7 

Prepared in cooperation with the National Park Service

Denitrification Rates in Marsh Soils and Hydrologic and 

Water Quality Data for Northeast Creek and Bass Harbor 

Marsh Watersheds, Mount Desert Island, Maine

Open-File Report 2011–1252

U.S. Department of the Interior

U.S. Geological Survey
Cover. Denitrification measurement site in Bass Harbor Marsh, Acadia National Park, Mount Desert Island, Maine showing chambers installed in 

the marsh soil used to measure the accumulation of nitrous oxide (N2O) in the chamber headspace. The batteries shown are used to power small fans 

in each chamber to maintain well mixed gases in the chambers.
In cooperation with the National Park Service

Denitrification Rates in Marsh Soils and Hydrologic and 

Water-Quality Data for Northeast Creek and Bass Harbor 

Marsh watersheds, Mount Desert Island, Maine

By Thomas G. Huntington, Charles W. Culbertson, and John H. Duff

Open-File Report 2011–1252

U.S. Department of the Interior

U.S. Geological Survey
ii

U.S. Department of the Interior

KEN SALAZAR, Secretary

U.S. Geological Survey

Marcia K. McNutt, Director

U.S. Geological Survey, Reston, Virginia 2011

For product and ordering information:

World Wide Web: http://www.usgs.gov/pubprod

Telephone: 1-888-ASK-USGS

For more information on the USGS—the Federal source for science about the Earth,

its natural and living resources, natural hazards, and the environment:

World Wide Web: http://www.usgs.gov

Telephone: 1-888-ASK-USGS

Suggested citation:

Huntington, T.G., Culbertson, C.W., and Duff, J.H., 2011, Denitrification rates in marsh soils and hydrologic and water 

quality data for Northeast Creek and Bass Harbor Marsh watersheds, Mount Desert Island, Maine: U.S. Geological 

Survey Open-File Report 2011–1252, 28 p. at http://pubs.usgs.gov/of/2011/1252/. 

Any use of trade, product, or firm names is for descriptive purposes only and does not imply 

endorsement by the U.S. Government.

Although this report is in the public domain, permission must be secured from the individual 

copyright owners to reproduce any copyrighted material contained within this report.
iii

Acknowledgements

The authors are grateful for the support they received from several people that contributed to the successful completion of this 

study. Staff at Acadia National Park (David Manski and Bill Gawley) helped facilitate this study from the initial conceptualization 

through its completion. Staff at the U.S. Geological Survey, Maine Water Science Center (James Caldwell, Charles Schalk, 

Laura Flight, and Greg Stewart) provided assistance with groundwater level data processing and interpretation, overall data 

review, and data uploading into the Survey’s National Water Information System database. Lynn Bartsch and James Caldwell, 

both with the U.S. Geological Survey, provided technical reviews that improved the report. 
iv

Contents

Abstract ......................................................................................................................................................................... 1

Introduction.................................................................................................................................................................... 1

Description of Study Area .......................................................................................................................................... 3

Purpose and Scope ................................................................................................................................................... 4

Data Collection Methods................................................................................................................................................ 7

Denitrification Methods............................................................................................................................................... 7

Field Denitrification Methods .................................................................................................................................. 7

Laboratory Denitrification Methods......................................................................................................................... 9

Laboratory Methods and Computation of N2O Release Rate................................................................................14

Water Quality Data Collection Methods ....................................................................................................................14

Field Methods........................................................................................................................................................15

Laboratory Methods ..............................................................................................................................................16

Ancillary Data Collection Methods ............................................................................................................................20

Field Denitrification Data...............................................................................................................................................20

Laboratory Denitrification Data .....................................................................................................................................21

Water-Quality Data .......................................................................................................................................................22

Quality-Control Samples...............................................................................................................................................22

Ancillary Data ...............................................................................................................................................................23

References Cited..........................................................................................................................................................25

Figures

Figure 1. Shaded relief site map showing locations of the study areas that include Northeast Creek (NEC) and 

Bass Harbor Marsh (BHM) watersheds on Mount Desert Island, Maine........................................................................ 4

Figure 2. Map showing the Northeast Creek watershed area, Mount Desert Island, Maine, and surface-water 

sampling locations on the major tributaries and the location of a groundwater monitoring well..................................... 5

Figure 3. Map showing the location of Bass Harbor Marsh watershed, Mount Desert Island, Maine, and surfacewater sampling locations on the major tributaries.......................................................................................................... 6

Purpose and Scope ....................................................................................................................................................... 7

Figure 4. Aerial photograph showing the locations of denitrification measurement sites and groundwater sampling 

wells in Northeast Creek watershed, Mount Desert Island, Maine................................................................................. 9

Figure 5. Aerial photograph showing the locations of denitrification measurement sites and groundwater sampling 

wells in Bass Harbor Marsh watershed, Mount Desert Island, Maine.......................................................................... 10

Figure 6. Graphs showing A, Water level in Fresh Meadow Marsh, Northeast Creek watershed, Mount Desert 

Island, Maine and discharge at Otter Creek USGS station ID 01022840 and B, daily precipitation at McFarland Hill 

Station, near Bar Harbor, Mount Desert Island, Maine. ............................................................................................... 24

Tables

Table 1. Dates of field (in situ) denitrification rate measurements, locations of measurements and treatments 

applied on those measurement dates.......................................................................................................................... 11

Table 2. Surface water and groundwater water-quality monitoring stations in Northeast Creek and Bass Harbor 

Marsh watersheds sampled in 2008 and 2009 ............................................................................................................ 16

Table 3. Constituent name, U.S. Geological Survey National Water Information System parameter codes 

and method codes, and minimum reporting limits for temperature, specific conductance, dissolved oxygen, and 

nutrients ................................................................................................................................................................. 18
v

Tables 4–27 (links to Excel files tables 4–27 are in text) 

Table 4. Denitrification rate measurements for June 2008

Table 5. Denitrification rate measurements for July 2008

Table 6. Denitrification rate measurements for August 2008

Table 7. Denitrification rate measurements for September 2008

Table 8. Denitrification rate measurements for October 2008

Table 9. Denitrification rate measurements for June 2009

Table 10. Denitrification rate measurements for July 2009

Table 11. Denitrification rate measurements for August 2009

Table 12. Denitrification rate measurements for September 2009

Table 13. Water, organic matter, and dry weight fractions (by weight) of marsh soils.

Table 14. Laboratory incubation denitrification rate measurements for May 5, 2009

Table 15. Laboratory incubation denitrification rate measurements for May 7, 2009

Table 16. Laboratory incubation denitrification rate measurements for May 13, 2009

Table 17. Laboratory incubation denitrification rate measurements for May 15. 2009

Table 18. Laboratory incubation denitrification rate measurements for May 20, 2009

Table 19. Laboratory incubation denitrification rate measurements for May 22, 2009

Table 20. Laboratory incubation denitrification rate measurements for May 27, 2009

Table 21. Laboratory incubation denitrification rate measurements for May 29, 2009

Table 22. Laboratory incubation denitrification rate measurements for June 2, 2009

Table 23. Nutrient water-quality data for stream water samples

Table 24. Field measurements of temperature and specific conductance for stream water samples

Table 25. Nutrient water-quality data for groundwater samples collected from NEC watershed

Table 26. Nutrient water-quality data for groundwater samples collected from BHM watershed

Table 27. Field measurements of temperature and specific conductance for groundwater samples
vi

Conversion Factors

Inch/Pound to SI

Multiply By To obtain

Length

inch (in.) 2.54 centimeter (cm)

mile (mi) 1.609 kilometer (km)

inch (in.) 25.4 millimeter (mm)

foot (ft) 0.3048 meter (m)

inch (in.) 3.937 * 10-5 micron (μ)

Area

acre 4,047 square meter (m2)

square mile (mi2) 2.590 square kilometer (km2) 

square foot (ft2) 0.09290 square meter (m2)

Volume

gallon (gal) 3.785 liter (L) 

ounce, fluid (fl. oz)) 29.57 milliliter (mL) 

cubic inch (in3) 16.39 cubic centimeter (cm3) 

Mass

ounce, avoirdupois (oz) 28.35 gram (g) 

ounce, avoirdupois (oz) 0.02835 milligram (mg) 

Application rate

ounce, avoirdupois (oz) per square 

yard [(oz/ yd2] 

0.02949 grams per square meter

(g/ m2) 

Temperature in degrees Celsius (°C) may be converted to degrees Fahrenheit (°F) as follows:

°F=(1.8×°C)+32

Temperature in degrees Fahrenheit (°F) may be converted to degrees Celsius (°C) as follows:

°C=(°F-32)/1.8

Vertical coordinate information is referenced to the North American Vertical Datum of 1988 (NAVD 88).

Horizontal coordinate information is referenced to the North American Datum of 1983 (NAD 83).

Altitude, as used in this report, refers to distance above the vertical datum.

Specific conductance is given in microsiemens per centimeter at 25 degrees Celsius (μS/cm at 25 °C).

Concentrations of chemical constituents in water are given either in milligrams per liter (mg/L) or micrograms per liter (μg/L).
vii

List of Acronyms

ANP Acadia National Park 

BHM Bass Harbor Marsh

DIN dissolved inorganic nitrogen 

GC-ECD gas chromatography electron capture detection 

NADP National Atmospheric Deposition Program 

NEC Northeast Creek

NETN Northeast Temperate Network 

NOAA National Oceanic and Atmospheric Administration 

NWQL National Water Quality Laboratory 

OM organic matter 

RPD relative percent differences 

SE standard error

UHP ultra high purity 

USGS U.S. Geological Survey

WHOI Woods Hole Oceanographic Institution 

1 

Denitrification Rates in Marsh Soils and Hydrologic 

and Water-Quality Data for Northeast Creek and Bass 

Harbor Marsh watersheds, Mount Desert Island, Maine

by Thomas G. Huntington, Charles W. Culbertson, and John H. Duff

Abstract

Nutrient enrichment from atmospheric deposition, agricultural activities, wildlife, and 

domestic sources is a concern at Acadia National Park because of the potential problem of waterquality degradation and eutrophication in estuaries. Water-quality degradation has been observed 

at the park’s Bass Harbor Marsh estuary but minimal degradation is observed in the Northeast 

Creek estuary. Previous studies at Acadia National Park have estimated nutrient inputs to 

estuaries from atmospheric deposition and surface-water runoff and have identified shallow 

groundwater as an additional potential nutrient source. Previous studies at Acadia National Park 

have assumed that a certain fraction of the nitrogen input was removed through microbial 

denitrification, but rates of denitrification (natural or maximum potential) in marsh soils have not 

been determined. The U.S. Geological Survey, in cooperation with Acadia National Park, 

measured in situ denitrification rates in marsh soils in Northeast Creek and Bass Harbor Marsh 

watersheds during the summer seasons of 2008 and 2009. Denitrification was measured under 

ambient conditions and following inorganic nitrogen and glucose additions. Laboratory 

incubations of marsh soils with and without acetylene were conducted to determine average 

ratios of nitrous oxide to nitrogen produced during denitrification. Surface water and 

groundwater samples were analyzed for nutrients, specific conductance, temperature, and 

dissolved oxygen. Water levels were recorded continuously during the growing season in the 

Fresh Meadow Marsh in the Northeast Creek watershed.

Introduction

Acadia National Park (ANP) was created to protect the natural beauty of the only rocky 

headlands along the U.S. Atlantic coast. Fundamental to this purpose is protection of the 

ecological, scientific, and scenic attributes of Acadia’s coastal ecosystems. Over 10 percent of 

the Park’s land area is classified as wetland (Calhoun and others, 1994). A diverse array of 

wetland types are represented, including forested and scrub-shrub wetlands, freshwater emergent 

marshes, peatlands, and salt marshes (Roman and others, 2001). Acadia’s estuaries provide 

unique fish and wildlife habitats within the Park. The estuaries are valuable as nursery grounds 

for coastal fish species and foraging habitat for wetland and aquatic birds. Estuaries also act as 

“filters” for water that ultimately enters the nearshore zone, thus helping to maintain coastal 

water quality (Herbert, 1999). Tidally exported productivity from salt marsh or wetland plant 

communities can have a profound effect on estuarine productivity, making coastal salt marshes 
2 

and wetlands among the most productive ecosystems in North America. Changes in sediment 

physicochemical conditions related to nutrient enrichment or perturbations due to human activity 

can alter the pattern of plant zonation, whereby highly competitive invasive plant species can 

potentially displace native plant species (Burke and others, 2002). Shoreline development and 

agricultural runoff also threatens the sustainability of salt marsh ecosystems throughout the 

northeastern U.S. (Silliman and Bertness, 2004; Fitch and others, 2009). 

Acadia’s Water Resources Management Plan (Acadia National Park, 2000) identifies 

accelerated rates of freshwater and coastal marine eutrophication as a priority water quality issue 

and one of the Park’s most important resource management challenges. Residential development 

outside the park boundary is increasing at an unprecedented rate. For example, between 1981 

and 2001 the number of homes in the combined drainage basins of Aunt Betseys Creek, French 

Hill Brook, Old Mill Brook, and Stony Book in the Northeast Creek (NEC) watershed increased 

from 83 to 279 (Nielsen 2002a, 2002b). Recent studies show that Acadia’s Bass Harbor Marsh 

(BHM) estuary is already tending towards a eutrophic state (Doering and others, 1995; Kinney 

and Roman, 1998). Evidence from many other Atlantic Coast estuarine systems indicates that 

land clearing and subsequent septic system installation and lawn fertilizer application are likely 

to increase nutrient loads of both groundwater and surface water entering the estuary (Valiela 

and others, 1990, 2000; Howes and others, 1996; Bricker and others, 1999; Cloern, 2001; 

National Research Council, 2000; Valiela and Bowen, 2002; Tobias and others, 2001; Harvey 

and Odum, 1990). 

The chief nutrient of concern for eutrophication within Acadia’s estuaries is nitrogen. 

Nitrogen inputs to the NEC and BHM watersheds and estuaries are derived from several sources 

including atmospheric deposition, septic effluent, tidal inputs, urban/agricultural runoff, and 

wildlife. Net mineralization of soil organic nitrogen can also contribute dissolved nitrogen 

species. Previous studies have quantified inputs from atmospheric deposition and surface water 

for NEC (Nielsen, 2002b), and BHM (Doering and others, 1995), but inputs from groundwater 

and tidal sources have not been determined. Rates of denitrification (natural or maximum 

potential) in marsh soils have not been determined in previous studies at ANP. 

Previous studies at Acadia National Park have assumed that a certain fraction of the 

nitrogen input was removed through microbial denitrification, but rates of denitrification (natural 

or maximum potential) in marsh soils have not been determined. Denitrification is the process 

of microbial conversion of nitrate to nitrogen (N2) and nitrous oxide (N2O) gases. Denitrification 

rate is measured indirectly as the N2O production rate. In the NEC and BHM watershed we 

assume that most denitrification occurs in the marsh soils and estuarine sediments as has been 

observed in other coastal wetlands (Seitzinger and Nixon, 1985; Seitzinger 1988; Day and 

others, 1989). Denitrification rates increase with increasing nitrogen loading (Seitzinger and 

others, 1984; Seitzinger and Kroeze, 1998; Wigand and others, 2004) and have been shown to be 

seasonally and diurnally dependent, governed principally by temperature, supply of nitrate, and 

availability of organic carbon (Smith and others, 1985; Jorgensen and Sorenson, 1988). 

Denitrification rates are highly variable and must be measured in situ to determine ambient rates 

and maximum potential rates.
3 

Description of Study Area

Northeast Creek (NEC) is a small microtidal estuary in the northeastern part of Mount 

Desert Island, Maine, that is experiencing increasing residential development in the watershed 

(fig. 1). The NEC watershed [26.02 square kilometers (km2)] is composed of several subwatersheds (fig. 2). A partial flow restriction near the estuary outlet results in tidal inundation of 

most wetland sediments only during high spring tides. Recent studies have suggested that deep 

groundwater in the underlying fractured bedrock is not a significant term in the freshwater 

budget for NEC (Nielsen, 2002a). Shallow groundwater discharge is a potential mechanism for 

the delivery of dissolved nitrogen species from domestic septic systems and fertilizer to estuaries 

that can result in eutrophication (Valiela and others, 1990; Reay and others, 1992; Portnoy and 

others, 1998; Valiela and Bowen, 2002). Shallow groundwater can enter the wetland, and 

eventually the estuary, through seepage zones at the upland/wetland boundary or it can enter the 

estuary directly through the creek bottom/banks (Howes and others, 1996; Portnoy and others, 

1998). Culbertson and others (2007) identified groundwater seeps in NEC and BHM using aerial 

thermal imagery and continuous and discrete measurements of temperature and specific 

conductance in selected seeps, but their relative contribution to total inputs is not known. 

The BHM watershed (21.78 km2

), in the southwestern part of Mount Desert Island, is 

composed of several sub-watersheds (fig. 3), the largest of which, Marshall Brook, lies outside 

Park boundaries and is substantially impacted by residential and commercial development, and a 

former landfill. Signs of eutrophication in BHM estuary, the largest tidal marsh estuary on the 

island, were documented more than a decade ago (Doering and others, 1995). Evidence of a 

continued eutrophication response to nutrient inputs from within the watershed and from oceanic 

sources to BHM is noted in more recent studies (Kinney and Roman, 1998; Farris and Oviatt, 

1999). A 1999–2000 study (Nielsen and others, 2002) corroborated the findings of Doering and 

others (1995) that freshwater dissolved inorganic nitrogen (DIN) comprises a significant portion 

of the total nitrogen load to BHM from Marshall and Heath Brooks; organic N comprised most 

of the total N pool in the other tributaries feeding BHM estuary (Doering and others, 1995). 
4 

Figure 1. Shaded relief site map showing locations of the study areas that include Northeast Creek (NEC) 

and Bass Harbor Marsh (BHM) watersheds on Mount Desert Island, Maine.
5 

Figure 2. Map showing the Northeast Creek watershed area, Mount Desert Island, Maine, and surface-water 

sampling locations on the major tributaries and the location of a groundwater monitoring well.
6 

Figure 3. Map showing the location of Bass Harbor Marsh watershed, Mount Desert Island, Maine, and 

surface-water sampling locations on the major tributaries.
7 

Purpose and Scope

This report provides the data collected by the U.S. Geological Survey(USGS), in 

cooperation with Acadia National Park, as part of a study to measure in situ denitrification rates 

in marsh soils in Northeast Creek and Bass Harbor Marsh watersheds during the summer seasons 

of 2008 and 2009. Field and laboratory data are also provided that report rates of N2O release 

from soils following additions of various rates of nitrate and with and without the addition of 

glucose. Laboratory incubation data are also provided that report rates of N2O production with 

and without the acetylene block that will permit calculation of the ratio of N2 to N2O produced 

during denitrification in future studies. These ratios are not presented in this open file report. 

Surface water and groundwater nutrient chemical analysis data and specific conductance, 

temperature, and dissolved oxygen data are also included in this report. This report also provides 

groundwater level data that was recorded continuously during the growing season in Fresh 

Meadow Marsh in the Northeast Creek Watershed. 

Data Collection Methods

Data collection methods are subdivided into three sections. The first section describes the 

methods used for measuring rates of N2O release in the field and the laboratory, and it describes 

the laboratory determination of N2O by gas chromatography. The second section describes the 

methods used for water quality sampling, filtration, and laboratory methods of chemical analysis. 

The final section describes the methods used for recording water level continuously. 

Denitrification Methods

Methods for measuring rates of denitrification are described in two sections. The first 

section describes the static chamber technique used for in situ (field) measurement, the 

measurement schedule, and the treatments that were applied to assess maximum denitrification 

rate (denitrification potential). The second section describes the laboratory incubation 

experiment, the laboratory determinations of N2O gas, and the calculation of denitrification rates.

Field Denitrification Methods

In situ denitrification measurements were made using a static chamber technique where 

denitrification was estimated from the increase in N2O concentration in the headspace of a 

chamber placed on the marsh sediment surface (Oremland and others, 1984; Meding and others, 

2001, Groffman and others, 2006). The flux chambers were constructed from plastic buckets that 

were 28.4 centimeters (cm) in internal diameter. The chambers were equipped with batteryoperated fans to mix the gas in the headspace and rubber septa for gas sampling. The chambers 

were installed by pressing them into the marsh soil surface to a depth of 10 cm to ensure an 

effective seal so that the N2O released from the soil into the headspace could not escape to the 

ambient air or be mixed with ambient air. The volume of the headspace was recorded during 

each incubation. Gas samples for N2O analysis were collected from the chambers through rubber 

septa via glass syringes at various time intervals following installation and sealing of the 

chambers. Gas samples were stored in glass vacutainers (vials) sealed with Teflon lined butyl 

rubber stoppers. The static chamber technique for measuring soil respiration has been described 

in Huntington and others (1998). 
8 

Denitrification measurements (N2O release rate) were made in marsh soils in NEC and 

BHM (figs. 4 and 5) on the dates shown in table 1 that lists the locations of the flux 

measurements, whether they were made under ambient conditions or with the additions of 

nitrate, glucose, or both. Table 1 also reports the rates of nitrate additions where nitrate 

treatments were applied. Nitrate, glucose, and acetylene treatment additions were performed after 

the chambers had been installed into the soil to ensure that the additions were confined to the soil 

below the 28.4-cm diameter chambers to a depth of 10 cm. Treatment solutions of sodium nitrate 

(NaNO3, minimum purity 99 percent, EMD Chemicals, USA) and glucose (minimum 

99 percent purity, ACROS Organics, USA) were made from concentrated solutions prepared in 

the laboratory and diluted in the field with shallow groundwater that was pumped from 

piezometers at the denitrification measurement sites. One liter (L) of a solution of the indicated 

nitrate or nitrate plus glucose concentration was added through an opening in the top of the 

chamber using a funnel. These 1-L additions of 0, 5, 15, 25, 50, 75, 100, 250, 500, and 

1,000 millimolar nitrate (mM NO3) were equivalent to nitrogen addition rates of 0, 1.1, 3.3, 5.5, 

11, 17, 22, 55, 110, and 221 grams nitrogen per square meter (g N m-2) respectively. The ‘0’ 

nitrate with no glucose addition treatments are referred to as ‘ambient’ in this report. For 

treatments where glucose was added, the 1-L treatment solutions contained 75 millimolar (mM) 

glucose, equivalent to 213 grams per square meter (g m-2) when applied to each chamber. After 

the treatment addition, a rubber septum was installed in the top of each chamber to ensure a gas 

tight seal and the fans were started to mix the chamber headspace gases. Acetylene (technical 

grade, 99.5 percent purity, obtained from Maine Oxy) was bubbled gently into one liter solutions 

for 2 minutes to saturate the solutions prior to putting these saturated solutions into the chambers. 

In most cases, two or three replicate chambers were used to determine average N2O 

production rates at a given location and for a given treatment. For all experimental trials with 

three or more replicate samples, we calculated the standard error (SE) as a percent of the mean 

value (SE percent) recorded in the data tables. For the first time period (T0 to T1), 90 percent of 

the ambient treatments had SE percent in the range of 2 to 120 percent and the median 

SE percent was 11 percent. For the first time period (T0 to T1), 90 percent of the nitrate addition 

treatments had SE percent in the range of 0.2 to 45 percent and the median SE percent was 

9.5 percent. Median SE percent increased during the second time period (T1 to T2) to 52 percent 

in the ambient trials and 16 percent in the nitrate addition trials. There were six denitrification 

measurement sites in NEC (fig. 4) and six in BHM (fig. 5) where repeated measurements were 

made over the growing seasons in 2008 and 2009. Flooding during both years limited the 

number of denitrification measurements that could be made in NEC. Denitrification 

measurements could not be made with the chamber that we used when there was more than 5 cm 

of standing water over the marsh soil surface.
9 

Figure 4. Aerial photograph showing the locations of denitrification measurement sites and groundwater 

sampling wells in Northeast Creek watershed, Mount Desert Island, Maine.
10

Figure 5. Aerial photograph showing the locations of denitrification measurement sites and groundwater 

sampling wells in Bass Harbor Marsh watershed, Mount Desert Island, Maine.
11

Table 1. Dates of field (in situ) denitrification rate measurements, locations of measurements and treatments 

applied on those measurement dates.

[NEC, Northeast Creek watershed, GW, groundwater, locations 1 through 6 in each watershed, BHM, Bass Harbor 

Marsh; g n m-2, grams nitrogen per square meter; g glucose m-2, grams glucose per square meter]

Date Start Location Ambient1 Nitrate2 

(g N m-2) Acetylene Glucose2

 

(g glucose m-2) 

Nitrate +

glucose

May 2008 5/29 NEC GW1-2 X

5/30 NEC GW3-4 X

June 2008 6/10 NEC GW1-3 X X

6/11 NEC GW4,6 X

6/12 NEC GW5 X

July 2008 7/15 NEC GW1, GW3 X

7/16 NEC GW1, GW3 1.1

7/16 NEC GW4 X

7/17 NEC GW2 FC1-3 X

7/17 NEC GW2 FC4-6 X 1.1

7/17 NEC GW4 FC4-6 1.1

7/17 NEC GW5 FC1-3 X

7/17 NEC GW5 FC5-6 1.1

7/18 NEC GW6 FC1-4 X

7/18 NEC GW6 FC5-6 1.1

Aug. 2008 8/13 NEC GW4, GW2 X 1.1 X

8/14 NEC GW4 11 X

Sept 2008 9/10 BHM GW3 X

9/11 BHM GW3 5.5,11,16.5,22

9/12 BHM GW3,GW2 5.5 X

9/15 BHM GW1 5.5 X

9/15 BHM GW4 5.5 X

9/16 BHM GW5 3.3

Oct 2008 10/30 BHM GW6 22,55,110,

220

X

June 2009 6/15 BHM GW1-6 X

6/16 BHM GW3 1.1,5.5,11,22,

55,110

213 X

6/17 BHM GW2 1.1,5.5,11,22,

55,110

213 X
12

Table 1. Dates of field (in situ) denitrification rate measurements, locations of measurements and treatments 

applied on those measurement dates.—Continued

[NEC, Northeast Creek watershed, GW, groundwater, locations 1 through 6 in each watershed, BHM, Bass Harbor 

Marsh; g n m-2, grams nitrogen per square meter; g glucose m-2, grams glucose per square meter]

Date Start Location Ambient1 Nitrate2 

(g N m-2) Acetylene Glucose2

 

(g glucose m-2) 

Nitrate +

glucose

July 2009 7/13 BHM GW1-6 X

7/14 BHM GW4 1.1,5.5,11,22,

55,110

213 X

7/15 BHM GW6 1.1,5.5,11,22,

55,110

213 X

7/16 NEC GW1-6 X 213

Aug 2009 8/25 BHM GW1-6 X 213

8/26 BHM GW5 X 1.1,5.5,11,22

44

213 X

Sep 2009 9/21 NEC GW1-6 X 213

9/22 NEC GW4 X 1.1,5.5,11,22

44

213 X

1 An “X” indicates that measurements on this date included ambient (no treatment additions) denitrification rate 

measurements, or, in the case of acetylene an “X” indicates that measurements on this date included the addition of 

acetylene.

2 Nitrate and glucose were added in one liter of solution through the top of the chamber after the chambers were 

installed in the marsh surface immediately prior to sealing the chamber and beginning the incubation. 
13

Laboratory Denitrification Methods

During denitrification both N2O and N2 are produced, but N2 is usually the primary byproduct (Meding et al., 2001). The N2-to-N2O ratio can be quite variable and dependent upon 

landscape position (Meding et al., 2001), sediment properties, and availability of nitrate (for 

example Weier, 1993; Parton and others, 1996). To ensure that future studies could determine 

the site-specific N2-to-N2O ratios, we conducted classical laboratory acetylene inhibition 

laboratory denitrification assays (Oremland and others, 1984; Duff and others, 1996) on 

sediment samples collected from each field measurement location. Marsh soil was collected 

using a hand auger to a depth of 1.25 meter and the material was stored at 4°C until use. Large 

root fragments were removed and the soil material was ground up using a hand-operated meat 

grinder to facilitate mixing and to allow subsequent subsampling of representative fractions of 

the original sample. Soils were stored chilled at 4°C until use. Approximately 20 grams (g) of 

fresh soil was weighed into each 133.88 cubic centimeters (cm3) glass flask. Groundwater was 

also obtained at NEC and BHM using a peristaltic pump to pump from shallow piezometers for 

use in the lab incubations. This groundwater was stored chilled at 4°C until use. The 

groundwater from each location was used to make up solutions containing various concentrations 

of nitrate on the same day that these treatments were used in the laboratory incubation 

experiments. Twenty milliliter (mL) of each appropriate solution were added to each flask and 

then the flasks were sealed. These 20-mL nitrate additions of 5, 10, 50, 75, 100, 200, 500, and 

1,000 mM NO3 were equivalent to nitrogen addition rates of 0.01, 0.14, 0.7, 1.05, 1.4, 3.5, 7, 

14 milligrams of nitrogen per gram (mg N g-1) fresh soil in the incubation flask (respectively). 

Deionized water was used to make a stock solution of 1 molar (M) glucose. Syringes were used 

to add 0.4 mL of the 1 M glucose solution to each flask that received the glucose treatment for a 

final glucose concentration of 19.6 mM glucose in the solution added to the fresh marsh soil 

(equivalent to 3.6 milligrams of glucose per gram (mg glucose g-1) fresh soil fresh soil in the 

incubation flask). The screw cap flask lids were Teflon lined and were equipped with a rubber 

septum to provide for syringe additions of glucose, sparging ultra high purity (UHP) He, addition 

of acetylene, or sampling using syringes. 

After sealing, all flasks were sparged with UHP for 20 minutes. For those treatments that 

received acetylene, 14 cm3 of UHP He was first removed by syringe and then 14 cm3 of 

acetylene was added to achieve a final headspace concentration of approximately 15 percent 

acetylene (technical grade, 99.5 percent purity, obtained from Maine Oxy) by volume. All flasks 

were then placed on a mechanical shaker and shaken for 5 minutes. After this initial equilibration 

4- cm3 gas samples were withdrawn from each flask and transferred to 3-mL vacutainers to 

determine N2O concentration at time zero (T0). Two gas bags were prepared; one contained 

UHP He and the other contained 15 percent acetylene (by volume) in UHP He. Four cm3 of 

“make up” gas from the appropriate bag were added to all flasks after collecting T0 samples to 

restore the original headspace gas volume and pressure. After the addition of the appropriate 

“make up” gas for each treatment, the flasks were shaken continuously until the subsequent gas 

sample collections (e.g. T1, T2, T3, or T4). During each subsequent sampling, 4 cm3 of gas was 

collected from the headspace by syringe and transferred to a 3-mL vacutainer and 4 mL of make 

up gas was added back to each flask. The gas samples in vacutainers (glass vials) sealed with 

Teflon lined butyl rubber stoppers were shipped by Federal Express in cardboard containers to a 

USGS laboratory specializing in gas analysis (Menlo Park, Calif.) for determination of 

N2O concentration. 
14

In most laboratory assays where nitrate was added, two, and sometimes three replicate 

chambers were used to determine average N20 production rates in laboratory assays. None of the 

laboratory assays included three replicate samples under ambient conditions. For all laboratory 

assays with three replicate samples, we calculated the standard error as a percent of the mean 

value (SE percent) recorded in the data tables. For the first time period (T0 to T1) 90 percent of 

the nitrate addition treatments had SE percent in the range of 2 to 97 percent and the median SE 

percent was 17 percent. Median SE percent decreased during the second time period (T1 to T2) to 

12 percent in these nitrate addition trials. 

Subsamples of fresh, ground, and homogenized marsh soils were analyzed at the Maine 

Water Science Center Laboratory to determine the water fraction and the organic matter (OM) 

fraction. Fresh soil samples were weighed and then oven dried at 105°C until they reached a 

constant weight and then they were reweighed to gravimetrically determine the water fraction 

(g water per g oven dry soil). These samples were then ashed in a muffle furnace at 500°C until 

they reached a constant weight and then they were reweighed to gravimetrically determine the 

OM content (g OM per g oven dry soil) (Davies, 1974). 

Laboratory Methods and Computation of N2O Release Rate

Gas samples were analyzed by Ni-63 electron capture detection gas chromatography 

(GC-ECD) using standard methods (Oremland and others, 1984; Duff, 1996, Aelion and Shaw, 

2000). The laboratory reported the N2O concentrations from the vacutainers (headspace at each 

sampling time) in units of nanomoles per milliliter (nmoles mL-1) N2O. The gas chromatograph 

is typically calibrated with Scott mini mix calibration standards twice during each run with a 

minimum of five concentrations that bracket the samples. Single point calibrations 

(0.0446 nmoles N20 per injection) are run approximately every 20 samples. The standard curves 

typically span from 0.00446 to 2.23 nmoles N2O per injection depending on the sample range 

with r2

s >0.99. The relative standard deviation of repeated standards typically averages 

3.8 percent with a standard deviation of +/-1.9 percent over the standard range. Laboratory air 

analyses are also run (0.00156 nmoles N2O). The relative standard deviation is significantly 

higher (25.6 precent) since laboratory air concentrations can not be controlled. During all 

laboratory sample analysis runs, N2O-free calibration gas is substituted for N2O standards or 

environmental samples to determine a baseline reading.

In situ N2O release rates were determined as nmoles m-2 hr-1 using the chamber 

headspace volume, the initial and subsequent headspace concentrations, the area of the marsh 

surface covered by the chamber (m-2), and the duration of the incubation (hour (hr)):

N2O release rate = [ [T2Conc. * vol - T1Conc. * vol ] * area-1] * incubation time-1 (1)

The N2O release rates reported herein did not account for the N2O produced during the 

lab incubations that was retained in the liquid phase. About 17 percent of the total N2O 

produced was retained in the liquid phase based on a simple Henry’s Law calculation for 

incubations conducted at 25oC using a Bunsen absorption coefficient of 0.544 as described in 

Tiedge (1983). 

Water Quality Data Collection Methods

Water quality samples were collected from tributaries that flow into the NEC and BHM 

estuaries (figs. 2 and 3; table 2) on an approximately monthly schedule during the 2008 and 2009 

growing seasons. These samples were analyzed for the constituents shown in table 3. Samples 
15

were collected from shallow groundwater in NEC and BHM marsh soils at the locations where in 

situ denitrification measurements were made during the 2008 and 2009 growing seasons 

(figs. 4 and 5). Various quality-assurance samples were also collected to determine bias and 

precision associated with the sample data.

Field Methods

For the surface water quality sampling sites on the NEC and BHM tributaries, a 

peristaltic pump was used to pump water directly from a fast flowing section of the tributary and 

through a cartridge filter (0.45 micron polyethersulfone membrane filter) and into 125-mL amber 

polyethylene bottles at each measuring station. For the groundwater samples, all samples were 

collected by pumping water from shallow piezometers that were installed in the NEC and BHM 

marsh soils at the locations where denitrification measurements were made. The piezometers 

were commercial PVC, 5-cm diameter, 1-meter length, and screened over their entire length, 

except within 15-cm of each end. The screen size was 0.0254 cm and the bottoms of the 

piezometers were equipped with a conical 5-cm well point. For installation, holes were augured 

with a hand auger using a 3.75-cm diameter bit to ensure a tight seal between the piezometers 

and the surrounding marsh soil. Piezometers were installed with 10-cm of unscreened pipe above 

ground surface. Caps were installed to prevent precipitation from entering the piezometers. 

Samples were pumped with a peristaltic pump or a vacuum pump into 250-mL polyethylene 

bottles. All polyethylene bottles were prewashed and rinsed profusely prior to sample collection. 

Groundwater samples were filtered on site with syringe cartridge filters, or were filtered in a 

laboratory within 6 hours using a peristaltic pump and a cartridge filter or a syringe and a 

cartridge filter. All filters were 0.45 micron pore size and all samples were filtered into 125-mL 

amber polyethylene bottles and chilled immediately.

All amber polyethylene sample bottles were acid washed and rinsed 3 times with filtered 

sample prior to sample collection. Samples were chilled on ice from the time of collection until 

received at the Maine Water Science Center in Augusta, Maine within 36 to 48 hours of 

collection. All samples were frozen in Augusta and stored frozen before overnight shipment to 

the Woods Hole Oceanographic Institute, Nutrient Analytical Facility (WHOI) for analysis. All 

samples were maintained frozen at WHOI until they were analyzed. All samples were analyzed 

for all constituents at the WHOI Nutrient Analytical Facility.

Field values for specific conductance and water temperature were measured at the time of 

sample collection using an Orion Model 122 meter or a YSI EC 300 meter (Wilde, variously 

dated). The meter was calibrated prior to each use using standard solutions for specific 

conductance. For surface water measurements the Orion or YSI probe was placed directly in a 

fast flowing section of the stream and allowed to equilibrate before recording the value. For 

groundwater measurements samples were pumped into a beaker (if the peristaltic pump was 

used) and the Orion or EC 300 probe was placed into the beaker while water was pumped in and 

allowed to equilibrate before readings were recorded. If the hand vacuum pump were used, water 

was first pumped into a vacuum flask and was then poured into a beaker and the Orion or YSI 

probe was placed into the beaker and readings were recorded within 30 seconds. Dissolved 

oxygen was determined using a Chemetrics field test kit employing the indigo carmine 

colorimetric method (ASTM D 887–87) (Gilbert and others, 1982). 
16

Table 2. Surface water and groundwater water-quality monitoring stations in Northeast Creek and Bass Harbor Marsh watersheds 

sampled in 2008 and 2009.

[mi2

, square miles; Dr., Drive, nr, near; ME, Maine, latitude and longitude determined with Global Positioning System (GPS) (+/- 2 to 5 meters); NEC, 

Northeast Creek watershed, GW, groundwater, locations 1 through 6 in each watershed, BHM (Bass Harbor Marsh watershed) Str., stream; Br. Brook]

Station Name

USGS

station

number

Latitude Longitude Drainage 

area in mi2

Number of 

samples

Surface water sites

Aunt Betsey’s Brook near Bar Harbor, ME 01022815 44 24 22.0 68 19 10.0 0.63 11

French Hill Brook near Bar Harbor, ME 01022817 44 24 23.0 68 18 44.0 0.56 11

Old Mill Brook at Old Norway Dr. nr Bar Harbor, ME 01022800 44 23 55.0 68 17 14.0 1.55 12

Marshall Brook near Southwest Harbor, ME 01022890 44 16 29.0 68 21 05.0 1.97 7 

Lurvey Brook near Southwest Harbor, ME 01022892 44 16 44.0 68 21 28.0 0.105 6 

Heath Brook near Tremont, ME 01022895 44 16 40.0 68 22 05.0 0.91 8 

Buttermilk Brook near Southwest Harbor, ME N/A 44 15 59.6 68 20 38.5 1 

Stony Brook below Hamilton Pond nr Bar Harbor, ME 01022810 44 25 28.0 68 17 29.0 2.66 1 

Adams Brook near Southwest Harbor, ME N/A 44 14 52.3 68 20 22.4 1 

Adams Brook near Southwest Harbor, ME N/A 44 14 35.3 68 19 53.0 1 

Adams Brook near Southwest Harbor, ME N/A 44 14 37.4 68 19 58.3 1 

Adams Brook near Southwest Harbor, ME N/A 44 14 45.0 68 20 09.2 1 

Aunt Betsey’s Estuary nr Bar Harbor, ME N/A 44 24 38.7 68 18 45.8 N/A 1 

Bass Harbor Marsh Estuary, nr SW Harbor, ME2 N/A 44 15 35.2 68 20 42.4 N/A 1 

Bass Harbor Marsh Estuary, nr SW Harbor, ME3 N/A 44 15 35.2 68 20 42.4 N/A 1 

Intermittent Str. draining to Aunt Betsey’s Br. nr GW2 N/A 44 24 40.0 68 18 32.0 N/A 1 
17

Groundwater sites1

NEC GW1 442439068184501 44 24 39.1 68 18 45.2 N/A 9

NEC GW2 442437068184201 44 24 36.5 68 18 42.6 N/A 7

NEC GW3 442439068184601 44 24 38.7 68 18 45.8 N/A 8

NEC GW4 442437068184401 44 24 37.0 68 18 44.3 N/A 8

Groundwater sites1—Continued

NEC GW5 442504068184401 44 25 03.6 68 18 44.7 N/A 7 

NEC GW6 442439068184602 44 24 39.2 68 18 46.2 N/A 3 

BHM GW1 441533068202701 44 15 32.5 68 20 27.7 N/A 4 

BHM GW2 441532068202901 44 15 32.0 68 20 29.8 N/A 4 

BHM GW3 441532068203101 44 15 32.1 68 20 31.1 N/A 7 

BHM GW4 441532068203301 44 15 32.1 68 20 33.2 N/A 4 

BHM GW5 441538068204701 44 15 37.9 68 20 47.4 N/A 4 

BHM GW6 441535068204201 44 15 35.2 68 20 42.4 N/A 6 

1

For groundwater sites the numbers of samples collected refers only to the pre-treatment (ambient) samples collected

2

sampled collected at water surface

3

sample collected 20 cm below water surface
18

Table 3. Constituent name, U.S. Geological Survey National Water Information System parameter codes and 

method codes, and minimum reporting limits for temperature, specific conductance, dissolved oxygen, and nutrients.

[°C, degrees Celsius;, na, not applicable; μs/cm, micro Siemens per centimeter; mg/L, milligrams per liter; 

N, nitrogen; SiO2, silica, dissolsved; P, phosphorus;] 

Parameter code1 Constituent name Method code

Minimum or 

laboratory 

reporting limit

00010 Temperature, water (°C) na 0.01 °C

00094 Specific conductance, field (μs/cm at 25 °C) na 1.0 μs/cm

00300 Oxygen, dissolved (mg/L) na 0.1 mg/L

00608 Ammonium (mg/L as N) 00623 0.01 mg/L

00955 Silica, dissolved (mg/L as SiO2) CL151 0.5 mg/L

00671 Phosphorus, orthophosphate, dissolved (mg/L as P) 00048 0.006 mg/L

00618 Nitrate + Nitrite (mg/L as aN) CL050 0.10 mg/L

62854 Nitrogen, total, dissolved (mg/L and N) CL063 0.01 mg/L

1

For groundwater sites the numbers of samples collected refers only to the pre-treatment (ambient) samples 

collected.

2

Sampled collected at water surface.

3

Sample collected 20 cm below water surface.
19

Laboratory Methods

Ammonia was analyzed colorimetrically by the indophenol method Lachat QuickChem 

Method 31-107-060-1-B (Scheiner, 1976; method 4500-NH3-F, p. 4.108-4.109, American Public 

Health Association, 1998). Silicate was analyzed colorimetrically by using Lachat QuickChem 

Method 31-114-27-1-C (method 4500-SiO2-F, p.4-160 and 4-161, American Public Health 

Association, 1998). Phosphate was analyzed colorimetrically by Lachat QuickChem Method 

31-115-01-1-H (Murphy and Riley, 1962; method 4500-P-E, p.4–146 and 4–147, American 

Public Health Association, 1998). Nitrate plus nitrite were analyzed colorimetrically by using 

Lachat QuickChem Method 31-107-04-1-E (Wood and others 1967; method 4500-NO3-F, 

p. 4–118 and 4–119, American Public Health Association, 1998). The concentration of nitrite in 

these waters is expected to be below detection and therefore throughout the report this analysis is 

referred to as nitrate. Total dissolved nitrogen was analyzed colorimetrically following alkaline 

persulfate digestion (D’Elia and others, 1977; method 4500-Norg, p.4–102 and 4–103, American 

Public Health Association, 1998). 

The reliability of the chemical data was ensured by the preparation and analysis of 

several types of quality-control samples. These quality-control samples include 1 field blank and 

10 replicate samples under ambient conditions and, and 3 replicate samples of groundwater 

following a nitrate addition treatment. Four replicate samples were also sent to the USGS 

National Water Quality Laboratory (NWQL), in Denver, Colorado, where they were analyzed 

for nitrate plus nitrate, ammonia, and total dissolved nitrogen. Additionally, reasonable range 

checks were performed by comparing laboratory-determined chemical data for the sampled 

tributaries with results reported for these tributaries that were sampled during previous studies. 

A field blank is used to test for positive bias that can result from contamination during 

any stage of sample collection, processing, or analysis. A field blank was prepared from 

deionized water that was produced by a laboratory-grade water purification system that uses a 

sequence of ion exchange resin columns. The resulting purified water has a specific conductance

between 1 and 3 microsiemens. One field blank was collected during sampling at base-flow 

conditions in October 2008. During collection of base-flow samples, the field blank was 

collected by transferring blank water from the holding bottle to the sample-collection bottle. It 

was then processed, stored, and shipped to the analytical laboratory in a manner consistent with 

the collection of other environmental samples. 

Replicate samples are samples thought to be identical in composition to the 

environmental samples. Replicate samples provide a measure of bias and variability for the 

method of sample collection, sample processing (splitting, filtering, and preservation), and 

laboratory analysis. Replicate samples were collected from tributaries (Aunt Betseys Brook, 

Marshall Brook, French Hill Brook, Old Mill Brook, Heath Brook and Adams Brook) during 

base-flow conditions in September 2008, October 2008, and August 2009. One replicate sample 

was collected from a groundwater sampling location (NEC GW2) in Fresh Meadow Marsh, 

Northeast Creek in August 2008. Three replicate samples were collected from a groundwater 

sampling location (NEC GW4) in Fresh Meadow Marsh, Northeast Creek in August 2008 

following the addition of 1 liter of a 50 mM nitrate solution to the marsh surface.

In addition to the quality-assurance samples collected during this project, WHOI and 

NWQL routinely analyze various quality-control samples, including laboratory reagent blanks, 

interference check solutions, laboratory control samples, standard reference materials, laboratory 

reagent spike samples, and laboratory duplicate samples (Furlong and others, 2001; Garbarino 

and others, 2006; written communication, Paul Henderson, WHOI, 2011). Both laboratories also 
20

participate in participate in laboratory-intercalibration (blind sample) programs including 

GEOTRACES [http://www.obs-vlfr.fr/GEOTRACES/index.php/science/intercalibration] to test 

and track method performance. The WHOI laboratory uses the QUASIMEME project (Quality 

Assurance of Information for Marine Environmental Monitoring in Europe) standardization 

protocols (Cofino and Wells, 1994). 

Ancillary Data Collection Methods

Water level was recorded continuously at one site (USGS Station ID No. 

442439068184502) in Fresh Meadow Marsh, Northeast Creek (fig. 2) using standard U.S. 

Geological Survey methods. The well was equipped with a Design Analysis DH21 submersible 

pressure transducer programmed to record water level and water temperature every 

fifteen minutes.

Soil temperature was recorded periodically at each of the principle denitrification 

measurement sites. Soil temperature was determined at a soil depth of 15 cm below the surface 

with a glass thermometer.

Daily precipitation data were obtained from the National Park Service (NPS at Acadia 

National Park). The NPS maintains a National Atmospheric Deposition Program (NADP) 

monitoring site that uses a Belfort tipping bucket rain gage to record daily precipitation data at 

McFarland Hill on Mount Desert Island. The NADP program is administered by the Illinois 

State Water Survey, Champaign, Ill. Data can be accessed from the URL 

http://nadp.sws.uiuc.edu/. 

Tidal data were obtained from National Oceanographic and Atmospheric Administration 

(NOAA) data files [http://tidesandcurrents.noaa.gov] accessed on 12/20/2009 for Frenchman’s 

Bay, Bar Harbor, Maine on Mount Desert Island. Stream-water discharge records were obtained 

for Otter Creek near Bar Harbor, Maine (USGS station ID 01022840). Streamwater data was 

obtained from the USGS National Water Information System (NWIS) 

[http://waterdata.usgs.gov/me/nwis/uv/?site_no=01022840] accessed on 4/1/2010. 

Field Denitrification Data

In these descriptions of data obtained in field experiments nitrate additions of 1-L of 0, 5, 

15, 25, 50, 75, 100, 250, 500, and 1,000 mM NO3 were equivalent to nitrogen addition rates of 

0, 1.1, 3.3, 5.5, 11, 17, 22, 55, 110, and 221 N m-2 respectively. Field denitrification 

measurements under ambient conditions (no treatments) were conducted in NEC watershed 

during June 10 through June 12, 2008 at six locations, and at two of those locations (GW1 and 

GW2) acetylene was added as a treatment (table 4). Field denitrification measurements were 

conducted in NEC watershed during July 15 through July 19, 2008 at six locations and included 

ambient and 5 mM nitrate additions to all locations (table 5). Field denitrification measurements 

were conducted in NEC watershed during August 13 through August 15, 2008 at two locations 

(GW2 and GW4) and included ambient, and 5, and 50 mM nitrate additions to both locations 

(table 6). Field denitrification measurements were conducted in BHM watershed during 

September 13 through September 16, 2008 at six locations. During these September 2008 

measurements, ambient, and 25, 50, 75, and 100 mM nitrate additions were conducted on GW3 

(table 7). In addition, during these September 2008 measurements, sites GW1, GW2, GW3 and 

GW4 received 25 mM nitrate treatments and 25 mM nitrate plus acetylene treatments, and GW5 

and GW6 received 15 mM nitrate treatments (table 7). Field denitrification measurements were 
21

conducted in BHM watershed during October 30 through October 31, 2008 at one location 

(GW6). During this October 2008 measurement, treatments included 100, 250, 500, and 

1000 mM nitrate with and without acetylene (table 8).

Field denitrification measurements under ambient conditions were conducted in BHM 

watershed during June 15 through June 16, 2009 at six locations (table 9). Field denitrification 

measurements were conducted in BHM watershed during June 16 through June 17, 2009 at the 

GW3 location that included 5, 25, 50, 100, 250, and 500 mM nitrate with and without 75 mM 

glucose and one treatment with 75 mM glucose only (table 9). Field denitrification 

measurements were conducted in BHM watershed during June 17 through June 18, 2009 at the 

GW2 location that included 5, 25, 50, 250, and 500 mM nitrate with and without 75 mM glucose 

and one treatment with 75 mM glucose only (table 9). Field denitrification measurements under 

ambient conditions were conducted in BHM watershed on July 13, 2009 at six locations 

(table 10). Field denitrification measurements were conducted in BHM watershed during July 

14 through July 16, 2009 at the GW4 and GW6 locations that included 5, 25, 50, 100, 250, and 

500 mM nitrate with and without 75 mM glucose (table 10). Field denitrification measurements 

under ambient conditions and with 75 mM glucose additions were conducted in NEC watershed 

on July 16, 2009 at six locations (table 10). Field denitrification measurements under ambient 

conditions and with 75 mM glucose additions were conducted in BHM watershed during August 

25 through August 26, 2009 at six locations (table 11). Field denitrification measurements were 

conducted in BHM watershed during August 26 through August 27, 2009 at the GW5 location 

that included ambient conditions; 75 mM glucose; and 5, 25, 50, 100, and 200 mM nitrate with 

and without 75 mM glucose (table 11). Field denitrification measurements under ambient 

conditions and with 75 mM glucose additions were conducted in NEC watershed during 

September 21 through September 22, 2009 at six locations (table 12). Field denitrification 

measurements were conducted in NEC watershed during September 22 through September 23, 

2009 at the GW4 location that included ambient; 75 mM glucose; and 5, 25, 50, 100, and 

200 mM nitrate with and without 75 mM glucose (table 12). 

Laboratory Denitrification Data

In these descriptions of data obtained in laboratory experiments nitrate additions of 

20 -mL of 5, 10, 50, 75, 100, 200, 500, and 1,000 mM NO3 were equivalent to nitrogen addition 

rates of 0.01, 0.14, 0.7, 1.05, 1.4, 3.5, 7, and 14 mg N mg N g-1 fresh soil respectively. The 

gravimetric analysis of fresh soil samples to determine the dry weight and organic matter fraction 

for the denitrification measurement sites in NEC and BHM are reported in 

table 13. On May 5, 2009, soils from NEC (GW1 and GW2) and BHM (GW1) were incubated 

with 0 and 50 mM NO3 with and without acetylene and denitrification rates were determined 

(table 14). On May 7, 2009, soils from NEC (GW2, GW3, GW4, and GW5) were incubated with 

50 mM NO3 with and without acetylene and denitrification rates were determined (table 15). On 

May 13, 2009, soils from NEC (GW6) and BHM (GW2, GW3, and GW4) were incubated with 

50 mM NO3 with and without acetylene and denitrification rates were determined (table 16). On 

May 15, 2009, soils from BHM (GW5 and GW6) were incubated with 50 mM NO3 with and 

without acetylene and denitrification rates were determined (table 17). On May 20, 2009, soils 

from NEC (GW4) were incubated with 0, 5, 10, 50, 75, 100, 250, 500, 750, and 1000 mM NO3

with and without acetylene and with and without glucose and denitrification rates were 

determined (table 18). On May 22, 2009, soils from NEC (GW2) and BHM (GW2) were 

incubated with 0, 75, and 500 mM NO3 with and without acetylene and with and without 
22

glucose and denitrification rates were determined (table 19). On May 27, 2009, soils from NEC 

(GW5) and BHM (GW3) were incubated with 0, 75, and 500 mM NO3 with and without 

acetylene and with and without glucose and denitrification rates were determined (table 20). On 

May 29, 2009, soils from NEC (GW1) and BHM (GW5) were incubated with 0, 75, and 500 mM 

NO3 with and without acetylene and with and without glucose and denitrification rates were 

determined (table 21). On June 2, 2009, soils from NEC (GW3) and BHM (GW1) were 

incubated with 0 and 75 mM NO3 with and without acetylene and with and without glucose and 

denitrification rates were determined (table 22).

Water-Quality Data

Water quality samples were collected periodically during May through October in 2008 

and 2009 from the sites listed in table 2. The surface water nutrient water-quality data are 

reported in table 23 and the field measurements of temperature and specific conductance are 

reported in table 24. The groundwater nutrient water-quality data for NEC and BHM are 

reported in tables 25 and 26 and the field measurements of temperature, specific conductance, 

and dissolved oxygen are reported in table 27. 

Quality-Control Samples

Trace amounts of ammonium, nitrate plus nitrite, phosphate, silicate, and total dissolved 

nitrogen were detected in the field blank. The concentration for these constituents measured in 

the field blank was one or more orders of magnitude less that the lowest concentration found in 

all of the environmental samples, and at least one order of magnitude lower than the minimum 

reporting limit for each constituent.

Field replicate samples were analyzed by comparing the relative percent differences 

(RPDs) of the results for each pair of samples where the same analyte was detected in both 

samples. The RPD was calculated as the absolute difference between replicate analyses divided 

by the average of the same two values. The RPDs for total dissolved nitrogen and for phosphate 

varied by 7 percent or less for the 3 replicate samples collected from tributaries that were 

analyzed in the WHOI laboratory. The RPDs for ammonium and silicate varied by between 3 

and 53 percent for the same tributary replicate samples. The RPDs for nitrate plus nitrite for 

these same tributary replicate samples were higher, varying between 29 percent and 141 percent. 

We can not explain these higher RPDs for nitrate plus nitrite in these tributary samples, 

especially since the RPDs for total dissolved nitrogen for these samples varied by only 7 percent 

or less. Previously published analyses for these streams (Nielsen and Kahl, 2007; Culbertson and 

others, 2007) and analyses for these streams conducted for the Northeast Temperate Network 

(NETN) (unpublished data, written communication, Bill Gawley, National Park Service) are 

consistent with the higher of the two replicate analyses for nitrate plus nitrate. These replicate 

nitrate plus nitrite concentrations are relatively low and precision is usually lower for lower 

analyte concentrations.

Four separate samples from tributaries were analyzed in both the WHOI and NWQL 

laboratories for ammonium, nitrate plus nitrite, and total dissolved nitrogen. It was not possible 

to calculate RPDs for ammonium and nitrate because the concentrations of these constituents 

generally were below the minimum reporting limit for the NWQL laboratory. The RPD for total 

dissolved nitrogen varied between 81 and 138 percent and the NWQL-determined concentrations 
23

were always higher than those determined by the WHOI laboratory. The WHOI-determined 

concentrations appear to be more consistent with previously published vales based on the 

calculated ratios of total dissolved nitrogen to nitrate plus nitrite. The NWQL concentrations 

result in ratios of total dissolved nitrogen to nitrate plus nitrite that are unprecedentedly high. 

The RPDs for nitrate plus nitrite determined for the untreated groundwater environmental sample 

and field replicate (NEC GW2) and among the 3 replicate groundwater samples collected at NEC 

GW4 following application of the 50 mM nitrate treatment were less than 1 percent.

Samples of gas collected from the free atmosphere above the marsh and from the 

chamber headspace immediately after placement and sealing on the marsh surface were 

analyzed for N2O and in all cases determined N2O concentrations were at least one order of 

magnitude below the lowest calibration standard indicating only trace amounts of N2O were 

present. In all cases, when N2O-free calibration gas was substituted for N2O standards or 

environmental samples during laboratory gaschromatographic analysis no N2O was detected.

Ancillary Data

Water level (in feet relative to land surface) for Fresh Meadow Marsh in Northeast Creek 

from May 2009 through December 2009 is shown in figure 6A. Discharge (cubic feet per second 

(ft3/s)) at Otter Creek on Mount Desert Island from May 2009 through December 2009 is also 

shown in figure 6A. Daily precipitation (inches) at McFarland Hill Station, near Bar Harbor, 

Mount Desert Island, from May 2009 through December 2009 is shown in figure 6B. Soil 

temperature (oC) recorded at 15 cm soil depth during denitrification measurements is reported in 

tables 4 through 12. 
24

Figure 6. Graphs showing A, Water level in Fresh Meadow Marsh, Northeast Creek watershed, Mount 

Desert Island, Maine and discharge at Otter Creek USGS station ID 01022840 and B, daily precipitation at 

McFarland Hill Station, near Bar Harbor, Mount Desert Island, Maine.
25

References Cited

Acadia National Park, 2000, Water Resources Management Plan, Kahl,S., Manski, D., Flora, M., 

and Houtman, N., eds. NPS D–209, April 2000, 102 p. 

Aelion, C.M., and Shaw, J.N., 2000, Denitrification in South Carolina (USA) Coastal Plain 

Aquatic Sediments: Journal of Environmental Quality, v. 29, p. 1696–1702. 

American Public Health Association, 1998, Standard methods for the examination of water and 

wastewater (20th ed.): Washington, D.C., American Public Health Association. 

Bricker, S., Clement, C.G., Pirhalla, D.E., Orlando, S.P., and Farrow, D.R.G., 1999, National 

estuarine eutrophication assessment of nutrient enrichment in the Nations' estuaries, NOAA, 

Natl. Ocean Serv., Special Projects Office and the Natlional Centers for Coastal Ocean 

Science, Silver Spring, MD, 71pp. 

Burke, D.J., Hamerlynck, E.P., and Hahn, D., 2002, Interactions among plant species and 

microorganisms in salt marsh sediments: Applied and Environmental Microbiology, v. 68, 

no. 3, p. 1,157–1,164. 

Calhoun, A.J.K., Cormier, J.E., Owen, R.B.J., Roman, C.T., and Tiner, R.W.J., 1994, The 

wetlands of Acadia National Park and vicinity: Maine Agriculture and Forest Experiment 

Station Miscellaneous Report Publication 721. University of Maine, Orono. 

Cloern, J.E., 2001, Our evolving conceptual model of the coastal eutrophication problem: 

Marine Ecology Progress Series, v. 210, p. 223–253. 

Cofino, W.P., and Wells, D.E., 1994, Design and evaluation of the QUASIMEME laboratory 

performance studies: a test case for robust statistics: Marine Pollution Bulletin, v. 29, 

p. 149–158. 

Culbertson, C.W., Huntington, T.G., and Caldwell, J.M., 2007, Nutrient enrichment in estuaries 

from discharge of shallow groundwater, Mount Desert Island, Maine: U.S. Geological Survey 

Scientific Investigations Report 2007–5188, 34 p. 

Davies, B.E., 1974, Loss-on-ignition as an estimate of soil organic matter.: Soil Science Society 

of America Proceedings, v. 38, p. 150–151. 

Day, J.W., Jr., Hall, C.A.S., Kemp, W.M., and Yanez-Arancibia, A., 1989, Estuarine Ecology, 

John Wiley and Sons, New York. 558 pp. 

D'Elia, C.F., Stuedler, P.A., and Corwin, N., 1977, Determination of total nitrogen in aqueous 

sampling using persulfate digestion: Limnology and Oceanography, v. 22, p. 760–764. 

Doering, P.H., Roman, C.T., Beatty, L.L., Keller, A.A., and Oviatt, C.A., 1995, Water quality 

and habitat evaluation of Bass Harbor Marsh, Acadia National Park, Maine. Technical Report

NPS/NESORNR/NRTR/95–31. 

Duff, J.H., Pringle, C.M., and Tiska, F.J., 1996, Nitrate reduction in sediments of lowland 

tropical streams draining swamp forest in Costa Rica: An ecosystem perspective: 

Biogeochemistry, v. 33, p. 179–196. 

Farris, C.N., and Oviatt, C.A., 1999, Changes in metabolic rates under fluctuating salinity 

regimes for two subtidal estuarine habitats: Estuaries, v. 22, p. 126–137. 

Fitch, R., Theodose, T., and Dionne, M., 2009, Relationships among Upland Development, 

Nitrogen, and Plant Community Composition in a Maine Salt Marsh: Wetlands, v. 29, no. 4, 

p. 1,179–1,188. 

Furlong, E.T., Anderson, B.D., Werner, S.L., Soliven, P.P., Coffey, L.J., and Burkhardt, M.R., 

2001, Methods of analysis by the U.S. Geological Survey National Water Quality 

Laboratory—Determination of pesticides in water by graphitized carbon-based solid-phase 
26

extraction and high-performance liquid chromatography/mass spectrometry: U.S. Geological 

Survey Water-Resources Investigations Report 01–4134, 73 p. 

Garbarino, J.R., Kanagy, L.K., and Cree, M.E., 2006, Determination of elements in naturalwater, biota, sediment and soil samples using collision/reaction cell inductively coupled 

plasma-mass spectrometry: U.S. Geological Survey Techniques and Methods, book 5, sec. B, 

chap. 1, 88 p. 

Gilbert, T.W., Behymer, T.D., and Castañeda, H.B., 1982, Determination of dissolved oxygen in 

natural and wastewaters: American Laboratory, v. 14, no. 3, p. 119–134. 

Groffman, P.M., Altabet, M.A., Böhlke, J.K., Butterbach-Bahl, K., David, M.B., Firestone, 

M.K., Giblin, A.E., Kana, T.M., Nielsen, L.P., and Voytek, M.A., 2006, Methods for 

measuring denitrification: Diverse approaches to a difficult problem: Ecological Applications, 

v. 16, no. 6, p. 2,091–2,122. 

Harvey, J.W., and Odum, W.E., 1990, The influence of tidal marshes on upland groundwater 

discharge to estuaries: Biogeochemistry, v. 10, p. 217–236. 

Herbert, R.A., 1999, Nitrogen cycling in coastal marine ecosystems: FEMS Microbiology 

Review, v. 23, p. 563–590. 

Howes, B.L., Weiskel, P.K., Goehringer, D.D., and Teal, J.M., 1996, Interception of freshwater 

and nitrogen transport from uplands to coastal waters-The role saltmarshes, in Nordstrom, 

K.F., and Roman, C.T., eds., Estuarine Shores--Evolution, environments and human 

alterations,: New York, Wiley, p. 287–310. 

Huntington, T.G., Harden, J.W., Dabney, S.M., Marion, D.A., Alonso, C., Sharpe, J.M., and 

Fries, T.L., 1998, Soil, environmental, and watershed measurements in support of carbon 

cycling studies in northwestern Mississippi: U.S. Geological Survey, Open File Report 

98–501, 91 p. 

Jorgensen, K.S., and Sorenson, J., 1988, Two annual maxima of nitrate reduction and 

denitrification in estuarine sediments (Norsminde Fjord, Denmark): Marine Ecology Progress 

Series, v. 94, p. 267–274. 

Kinney, E.H., and Roman, C.T., 1998, Response of primary producers to nutrient enrichment in 

a shallow estuary: Marine Ecology Progress Series, v. 163, p. 89–98. 

Koch, M.S., Maltby, E., Oliver, G.A., and Bakker, S.A., 1992, Factors controlling denitrification 

rates of tidal mudflats and fringing salt marshes in south-west England: Estuarine, Coastal and 

Shelf Science, v. 34, no. 5, p. 471–485. 

Meding, S.M., Morris, L.A., Hoover, C.M., Nutter, W.L., and Cabrera, M.L., 2001, 

Denitrification at a long-term forested land treatment system in the Piedmont of Georgia: 

Journal of Environmental Quality, v. 30, p. 1411–1420. 

Murphy, J., and Riley, J.P., 1962, A modified single solution method for determination of 

phosphate in natural waters: Analytica Chemica Acta, v. 27, no. 31–36. 

Nielsen, M.G., 2002a, Water budget for and nitrogen loads to Northeast Creek, Bar Harbor, 

Maine: U.S. Geological Survey Water-Resources Investigations Report 02–4000, 32 p. 

Nielsen, M.G., 2002b, Estimating quantity of water in fractured bedrock units on Mount Desert 

Island, and estimated ground-water use, recharge, and dilution of nitrogen in septic waste in 

the Bar Harbor area, Maine: U.S. Geological Survey Open-File Report 02–435, 45 p. 

Nielsen, M.G., Caldwell, J.M., Culbertson, C.W., and Handley, M., 2002, Hydrologic data 

collected in small watersheds on Mount Desert Island, Maine: 1999–2000. U.S. Geological 

Survey Open-File Report 02–416, 47 p. 
27

Nielsen, M.G., and Kahl, J.S., 2007, Nutrient export from watersheds on Mt. Desert Island, 

Maine, as a function of land use and fire history: Environmental Monitoring and Assessment, 

v. 126, p. 81–96. 

NRC, 2000, Clean Coastal Waters: Understanding and Reducing the Effects of Nutrient 

Pollution. National Academy Press, Washington, DC., 428 pages. 

Oremland, R.S., Umberger, C., Culbertson, C.W., and Smith, R.L., 1984, Denitrification in San 

Francisco Bay intertidal sediments: Applied and Environmental Microbiology, v. 47, no. 5, 

p. 1,106–1,112. 

Parton, W.J., Mosier, A.R., Ojima, D.S., Valentine, D.W., Schimel, D.S., Weier, K., and 

Kulmala, A.E., 1996, Generalized model for N2 and N2O production from nitrification and 

denitrification: Global Biogeochemical Cycles, v. 10, no. 3, p. 401–412. 

Portnoy, J.W., Nowicki, B.L., Roman, C.T., and Urish, D.W., 1998, The discharge of nitratecontaminated groundwater from developed shoreline to marsh-fringed estuary: Water 

Resources Research, v. 34, no. 11, p. 3,095–3,104. 

Reay, W.G., Gallagher, D.L., and Simmons, G.M.J., 1992, Groundwater discharge and its impact 

on surface water quality in a Chesapeake Bay inlet: Journal of the American Water Resources 

Association, v. 28, p. 1,121–1,134. 

Roman, C.T., LaBash, C., Weber, J., and Rooney, S., 2001, Wetland and deepwater habitats, 

Northeast Creek (Mount Desert Island, Maine), Technical Report with Vegetation Map, U.S. 

National Park Service, Fort Collins, CO. 

Scheiner, D., 1976, Determination of ammonia and Kjeldahl nitrogen by indophenol method: 

Water Research, v. 10, p. 31–36. 

Seitzinger, S.P., 1988, Denitrification in freshwater and coastal marine ecosystems: ecological 

and geochemical significance: Limnology and Oceanography, v. 33, p. 702–724. 

Seitzinger, S.P., and Kroeze, C., 1998, Global distribution of nitrous oxide production and N 

inputs in freshwater and coastal marine ecosystems: Global Biogeochemical Cycles, v. 12, 

p. 93–113. 

Seitzinger, S.P., and Nixon, S.W., 1985, Eutrophication and the rate of denitrification and nitrous 

oxide production in coastal marine sediments: Limnology and Oceanography, v. 30, 

p. 1,332–1,339. 

Seitzinger, S.P., Nixon, S.W., and Pilson, M.E.Q., 1984, Denitrification and nitrous oxide 

production in a coastal marine ecosystem: Limnology and Oceanography, v. 29, p. 73–83. 

Silliman, B.R., and Bertness, M.D., 2004, Shoreline development drives invasion of Phragmites 

australis and the loss of plant diversity on New England salt marshes.: Conservation 

Biology v. 18, no. 5, p. 1,424–1,434. 

Smith, C.J., Delaune, R.D., and Patrick, W.H., 1985, Fate of riverine nitrate entering an estuary: 

I Denitrification and nitrogen burial: Estuaries, v. 8, p. 15–21. 

Tiedge, J.M., 1983 Denitrification. In A.L. Page and others, (eds.) Methods of soil analysis, 

Part 2. 2d ed. Agronomy 9:1011–1026. 

Tobias, C.R., Harvey, J.W., and Anderson, I.C., 2001, Quantifying groundwater discharge 

through fringing wetlands to estuaries: Seasonal variability, methods comparison, and 

implications for wetland-estuary exchange: Limnology and Oceanography, v. 46, p. 604–615. 

Valiela, I., and Bowen, J.L., 2002, Nitrogen sources to watersheds and estuaries: role of land 

cover mosaics and losses within watersheds: Environmental Pollution, v. 118, no. 2, 

p. 239–248. 
28

Valiela, I., Cole, M.L., McClelland, J., Hauxwell, J., Cebrian, J., and Joye, S., 2000, Role of Salt 

Marshes as Part of Coastal Landscapes, in Weinstein, M.P., and Kreeger, D.A., eds., Concepts 

and Controversies in Tidal Marsh Ecology: Dordrecht, Kluwer, p. 23–28. 

Valiela, I., Costa, J., Foreman, K., Teal, J.M., Howes, B., and Aubrey, D., 1990, Transport of 

groundwater-borne nutrients from watersheds and their effects on coastal waters: 

Biogeochemistry, v. 10, p. 177–197. 

Weier, K.L., Doran, J.W., Power, J.F., and Walters, D.T., 1993, Denitrification and the 

dinitrogen/nitrous oxide ratio as affected by soil water, available carbon, and nitrate: Soil 

Science Society of America Journal, v. 57, no. 1, p. 66–72. 

Wigand, C., McKinney, R.A., Chintala, M.M., Charpetier, M.A., and Groffman, P.M., 2004, 

Denitrification enzyme activity of fringe salt marshes in New England (USA): Journal of 

Environmental Quality, v. 33, p. 1,144–1,151. 

Wilde, F.D., variously dated, Field Measurements: U.S. Geological Survey Techniques of WaterResources Investigations, book 9, chap. A6, accessed april 20, 2011, at 

http://pubs.water.usgs.gov/twri9A/. 

Wood, E.D., Armstrong, F.A.J., and Richards, F.A., 1967, Determination of nitrate in sea water 

by cadmium-copper reduction to nitrite: Journal of the Marine Biological Association of the 

United Kingdom, v. 47, p. 23–31. 
Prepared by the Pembroke Publishing Service Center.

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Maine Water Science Center

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Huntington and others—Denitrification Rates in Marsh Soils and Hydrologic Data for Two Watersheds, Mount Desert Island, Maine —Open-File Report 2011-1252