Aquatic Sciences

, Volume 79, Issue 4, pp 891–908 | Cite as

Molecular composition and bioavailability of dissolved organic nitrogen in a lake flow-influenced river in south Florida, USA

  • Oliva Pisani
  • Joseph N. Boyer
  • David C. Podgorski
  • Cassondra R. Thomas
  • Teresa Coley
  • Rudolf Jaffé
Research Article


Dissolved organic nitrogen (DON) represents a large percentage of the total nitrogen in rivers and estuaries, and can contribute to coastal eutrophication and hypoxia. This study reports on the composition and bioavailability of DON along the Caloosahatchee River (Florida), a heavily managed system receiving inputs from Lake Okeechobee as well as agricultural and urban runoff from the surrounding watershed. Water samples were collected bimonthly for 1 year beginning December 2014 at three stations along the river. Treatments included 28-day dark incubations with and without prior photo-irradiation. Concentrations of DON, ammonium, nitrate–nitrite, total hydrolyzable amino acids (THAA), and urea, as well as bacterial numbers, leucine aminopeptidase activity, and fluorescent optical properties were measured. Ultra-high resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) was used to characterize the molecular composition of DON before and after incubation for selective samples. The total dissolved N pool was dominated by DON (61–99%), with low inorganic N (1–39%), and small amounts of THAA-N (0.1–23%) and urea-N (0.6–3.2%). The mean percentage of biologically available DON (BDON) for the study was 15% (−12–61% range) with highest values occurring when water inputs from Lake Okeechobee were the most dominant freshwater source. FT-ICR MS analysis revealed the presence of a wide range of N-containing formulas and the generation of aliphatic and ‘peptide-like’ structures likely due to microbial alteration of the carbon skeleton of DON compounds. Effects of light exposure prior to incubation did not have a measurable effect on %BDON but did affect bacterial biomass and DON composition. These findings may help predict nutrient loading effects to the Caloosahatchee River estuary and may aid in understanding wetland potential as a treatment technology for removing N in this and other freshwater systems sensitive to N loading.


Estuary Eutrophication Amino acids Urea Optical properties FT-ICR MS 



This work was funded through the South Florida Water Management District (SFWMD Contract #4600003105) and supported, in part, by a Section 319 Nonpoint Source Management Program Implementation grant from the U.S. Environmental Protection Agency through an agreement/contract with the Nonpoint Source Management Section of the Florida Department of Environmental Protection. The authors thank the field crew from the SFWMD for logistical support, and the National High Magnetic Field Laboratory at Florida State University for the mass spectrometric analyses (NSF DMR-1157490). Additional support from the George Barley Endowment and the FCE-LTER program (Grant No. DEB-1237517) is acknowledged. This is contribution number 832 from the Southeast Environmental Research Center in the Institute of Water & Environment at Florida International University.

Supplementary material

27_2017_540_MOESM1_ESM.docx (62 kb)
Supplementary material 1 (DOCX 62 KB)


  1. Amado AM, Cotner JB, Cory RM, Edhlund BL, McNeill K (2014) Disentangling the interactions between photochemical and bacterial degradation of dissolved organic matter: amino acids play a central role. Environ Microbiol. doi: 10.1007/s00248-014-0512-4 Google Scholar
  2. Amon RMW, Benner R (1996) Bacterial utilization of different size classes of dissolved organic matter. Limnol Oceanogr 41:41–51CrossRefGoogle Scholar
  3. Amon RMW, Fitznar HP, Benner R (2001) Linkages among the bioreactivity, chemical composition, and diagenetic state of marine dissolved organic matter. Limnol Oceanogr 46:287–297CrossRefGoogle Scholar
  4. Antonini GA, Fann DA, Roat P (2002) A historical geography of southwest Florida waterways, volume 2, Placida Harbor to Marco Island. National Seagrant College Program, Silver SpringGoogle Scholar
  5. Bailey N, Magley W, Mandrup-Poulsen J, O’Donnell K, Peets R (2009) Nutrient TMDL for the Caloosahatchee Estuary (WBIDs 3240 A, 3240B, and 3240C). Florida Department of Environmental Protection. TMDL Report, p 119Google Scholar
  6. Benner R, Kaiser K (2011) Biological and photochemical transformations of amino acids and lignin phenols in riverine dissolved organic matter. Biogeochem 102:209–222CrossRefGoogle Scholar
  7. Berman T, Bronk DA (2003) Dissolved organic nitrogen: a dynamic participant in aquatic ecosystems. Aquat Microb Ecol 31:279–305CrossRefGoogle Scholar
  8. Bernal S, Butturini A, Sabater F (2005) Seasonal variations of dissolved nitrogen and DOC:DON ratios in an intermittent Mediterranean stream. Biogeochem 75:351–372CrossRefGoogle Scholar
  9. Blakney GT, Hendrickson CL, Marshall AG (2011) Perdator data station: a fast data acquisition system for advanced FT-ICR MS experiments. Int J Mass Spectrom 306:246–252CrossRefGoogle Scholar
  10. Borsuk ME, Stow CA, Reckhow KH (2004) Confounding effect of flow on estuarine response to nitrogen loading. J Environ Eng 130:605–614CrossRefGoogle Scholar
  11. Bronk DA, Ward BB (1999) Gross and net nitrogen uptake and DON release in the euphotic zone of Monterey Bay, California. Limnol Oceanogr 44:573–585CrossRefGoogle Scholar
  12. Bronk DA, Glibert PM, Ward BB (1994) Nitrogen uptake, dissolved organic nitrogen release, and new production. Science 265:1843–1846CrossRefPubMedGoogle Scholar
  13. Burley SK, David PR, Lipscomb WN (1991) Leucine aminopeptidase: Bestatin inhibition and a model for enzyme-catalyzed peptide hydrolysis. Proc Natl Acad Sci USA 88:6916–6920CrossRefPubMedPubMedCentralGoogle Scholar
  14. Bushaw KL, Zepp RG, Tarr MA, Schulz-Jander D, Bourbonniere RA, Hodson RE, Miller WL, Bronk DA, Moran MA (1996) Photochemical release of biologically available nitrogen from aquatic dissolved organic matter. Nature 381:404–407CrossRefGoogle Scholar
  15. Buzzelli C, Wan Y, Doering PH, Boyer JN (2013) Seasonal dissolved inorganic nitrogen and phosphorus budgets for two sub-tropical estuaries in south Florida, USA. Biogeosciences 10:6721–6736CrossRefGoogle Scholar
  16. Caccia VG, Boyer JN (2007) A nutrient loading budget for Biscayne Bay, Florida. Mar Pollut Bull 54:994–1008CrossRefPubMedGoogle Scholar
  17. Castell JV, Cervera M, Marco R (1979) A convenient micromethod for the assay of primary amines and proteins with fluorescamine. A reexamination of the conditions of reaction. Anal Biochem 99:379–391CrossRefPubMedGoogle Scholar
  18. Chen M, Jaffé R (2014) Photo- and bio-reactivity patterns of dissolved organic matter from biomass and soil leachates and surface waters in a subtropical wetland. Water Res 61:181–190CrossRefPubMedGoogle Scholar
  19. Chróst RJ, Velimirov B (1991) Measurement of enzyme kinetics in water samples: effect of freezing and soluble stabilizer. Mar Ecol Prog Ser 70:93–100CrossRefGoogle Scholar
  20. Conley DJ, Paerl HW, Howarth RW, Boesch DF, Setzinger SP, Havens KE, Lancelot C, Likens GE (2009) Eutrophication: time to adjust expectations—response. Science 324:724–725CrossRefGoogle Scholar
  21. Cory RM, McKnight DM (2005) Fluorescence spectroscopy reveals ubiquitous presence of oxidized and reduced quinones in dissolved organic matter. Environ Sci Technol 39:8142–8149CrossRefPubMedGoogle Scholar
  22. Del Vecchio R, Blough NV (2002) Photobleaching of chromophoric dissolved organic matter in natural waters: Kinetics and modeling. Mar Chem 78:231–253CrossRefGoogle Scholar
  23. Ding Y, Watanabe A, Jaffé R (2014) Dissolved black nitrogen (DBN) in freshwater environments. Org Geochem 68:1–4CrossRefGoogle Scholar
  24. Dittmar T, Koch B, Hertkorn N, Kattner G (2008) A simple and efficient method for the solid-phase extraction of dissolved organic matter (SPE-DOM) from seawater. Limnol Oceanogr Meth 6:230–235CrossRefGoogle Scholar
  25. Doering PH, Chamberlain RH (1999) Water quality and source of freshwater discharge to the Caloosahatchee Estuary, Florida. J Am Water Resour Assoc 35:793–806CrossRefGoogle Scholar
  26. Doering PH, Chamberlain RH, Haunert KE (2002) Using submerged aquatic vegetation to establish minimum and maximum freshwater flows to the Caloosahatchee Estuary, Florida. Estuaries 25:1343–1354CrossRefGoogle Scholar
  27. Doering PH, Haunert KE, Qiu C, Coley T (2011) Caloosahatchee River estuary and southern Charlotte Harbor. 2010 South Florida Environmental Report, volume 1, Chap 12, p 189Google Scholar
  28. Findlay SEG, Sinsabaugh RL (2003) Aquatic ecosystems: interactivity of dissolved organic matter. Academic Press, San DiegoGoogle Scholar
  29. Frost PC, Benstead JP, Cross WF, Hillebrand H, Larson JH, Xenopoulos MA, Yoshida T (2006) Threshold elemental ratios of carbon and phosphorus in aquatic consumers. Ecol Lett 9:774–779CrossRefPubMedGoogle Scholar
  30. Garcia JC, Ketover DJ, Loh AN, Parsons ML, Urakawa H (2015) Influence of freshwater discharge on the microbial degradation processes of dissolved organic nitrogen in a subtropical estuary. A van Leeuw J Microb 107:613–632CrossRefGoogle Scholar
  31. Glibert PM, Harrison J, Heil C, Seitzinger S (2006) Escalating worldwide use of urea—a global change contributing to coastal eutrophication. Biogeochem 77:441–463CrossRefGoogle Scholar
  32. Green NW, Perdue EM, Aiken GR, Butler KD, Chen H, Dittmar T, Niggemann J, Stubbins A (2014) An intercomparison of three methods for the large-scale isolation of oceanic dissolved organic matter. Mar Chem 161:14–19CrossRefGoogle Scholar
  33. Hansen AM, Kraus TEC, Pellerin BA, Fleck JA, Downing BD, Bergamaschi BA (2016) Optical properties of dissolved organic matter (DOM): effects of biological and photolytic degradation. Limnol Oceanogr 61:1015–1032CrossRefGoogle Scholar
  34. Helms JR, Stubbins A, Ritchie JD, Minor EC, Kieber DJ, Mopper K (2008) Absorption spectral slopes and slope ratios as indicators of molecular weight, source, and photobleaching of chromophoric dissolved organic matter. Limnol Oceanogr 53:955–969CrossRefGoogle Scholar
  35. Hendrickson J, Trahan N, Gordon E, Ouyang Y (2007) Estimating relevance of organic carbon, nitrogen, and phosphorus loads to a blackwater river estuary. J Am Water Resour Assoc (JAWRA) 43:264–279CrossRefGoogle Scholar
  36. Ikenberry C, Bhimani S, McDonald S, Jenkins D, Dennis G, Gray R (2002) Bioavailability of dissolved organic nitrogen in WWTP effluent: a Truckee River case study. In: Proceedings of the Water Environment Federation, WEFTEC 2002: Session 61–70, pp 575–585Google Scholar
  37. Jaffé R, McKnight D, Maie N, Cory R, McDowell WH, Campbell JL (2008) Spatial and temporal variations in DOM composition in ecosystems: the importance of long-term monitoring of optical properties. J Geophys Res. doi: 10.1029/2008JG000683 Google Scholar
  38. Jørgensen L, Markager S, Maar M (2014) On the importance of quantifying bioavailable nitrogen instead of total nitrogen. Biogeochem 117:455–472CrossRefGoogle Scholar
  39. Kaiser NK, Quinn JP, Blakney GT, Hendrickson CL, Marshall AG (2011) A novel 9.4 T FTICR mass spectrometer with improved sensitivity, mass resolution, and mass range. J Am Soc Mass Spectrom 22:1343–1351CrossRefPubMedGoogle Scholar
  40. Kaushal SS, Lewis Jr WM (2005) Fate and transport of dissolved organic nitrogen in minimally disturbed streams of Colorado, USA. Biogeochem 74:303–321CrossRefGoogle Scholar
  41. Kiersztyn B, Suida W, Chrøst RJ (2012) Persistence of bacterial proteolytic enzymes in lake ecosystems. FEMS Microbiol Ecol 80:124–134CrossRefPubMedGoogle Scholar
  42. Kim S, Kramer RW, Hatcher PG (2003) Graphical method for analysis of ultrahigh-resolution broadband mass spectra of natural organic matter, the van Krevelen diagram. Anal Chem 75:5336–5344CrossRefPubMedGoogle Scholar
  43. Knicker H (2010) “Black nitrogen”—an important fraction in determining the recalcitrance of charcoal. Org Geochem 41:947–950CrossRefGoogle Scholar
  44. Kragh T, Sondergaard M, Tranvik L (2008) Effect of exposure to sunlight and phosphorus-limitation on bacterial degradation of coloured dissolved organic matter (CDOM) in freshwater. FEMS Microbiol Ecol 64:230–239CrossRefPubMedGoogle Scholar
  45. Lechtenfeld OJ, Kattner G, Flerus R, McCallister SL, Schmitt-Kopplin P, Koch BP (2014) Molecular transformation and degradation of refractory dissolved organic matter in the Atlantic and Southern Ocean. Geochim Cosmochim Acta 126:321–337CrossRefGoogle Scholar
  46. Leenheer JA, Croue J (2003) Characterizing dissolved organic matter. Environ Sci Technol 37:18 A–26 ACrossRefGoogle Scholar
  47. Liu Z, Choudhury SH, Xia M, Holt J, Wallen CM, Yuk S, Sanborn SC (2009) Water quality assessment of coastal Caloosahatchee River watershed, Florida. J Environ Sci Health A 44:972–984CrossRefGoogle Scholar
  48. Lusk MG, Toor GS (2016a) Dissolved organic nitrogen in urban streams: biodegradability and molecular composition studies. Water Res 96:225–235CrossRefPubMedGoogle Scholar
  49. Lusk MG, Toor GS (2016b) Biodegradability and molecular composition of dissolved organic nitrogen in urban stormwater runoff and outflow water from a stormwater retention pond. Environ Sci Technol 50:3391–3398CrossRefPubMedGoogle Scholar
  50. Maie N, Parish KJ, Watanabe A, Knicker H, Benner R, Abe T, Kaiser K, Jaffé R (2006) Chemical characteristics of dissolved organic nitrogen in an oligotrophic subtropical coastal ecosystem. Geochim Cosmochim Acta 70:4491–4506CrossRefGoogle Scholar
  51. Maie N, Pisani O, Jaffé R (2008) Mangrove tannins in aquatic ecosystems: their fate and possible influence on dissolved organic carbon and nitrogen cycling. Limnol Oceanogr 53:160–171CrossRefGoogle Scholar
  52. McKnight DM, Boyer EW, Westerhoff PK, Doran PT, Kulbe T, Andersen DT (2001) Spectrofluorometric characterization of dissolved organic matter for indication of precursor organic material and aromaticity. Limnol Oceanogr 46:38–48CrossRefGoogle Scholar
  53. Moran MA, Zepp RG (1997) Role of photoreactions in the formation of biologically labile compounds from dissolved organic matter. Limnol Oceanogr 42:1307–1316CrossRefGoogle Scholar
  54. Obernosterer I, Benner R (2004) Competition between biological and photochemical processes in the mineralization of dissolved organic carbon. Limnol Oceanogr 49:117–124CrossRefGoogle Scholar
  55. Osborne DM, Podgorski DC, Bronk DA, Roberts Q, Sipler RE, Austin D, Bays JS, Cooper WT (2013) Molecular-level characterization of reactive and refractory dissolved natural organic nitrogen compounds by atmospheric pressure photoionization coupled to Fourier transform ion cyclotron resonance mass spectrometry. Rapid Commun Mass Spectrom 27:851–858CrossRefPubMedGoogle Scholar
  56. Podgorski DC, McKenna AM, Rodgers RP, Marshall AG, Cooper WT (2012) Selective ionization of dissolved organic nitrogen by positive ion atmospheric pressure photoionization coupled with Fourier transform ion cyclotron resonance mass spectrometry. Anal Chem 84:5085–5890CrossRefPubMedGoogle Scholar
  57. Porter KG, Feig Y (1980) The use of DAPI for identifying and counting aquatic microflora. Limnol Oceanogr 25:943–948CrossRefGoogle Scholar
  58. Redfield AC (1958) The biological control of chemical factors in the environment. Am Sci 46:205–221Google Scholar
  59. Revilla M, Alexander J, Glibert PM (2005) Urea analysis in coastal waters: comparison of enzymatic and direct methods. Limnol Oceanogr Method 3:290–299CrossRefGoogle Scholar
  60. Rudnick DT, Chen Z, Childers DL, Boyer JN, Fontaine TD (1999) Phosphorus and nitrogen inputs to Florida Bay: the importance of the Everglades watershed. Estuaries 22:398–416CrossRefGoogle Scholar
  61. Savory JJ, Kaiser NK, McKenna AM, Xian F, Blakney GT, Rodgers RP, Hendrickson CL, Marshall AG (2011) Parts-per-billion Fourier transform ion cyclotron resonance mass measurement accuracy with a “walking” calibration equation. Anal Chem 83:1732–1736CrossRefPubMedGoogle Scholar
  62. Schindler DW, Hecky RE, Findlay DL, Stainton MP, Parker BR, Paterson MJ, Beaty KG, Lyng M, Kaisan SEM (2008) Eutrophication of lakes cannot be controlled by reducing nitrogen input: results of a 37-year whole-ecosystem experiment. P Natl Acad Sci USA 105:11254–11258CrossRefGoogle Scholar
  63. Scully NM, Maie N, Dailey S, Boyer JN, Jones RD, Jaffé R (2004) Early diagenesis of plant-derived dissolved organic matter along a wetland, mangrove, estuary ecotone. Limnol Oceanogr 49:1667–1678CrossRefGoogle Scholar
  64. See JH, Bronk DA, Lewitus AJ (2006) Uptake of Spartina-derived humic nitrogen by estuarine phytoplankton in nonaxenic and axenic culture. Limnol Oceanogr 51:2290–2299CrossRefGoogle Scholar
  65. Seidel M, Yager PL, Ward ND, Carpenter EJ, Gomes HR, Krusche AV, Richey JE, Dittmar T, Medeiros PM (2015) Molecular-level changes of dissolved organic matter along the Amazon River-to-ocean continuum. Mar Chem 117:218–231CrossRefGoogle Scholar
  66. Seitzinger SP, Sanders R (1997) Contribution of dissolved organic nitrogen from rivers to estuarine eutrophication. Mar Ecol Prog Ser 159:1–12CrossRefGoogle Scholar
  67. Seitzinger SP, Sanders RW, Styles R (2002) Bioavailability of DON from natural and anthropogenic sources to estuarine plankton. Limnol Oceanogr 47:353–366CrossRefGoogle Scholar
  68. Sinsabaugh RL, Follstad Shah JJ (2012) Ecoenzymatic stoichiometry and ecological theory. Annu Rev Ecol Evol S 43:313–343CrossRefGoogle Scholar
  69. Sleighter RL, Cory RM, Kaplan LA, Abdulla HA, Hatcher PG (2014) A coupled geochemical and biogeochemical approach to characterize the bioreactivity of dissolved organic matter from a headwater stream. J Geophys Res Biogeosci 119:1520–1537CrossRefGoogle Scholar
  70. Solórzano L, Sharp JH (1980) Determination of total dissolved phosphorus and particulate phosphorus in natural waters. Limnol Oceanogr 25:754–758CrossRefGoogle Scholar
  71. Spencer RGM, Guo W, Raymond PA, Dittmar T, Hood E, Fellman J, Stubbins A (2014) Source and biolability of ancient dissolved organic matter in glacier and lake ecosystems on the Tibetan Plateau. Geochim Cosmochim Acta 142:64–74CrossRefGoogle Scholar
  72. Stepanauskas R, Leonardson L, Tranvik LJ (1999) Bioavailability of wetland-derived DON to freshwater and marine bacterioplankton. Limnol Oceanogr 44:1477–1485CrossRefGoogle Scholar
  73. Stepanauskas R, Laudon H, Jorgensen NOG (2000) High DON bioavailability in boreal streams during a spring flood. Limnol Oceanogr 45:1298–1307CrossRefGoogle Scholar
  74. Stepanauskas R, Jørgensen NOG, Eigaard OR, Žvikas A, Tranvik LJ, Leonardson L (2002) Summer inputs of riverine nutrients to the Baltic Sea: bioavailability and eutrophication relevance. Ecol Monogr 72:579–597CrossRefGoogle Scholar
  75. Sterner RW, Elser JJ (2002) Ecological Stoichiometry: the biology of elements from molecules to the biosphere. Princeton University Press, PrincetonGoogle Scholar
  76. Tranvik LJ, Berlitsson S (2001) Contrasting effects of solar UV radiation on dissolved organic sources for bacterial growth. Ecol Lett 4:458–463CrossRefGoogle Scholar
  77. Vahatalo AV, Wetzel RG (2008) Long-term photochemical and microbial decomposition of allochthonous organic carbon in a large humic lake. Limnol Oceanogr 53:1387–1392CrossRefGoogle Scholar
  78. Varela MM, Barquero S, Bode A, Fernández E, González N, Teira E, Varela M (2003) Microplanktonic regeneration of ammonium and dissolved organic nitrogen in the upwelling area of the NW of Spain: relationships with dissolved organic carbon production andphytoplankton size-structure. J Plankton Res 25:719–736CrossRefGoogle Scholar
  79. Wagner S, Dittmar T, Jaffé R (2015) Molecular characterization of dissolved black nitrogen via electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry. Org Geochem 79:21–30CrossRefGoogle Scholar
  80. Wang H, Wang H (2009) Mitigation of lake eutrophication: Loosen nitrogen control and focus on phosphorus abatement. Prog Nat Sci 19:1445–1451CrossRefGoogle Scholar
  81. Wetland Solutions (2012) Evaluation of total nitrogen reduction options for the C-43 Water Quality Treatment Area Test Facility, Final Task 2 Report. Prepared for the South Florida Water Management DistrictGoogle Scholar
  82. White JR, Reddy KR (2003) Nitrification and denitrification rates of Everglades wetland soils along a phosphorus-impacted gradient. J Environ Qual 32:2436–2443CrossRefPubMedGoogle Scholar
  83. Wiegner TN, Seitzinger SP, Glibert PM, Bronk DA (2006) Bioavailability of dissolved organic nitrogen and carbon from nine rivers in the eastern United States. Aquat Microb Ecol 43:277–287CrossRefGoogle Scholar
  84. Wymore AS, Rodríguez-Cardona B, McDowell WH (2015) Direct response of dissolved organic nitrogen to nitrate availability in headwater streams. Biogeochem 126:1–10CrossRefGoogle Scholar
  85. Zsolnay A, Baigar E, Jimenez M, Steinweg B, Saccomandi F (1999) Differentiating with fluorescence spectroscopy the sources of dissolved organic matter in soils subjected to drying. Chemosphere 38:45–50CrossRefPubMedGoogle Scholar

Copyright information

© Springer International Publishing 2017

Authors and Affiliations

  • Oliva Pisani
    • 1
    • 5
  • Joseph N. Boyer
    • 2
  • David C. Podgorski
    • 3
  • Cassondra R. Thomas
    • 4
  • Teresa Coley
    • 4
  • Rudolf Jaffé
    • 1
  1. 1.Department of Chemistry and Biochemistry, Southeast Environmental Research CenterFlorida International UniversityMiamiUSA
  2. 2.Department of Environmental Science and Policy, Center for the EnvironmentPlymouth State UniversityPlymouthUSA
  3. 3.Department of Earth, Ocean and Atmospheric ScienceNational High Magnetic Field LaboratoryTallahasseeUSA
  4. 4.Coastal Ecosystems SectionSouth Florida Water Management DistrictWest Palm BeachUSA
  5. 5.Southeast Watershed Research LaboratoryUSDA-ARSTiftonUSA

Personalised recommendations