighed, the difference in weight representing total volatile solids. Total volatile solids represents the portion of the residue that is composed of organic molecules. See http://www.nemi.gov; method number 160.4.turbid- measured by use of a nephelometer. III. Big Spring Sediment Coring Methods A. Field Methods- collecting sediment coresSediment core samples taken with WDNR piston core samplerB. Sediment Analysis- HydrometerDocumentation: Robertson, G.P., Coleman, D.C., Bledsoe, C.S. and Sollins, P., 1999. Standard Soil Methods for Long-Term Ecological Research. Oxford University Press, New York, 462 pp.Hydrometer Analysis- procedure used to determine percent clay:<p style="margin-left:.25in;">1. Dry the sample in a pre-weighed aluminum pan for at least 24 hr at 105 C. Make sure sample is completely dry before weighing.<p style="margin-left:.25in;">2. Weigh the dried sample, then ash for at least 8 hr at 550 C. Make sure to break up any large clumps before ashing.<p style="margin-left:.25in;">3. Weigh the ashed sample, then crush any aggregates with a pestal. Mix sample thoroughly.<p style="margin-left:.25in;">4. Transfer 40g, plus or minus one gram, of the sample into a 500mL wide mouth bottle<p style="margin-left:.25in;">5. Add 10g of sodium hexametaphosphate to the bottle.<p style="margin-left:.25in;">6. Add approx 200mL of deionized water to bottle. Shake vigorously with hand.<p style="margin-left:.25in;">7. Stir samples on shaker table for at least 8 hr at speed 40. Putting them in a box and fastening with bungee cords works best.<p style="margin-left:.25in;">8. Transfer sample to 1L cylinder, making sure to get all of sample out of bottle. Fill cylinder with deionized water up to the 1L mark.<p style="margin-left:.25in;">9. Prepare a blank cylinder by adding 10g of sodium hexametaphosphate and filling to 1L.<p style="margin-left:.25in;">10. Allow all cylinders to equilibrate to room temperature ( approx 30 min).<p style="margin-left:.25in;">11. Starting with the blank cylinder, put stopper into cylinder and shake end-over-end for approx 5 min. Rinse stopper. Repeat this step for all cylinders, rinsing stopper between cylinders.<p style="margin-left:.25in;">12. Record the time that you stopped shaking each cylinder.<p style="margin-left:.25in;">13. At 1.5 hr from time of shaking, record temperature and hydrometer level of the blank cylinder. Then record the 1.5 hr hydrometer level for each successive cylinder.<p style="margin-left:.25in;">14. At 24 hr from time of shaking, record temperature and hydrometer level of the blank cylinder. Then record the 24 hr hydrometer level for each successive cylinder. Sieve Analysis- procedure used to determine quantity of sand and silt<p style="margin-left:.25in;">1. After hydrometer analysis, pour the entire sample into the .063mm sieve. Rinse the sample thoroughly until all the clay is out. Try to break up any clay clumps you see.<p style="margin-left:.25in;">2. Transfer the sample to a pre-weighed and labeled aluminum pan. You will probably need to backwash the sieve to get the entire sample out. You can use a syringe to pull water from the pan if it gets too full. Dry the sample for 48 hours at 50-60C.<p style="margin-left:.25in;">3. Before transferring the dried sample to the sieves, make sure you pre-weigh the sieves and put their weight on the data sheet. You will need to do this before every sample as you might not get all the sample out of the sieves from the previous sample. Stack the sieves in the following order, top to bottom : 4mm, 2mm, 1mm, 0.5mm, 0.25mm, 0.125mm, 0.063mm, and pan. Pour the sample into the top sieve. Place the lid on, located on sieve shaker, and put the stack of sieves into the sieve shaker. Fasten the tie downs. Set shaker for 3 minutes. <p style="margin-left:.25in;">4. Remove stack of sieves from shaker. It&rsquo;s ok to leave the pan behind temporarily as it might be tight. Weigh each sieve and record the weight in the data sheet. If you see any clay clumps, break them up with your fingers and re-shake the stack a little, using hands is okay.<p style="margin-left:.25in;">5. Dump the sample out in the trash and clean the sieve with the brush. At the end of the day it might be necessary to backwash the sieves with water and dry overnight in the oven. <p style="margin-left:.25in;"> Calculations:1. percent clay was determined by the hydrometer analysis- P1.5, P24, X1.5, X24, and m are the variables that were calculated to determine percent clay by the hydrometer analysis.P1.5= ((sample hydrometer reading at 1.5 hours- blank hydrometer reading at 1.5 hours)/ (sample weight)) multiplied by 100.P24= ((sample hydrometer reading at 24 hours- blank hydrometer reading at 24 hours)/ (sample weight)) multiplied by 100X1.5= 1000*(.00019*(-.164* (sample hydrometer reading at 1.5 hours)+16.3)² *8100X24=1000*(.00019*(-.164* (sample hydrometer reading at 24 hours)+16.3)² *8100m= (P1.5-P24)/(ln(X1.5/X24))percent clay = m * ln(2/X24)) + P24clay (grams) = total weight * ( percent clay/ 100)2. percent Sand and percent Silt were determined based on the results of the sieve analysis which determined the grams of sand and silt.percent sand= total weight * (percent sand/ 100)percent silt= total weight * (percent silt/ 100)3. Othersorganic matter (grams) was calculated in this analysis as dry weight (grams) &ndash; ashed weight (grams)percwnt organic matter was calculated as ((organic matter (grams))/(total dry weight (grams)) multiplied by 100 C. Sediment Chemical Analysis1. SRP/ NaOH-PChemical analysis was done according to the protocol outlined in Pionke and Kunishi (1992). Each sample was first centrifuged and separated into aqueous and sediment fractions. The sediment fraction was then dried. The aqueous fraction was analyzed for soluble reactive phosphorus (srp) by automated colorimetry Nemi Method Number 365.4; see http://www.nemi.gov. NaOH P was then determined by NaOH extractions as described in Pionke and Kunishi (1992). Documentation: Pionke HB, Kunishi HM (1992) Phosphorus status and content of suspended sediment in a Pennsylvania watershed. Soil Sci 153:452&ndash;462.2. NH4 / KCl-NH4 The exact procedure that was used to analyze samples for ammonium is unknown. However, it is known that a KCl extraction was used. The KCl-NH4 was calculated as the concentration of ammonium in milliGramsPerLiter divided by the sediment weight in grams. 3. NO3 / KCl-NO3The exact procedure that was used to analyze samples for nitrate is also unknown. Again, it is known that a KCL extraction was used. The KCl-NO3 was calculated as the concentration of nitrate in milliGramsPerLiter divided by the sediment weight in grams.Note: The same sediment sample was used to measure ammonium and nitrate IV. Big Spring Creek Longitudinal Profile A standard longitudinal stream profile was conducted at Big Spring Creek, WI (wbic=176400) on unknown date(s). It is speculated that the profile was done during the summer of 2005, during which the rest of the data for Big Spring Creek was collected. Measurements for the profile began at the Big Spring Dam site (43.67035,-89.64225), a dam which was subsequently removed. The first (x_dist, y_dist) of (2.296, 5.57) corresponds to the location where the stream crosses Golden Court Road, whereas the second coordinate pair of (-2.615, -36.303) corresponds to the point below the previous Big Spring Creek Dam site. The third (x_dist, y_dist) of (-9.472, 7.681) corresponds to the top of the dam gates and is assigned a distance=0 as it is the starting point.

Methods

Inorganic and organic carbon: Samples for inorganic and organic carbon are collected together with a peristaltic pump and tubing and in-line filtered, if necessary, (through a 0.40 micron polycarbonate filter) into glass, 24 ml vials (that are compatible with the carbon analyzer autosampler), and capped with septa, leaving no head space. The samples are stored refrigerated at 4 degrees Celsius until analysis, which should occur within 2-3 weeks. The detection limit for inorganic carbon is 0.15 ppm, and the analytical range for the method is 60 ppm. The detection limit for organic carbon is 0.30 ppm and the analytical range for the method is 30 ppm. Method Log: Prior to May 2006 samples, inorganic carbon was analyzed by phosphoric acid addition on an OI Model 700 Carbon Analyzer. From May 2006 to present, inorganic carbon is still analyzed by phosphoric acid addition, but on a Shimadzu TOC-V-csh Total Organic Carbon Analyzer. Method Log: Prior to May 2006 samples, organic carbon was analyzed by heated persulfate digestion on an OI Model 700 Carbon Analyzer. From May 2006 to present, Organic carbon is analyzed by combustion, on a Shimadzu TOC-V-csh Total Organic Carbon Analyzer. Dissolved reactive silica Samples for silica are collected with a peristaltic pump and tubing and in-line filtered (through a 45 micron polycarbonate filter) into 120 ml LDPE bottles and acidified to a 1percent HCl matrix by adding 1 ml of ultra pure concentrated HCl to 100 mls of sample. For every sample acidification event, three acid blanks are created by adding the same acid used on the samples to 100 mls of ultra pure water supplied from the lab. Once acidified, the samples are stable at room temperature until analysis, which should occur within one year. Until acidification, the samples should be refrigerated at 4 degrees Celsius. Dissolved reactive silica is determined by the Heteropoly Blue Method and the absorption is measured at 820 nm. The detection limit for silica is 6 ppb and the analytical range is 15000 ppb. Method Log These determinations were performed manually using a Bausch and Lomb Spectrophotometer from the beginning of the project until April 1984. From 1984 through 2005, dissolved reactive silica was determined on a Technicon Auto Analyzer II. From January 2006 to present, samples are run on an Astoria-Pacific Astoria II Autoanalyzer. total and dissolved nitrogen and phosphorus Samples for total and dissolved nitrogen and phosphorus analysis are collected together with a peristaltic pump and tubing and in-line filtered, when necessary, (through a 45 micron polycarbonate filter) into 120 ml LDPE bottles and acidified to a 1percent HCl matrix by adding 1 mL of ultra pure concentrated HCl to 100 mls of sample. For every sample acidification event, three acid blanks are created by adding the same acid used on the samples to 100 mls of ultra pure water supplied from the lab. Once acidified, the samples are stable at room temperature until analysis, which should occur within one year. Until acidification, the samples should be refrigerated at 4 degrees Celsius. The samples must first be prepared for analysis by adding an NaOH–Persulfate digestion reagent and heated for an hour at 120 degrees C and 18-20 psi in an autoclave. The samples are analyzed for total nitrogen and total phosphorus simultaneously by automated colorimetric spectrophotometry, using a segmented flow autoanalyzer. Total nitrogen is determined by utilizing the automated cadmium reduction method, as described in Standard Methods, where the absorption is monitored at 520 nm. The detection limit for total and dissolved nitrogen is approximately 21 ppb and the analytical range for the method extends to 2500 ppb. The detection limit for total phosphorus is approximately 3 ppb and the analytical range for the method extends to 800 ppb. Method Log: Prior to January 2006 samples, total nitrogen was determined on a Technicon segmented flow autoanalyzer. From 2006 to present, total nitrogen is determined by an Astoria-Pacific Astoria II segmented flow autoanalyzer.

Methods

Site SelectionBecause lakes are a dominant feature of the region and stream characteristics could potentially differ based on their hydrologic connections to lakes, we classified streams into three categories as a function of their hydrologic connections to lakes. The first category was streams that had no lakes within the drainage network upstream of the sampling location. The second category was streams that originated from headwater lakes (i.e., no stream inlet but a stream outlet) and the headwater lake was the only lake in the drainage network above the sampling location. The final category had at least a single drainage lake (i.e., a lake with both stream inlet(s) and outlet) in the drainage network above the sampling location. We then used these categories to select sampling sites using a stratified random design for a variety of chemical and physical characteristics.All streams identified on 1:24,000 7.5 inch USGS topographical maps that crossed access points were selected as potential sampling locations and assigned to one of the three stream types. A stream could be classified by more than a single category depending on the sampling location within the drainage network. However, a single drainage network was never sampled more than once to ensure sample independence. Of the 500 possible sampling locations, 52 sites were selected and sampled.SamplingAll streams were sampled 7-10 channel widths upstream of an access point to minimize any influences caused by culverts and other features. Water samples were collected from the center of the channel using a peristaltic pump. Stream discharge was measured after Gore (2007) using cross sectional area and water velocity.Chemical AnalysesAll samples for both studies were collected and processed following the North Temperate Long Term Ecological Research (NTL-LTER) protocols (http://lter.limnology.wisc.edu). Filtering was done in the field using an in-line 0.45 μm membrane filter. All samples were stored on ice and returned to the laboratory where they were preserved according to NTL-LTER protocols. Acid neutralizing capacity (ANC) was determined by Gran titration (APHA 2005). DOC was measured on a Shimadzu TOC-V carbon analyzer. Total nitrogen and phosphorus (unfiltered, TN and TP; filtered, TDN and TDP), nitrate+nitrite (NO3-N), and ammonium (NH4-N) were quantified with an Astoria-Pacific segmented flow auto-analyzer. Soluble reactive phosphorus (SRP) in streams was measured colormetrically on a Beckman DU-800 spectrophotometer (APHA 2005). Anions (Cl- and SO4 2-) were measured using a Dionix DX-500 ion chromatograph and cations (Ca, Mg, Na, K, Fe, K, and Mn) on a Perkin Elmer ICP mass spectrometerDissolved inorganic carbon (DIC) and pH were quantified differently in the lakes and stream data sets. For the lakes data, DIC was determined with a Shimadzu TOC-V carbon analyzer, whereas DIC for the streams dataset was determined by headspace equilibration of acidified water samples in the field and direct measurement of carbon dioxide (CO2) gas on a Shimadzu gas chromatograph (Cole et al. 1994). pH measurements for the lakes dataset were quantified on non-air equilibrated samples in the lab with a Accumet 950 pH meter while direct measurements were taken in the field for the streams dataset using a hand-held Orion model 266 pH meter that was allowed to equilibrated about 20 min in the center for the stream channel.Several variables presented in this study were determined from calculations based on measured values. In streams, dissolved organic nitrogen and phosphorus (DON and DOP, respectively) were determined by the difference between inorganic nutrients and total dissolved nutrients (e.g., DOP = TDP-SRP). We were unable to determine DON in lakes due to the lack of inorganic nitrogen data. It was assumed that DOP approximately equals TDP in lakes because dissolved inorganic phosphorus concentrations in the region are typically below detection limits in the epilimnion during the summer months and consequently not quantified (NTL-LTER unpublished data).

Methods

Samples for nitrateornitrite and ammonium are collected together with a peristaltic pump and tubing and in-line filtered (through a 40 micron polycarbonate filter) into new, 20 ml HDPE plastic containers with conical caps. The samples are stored frozen until analysis, which should occur within 6 months. The samples are analyzed for nitrateornitrite (and ammonium) simultaneously by automated colorimetric spectrophotometry, using a segmented flow autoanalyzer. Nitrateornitrite is determined by utilizing the automated cadmium reduction method, as described in Standard Methods, where the absorption is monitored at 520 nm.The detection limit for nitrateornitrite is approximately 2 ppb and the analytical range for the method extends to 4000 ppb.Ammonium is determined by utilizing the Berthelot Reaction, producing a blue colored indophenol compound, where the absorption is monitored at 660 nm.The detection limit for ammonium is approximately 3 ppb and the analytical range for the method extends to 4000 ppb.Method Log: Prior to January 2006 samples, nitrateornitrite was determined on a Technicon segmented flow autoanalyzer. From 2006 to present, nitrateornitrite is determined by an Astoria-Pacific Astoria II segmented flow autoanalyzer.Samples for total and dissolved phosphorus and nitrogen analysis are collected together with a peristaltic pump and tubing and in-line filtered, when necessary, (through a 40 micron polycarbonate filter) into 120 ml LDPE bottles and acidified to a 1percent HCl matrix by adding 1 mL of ultra pure concentrated HCl to 100 mls of sample. For every sample acidification event, three acid blanks are created by adding the same acid used on the samples to 100 mls of ultra pure water supplied from the lab. Once acidified, the samples are stable at room temperature until analysis, which should occur within one year. Until acidification, the samples should be refrigerated at 4 degrees Celsius.The samples must first be prepared for analysis by adding an NaOH&ndash;Persulfate digestion reagent and heated for an hour at 120 degrees C and 18-20 psi in an autoclave.The samples are analyzed for total nitrogen and total phosphorus simultaneously by automated colorimetric spectrophotometry, using a segmented flow autoanalyzer. Total phosphorus is determined by forming a phosphoantimonylmolybdenum complex and the absorption is monitored at 880 nm.The detection limit for total phosphorus is approximately 3 ppb and the analytical range for the method extends to 800 ppb.Method Log: Prior to January 2006 samples, total phosphorus was determined on a Technicon segmented flow autoanalyzer. From 2006 to present, total phosphorus is determined by an Astoria-Pacific Astoria II segmented flow autoanalyzer.

Methods

Methods for 1984-1990 were described by Carpenter and Kitchell (1993) and methods for 1991-1997 were described by Carpenter et al. (2001).Carpenter, S.R. and J.F. Kitchell (eds.). 1993. The Trophic Cascade in Lakes. Cambridge University Press, Cambridge, England.Carpenter, S.R., J.J. Cole, J.R. Hodgson, J.F. Kitchell, M.L. Pace,D. Bade, K.L. Cottingham, T.E. Essington, J.N. Houser and D.E. Schindler. 2001. Trophic cascades, nutrients and lake productivity: whole-lake experiments. Ecological Monographs 71: 163-186.Additional methods are described in the Cascade Methods manual, which can be found here:http://c13.valuemembers.net/Pages/methods_09.html