the freshwater dissolved organic matter fluorescence–total organic carbon relationship

HYDROLOGICAL PROCESSESHydrol. Process. 21, 2093–2099 (2007)Published online 4 June 2007 in Wiley InterScience(www.interscience.wiley.com) DOI: 10.1002/hyp.6371

The freshwater dissolved organic matter fluorescence–totalorganic carbon relationship

Susan A. Cumberland and Andy Baker*School of Geography, Earth and Environmental Sciences, The University of Birmingham, Birmingham B15 2TT, UK

Abstract:

The fluorescent properties of dissolved organic matter (DOM) enable comparisons of humic-like (H-L) and fulvic-like (F-L)fluorescence intensities with dissolved organic carbon (DOC) in aquatic systems. The fluorescence-DOC relationship differedin gradient, i.e. the fluorescence per gram of carbon, and in the strength of the correlation coefficient. We compare thefluorescence intensity of the F-L and H-L fractions and DOC of freshwater DOM in north Shropshire, England, featuring ariver, wetland, spring, pond and sewage DOM sources. Correlations between fluorescence and DOC varied between samplesites. Wetland water samples for the F-L peak gave the best correlation, r D 0Ð756; the lowest correlation was from finaltreated sewage effluent, r D 0Ð167. The relationship between fluorescence and DOC of commercially available InternationalHumic Substances Society standards were also examined and they generally showed a lower fluorescence per gram of carbonfor the F-L peak than the natural samples, whereas peat wetland DOM gave a greater fluorescence per gram of carbon thanriver DOM. Here, we propose the strength of the fluorescence–DOC correlation to be a useful tool when discriminatingsources of DOM in fresh water. Copyright 2007 John Wiley & Sons, Ltd.

KEY WORDS dissolved organic carbon; dissolved organic matter; fluorescence; wetlands; rivers

Received 20 December 2005; Accepted 22 May 2006

INTRODUCTION

Humic substances occur in every natural water samplethat has ever been analysed for them (Malcolm, 1985).Dissolved humic substances can be defined as passingthrough a 0Ð45 µm membrane, and are made up of humicand fulvic substances, the latter being present in largerproportion and characterized as being soluble under allpH conditions (Aiken, 1985). Freshwater dissolved humicsubstances are derived from dead and decaying detritus,aquatic plants, animals and debris from overhanging veg-etation, and tend to be of a younger state than that ofsoil humic substances (Hongve, 1999). River aquatic ful-vic substances are derived from plant and tree residues,which contain more phenolic and lignin-derived organiccompounds than those found in soil (Chen et al., 2003).Historically, much work has gone into the determinationand separation of these humic and fulvic substances, andyet still relatively little is understood about the molec-ular distribution and fluorescent characteristics of thesemolecules (Alberts and Takacs, 2004). Analysis of freeze-dried fulvic acid using 13C nuclear magnetic resonancespectroscopy demonstrates that it contains a number ofgroups, such as aliphatic carbon atoms, carbohydrates,olefinic carbon atoms, aromatic carbons atom, carboxylester amide carbon atoms and ketonic and aldehydeiccarbon atoms (Klapper et al., 2002; Sierra et al., 2005).They are responsible for water colour (Hongve, 1999) and

* Correspondence to: Andy Baker, School of Geography, Earth andEnvironmental Sciences, The University of Birmingham, BirminghamB15 2TT, UK. E-mail: [email protected]

sometimes pH due to the carboxylic acid groups attachedto them (Klapper et al., 2002). The aromatic and car-boxylic fractions provide the fluorescence of humic andfulvic substances (Senesi et al., 1989); fulvic substancesare more highly fluorescent than humic substances andhumic fluorescence intensity is more pH dependent(Goslan et al., 2004; Sierra et al., 2005). Owing to theambiguous characteristics of these substances, we feelthat it is more appropriate to refer to them, hereafter, ashumic-like (H-L) and fulvic-like (F-L) substances.

In freshwaters, it has been suggested that dissolvedorganic carbon (DOC) is usually dominated by the H-Land F-L fractions, making up 50–75% of the total (ofwhich colloidal matter comprises up to 20% of the totalDOC; Hope et al., 1994). However, the proportion of H-Land F-L substances in the DOC is variable. For example,about 50% of uncoloured (USA) stream water consistsmostly of humic substances and 90% of these humicsubstances found in dissolved organic matter (DOM)are the F-L fraction (Malcolm, 1985). In the Amazonbasin, humic substances are attributed to as much as60% of the aquatic total organic carbon (TOC; Patel-Sorrentino et al., 2002). In stream/aquatic systems, DOClosses are possible due to biotic processes such as biofilmrespiration and absorption onto algae and abiotic uptakeonto mineral surfaces (Dawson et al., 2001). Losses ofCO2 from volatile DOC fractions by degassing to theatmosphere, as well as variable DOM inputs throughoutand between catchments, can also lead to this variability.

The presence of fluorescent DOM enables the compar-ison of H-L and F-L fluorescence intensity with DOC.

Copyright 2007 John Wiley & Sons, Ltd.

2094 S. A. CUMBERLAND AND A. BAKER

There are many potential benefits of using fluorescence asa tool for determining and measuring DOC in aquatic sys-tems. It is a relatively quick process, with a typical scantaking approximately 1 min; this is valuable, as labileDOM can degrade over time. The method is non-intrusiveand thus does not interfere with the molecular structure(Klabitz et al., 2000; Baker, 2002). However, given thevariable fraction of fluorescent DOM as part of the totalDOC pool, it is essential to investigate the fluorescenceintensity–DOC relationship. Therefore, we compare thefluorescence intensity of the F-L fraction and DOC offreshwater DOM. The study site that this paper focuseson, Norton in Hales, in north Shropshire, England, is asite that features river, wetland, spring, pond and sewageDOM sources all within a small location (approximately500 m), covering a wide range of variation in aquatic andsoil-derived water samples.

SITE DESCRIPTION AND METHODOLOGY

Norton in Hales [UK National Grid Reference SJ706 384], north Shropshire, UK, is situated along theRiver Tern, a tributary of the River Severn, 16 kmfrom its source. The River Tern is a lowland catchment,primarily groundwater fed from red sandstone overly-ing the Permo-Triassic Sherwood sandstone. Its catch-ment boundary, thus, is not defined by its topography(described by Clay et al. (2004)). This amounts to avery stable base flow in the River Tern, showing min-imal response to normal rainfall events. Local land useis generally under semi-intense mixed cereal/dairy/beeffarming.

Figure 1 illustrates the diversity of the sample siteswithin a small area. In-stream water samples werecollected (river sites A, C, E and H), as a transect downthe river starting above the wetland (A) and finishingbelow the sewage treatment outflow pipes (H). Betweenthese points, samples were also taken from the sewage

treatment works (STW) outlet itself (G), a groundwater-fed pond overflow, POND (B), a wetland dischargesite (D) (not always connected to the main river) anda groundwater spring diverted through small concreteconduit (F). Additionally, three samples were taken fromthe wetland: surface water (I), 0–45 cm depth (J), and50–100 cm depth (K). To obtain samples from the deeperpeat, two perforated drain pipes, about 50 cm apart, weresunk (to 50 cm deep and 1 m deep, with the latter beingperforated from 50 to 100 cm) and sealed at both ends, toreduce oxidation and contamination from other externalsources. The sample was obtained by unscrewing a lidand dipping for the sample, throwing away the first fewdips so that the sample was taken from the middle of theperforated column depth. The wells were then completelyemptied before resealing. Thus, water in tubes was notallowed to stagnate, but perforation allowed ‘flow’ ofthe wetland and was representative of (actual) bog water.These samples (D, I, J and K) were grouped together toform BOG.

Water samples were collected fortnightly from Nor-ton in Hales from March to October 2005. Plastic bot-tles (500 ml), pre acid washed (10% HCl, distilled-waterrinsed), were used to ensure carbon- and fluorescence-free blanks (Bolton, 2004). Bottles were also pre-rinsedwith sample before filling. On return to the laboratory(same day) the samples were then filtered through pre-rinsed Whatman GF/C filter papers under vacuum. Sam-ples were kept in the refrigerator overnight between 4 and6 °C they were then analysed for fluorescence and TOCthe next day, with DOC analysis completed with 36 h ofcollection (on two occasions this was a day later due tomachine downtime); this was critical, as the fluorescenceproperties of a sample can alter over time (Baker, 2002).

DOC was measured using a Shimadzu 5050 carbonanalyser from filtered and unfiltered (not reported here)samples. DOC was obtained by measuring the total car-bon and subtracting the measured inorganic carbon andcarbonates (dissolved CO2 and CO2�

3 ). As other studies

H GF

B

E

J K

I

C

A

Ponddrain

D

N

Sewage Works

Key

Telford BirminghamSJ 707 385

RiverTern

NORTON INHALES

Norton-in-Hales

0 50 100 metres

Bog

Water

Figure 1. Location of sample sites

Copyright 2007 John Wiley & Sons, Ltd. Hydrol. Process. 21, 2093–2099 (2007)DOI: 10.1002/hyp

FRESHWATER DOM FLUORESCENCE–TOC RELATIONSHIP 2095

have demonstrated, some methodologies suggest purgingwith acid to remove the inorganic fraction (Clark et al.,2002). This was not executed in this case because thetotal carbon and inorganic carbon concentrations are sig-nificantly different from each other, giving differencingerrors of <1 mg l�1 for DOC. Calibration was with cer-tified organic and inorganic standards from Reageacon,and samples were analysed in triplicate and the meantaken from the best two (CV <2%). Glass vials werewashed in 4% Decon 90 and 10% HCl and pre-rinsedwith ultra-pure water and sample.

A fluorescence scan for the organic fraction of the sam-ple was obtained from filtered samples within 48 h ofsample collection. DOM fluorescence was measured in4 ml capacity cuvettes using a Varian Cary Eclipse flu-orescence spectrophotometer equipped with a multicellholder with Peltier temperature controller to enable themeasurement of excitation–emission matrices (EEMs)at precisely controlled (š0Ð1 °C) temperatures at 20 °C.Each EEM was generated by scanning excitation wave-lengths from 200 to 400 nm at 5 nm steps and detectingthe emitted fluorescence between 280 and 500 nm at2 nm steps. Scan speed was 9600 nm min�1, permit-ting collection of a complete EEM in ¾60 s. For allEEMs, two fluorescence peaks were identified that werealways detectable. These were (1) fluorescence excitedbetween 300 and 340 nm excitation and emitted between400 and 460 nm, and (2) fluorescence excited between220 and 250 nm excitation and emitted between 400 and460 nm, both fluorophores attributed to F-L and H-L sub-stances (Baker and Inverarity, 2004). For each peak, theexcitation and emission wavelengths and the intensity ofemitted fluorescence were recorded. To calibrate the flu-orescence intensity, we also measured the strength of theRaman signal at excitation 348 nm (emitted between 395and 400 nm) and all results are standardized to a meanRaman peak of 20 intensity units. Our results can becompared with a quinine sulphate standard: 32Ð5 intensityunits are equivalent to one quinine sulphate unit (1 µg l�1

in 0Ð1 M H2SO4). Samples were not diluted, with theexception of one intensely fluorescent BOG sample, forwhich a ð2 dilution, to bring to scale, was used.

To be able to put our sample sites in a wider con-text, baseline geochemical and nutrient data have alsobeen collected (Table I). The pH was measured in thefield using a Jenway hand-held meter. Determinationof nutrients (nitrate, ammonium and phosphate) wasundertaken at the University of Reading using a Chem-Lab continuous flow autoanalyser. Samples were thenmicrowave digested (CEM Corporation) using the per-sulphate method as described by Johnes and Heathwaite,(1992) and then reanalysed to determine the organicfraction.

RESULTS

Table I presents summary fluorescence and DOC results,together with secondary geochemical and nutrient data.

Data from sites A, C, E, and H are combined to form‘RIVER’; sites D, I, J, and K are combined to form‘BOG’. The treated final effluent (STW), pond site B(POND) and groundwater site F (SPRG) were analysedwithout grouping.

For all groups, the excitation and emission wavelengthsand intensities of H-L and F-L fluorescence, as wellas DOC concentrations, fall within the range reportedfrom other rivers within the UK (Baker and Inverarity,2004; Baker and Spencer, 2004; Worrall et al., 2004),suggesting that DOM in the River Tern is typical ofrivers in the UK. Intensities for the river F-L peak rangedbetween 113 and 424 a.u., with a mean of 172 a.u. anda standard deviation of 69 a.u. Intensities for the BOGF-L peak ranged between 77 and 951 a.u., with a meanof 409 a.u. and a standard deviation of 208 a.u.

EEM fluorescence intensity values of the F-L peaks(fluorophores) for all samples were compared with mil-ligrams per litre DOC. The fluorescence per milligram perlitre of DOC is presented in Table I, and the correlationcoefficients between F-L fluorescence intensity and DOCin Figure 2, together with the fluorescence–DOC rela-tionship of commercially available International HumicSubstances Society (IHSS) standards: Nordic fulvic acid(referred to hereafter as Nordic FA), Suwanee River ful-vic acid (SRFA), Suwanee River humic acid (SRHA)and Suwanee River natural organic matter (SRNOM). Ingeneral, the Norton in Hales samples showed a higher flu-orescence per milligram of carbon for the fulvic peak thanthe IHSS standards, of which the Nordic FA showed themost fluorescence per gram of carbon of the IHSS stan-dards, then SRFA, then SRNOM, then SRHA displayingthe least intensity per unit carbon.

Correlations (product for normally distributed data;rank for non-normally distributed data) between fluores-cence and DOC varied between sample sites. BOG gavethe best correlation (r D 0Ð756), then POND (r D 0Ð657),SPRG (r D 0Ð631) and RIVER (r D 0Ð409), with theSTW giving the lowest correlation (r D 0Ð14). That BOGshowed less scatter than SPRG, POND and RIVER couldbe explained by its early stage of decomposition andrelative similarity in source and composition of the fluo-rescent molecules. The weak correlation observed in thesewage outlet could be attributed to other organic fluo-rescence chemicals, such as fluorescent whitening agentsthat are present in municipal sewage waste, and the lowgradient due to the relative insignificance of the F-Lfraction as a proportion of DOC compared with otherfluorophores.

Results for the H-L fluorophores for all samples werealso compared against DOC (Table I and Figure 3).H-L fluorophores generally had similar correlations withDOC than the F-L fluorescence. This is despite thefluorescence intensity for the H-L fluorophores beingmore dependent on pH (Patel-Sorrentino et al., 2002;Sierra et al., 2005), with fluorescence intensity increasingwith increasing pH (Mobed et al., 1996). The H-L peaksfor the BOG samples were harder to locate, as theyappear close or within the Rayleigh–Tyndall scatter line



Tabl

eI.

Fluo

resc

ence

,or

gani

can

din

orga

nic

carb

on,

nutr

ient

and

pHda

tafo

rth

efiv

esa

mpl

egr

oups

F-L

H-L

DO

C�m

gl�1

�In

t./D

OC

Org

anic

�mg

l�1�

C/N

pH

Ex.

(nm

)E

m.

(nm

)In

t.(a

.u.)

Ex.

(nm

)E

m.

(nm

)In

t.(a

.u.)

TC

ICD

OC

TO

Na

NO

2-N

NH

4-N

PO4-P

NP

All

data

Min

.30

040

626

219

408

7412

Ð960Ð3

00Ð2

90Ð0

00Ð0

00Ð0

00Ð0

00Ð4

9�0

Ð016Ð3

1M

ax.

344

450

951

255

461

1558

122Ð3

086

Ð7341

Ð6228

Ð920Ð3

25Ð6

54Ð6

74Ð8

90Ð7

18Ð1

5M

ean

331

425

220

237

428

366

48Ð47

37Ð53

10Ð95

20Ð14

8Ð05

0Ð05

0Ð51

0Ð52

2Ð10

0Ð09

1Ð28

7Ð30

SD8

816

47

722

517

Ð5713

Ð468Ð6

47Ð4

70Ð0

71Ð0

91Ð2

11Ð2

90Ð1

70Ð4

6N

o.29

229

229

229

129

129

114

814

814

829

2929

2929

2948

Riv

er Min

.31

541

711

323

541

421

723

Ð8318

Ð012Ð7

47Ð6

00Ð0

20Ð0

00Ð1

30Ð7

9�0

Ð016Ð8

2M

ax.

340

440

424

254

444

600

59Ð01

42Ð92

39Ð87

8Ð83

0Ð07

1Ð58

0Ð33

4Ð23

0Ð25

7Ð81

Mea

n33

042

717

223

942

830

942

Ð9434

Ð048Ð8

919

Ð387Ð6

40Ð0

40Ð4

10Ð1

82Ð0

70Ð0

71Ð1

17Ð5

4SD

75

695

688

6Ð23

5Ð51

7Ð54

2Ð33

0Ð02

0Ð65

0Ð09

1Ð42

0Ð10

0Ð27

No.

6464

6464

6464

6464

6412

1212

1212

1215

BO

G Min

.32

141

677

219

416

155

12Ð96

0Ð30

1Ð37

0Ð00

0Ð00

0Ð00

0Ð00

0Ð64

0Ð01

6Ð31

Max

.34

445

095

125

046

115

5812

2Ð386

Ð7341

Ð625Ð5

90Ð0

65Ð6

50Ð2

92Ð6

10Ð0

97Ð7

7M

ean

334

427

409

231

429

645

63Ð16

46Ð56

16Ð60

24Ð66

1Ð17

0Ð02

0Ð87

0Ð08

1Ð28

0Ð03

8Ð15

7Ð00

SD5

820

85

928

624

Ð4519

Ð899Ð9

02Ð7

50Ð0

31Ð8

80Ð1

11Ð2

40Ð0

80Ð4

6N

o.49

4949

4848

4949

4949

99

99

99

20

STW M

in.

320

406

177

240

420

284

22Ð42

15Ð91

4Ð79

14Ð87

0Ð18

0Ð17

2Ð21

3Ð72

0Ð25

7Ð01

Max

.34

042

631

125

043

640

746

Ð9735

Ð3317

Ð9528

Ð920Ð3

20Ð7

44Ð6

74Ð8

90Ð7

17Ð5

0M

ean

333

419

249

245

426

340

38Ð99

26Ð44

12Ð55

19Ð87

17Ð30

0Ð19

0Ð34

2Ð88

3Ð09

0Ð37

0Ð71

7Ð30

SD5

442

34

377Ð0

95Ð6

43Ð4

012

Ð990Ð1

40Ð3

22Ð2

32Ð1

30Ð3

10Ð2

3N

o.16

1616

1616

1616

1616

33

33

33

5

BM

in.

300

416

6823

041

615

929

Ð1625

Ð003Ð8

42Ð0

50Ð0

00Ð0

00Ð0

00Ð6

40Ð0

17Ð6

1M

ax.

330

436

112

245

434

255

51Ð08

40Ð68

14Ð65

2Ð06

0Ð08

0Ð04

0Ð01

0Ð89

0Ð00

8Ð15

Mea

n31

842

689

234

424

200

41Ð76

34Ð21

7Ð55

11Ð77

2Ð06

0Ð04

0Ð02

0Ð01

0Ð77

0Ð02

3Ð64

7Ð90

SD10

514

45

345Ð4

44Ð1

43Ð1

81Ð1

90Ð0

40Ð0

20Ð0

00Ð4

50Ð0

10Ð2

0N

o.14

1414

1414

1414

1414

22

22

22

5

FM

in.

315

412

4023

541

277

35Ð30

29Ð75

0Ð29

12Ð79

0Ð02

0Ð02

0Ð14

2Ð06

�0Ð01

6Ð82

Max

.34

042

816

025

143

826

059

Ð1146

Ð9016

Ð9618

Ð930Ð0

40Ð0

70Ð1

63Ð0

30Ð0

67Ð7

3M

ean

331

420

6524

442

411

241

Ð4736

Ð624Ð8

513

Ð4516

Ð790Ð0

30Ð0

40Ð1

52Ð6

00Ð0

20Ð2

97Ð2

1SD

65

285

847

6Ð20

5Ð14

4Ð31

8Ð86

0Ð02

0Ð03

0Ð07

1Ð36

0Ð03

0Ð38

No.

1616

1616

1616

1616

163

33

33

34

aTo

tal

oxid

ized

nitr

ogen

(nitr

ate

and

nitr

ite).



0 10 20 30 40 50 60

Inte

nsity

Inte

nsity

Inte

nsity

0

200

400

600

800

1000

0 10 20 30 40 50 600 10 20 30 40 50 600

200

400

600

800

1000

mgL−1 TOC

0 10 20 30 40 50 600

200

400

600

800

1000

0 10 20 30 40 50 60

r = 0.409 r = 0.756

r = 0.631 r = 0.657

r = 0.167

mgL−1 TOC

0 10 20 30 40 50 60

NORDIC r = 0.992SRFA r = 0.997SRHA r = 0.994SRNOM r = 0.998

(a) (b)

(c) (d)

(e) (f)

Figure 2. Comparisons of the fluorescence of F-L peak versus DOCfor: (a) RIVER, y D 7Ð92x C 103Ð15; (b) BOG, y D 15Ð87x C 121Ð03;(c) SPRG, y D 4Ð15x C 45Ð8; (d) POND, y D 2Ð89x C 67Ð77; (e) STW,

y D 2Ð09x C 219Ð82; (f) IHSS samples

(Ex/Em 230/460 nm). As for the F-L fluorescence, theNorton in Hales samples showed a higher fluorescenceper milligram of carbon for the H-L peak than the IHSSstandards.

DISCUSSION

Figures 2 and 3 demonstrate that H-L and F-L fluo-rophores have a correlation that varies with sample site.The strongest correlations are in the BOG and PONDsamples, where we hypothesize that natural H-L and F-Lmaterial dominates the DOM pool. In contrast, the weak-est relationship occurs in the final treated effluent, a watersource where H-L and F-L materials make up a relativelysmall component of DOM. These results match thoseof Baker (2002), investigating a small urban catchment.There, the F-L–DOC correlation for the whole catchmentwas r D 0Ð68, again with stronger correlations in the trib-utary containing greater proportions of natural DOM andpoorer correlations in the subcatchments where anthro-pogenic influences were stronger (Table II). We suggestthat the strength of the F-L–DOC correlation could beused as an indicator of anthropogenic influence on DOM.However, within-stream processing of DOM and mixingof DOM from different sources also has to be taken intoconsideration. For example, Clark et al. (2002) observedremarkable correlations (0Ð999) with DOM fluorescence

0 10 20 30 40 50 60 0 10 20 30 40 50 60

0 10 20 30 40 50 60 0 10 20 30 40 50 60

0 10 20 30 40 50 60 0 10 20 30 40 50 60

Inte

nsity

Inte

nsity

Inte

nsity

0

200

400

600

800

1000

0

200

400

600

800

1000

mgL−1 TOC

0

200

400

600

800

1000

mgL−1 TOC

NORDIC r = 0.985SRFA r = 0.961SRHA r = 0.984SRNOM r = 0.956

r = 0.609

r = 0.463

r = 0.630

r = 0.504

r = 0.277

(a) (b)

(c) (d)

(e) (f)

Figure 3. Comparison of the fluorescence of H-L peak versus DOCfor: (a) RIVER, y D 9Ð42x C 224Ð62; (b) BOG, y D 14Ð42x C 377Ð3 (onedata point is off scale, DOC D 16Ð5, Int. D 1558, not shown tofacilitate equal scaling); (c) SPRG, y D 6Ð8621x � 80Ð69; (d) POND,y D 6Ð66x C 150Ð88; (e) STW, y D 2Ð919x C 299Ð69; (f) IHSS samples

and DOC when they went from a fresh water to a salinewater (Shark Head River to Florida Bay). Burdige et al.(2004) also reported good correlations from contrastingsites in the Chesapeake Bay, USA, with r D 0Ð92. Incontrast, the scatter observed in the BOG, SPRG andRIVER TOC–fluorescence intensity relationships indi-cates the variability and instability of natural DOM whensampled closer to its source, as originally hypothesized inthe river continuum concept (Vannote et al., 1980). Daw-son et al. (2001) showed changes in carbon fluxes in anupland stream (CO2 degassing) and that the source of thenatural organic matter (NOM) may be unstable due to itsstate of decomposition and tributary contribution. By thetime DOM reaches the estuary, within-stream process-ing and mixing of DOM from different sources leads toa final DOC–fluorescence relationship that is dominatedby the stable, recalcitrant fluorescent fraction, leading toa stronger correlation.

The F-L fluorescence per gram of carbon of our sam-ples (Table I and Figure 4) was shown to vary betweensample sites. The BOG samples, with arguably the fresh-est DOM, have a greater fluorescence per gram of carbonthan the river samples. F-L fluorescence per gram ofcarbon was also observed to be greater than the F-LIHSS standards, especially for the BOG samples site. Forthe H-L fluorescence per gram of carbon, this differencebetween our field samples and humic IHSS standards was



Table II. Comparison of the correlation coefficient between TOC and F-L fluorescence intensity with other studies

Study Site ID Description Correlation coefficient Comments

Ouseburn (Baker, 2002)Tributaries 4 Harey Burn 0Ð45 Impacted, sewerage failures

11 Brunton tributary 0Ð78 Unimpacted, agricultural land cover17 Gosforth Park tributary 0Ð53 Impacted from occasional CSO discharges21 The Letch 0Ð75 Unimpacted, suburban and agriculture

22a Town Moor Tributary 0Ð78 Unimpacted, grassland and suburban6 Abbotswood Burn �0Ð20 Impacted, cross connected sewers8 Airport tributary �0Ð30 Impacted, airport deicer

Main stream 8 Woolsington 0Ð82 Unimpacted, agricultural land cover10 Brunton Bridge �0Ð34 Downstream tributaries 6 & 813 Red House Farm �0Ð17 Downstream of tributary 1116 Great N Road �0Ð29 Downstream of tributary 1722 Castle Dean gauge 0Ð42 Downstream of tributaries 21 & 22

River Tern (this study) RIVER 0Ð87BOG 0Ð76SPRNG 0Ð63POND 0Ð66STW FTE 0Ð17

Clark et al. (2002) Florida Bay 0Ð99 EstuaryBurdige et al. (2004) Chesapeake Bay 0Ð92 Estuary

mgL−1 DOC

0 10 20 30 40 50 60

Ful

vic

- lik

e In

tens

ity

0

200

400

600

800

1000

RIVERPONDSPRGBOGSTWNordic FASRFASRHASRNOM

Figure 4. DOC versus F-L fluorescence intensity for all samples, showingthe different fluorescence intensities per gram of carbon for different

carbon sources

even greater. These results suggest that a fluorescent frac-tion is lost in the IHSS standard preparation procedure,especially the humic (SRHA) standard. This is in agree-ment with previous investigations that have suggestedthat no ideal system is available for isolating pure hypo-thetical humic substances from a water sample, and thatcertain changes in the structural composition of the DOMtake place during each isolation procedure based on achemically assisted sorption–desorption technique (Peu-ravuori et al., 2005). Studies that use IHSS standards (e.g.Mobed et al., 1996; Klapper et al., 2002; Chen et al.,2003; Alberts and Tackacs, 2004; Sierra et al., 2005), and

in particular humic standards to quantify carbon fluxesfrom DOM fluorescence properties, could be consider-ably overestimating carbon quantities.

CONCLUSIONS

We demonstrate different fluorescence intensity–DOCrelationships for H-L and F-L fluorescence centres,depending on the source of DOM. These relationshipsdiffer in both gradient, i.e. the fluorescence per gramof carbon, and in the strength of the correlation coef-ficient. Strongest correlations are observed at sites wherenatural DOM dominates the DOC pool, whereas greaterfluorescence per gram of carbon is observed in our peatwetlands than with within-river DOM. IHSS standards,in particular humic extracts, have significantly lower flu-orescence per gram of carbon than our field samples.Assuming that all catchments have sufficient variabilityin both DOC and fluorescence, we propose the strengthof the fluorescence–TOC correlation to be another usefultool when discriminating sources of NOM in freshwaters.Even when there is little enough variability of DOC andfluorescence, our results show significantly different flu-orescence per gram of carbon between sources. Analysesof both fluorescence and DOC potentially fill a gap inthe available data required to identify the mechanisticprocess, and magnitudes of, aquatic carbon fluxes to theestuary (Eatherall et al., 1998).

ACKNOWLEDGEMENTS

We would like to thank Andy Moss, Richard Johnson andIan Morrissey for excellent laboratory and field support,Kevin Burkhill (Birmingham University) for Figure 1and Martin Heaps and Geoff Warren for laboratoryanalysis at Reading University. The project was funded



by the NERC LOCAR programme (to SC) and fundingfrom a Philip Leverhulme Prize (to AB). The manuscriptbenefited from the helpful comments of three anonymousreferees.

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the freshwater dissolved organic matter fluorescence–total organic carbon relationship

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